1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

LABORATORY INVESTIGATION FOR PRIMARY Note:

HEMOSTASIS • The main difference between of the Duke’s and
PRIMARY HEMOSTASIS Ivy’s method would be the site where the
• Role of platelets and blood vessels in response puncture is made.
to vascular injury • Duke’s method and Ivy’s method are the most
LABORATORY TESTS common method used to assess bleeding time.
1 Bleeding Time (BT)
2 Capillary Fragility Test (CFT) CAPILLARY FRAGILITY TEST
3 Clot Retraction Test (CRT) • Measurement of the stability of capillaries under
4 Platelet Tests (Quantitative & Qualitative) condition of increased pressure or hypoxia
BLEEDING TIME (BT) o We measure how stable will your capillaries
be when subjected to increased pressure or
• In-vivo measurement of platelet adhesion and
hypoxia
aggregation on the locally injured vascular
o Usually, we induce increased pressure or
endothelium
hypoxia by using a bp cuff
• How it is performed?
o puncturing the skin and right after, let the DIFFERENT METHODS FOR CAPILLARY FRAGILITY TEST
blood ooze out • RUMPEL LEEDE TEST
• BT is basically the length of time for the blood to § 100mmHg for 5 minutes
ooze out • QUICK’S TEST
• Endpoint of BT: when the blood stops to ooze out § Inflate between the systolic and diastolic
PRECAUTIONS pressures for 5 minutes
• ASPIRIN INTAKE • GOTHLIN’S TEST
§ can prolong the bleeding time § Both arms inflated at 35mmHg for 15 minutes
§ aspirin inhibits thromboxane A2 (txA2) Notes:
synthesis The difference of these 3 methods would be the
o txA2: protein that facilitates and pressure applied to the bp cuff (pressure set) or the
promotes platelet aggregation time it takes or the time how long would the bp cuff
§ txA2 inhibition by aspirin leads to less platelet stay or left in your arm.
aggregation→ Prolonged bleeding time
• Increased/Prolonged Bleeding Time
§ caused by either impaired platelet function
(aggregation/adhesion) or vascular disorders
that may make your blood vessels fragile
DIFFERENT METHODS IN PERFORMING BLEEDING TIME
• DUKE’S METHOD
§ The puncture is usually made on the earlobe
§ We only puncture the earlobe once • PETECHIAE
§ 1-3 minutes § In the capillary fragility test, we will wait and
• IVY’S METHOD see for the formation of petechiae.
§ The puncture is usually made on the forearm § These are tiny spots of blood in your skin
§ We puncture the forearm or we make 2-3 caused by ruptured capillaries.
punctures in the forearm § When the capillaries rupture, the tendency is
§ We need to place a bp cuff in the arm of the blood will leak out into your extracellular
patient and then this bp cuff should be spaces and eventually to the skin forming tiny
inflated to a pressure of 40mmHg patches.
§ 1-7 minutes HOW DOES CAPILLARY FRAGILITY TEST ASSESS
• OTHER METHODS PLATELET FUNCTION AND PLATELET NUMBER?
§ Macfarlane’s Method • As mentioned in the previous slide, we assess the
§ Mielke’s Method (9 X 1mm) stability of capillaries. If capillaries are subjected
§ Simplate Method (5 X 1 mm) to high pressure, and even high apoxia, the
tendency is that they would easily break. If a

T R AN S F O R M E R S 1
1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

blood vessel breaks, the body would activate its • Once we get the result, we multiply it by one
primary hemostasis mechanism, so there would hundred (100)
be the formation of a platelet plug. volume of serum expressed
% Clot Retraction = × 100
§ If there is a ruptured vessel, the normal volume of whole blood

response of the body is that it would form an

NORMAL VALUE FOR CLOT RETRACTION TIME
initial platelet plug to seal of the injured vessel,
• NV- 44-64% after 2 hours
so once this injured vessel is sealed, then
NORMAL CLOT
blood will no longer leak out.
• If you have normal platelet count and normal
• If you have a normal amount of platelet, and
platelet function, you would produce a normal
they are functioning normally, upon subjecting
clot.
capillaries to high pressure what will happen?
• Its appearance is firm, elastic, dry
Ø Since the platelets are normally
• When laid or placed on a flat surface (should
functioning, what will happen is that
have sufficient turgor to maintain its shape)
there would only be FEW petechiae
would maintain its shape
will appear to your arm, due to the
• From the procedures of performing CRT, the
platelets being able to seal the
quantitative method, we put an applicator stick
ruptured capillaries.
inside the tube.
• What would happen if you have platelets that
• A normal clot would adhere to the applicator
are poorly functioning?
stick and should not fall off.
• The platelets will not be able to seal the injured
vessel, and would lead to a formation of a lot of
petechiae in your arm.

CLOT RETRACTION TIME

• Obsolete test used to assess platelet as well
• It has two methods
DIFFERENT METHODS IN PERFORMING BLEEDING TIME
QUALITATIVE METHOD DEFECTIVE CLOT
HIRSCHBOECK OR CASTOR OIL METHOD • Thrombocyte Deficiencies
• uses a glass slide and only a drop of blood Clot non-retractile or clots poorly
• dimpling phenomenon or extrusion of serum on § Thrombocytopenia - decrease number of
the surface of the drop of clotted blood platelets
• NV: 15-45 mins § Thrombasthenia - weak or poorly functioning
QUANTITATIVE METHOD platelets
MACFARLANE’S METHOD • Fibrinogen Deficiencies
• uses a graduated tube or a centrifuge tube with o In cases of fibrinogen deficiencies, the blood
graduations and a large volume of blood will not clot.
§ In the graduated tube, we use an applicator § Afibrinogenemia- blood does not clot
stick in the middle § Hypofibrinogenemia- clot is
• The clot attached to the stick is removed small/increased red cell fall out
• Serum extracted from the clot is measured • Delayed Clotting
• Retraction is expressed in terms of serum o In cases of coagulation factor deficiencies,
obtained from the original volume of blood therefore, you will also have a slow clotting,
COMPUTATION FOR CLOT RETRACTION TIME your blood will slowly clot, it will not clot
How do we compute for the Clot Retraction Time? within the allotted two hours for the test.
• To compute for the percent (%) clot retraction, Slow clotting with slow sedimentation of red cells
we must first measure the volume of serum § Hemophilloid state
expressed § Presence of circulating anticoagulants
• Once we compute for the volume of the serum
express, we divide it by the volume of the whole
blood that we used

T R AN S F O R M E R S 2
1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

§ Take note: “We do not add all four in one

preparation so we prepare let’s say 4 tubes
and we add PRP in each of these tubes. After
we prepare 4 tubes with PRP, in each tube,
we will then add the different agonist (Tube 1:
ADP, Tube 2: Collagen; Tube 3: Epinephrine;
Tube 4: Ristocetin). After preparing these
mixture…
§ Measure the amount of LIGHT TRANSMITTED
using your Aggregometer.
• Again, the clot retraction time this is mainly § Take note: “If there is increased platelet
dependent or proportional to the number aggregation, then there will be more light
and the quality of your platelets so the clot transmitted from the solution. MORE PLATELET
retraction time it cannot assess or it is not AGGREGATING, THERE WILL BE LIGHTER
associated with fibrinogen concentration TRANSMITTED. SO diba ganito yan, PRP
and not associated as well with packed red (Platelet Rich Plasma) so meaning your
cell volume. Also, if you have increased plasma is still filled with platelet. SO maraming
fibrinolytic activity, then you will have an maraming platelet pa yung plasma mo. SO
impaired clot retraction. dun sa platelet rich plasma mo, you added
an aggregating agent. So, what would
PLATELET TESTS happen? This aggregating agent will
Divided into quantitative and qualitative tests. promote your platelets to clump up together.
QUANTITATIVE TESTS: DIRECT AND INDIRECT PLATELET Kapag nag clump/aggregate ung platelets
COUNT mo, what would happen? Eventually your
Involved platelet counting. platelets will aggregate nga and it will clear
1 Rees and Ecker off the plasma. Kasi nga mawawala na ung
2 Fonio’s Method platelets dahil mag kukumpulan sila. So, your
3 Brecker-Cronkite plasma would become clear. The more na
4 Dameshek nagaaggregate ung platelets mo, the more
na nagiging clear ang solution (more light is
QUALITATIVE TESTS: PLATELET FUNCTION TESTS
passing through the solution).
We assessed the platelet aggregation and platelet
adhesiveness.
1 Platelet Aggregation Test
2 Platelet Adhesiveness Test
QUANTITATIVE TESTS INDICATION OF PLATELET AGGREGATION TEST
DIRECT METHODS • Can be used to diagnose some or certain
SPECIMEN: Diluted blood diseases.
METHOD: Rees and Ecker, Brecker-Cronkite • So, for example, your Von Willebrand Disease
(VWD) and Bernard-Soulier syndrome (BSS).
INDIRECT METHODS
SPECIMEN: Blood smear
METHOD: Fonio’s Method and Dameshek Method
QUALITATIVE TESTS
PLATELET AGGREGATION TEST
• “We measure how well your platelet aggregate
in the presence of an aggregating agent”
• HOW IS THIS TEST PERFORMED?
§ First, prepare a platelet rich plasma (PRP)…
§ Then add Aggregating Agent… (or an
agonist. These agonists are your ADP,
Collagen, Epinephrine and Ristocetin.

T R AN S F O R M E R S 3
1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

VWD and BSS Glanzmann’s venipuncture.

Thrombasthenia
• Normal: ACE • Normal: Ristocetin Why is there lower platelet count in the glass bead
• Abn: Ritocetin • Abn: ACE collecting system?
• Normal aggregation • There would be • Because your platelet should adhere to the glass
in ADP collagen and normal aggregation beads kaya when you perform platelet count in
epinephrine but in ristocetin but this glass bead collecting system, dapat mas
would have abnormal mababa ang platelet count.
abnormal results in aggregation in ADP
ristocetin. collagen and LABORATORY DEMONSTRATION OF TESTS FOR PRIMARY
epinephrine. HEMOSTASIS
PLATELET ADHESION TEST BLEEDING TIME (BT)
(SALZMAN METHOD) • Traditional test
• Usually performed using salsman’s method. • To assess platelet function and their interaction
• HOW IS THIS TEST DONE? with vessel walls during injury
a. TWO BLOOD SPECIMENS
a. Routine venipuncture
b. Glass bead collecting system
b. PLATELET COUNT
a. Platelet count without bead
b. Platelet count with bead

o First, we have to collect 2 blood specimens.

1 blood sample using routine venipuncture
and the other blood sample using glass MATERIALS NEEDED
bead collecting system.
• Sterile Cotton & Alcohol – to sterilize puncture
o After collecting these two-blood sample, we
site
then perform platelet count in each. (1
platelet count from the sample from routine
venipuncture and another platelet count
from the sample in the glass bead collecting
system.)
COMPUTATION FOR % PLATELET ADHESIVENESS

#$% *+. -// − #$% *+. -/ 12345

% #$% &'( = 6 788
#$% *+. -// 12345 • Sphygmomanometer (Blood Pressure Cuff –
application of constant pressure & Stethoscope)
o So, after performing the platelet count, we • Sterile disposable lancet – create puncture
can now compute for the percent platelet wounds
adhesiveness using this formula. • Circular filter paper – blotting of blood
NORMAL VALUE FOR PLATELET ADHESIVENESS
• 26 to 60% PLT ADH
• Between the sample from the routine
venipuncture and the glass bead collecting
system, there should be lesser platelet count in
the glass bead collection system in normal
conditions.

Take note:
• Normal condition: Lower Platelet count in glass • Stopwatch – record the time from the point at
bead collecting system kaysa sa routine which bleeding starts up until it completely stops.

T R AN S F O R M E R S 4
1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

IVY’S METHOD • Avoid touching the wound with filter paper while
• First, place the BP cuff on the patient’s arm just doing so. Once bleeding stopped, stop the
above the elbow. timer. Release BP cuff and record the bleeding
time of each puncture site

• Inflate at a pressure of 40 mmHg, which is

maintained throughout the test.
• Final result to be reported would be the average
of the 2 recorded bleeding times
Note: In presence of vascular or platelet disorders,
prolonged results may be observed

• Cleanse the puncture site (forearm) with 70%

alcohol. Specific puncture site should be 3 finger
width below the bend of the elbow.

• NORMAL VALUE: < 5 minutes

• Results in the Lab demonstration:
Site
1st puncture 1 min
2nd puncture 1 min
• Grasp the underside of the arm firmly and use
1 min + 1 min = 2 mins
the lancet to make two 3mm deep, puncture ! #$%&
• = 7 9:; (3=2. )
wounds. !
Note: Avoid any subcutaneous veins while creating
a puncture CAPILLARY FRAGILITY TEST (CFT)
• Aka “the tourniquet test”
• Non-specific method for evaluating bleeding
tendencies
• Measures the fragility of capillary walls when
placed under increased pressure controlled by a
BP cuff around the arm

• When blood starts to ooze out, start the timer.

Note: Blot both areas with different filter papers
every 30 seconds.

T R AN S F O R M E R S 5
1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

MATERIALS NEEDED
• BP Cuff – only equipment needed

Results: no petechiae seen = Negative

RUMPEL-LEEDE’S METHOD
QUICK’S METHOD • This is the second method of CFT.
• This method will require the patient’s blood
pressure before proceeding with the test proper.

• The BP cuff is inflated to exactly 100 mmHg.

• Start by taking the blood pressure reading. • Repeat the same steps after 5 minutes. (Examine
the area and count the petechiae present).
• Results: no petechiae seen = Negative
Note: The appearance of petechiae indicates that
small amounts of blood have escaped into the skin
which is why they are sometimes referred to as
hemorrhagic spots.
CFT POSITIVE RESULTS
• Inflate the blood pressure cuff, halfway between
the obtained systolic and diastolic pressures in
the previous step. Make sure to not exceed a
pressure of 100 mmHg.
• In this case, BP of the patient is 110/90, so it’s
halfway would be at 100 mmHg.
Patient’s BP 110/90
Halfway of systolic 100 mmHg
and diastolic

• After 5 minutes, remove the BP cuff and examine

the area for petechiae (small red spots). Count • If the capillaries are abnormally fragile, then
those petechiae in the two-inch space below more of these petechiae are expected to
the BP cuff and then record the results. appear. Both of these may indicate prolonged
bleeding tendencies.

T R AN S F O R M E R S 6
1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

FINDINGS RESULT
<10 petechiae on the Normal/Negative
forearm 5 minutes after
the test
>10 petechiae Positive
NUMBER OF PETECHIAE SCORE
A few on anterior 0-10 1+
forearm
Many on anterior 10-20 2+ • Sample will then be transferred to a graduated
forearm centrifuge tube.
Many on whole arm 20-50 3+ • The total of blood to be used in this test should
and top of hand also be noted in order to compute for the
Confluent petechiae 50 and 4+ degree of blood retraction later on.
on all areas of arm above
and top of hand
CLOT RETRACTION TIME (CRT)
• Tests normally affected by platelet number and
platelet function.
• Due to the action of platelets, the clot formed
after coagulation undergoes retraction, which
causes the extrusion of serum from the clot.
• In the presence of a decreased platelet count or
impaired platelet function, poor clot retraction is • After the transfer, place the applicator stick at
expected to occur. the center of the tube.

MATERIALS NEEDED
• Incubate in the water bath for 2 hours at 37
degrees Celsius.

• To assess this process, we will be needing a

venipuncture set for sample collection, a
graduated centrifuge tube, an applicator stick,
and a water bath for incubation. • 2 hours later...
PROCEDURE • Remove the applicator stick together with the
• 5 mL of whole blood is to be collected first via clot, which is expected to have adhered to the
venipuncture. stick during incubation.
• Please remember that normal clot retraction
should begin within 30 seconds after the blood
has been drawn.

T R AN S F O R M E R S 7
1 Laboratory Tests for Primary Hemostasis Hematology 2 (LAB)

1. Test includes description of clot, degreeof

clot retraction, serum expressed, and RBC
fallout.
2. The following are observed in a red top tube
if the fibrin clot and hematocrit level are
normal:
• Clot occupies 40% to 60% of the original
sample volume.
§ If normal clot retraction took place,
• Measure the volume of serum extruded from the then the clot is expected to occupy
clot and proceed to computing for the degree about 40% to 60% of the original
of clot retraction. volume in the tube.
• The remaining 40% to 60% consists of
serum and red cell fallout.
§ With the rest, occupies the expressed
serum and the red cell fallout.
• Red cell fall-out is usually <5% of the
original volume.
§ The amount of red cell which fall out
of the clot and settle at the bottom
of the tube is usually on less than 5%
of the original volume.
Ø Abnormalities in terms of appearance of the clot
CRT COMPUTATION
or the degree of clot retraction may be
• This is done by divided the volume of serum by
observed in the presence of platelet
the total volume of the blood used in the test.
abnormalities or fibrinogen deficiencies.
Volume of blood 3 mL
obtained
Total volume of blood 5 mL
3 @A
C 100 = F8%
5 @A
• The results represents the degree of clot
retraction which is expressed in percentage.

CRT RESULTS

• Clot description in addition to the degree may

also included.

T R AN S F O R M E R S 8
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Hematology 2
AY 2021 - 2022

HERMANSKY-PUDLAK SYNDROME

Submitted by:
Group 3 - 3B
AQUINO, Reanna Gene
CARREON, Mariane Irish
CRUZ, Francheska Angela
MENDOZA, Mickael Erika
NAMOCATCAT, Kasandra
OCAMPO, Marc Robert
SAMSON, Krystelle
SANCHEZ, Irvina
 GROUP 3 CASE PRESENTATION

Case 5:
An 8-year-old girl presented herself in the emergency room due to recurrent epistaxis. Upon
history taking, it was noted that the patient had a history of other bleeding events and
inflammatory bowel disease (IBD). Physical examination indicated that the patient had visual
impairment in the form of nystagmus and hypopigmentation of the skin and hair. There was no
significant presence of purpura or ecchymosis on any part of her body. Laboratory tests were
ordered and the following were the results:

RBC count 3 x 106/uL

WBC count 9 x 109/L

Hemoglobin 10 g/dL

Prothrombin time (PT) Normal

Partial thromboplastin time (PTT) Normal

Bleeding time (BT) Prolonged

Platelet count 276 x 109/L

1. What is the probable diagnosis of the patient? Give etiology and pathophysiology of
the disorder.

The patient may be experiencing Hermansky-Pudlak Syndrome (HPS), an autosomal

recessive disorder that is identified by two manifestations which are loss of pigmentation and
visual impairment. Both genders are affected by this disorder. However, it is common among
northwest Puerto Ricans since they are believed to be carriers of HPS Type 1 gene.
Moreover, this rare disorder is ranked as the third common form of albinism.

Ten genes are associated with HPS. These include HPS1, AP3B1, HPS3, HPS4, HPS5,
HPS6, DTNBP1, BLOC1S3, PLDN, and AP3D1. These protein-encoding genes form the
 Biogenesis of Lysosome-related Organelles Complexes (BLOCs), which synthesize
lysosome-related organelles (LROs), such as melanosomes and platelet dense granules. A
mutation in one of these genes lead to defective LROs resulting in the clinical manifestations
of HPS including oculocutaneous albinism, visual impairment, and bleeding diathesis.

Melanosomes are intracellular organelles produced by melanocytes which are responsible

for the production and storage of melanin pigment. Defective melanosomes are not able to
properly synthesize and distribute melanin to the eyes, skin, and hair, resulting in
hypopigmentation. Melanin is crucial in the development of the visual system, therefore,
individuals with HPS display different eye abnormalities such as visual acuity, nystagmus,
strabismus, and photophobia. Meanwhile, the absence or deficiency of platelet dense
granules, which contain adenine nucleotides, serotonin, and calcium, cause changes in
platelet function. Secretion of platelet cytoplasmic granules such as alpha and dense
granules are essential for normal platelet aggregation, thus, individuals affected with HPS
experience prolonged bleeding due to the slow formation of the initial platelet plug.

2. What are the clinical signs and symptoms unique to the disorder that led you to that
diagnosis?

Patients with Hermansky Pudlak Syndrome (HPS) often present distinct clinical signs and
symptoms such as oculocutaneous albinism (reduced melanin pigment in the eyes, skin and
hair) and bleeding diathesis (tendency to bleed or bruise easily).

Hypopigmentation of eyes, skin and hair

● HPS is an autosomal recessive disorder that is characterized by a reduction or complete

lack of melanin pigment because the LRO’s within the melanocytes are not able to
produce and distribute the melanin thus, resulting in light/blonde hair color, white/olive
skin color that does not tan and fully translucent irises. Most children with HPS also
experience nystagmus (uncontrolled eye movement) and generally slows down as they
grow older and is commonly seen when tired or anxious due to the increased crossing of
the optic nerve fibers. This can also lead to reduced vision and depth perception. It is
also common to be light sensitive as there is little pigment that protects the eyes from
sunlight.
 Bleeding diathesis

● Platelets that circulate in the bloodstream help in the clot formation. However, in cases of
HPS, the platelets do not function well as they lack dense granules which affects the
secondary aggregation response. Due to this, they are prone to bruising as well as
frequent or heavy nose bleeds (epistaxis). Other bleeding problems such as gingival
bleeding, postpartum hemorrhage, colonic bleeding, and prolonged bleeding during
menstruation or after tooth extraction, circumcision, or other surgeries can also occur.

In some individuals with HPS, such as with the patient presented in the case, inflammatory
bowel disease is observed. About 15% of HPS patients develop colitis (inflammation) in the
intestines. This is due to a bleeding granulomatous colitis that is similar to Crohn's disease.
Patients with HPS-1, HPS-3, HPS-4 and HPS-6 mutations are said to more likely develop
involvement of the gastrointestinal tract by a granulomatous colitis which is a severe form of
inflammatory bowel disease. This granulomatous reaction involves the intracellular
deposition of ceroid in the reticuloendothelial system, a compound believed to be derived
from lipid peroxidation. These HPS genes provide an instruction to form a protein complex
called the Biogenesis of Lysosome-related Organelles Complex 3 (BLOC-3), which is known
to trigger the activation of Rab GTP-ase proteins such as Rab 32 and Rab38. These
proteins co-locate and interact with the protein LRRK2 to transport vesicles and endosomes
that plays a vital role in the traffic and biogenesis of melanosomes and lysosomes. Hence, in
the case of HPS, this system is disordered accounting for the characteristics of albinism,
thus, showing the clinical presentation of the patient’s visual impairment in the form of
nystagmus and hypopigmentation in her skin and hair. Moreover, due to the abnormal
synthesis and function of lysosomes, the patient is unable to eliminate ceroid leading to their
accumulation in the cells of the lungs and intestines which results in pulmonary fibrosis and
colitis, respectively. HPS is known to develop at any age, thus, it can also be seen in
younger children.

3. Give hematological and other laboratory tests for diagnosis of the disorder.

When testing the platelets of a person with HPS, the dense granules will be then observed
under the electron microscope to see if the chemical substances (ADP, serotonin, calcium,
ATP, phosphate) are present in the outer membrane of the platelet. If these chemical
substances are present, a 'chocolate chip cookie' like appearance can be seen and these
 substances aid in the process of clotting. If a person has HPS, these granules (Dense
bodies) are not present which means there are no chemical substances that are present,
thus, platelets display a ‘Swiss cheese’ appearance. Patients with HPS show lack of
granulophysin/CD63 in their platelets which is a component of Dense bodies and a
lysosomal membrane marker. Platelet electron microscopy is said to be the most accurate
test for the diagnosis of HPS.

Also when testing for the platelet aggregation, impaired secondary aggregation response
can be observed as well. In addition, when bleeding time is tested the bleeding is prolonged.
But when Prothrombin time (PT), partial thromboplastin time (aPTT), and platelet counts are
tested results are normal. Other tests include Genetic linkage analysis which is
recommended for patients with Oculocutaneous albinism (OCA) and bleeding tendencies to
rule out the causative mutation. Genetic testing is highly advised to determine the specific
subtype in patients with HPS, as this can alter follow-up needs and prognosis. Serial single
gene testing which targeted analysis for the gene HPS1 (pathogenic variant) can be
recommended for Northwestern Puertorican Ancestry Individuals. A more comprehensive
genomic testing including exome and genome sequencing can be performed particularly due
to the large number of genes associated with the syndrome. This testing provides a wider
view as it can suggest a diagnosis not previously considered such as mutation of a different
gene that resulted in the same clinical manifestation to HPS.

4. What are the abnormal laboratory findings? Explain why such tests are abnormal.

The abnormal laboratory findings, in this case, are the RBC count (low), hemoglobin (low),
and bleeding time (prolonged).

The prolonged bleeding time is due to reduced platelet aggregation function resulting from
the deficient or absence of platelet-dense granules caused by a mutation in the genes
encoding for the formation of Lysosome-related organelles (LROs) in Hermansky-Pudlak
Syndrome (HPS). These dense granules contain ADP, ATP, serotonin, and calcium, which
are released to attract other platelets and constitute the secondary aggregation response.
However, in HPS, these dense granules are missing or poor; therefore, no substances are
released, resulting in the inability of blood to clot properly, thereby prolonged bleeding time.
Since the patient has a prolonged bleeding time, RBC count and hemoglobin level will also
decline. Low red blood cells due to excessive bleeding result since the loss of red blood
cells surpasses the formation of new red blood cells. Furthermore, fewer red blood cells will
also have fewer hemoglobin levels as hemoglobin is the main protein of red blood cells that
carry and deliver oxygen to all cells in the body.
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Cheng, H. (2014). Hermansky-Pudlak Syndrome. Retrieved March 5, 2022 from

https://dermnetnz.org/topics/hermansky-pudlak-syndrome

Girot, P. Le Berre, C., & Bourreille A. (2019). Crohn’s-like acute severe colitis associated
with Hermansky-Pudlak syndrome: A case report. World J Gastroenterol 2019.
https://www.wjgnet.com/1007-9327/full/v25/i8/1031.htm

Huizing, M. (2021). Hermansky-Pudlak syndrome. GeneReviews® [Internet]. Retrieved

March 5, 2022, from https://www.ncbi.nlm.nih.gov/books/NBK1287/

Huizing, M., Malicdan, M. C. V., Gochuico, B. R., & Gahl, W. A. (1993).

Hermansky-pudlak syndrome. In M. P. Adam, H. H. Ardinger, R. A. Pagon, S. E. Wallace, L. J.
Bean, K. W. Gripp, G. M. Mirzaa, & A. Amemiya (Eds.), GeneReviews®. University of
Washington, Seattle. http://www.ncbi.nlm.nih.gov/books/NBK1287/

Hurford, M. T., & Sebastiano, C. (2008). Hermansky-pudlak syndrome: report of a case

and review of the literature. International journal of clinical and experimental pathology, 1(6),
550–554. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2480580/?report=classic

National Organization for Rare Disorders (NORD). (2018). Hermansky Pudlak

Syndrome. Retrieved March 3, 2022, from
https://rarediseases.org/rare-diseases/hermansky-pudlak-syndrome/#:~:text=HPS%20is%20inh
erited%20as%20an,are%20responsible%20for%20this%20disorder.

Omar, A. (2021). Hermansky-Pudlak Syndrome Workup: Laboratory Studies, Imaging

Studies, other tests. Hermansky-Pudlak Syndrome Workup: Laboratory Studies, Imaging
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 Sara. (2016). Diagnosing HPS: Hermansky-Pudlak Syndrome Network.
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https://www.hpsnetwork.org/hps-information/diagnosing-hps/

Salvaggio, H.L., Graeber, K.E., Clarke, L.E., Schlosser, B.J., Orlow, S.J., & Clarke, J.T.
(2014). Mucocutaneous Granulomatous Disease in a Patient With Hermansky-Pudlak
Syndrome. JAMA Dermatology, 150(10), 1083-1087. doi:10.1001/jamadermatol.2014.147

Suzanne Lareau, RN, MS, Bonnie Fahy, RN, MN, Donna Appell, RN, Ganesh Raghu
MD. American Thoracic Society. What is Hermansky-Pudlak Syndrome? Retrieved from March
3, 2022, from
https://www.thoracic.org/patients/patient-resources/resources/hermansky-pudlak-syndrome.pdf

Velázquez-Díaz, P., Nakajima, E., Sorkhdini, P., Hernandez-Gutierrez, A., Eberle, A.,
Yang, D., & Zhou, Y. (2021). Hermansky-pudlak syndrome and lung disease: Pathogenesis and
therapeutics. Frontiers in Pharmacology, 12, 644671. https://doi.org/10.3389/fphar.2021.644671
 ANGELES UNIVERSITY FOUNDATION
COLLEGE OF ALLIED MEDICAL PROFESSIONS
DEPARTMENT OF MEDICAL TECHNOLOGY

Case Study #3
Hematology 2 (Laboratory)

Diagnosis: May-Hegglin Anomaly

GROUP 3 | BSMT - 3F

Submitted by:
Chu, Lin Dominic
Guiao, Jemima
Halili, Christian Nicole
Moreno, Uriel
Ponio, Mica Alyssa
Ramos, Jan Marie
Reyes, Klouja Chantal Erica Alessandra

1
I. Summary of the Case

A 43-year-old woman arrived at her primary care physician for a routine medical exam.
She had no major complaints except for mild gingival bleeding and weaker hearing. Her
medical record shows years of chronic stable thrombocytopenia, occasional mild bleeding,
and easy bruising but she had no history of severe hemorrhage or transfusions despite
delivering a baby through C-section 10 years ago.

II. Lab Results

The results of the laboratory tests are as follows:

TESTS RESULTS NORMAL RANGE

RBC count 5 x 10^6/uL 3.80 - 5.20 million/cubic mm

(NORMAL)

WBC count 10.4 x 10^9/L 3.6 - 10.6 x 10"/uL

(NORMAL)

Hemoglobin 14 g/dL 12 - 16 gm/dl

(NORMAL)

Hematocrit 42% 37 - 52 %
(NORMAL)

Platelet count 55 x 10^9/L 157 - 371 x10^9/L

(LOW)

MPV 15 fL 7 – 12 fL
(HIGH)

Peripheral blood smear shows normocytic normochromic red blood cells. Large
platelets are also observed while leukocytes are normal except for the appearance of large
crystalline inclusions on granulocytes and monocytes.

2
III. Answers to the Guide Questions
1. What is the probable diagnosis of the patient?
a. Etiology of the disorder.

May - Hegglin Anomaly, which falls under the MYH-9-related Thrombocytopenia

Syndromes, is the most likely diagnosis for the patient. May - Hegglin Anomaly is a rare,
inherited blood platelet disorder characterized by the triad of thrombocytopenia, abnormally
large platelets, and inclusion (Döhle-like) bodies in leukocytes (National Organization for Rare
Disorders, Inc., 2013). This condition is caused by a mutation in the nonmuscle myosin heavy
chain 9 gene (MYH9) on chromosome 22q12-13. This MYH9 gene is considered vital in the
formation of a protein, Myosin 9, which is a subunit of the myosin IIA or the protein that
regulates platelet maturation and their release (Turgeon, 2018). This myosin II can be further
divided into three forms: myosin IIA, myosin IIB and myosin IIC, which all contribute to cell
movement, maintenance of cell structures and cytokinesis. Thus, the modifications in this
specific MYH9 gene results in the disordered production of myosin heavy chain type IIA, which
affects megakaryocyte maturation and platelet fragmentation, subsequently affecting the overall
production of platelets (Keohane, Walenga, & Otto, 2019).

May-Hegglin anomaly is also an autosomal dominant disorder determined by the

combination of genes for a particular trait on the chromosomes received from the father and the
mother. Therefore, the abnormal gene can be inherited from either parent or result from a new
mutation in the affected individual (Kanwar, 2019). Dominant genetic disorders can still manifest
even if only a single gene copy is abnormal. As a result, regardless of the sex of the resulting
child, the risk of passing the faulty gene from affected parent to children is 50% for each
pregnancy.

b. Pathophysiology of the disorder.

The causation of MYH9 gene mutation for May-Hegglin anomaly (MHA) existence roots
for the dysfunctional non muscle myosin heavy chain class IIA (NMMHC-IIA) production
(Keohane, Otto & Walenga, 2019). From McPherson and Pincus (2017), this specific mutation in
MHA alters the assembly and stability of myosin, leading to a defective megakaryocyte
maturation. The result of this faulty maturation process then, is an insufficient bone marrow
megakaryocytes or megakaryocytic hypoplasia, as manifested in this congenital disorder

3
(Keohane, Otto & Walenga, 2019). A decreased quantity of megakaryocytes also corresponds
to a reduced platelet count, since the formation of platelets originates from the shedding of
megakaryocyte cytoplasm. Thus, thrombocytopenia is experienced by patients with MHA such
as in the case study. The medical record of the 43-year-old woman shows having chronic stable
thrombocytopenia for years, which coheres with her platelet count of 55X10^9/L only.

The role of platelets mainly involves hemostasis or the stoppage of blood, as a response
to an injury or break in the blood vessels. However, if there is a decreased number of platelets,
its function cannot be executed properly, resulting in hemorrhage or excessive bleeding and
bruising easily (Cleveland Clinic, 2020). Depending on the degree of thrombocytopenia,
individuals with MHA may suffer from severe to mild bleeding tendencies (Keohane, Otto &
Walenga, 2020). With severe hemorrhage, the need of platelet transfusion is highly required.
Mild bleeding, on the other hand, doesn’t compel the same treatment such as in this instance.
Aside from bleeding, other clinical features of the patients such as renal failure, loss of hearing
and congenital cataracts, can also arise depending on where the mutations in the MYH9 gene
are located. (Hanson, 2020). But, in general, MHA-affected persons are often asymptomatic
(Kanwar & Yaish, 2019).

As a MHY9 thrombocytopenia disorder, it can be differentiated by other MYH9-related

thrombocytopenia syndromes such as Fechtner syndrome, Sebastian syndrome and Epstein
syndrome through clinical manifestations. All of these disorders have macrothrombocytopenia,
but these are distinguished by other characteristics such as nephritis, sensorineural deafness,
and cataracts.(McPherson & Pincus, 2017). The three stated features are present in Fechtner
syndrome, whereas, none of these indicators are observed in both, May Hegglin anomaly and
Sebastian syndrome. To discern Epstein syndrome, its symptoms include renal disease and
deafness only.

Specifically, the low platelet count of MHA is termed macrothrombocytopenia as it

includes the presence of large sized platelets. This rare autosomal dominant disorder may be
manifested in a triad: thrombocytopenia, enlarged platelets and inclusion bodies (Kanwar &
Yaish, 2019). Based on Keohane, Otto and Walenga (2020), the peripheral blood film of MHA
consists of large platelets, and neutrophils and monocytes with Döhle-like bodies. The presence
of these large platelets signifies these are prematurely released from the bone marrow, as a
result of an increased demand in the circulation. According to Hanson (2020), the gene mutation
also contributes in stimulating the release of the premature platelets.

4
 2. Which of the given information in the case supports your diagnosis?

As an autosomal dominant disorder, higher probability of inheriting MHA can be the case
for individuals with an infected parent. In terms of clinical features, this disease is characterized
by various degrees of thrombocytopenia associated with bleeding and purpura. For this reason,
the manifestations of the female patient coheres with such a condition since, as stated in her
medical profile, she’s been experiencing chronic stable thrombocytopenia, occasional mild
bleeding, and bruising. Therefore, it can be a potential evidence that she has MHA due to the
common symptoms that have appeared (Kanwar, 2019). The demonstration of gingival bleeding
and easy bruising can also be attributed to the low platelet count or thrombocytopenia
associated with the defective megakaryocyte maturation and fragmentation from the MYH9
gene mutation (Untanu, 2021). In addition, the patient also experiences hearing loss, which is
manifested in people with the disorder.

Furthermore, another piece of evidence can be sourced from the laboratory findings.
Despite having normal red blood cells (RBCs), proof that the patient has MHA is due to the
presence of large platelets and inclusions (Döhle-like bodies) on granulocytes such basophils,
eosinophils, neutrophils, and monocytes. These inclusions are composed of myosin heavy
chains, hence, their development can be caused by the alteration of MYH9 gene. Moreover, in
platelet morphology, the mean platelet volume (MPV) of the patient is increased by 15
femtoliters (fL). According to Osborn (2018), an increased MPV signifies that platelets are larger
in contrast to the average size, thus correlating with MPV of those who have MHA, which
ranges from 15 to as high as 30 fL.

3. Give hematological or other laboratory tests to confirm the diagnosis of the

disorder.

Identification of a causative mutation in the MYH9 gene is a definitive diagnosis of

May-Hegglin Anomaly disorder, as it is part of the MYH9-related macrothrombocytopenia
disorders. To differentiate from the other MYH9-related disorders and to confirm the diagnosis of
the May-Hegglin Anomaly disorder, May-Grünwald-Giemsa-stained peripheral blood smear is
highly suggested to be performed to identify inclusion bodies present (Greer et al., 2013).
May-Hegglin Anomaly has well-defined, pyroninophilic, and basophilic inclusions found in
neutrophils, basophils, eosinophils, and monocytes. Additionally, the inclusions of MHA
resemble the appearance of Dohle bodies, but more often appear elongated, discrete, and

5
larger. Ultrastructural analysis can also be performed to further differentiate the appearance of
the inclusions between the MYH9-related disorder (Greer et al., 2013).

May-Hegglin Anomaly is also characterized by having a positive result in tourniquet test,

prolonged bleeding and clot retention time, increased mean platelet volume, decreased platelet
count, increased platelet serotonin, and increased platelet spreading while shown normal results
in platelet aggregation and WBC count (Greer et al., 2013).

4. How is the patient’s condition managed?

In general, May-Hegglin Anomaly is considered a disease that does not usually show
clinically significant bleeding problems, thus, usually discovered by chance (Untanu, 2021). The
severity of bleeding depends on the degree of thrombocytopenia acquired. For cases of mild
bleeding, treatment is not required whereas platelet transfusions are administered for those
suffering from severe bleeding. But, prophylactic platelet transfusions are not routinely used
during surgery and delivery.
To address low platelet count, intravenous desmopressin is usually used preoperatively
as a nonspecific agent to improve hemostasis. While medications such as corticosteroids and
the surgery of splenectomy are ineffective with this disease (Kanwar,2019).
If MHA is diagnosed among pregnant women, episodes of bleeding may be experienced
thus, there is a need for monitoring of abnormal bleeding and/or hemorrhages.
Lastly, genetic counseling may be necessary for those who have both mild and severe
cases to ease the worry or fear of the patient and their families (National Organization for Rare
Disorders, Inc., 2017) .

IV. Reference:
Kanwar, V. (2019). May-Hegglin Anomaly: Practice Essentials, Background, Pathophysiology.
EMedicine. Retrieved from https://emedicine.medscape.com/article/956447-overview
Keohane, E. M., Walenga, J. M., & Otto, C. N. (2019). Rodak’s Hematology : Clinical principles
and Applications (6th ed.). St. Louis, Missouri.: Elsevier.
Mary Louise Turgeon. (2018). Clinical hematology : theory and procedures. Philadelphia, Pa:
Wolters Kluwer.Mcpherson, R. A., & Pincus, M. R. (2017).

6
Henry’s clinical diagnosis and management by laboratory methods (23rd ed.). St. Louis,
Missouri: Elsevier.
National Organization for Rare Disorders, Inc. (2013). May Hegglin Anomaly. Retrieved from
NORD (National Organization for Rare Disorders) website:
https://rarediseases.org/rare-diseases/may-hegglin-anomaly/
Untanu, R. V., & Vajpayee, N. (2021). May Hegglin Anomaly. PubMed; StatPearls Publishing.
https://www.ncbi.nlm.nih.gov/books/NBK441952/
McPherson, R. & Pincus, M. (2017). Henry’s Clinical Diagnosis and Management by Laboratory
Methods (23rd Edition). St.Louis, Missouri: Elsevier Inc.
Rodak, B. F. & Keohane, E. M. (2012). Hematology: Clinical principles and applications (6th
ed.). St.Louis, Missouri: Elsevier Saunders.
Cleveland Clinic. (2020, November 30). Thrombocytopenia. Retrieved from
https://my.clevelandclinic.org/health/diseases/14430-thrombocytopenia
Hanson, M. J. (2020, March 6). Case Study: Understanding May-Hegglin Anomaly. Retrieved
from
https://www.clinicaladvisor.com/home/topics/hematology-information-center/case-study-u
nderstanding-may-hegglin-anomaly/
Greer, J., Arber, D., & Glader, B. (2013). Wintrobe’s Clinical Hematology (13th ed.).
Philadelphia, Pa: Lippincott Williams & Wilkins - Wolters Kluwer.
Kanwar, V. & Yaish, H. (2019, February 28). May-Hegglin Anomaly. Retrieved from
https://emedicine.medscape.com/article/956447-overview

National Organization for Rare Disorders, Inc,. (2017). May hegglin anomaly. Retrieved, from
NORD (National Organization for Rare Disorders) website :
https://rarediseases.org/rare-diseases/may-hegglin-anomaly/

7
 ANGELES UNIVERSITY FOUNDATION
ANGELES CITY
COLLEGE OF ALLIED MEDICAL PROFESSIONS
DEPARTMENT OF MEDICAL TECHNOLOGY

MTHEMA2: HEMATOLOGY 2 (LABORATORY)

MODULE 4 CASE ANALYSIS

Glanzmann
Thrombasthenia

Submitted by: GROUP#5

Cambe, Crystal Mae
De Jesus, Ma. Allaysa
Elizalde, Angela Leslie
Gatbonton, Kiel Patrick
Gelido, Jacklyn
Pangan, Vincent Earl
Quizon, Queeny Rei

Submitted to:
MTHEMA2 Instructors

March 09, 2022

Case 4:
A 28-year-old female patient experienced excessive bleeding after undergoing
gastrointestinal surgery. Once the bleeding was under control, further examinations were
conducted to determine its potential cause. Upon physical examination, petechiae and
subcutaneous hematoma were observed. Her medical history revealed frequent episodes of
epistaxis and increased bruising tendencies in previous years. Laboratory examination revealed
the following findings:

TEST RESULT Normal Value INTERPRETATION

Male: 13.5–18.0g/dL
Hemoglobin 10.5 g/dL DECREASED
Female: 12.0–15.0 g/dL

Male: 40–54%
Hematocrit 30% DECREASED
Female: 35-49%

150-450 x 109/L
Platelet count 190 x 109/L NORMAL
150,000 to 450,000/uL

Duke’s Method:
1-3 minutes
Bleeding time 15 minutes PROLONGED
Ivy’s Method: 1-7
minutes

Prothrombin time 12 seconds 12.6–14.6 seconds NORMAL

APTT 27 seconds 25–35 seconds NORMAL

Additional testing revealed no abnormalities in platelet morphology. Aggregation studies were
also performed and results showed impaired aggregation with ADP and epinephrine, slightly
abnormal aggregation with collagen, and normal response to ristocetin.

GUIDE QUESTIONS:

1. What is the most probable diagnosis of the patient? Explain why you came up with this
diagnosis.
2. Discuss the disorder’s etiology and pathophysiology. Indicate how it is related to
hemostasis as well.
3. What clinical and laboratory findings are usually associated with this disorder?
4. What treatment is given to patients with this disorder?

ANSWERS:
1. What is the most probable diagnosis of the patient? Explain why you came up with
this diagnosis.

The 28-year-old female patient's common signs, symptoms, and laboratory findings
support the most possible diagnosis of Glanzmann thrombasthenia (GT). Excessive
bleeding is a common sign of GT, and it is seen with the patient after gastrointestinal
surgery. Physical examination also reveals that the patient has visible petechiae and
hematoma. Her medical history also cites frequent episodes of epistaxis and the presence of
bruises. Her laboratory findings show GT characteristic features such as normal platelet
count and morphology, as well as a lack of aggregation with almost all activating agents
except ristocetin. The findings of these aggregation studies strongly suggest that
Glanzmann thrombasthenia is the current platelet aggregation disorder of the patient.

2. Discuss the disorder’s etiology and pathophysiology. Indicate how it is related to

hemostasis as well.

2.a. Etiology and Pathophysiology of Glanzmann Thrombasthenia (GT)

 Retrieved from: https://step2.medbullets.com/heme/121688/glanzmann-thrombasthenia#

ETIOLOGY:

Glanzmann’s Thrombasthenia (GT) is a type of rare blood clotting disorder that may be
inherited by an individual in an autosomal recessive pattern. This type of disorder is caused
by platelet membrane glycoprotein IIb/IIIa (GP IIb/IIIa) complex deficiency or abnormality.
GP IIb/IIIa is a type of membrane receptor that has the ability to bind to fibrinogen, von
Willebrand factor (vWF), fibronectin, and other adhesive ligands. In order for these proteins
to be expressed on the platelet’s surface, GP IIb/IIIa complex proteins should be produced
and put together into a complex. However, in this condition, due to the mutations on the
ITGA2B and ITGB3 genes seen on chromosome 17 that codes for GPIIb or GPIIIa, there is
no production of either of the two proteins resulting to the formation of abnormal platelets
making platelets unable to form platelet plug when bleeding occurs which results to
excessive bleeding.

PATHOPHYSIOLOGY:

Glanzmann Thrombasthenia (GT) defects in glycoprotein IIb/IIIa (GP IIb/IIIa) complex

either will it be qualitatively or quantitatively by which adhere to arginine-glycine-aspartate
peptide sequence of the fibrinogen and adhesively soluble ligands, like fibronectin and von
Willebrand factor (vWF). The mentioned soluble ligands and activation of integrin αIIb or β3
are important for platelet aggregation.
 As an addition, GP IIb/IIIa (integrin αIIb or β3) is known to be a heterodimeric receptor
that occurs in large volume in platelet plasma membranes. This integrin composed a low
affinity for ligands during resting state. Through exposure to subendothelial matrix (soluble
agonists) during platelet activation, an “inside-out” cellular signaling will generate
conformational changes or evolvement in GP IIb/IIIa.

2.b. Correlation of GP IIb/IIIa in Primary Hemostasis.

Furthermore, due to GP IIb/IIIa conformational change, this will now allow high affinity
binding to fibrinogen by which will now play an important role to serve as a “bridge” for the 2
adjacent platelets to be together along in the presence of calcium, therefore will initiate
platelet plug formation and platelet aggregation.

In relation to hemostasis, a platelet plug must be formed on the site of injury in order to
stop the blood from leaking out of the blood vessel which occurs during primary hemostasis.
vWF will be exposed on the injured site which allows the platelet attachment. Platelets will
then adhere to the injured site through the vWF. Once platelet adhesion occurs, their shapes
will then change and will start secreting their granules to recruit more platelets to form
platelet plugs. Since GP IIb/IIIa is responsible for binding vWF, deficiency on this complex
will prevent the platelets from attaching to the vWF present on the injured site therefore
platelet or hemostatic plug will not be formed.

3. What clinical and laboratory findings are usually associated with this disorder?

Physical/Clinical Findings:

The disease in suspect, Glanzmann thrombasthenia, has a wide variety of symptoms

that commonly affect the diseased individuals. As stated by the Genetic and Rare Diseases
Information Center (GARD), the symptoms may differ based on each individual but are
categorized into several groups according to their commonality. First, 80-99% of patients
have prolonged bleeding time, and spontaneous or recurrent epistaxis (nosebleeds).
Second, gingival (gum) bleeding, prolonged bleeding after surgery, and easy bruising are
said to be present in 30-79% of patients. Besides these symptoms, other uncommon
manifestations may also occur in patients such as ecchymosis, gastrointestinal hemorrhage,
macroscopic hematuria, hematomas, purpura, menometrorrhagia (uterine bleeding), etc.
 Relating these symptoms to the patient, she is likely suffering from Glanzmann’s
thrombasthenia. To support this, she has several symptoms that are associated with the
disorder. First of the signs is the excessive bleeding after the GI surgery. Next, based on her
history, she suffers from the symptoms of frequent epistaxis, and bruising tendency. Finally,
as revealed in her physical exam, she presented petechiae and hematoma. Therefore, her
physical findings support the suspected condition of Glanzmann thrombasthenia.

Common Clinical Signs and Symptoms Patient’s Clinical Signs and Symptoms

Recurrent Epistaxis (80-99%) Recurrent Epistaxis

Gingival Bleeding (30-79%) Hematoma

Prolonged Bleeding after surgery (30-79%) Petechiae

Easy bruising (30-79%) Prolonged Bleeding after surgery

Easy bruising

Laboratory Findings:

Glanzmann thrombasthenia(GT) is a bleeding disorder characterized

by prolonged bleeding time/excessive bleeding. In normal hemostasis,
The formation of platelet plug is induced to prevent excessive bleeding,
however, in the condition of GT the coagulation is delayed due to the
Decrease
mutations of the gene GPIIb/IIIa (ITG αIIbβ3) which provides
Hemoglobin and
instructions to produce two parts of protein receptor, integrin
Hematocrit
alphaIIb/beta3 (αIIbβ3) which helps in the clot formation of the platelet
Levels
to bind and aggregate to another platelet and form a platelet plug.
The 28-year-old female patient experience excessive bleeding after
undergoing gastrointestinal surgery, causing a decreased level of
hemoglobin and hematocrit

Normal Platelet count and morphology are normal. The condition GT is a

Platelet count qualitative type of platelet disorder that causes a bleeding disorder due
and Platelet to the defect on platelet function, aggregation.
Morphology

Bleeding time test is an in-vivo assessment of platelet adhesion and

aggregation. A standardized puncture on the earlobe (Duke’s method)
or on the forearm (Ivy’s method).
In the condition of GT, coagulation is delayed, due to the mutations of
Prolonged the gene GPIIb/IIIa (ITG αIIbβ3) which provides instructions to
Bleeding Time produce two parts of the protein receptor known as integrin
alphaIIb/beta3 (αIIbβ3). This protein helps in clot formation to bind the
platelets together. Furthermore, due to the deficiency of this protein
integrin alphaIIb/beta3 (αIIbβ3) platelet cannot bind together to form a
clot which leads to prolonged bleeding

Normal PT is utilized to measure the coagulation factors in the common and

Prothrombin Time extrinsic pathway. In the condition of GT, the coagulation is normal
(PT) since the defect is found on the platelet's function.

Normal Activated Similar to the PT, activated partial thromboplastin time (APTT) is a
Partial screening test for the coagulation factors found in the common and
Thromboplastin intrinsic pathway. In the condition of GT, the coagulation is normal
Time (APTT) since the defect is found on the platelet's function.

Ristocetin Normal In the condition GT, all aggregating

agents (agonist) are abnormal except
Collagen Abnormal
with the agonist ristocetin, due to the

Epinephrine Abnormal defect of the glycoprotein IIb/IIIa

ADP Abnormal

4. What treatment is given to patients with this disorder?

 Glanzmann Thrombasthenia is a genetic disorder that cannot be cured, it will persist
throughout the patient’s life, and it can only be managed. Therapeutic procedures for GT
may be divided into three methods, (1) blood transfusion, (2) administration of drugs, and (3)
symptomatic treatment. These treatments are not done on a regular basis, they are usually
done before surgical operations, managing bleeding after an accident, and during
spontaneous bleeding episodes.

Blood transfusions, specifically platelet transfusions are administered when local

treatments and/or anti-fibrinolytics fail to control bleeding in patients with GT and as a
prophylaxis for patients undergoing major surgery. Platelet transfusion is considered a
standard treatment in individuals with GT since it provides for partial correction of the
functional impairment of the platelets. However, a disadvantage of this procedure is that
alloantibodies may be formed against the transfused platelets which can cause relative or
absolute platelet refractoriness, which accelerates platelet breakdown and reduces the
benefits of subsequent platelet transfusions. To avoid potential alloimmunization,
HLA-matched platelets are advised.

If platelet transfusion is deemed ineffective to the patient, an alternative treatment is the

administration of the drug NovoSeven RT, a recombinant factor VIIa product. This drug was
approved by the US FDA in 2014 to be used for the treatment of bleeding episodes and
perioperative care of GT patients who are unresponsive to platelet transfusions, whether or
not anti-platelet alloantibodies are present.

For the management of symptoms, nasal packing or application of foam soaked with
thrombin may typically be used to treat nosebleeds. Gum bleeding can be avoided by
maintaining a regular dental routine. In cases of menorrhagia, it is initially treated with
anti-fibrinolytic drugs, if these drugs are ineffective, hormone therapy with progesterone
alone or progesterone combined with estrogen should be administered to suppress
menstrual periods. Platelet transfusions or rFVIIa in conjunction with an
anti-fibrinolytic may be given before delivery to gestational patients who are at risk of
primary and secondary postpartum hemorrhage, as well as fetal bleeding.

Here is a table summarizing the treatments that are given to patients with Glanzmann
Thrombasthenia,
 Treatment Purpose

Platelet transfusion For persistent bleeding or before a major surgery

Alternative treatment for patients who do not respond to

rFVIIa (NovoSeven RT) blood transfusion, and when anti-fibrinolytic and local
measures have failed

Local measures:
Compression
Gelatin sponges
Fibrin sealants
For minor and moderate bleeding such as gingival
Topical thrombin
bleeding, epistaxis, and menorrhagia.

Anti-fibrinolytic agents:
Tranexamic acid
Epsilon aminocaproic acid
References:

Ali, Z.A. (2021). Glanzmann Thrombasthenia. Retrieved from

https://emedicine.medscape.com/article/200311-overview

Botero, J. P., Lee, K., Branchford, B. R., et al (2020). Glanzmann thrombasthenia: genetic basis
and clinical correlates. https://www.haematologica.org/article/view/9325

Coller, B. S., Rockefeller, D., Adler, A., & Adler, F. (n.d.). Glanzmann Thrombasthenia. NORD
(National Organization for Rare Disorders).
https://rarediseases.org/rare-diseases/glanzmann-thrombasthenia/

Genetic and Rare Diseases Information Center (2011). Glanzmann Thrombasthenia. Retrieved
from https://rarediseases.info.nih.gov/diseases/2478/glanzmann-thrombasthenia

Glanzmann Thrombasthenia. NORD (National Organization for Rare Disorders). (2018, March
14). https://rarediseases.org/rare-diseases/glanzmann-thrombasthenia/

Keohane, E. M., Walenga, J. M., &amp; Otto, C. N. (2020). Rodak's hematology: Clinical
principles and applications (6th ed.). Elsevier.

Rodak, B. F., Fritsma, G. A., & Keohane, E. M. (2012). Hematology: Clinical Principles and
applications (4th ed.). Elsevier Saunders.

Solh, M., Solh, T., & Botsford, A. (2015). Glanzmann’s thrombasthenia: pathogenesis, diagnosis,
and current and emerging treatment options. Journal of Blood Medicine, 219.
https://doi.org/10.2147/jbm.s71319
 ANGELES UNIVERSITY FOUNDATION
COLLEGE OF ALLIED MEDICAL PROFESSIONS
DEPARTMENT OF MEDICAL TECHNOLOGY

Case Study #3
Hematology 2 (Laboratory)

Diagnosis: May-Hegglin Anomaly

GROUP 3 | BSMT - 3F

Submitted by:
Chu, Lin Dominic
Guiao, Jemima
Halili, Christian Nicole
Moreno, Uriel
Ponio, Mica Alyssa
Ramos, Jan Marie
Reyes, Klouja Chantal Erica Alessandra

1
I. Summary of the Case

A 43-year-old woman arrived at her primary care physician for a routine medical exam.
She had no major complaints except for mild gingival bleeding and weaker hearing. Her
medical record shows years of chronic stable thrombocytopenia, occasional mild bleeding,
and easy bruising but she had no history of severe hemorrhage or transfusions despite
delivering a baby through C-section 10 years ago.

II. Lab Results

The results of the laboratory tests are as follows:

TESTS RESULTS NORMAL RANGE

RBC count 5 x 10^6/uL 3.80 - 5.20 million/cubic mm

(NORMAL)

WBC count 10.4 x 10^9/L 3.6 - 10.6 x 10"/uL

(NORMAL)

Hemoglobin 14 g/dL 12 - 16 gm/dl

(NORMAL)

Hematocrit 42% 37 - 52 %
(NORMAL)

Platelet count 55 x 10^9/L 157 - 371 x10^9/L

(LOW)

MPV 15 fL 7 – 12 fL
(HIGH)

Peripheral blood smear shows normocytic normochromic red blood cells. Large
platelets are also observed while leukocytes are normal except for the appearance of large
crystalline inclusions on granulocytes and monocytes.

2
III. Answers to the Guide Questions
1. What is the probable diagnosis of the patient?
a. Etiology of the disorder.

May - Hegglin Anomaly, which falls under the MYH-9-related Thrombocytopenia

Syndromes, is the most likely diagnosis for the patient. May - Hegglin Anomaly is a rare,
inherited blood platelet disorder characterized by the triad of thrombocytopenia, abnormally
large platelets, and inclusion (Döhle-like) bodies in leukocytes (National Organization for Rare
Disorders, Inc., 2013). This condition is caused by a mutation in the nonmuscle myosin heavy
chain 9 gene (MYH9) on chromosome 22q12-13. This MYH9 gene is considered vital in the
formation of a protein, Myosin 9, which is a subunit of the myosin IIA or the protein that
regulates platelet maturation and their release (Turgeon, 2018). This myosin II can be further
divided into three forms: myosin IIA, myosin IIB and myosin IIC, which all contribute to cell
movement, maintenance of cell structures and cytokinesis. Thus, the modifications in this
specific MYH9 gene results in the disordered production of myosin heavy chain type IIA, which
affects megakaryocyte maturation and platelet fragmentation, subsequently affecting the overall
production of platelets (Keohane, Walenga, & Otto, 2019).

May-Hegglin anomaly is also an autosomal dominant disorder determined by the

combination of genes for a particular trait on the chromosomes received from the father and the
mother. Therefore, the abnormal gene can be inherited from either parent or result from a new
mutation in the affected individual (Kanwar, 2019). Dominant genetic disorders can still manifest
even if only a single gene copy is abnormal. As a result, regardless of the sex of the resulting
child, the risk of passing the faulty gene from affected parent to children is 50% for each
pregnancy.

b. Pathophysiology of the disorder.

The causation of MYH9 gene mutation for May-Hegglin anomaly (MHA) existence roots
for the dysfunctional non muscle myosin heavy chain class IIA (NMMHC-IIA) production
(Keohane, Otto & Walenga, 2019). From McPherson and Pincus (2017), this specific mutation in
MHA alters the assembly and stability of myosin, leading to a defective megakaryocyte
maturation. The result of this faulty maturation process then, is an insufficient bone marrow
megakaryocytes or megakaryocytic hypoplasia, as manifested in this congenital disorder

3
(Keohane, Otto & Walenga, 2019). A decreased quantity of megakaryocytes also corresponds
to a reduced platelet count, since the formation of platelets originates from the shedding of
megakaryocyte cytoplasm. Thus, thrombocytopenia is experienced by patients with MHA such
as in the case study. The medical record of the 43-year-old woman shows having chronic stable
thrombocytopenia for years, which coheres with her platelet count of 55X10^9/L only.

The role of platelets mainly involves hemostasis or the stoppage of blood, as a response
to an injury or break in the blood vessels. However, if there is a decreased number of platelets,
its function cannot be executed properly, resulting in hemorrhage or excessive bleeding and
bruising easily (Cleveland Clinic, 2020). Depending on the degree of thrombocytopenia,
individuals with MHA may suffer from severe to mild bleeding tendencies (Keohane, Otto &
Walenga, 2020). With severe hemorrhage, the need of platelet transfusion is highly required.
Mild bleeding, on the other hand, doesn’t compel the same treatment such as in this instance.
Aside from bleeding, other clinical features of the patients such as renal failure, loss of hearing
and congenital cataracts, can also arise depending on where the mutations in the MYH9 gene
are located. (Hanson, 2020). But, in general, MHA-affected persons are often asymptomatic
(Kanwar & Yaish, 2019).

As a MHY9 thrombocytopenia disorder, it can be differentiated by other MYH9-related

thrombocytopenia syndromes such as Fechtner syndrome, Sebastian syndrome and Epstein
syndrome through clinical manifestations. All of these disorders have macrothrombocytopenia,
but these are distinguished by other characteristics such as nephritis, sensorineural deafness,
and cataracts.(McPherson & Pincus, 2017). The three stated features are present in Fechtner
syndrome, whereas, none of these indicators are observed in both, May Hegglin anomaly and
Sebastian syndrome. To discern Epstein syndrome, its symptoms include renal disease and
deafness only.

Specifically, the low platelet count of MHA is termed macrothrombocytopenia as it

includes the presence of large sized platelets. This rare autosomal dominant disorder may be
manifested in a triad: thrombocytopenia, enlarged platelets and inclusion bodies (Kanwar &
Yaish, 2019). Based on Keohane, Otto and Walenga (2020), the peripheral blood film of MHA
consists of large platelets, and neutrophils and monocytes with Döhle-like bodies. The presence
of these large platelets signifies these are prematurely released from the bone marrow, as a
result of an increased demand in the circulation. According to Hanson (2020), the gene mutation
also contributes in stimulating the release of the premature platelets.

4
 2. Which of the given information in the case supports your diagnosis?

As an autosomal dominant disorder, higher probability of inheriting MHA can be the case
for individuals with an infected parent. In terms of clinical features, this disease is characterized
by various degrees of thrombocytopenia associated with bleeding and purpura. For this reason,
the manifestations of the female patient coheres with such a condition since, as stated in her
medical profile, she’s been experiencing chronic stable thrombocytopenia, occasional mild
bleeding, and bruising. Therefore, it can be a potential evidence that she has MHA due to the
common symptoms that have appeared (Kanwar, 2019). The demonstration of gingival bleeding
and easy bruising can also be attributed to the low platelet count or thrombocytopenia
associated with the defective megakaryocyte maturation and fragmentation from the MYH9
gene mutation (Untanu, 2021). In addition, the patient also experiences hearing loss, which is
manifested in people with the disorder.

Furthermore, another piece of evidence can be sourced from the laboratory findings.
Despite having normal red blood cells (RBCs), proof that the patient has MHA is due to the
presence of large platelets and inclusions (Döhle-like bodies) on granulocytes such basophils,
eosinophils, neutrophils, and monocytes. These inclusions are composed of myosin heavy
chains, hence, their development can be caused by the alteration of MYH9 gene. Moreover, in
platelet morphology, the mean platelet volume (MPV) of the patient is increased by 15
femtoliters (fL). According to Osborn (2018), an increased MPV signifies that platelets are larger
in contrast to the average size, thus correlating with MPV of those who have MHA, which
ranges from 15 to as high as 30 fL.

3. Give hematological or other laboratory tests to confirm the diagnosis of the

disorder.

Identification of a causative mutation in the MYH9 gene is a definitive diagnosis of

May-Hegglin Anomaly disorder, as it is part of the MYH9-related macrothrombocytopenia
disorders. To differentiate from the other MYH9-related disorders and to confirm the diagnosis of
the May-Hegglin Anomaly disorder, May-Grünwald-Giemsa-stained peripheral blood smear is
highly suggested to be performed to identify inclusion bodies present (Greer et al., 2013).
May-Hegglin Anomaly has well-defined, pyroninophilic, and basophilic inclusions found in
neutrophils, basophils, eosinophils, and monocytes. Additionally, the inclusions of MHA
resemble the appearance of Dohle bodies, but more often appear elongated, discrete, and

5
larger. Ultrastructural analysis can also be performed to further differentiate the appearance of
the inclusions between the MYH9-related disorder (Greer et al., 2013).

May-Hegglin Anomaly is also characterized by having a positive result in tourniquet test,

prolonged bleeding and clot retention time, increased mean platelet volume, decreased platelet
count, increased platelet serotonin, and increased platelet spreading while shown normal results
in platelet aggregation and WBC count (Greer et al., 2013).

4. How is the patient’s condition managed?

In general, May-Hegglin Anomaly is considered a disease that does not usually show
clinically significant bleeding problems, thus, usually discovered by chance (Untanu, 2021). The
severity of bleeding depends on the degree of thrombocytopenia acquired. For cases of mild
bleeding, treatment is not required whereas platelet transfusions are administered for those
suffering from severe bleeding. But, prophylactic platelet transfusions are not routinely used
during surgery and delivery.
To address low platelet count, intravenous desmopressin is usually used preoperatively
as a nonspecific agent to improve hemostasis. While medications such as corticosteroids and
the surgery of splenectomy are ineffective with this disease (Kanwar,2019).
If MHA is diagnosed among pregnant women, episodes of bleeding may be experienced
thus, there is a need for monitoring of abnormal bleeding and/or hemorrhages.
Lastly, genetic counseling may be necessary for those who have both mild and severe
cases to ease the worry or fear of the patient and their families (National Organization for Rare
Disorders, Inc., 2017) .

IV. Reference:
Kanwar, V. (2019). May-Hegglin Anomaly: Practice Essentials, Background, Pathophysiology.
EMedicine. Retrieved from https://emedicine.medscape.com/article/956447-overview
Keohane, E. M., Walenga, J. M., & Otto, C. N. (2019). Rodak’s Hematology : Clinical principles
and Applications (6th ed.). St. Louis, Missouri.: Elsevier.
Mary Louise Turgeon. (2018). Clinical hematology : theory and procedures. Philadelphia, Pa:
Wolters Kluwer.Mcpherson, R. A., & Pincus, M. R. (2017).

6
Henry’s clinical diagnosis and management by laboratory methods (23rd ed.). St. Louis,
Missouri: Elsevier.
National Organization for Rare Disorders, Inc. (2013). May Hegglin Anomaly. Retrieved from
NORD (National Organization for Rare Disorders) website:
https://rarediseases.org/rare-diseases/may-hegglin-anomaly/
Untanu, R. V., & Vajpayee, N. (2021). May Hegglin Anomaly. PubMed; StatPearls Publishing.
https://www.ncbi.nlm.nih.gov/books/NBK441952/
McPherson, R. & Pincus, M. (2017). Henry’s Clinical Diagnosis and Management by Laboratory
Methods (23rd Edition). St.Louis, Missouri: Elsevier Inc.
Rodak, B. F. & Keohane, E. M. (2012). Hematology: Clinical principles and applications (6th
ed.). St.Louis, Missouri: Elsevier Saunders.
Cleveland Clinic. (2020, November 30). Thrombocytopenia. Retrieved from
https://my.clevelandclinic.org/health/diseases/14430-thrombocytopenia
Hanson, M. J. (2020, March 6). Case Study: Understanding May-Hegglin Anomaly. Retrieved
from
https://www.clinicaladvisor.com/home/topics/hematology-information-center/case-study-u
nderstanding-may-hegglin-anomaly/
Greer, J., Arber, D., & Glader, B. (2013). Wintrobe’s Clinical Hematology (13th ed.).
Philadelphia, Pa: Lippincott Williams & Wilkins - Wolters Kluwer.
Kanwar, V. & Yaish, H. (2019, February 28). May-Hegglin Anomaly. Retrieved from
https://emedicine.medscape.com/article/956447-overview

National Organization for Rare Disorders, Inc,. (2017). May hegglin anomaly. Retrieved, from
NORD (National Organization for Rare Disorders) website :
https://rarediseases.org/rare-diseases/may-hegglin-anomaly/

7
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Kasabach-Merritt Syndrome

De Vega, Audrey

Dimabuyu, Jeom Arldein

Dizon, Fedeleen

Galang, Atria Therese

Garcia, Ella Cristine

Manuyag, Christine

Mercado, Alexandrea

Muñoz, Pamela Jasmine

BSMT 3 - F
Case Presentation

Case 2:
A 3-month-old male infant was rushed to the emergency department of Eunsang University
Medical Center due to recurrent and uncontrolled epistaxis. The infant appears lethargic and pale,
with shortness of breath. While examining the patient, the attending physician, Dr. Ko Seung-Tak
noticed several petechial rashes and purpura in the infant’s lower extremities with some on the
upper extremities. Further examination revealed several rubbery birthmarks in the abdomen,
some on the thighs, and a large one at the back near the left shoulder. Dr. Ko ordered a complete
laboratory workup including coagulation, hematologic, and immunologic tests. The results are the
following:

LAB TEST RESULTS

Platelet count 98 x 10^9/L

Prothrombin Time 29 seconds

Activated Partial Thromboplastin Time 58 seconds

D-Dimer 488 ng/mL

Coomb’s test Negative

Hemoglobin 8.2 g/L

Hematocrit 27%

Serum lactate dehydrogenase Increased

Serum bilirubin Increased

*both plasma and serum appears reddish even after repeat collection
GUIDE QUESTIONS:

1. What is the probable diagnosis of the patient? Give the etiology and pathophysiology of
the disorder. Does the disease have another name?

Based on the given case above, the probable diagnosis of the patient is Kasabach-Merritt
syndrome (KMS) which is life –threatening and has a reported high mortality rate. This disorder
is termed as the hemangioma thrombocytopenia syndrome or congenital hemangioma since
it was believed to be associated with thrombocytopenic purpura and enlarged capillary
hemangioma. It is caused by two rare vascular tumors, usually benign tumors, namely the
kaposiform hemangioendothelioma (KHE) and tufted angioma (TA) ; however, the cause of
these tumors are still unknown. This type of vascular disorder is a hereditary vascular disease
characterized by a rapid growing vascular tumor, thrombocytopenia (low platelets),
microangiopathic hemolytic anemia (destruction of red blood cells), and consumptive
coagulopathy (impaired clotting). KMS is commonly known to occur in infants and young
children.

In terms of its pathophysiology, both the KHE and TA contribute to the occurrence of KMS.
The presence of these tumors is an abnormality in endothelium that allows platelets to adhere
and aggregate. Activation of coagulation cascade will then follow as well as deposition of fibrin.
As the clot forms within the blood vessel, there would be destruction of red blood cells passing
through it. Because of this, there will be Disseminated Intravascular Coagulation (DIC) which
causes patients to be prone to bleeding diathesis since platelets and coagulation factors are
already consumed in the clot. Moreover, the involvement of platelet consumption, coagulation
cascade and microangiopathic red cell destruction takes place within the hemangioma manifested
as a rubbery birthmark most commonly seen in the face, scalp, chest or back. Death from this
condition is usually due to delayed diagnosis, cardiac failure, and life-threatening hemorrhage,
vascular and retroperitoneal lesions.

2. What are the clinical signs and symptoms unique to the disorder that led you to that
diagnosis?

The 3-month-old male infant has Kasabach-Merritt syndrome (KMS) because of the
following clinical and physical signs and symptoms that are evident in the given case. The first
reason is that the patient had recurring and uncontrollable epistaxis, which could be caused by
low platelet counts or thrombocytopenia, as seen by his platelet count lab result and a prolonged
bleeding that correlates also with his PT and aPTT test results. Small-vessel bleeding in the skin,
such as petechiae and purpura, was also noticed in the patient's lower and upper limbs is also
due to thrombocytopenia. Secondly, the patient is described as lethargic, pale, and having trouble
breathing. In addition, the patient has low hemoglobin and hematocrit, and a high serum lactate
dehydrogenase level. This may be because of the infant suffering from microangiopathic
hemolytic anemia, which is one of the most well-known features of KMS. Lastly, the presence of
giant cavernous hemangioma (vascular tumor) is also one of the unique clinical signs observed
in patients with KMS. Which is reported in the case, as numerous rubbery birthmarks in the
abdomen, some on the thighs, and a huge one on the left shoulder are seen on the patient.
Therefore, after a thorough analysis with these stated signs and symptoms and laboratory results,
we concluded that the patient is suffering from Kasabach-Merritt syndrome.

3. What is the significance of each of the tests in the table? Are they related to the main
condition? Why or why not? Explain this for each of the given tests.

Platelet Count
The number of platelets in the blood can be affected by many diseases. Platelets may be
counted to monitor or diagnose diseases, or to look for the cause of too much bleeding or clotting.
Therefore, this test is related to Kasabach-Merritt Syndrome (KMS) because this syndrome is
characterized by a coagulopathy with features of profound low platelets or thrombocytopenia.

Patient Result 98x10^9/L

Normal Value 150-450x10^9/L

Interpretation decreased

Additional Information: normal to decreased

Plt ct result in individuals w/ vascular
disorders

Prothrombin Time
This is performed to detect coagulation factor deficiencies, and is also used to monitor
treatment with oral anticoagulation therapy (coumadin, warfarin, etc.) as well. The prothrombin
time evaluates one’s ability to clot, and since KMS is a rare consumptive coagulopathy, such a
test would be helpful in diagnosing the patient.

Patient Result 29 sec

Normal Value 10-13 sec

Interpretation prolonged (abnormal coagulation mechanism)

Activated Partial Thromboplastin Time

This test, together with the prothrombin time, are both performed to detect any abnormality
in the coagulation mechanism of the body. Hence, similarly to what was mentioned from the
previous laboratory test, these tests could aid the physician in coming up with a definitive
diagnosis for the patient due to the fact that KMS is considered as a rare but life-threatening
coagulopathy of infancy.

Patient Result 58 sec

Normal Value <35 sec

Interpretation prolonged (abnormal coagulation mechanism)

D-dimer Test
The D-dimer test measures the amount of D-dimer, which is a type of protein that the body
produces to break down blood clots. It is also only detected if fibrinolysis occurs. This test is used
to monitor Disseminated Intravascular Coagulation (DIC) and to screen thromboembolic
diseases. If the patient is diagnosed with Kasabach-Merritt Syndrome, an elevated D-dimer is
usually seen in their laboratory results. It is due to vascular malformations, resulting in endothelial
wall abnormalities that cause abnormal blood flow. Thus, increasing blood clots.

Patient Result 488 ng/mL

Normal Value 0-240 ng/mL

Coomb’s Test
Coomb's test is performed to look for certain antibodies that attack RBCs. The main
principle of this test is that antihuman globulins obtained from the immunized nonhuman species
will bind to human globulins such as the IgG or complement by either free in serum or attached
to antigens on RBCs. For a patient with KMS, Coomb's test is usually negative since there is no
anti-immune process detected. Moreover, due to these antibodies that indicate a condition called
hemolytic anemia, the blood does not contain enough RBCs because they have been destroyed.

Patient Result Negative

Hemoglobin
One of the triad in diagnosing or ruling Kasabach-Merritt Syndrome is having
thrombocytopenia (low platelet count). If the bone marrow does not make enough RBCs and
platelets, this can cause a drop in hemoglobin. Usually, having a low platelet count is associated
with a low hemoglobin level. Furthermore, in cases of KMS, a decrease in hemoglobin level is
also due to the destruction of RBCs.

Patient Result 8.2 g/L

Normal Value 10.4 - 16.0 g/dL

Hematocrit
Hematocrit is essential in examining our blood for diagnosis of disease; this is used in
determining the capability of the body to deliver oxygen. This test is used to diagnose different
diseases like anemia, Chronic Kidney Disease, lung and heart disease, and many more.

Based on the laboratory results of the patient, the hematocrit value is reduced up to 27%.
Since the baby is diagnosed with Kasabach-Merritt Syndrome, the result of the Hematocrit level
will be likely to be reduced. The patient has encountered epistaxis, which is due to blood loss.
Hemorrhage will lead to a decreased result of the Hematocrit level.

Patient Result 27%

Interpretation Decreased

Normal Value (1 month- 1 year old) 32-51%

Serum Lactate dehydrogenase

This test is used to assess the level of lactate dehydrogenase or lactic acid
dehydrogenase. Due to RBC destruction, the lactate dehydrogenase and hemoglobin will be
released into the circulation. Later on, hemoglobin will be converted to indirect bilirubin. Increased
result of Serum Lactate dehydrogenase is due to microangiopathic hemolytic anemia or the
RBC destruction.
 Patient Result Increased

Normal Value (For Infants) 160 to 450 units per liter

Serum Bilirubin
Serum bilirubin blood test is used to check if the liver of the patient is functioning properly.
This test is used for the detection and diagnosis of newborn jaundice. Newborns are prone to
jaundice because their maturity of the liver of the newborns are not met yet. This is why newborns
are having a hard time getting rid of bilirubin. After the destruction of RBC, Hemoglobin is
released, and then conversion of hemoglobin to bilirubin will take place, causing increased serum
bilirubin level.

Patient Result Increased

Normal Value (For Infants) 1.0-12.0 mg/dL

References:

Coombs' test. Johns Hopkins Lupus Center. (2019, March 27). Retrieved March 9, 2022, from
https://www.hopkinslupus.org/lupus-tests/clinical-tests/coombs-test/

Elkoundi, Samali, Kartite, Tbouda, Bensghir, & Haimeur. (2016). Anesthestic management of
Kassabach-Meritt phenomenon in an adult: Case report. BMC Anesthesiology, 16(1).
https://doi.org/10.1186/s12871-016-0278-y

Huang, Zhou, Han, Li, & Wang. (2019). Successful treatment of an adult with Kasabach-Merritt
syndrome using thalidomide, vincristine, and prednisone. Journal of International Medical
Research, 47(4), 1810–1814. https://doi.org/10.1177/0300060519830242

Hall, G. W. (2001). Kasabach-Merritt Syndrome: Pathogenesis and management. British Journal

of Haematology, 112(4), 851–862. https://doi.org/10.1046/j.1365-2141.2001.02453.x

Lewis, D., & Vaidya, R. (2022, January 19). Kasabach Merritt Syndrome. Nih.gov; StatPearls
Publishing. https://www.ncbi.nlm.nih.gov/books/NBK519053/

Kasabach-Merritt phenomenon. NORD (National Organization for Rare Disorders). (2020, April
7). Retrieved March 6, 2022, from https://rarediseases.org/rare-diseases/kasabach-merritt-
phenomenon/

Kasabach-merritt syndrome. (2012). Owlapps.

http://www.owlapps.net/owlapps_apps/articles?id=5108362&lang=en

Keohane, E. M., Smith, L. J., & Walenga, J. M. (2016). Rodak’s hematology: clinical principles
and applications (5th ed). St. Louis, Missouri: Elsevier Saunders.

Mahajan, Margolin, & Iacobas. (2017). Kasabach-Merritt Phenomenon: Classic presentation and
Management Options. Clinical Medicine Insights: Blood Disorders, 10.
https://doi.org/10.1177/1179545x17699849

Osman, N. (2013). Kasabach – Merritt syndrome: A case report. Retrieved from:

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4949964/

Yadav, D., Maheshwari, A., Aneja, S., Seth, A., & Chandra, J. (2011). Neonatal Kasabach-Merritt
phenomenon. Indian journal of medical and paediatric oncology : official journal of Indian
 Society of Medical & Paediatric Oncology, 32(4), 238–241. https://doi.org/10.4103/0971-
5851.95150

Habringer, S., Boekstegers, A., Weiss, L., Hopfinger, G., Meissnitzer, T., Melchardt, T., Egle, A.
and Greil, R. (2014), Kasabach-Merritt phenomenon in hepatic angiosarcoma. Br J
Haematol, 167: 716-718. https://doi.org/10.1111/bjh.13049
ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

EHLERS-DANLOS
SYNDROME

Submitted by:

Group 5
Butiu, Dana
Canlas, Hidekoh
Capati, Angelica
Dabu, Christina
Lay, Jason
Marquez, Timothy
Ong, Johniel May
Yco, Mia

BSMT 3B

Submitted to:

Mrs. Genevieve A. Dizon

March 2022
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

CASE 1
A female patient in her early 50’s consulted a doctor regarding several physical conditions that
have manifested throughout the years. The patient is slightly overweight with smooth and
velvety skin all over her body that becomes loose, especially around the elbows and knees. The
conjunctiva is pale, the sclera bluish-white, the pupils are normal in size. The little finger was
able to extend up to the wrist. No dislocation of joints was noted. Different laboratory tests are
as follows:

LABORATORY FINDINGS
Parameter Result Reference Value Interpretation
1 Hemoglobin 12g/dL Female: 12-15 g/dL Normal
2 Platelet count 333,000/cumm 150,000-450,000cumm Normal
3 Bleeding Time 12 minutes 1-3 minutes Prolonged
(Duke’s Method)
4 Clotting Time 3 minutes 2-4 minutes Normal
(Drop Method)
5 Blood pressure 138/90mmHg 120/80 High

1) Enumerate the abnormal physical findings.

a. Female patient in early 50’s with several physical conditions that have manifested
throughout the years.

EDS is a hereditary connective tissue disorder, wherein mildly affected

individuals are usually diagnosed upon reaching early adulthood due to the gradual
development of the disease. This condition takes time before observable signs and
symptoms are manifested (Tidy, 2017) and it occurs throughout the lifetime. Moreover,
EDS is progressive during the lifetime and is affected by different factors, including age,
gender, weight, and sex hormones among females, which have been seen to influence
the probability of developing the disease. As such, milder types of the disease and their
conditions are commonly manifest in females (Ehlers Danlos Society, 2017).
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

b. Smooth and velvety skin all over her body that becomes loose (elbows and knees).

Under normal conditions, the body has connective tissues such as collagen fibers
that provide structural support and strength to the skin, blood vessels, muscles, tendons,
or ligaments. When there is a weak connective tissue caused by defective collagen
synthesis, the blood vessels and the skin is observed to be unusually soft, smooth,
loose, or stretchy with a velvety appearance. These characteristics may be attributed to
the extreme atrophic scars that are thin and wide, as well as the delicate blood vessels
in the skin surface which are prone to damage. In addition, deposited hemosiderin
spreads along the shins and extensor surfaces of knees and elbows, where the skin can
stretch more than normal (Bowen, 2017). These findings experienced by the patient, in
this case, are some of the distinguishing features of EDS.

c. Little finger was able to extend up to the wrist

Due to the abnormal structural formation of connective tissue, distal joint

hypermobility often affects large and small joints. Since the patient in the case has weak
connective tissue, the collagens are usually hyperelastic and flexible, resulting in a
higher than normal degree and range of movement of joints. It is the reason why finger
hyperreflexia occurs, where the patient can abnormally stretch her little finger up to the
wrist to such an extent without any pain felt. In line with this, women patients with
Ehlers-Danlos Syndrome (EDS) are often observed to have significantly high rates of
hypermobility of elbow and knee joints compared to men (Reuter & Fichthorn, 2019).

d. Conjunctiva is pale, the sclera bluish-white, the pupils are normal in size.

Individuals with EDS are more likely to have pale conjunctiva with thinner sclera
than the normal, or a thinning of the sclera, which may have been caused by the defects
in the collagen formation in connective tissues around the eyes. This appearance of the
eyes is indicative of fragile eyeballs. As a result, the sclera or the lining area around the
pupil is seen to be blue rather than white because the choroid layer shines through with
a blue-grey color even though the size of the pupil is normal.
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

e. No dislocation of joints was noted.

Since the patient has manifested the signs and symptoms throughout her years,
recurrent or repeated dislocations around the shoulders and ankles may or may not be
present in diagnosing the patient with EDS.

What disorder may be attributed to them?

The disorder attributed to the findings is Ehlers-Danlos syndrome (EDS). EDS is a group
of inherited connective tissue disorders characterized by abnormalities in collagen structure,
synthesis, and or processing. Collagen is one of the body's most important structural proteins
used to strengthen connective tissue and provide flexibility where it is needed. EDS has 13
subtypes and can be inherited in an autosomal dominant, recessive, or X-linked pattern. It is
caused by several gene mutations such as COL5A2, COL5A1, COL1A1, TNXB, PLOD1,
FKBP14 COL1A2, COL3A1, and ADAMTS2, depending on the subtype.

Joint hypermobility and soft, velvety skin that is stretchy and easily bruised are the most
common complications seen in EDS. Soft and velvety skin is the texture of the skin exhibited by
people with EDS. This clinical feature is due to the depletion of collagen fibrils, resulting in a thin
dermis. Connective tissues typically hold everything in place, such as the tissues or organs ,
providing support. However, people with EDS have weak connective tissues due to the
insufficient synthesis of collagen, causing the skin to stretch more than usual and joints to be
loose and flexible (hypermobility). Easy bruising is caused by fragile blood vessels, leading to a
higher risk of major bleeding events. Conjunctiva that is pale and the sclera is bluish-white is
observed in patients with EDS because of the abnormalities in collagen production in the
connective tissues around the eyes. Lastly, the sclera appears blue rather than white because
the choroid layer shines through with a blue-grey appearance.

2) How is this disorder connected to hemostasis?

As mentioned, collagen is a tough, fibrous protein that serves as one of the major structural
components of the body, abundantly seen in bones, tissues, skin, and blood vessels. It provides
support to the blood vessels, specifically in the layers of connective tissue. In line with this, EDS
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

is a heterogeneous group of related disorders affecting the genes involved in collagen

synthesis. The genetic mutation may lead to the fragility of connective tissues, significantly
affecting the strength and elasticity, a vital property for the body's structure. As a result, clinical
manifestations of EDS are presented as having a ‘velvety,’ ‘loose’ or ‘hyper extensive’ skin,
‘hypermobile’ joints, fragile blood vessels, and easy bruising tendency since patients may
possess either collagen with poor strength or insufficient amount of structurally normal collagen.

In the context of hemostasis, collagen bands are one of the pro-coagulant properties of the
damaged vascular intima that may promote platelet adhesion. The latter occurs when
subendothelial collagen is exposed to the blood circulation due to an injury (e.g., trauma) which
promotes activation and binding of platelets and von Willebrand factor. If collagen is insufficient
or dysfunctional, such as in the case of EDS, the succeeding platelet function (adhesion,
secretion, and aggregation) involved in the primary hemostasis will not further proceed to the
next components of hemostasis, resulting in excessive bleeding/hemorrhage. Moreover,
collagen insufficiency or alteration may cause the thinning of the said lining, which may
contribute to the fragility of the blood conduits resulting in an increased bruising tendency of the
patients.

With this, the most remarkable of all the laboratory findings for the given case is the bleeding
time. Other parameters (e.g., hemoglobin, clotting time) are expected to be normal in the case
of EDS as these are not directly affected by the abnormality in collagen. For instance, clotting
time is used only to assess the ability of the patient to form blood clots and is performed in-vitro.
This procedure is done with a glass slide or tube, and collagen is not a factor considered in this
condition; hence, abnormalities like in EDS will not affect the blood clot formation resulting in a
normal clotting time.

3) If CFT and PFT are performed, what are the expected results? Explain why.

In the given case, 138/90 mmHg was obtained as the systolic and diastolic pressures of the
patient. It is probable that blood pressure was obtained for the Capillary Fragility Test (CFT) via
Quick’s Method. CFT assesses the stability of capillaries by subjecting them to increased
pressure and afterward describing the number of petechiae formed on the patient’s skin.
Normally, only a few petechiae will be observed if the platelet count and function are both
 ANGELES UNIVERSITY FOUNDATION
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sufficient. These enable the sealing of ruptured vessels after being subjected to high pressure
as a result of primary hemostasis, which is a normal body response that causes the formation of
a temporary platelet plug to block the tears and breaks on injured blood vessels to prevent
blood from leaking out. In contrast, if the platelets are neither sufficient nor functional, or if the
blood vessel itself is highly permeable or unstable, numerous petechiae will be formed in the
skin of the patient due to the continuous leakage of blood. However, CFT results are not only
dependent on platelet adequacy but also on blood vessel stability.

Individuals with EDS usually have blood vessels that easily rupture. This condition can
be attributed to the common clinical feature of the syndrome: genetic abnormalities of the
collagen fibers that normally provide support to the vascular tissues. When the collagen is weak,
or when the fibers are abnormal in production, structure, or cross-linking, patients will have an
increased susceptibility to blood vessel injuries. Therefore, the EDS patient is expected to have
a positive CFT result or increased capillary fragility. This would be marked by the numerous
petechiae produced as the capillaries easily break due to defective collagen. In the given case,
the fragility of the patient’s blood vessels is even exacerbated by the hypertensive condition of
the patient. Having a high blood pressure can further damage her already weak blood vessels,
and induce more complications with her EDS. The high degree of blood vessel fragility
experienced by the EDS patient, therefore, cannot be compensated by the normal platelet count
and function alone, resulting in the formation of numerous petechiae.

Conversely, platelet function tests, namely, platelet aggregation test and platelet
adhesiveness test, will be expected to demonstrate normal results among EDS patients. First,
the platelet aggregation test measures how well platelets clump together upon the addition of
agonists or aggregating agents such as collagen and ristocetin. The degree of aggregation is
directly proportional to the light transmittance of the solution, which can be detected by an
aggregometer. Second, the platelet adhesiveness test exhibits the ability of the platelets to
adhere to glass surfaces which, consequently, may be indicative of their ability to adhere to the
blood vessel walls. Platelet count is done in two samples: one without glass beads and one with
glass beads. Normal adhesion is observed when the sample with glass beads yields lower
counts.
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

Since these tests are performed to examine the properties of platelets, they are not
affected by the mutation or abnormality that causes EDS. This condition primarily lies in the
structural integrity of blood vessel walls due to defective collagen and not the platelets
themselves. To illustrate, vascular injury normally induces the adhesion of platelets to the
exposed underlying collagen fibers in the subendothelium. This typically occurs within 1 to 2
minutes following an injury in the endothelium. Afterwards, the platelets aggregate or clump
together. However, these roles of platelets in primary hemostasis are impeded by the blood
vessel fragility due to the weakness of the subendothelial collagen. In such conditions,
interaction with the defective blood vessels results in the inefficient execution of platelet
adhesion and aggregation and, ultimately, in the failure of platelet plug formation. However,
testing in-vitro, such as through platelet function tests, enables the ability of platelets to adhere
and aggregate to be different (i.e., normal) since they are not interacting with the defective
collagen within ruptured blood vessels.

4) What treatments can be given to the patient?

Although there is currently no cure for Ehlers-Danlos Syndrome, lifelong treatment and
management is necessary to alleviate the signs and symptoms and prevent life-threatening
complications. EDS has various subtypes; hence, the treatment and management depends on
the affected body systems. The primary aspects of the treatment and management consist of
work-up in the cardiovascular system, physical therapy, management of pain, and psychological
follow-up. Surgery could also be an option for some patients with EDS, but assessment with the
severity and the type of EDS must be considered (GARD, 2017).

Physical therapy and low-resistance exercise are usually recommended for patients who are
prone to joint dislocations. Through these activities, people with EDS can strengthen their
muscles and enhance the stability of their joints. In addition, assistive devices like wheelchairs,
braces, or scooters may also be useful as their supplementary support. Moreover, prescriptions
of pain medication and high blood pressure such as beta-blockers and celiprolol, are also
recommended to manage the severe muscle and bone pain, as well as to manage the blood
pressure of the patient and lessen the fragility of the arteries as well as reduce the stress on the
collagen fibers within the arterial walls, respectively. Likewise, intake of Ascorbic acid or Vitamin
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

C aids in the reduction of bruise development and Vitamin D for the maximization of bone
density.

Routine screening for high blood pressure and arterial diseases must be initiated and
early treatment must be applied since hypertension increases the risks of vascular fragility and
complications. Furthermore, regular eye check-up must also be done since people with EDS are
at risk for eye complications such as detachment of retina, glaucoma, and globus rupture.
Psychological counseling is also necessary to help patients be aware and easily cope with the
changes that are bound to happen parallel to their disease, as well as to provide support and
empowerment while adapting to the said changes (Toucedo et. al., 2020).
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

References

Bowen, J. M., Sobey, G. J., Burrows, N. P., Colombi, M., Lavallee, M. E., Malfait, F., &
Francomano, C. A. (2017). Ehlers-Danlos syndrome, classical type. American Journal of
Medical Genetics Part C: Seminars in Medical Genetics, 175(1), 27–39.
https://doi.org/10.1002/ajmg.c.31548

Cedars Sinai. (2021). Ehlers-Danlos Syndrome (EDS). Retrieved from

https://www.cedars-sinai.org/health-library/diseases-and-conditions/e/ehlers-danlos-synd
rome-eds.html

Chen, Y., Li, J. & Atalay, A. (2021). Ehlers-Danlos Syndrome. Retrieved from
https://now.aapmr.org/ehler-danlos-syndrome/

Driscoll, D. (n.d.). Ocular Complications of Ehlers Danlos Syndrome. Total Eye Care. Retrieved
from https://totaleyecare.com/ocular-complications-ehlers-danlos-syndrome/

Ehlers danlos syndromes - NORD (national organization for rare disorders). (2015, February
11). Retrieved from NORD (National Organization for Rare Disorders) Retrieved from
https://rarediseases.org/rare-diseases/ehlers-danlos-syndrome/

Genetic and Rare Disease Information Center. (2017). Ehlers-Danlos Syndromes. Retrieved
from https://rarediseases.info.nih.gov/diseases/6322/ehlers-danlos-syndromes

Keohane, E., Smith, L., & Walenga, J. (2016). Rodak’s hematology: Clinical principles and
applications (5th ed.). St. Louis: Elsevier Saunders.

Malfait, F., Francomano, C., Byers, P., Belmont, J., Berglund, B., Black, J., Giunta, C. (2017).
The 2017 international classification of the Ehlers-Danlos syndromes. American Journal
of Medical Genetics Part C: Seminars in Medical Genetics, 175(1), 8–26.
https://doi.org/10.1002/ajmg.c.31552

National Organization for Rare Disorders (2017). Ehlers Danlos Syndromes. Retrieved from
https://rarediseases.org/rare-diseases/ehlers-danlos-syndrome/
 ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions

Reuter, P. R., & Fichthorn, K. R. (2019). Prevalence of generalized joint hypermobility,

musculoskeletal injuries, and chronic musculoskeletal pain among American university
students. PeerJ Publishing, 7, e7625. https://doi.org/10.7717/peerj.7625

Tidy, C. (2017). Ehlers-Danlos Syndrome. Patient.info. Retrieved from

https://patient.info/bones-joints-muscles/ehlers-danlos-syndrome-leaflet

Toucedo, I. & et. al (2020). Psychosocial Influence of Ehlers-Danlos Syndrome in Daily Life of
Patients: A Qualitative Study. Retrieved from:
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7503231/
HEMATOLOGY 2 LAB: CASE PRESENTATION CJLN, BSMT-3C

MODULE 3: PLATELET AND VASCULAR DISORDERS ▪ GIT bleeding

E.) LABORATORY TESTS
CASE NO.1: EHLERS-DANLOS SYNDROME • Primary Hemostasis Tests
Edvard Ehlers and Henri-Alexandre Danlos 1. Bleeding Time- prolonged bleeding
QUESTION AND ANSWER time; formation of temporary
A.) ETIOLOGY platelet plug does not proceed
• RECESSIVE 2. Capillary Fragility Test- affected;
• DOMINANT- the greatest number of positive due to weak vessels
population 3. Clot Retraction Time- normal; do not
• X-LINKED- rarest test for skin/blood vessel function
B.) VARIETIES OF EDS 4. Platelet Function Test- not affected;
• CLASSICAL TYPE- dominant; COL5A1, normal- platelets are normal in this
COL5A2, COL1A1 disease
• CLASSICAL-LIKE TYPE: COL1A1 5. Coagulation test- Normal
• Autosomal recessive: 6. Reptilase- Normal
▪ TNXB gene- Tenascin-X (flexibility) 7. aPTT- Normal
▪ TNXA gene- pseudo gene 8. Coagulation mixing studies- Normal
(complicates testing for classical-like F.) TREATMENT
EDS) • No cure
• VASCULAR TYPE-dominant • Supportive care:
▪ COL3A1 gene ▪ Physical therapy
▪ COL1A1 ▪ Assistive devices: braces, wheel
• HYPERMOBILITY TYPE- dominant chairs
▪ No genes associated
B.) S/SX (recessive and dominant)
• Childhood CASE NO. 2: KASABACH-MERRITT SYNDROME
C.) COLLAGEN “Hemangioma Thrombocytopenia Syndrome”
• Subendothelial of BV QUESTION AND ANSWER
• Weak CTs (skin and blood vessels)= bruising A.) UNIQUE FEATURES OF KMS
and bleeding TRIAD:
• Type I- skin, tendons, ligaments, bones • *Vascular tumor- Hemangiomas (root cause)
• Type II- cartilage ▪ Trap the platelets= activates and
• Type III- reticular fibers (supporting mesh for aggregates
soft organs) ▪ Leads to excessive clot formation in
• Type IV- Basement membrane/ lamina the blood vessels= bleeding
(epithelial cells- skin) tendencies
▪ Unnecessary clot formation=
• Type V- Different cell surfaces: hair, placenta,
Disseminated Intravascular
and those in type I
Coagulation
D.) DISTINCT CHARACTERISTICS
• Thrombocytopenia
• Skin hyper elasticity- “lastic man”
▪ Prone to bruising, easy wounding, • Bleeding
poor healing, prone to scars B.) WHY IS IT THE DIAGNOSIS?
• Joint hypermobility • Rubbery birth mark (Hemangioma)- right at
▪ Joint laxity- extends joints to its birth
fullest capacity without pain C.) LABORATORY TESTS
• Extensive bruising • Platelet count- decreased; trapped platelets/
▪ Fragile blood vessels, severity of consumption of platelets
bleeding (up to arterial rupture) • PT and aPTT- abnormal coagulopathy;
▪ Pregnant women: post-partum abnormal activation of platelets/ clot
bleeding
HEMATOLOGY 2 LAB: CASE PRESENTATION CJLN, BSMT-3C
formation (coagulation factors are • Defective maturation and poor
consumed) fragmentation of megakaryocytes
• Fibrinogen levels- decreased; consumptive ▪ Poor fragmentation= bigger platelets
coagulopathy are produced= reduced in production
• D-Dimer- elevated; vascular malformations number
▪ Purpose: D-dimer- by-product of clot • Problem: non-functional myosin 9 (MyH9
lysis gene mutation)
▪ Elevated D-Dimer: uncontrolled D.) OTHER MORPHOLOGIC ABNORMALITIES ASIDE
thrombosis= KMS’ uncontrolled FROM SIZE (PLATELETS)
blood formation; • No other morphologic abnormalities for
▪ Excessive thrombosis= Increased platelets= normal microscopic appearance
fibrinolysis= Increased D-Dimer E.) LABORATORY TESTS
• COOMB’s Test- Negative
▪ Microangiopathic hemolytic anemia:
non-immune hemolysis; red blood
cell destruction
▪ Red cell destruction: red cells will
squeeze through the clot blocking the
blood vessel= red cell rupture= RBC
hemolysis
• Hemoglobin and hematocrit- decreased;
slight hemolytic anemia
• Platelet Function Test- normal
• Serum Lactate Dehydrogenase- increased
• Clot Retraction Time- prolonged
• Bleeding Time- Prolonged
• Platelet Aggregation Test- normal; since it is
• Clot Retraction Time- Poor; decreased
a quantitative disorder (function is not solely
platelets/ thrombocytopenia
affected)
• Platelet aggregation studies- normal
• Platelet Retraction Test- normal
• PT and aPTT- irrelevant; these are for
secondary hemostasis
CASE NO. 3: MAY-HEGGLIN ANOMALY • Platelet Count- decreased
Discovered by: Richard May • Tourniquet Test- positive
Improved by: Dr. Robert Hegglin F.) DOHLE BODIES (round/oval) VS DOHLE-LIKE
QUESTION AND ANSWER BODIES (spindle-shaped)
A.) ETIOLOGY • Congenital/ inherited: Dohle-like bodies
• Autosomal dominant disorder (50% for every • Acquired: True Dohle bodies
pregnancy) G.) WHAT CAUSES THEIR APPEARANCE IN A SMEAR?
• Thrombocytopenia • True Dohle bodies- infections, burns, and
• Abnormally large platelets drugs
• Inclusion (Dohle-like) bodies in leukocytes H.) NEUTROPHILS IN DOHLE-LIKE BODIES
B.) MACROTHROMBOCYTOPENIA • Normal and function properly
I.) TREATMENT
• Mild Bleeding cases- Do not require
treatment
• Severe Bleeding cases- Requires treatment
• Genetic counselling- patients with history;
• May-Hegglin- large, spindle-shaped ease worries and fear of the family
• Sebastian- spherical
• Fechtner- spherical
• Epstein- absent or very faint
C.) THROMBOCYTOPENIA
HEMATOLOGY 2 LAB: CASE PRESENTATION CJLN, BSMT-3C
• mucocutaneous bleeding (gingival) and
spontaneous bruising.
▪ Onset: Starts at birth (purpura and
hematoma)
▪ Severity lessens as the age increases;
much severe at early age
F.) DIAGNOSIS
• GT has general symptoms
• Definite diagnosis: Laboratory tests
▪ PLT count- Normal; the problem is
platelet function (aggregation)
▪ PLT morphology- Normal
▪ Platelet Aggregation Test- (ADP,
CASE NO. 4: GLANZMANN THROMBASTHENIA (GT) Collagen, Epinephrine: ABNORMAL)
QUESTION AND ANSWER ▪ *Ristocetin- NORMAL
A.) ETIOLOGY ▪ Bleeding Time- Prolonged
• Rare blood clotting disorder ▪ Clot Retraction Time- poor;
• Deficiency in GpIIb-IIIa complex; mutation in abnormal aggregation
ITGA2B and ITGB3 ▪ PT and aPTT- Normal; for secondary
▪ Binds to fibrinogen; aggregates hemostasis only = not affected
platelets together
▪ Von Willebrand Factor (vWF)
▪ Fibronectin
▪ Other adhesive ligands
• Abnormal platelet function
B.) INHERITED OR ACQUIRED?
• Inherited (mostly observed): autosomal CASE NO. 5: HERMANSKY-PUDLAK SYNDROME
recessive QUESTION AND ANSWER
• May also be acquired A.) AUTOSOMAL RECESSIVE DISORDER
• ITGA2B - Integrin Subunit Alpha 2b • Both parents are carriers; 25% chance to pass
• ITGB3 - Integrin Subunit Beta 3 the genes to the offspring
C.) PRIMARY HEMOSTASIS B.) ETIOLOGY
• Platelet aggregation • 10 associated genes
▪ Normal: platelet to platelet • Most common and severe form: HPS1 gene
interaction which is mediated by (Type 1 HPS)
ADP, fibrinogen, GpIIb-IIIa • TRIAD: Oculocutaneous Albinism, Dense
▪ Abnormal: problems in the granule deficiency, and Accumulation of
aforementioned factors Ceroid lipofuscin
▪ Glanzmann: leads to excessive • Oculocutaneous Albinism -most common
bleeding; deficiency/lack of GpIIb-IIIa feature of HPS; hypopigmentation in eyes,
complex = NO RECEPTOR, NO skin, and hair
AGGREGATION C.) OTHER ASSOCIATED DISEASE TO HPS
▪ Fibrinogen- target of GpIIb-IIIa • Inflammatory Bowel Disease- Ceroid
complex lipofuscin
D.) 3 SUBTYPES OF GT • LUNGS (other organ affected)- Pulmonary
• Type I: less than 5% of normal/ almost fibrosis
completely absent D.) LYSOSOME-RELATED ORGANELLES (LROs)
• Type II: 5% and 20% of normal/ partial • Share similar characteristics with lysosomes
deficiency • Found in certain cells only: melanocytes
• Type III: normal amount, dysfunctional (melanin), platelets, pneumocytes (lung
receptors (qualitative) cells)
E.) S/Sx:
HEMATOLOGY 2 LAB: CASE PRESENTATION CJLN, BSMT-3C
• Abnormal in melanocytes= • Ecchymosis- bruising
hypopigmentation in HPS • Epistaxis- nose-bleeding
• Abnormal in platelets= platelet aggregation • Gingivitis- recurrent inflammation of the
and prolonged bleeding in HPS gums
• Abnormal in pneumocytes= pulmonary
fibrosis D.) GLYCOPROTEINS ASSOCIATES WITH BSS
E.) IMPORTANCE OF PLATELET DENSE GRANULES in
HPS
• Secretory granules
• ADP
F.) LABORATORY FINDINGS
ELECTRON MICROSCOPE
• Chocolate chip cookie- NORMAL; with dense
granules
• Butter cookie appearance- HPS; without
dense granules
▪ Observed in HPS patients
• GPIB alpha – C#17
• Swiss cheese appearance- due to open
canalicular system; doorway for platelet • GBIB beta – C#22
products’ release • BPIX & GPV – C#3
G.) LABORATORY TESTS E.) GIANT PLATELETS
• Bleeding Time- prolonged • Shapes of platelets
F.) LABORATORY RESULTS
• Low RBC count
• Hemoglobin and hematocrit-
• Low hemoglobin
decreased/slightly; due to epistaxis and
• pT and APTT- normal; HPS is qualitative
bleeding (nose, gingival, purpura, petechiae)
• Platelet count- normal; platelet functions are
• Platelet Count- decreased;
affected
MACROTHROMBOCYTOPENIA
• Bleeding Time- prolonged; No binding sites=
no clot formation
CASE NO. 6: BERNARD-SOULIER SYNDROME • pT and APTT- normal; these are for
Discovered by: Jean Bernard and Jean-Pierre Soulier secondary hemostasis
QUESTION AND ANSWER • Platelet Aggregation Studies- All agonists
A. ETIOLOGY (ADP, Collagen, Arachidonic acid,
• Rare disorder in platelet adhesion Epinephrine) are Normal
• MUTATIONS: • Ristocetin- DECREASED/ ABNORMAL
▪ GP1BA G.) COMPARISON WITH SIMILAR DISORDERS
▪ GP1BB • Glanzmann Thrombasthenia- platelet
▪ GP9 aggregation
• Lack of GPIB-V-IX • Von Willebrand Factor Disease- vWF
▪ Marked bleeding diathesis: impedes • Bernard-Soulier Syndrome- platelet ashesion
the interaction of the complex with H.) SPECIMEN OF CHOICE
vWF • Citrated plasma
▪ Impaired clot formation I.) TREATMENT
▪ Excessive bleeding of a patient • No specific treatment
B.) INHERITANCE PATTERNS • Manageable
• *Autosomal Recessive- more common; ▪ Platelet transfusion (platelet
mutations in both alleles- biallelic BSS-bBSS apheresis)
• FOLLOW RODAKS • Desmopressin (DDVAP)- medications
• Autosomal Dominant- rare; mutations in one • INCREASE THE RISK OF BLEEDING:
allele- monoallelic BSS-mBSS ▪ Anti-histamines
C.) TRIAD OF SYMPTOMS ▪ Non-steroidal drugs
HEMATOLOGY 2 LAB: CASE PRESENTATION CJLN, BSMT-3C
▪ Anti-inflammatory
▪ Certain antibodies
▪ Drugs: Cocaine
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

SECONDARY HEMOSTASIS formation of fibrin strand. Repeat with the

CLOTTING TIME second drop and then the third drop.
• Clotting time is the interval between the moment 6. Stop the timer immediately as soon as fibrin
when bleeding starts at the moment when the strands are seen clinging at the tip of the lancet
fibrin clot threads is first seen. Clotting time on the third drop
depends on the availability of coagulation REFERENCE RANGE: 2-4 minutes
factors. EXAMPLE OF FIBRIN STRAND:
• When the blood vessel ruptures, in a few minutes
blood loses its fluidity and set into a semisolid
mass call clot. This process is called blood
coagulation.
• In clotting time, we check here the presence of
a Visible platelet plug prints, so if the platelet plug
prints is visible, that’s when we report the time on
when the platelet plug prints are seen.
CLOTTING TIME METHODS
MICRO-METHOD
• SLIDE METHOD/DROP METHOD
MATERIALS:
Ø Blood lancet - As seen in the picture above, if the fibrin
Ø Glass slide strand is formed such as that of the image,
Ø Timer/Stopwatch you can now note the time of the formation
Ø Cotton of the strand taking place.
Ø 70% ethyl alcohol B. GLASS CAPILLARY/WRIGHTS METHOD
• GLASS CAPILLARY METHOD/WRIGHT’S METHOD PROCEDURE
MATERIALS: 1. Apply 70% ethyl alcohol to the puncture site to
Ø Capillary tube, non-heparinized disinfect the puncture site using a cotton swab.
Ø Blood lancet Allow it to dry naturally
Ø Cotton 2. Prick the finger with usual aseptic precautions.
• 70% Ethyl Alcohol Immediately start the timer upon seeing the first
MACRO-METHOD drop of blood.
• LEE AND WHITE METHOD 3. Dip one end of the capillary tube into the blood
MATERIALS: drop gently without pressure.
Ø Glass test tubes, 3 pieces (13x100nm) 4. Allow to fill the capillary with blood by lowering
Ø Syringe (G21/23 needle) the end of fitted capillary
Ø Water Bath 37 degrees Celsius • Do not stuck the blood around % of its length
• Timer/ Stopwatch undipped
5. After about 2 minutes, start snapping off small
PROCEDURES lengths of the tube, at interval of 15 seconds,
MICRO-METHOD each time noting whether the fibrin thread is
A. SLIDE/DROP METHOD PROCEDURE formed between the snapped ends
• Putulin yung capillary tube into smaller pieces
1. Disinfect site of puncture with 70% ethyl alcohol.
and check if there is a formation of fibrin
Air dry.
strand/thread
2. Puncture to a depth of 3mm using a blood
6. Repeat breaking at regular time intervals, until
lancet
fibrin thread appears at the broken end of
3. Start timer as soon as the first drop of blood
capillary tube. Do not pull away the catted
disappears.
pieces
4. Transfer the three drops of blood onto a clean
record the time interval between pricking finer and
glass slide. Careful not to touch the skin.
first appearance of fibrin thread at the broken ends
5. Pass the tip of the lancet through the first drop of
of capillary tube. That is clotting time of blood
blood every 30 seconds and note for the
REFERENCE RANGE: 4-9 minutes

T R AN S F O R M E R S 1
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

EXAMPLE OF FIBRIN STRAND: 8. The sample in tube 1 is expected to clot first

since it contains the least amount of tissue fluid
among all three tubes
• Kasi eto yung unang sample na na-aspirate
Record the coagulation time as the time that
lapsed from the withdrawal of blood up to the
clotting of blood in tube 3.
REFERENCE RANGE: 7-15 minutes
PLASMA RECALCIFICATION TIME
• A modification of Lee and White Method is
recalcification time of plasma
• More sensitive method than the coagulation
• here is an example of how to snap the
time of whole blood
capillary tube.
• May reveal abnormality which is NOT
• If fibrin thread is already present sa
detectable by the determination of the clotting
pinagputulan na capillary tube, i-nonote na
time of the venous blood
‘yung time when the fibrin thread appeared.
• The activated recalcification time makes use of
This will be interpreted as the clotting time from the
0.25M CaCl2 as activator
very beginning of the procedure (finger prick) and
SPECIMEN:
up to the formation of the thread between the ends
• Citrated Plasma
of the snapped capillary tube
• Platelet-Rich Plasma/Platelet Poor Plasma or
MACRO-METHOD both
C. MODIFIED LEE AND WHITE CLOTTING TIME REFERENCE RANGE:
PROCEDURE • Less than 50 seconds
PRINCIPLE: the coagulation of blood is the length of
time required for a measured amount of blood to ACTIVATED COAGULATION TIME OF WHOLE BLOOD
clot under certain conditions. The test is based on
PRINCIPLE:
the fact that when venous blood is put into a glass
• Whole blood contains all the components
tube (foreign surface), it will form a solid clot. The
necessary to produce a clot when removed
time required for this response is a measure of overall
from the veins and put into a glass tube. By
intrinsic and common pathway coagulation
adding an activator and keeping the blood
1. Label 3 uniformly sized test tubes as tubes 1, 2,
at a constant 37 deg. cel.
and 3
A more reliable and rapid screen of the intrinsic and
2. Draw 4 mL of blood via venipuncture
common pathway is achieved
3. Note the time at which blood enters the syringe
REAGENTS:
4. Carefully dispense 1 mL of blood to tube 3, then
• Two evacuated tubes containing 12 mg of
1 mL to tube 2, and another 1 mL to tube 1
diatomite are needed
• Sequence: tube 3 à 2 à 1
Diatomite – an activator present or needed in
• Pabaliktad yung sequence kasi mas nauuna
activated coagulation time of whole blood
yung last tube na ma-fill
5. Incubate all tubes in a water bath that is EQUIPMENT/MATERIALS:
maintained at 37 degrees Celsius • A portable heat block
6. By doing a tilt tube method, check for clot • Thermometer
formation on tube 1. Repeat every 30 seconds • Two stopwatch
until the blood inside the tube does not spill out Why two? Because evacuated tube that are to be
or drip incubated are TWO.
7. After the blood in tube 1 has clotted, proceed REFERENCE RANGE:
to checking clot formation in tube 2 and repeat • 75 – 120 seconds
above procedure with tube 3. NOTE: Target range during heparin therapy is 140 –
• Sequence of checking: tube 1 à 2 à 3 185 seconds (the time is changed when the patient
is in heparin therapy)

T R AN S F O R M E R S 2
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

PARTIAL THROMBOPLASTIN TIME ADVANTAGES:

PRINCIPLE: • It is more sensitive with Stanley Brown
• The APTT measure all factors except VII and Method
XIII. Maximum activation of the contact • Fresh specimens are not necessary, and the
factors is accomplished by addition of the method can be used for mailed samples for
activators. Phospholipid is supplied to blood
substitute for platelet factor 3 (PF3). From this The method is not affected by heparin
point, the APTT is essentially the same as a C. OWREN’S THROMBOTEST METHOD
recalcification time. • For the control of coumarin anticoagulant
SPECIMEN REQUIREMENTS: therapy, this is considered as the most sensitive
• Citrated Platelet-poor plasma test.
REAGENTS: • Ginagamit po siya for the control of coumarin
• Phospholipid with activators (APTT reagent) anticoagulant therapy
and 0.025 M CaCl2 (or as recommended by D. FIBROMETER METHOD
reagent manufacturer) are used. • It is an electromechanical semiautomated
EQUIPMENT/MATERIALS: instrument that has been used extensively in
• 12x75 mm glass tubes one-stage prothrombin method of Quick.
• Heat block (Incubator) • Fibrometer ginamait po siya sa one-stage
• Pipets prothrombin method of Quick.
REFERENCE RANGE: E. MICROMETHOD (PROTIME)
• 20-45 seconds
• This is microtechnique employed for children
PLASMA PROTHROMBIN TIME and the method uses micropipettes; the
• Measures the EXTRINSIC and COMMON principle of the test is similar with one-stage
pathway of coagulation prothrombin time or Stypven Time
o EXTRINSIC Factor: III and VII • Gumagamit ng beaker
o COMMON: I, II, V, X, AND XIII STYPVEN TIME (RUSSEL VIPER VENOM TIME)
• It is used to monitor oral anticoagulant therapy
• This venom is capable of bypassing the factor VII
(warfarin, Coumadin, Heparin, Coumarin)
and directly activating factor X to Xa
• Can detect prothrombin, fibrinogen, Factor V,
§ In this Stypven time, binabypass niya po si
VII, and X deficiencies
factor VII inactivate na niya agad si factor X
SPECIMEN:
kase ine-extrinsic (?) si factor VII will activate
• Sodium Citrate Tube (Light Blue top) factor IX and X, sa RUSSEL VIPER VENOM,
METHODS OF PROTIME binabypass niya na si factor VII ina
A. ONE-STAGE METHOD OF QUICK (STANLEY automatically activating the factor X.
BROWN METHOD) • It is used to distinguish deficiencies
PRINCIPLE: of Factor X and those of Factor VII. It is also used
Tissue thromboplastin and calcium added to to detect deficiencies in prothrombin,
plasma react to fibrinogen to form a clot. The fibrinogen, and Factors V and X. It differs from
thromboplastin added to the plasma takes the prothrombin time in the in those deficiencies in
place of the juice formation of extrinsic Factor VII are not detected
thromboplastin. The prothrombin time is therefore § Ang pinagkaiba nila ni prothrombin time
prolonged if there is a deficiency of factors V, VII, or yung deficiency ni Factor VII ay hindi niya na
X or a very severe deficiency of Factor I and II dedetect kase it is capable of bypassing the
NORMAL VALUE: factor VII.
• 10 to 15 seconds • Uses snake venom (Vipera russelli)
Depends on the reagent and machine. • Common name: East Indian Viper
B. TWO-STAGE PROTHROMBIN AND REAGENT: platelin + chloride
PROCONVERTIN TEST (OWREN AND AAS) SPECIMEN: Platelet Poor Plasma (PPP)
• It offers combined estimation of the levels of Determines problem in common pathway
prothrombin and Proconvertin NORMAL VALUE: 6-7 seconds

T R AN S F O R M E R S 3
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

PROTHROMBIN ACTIVITY OR INDEX

• Reported in percentage (%), with 100% as the
maximum level
!"#$%&' (%* +',#*-+ #. ,#*/"#0)
• PT Activity (%) = × 100
!"#$%&' (%* +',#*-+ #. 23/%'*/)
INTERNATIONAL NORMALIZED RATIO
• Method of reporting proposed to monitor
patients in oral anticoagulant therapy
• The prothrombin time ratio had the test been
performed using international standard
thromboplastin reagent
• Results are reported as International Normalized
Ratio (INR)
o INR - Patient PT x 100 Control PT
• The closer the ISI is to 1, the more sentistive the
reagent is
• The higher the ISI, the less sensitive the reagent is • In substitution studies:
• NORMAL INR = 1.1 1. First determine if the given is extrinsic,
intrinsic, or common coagulation pathway.
INR CONDITIONS
2. Check which tests are abnormal (PT, APTT,
2.0 - 2.5 DVT, High risk surgery
TT)
2.0 - 3.0 Hip surgery, Femur fracture
With this, we would know what factor is defecient
2.0 - 3.0 DVT, Pulmonary embolism,
SERUM PROTHROMBIN TIME/PROTHROMBIN
Transient ischemic attack
CONSUMPTION TEST (PCT)
2.5 - 3.5 Mechanical/ prosthetic heart
• Best considered as a test of platelet
valves
phospholipid activity.
2.0 - 4.5 Recurrent DVT and pulmonary
• If the prothrombin time and the PTT are normal,
embolism, Myocardial infarction,
a short PCT will indicate deficiency of PF3 due to
Arterial disease
thrombocytopenia or thrombopathia
• This ISI is a calibration parameter that defines the
• PF3 – responsible in interaction with platelet
responsiveness of the reagent relative too a
phospholipid surfaces
World Health Organization (WHO) International
THROMBOPLASTIN GENERATION TEST
Reference Preparation, which by definitiion has
• Bigg’s and Macfarlane Method
an ISI of 1.0
• Hick’s-Pitney Kaolin Modification model
SUBSTITUTION TEST/MIXING STUDIES
• used if the PT or APTT is high
LABORATORY EVALUATION FOR THROMBIN
• adopted if primarily tests like PT or APTT are
abnormally prolonged and indicate a factor THROMBIN TIME
deficiency. The patient’s deficient plasma is • Test for the deficiency or inhibition of fibrinogen.
diluted 1:1 with a plasma or serum substitute and • Sa thrombin time, magaactivate ng fibrinogen.
the APTT or PT is repeated. Kapag naginhibit si fibrinogen order deficiency
• A correction if the original prolonged APTT or PT of fibrinogen in coagulation system, we can
indicates that the deficient factor has been check it through thrombin time.
added to patient’s plasma by substitution PRINCIPLE:
solutions as follows: Commercially prepared thrombin reagent is added
to citrated plasma, and the time required for clot
formation is measured.
NORMAL VALUE: 10-20 seconds
FIBRINDEX TEST
• Commercially available test wherein upon
addition of plasma containing fibrinogen,
thrombin produces clotting.

T R AN S F O R M E R S 4
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

PROCEDURE: LABORATORY DEMONSTRATION

0.5 CC Fibrindex + 0.5 CC plasma in a test tube, SECONDARY HEMOSTASIS
normally clot will be formed within 5-10 seconds • Normal coagulation is in order to stop bleeding
and to repair the vessel walls in the presence of
Addition of 0.5 CC fibrindex and 0.5 CC plasma then injury
observe mo if there is a formation of blood within the COAGULATION TIME
range of 5 to 10 seconds. • Coagulation is an important part of hemostasis
NORMAL CLOT TIME: 5-10 seconds wherein a damaged blood vessel wall is
FIT-TEST (IMMUNOLOGICAL TEST) covered by a fibrin clot to stop hemorrhage and
• Rapid slide test based on the agglutination of aid in the repair of the damaged vessel.
fibrinogen-coated red blood cells by the latex • This is why disorders in coagulation can lead to
anti-human fibrinogen reagent. Normally, either increased hemorrhage or thrombosis.
presence of fibrinogen is indicated by o Hence, various tests were developed
agglutination. to assess the body's coagulation
o In fit-test, ang tinitignan natin ditto ay mechanism and one of those is the
agglutination. So kapag nagkaroon ng coagulation time.
agglutination, there is a presence of MICROMETHOD/ MACROMETHOD/
fibrinogen. SLIDE METHOD TUBE METHOD
• Rapid test for fibrinogen determination.
FIBRINOGEN TITER METHOD
• Serial dilutions of plasma are diluted with
thrombin. The titer is the highest dilution in which
a fibrin clot can be seen, and is related to the
fibrinogen concentration and indirectly to the • Capillary
presence of fibrinogen is indicated by blood collection
agglutination • Venous blood
• Same as the fit-test. Presence of agglutination collection
ang ating hinahanap. So, gumagamit tayo ng
• Coagulation time may be determined via
serial dilution to determine the presence of
micromethod or the slide method and the
agglutination.
macromethod.
ASSAY OF PLASMA FIBRINOGEN
• Several accurate methods are now available for
MICROMETHOD/SLIDE METHOD
the quantitative assay of plasma fibrinogen.
MATERIALS
Fibrinogen is usually converted into fibrin which is
quantified by gravimetric, nephelometric, • Lancet
chemical, immunologic and precipitation
methods.
METHODS:
a. Ellis and Stransky Method
b. Stirland’s Method
c. Turbidimetric Method of Partfantjevet. Al
d. Ratnoff and Menzie Method
e. Fibrin Clot Method • Glass slide

§ these are the methods to quantify plasma

fibrinogen.

• Applicator stick
• Stopwatch

T R AN S F O R M E R S 5
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

PROCEDURE
1. A skin puncture is first performed using a
lancet.

5. When a thread-like fibrin clot is observed,

stop the timer then record the coagulation
time.

2. Once blood starts to ooze out, the timer

should then be started.

3. Three (3) drops of blood from the puncture

will then be placed in a single glass slide and
then just wait for clot formation to occur.

4. You may check for clot formation by

drawing the blood with the lancet every 30
seconds.

• NORMAL COAGULATION TIME: 2 to 4 minutes

T R AN S F O R M E R S 6
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

MACROMETHOD/TUBE METHOD
MATERIALS
• Venipuncture set

• Three test tubes 3. After collecting 4 mL of blood, transfer 1 mL

into tube 3, another 1 mL to tube 2, and
lastly, tube 1.

• Water bath

4. Incubate these tubes in the water bath

maintained at body temperature.

• Stopwatch
PROCEDURE
1. The tubes should first be labeled as tubes 1,
2, and 3.

5. Check for clot formation on tube 1 by tilting

the tube every 30 seconds. When clot has
formed in this tube, you may then check clot
formation in tube 2 followed by tube 3.
2. Proceed to the collection of 4 mL of venous
blood. Start the timer once blood enters the
syringe.

T R AN S F O R M E R S 7
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

MATERIALS

6. The sample in tube 1, which contains the

LAST portion of the collected blood is
expected to clot first since it contains the
least amount of tissue fluid among all three PRIMARY REAGENTS
tubes.

o Tissue Thromboplastin and Calcium

chloride solution
o Control plasma
PROCEDURE

7. Once clot is observed in tube 3, stop the

timer and record the coagulation time.

• NORMAL COAGULATION TIME: 7 to 15 minutes

• COAGULATION FACTOR DEFICIENCIES:
● To determine the Prothrombin time, we need to
prolonged
saturated plasma which may be obtained
• The specific coagulation factor deficient cannot through the centrifugation of saturated blood
be determined using this test. for 5 minutes.

PROTHROMBIN TIME (PT)

• This is performed to detect coagulation factor
deficiencies associated with the extrinsic and
common pathway, and to monitor treatment
with coumadin or warfarin
• Evaluates ability to clot
• In most cases the PT is usually reported together
with the International normalized ratio for the
standardization of the results

T R AN S F O R M E R S 8
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

● Proceed to add 0.2 mL of the reagent in both

tubes

● The plasma should be immediately separated

from the red cells by transferring it to a separate
test tube.

● When the 3 minutes of incubation has elapsed,

transfer 0.1 mL of the patient’s plasma to the
tube labeled as T and 0.1 mL of the control
plasma into the tube labeled as C, then start the
stopwatch.
● This is then to be incubated together with control
plasma in the water bath for 2-3 minutes at 37°C
degrees celsius.

● While waiting for the incubation step, you may • Mix both tubes and tilt them back and forth until
prepare 2 separate test tubes, one labeled as T clot formation is observed. Once this happens,
for test and the other labeled as C for control stop the timer and record the time for both
tubes

T R AN S F O R M E R S 9
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

RESULT: PROCEDURE
• The calcium chloride which will replace the
calcium removed via chelation, is first
incubated in the water bath for 5 minutes. This is
performed to pre-warm the said reagent.

● Result: both have clot formation

• The result of the plasma should be within the
• Next, to obtain the needle sample which is
normal limit ranging from 10 to 13 seconds for it
citrated plasma, the same centrifugation step is
to be considered valid. The said normal values
performed for APTT.
may vary depending on the reagent used by
the laboratory
INTERNATIONAL NORMALIZED RATIO (INR)
• Ensures that results from the PT test are the same
form one lab to another

ACTIVATED PARTIAL THROMBOPLASTIN TIME (APTT)

• This test can detect coagulation factor
• The separated plasma is then transferred to the
deficiencies associated with the intrinsic and
tube labeled T for test. And the control plasma
common pathway, as well as the presence of
to the tube labeled C for control.
circulating anticoagulants or inhibitors in the
blood.
MATERIALS
To perform this test, we will need a:

• 0.2 mL of the APTT reagent will then be added to

both of these tubes, which are to be mix
thoroughly.

• Venipuncture set – to collect citrated blood.

• Test tubes
• Water bath – for incubation
• Stopwatch
• Pipettes
• APTT reagent + Calcium chloride – as the
reagents • These tubes will then be incubated to water
• Control plasma bath for 5 minutes at 37 degree C.

T R AN S F O R M E R S 10
2 Laboratory Evaluation of Secondary Hemostasis Hematology 2 (LAB)

RESULTS:

• After 5 minutes, 0.2 mL of the pre-warmed Control: (+) clot formation

calcium chloride will then be added to both
tubes.

• Patient’s plasma/test: (+) clot formation

• NORMAL CLOT FORMATION: LESS THAN 35
SECONDS
• This test together with the prothrombin time, are
• The timer will be started immediately after. performed to detect any abnormality in the
• Both tubes which are still in the water bath, coagulation mechanism of the body.
should be tilted every after 5 seconds. And • Any problems in the said coagulation
should be removed after 20 seconds. Continue mechanism will result to prolonged results.
tilting both tubes.

• Stop the timer when clot formation is observed.

Which normally occurs in less than 35 seconds.

T R AN S F O R M E R S 11
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Hematology 2
2nd Semester, AY 2021-2022

GROUP 5
CASE STUDY
Hemophilia A

Group Members:
Dominguez, Noriz Ember A.
Gloria, Janelle Keith C.
Guinto, Angelica
Gumalot, Joshua C.
Malonzo, Hannah Sofia D.
Songco, Mharjonie Martin C.
Valerio, Sheila Emmanuelle T.
Villanueva, Victor Miguel

Date Submitted:

March 16, 2022

BSMT3-A
Case 1:
Sarah Crewe is a 7-year-old young girl who works as a maid in Miss Minchin’s orphanage due
to her debts and unpaid tuition fees. One time, she was tasked to peel potatoes and accidentally
cut her thumb and index finger. Few days after the incident, her cuts did not stop bleeding and
were still swollen. Due to this, Sarah’s friends, Becky and Lottie, decided to bring her to the
hospital.

On physical examination, the physician also found bruises on Sarah’s legs and arms. Sarah
mentioned that those bruises showed up after falling off a hill when she was fetching water. It
was also noted that Sarah’s father died due to a “bleeding disorder”. She was referred to the
outpatient laboratory for the following tests: BT and coagulation profile:
LABORATORY DATA RESULT REFERENCE RANGE

Bleeding Time 5 minutes 3–8 minutes

Prothrombin Time 14.5 secs 12–16 seconds

Activated Partial Thromboplastin 70.5 secs 20-38 seconds

Time

Thrombin Time 14 secs 12-18 seconds

Factor VIII activity 4.0% 100–300%

Factor IX activity 30.0% 11–55%

vwF antigen concentration 127% 60% to 153%

vwF activity 127.5% 43% to 138%

GUIDE QUESTIONS:

1. What is the most probable diagnosis of the patient? Explain why you answered this
disorder.
Based on the given clinical manifestations and laboratory results, the most probable
diagnosis of the patient is Hemophilia A, also known as the “Classic Hemophilia”.
 Hemophilia A is a genetic bleeding disorder that is caused by a deficiency in the coagulation
factor VIII. Factor VIII plays a vital role in the intrinsic pathway, it serves as a cofactor to
factor IX that helps in the activation of Factor X. The factor VIII/IX complex with the help of
thrombin greatly amplifies the activation of factor X at a rate of 10,000 times. Factor X
initiates the common pathway ultimately leading to the formation of blood clots. Overall the
deficiency of Factor VIII greatly delays the coagulation cascade that leads to the delay in the
formation of blood clots in response to injury such as experienced by the patient. Hemophilia
is a recessive sex-linked condition. The X chromosome contains the defective gene that
causes hemophilia. In general, it primarily affects males but in this case, Sarah who is a
female patient exhibited symptoms of the disease which is a rare case. In relation to this, it
is important to take note that her father died due to a bleeding disorder which is most
probably hemophilia A as well. In the inheritance pattern of hemophilia, 100% of females
born from a father that has hemophilia will be a carrier of the hemophilia gene. This is
because females have 2 X chromosomes, the disease is not caused by the presence of the
hemophilia gene on one of them because a functional gene that codes for factor production
compensates for the abnormal gene, making them a carrier. Females who have the gene
can pass it to their son who will have hemophilia, even if the mother is not affected. Female
carriers also may experience bleeding symptoms in some cases. According to the CDC, a
female who is a carrier sometimes can have symptoms of hemophilia. Another possibility on
how hemophilia can affect females is through imbalanced lyonization of the normal X
chromosome. Lyonization is also known as the inactivation of the X chromosome. To put it
simply the chromosome that does not have the hemophilia gene is inactivated or turned off,
therefore the chromosome that presents the hemophilia gene will be expressed. Moreover
imbalanced lyonization of the normal chromosome X explains why clotting factors vary from
one carrier to another.

2. Is the disorder described considered to be a mild, moderate or severe form of the

condition? Differentiate these 3 forms.
The laboratory tests are primarily utilized to diagnose Hemophilia A, this comprises the
platelet count, prothrombin time (PT), which are substantially regular in both findings, and
activated partial thromboplastin time (aPTT), which shows abnormal, indicating a prolonged
result. The prothrombin time (PT) exhibits usual values as the test evaluates the extrinsic
pathways that involve coagulation factors VII, X, V, II (prothrombin), and I (fibrinogen).
Whereas activated PTT assesses the intrinsic pathways, which involves the factors XII, XI,
IX, VIII, X, V, II, and I. The involvement of Factor VIII in the intrinsic pathway causes the
abnormal result. In connection to our case study, hemophilia A is due to the deficiency of the
coagulation factor VIII, and considering that the factor VIII is deficient in the disorder thus
other factors will have ordinary levels, hence the prothrombin time (PT) reveals a normal
while activated partial thromboplastin time (aPTT) demonstrated a prolonged result due to
the involvement of factor VIII under the intrinsic pathway. The severity of hemophilia A will
be determined by the intensity of the fundamental mutation that governs the factor's activity.

In accordance with the National Hemophilia Foundation, hemophilia A scopes from mild,
moderate, and severe, signs and symptoms also play a vital role in identifying the severity of
the disorder. In mild hemophilia A, factor VIII values range from 6% to 49%, accounting for
approximately 25% of hemophilia A cases. Mild hemophilia A patients primarily experience
bleeding after serious trauma/injury/surgery, and they rarely or never experience bleeding
complications. Furthermore, women with mild hemophilia A are more likely to have
excessive menstrual bleeding and significant bleeding after childbirth. In moderate
hemophilia A, factor VIII values range from 1% to 5%, which accounts for approximately
15% of cases wherein patients with this severity level experience prolonged bleeding that
tend to accompany trauma in minor surgery or injury and dental extraction, as well as rare
spontaneous bleeding episodes. Finally, for severe hemophilia A factor VIII ranges less
than 1% which accounts for roughly 60% of all hemophilia A cases. People with severe
hemophilia A experience frequent bleeding that may occur once or twice a week. Bleeding
may occur spontaneously in their muscles and joints. Prolonged bleeding could perhaps
happen as a consequence of trauma, injury, or surgery, but it can also occur for no apparent
cause.

In this case, we tend to diagnose Moderate Hemophilia A due to the non-stop bleeding
experience after the cut, bruises in her legs and arms, and to its family history wherein the
patient’s father died due to a “bleeding disorder”. Furthermore, the laboratory test
strengthened our diagnosis wherein there was an abnormality between her (aPTT) Activated
Partial Thromboplastin Time that shows (4.0%) which falls under moderate hemophilia A
wherein factor VIII levels ranges from 1% to 5%.
3. Is the disorder inherited or acquired? Explain briefly.

The disorder is inherited. Hemophilia A is inherited in an X-linked recessive pattern. This

means that the X chromosome, which is one of the two sex chromosomes contains the
genes linked to the disease, wherein one mutated copy of the gene in each cell is enough to
cause the disease in males (who have only one X chromosome). On the other hand, to
develop the condition in females (who have two X chromosomes), a mutation would have to
occur in both copies of the gene. In this inheritance pattern, females with hemophilia are
rare because it is unusual that they will have two mutated copies of this gene. The expected
outcome is 25% if the mother is a carrier and the father does not have the disease. Fathers
cannot pass on X-linked features to their sons, but his only X chromosome can be passed
down to all of his daughters, making them a carrier.

In this case, the female can develop hemophilia if she inherited the hemophilia alleles
from both parents, or if she inherits one hemophilia allele from her father, which means that
she has a 50% chance of being heterozygous. Additionally, female carriers have roughly half
the normal level of coagulation Factor VIII or IX, which is usually sufficient for normal blood
clotting. However, approximately 10% of carrier females have less than half of the normal
level of one of these coagulation factors such as in this case, putting them at risk for
irregular bleeding, especially after an injury.

CDC. (2021, June 29). How hemophilia is inherited. Centers for Disease Control and Prevention.
https://www.cdc.gov/ncbddd/hemophilia/inheritance-pattern.html.
4. What is the choice of treatment with this disorder?
Replacement therapy is the treatment of choice with hemophilia. For patients with
hemophilia A, concentrates of coagulation Factor VIII are slowly administered either through
injecting or infusing it into the patient's vein to help replace the amount of clotting factor that
is lost or lacking. There are two major types of clotting factor products that may be used for
replacement therapy of patients with hemophilia; one that is derived from human plasma
and the other one which is genetically engineered and is approved by the US Food and
Drug Administration (FDA). There are various clotting factor concentrates that are derived
from human plasma proteins. Although the risk of acquiring a disease from human
plasma-derived clotting factors is very little, blood products undergo different processes
including separation, testing for any potential virus, and treatment to prevent the spread of
bloodborne viruses. On the other hand, recombinant clotting factor concentrates may also
be used to further lessen the patient's risk of getting a disease since these factor
concentrates are genetically engineered and do not have plasma or albumin (recombinant
factors VIII and IX only). Patients with hemophilia may have replacement therapy either as
prophylactic or on-demand therapy, depending on the severity of the disease. On-demand
replacement therapy is usually performed on patients with mild or moderate hemophilia A,
wherein the patient will only have the said treatment as needed to stop the bleeding when it
happens. Prophylactic replacement therapy, however, is performed on a regular basis to
prevent bleeding and other complications. As demand therapy is less expensive and
intensive compared to prophylactic therapy, there is a possibility that bleeding can result in
serious problems (e.g. joint damage) before the patient could receive such treatment,
especially to patients who have severe hemophilia A. Home replacement therapy may be
given to these patients since the infusion of factor concentrates works effectively within one
hour of bleeding. When bleeding occurs, immediate treatment should be performed to
reduce the risks of organ damage or other serious complications. Also, family members of
children suffering from hemophilia must also learn to watch for signs of bleeding as children
sometimes disregard these signs to avoid discomfort when receiving treatment.

If replacement therapy is unavailable, other alternatives such as antifibrinolytic

medications, desmopressin, and/or bleeding site treatment may be applied to prevent
temporary clots from deteriorating. Antifibrinolytics can aid primarily in the mild severity form
of the disorder by slowing down the clot factor’s breakdown which controls bleeding of the
 bruise or injury. Examples of antifibrinolytics are tranexamic acid (TXA) and
epsilon-aminocaproic acid (EACA), which are usually taken orally in the form of a tablet or
liquid. The dosage of it is dependent on the medical condition to be prescribed by the doctor.
Moreover, desmopressin acetate (DDAVP®) is generally known as man-made vasopressin,
a medication done intravenously (children ≥ 12 yrs. of age and adults), orally, or via nasal
spray (children ≥ 4 yrs old and geriatric), is directed towards the control of urine amounts
produced by the kidney in cases of diabetes insipidus, but it can also aid in blood by
increasing the Factor VIII (anti-hemophilic factor) levels, which is responsible for
blood-clotting as encoded by the F8 gene in the intrinsic pathway. DDVAP® however may
present side effects (i.e, nausea, headache, and vomiting), especially in children with
increased sensitivity. Another brand of desmopressin is Stimate®, a nasal formulation spray
that is more specific to treat bleeding complications. Additionally, bleeding site treatment
through the use of pain medications, and undergoing physical therapy may alleviate the pain
and potential swelling of the injury. If the administered treatment fails to increase the activity
of Factor VIII in the body, it would suggest that the patient may have elevated levels of
Factor VIII inhibitors, wherein further testing such as Factor VIII assay and mixing studies is
highly advised in order to introduce new treatment.

References:
● CDC. (2021, June 29). How hemophilia is inherited. Centers for Disease Control and
Prevention. https://www.cdc.gov/ncbddd/hemophilia/inheritance-pattern.html.
● Hemophilia Inheritance Patterns. (n.d.). Cdc.Gov. Retrieved March 15, 2022, from
https://www.cdc.gov/ncbddd/hemophilia/course/Hemophilia_Patterns_v3.pdf.
● S;, N. (n.d.). Multifunctional roles of thrombin. Annals of clinical and laboratory science.
Retrieved March 15, 2022, from https://pubmed.ncbi.nlm.nih.gov/10528826/
● Inheritance pattern. Hemophilia Federation of America. (2021, January 4). Retrieved
March 15, 2022, from
https://www.hemophiliafed.org/home/understanding-bleeding-disorders/what-is-hemophil
ia/hemophilia-a/inheritance-pattern/
● TBH Creative, info@tbhcreative.com. (n.d.). Women with hemophilia. Indiana
Hemophilia & Thrombosis Center. Retrieved March 15, 2022, from
https://www.ihtc.org/women-with-hemophilia
● Factor VIII. Factor VIII - an overview | ScienceDirect Topics. (n.d.). Retrieved March 15,
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-viii
● Chaudhry, R. (2021, September 1). Physiology, coagulation pathways. StatPearls
[Internet]. Retrieved March 15, 2022, from
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● Douglass A Drelich, M. D. (2021, November 16). Hemophilia A (factor VIII deficiency).
Practice Essentials, Background, Pathophysiology. Retrieved March 15, 2022, from
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jXE6DX4QIC2fZvHQJX7jtl6vAoFG5KLjdDsl-Q_4
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hemophilia%20A%3A%206%25%20up,extraction%20results%20in%20prolonged%20bl
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● MedlinePlus. (2022, February 18). Sex-linked recessive.
https://medlineplus.gov/ency/article/002051.htm
● U.S. National Library of Medicine. (2020, August 18). Hemophilia: Medlineplus genetics.
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https://medlineplus.gov/genetics/condition/hemophilia/#inheritance
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https://www.hoacny.com/patient-resources/blood-disorders/what-hemophilia/how-hemop
hilia-treated
● CDC. Treatment of Hemophilia | CDC [Internet]. Centers for Disease Control and
Prevention. 2020 [cited 2022 Mar 15]. Available from:
https://www.cdc.gov/ncbddd/hemophilia/treatment.html
● Rush University. (2022). Hemophilia in Children - Medical Conditions | Rush System.
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https://www.rush.edu/kids/conditions/hemophilia-children
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Mayo Clinic.
https://www.mayoclinic.org/drugs-supplements/desmopressin-nasal-route/before-using/d
rg-20066827?p=1
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Hematology 2
2nd Semester, AY 2021-2022

HEMATOLOGY 2

CASE PRESENTATION

CASE NO. 2: VON WILLEBRAND DISEASE

GROUP 1 - BSMT3C

DUCUT, LYRON MARK D.

PORNOBI, AGAPE JOY

SIBLAWON, ANGELA

SORIANO, ERIN FRANCES

REYES, EUNICE RUSSELLE

TIGLAO, JANELLE

LEAÑO, REYCHEL

IGNACIO, ROMAE JONES

 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

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2nd Semester, AY 2021-2022

CASE 2:
A 20-year-old male patient was admitted to the hospital after experiencing excessive bleeding
after a tooth extraction. He also presented with dizziness and fatigue upon admission. His
history revealed previous events of recurrent bruising, epistaxis, and anemia. He also had a
family history of mild bleeding problems. Lab examinations were done, and the following results
were obtained. Diagnosis was verified by requesting for factor assays and multimer analysis.

TESTS RESULTS

Hemoglobin 14.0 g/dL (normal)

Hematocrit 41% (normal)

Platelet Count 285 x 103 /μL (normal)

PT 13 seconds (normal)

APTT 51 seconds (prolonged)

Bleeding Time 16 minutes (prolonged)

RIPA Decreased

GUIDE QUESTIONS:
1. What is the most probable diagnosis? Explain the findings which led to this
diagnosis.

Von Willebrand Disease (vWD) is the most probable diagnosis of the patient. vWD is
caused by any one of dozens of germline mutations that result in quantitative or structural
abnormalities of Von Willebrand Factor (vWF). Both quantitative and structural abnormalities
lead to decreased adhesion by platelets to injured vessel walls, causing impaired primary
hemostasis which in this case results in a prolonged or excessive bleeding of the patient.
This condition is characterized by abnormal platelet function expressed in prolonged
bleeding time associated with surgical or oral procedure and may be accompanied by a
decreased factor VIII procoagulant activity. Structural (qualitative) or quantitative vWF
 Angeles University Foundation
College of Allied Medical Professions
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Hematology 2
2nd Semester, AY 2021-2022

abnormalities reduce platelet adhesion, which leads to mucocutaneous hemorrhage of

varying severity such as epistaxis and excessive bleeding that was aforementioned after a
tooth extraction.
Symptoms vary over time because vWF production and release are susceptible to a
variety of physiologic influences. Severe quantitative vWF deficiency creates in addition
factor VIII deficiency owing to the inability to protect nonbound factor VIII from proteolysis.
When factor VIII levels decrease to less than 30% of normal, anatomic soft tissue bleeding
accompanies the typical mucocutaneous bleeding pattern of vWD resulting in the laboratory
test to have prolonged bleeding time and activated prothrombin time (APTT). In addition, the
primary function of vWF is to mediate platelet adhesion to subendothelial collagen in areas
such as capillaries and arterioles. Since vWF is deficient, platelet adhesion cannot normally
occur which also results in prolonged bleeding time especially in the mucocutaneous area.
The patient’s clinical signs and symptoms include excessive bleeding, dizziness,
fatigue, recurrent bruising, epistaxis, and anemia are common signs and symptoms of vWD.
It is because of quantitative or structural abnormalities that leads to insufficient level of
circulating vWF, factor VIII and a low level of ristocetin cofactor activity. The laboratory
findings of patients shows a normal hemoglobin, hematocrit, platelet count and prothrombin
time (PT), a prolonged bleeding time and activated prothrombin time (APTT), and a low level
of ristocetin which is compared to the vWF laboratory findings that confirms the diagnosis.

2. What is the main hemostatic abnormality in this disease? Explain how the
abnormality affects normal hemostasis.

The platelet adhesion in primary hemostasis is disrupted in Von Willebrand Disease

(vWD) due to a faulty Von Willebrand Factor (vWF) interaction between platelets and the
vessel wall. In normal hemostasis, vasoconstriction is the initial response when a vessel is
injured, it is predominantly mediated by endothelin-1, which is generated by damaged
endothelium and constricts the damaged arterial, reducing blood flow in the site.
Subendothelial collagen, vWF, ATP, and inflammatory mediators are exposed when the
endothelium is damaged. Platelets will adhere to the injured area through the Von
Willebrand factor, which attaches to the endothelium and the GPIB receptor on platelets.
 Angeles University Foundation
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Hematology 2
2nd Semester, AY 2021-2022

The platelet receptor GPIB will bind to the von Willebrand factor present in the blood
vessels.
After the platelets have adhered to the injured vessel wall, platelet activation will
follow. Thrombin will trigger the production of Cytoplasmic granules (alpha and dense
granules) such as Serotonin, platelet activating factor and adenosine diphosphate (ADP).
These granules aid or assist in the recruitment of other platelets in order for aggregation to
take place. ADP is stored in the dense granules of platelets and is responsible for the
conformational change of another platelet receptor (GPIIb complex). When ADP is released,
it interacts with platelet membrane P2Y1 and P2Y12 receptors. P2Y1 creates a change in
the morphology of the pseudopod and facilitates platelet aggregation. P2Y12 is a key factor
in the clotting cascade's activation. When ADP attaches to its receptors, it helps to reveal
the GPIIb-IIIa complex's binding site, allowing it to be activated.
After platelets have been activated, platelet aggregation begins. The Gp IIb/IIIa
receptors bind to vWF and fibrinogen once activated. Fibrinogen is located in the circulation
and binds platelets' Gp IIb/IIIa receptors, allowing them to communicate with one another.
This results in the formation of a weak platelet plug. However, due to the deficient, absent,
or defective Von Willebrand Factor, the platelets cannot adhere together properly or glue
themselves appropriately to the blood vessel walls. The clot will take longer to form or will
form improperly as a result, and the bleeding will take longer to stop. This can result in
significant, difficult-to-stop bleeding. Although uncommon, the bleeding can cause injury to
joints or internal organs, as well as be life-threatening.
Additionally, vWD is caused by defects in the vWF. This typically affects the primary
hemostasis, wherein it results in mucocutaneous bleeding tendency simulating a platelet
function defect. The vWF functions to initiate platelet adhesion to the injured vessel wall and
it serves as a carrier for Factor VIII in the plasma. In the primary hemostasis, vWF binds to
the platelet glycoprotein 1b and links to the injured vessel and other platelets. While in the
secondary hemostasis, the vWF binds to the Factor VIII and protects it from being removed
early.
As mentioned above, vWD also affects secondary hemostasis. vWF is a cofactor in
the adhesion of platelets to the exposed subendothelium and protects factor VIII from
inactivation, preventing it from being broken down (metabolized) before reaching the site of
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Hematology 2
2nd Semester, AY 2021-2022

damage. In cases of vWD wherein there is a deficiency or defective vWF, it fails to act as a
glue to hold platelets together at the site of injury to a blood vessel. Consequently, platelets
do not adhere to the blood vessel wall and the blood clot breaks down prematurely. In some
situations, the lack of vWF results in decreased amounts of factor VIII in the blood, causing
blood clots to take an unusually lengthy time to form. These abnormal activities impact
bleeding time results, with prolonged activated partial thromboplastin time (APTT).

3. What are the different types and subtypes of this disease? Differentiate them in terms
of their etiology/pathophysiology and laboratory findings.

TYPES SUBTYPES ETIOLOGY / PATHOPHYSIOLOGY

Type 1 It is the most common and mildest form of vWD that is characterized by a
[75%] quantitative defect wherein the individual affected has low levels of vWF in
their bloodstream. Consequently, since vWF is also responsible for protecting
factor VIII , patients with this type of vWD also show deficiency in this protein (↓
vWF = ↓ factor VIII). It is often caused by autosomal dominant frameshifts,
nonsense mutations, or deletion mutations in the gene expression of the vWF
gene which leads to an impaired intracellular transportation of its sub-units.
Patients with this type of vWD often present mild to moderate bleeding.
Menorrhagia among women is a common symptom that leads to the diagnosis
of this disease.

Type 2 It is characterized more by qualitative defects wherein the individual may

have normal or moderately decreased levels of vWF, but with reduced
function in blood clot formation due to functional or structural defects of the
vWF multimers.

Subtype 2A It is caused by well-known autosomal dominant point

[10-20%] mutations in the D1 and A2 structural domains of the vWF
molecules. Due to these mutations, VWF becomes more
 Angeles University Foundation
College of Allied Medical Professions
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Hematology 2
2nd Semester, AY 2021-2022

susceptible to ADAMTS-13 proteolysis which provides

instructions on how to produce enzymes that regulate blood
clotting, resulting in a predominance of small molecular
weight plasma multimers. There is abnormality in the size of
vWF multimeters, wherein it may be too small or too big,
thereby not allowing the platelets to adhere properly and
form a platelet plug. The defective assembly of vWF
multimers in the golgi apparatus can lead to the deficiency
of high & intermediate-molecular-weight multimers in the
circulation which works most effectively in platelet adhesion
compared to small-molecular-weight multimers. Patients with
this type of VWD present normal or slightly reduced vWF:Ag
levels with a markedly reduced vWF.

Subtype 2B These are caused by mutations in the A1 domain,

[>5%] increasing the affinity of vWF for platelet GPIb/IX/V which
is its usual binding location. It constitutes “gain-of-function”
mutations. The vWF multimers are not in the right size, but
due to a decreased level of large multimers in the plasma
and a markedly increased proteolysis, this causes vWF to
become too active. Hence, in this subtype, the vWF
attaches to the platelets at the wrong time or when they
are not supposed to. As a normal response, the body
removes the platelets attached to vWF, resulting in
decreased levels of platelets and VWF in the blood then
shortage when needed to form a clot when an actual injury
occurs.

Subtype 2M The vWF is unable to attach to the platelets as it normally

should when an injury occurs, rendering decreased platelet
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Hematology 2
2nd Semester, AY 2021-2022

receptor binding and inability to form a good platelet plug,

despite having normal secretion and assembly of vWF
multimers. It is caused by a mutation that impairs the ability
of vWF to bind to platelets leading to decreased exposure of
vWF to ADAMTS-13.

Subtype 2N It is referred to as the Normandy variant since this type of

[>5%] VWD is first described in Normandy, France. In this type of
VWD, the vWF can normally attach to platelets (normal
vWF:Ag) but has decreased affinity for factor VIII
(deficient), another protein that is also needed for blood to
clot. The factor VIII binding site is impaired by an
autosomal missense mutation. Since vWF cannot bind and
protect factor VIII, the body will begin to remove it from the
bloodstream, leading to a decreased level of factor VIII.
This subtype is also known as autosomal hemophilia due
its nearly identical clinical symptoms from hemophilia,
except that VWD Type 2N affects both men and women. It is
suspected from a woman experiencing anatomic bleeding
symptoms, alongside being diagnosed with hemophilia. On
the other hand, VWD Type 2N is considered when a male
patient who is misdiagnosed with hemophilia A fails to react
to factor VIII concentrate therapy.
Since there are lower levels of factor VIII than of vWF, it can
be misdiagnosed as a case of hemophilia since they have
similar symptoms. So it is important to take note that in this
type of VWD (autosomal, affects males and females), the
problem involves the inability of vWF to protect factor VIII
and not factor VIII itself, such as in the case of hemophilia
(x-linked, affects females only).
 Angeles University Foundation
College of Allied Medical Professions
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Hematology 2
2nd Semester, AY 2021-2022

Type 3 It is the rarest, yet most severe form of vWD that is characterized by an
[3%] autosomal recessive (missense or nonsense) mutation, resulting in
extremely low levels or absence of vWF (undetectable) and deficiency of
factor VIII in the blood. These deficiencies impairs both stages of hemostasis
which causes the body to be unable to properly form a platelet plug and a
stable fibrin clot, hence inducing the most severe symptoms of this disease.
Severe bleeding may occur in the form mucocutaneous and anatomic
hemorrhage with severe mucosal bleeding or epistaxis, bleeding from the
mouth, or even bleeding between their joints and muscles. Women with this
type of vWD tend to have a prolonged menstrual period along with a heavy
flow. Due to the severity of these symptoms, type 3 vWD is almost often
diagnosed at a young age.

LAB TYPE SUBTYPE SUBTYPE SUBTYPE SUBTYPE TYPE

TEST 1 2A 2B 2M 2N 3

VWF: Low Low Low Low Normal Very low

Ristocetin or absent
cofactor, VWF:
Collagen
binding, VWF:
Immunoactivit
y, VWF: GPIbR,
VWF: GPIbM

VWF: Antigen Low Normal to Normal to Normal Normal Very low

slightly slightly or absent
decreased decreased

VWF activity to >0.5 <0.5 <0.5 <0.5 >0.5 N/A

VWF: Ag ratio
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Platelet count Normal Normal Decreased Decreased Normal Normal

Partial Normal to Normal Normal Normal to Normal to Prolonged

thromboplastin slightly slightly slightly
time (PTT) prolonged prolonged prolonged

RIPA Decreased Decreased Increased Decreased Normal Absent

Factor VIII Slightly Normal Normal Normal Low <10

activity low units/dL

VWF multimers Normal Large and Large Normal Normal All forms
pattern intermedia forms pattern pattern absent
te forms absent
absent

4. What specific tests may be performed to definitively diagnose this disease? You may
include tests that were mentioned in the case and those that were not mentioned.

Von Willebrand disease (vWD) is considered the most prevalent hereditary bleeding
disorder but diagnosis may also be the hardest. Mild mucosal bleeding is its most common
clinical symptom, while surgical bleeding can be exhibited with some challenges, and joint
bleeding may be experienced in the most severe cases. Information regarding family histories
either of diagnosed VWD or of bleeding symptoms is beneficial.
A number of tests of von Willebrand factor (vWF) quantity and function, as well as factor
VIII activity, are required for laboratory diagnosis. A combination of blood tests is required to
assess the function of Von WIllebrand factor (vWF). The Complete Blood Count (CBC),
Activated Partial Thromboplastin Time (APTT) Test, Prothrombin Time (PT) Test, and Fibrinogen
Test are all screening tests that are used to determine how well the blood clots. These
screening tests are done first to determine if blood clotting is functioning properly and levels of
clotting factors found in blood fluctuate over time due to the changes that happen when the
body responds to illness, pregnancy, and stress.
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Hematology 2
2nd Semester, AY 2021-2022

The following are some instances of screening tests for vWD that may reveal the
presence of a bleeding condition, but more detailed testing is required to determine the kind
of bleeding disorder:
Complete blood count (CBC) - It is a common test that measures the amount of
hemoglobin and types of white blood cells and platelets found in blood. Usually CBC has a
normal result among patients with vWD. However, due to unusually heavy bleeding it may
lead to low red blood cell count. In our case, the patient shows a normal result of
hemoglobin, hematocrit, and platelet count. However, bleeding time is prolonged which
indicates there is a problem with the blood clot of the patient.
● Activated Partial thromboplastin time (APTT) test - It is a test that measures the
length of time that bloods coagulate. Also, measures the coagulation ability of factor VIII,
IX, XI, and XII. A result of longer clotting time will show if any of these factors are proven
deficient in the patient's body.
● Prothrombin time - This test measures primarily clotting ability of factor I, II, V, VII, and
X. In which, it also measures the time it takes for a blood to clot. Patients with vWD show
a normal result. (Extrinsic)
● Fibrinogen test - Also known as clotting factor I in which is a test that helps the
physician to assess the ability to form a blood clot. This test is usually conducted when a
patient has abnormal results of PT or APTT or it can act as a combination test with other
blood clotting tests. Results for patients with vWD may have a result of normal
fibrinogen.
● vWF antigen - is a screening test that evaluates the protein level of vWF and it can be
performed by using ELISA or automated latex immunoassay. However, it only
determines the presence of vWF but not the function of it. Therefore, it requires
additional testing to assess the dysfunction of vWF.
● vWF ristocetin cofactor activity - It is considered widely and commonly used in
assessing the function of vWF in attaching with GPIbα.
● Factor VIII activity - This test is considered an important factor to assess for suspected
vWD because vWF is a carrier protein for Factor VIII, which is included in the first
workup. Under the normal physiologic conditions the ratio of Factor VIII activity is around
1. However, decreased level will be found for type 2N and type 3 VWD will be <10 IU/dL.
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2nd Semester, AY 2021-2022

To confirm which bleeding disorder is there, these are some examples for specific
tests for patient who are suspected with vWD:

● Ristocetin - Induced platelet aggregation (RIPA) - it is consider an in - vitro test that

evaluates the presence of platelet glycoprotein (GP) Ibα - V - IX complex and vWF
interaction and it is commonly conduct using platelet - rich plasma (PRP). A specimen of
5 - 10 blood in citrate is used. In type 2B vWD and platelet - type vWD, unexpected
aggregation to low - dose ristocetin with range value of 0.6 mg/dL is observed. In
addition, vWF-platelet bindings are necessary to distinguish between vWD variants.
● vWF gene sequencing - Due to the huge size of the vWF gene and the high intensity of
normal variations, genetic diagnosis is limited with vWF. If a confirmatory test is
unavailable or difficult to interpret the result, DNA sequencing can be performed for type
2 vWD. Mutations in the vWF A1 domain are seen in type 2B and type 2M patients with
platelet binding defects. Mutations in the VWF D′ and D3 domains are observed in type
2N, where the defect involves FVIII binding. Defects in type 2A can occur in the A2
domain (near the AD-AMTS13 cleavage site) or in the N or C terminal multimerization
domains. Types 1 and 3 VWD defects can be discovered across the vWF gene, although
new sequence changes should be evaluated with caution because not all variants are
linked to VWD.
● vWF-FVIII binding (VWF:FVIIIB) - tests the capacity of vWF extracted from patient
plasma to bind recombinant Factor VIII and is performed to confirm a diagnosis of type
2N vWD, which is characterized by a particular deficiency in Factor VIII binding.
● vWF multimer distribution - a normal multimer distribution indicates the existence of
type 2M vWD, while a loss of high molecular weight multimers indicates the presence of
type 2A or type 2B vWD. Agarose gel electrophoresis is commonly used for multimer
analysis, however it is not conducted by all laboratories as it is time consuming.
● vWF collagen binding (VWF:CB) - vWF testing uses collagen binding in two ways. It
may be used to identify large molecular weight multimers, and help identify type 2M vWD
as they have a unique collagen-binding deficiency.
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● vWF–platelet binding (VWF:PB) - an increase in spontaneous platelet binding confirms

Type 2B vWD. This assay will show higher binding in type 2B vWD but no increase in
type 2A vWD with the use of commercial platelets and patient plasma.
● vWF propeptide (VWFpp) - while this method may not be accessible in all laboratories,
observing an increase in the VWFpp/VWF:Ag ratios may indicate the presence of type
1C, or clearance defects, and some type 2 vWD variations
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REFERENCES:
Bharati, K. P., & Prashanth, U. R. (2011). Von Willebrand disease: an overview. Indian journal of
pharmaceutical sciences, 73(1), 7–16. https://doi.org/10.4103/0250-474X.89751

Centers for Disease Control and Prevention. (2020, October 26). VWD diagnosis. Centers for
Disease Control and Prevention. Retrieved March 16, 2022, from
https://www.cdc.gov/ncbddd/vwd/diagnosis.html#:~:text=The%20blood%20tests%20that%20a,o
f%20VWF%20in%20the%20blood

Centers for Disease Control and Prevention. (2021, April 1). What is von willebrand disease?
Centers for Disease Control and Prevention. Retrieved March 16, 2022, from
https://www.cdc.gov/ncbddd/vwd/facts.html

Clinlab Navigator. ClinLabNavigator. (n.d.). Retrieved March 16, 2022, from

http://www.clinlabnavigator.com/ristocetin-cofactor.html

Hemophilia of Georgia. (n.d.). Types of von willebrand disease. Retrieved March 16, 2022, from
https://www.hog.org/handbook/article/1/7/types-of-von-willebrand-disease

Ristocetin-Induced Platelet Agglutination [RIPA]. Sang Medicine . (n.d.). Retrieved March 16,
2022, from
https://practical-haemostasis.com/Factor%20Assays/vwf/VWF%20Functional%20Assays/ripa_t
est.html

Ristocetin-induced platelet aggregation. RCPA. (n.d.). Retrieved March 16, 2022, from
https://www.rcpa.edu.au/Manuals/RCPA-Manual/Pathology-Tests/R/Ristocetin-induced-platelet-
aggregation
 Angeles University Foundation
College of Allied Medical Professions
Department of Medical Technology

Hematology 2
2nd Semester, AY 2021-2022

Roberts, J. C., & Flood, V. H. (2015, May). Laboratory diagnosis of Von Willebrand disease.
International journal of laboratory hematology. Retrieved March 16, 2022, from
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5600156/

Rodak, B. F. (2016). Hematology: Clinical Principles and Application (5th ed.). Elsvier Saunders.

StatPearls. (2021, September 8). Physiology, clotting mechanism. StatPearls. Retrieved March
16, 2022, from https://www.statpearls.com/ArticleLibrary/viewarticle/19639#:~:text=Primary
%20hemostasis%20is%20the%20formation,platelet%20activation%2C%20and%20platele
t%20aggregation

Yawn, B. P., Nichols, W. L., & Rick, M. E. (2009, December 1). Diagnosis and management of
von willebrand disease: Guidelines for primary care. American Family Physician. Retrieved
March 16, 2022, from https://www.aafp.org/afp/2009/1201/p1261.html
ANGELES UNIVERSITY FOUNDATION
College of Allied Medical Professions
Department of Medical Technology

Case Analysis:
Vitamin K Deficiency

Group 3 | BSMT 3D

Gatchalian, Beatriz
Manalansan, Darcee Lienne
Ponce, Kaila Therese
Romero, Reianne
Salac, Jose Conrado
Santos, Marc Anjelo
Sotto, Charmaine
Valenzuela, Kathlyn Joy

March 2022
Case 3:

A 1-month-old baby was rushed to the emergency department due to constant bruising and
bloody stool. The patient was born through a C-section and there were no reported
complications. He was also exclusively breastfed. However, the mother had noted that the baby
had become paler and more irritable a few weeks after birth.

The patient was responsive during the initial examination during admission and his vital signs
were normal. The results of the requested hematology and coagulation tests are as follows:

Test Result Normal Value (Keohane, Smith,

Walenja, 2016)

WBC Count 10 x 10 ^9/L 6.0-18.0 x 109/L

RBC Count 3.5 x 10^12/L 3.40-5.00 x 1012/L

Hemoglobin 9.5 g/dL 10.6-16.4 g/dL

Hematocrit 25% 32-50%

PT Prolonged 10-13 seconds

APTT Slightly prolonged 25-35 seconds

Mixing studies Corrected with addition of

 normal plasma and aged serum,
not corrected with adsorbed
plasma

Thrombin time Normal Less than 20 seconds

QUESTIONS:

1. What is the probable diagnosis of the patient? Give the etiology and pathophysiology
of the disorder.
A. Etiology

Given the signs and symptoms, age, and test results of the patient, the probable
diagnosis is Vitamin K Deficiency (VKD). This condition is classified under Acquired
Coagulopathies wherein it is caused by the absence of Vitamin K-dependent clotting factors
(also known as Prothrombin Factors), namely the Factors II, VII, IX, and X (Keohane, Otto, &
Walenga, 2020). A deficiency of these factors may lead to significant bleeding, poor bone
development, osteoporosis, and an increased risk of cardiovascular disease (Eden & Coviello,
2022). Aside from that, other causes include insufficient uptake in the diet of an individual,
intake of certain drugs or antagonists (such as Coumadin) that results in disruption in the normal
flora that contributes to the formation of vitamin K, and malabsorption. This condition can affect
normal healthy adults, however, it is more commonly associated with infants as they are
constitutionally Vitamin K deficient upon birth due to the sterility of their intestines as well as the
reduced concentration of vitamin K in breastmilk (Eden & Coviello, 2022; Keohane, Otto, &
Walenga, 2020). As cited by Eden & Coviello (2022) and University of Rochester Medical
Center, VKDB in newborns is categorized into three based on the time of its manifestation: Early
VKDB (0-24 hours after birth), Classic VKDB (1-7 days after birth), and Late VKDB (1-12 weeks
up to 6 months after birth).

TYPES OF WHEN IT OCCURS CHARACTERISTICS

VKDB

EARLY 0-24 hours after birth ● Severe

● Infants whose mother used certain medications
= interference with how the body uses Vitamin
K

CLASSICAL 1-7 days after birth ● Bruising

● Bleeding from the umbilical cord
 LATE 2-12 weeks after ● Intracranial hemorrhage (30-60%)
birth, up to 6 months ● Breastfed only infants with no Vit K shot
● Warning bleeds (rare)

B. Pathophysiology

Vitamin K is a group of chemicals with a typical naphthoquinone structure that comes in

two forms: vitamin K1 (phylloquinone) and vitamin K2. The former can be acquired from a
variety of foods such as plant oils, green leafy vegetables, and the like, whereas the latter, also
known as menaquinones, is produced by bacteria in the colon but can also be taken from meat,
cheese, or liver.

Vitamin K also serves as a cofactor for the enzyme 𝜸-glutamyl carboxylase. The stated
enzyme is in charge of post-translational carboxylation, which involves the conversion of
glutamate residues in proteins to 𝜸-carboxyglutamate (Gla), a step mediated by the 𝜸-glutamyl
carboxylase. Gla residues are critical for protein function because they enhance or permit
calcium-binding and proper assembly of vitamin K-dependent clotting components on
phospholipid membranes. Some coagulation factors, particularly factors II (prothrombin), VII, IX,
and X, are Gla-containing proteins that rely on vitamin K; thus, a vitamin K deficiency can result
in clinically significant bleeding. The reason for this is because when there is a vitamin K deficit,
undercarboxylated proteins called protein-induced by vitamin K absence (PIVKA) develop,
which affects or inhibits the function of vitamin K dependent-proteins in calcium-binding as well
as normal phospholipid membrane assembly. Increased PIVKA-II is a useful biomarker of
vitamin K deficiency because the amount of aberrant prothrombin is increased due to the
deficiency. Furthermore, due to a lack of vitamin K, there will be a decrease in the proteins C, S,
and Z, which are involved in blood coagulation. This type of deficiency is commonly seen in
babies, and it is most likely caused by inadequate consumption of the vitamin. On the contrary,
this can occur at any age due to a reduction in vitamin K absorption.

Vitamin K levels in neonates are low and occasionally undetectable; as a result,

vitamin-K-dependent coagulation levels are equally low. The most likely cause or
pathophysiology of vitamin K shortage in babies is that vitamin K does not easily transfer
through the placenta, and the amount of phylloquinone measured in the umbilical cord blood is
quite low. Following that, newborns and babies have a sterile gut, which prevents vitamin K
production. It will take some time for bacteria like Bacteroides fragilis to develop, which is
required for vitamin K2 production. Lastly, because there are passive antibodies taken from the
mother, breastfeeding may cause a delay in the establishment of normal flora in the gut.
Furthermore, Lactobacilli, a kind of bacterium that does not synthesize vitamin K, are found in
breastmilk, affecting vitamin K2 formation. Infants are more likely to have vitamin K deficiency,
which can lead to bleeding if not treated; hence the name vitamin K deficiency bleeding (VKDB)
or formerly known as hemorrhagic disease of the newborn (HDN), when the cause of the
coagulopathy is a vitamin K deficit. This disease or deficit causes bleeding in the umbilicus,
gastrointestinal tract (GIT), mucous membrane, and venipuncture or circumcision sites in
infants. (Eden & Coviello, 2022; Keohane, Otto, & Walenga, 2020)
2. Which of the given information in the case supports your diagnosis? Explain the
significance of each abnormal laboratory test result to the disease.

A. Clinical Manifestations

Newborns are congenitally vitamin K deficient because of their sterile intestines wherein
the bacteria that will produce the said type of vitamin is not yet present. It is also caused by the
limited concentrations of vitamin K in the human breast milk and only small amounts of it are
received in the placenta from their mother. In addition to this, in newborns, although normal, the
vitamin K-dependent clotting factors' activity levels are low. With this, vitamin K deficiency is
usually presented with a history of bleeding and bruises due to the impaired function of the
secondary hemostasis. The baby becoming paler a few weeks after birth also supports the
clinical manifestations of the diagnosis due to bleeding that leads to blood loss which may be
associated with anemia. The baby being irritable may be a sign that there is bleeding present in
the brain. Therefore, the clinical manifestations which involve constant bruising and bloody stool
of the patient are information that supports the diagnosis of Vitamin K deficiency (Keohane
et.al., 2016).

B. Laboratory Results

The patient's results in the coagulation tests, specifically the PT, APTT, and mixing
studies, support the diagnosis of Vitamin K Deficiency.

Prothrombin time (PT) measures the extrinsic which includes the factors III and VII, and
the common pathway that has the factors I, II, V, and X. The prolonged result of PT in the
patient may indicate that it is due to a deficiency in one or more factors from the pathways. This
means that the deficiency of Vitamin K in the baby’s body contributes to the absence of
coagulation factors which are needed for blood clotting to stop the bleeding. A prolonged PT
with or without a prolonged APTT supports the clinical suspicion of Vitamin K deficiency. In
addition, activated partial thromboplastin time (APTT) measures the integrity of the intrinsic and
common pathways. Slightly prolonged results in the patient's APTT again may suggest that
there is a deficiency in the factors included in the mentioned pathways but it proves that the
case is not yet severe wherein if both the APTT and PT are prolonged, it means the case is
already complicated or at severe state.

More so, mixing studies using adsorbed plasma, is used for testing various coagulation
disorders. It lacks the clotting factors II (prothrombin), VII, IX, and X which are known to be
vitamin K-dependent clotting factors. In the patient's result, it stated that the adsorbed plasma is
not corrected which means that it lacks these factors (II, VII, IX, and X). The absence of the
important factors will prevent the activity of clotting that leads to being prone to experience
bruising. According to Johnson, 2020, a bloody stool can be observed because of intestinal
bleeding that can’t be stopped by the secondary hemostasis. Therefore, due to the deficiency, it
strongly suggests that the diagnosis is Vitamin K deficiency.

Lastly, the laboratory results yielded decreased hemoglobin and hematocrit levels and it
was mentioned by the mother that the patient became paler and more irritable after birth. These
may suggest that the patient has iron-deficiency anemia as well. Iron-deficiency anemia is most
common during late pregnancy, especially in infants who only feed on milk, which is low in iron.
(Wang, M. 2016) Vitamin K deficiency in itself does not cause anemia. However, its symptoms,
which include bleeding, may be associated with anemia. The hemoglobin level resulting in a
decreased amount was contributed by the blood loss of the body which can be observed on the
bloody stool. The hematocrit level is also decreased because of bleeding making the blood
diluted thus the percentage of the total healthy red blood cells present in the blood has dropped.
But the red blood cells being normal does not agree with the abnormal levels of hemoglobin and
hematocrit thus concluding that Vitamin K deficiency is absolutely associated with
Iron-deficiency anemia also known as hypochromic anemia. This means that there are enough
red blood cells in the baby’s body but iron is insufficient to carry the needed oxygen for
circulation making the red blood cells unhealthy. (Wang, 2016)

3. Give hematological or other laboratory tests to confirm the diagnosis of the disorder.

For further confirmation of the diagnosis of the disorder, the following hematological or
laboratory tests may be used:

A. Clotting Time, Thrombin time, and Platelet count

The clotting time may also be useful in determining the Vitamin K deficiency. The
results would be increased or prolonged because of the defect among the coagulation
factors. Whereas, the platelet count results are normal as well as the thrombin time
because the fibrinogen level in Vitamin K deficiency is normal (Kher & Verma, 2021).

B. Peripheral Blood Smear and Ferritin Level

Hematological examination of the patient with Vitamin K deficiency may reveal
anemia with hypochromia, anisocytosis, and poikilocytosis. As indicated, the RBC count
 is normal but the hematocrit and hemoglobin are significantly decreased which is why
the patient may have anemia. This is clinically manifested by bloody stool that may be
due to the gastrointestinal tract (GIT) bleeding indicating problems in the secondary
hemostasis. Furthermore, measurement of the ferritin level may be useful and the
expected result would be low levels of ferritin (iron-storing blood protein) (Kumar, Abbas,
& Aster, 2015).

C. PIVKA-II level estimation: HPLC, ELISA, Immuno-electrophoresis

PIVKA-II are proteins that are induced from the absence of Vitamin K. These
proteins are recognized as a sensitive biomarker in the diagnosis of the said disorder
among newborns and adults. Thus, PIVKA-II may have minimal variations due to
different factors such as age that may affect the concentration of vitamin K plasma and
serum. Among individuals who consume less than 60 mcg of vitamins a day may result
in an increased PIVKA-II level. Then, after birth, 10%-50% of newborns are observed to
have elevated PIVKA-II levels (Eden & Coviello, 2021). The presence of these proteins
is an indication of deficiency in vitamin K, hence regarded as abnormal. Their presence
could be evaluated using HPLC, ELISA, and Immuno-electrophoresis. Note that it may
disappear on the 5th day of vitamin K administration (Kher & Verma, 2021).

D. Others: Measurement of Phylloquinone

For further confirmation of the disorder, the measurement of phylloquinone in
serum is done. It is considered as the most commonly used marker in determining
Vitamin K status thus, exhibiting the abundance of vitamin K among the individual. With
 regards to, a <0.15 ug/L of phylloquinone concentration is suggestive for deficiency
(Card et.al., 2019).

4. How is the patient’s condition managed/treated?

Vitamin K deficiency can be prevented with Vitamin K administration. It can help prevent
hemorrhagic disorders in newborns. This can be done by delivering it orally or intramuscularly.
Infants predisposed to early HDN due to maternal drug use should be given 1 mg of vitamin K
at birth and may be done intravenously. The infant may be delivered via cesarean section to
avoid the trauma of vaginal delivery. High-risk mothers may be given vitamin K orally 7-10 days
before delivery to help prevent it (Behera & Kulkarni, 1998).

The treatment for early and classic forms of infant hemorrhagic disease is an oral dose
of vitamin K with a 2mg dosage repeated every 2-4 weeks and every 6-8 weeks. Parenteral
vitamin K is not as effective as oral vitamin K in the late stages of the condition; therefore,
newborns are given a single intramuscular dose of 0.5-1 mg. The dosage of vitamin K varies
depending on the weight of the infants. Intramuscular injection of a single 0.5 mg dose is
administered to infants weighing less than 1500 gm, and a single dose of 1 mg is given to
newborns weighing more than 1500 gm. To prevent late vitamin K deficiency hemorrhage, all
breastfed newborns with diarrhea and malabsorption require an additional post-natal dosage of
vitamin K. 10-20 mL/kg of fresh frozen plasma should also be given in the case of
life-threatening hemorrhages. More than 20% of blood loss and shock symptoms necessitate
emergency blood transfusions (Kher & Verma, 2021).
 Summarized under Table 1 are the different treatment or management procedures done
for Vitamin K deficiency (Eden & Coviello, 2021).

Prophylaxis in newborns Intramuscular injection

● 1 mg of vitamin K1 within 1 hour of birth
● Preferable for efficacy

Orally
● 2 mg of Vitamin K1 at birth at 4-6 days and
at 4-6 weeks
● 2 mg of Vitamin K1 at birth and
consecutive weekly dose of 1 mg for 3
months

VKDB Slow intravenous or subcutaneous infusion:

● 1-2 mg of Vitamin K1
● Severe bleeding: 10-15 mL/kg of fresh
frozen plasma

Deficiency due to malabsorption ● Daily administration of 0.3-15 mg/day of

Vitamin K1
● Ineffective oral dosing: Parenteral vitamin
K1 should be considered

VKCFD Oral dose by intravenous infusion:

● 10 mg of Vitamin K1 2-3x per week
● Severe bleeding/surgery: 15-20 mL/kg of
fresh frozen plasma, Prothrombin Complex
Concentrates and recombinant Factor VII

Nutritional deficiency in adults By diet or oral supplementation:

● At least 120 and 90 ug/day for men and
 women

Chronic conditions An increased dosage of oral Vitamin K1 and K2

may be helpful. However, there are no present
guidelines available.

Table 1. Treatment/Management for other types of VKD

References:

1. Keohane E.M, Otto C.N, and Walenga J.M. (2020). Rodak’s Hematology: Clinical

Principles and Applications (6th ed.). Canada: Elsevier Inc.

2. Eden, R. E., & Coviello., J. M. (2021, July 26). Vitamin K Deficiency. Retrieved from
https://www.ncbi.nlm.nih.gov/books/NBK536983/#:~:text=Vitamin%20K%20Deficiency%
20Bleeding%20
3. Kher, P., & Verma, R. P. (2021, July 03). Hemorrhagic Disease Of Newborn. Retrieved
from https://www.ncbi.nlm.nih.gov/books/NBK558994/
4. Kumar, V., Abbas, A. K., & Aster, J. C. (2020). Robbins & Cotran pathologic basis of
disease (10th ed.). Elsevier
5. Card, D. J., Gorska, R., & Harrington, D. J. (2019, December 20). Laboratory
assessment of vitamin K status. Retrieved from
https://pubmed.ncbi.nlm.nih.gov/31862867/
6. Behera, M., & Kulkarni, S. (1998). VITAMIN ‘K’ DEFICIENCY HAEMORRHAGIC DISEASE
OF NEW-BORN AND PRESENT CONTROVERSIES. Medical Journal Armed Forces India,
54(2), 143-145. doi: 10.1016/s0377-1237(17)30506-3
7. University of Rochester Medical Center (n.d.). Vitamin K Deficiency Bleeding in the
Newborn. Retrieved from
https://www.urmc.rochester.edu/encyclopedia/content.aspx?ContentTypeID=90&Content
ID=P02372#:~:text=Babies%20are%20normally%20born%20with,severe%20bleeding%
20or%20a%20hemorrhage
8. Lippi, G., and Franchini, M. (2011). Vitamin K in neonates: facts and myths. Blood
transfusion = Trasfusione del sangue, 9(1), 4–9. https://doi.org/10.2450/2010.0034-10
9. Wang, M. (2016). Iron Deficiency and Other Types of Anemia in Infants and Children.
Iron Deficiency and Other Types of Anemia in Infants and Children - American Family
Physician (aafp.org)
10. Blood clotting process. (2018, September 3). Retrieved March 14, 2022, from
News-Medical.net website:
https://www.news-medical.net/health/Blood-Clotting-Process.aspx
11. CDC. (2021, January 20). What is Vitamin K Deficiency Bleeding? Retrieved March 14,
2022, from Centers for Disease Control and Prevention website:
https://www.cdc.gov/ncbddd/vitamink/facts.html
12. Hematocrit test. (n.d.). Retrieved March 14, 2022, from Mayoclinic.org website:
https://www.mayoclinic.org/tests-procedures/hematocrit/about/pac-20384728
13. Hemoglobin - complete blood count. (n.d.). Retrieved March 14, 2022, from Rnceus.com
website: https://www.rnceus.com/cbc/cbchg.html
14. Perfectionalis, M. [MedicosisPerfectionalis]. (2020, July 23). Vitamin K deficiency |
Hemorrhagic Disease of the Newborn. Retrieved March 14, 2022, from
https://www.youtube.com/watch?v=q2YKVtpwJWc
15. Vitamin K Deficiency. (n.d.-a). Retrieved March 14, 2022, from MSD Manual
Professional Edition website:
https://www.msdmanuals.com/professional/nutritional-disorders/vitamin-deficiency,-depe
ndency,-and-toxicity/vitamin-k-deficiency
16. Vitamin K Deficiency. (n.d.-b). Retrieved March 14, 2022, from MSD Manual Consumer
Version website:
https://www.merckmanuals.com/home/disorders-of-nutrition/vitamins/vitamin-k-deficiency
17. (N.d.). Retrieved March 14, 2022, from Sciencedirect.com website:
https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/vitami
n-k-deficiency#:~:text=The%20classic%20Gla%2Dcontaining%20proteins,has%20a%20
role%20in%20coagulation.
 ANGELES UNIVERSITY FOUNDATION
Angeles City
COLLEGE OF ALLIED MEDICAL PROFESSIONS
DEPARTMENT OF MEDICAL TECHNOLOGY

CASE 4:
Disseminated Intravascular
Coagulation

Jihan Larkin Cruz

Dolorie Mae Hernandez

Mara Joyce Ituriaga

Gabriel Joey Licup

Aubrey Rose Lugay

Gianne Frangel Montemayor

Jeyanna Valdez

Deina Pamela Valerio

BSMT 3-F
 I. CASE PRESENTATION

Case 4:

A 40-year-old female was admitted to a hospital with a working diagnosis of Septic Shock. The
patient complains of bloody diarrhea approximately 5 times per day, abdominal pain, and
nausea for the past 4 days. He also complains of intermittent fevers and dry mouth. Preliminary
lab results include excessive hematochezia and a positive blood culture for Gram Negative
organisms. Other Laboratory results are as follows:

WBC 25 x 103/UL

RBC 3.8x106/UL

Hct 20%

PC 50,000/UL

PT value more than upper range

of machine

D-Dimer Positive

Fibrinogen 50 mg/dL

II. GUIDE QUESTIONS AND ANSWERS

1. What is the most probable Diagnosis of the patient? What led you to this conclusion?

Based on the given case, the most probable diagnosis of the patient is Disseminated
Intravascular Coagulation (DIC). DIC is also known as defibrination syndrome or consumption
coagulopathy and is an acquired clinicobiological condition characterized by widespread and
uncontrollable blood coagulation activation, which leads to fibrin deposition within the
vasculature, organ dysfunction, clotting factor and platelet consumption, and life-threatening
bleeding. It is often seen in severe sepsis and septic shock. Moreover, infections, usually
caused by Gram-negative microorganisms, may also trigger DIC through the production of
endotoxins that expose collagen.

Its clinical manifestations include fever, hypotension, hemorrhage, thrombosis, and

shock. However, these manifestations are often masked by the symptoms of the underlying
condition of the patient. The signs and symptoms experienced by the patient which include
bloody diarrhea, abdominal pain, nausea, intermittent fevers, dry mouth as well as her
preliminary laboratory results are all related to sepsis and bacterial infection. Furthermore, the
other laboratory findings presented such as increased WBC count, normal RBC count,
decreased hematocrit levels, decreased platelet count, prolonged prothrombin time, positive
D-dimer test, and decreased fibrinogen levels are indicative of DIC. The laboratory results
suggest that the patient has an acquired syndrome called Disseminated Intravascular
Coagulation (DIC).

2. Give the pathophysiology, clinical signs and symptoms of your diagnosis.

2.1 PATHOPHYSIOLOGY

2.1.1 Normal hemostasis vs. DIC

In normal conditions, formation of a temporary platelet plug will occur during primary
hemostasis and stabilization of that clot will occur at the secondary hemostasis. After the
formation of the stable fibrin clot, fibrinolysis will occur in order to dissolve it so that unnecessary
blood clotting will be prevented. However, in DIC, there is an abnormal spread of blood clots all
over the body.

Disseminated intravascular coagulation is a sequela of many different diseases. Among

the common “triggering events” that could make a person susceptible to acquiring DIC are
infections, extravascular trauma, cancer (especially leukemia), snakebites, intravascular
hemolysis, and inflammation. In the patient’s case, the event that complicates DIC is sepsis,
which is an infection in the blood commonly caused by bacteria. Klebsiella pneumoniae,
Escherichia coli, and Pseudomonas aeruginosa are some of the gram negative bacteria that
cause sepsis. DIC is explained to be paradoxical because in some parts there can be too much
clotting, and in other parts there is too much bleeding.
 Even though DIC has different causes, the pathway of the disease will always lead to
excessive thrombin formation. Hence, the main culprit is circulating thrombin, which activates
platelets, coagulation proteins, and fibrin formation.

2.1.2 Events that lead to excessive clotting

Coagulation or massive clot formation happens due to the release of bacterial

endotoxins. These endotoxins can release procoagulant factors that may put the patient in a
hypercoagulable state. These procoagulants can be in the form of tissue factor,
lipopolysaccharide and enzymes. In addition, endotoxins can activate cytokines that can affect
the coagulation pathway. As the endotoxins induce the expression of the tissue factor, the
extrinsic coagulation pathway is activated. Endothelial exposure to cytokines such as TNF-a
and IL-1b will lead to the activation of the intrinsic pathway. The overactivation of the intrinsic
and extrinsic pathway will also activate the common pathway and will lead to the production of
excessive thrombin. Because of this, the coagulation cascade goes into overdrive as a
response, resulting in extensive clot formation, which blocks blood vessels and causes
ischemia, necrosis and damage to organs. Aside from extensive clot formation, excess in
thrombin may also lead to the depletion of platelets in the circulation since thrombin can
stimulate platelets to undergo aggregation.

2.1.3 Events that lead to excessive fibrinolysis/bleeding

The activation of plasminogen starts the fibrinolytic process. In the common pathway,
fibrinogen is cleaved by thrombin, resulting in fibrin. In normal hemostasis, fibrin monomers
polymerize spontaneously to create an insoluble clot through the help of factor XIII.
Cross-linking strengthens the polymer, which binds plasminogen as it develops. However, a
portion of fibrin monomers fail to polymerize in DIC and circulate as soluble fibrin monomers in
the plasma. Platelets become entangled in these fibrin polymers and are exposed to thrombin,
resulting in platelet activation, which drives the coagulation system and leads to
thrombocytopenia. Also, as coagulation inhibitors such as protein C, protein S, and antithrombin
are consumed, control over the coagulation pathway is impaired. Soluble fibrin monomers, fibrin
polymers, and cross-linked fibrin activate plasminogen and convert it to plasmin.

Other factors that can cause the increased conversion of plasminogen to plasmin are the
coagulation factors XIa, XIIa, and kallikrein. Aside from the cytokines mentioned earlier,
cytokines such as TNF-a and IL-6 can participate in fibrinolysis by stimulating the release of
tissue plasminogen activator (TPA). Normally, plasmin acts in a specific area to digest only the
solid fibrin clot to which it is bound. In cases of DIC, the plasmin circulates in plasma and
digests all forms of fibrinogen and fibrin. Thus, increased fibrinolysis will lead to excessive
production of fibrin degradation products such as X, Y, D, E, and D-dimer.

The overall hemorrhagic outcome of DIC is influenced by activation of thrombin,

circulating plasmin, loss of control, and thrombocytopenia. This causes consumption of platelets
as well as coagulation factors.

2.2 CLINICAL SIGNS AND SYMPTOMS

The DIC symptoms can be chronic, acute or fulminant and are commonly masked by the
symptoms of underlying illness. The organ failure symptoms such as the renal function
impairment, adult respiratory distress syndrome, and central nervous system manifestations can
be caused by thrombosis in the microvasculature of major organs. The skin, bone, bone marrow
necrosis can also be observed. The Purpura fulminans can be observed among
meningococcemia, chickenpox, and spirochete infections.

SIGNS & SYMPTOMS OF EXCESSIVE SIGNS & SYMPTOMS OF BLEEDING

BLOOD CLOTTING

● Chest pain and shortness of breath INTERNAL BLEEDING

● Pain, redness, warmth, and swelling ● Presence of blood in urine and stool
in lower leg if blood clots form in deep ● Headaches, double vision and
vein seizures.
● Headache, speech changes, ● Low blood pressure
paralysis, and trouble speaking and
understanding if blood clot are from
blood vessel of brain
● Heart attack, lung and kidney
EXTERNAL BLEEDING
problems
● Bleeding following surgical cuts or
the insertion of a needle at the site of
the wound.
 ● Bleeding from the nose, gums, or
mouth, particularly after brushing your
teeth.
● Purpura
● Petechiae
● Bruising

Signs and Symptoms of DIC associated with sepsis from Escherichia coli:
● Bloody diarrhea
● Abdominal pain/ severe stomach cramps
● Nausea
● Intermittent fever
● Dry mouth
● Excessive hematochezia

3. What other laboratory results are expected with your diagnosis?

The diagnosis is done by combining the clinical presentations with any noted
abnormalities from the laboratory tests. Usually, DIC may be diagnosed by combining the
following tests: platelet count, PT, and assay for D-dimer and fibrinogen. Typically, a prolonged
coagulation test, thrombocytopenia, high levels of FDPs especially D-dimer, and decreased
fibrinogen are usual findings in DIC. Platelet count is decreased due to platelet consumption, PT
is prolonged due to consumption of coagulation factors, and fibrinogen is decreased while
D-dimer is increased due to increased fibrinolysis. Aside from these tests, we can also test for
aPTT and PTT, which will be prolonged because of the consumption of coagulation factors,
thrombin time and reptilase time which are prolonged due to decreased fibrinogen levels,
increased FDPs, and increased presence of soluble fibrin monomers. Peripheral Blood film
exam is also done and schistocytes are seen in half of the cases of DIC.

There are other specialized laboratory assays for diagnosing DIC:

● Protein C, Protein S, and AT activity assays which are used to check the
plasma and AT concentration that will have a value of <50% in DIC patients.
 These Proteins have a decreased value because they are consumed in the DIC
cases.
● Plasminogen, tissue plasminogen activator, and plasminogen activator
inhibitor-1 which is decreased due to increased fibrinolysis.
● Soluble fibrin monomer which is a hemagglutination assay that measures the
fibrin monomer. Supposedly, the fibrin monomer must be insoluble after
polymerization in order to establish a stable clot. DIC cases have a positive result
because during intravascular coagulation, the fibrin monomer may sometimes fail
to polymerize thus making it soluble. Soluble fibrin monomers are detected
because they circulate freely in the plasma.
● Factor assays for II (prothrombin), V, VIII, and X may also be done and usually,
there is a result of <30% for cases of DIC. However, these assays may provide
misleading or unreliable results since factor V and VIII are increased in cases of
inflammation.
● Thrombus precursor protein immunoassay which results to >3.5 ugm/L
● Detection of localized thrombosis markers which are the prothrombin
fragment 1+2, thrombin-antithrombin. In cases of DIC they are elevated and
these are used in the monitoring of DIC therapy.
● Serum FDP which results in >10 ug/mL. However, this assay is no longer
utilized.

4. Give different treatments applicable to your diagnosis.

Individuals with this condition should be treated at hospitals with applicable patient care
and subspecialty expertise such as blood banking, hematology, and surgery. DIC can result
from sepsis, infection, trauma, malignancies, and obstetric complications. Therefore, the
treatment for DIC should mainly focus on addressing the underlying disorder. If the
underlying disorder is addressed, the components of DIC will resolve on its own. For instance, if
the underlying disorder is infection, then proper administration of antibiotics should be done.

DIC can cause thrombosis due to systemic activation of platelets/ coagulation factors
and bleeding due to excessive consumption of platelets/ coagulation factors. The reduction in
platelet count is observed in the patient’s case. Platelet transfusion should be considered to
compensate for thrombocytopenia and resolve bleeding. Also, the patient presents a low
fibrinogen level which is a coagulation factor responsible for clotting. This can be corrected
through the administration of cryoprecipitate/ purified fibrinogen concentrates and fresh frozen
plasma. Cryoprecipitate replaces the fibrinogen while fresh frozen plasma elevates the levels of
other clotting factors. The administration of heparin (anti-coagulant) is also considered when the
patient exhibits thrombosis or pulmonary embolism such as in the case of slow evolving DIC.
However, it should be avoided for patients with bleeding or bleeding risk as this can only worsen
the problem.

III. PERTINENT INFORMATION

History of DIC

The first clinical observations related to DIC dates back to the 19th century. In 1834, M.
Dupuy performed an experiment wherein he injected brain tissue intravenously to animals and
led to widespread clots in their circulation. In 1865, the term “Trousseau syndrome” was
described by Armand Trousseau. The term was used to describe the increased tendency of
patients with advanced malignant disease to develop migratory thrombophlebitis (tumor-induced
chronic DIC). In 1965, Donald McKay published a book about DIC in which he first described
the disease as an “intermediary mechanism”.

Types of DIC

● Acute and uncompensated DIC: Initiating events such as septicemia, obstetric

emergencies, intravascular hemolysis, viremia, burns, acute inflammation, crush injuries,
aortic aneurysms, and cardiac disorders can lead to acute DIC. In this type, various
hemostatic components are decreased. Furthermore, it develops faster and can occur
within hours or days. The most noticeable sign is severe bleeding therefore, it can be
fatal in 25% to 50% of cases. Thus, immediate medical intervention is needed.
● Chronic DIC: Conditions that may complicate the chronic type of DIC are tissue
necrosis, liver and renal disease, chronic inflammation, vascular prostheses, vascular
tumors, adenocarcinoma, and several malignancies. The hemostatic components in this
type may have varying laboratory results such as slightly reduced, normal, and elevated.
Chronic DIC develops slower and there may not be noticable signs and symptoms.

Other diseases that resemble DIC

 ● Severe liver disease: Both DIC and severe liver disease present the same laboratory
findings. However, the decrease in the coagulation factors and platelet count in DIC is
due to “consumption”. In severe liver disease, the laboratory findings are mostly
attributed to “inadequate production”. Moreover, elevated D-dimers in DIC is due to the
increased fibrinolysis while in severe liver disease, it is due to the response to
inflammation. To differentiate them, patient history, signs, and symptoms must be
considered and liver enzyme assays may be performed.
● Dilutional coagulopathy: Dilutional coagulopathy occurs when there is massive
transfusion. Because of this, PT, PTT, and TT are prolonged and platelet count is
decreased. When the fluid that replaces the blood during transfusion does not contain
enough coagulation factors, coagulation factors and platelets will be consumed. Hence,
to differentiate it with DIC, it is important to take note of the patient history.
● Obstetric complications pre-eclampsia, eclampsia, acute fatty liver of pregnancy,
and HELLP ((hemolysis, elevated liver function tests, and low platelets) syndrome:
They resemble the laboratory findings of DIC and may be distinguished through patient
history, signs, and symptoms.
● Thrombotic thrombocytopenic purpura (TTP) and hemolytic-uremic syndrome
(HUS): They may have similar symptoms with DIC. However, PT, PTT, and TT are
normal in TTP and HUS. It is also important to note that TTP and HUS can arise without
any identifiable primary disorder while DIC arises from a primary condition.

IV. REFERENCES:

Boral, B. M., Williams, D. J., & Boral, L. I. (2016). Disseminated intravascular coagulation.
American Journal of Clinical Pathology, 146(6), 670–680.
https://doi.org/10.1093/ajcp/aqw195

Keohane, E. M., Walenga, J. M., & Otto, C. N. (2020). Rodak's hematology: Clinical principles
and applications. Elsevier.

Levi, M.M. (2020). Disseminated Intravascular Coagulation (DIC) Treatment & Management.
Medscape. https://emedicine.medscape.com/article/199627-treatment#d1
Moake, J. (2021). Disseminated Intravascular Coagulation. MSD manuals.
https://www.msdmanuals.com/professional/hematology-and-oncology/coagulation-disord
ers/disseminated-intravascular-coagulation-dic

Papageorgiou, C., Jourdi, G., Adjambri, E., Walborn, A., Patel, P., Fareed, J., Elalamy, I.,
Hoppensteadt, D., & Gerotziafas, G. T. (2018). Disseminated intravascular coagulation:
An update on pathogenesis, diagnosis, and therapeutic strategies. Clinical and Applied
Thrombosis/Hemostasis, 24(9_suppl). https://doi.org/10.1177/1076029618806424

Turgeon, M. L. (2012). Clinical hematology: theory and procedures. 5th edition. Philadelphia,
PA: Wolters Kluwer Health/Lippincott Williams & Wilkins.

U.S. Department of Health and Human Services. (2019, October 8). Disseminated intravascular
coagulation. National Heart Lung and Blood Institute. Retrieved from
https://www.nhlbi.nih.gov/health-topics/disseminated-intravascular-coagulation

U.S. Department of Health and Human Services. (n.d.). Disseminated intravascular coagulation.
National Heart Lung and Blood Institute. Retrieved March 15, 2022, from
https://www.nhlbi.nih.gov/health-topics/disseminated-intravascular-coagulation#:~:text=A
cute%20DIC%20is%20more%20severe,has%20no%20signs%20or%20symptoms.

van der Poll, T., & Levi, M. (2014). A short contemporary history of disseminated intravascular
coagulation. Seminars in Thrombosis and Hemostasis, 40(08), 874–880.
https://doi.org/10.1055/s-0034-1395155

Wada, H., Matsumoto, T., Yamashita, Y. (2014, February 20). Diagnosis and treatment of
disseminated intravascular coagulation (DIC) according to DIC guidelines.
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4267589/
 ANGELES UNIVERSITY FOUNDATION

COLLEGE OF ALLIED MEDICAL PROFESSIONS

DEPARTMENT OF MEDICAL TECHNOLOGY

ANGELES CITY

Presence of Circulating

Anticoagulants
(Case Analysis Written Report)

SUBMITTED BY:

BSMT - 3B

GROUP 2

LEADER:

JENKIN DANIELLE O. SALAC

MEMBERS OF THE GROUP:

BALUYUT, UMI

BARREDO, ALLAINA MARIE

ENCINARES, KIM AIRA

GOMEZ, ANNE LOUISE

MAGAT, EUGENE JOSHUA

QUITO, NOREEN

SANTILLAN, JARAIZA MIN

MARCH 16, 2022

 I. Case 5:

A 24-year-old female with history of COVID-19 infection was referred to the Research Institute
for Tropical Medicine (RITM) after presenting with persistent fever, painful swelling of the lower
extremities, urticaria-like skin lesions and rashes on the face and limbs accompanied with
severe itching, and painful joints. Blood tests were ordered and results showed that her liver
function tests were all normal, blood culture and urine culture turned out negative, and no
abnormalities were associated with her heart and abdomen. A peripheral blood smear was
requested and showed the presence of toxic granulation and schistocytes. Due to the difficulty
of ruling out her diagnosis, a specialist was assigned to her case and requested more laboratory
tests. The following are her other results:

Platelet count 115,000/mm3

LDH 2,000 IU/L

Direct Coombs Test Positive

ANA Positive

APTT Prolonged

PT Normal

II. ANSWERS TO GUIDE QUESTIONS

1. What is the probable diagnosis of the patient? What are the clinical manifestations
unique to the disorder?

The probable diagnosis indicative for our patient is under the presence of
circulation anticoagulants – Systemic Lupus Erythematosus. SLE is an autoimmune
disorder that is characterized by the loss of self-tolerance of autoreactive T and B cells
thus producing pathogenic autoantibodies that are labeled as antinuclear antibodies.
They are directed to destroy self-nuclear antigens. SLE affects a wide variety of organs
and too often, it causes red lesions on the skin.
Etiology of SLE may be implicated by several viruses such as our Epstein-Barr
Virus, cytomegalovirus, retrovirus and such and to this date, there are cases that reports
the manifestation of SLE following CoVid-19 in which the said virus may influence the
production of autoantibodies. Thus, our 24-year old patient may have acquired SLE
through her acquisition of CoVid-19 in the past.
Laboratory findings that are often seen for patients with this disorder are the
following: decreased platelet count, elevated levels of LDH, a positive Direct Coomb’s
Test and ANA. Also, there is a prolonged aPTT. Looking at the table, common features of
SLE match with our patient thus, we conclude that the patient's Diagnosis is indicative of
Systemic Lupus Erythematosus.
https://jmedicalcasereports.biomedcentral.com/articles/10.1186/s13256-020-02582-8
 Systemic Lupus Erythematosus has various clinical manifestations which are
difficult to differentiate from other diseases because some of its manifestations are too
general to use for diagnosis. According to John Hopkins Lupus Center, one of the
distinct characteristics of SLE is having a rash after a sun exposure, which is specifically
called the malar rash or butterfly rash. This is being called as butterfly rash due to the
appearance of butterfly-like redness/rash which covers from the bridge of the nose and
extends upto the cheeks. Other features include photosensitivity rash, and discoid
lesions. All of these may appear due to an exposure to sun of patients with SLE. Lastly,
the least common feature is alopecia, which is characterized by a scar in the scalp,
usually in discoid form and often causes hair loss.

2. Give the pathophysiology of the disease and its clinical signs and symptoms.

PATHOPHYSIOLOGY

Patients with specific illness as well as those with no evident underlying

condition, have pathologically acquired inhibitors of blood coagulation, often known as
circulating anticoagulants. One of the inhibitors that can be found in the presence of
circulating anticoagulant is lupus anticoagulant, which was originally discovered in
individuals with Systemic Lupus Erythematosus that is why it is called lupus. The term
"lupus anticoagulant" is misleading because it is found in persons without lupus and has
been linked to a number of different autoimmune disorders, neoplasias, infections, and
drug administration as well as it can be seen on normal individuals. Autoantibodies
called lupus anticoagulants (LAs) interact with the phospholipid surfaces of the reagents
used in the APTT test and sometimes in the prothrombin time test, causing the test
findings to be delayed or prolonged. Antiphospholipid antibodies are a type of antibody
that binds to phospholipids (APLs). LAs are typically identified when normal coagulation
testing reveals an unexpectedly prolonged APTT and, in some cases, PT can also be
found in the test but Lupus anticoagulant is the result of research or a laboratory
phenomenon. Despite the fact that laboratory tests indicate a problem with hemostasis,
most patients do not bleed and instead tend to be hypercoagulable.

Immunologically, the antiphospholipid antibody lupus anticoagulant is

immunoglobulin G, although it can also be immunoglobulin M or a combination of the
two. It interacts with the phospholipid surfaces of test reagents used in the APTT,
prolonging clotting times, but further studies conclude that a protein cofactor must be
associated with the phospholipid surface. Protein targets recognized by aPL include
b2-glycoprotein-1 (b2GP1), prothrombin, FV, FVII, PC, PS, TFPI, and heparin, and it is
also claimed that antiphospholipid antibodies increase the risk of thrombosis.
Pathophysiologic mechanisms include injury and activation of endothelial cells and
monocytes causing cells in the blood or in contact with the blood to acquire a
procoagulant phenotype; platelet activation; and inhibition of endogenous anticoagulant
and fibrinolytic mechanisms. Furthermore, aPLs can increase platelet aggregation and
encourage leukocytes to produce tissue factors.

SIGNS AND SYMPTOMS

 According to the National Heart, Lung, and Blood Institute, the signs and
symptoms associated with the presence of circulating anticoagulants such as lupus
anticoagulants in the body are linked to abnormal blood clotting, which can be
associated with the disorder Systemic Lupus Erythematosus. These blood clots may
travel to different parts of the body such as the arteries or veins of the brain, heart,
kidneys, lungs, and limbs. Due to this, the patients may develop thrombocytopenia which
can be associated with internal or external bleeding. Aside from these, people with this
condition may also develop rashes on the wrists and knees. Among women, the
occurrence of multiple miscarriages may also indicate the presence of circulating
anticoagulants.
The formation of blood clots in the body may manifest differently depending on
the area or location where it developed. Signs and symptoms associated with the
formation of blood clots in the arms or legs may lead to swelling, redness or
discoloration, and pain or numbness in these areas. The presence of blood clots in
areas such as the heart and lungs may lead to chest pain, heart valve problems,
excessive sweating, breathing difficulties, fatigue, and dizziness. Blood clots in the
stomach or kidneys may cause stomach pain, thigh pain, nausea, diarrhea or presence
of blood in the stool, and fever. While blood clots in the brain may lead to speech
changes, memory loss, and chronic headaches.
3. Give hematological and other laboratory tests for the diagnosis of the disorder.

Hematological and laboratory tests play an integral role in the diagnosis of

Circulating anticoagulant, specifically Lupus anticoagulant to enhance the sensitivity and
specificity in the determination of the said disease. There are eight of these tests which
are usually used for the diagnostic accuracy in testing LAC and two of these tests are
the most commonly used which are the Dilute Russell Viper Venom test and Silica-based
PTT. Nonetheless, to further ensure an efficient diagnosis- other laboratory assays are
also performed.

3.1 Dilute Russell Viper Venom Test (dRVVT)

This test is considered to be the most regarded LAC detection method which
actually activates factor X. It is usually paralleled with LAC-sensitive partial
thromboplastin time assay (PTT) which activates factor XII. Russell viper venom is used
in the dRVVT, which is mixed with a limited amount of diluted rabbit brain phospholipid.
The venom of the snake stimulates coagulation factor X, causing a fibrin clot to develop.
By interfering with the assembly of the prothrombinase complex, LA prolongs dRVVT,
which can be reversed by adding extra phospholipid. Concomitant anticoagulant
medication with heparin, warfarin, or direct thrombin inhibitors might cause test findings
to be falsely abnormal.

3.2 Silica-based PTT/ Silica Clot Time (SCT)

SCT is an integrated test system which has two components similar to the
dRVVT. To activate FXII in the intrinsic route, the reagent contains a colloidal silica
suspension. Patient plasma is introduced to a second reagent containing an enhanced
amount of phospholipid in the confirmatory phase of the experiment. The extra
phospholipid neutralizes the antibody by increasing the surface area available to the
intrinsic route and prothrombinase complexes, reducing clotting time. In certain
 investigations, this assay was found to be the most sensitive for detecting LA in patients
who met the clinical criteria.

3.3 Dilute Thromboplastin Time/ Tissue Thromboplastin Inhibitor

Activates factor VII
The Dilute Thromboplastin Inhibition test is based on the idea that diluting the
Thromboplastin reagent in a PT-based assay makes it more sensitive to a Lupus
Anticoagulant.

3.4 Hexagonal Phase Phospholipid Neutralization (STACLOT-LA)

A Staclot LA is considered as a two-part assay which is based on aPTT and is
performed with and without a source of phospholipid, HPE, or using a reagent which is
sensitiving for lupus anticoagulants. The HPE will play a role in neutralizing the lupus
anticoagulants that are present on the patient’s plasma which will result in a decreased
clotting time, which will then be compared with the same sample without the addition of
HPE.

3.5 Kaolin Clotting time (KCT)

Kaolin Clotting time has been regarded as the most sensitive test to detect
circulating anticoagulants. KCT is an aPTT test without the addition of phospholipid. The
principle behind this test is that it relies on plasma lipids and residual cell membrane
fragments to provide a surface of phospholipid for the coagulation reaction.
3.6 Platelet Neutralization Procedure
This test plays a role as a confirmatory test or exclusion test for the detection of
lupus anticoagulant inhibitors whenever it is used together with other coagulation tests. It
aids in the differentiation of inhibitors of specific factors such as the factor VIII inhibitor
from lupus anticoagulant inhibitors.

3.7 Taipan Venom Time

This test utilizes a reagent which is isolated from the venom of Taipan snakes,
specifically the Oxyuranus scutellatus. The venom consists of one prothrombin activator
which will aid in the conversion of prothrombin to thrombin through the presence of
calcium and phospholipid.

3.8 Textarin:Ecarin Ratio

This test is based on the two differential dependence of two snake venoms:
Pseudonaja textilis and Echis carinatus. The venom from Psudonaja textilis plays a role
in activating prothrombin which requires Calcium, Factor V and phospholipid. On the
other hand, venom from Echis carinatus has a role in activating prothrombin for the
formation of Meizothrombin without the presence of phospholipid. If there is a presence
of lupus anticoagulant, then the test is prolonged.

4. What are the abnormal laboratory findings? Explain why such tests are abnormal.

The abnormal laboratory findings in this case are:

● ANA test positive
● Prolonged aPTT
● Elevated LDH levels
 ● Positive Direct Coombs Test
● Decreased platelet count

Test Patient Result

Antinuclear Antibody Test Positive

The laboratory results showed a positive result in the ANA (Antinuclear antibody) test
which may indicate that the patient is suffering from an autoimmune disease called
Systemic Lupus Erythematosus (SLE). SLE is a type of systemic rheumatic disease
which is characterized by the presence of autoantibodies that are directed against
self-nuclear antigens, known as antinuclear antibodies. These autoantibodies are
considered as a hallmark of systemic rheumatic diseases, and may be directed to the
components of the surface, cytoplasm, nuclear envelope, or nucleus of the cell. Thus, a
positive ANA test in conjunction with the other laboratory tests indicates SLE. This
disease is also associated with the presence of lupus anticoagulants in the circulation
which supports the diagnosis, presence of circulating anticoagulants.

Test Patient Result

aPTT Prolonged

Acquired inhibitors of clotting proteins, also known as circulating anticoagulants,

inactivate or inhibit the usual procoagulant activity of coagulation factors. Inhibitors are
frequently characterized as specific, those directed against a coagulation factor, or
nonspecific, those directed against a complex of factors, such as the Lupus
Anticoagulants, an IgG immunoglobulin directed against a number of
phospholipid-protein complexes. LA is the most common coagulation inhibitor found in
SLE patients, and also the most common cause of prolonged aPTT. Lupus anticoagulant
or antiphospholipid antibody influences coagulation protein reactions, these inhibitors
are antibodies directed to epitopes of proteins bound to phospholipids, which prolongs
phospholipid-dependent clotting assays in vitro such as seen in aPTT.

Test Patient Result

Direct Coomb’s Test Positive

Direct Coomb’s Test, also known as antiglobulin testing, is an immunological procedure

used to detect the presence of antibodies that are stuck to the surface of RBCs. A
positive Coomb’s Test in this case denotes the presence of antibodies on the RBC’s
surface which is evident in cases of SLE wherein a large number of autoantibodies is
present. The specific antibody of interest in this test is the circulating lupus
anticoagulant, which is an antiphospholipid antibody that sticks to the surface or the
membrane of RBCs thus producing a positive Direct Coomb’s Test.
 Test Patient Result Reference value

Lactate Dehydrogenase 2,000 IU/L 105 to 333 IU/L

The highly elevated levels of lactate dehydrogenase (LDH) is due to the increase in LDH
isoenzymes present in the circulation. LDH is found in most of the body tissue, including
the heart, blood, kidneys, lungs, and brain. When these tissues are damaged, such as in
cases of multi-organ tissue damage, the LDH is released from within these tissues. This
causes an elevation in the LDH levels, which is evident in this case. Multi-organ tissue
damage caused by the presence of autoantibodies is a common occurrence in cases of
SLE. However, in order to determine the exact source of damage, a further
determination of the specific isoenzymes elevated is required.

Test Patient Result Reference Value

Platelet Count 115,000/mm3 150,000-450,000/mm3

Thrombocytopenia can be caused by many possible reasons, however in this case it is

most likely due to the presence of autoantibodies directed against the platelet surface
glycoproteins. The circulating lupus anticoagulants or antiphospholipid antibodies
causes an increased risk of blood clotting thus inducing unnecessary blood coagulation
which results in the decreased number of platelets.

III. PERTINENT INFORMATION

Lupus anticoagulants (LACs) are immunoglobulins that interfere with in vitro

phospholipid-dependent coagulation assays. Feinstein and Rapaport coined the phrase in 1972,
but the first observations of the phenomena were made more than 20 years earlier. LACs were
first identified in patients with systemic lupus erythematosus (SLE), but they are now known to
occur often in patients with different illnesses, as well as in people who have no obvious
underlying disease. Although LACs and anti-cardiolipin antibodies (ACAs) are currently
regarded as different entities, they are both members of a wide set of immunoglobulins that bind
to phospholipid-bound proteins such as 2-glycoprotein-I (2GPI) and prothrombin.

IV. CONCLUSION

The 24-year-old female patient has a condition wherein circulating anticoagulants such
as lupus anticoagulants, which is an antiphospholipid antibody, can be found in her body. This is
a condition that results in abnormal blood clotting within her blood vessels. The presence of
these anticoagulants can also be associated with conditions such as Systemic Lupus
Erythematosus and Antiphospholipid Syndrome. SLE is a condition characterized by the
presence of antinuclear antibodies, while APS is characterized by the presence of
antiphospholipid antibodies. However, both disorders can lead to the immune system attacking
its own tissues.
 The diagnosis was confirmed through the signs and symptoms exhibited by the patient,
as well as their laboratory results. Signs and symptoms observed were persistent fever, swelling
of the lower extremities, skin lesions and rashes, severe itching, and painful joints, all of which
are some of the common manifestations observed in the presence of circulating anticoagulants.
Aside from these, the laboratory findings such as Positive for Antinuclear Antibody Test,
Prolonged Activated Partial Thromboplastin Time, Elevated Lactate Dehydrogenase Levels,
Positive Direct Coombs Test, and a Decreased Platelet count, can all be indicative of the said
condition.

As mentioned, other laboratory tests that can also be utilized to diagnose this condition
includes Dilute Russel Viper Venom Test, Silica Clot Time, Tissue Thromboplastin Inhibitor,
Hexagonal Phase Phospholipid Neutralization, Kaolin Clotting time, Platelet Neutralization
Procedure, Taipan Venom Time, and Textarin: Ecarin Ratio. Among these tests, the Platelet
Neutralization Procedure may be used as a confirmatory test to detect lupus anticoagulant
inhibitors when other coagulation tests are performed.

REFERENCES

Antiphospholipid Antibody Syndrome | NHLBI, NIH. (n.d.). National Heart, Lung, and Blood

Institute. Retrieved March 13, 2022, from

https://www.nhlbi.nih.gov/health-topics/antiphospholipid-antibody-syndrome

Bartels, C. M. (2021, October 17). Which autoantibody tests are used in the diagnosis of

systemic lupus erythematosus (SLE)? Latest Medical News, Clinical Trials, Guidelines -

Today on Medscape. Retrieved March 15, 2022, from

https://www.medscape.com/answers/332244-19822/which-autoantibody-tests-are-used-i

n-the-diagnosis-of-systemic-lupus-erythematosus-sle

Cojocaru, M., Cojocaru, I. M., Silosi, I., & Vrabie, C. D. (2011). Manifestations of systemic lupus

erythematosus. Maedica, 6(4), 330–336. Retrieved from

https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3391953/#:~:text=The%20patients%20wit

h%20SLE%20may,or%20recurrent%20active%20SLE%20flares

Hexagonal phase phospholipid neutralization. (n.d.). Retrieved March 13, 2022, from

https://www.laboratoryalliance.com/assets/Uploads/testpdfs/Hexagonal_Phase_Phospho

lipid_Neutralization.pdf
Inoue, T. (1986). Serum lactate dehydrogenase and its isozymes in lupus nephritis. Archives of

Internal Medicine, 146(3), 548–552. https://doi.org/10.1001/archinte.146.3.548

John Hopkins Lupus Center. (N/A). Lupus-specific Skin Disease and Skin Problems. Retrieved

from

https://www.hopkinslupus.org/lupus-info/lupus-affects-body/skin-lupus/#:~:text=Malar%2

0Rash,the%20bridge%20of%20the%20nose

Mckenzie, S. B., Landis-Piwowar, K., & Joanne Lynne Williams. (2020). Clinical laboratory

hematology. Pearson.

MD, MPH, D. E. (2022, January 19). Patient education: Antiphospholipid syndrome (Beyond the

Basics). UpToDate. Retrieved March 15, 2022, from

https://www.uptodate.com/contents/antiphospholipid-syndrome-beyond-the-basics#H4

Mls, K. E. P., Smith, L., & (Ascp)Hcm, W. J. P. M. (2015). Rodak’s Hematology: Clinical

Principles and Applications (5th ed.). Saunders.

Richey, M. (n.d.). Understanding lupus lab tests and results: Rheumatology. Hospital for Special

Surgery. Retrieved March 15, 2022, from

https://www.hss.edu/conditions_understanding-laboratory-tests-and-results-for-systemic-

lupus-erythematosus.asp

Polonsky, M. (2018, September 17). Lupus Anticoagulants. Healthline. Retrieved March 13,

2022, from https://www.healthline.com/health/lupus-anticoagulant#symptoms

Practical-haemostasis.com. Kaolin Clotting Time [KCT]. (n.d.). Retrieved March 13, 2022, from

https://practical-haemostasis.com/Thromobophilia/APS/kct.html

Practical-haemostasis.com. Taipan Venom Time. (n.d.). Retrieved March 13, 2022, from

https://practical-haemostasis.com/Thromobophilia/APS/taipan_venom_time.html#:~:text
 =The%20Taipan%20venom%20time%20employs,test%20for%20a%20Lupus%20Antico

agulant.

Practical-haemostasis.com. Textarin:Ecarin Ratio. (n.d.). Retrieved March 13, 2022, from

https://practical-haemostasis.com/Thromobophilia/APS/textarin_ecarin_ratio.html#:~:text

=The%20two%20venom%20times%20are,presence%20of%20a%20Lupus%20Anticoag

ulant.

Test ID : PNP. PNP - Overview: Platelet Neutralization Procedure, Plasma. (n.d.). Retrieved

March 13, 2022, from

https://www.mayocliniclabs.com/test-catalog/overview/8866#:~:text=When%20a%20prol

onged%20activated%20partial,a%20lupus%20anticoagulant%20(LAC).

Turgeon, M. L. (2022). Clinical Hematology 5Ed: Theory & Procedures (31303rd ed.). Lww.

Win, N., Islam, S. I., Peterkin, M. A., &amp; Walker, I. D. (1997). Positive direct antiglobulin test

due to antiphospholipid antibodies in normal healthy blood donors. Vox Sanguinis, 72(3),

182–184. https://doi.org/10.1046/j.1423-0410.1997.7230182.x

FOR THE POWERPOINT PRESENTATION

Q1
Unique Clinical Manifestation
Malar/Butterfly rash

Photosensitivity rash

Discoid Lesions
Alopecia

Q2
PATHOPHYSIOLOGY
Lupus Anticoagulant
- Originally found on patients with SLE
- Found on persons;
- Without lupus
- Linked with various autoimmune disorders
- Neoplasias
- Infections
- Drug administration
- Normal individuals

Antiphospholipid Antibody “Lupus anticoagualant”

- Phospholipid surfaces of the reagents used in the APTT test (sometimes in PT)
- Identified when there is unexpected prolonged APTT (as well as PT)

In immunology: Found in IgM or IgG or can be in both

Further studies: Protein factor is required to associate with phospholipid surface

Protein recognized by APLs

- b2-glycoprotein-1 (b2GP1)
 - prothrombin
- FV
- FVII
- PC
- PS
- TFPI
- Heparin

Pathophysiologic Mechanism

- Injury and activation of endothelial cells and monocytes

- Platelet activation
- Inhibition of endogenous anticoagulant and fibrinolytic mechanisms
- Can increase platelet aggregation

SIGNS AND SYMPTOMS

➔ Blood Clotting
● Arteries or veins of the brain
● Heart
● Kidneys
● Lungs
● Limbs
➔ Thrombocytopenia
● Internal Bleeding
● External Bleeding
➔ Rashes on the wrists and knees
➔ Multiple Miscarriages

BLOOD CLOTTING

STOMACH OR
ARMS & LEGS HEART & LUNGS BRAIN
KIDNEYS

Chest pain
Stomach pain
Swelling Heart valve problems
Thigh pain Speech changes
Redness or Excessive sweating
Nausea Memory loss
discoloration
Breathing difficulties
Diarrhea or blood in Chronic headaches
Pain or numbness
stool
Fatigue
Fever
Dizziness

PICTURES:
Q3

LABORATORY TESTS

Dilute Russell Viper Venom Test (dRVVT) Most reliable detection method; Activates
Factor X

3.2 Silica-based PTT/ Silica Clot Time Activates Factor XII

(SCT)

Dilute Thromboplastin Time/ Tissue Activates Factor VII

Thromboplastin Inhibitor

Hexagonal Phase Phospholipid Neutralizes lupus anticoagulant

Neutralization (STACLOT-LA)

Kaolin Clotting time (KCT) Without the addition of phospholipid

Platelet Neutralization Procedure Differentiation of inhibitors

Taipan Venom Time Isolated from the venom of Taipan snake

Textarin:Ecarin Ratio Two venom snakes: Pseudonaja textilis and

Echis carinatus

The dilute Russell viper venom time (DRVVT) assay, regarded as the most reliable LAC detection method, activates factor X.
DRVVT is typically paralleled by an LAC-sensitive partial thromboplastin time assay (PTT) that acti- vates factor XII. The dilute
thromboplastin time test (DTT) activates factor VII.

Q4
 Test Patient Result

Antinuclear Antibody Test Positive

aPTT Prolonged

Direct Coomb’s Test Positive

Test Patient Result Reference value

Lactate Dehydrogenase 2,000 IU/L 105 to 333 IU/L

Platelet Count 115,000/mm3 150,000-450,000/mm3

 Angeles University Foundation
College of Allied Medical
Professions
Department of Medical Technology

Case Analysis
Afibrinogenemia

GROUP 5 | BSMT – 3E
Submitted by:
De Jesus, Nina
Dungca, Louise
Pineda, Kathleen
Salas, Carmela
Sampang, Nina
Timuat, Christine
Villarosa, Niña

1
I. Case Background

A 19-year-old female patient was admitted to the emergency department with acute
abdominal pain and nausea. Several years ago, she suffered from a massive bleeding and led
to a diagnosis of hemoperitoneum complicating an ovarian cyst. She was treated monthly with
fibrinogen concentrates, tranexamic acid (during menstrual periods), and oral contraception. A
few hours later, she exhibits tachycardia, increased abdominal pain, and hypotension. Her
hemoglobin level drops from 13.5 g/dL to 5g/dL. Result of coagulation test shows prolonged
PT, aPTT, TT, and bleeding time.

II. Answers to the Guide Questions

1. What is the probable diagnosis of the patient? Why?

Given the clinical manifestations and laboratory test results, the patient is most likely
suffering from afibrinogenemia, specifically, congenital afibrinogenemia. The diagnosis is
evidenced by the presence of acute abdominal pain and nausea that may be the consequence of
gastrointestinal hemorrhage, as well as the abnormal results for the performed coagulation tests,
specifically prothrombin time (PT), activated partial thromboplastin time (aPTT), and thrombin
time (TT). It is mentioned in the case that the patient suffered several years ago from a massive
bleeding, which is also seen in patients with congenital afibrinogenemia. The patient may also be
experiencing menorrhagia since fibrinogen concentrates, tranexamic acid, and oral contraception
were prescribed to treat her monthly. The prolonged results in the coagulation tests show that
there is a defect in secondary hemostasis as it is characterized by the total lack of fibrinogen or
coagulation factor I in the plasma, wherein a fibrin clot does not appear when the patient’s plasma
is tested. In a third of patients with afibrinogenemia, bleeding time is prolonged due to the inability
of platelets to aggregate without the presence of fibrinogen; however, in some afibrinogenemic
patients, sparse hemorrhagic episodes are observed due to the presence of functional von
Willebrand factor (VWF). This would then allow for normal platelet function, namely adhesion,
aggregation, and loose thrombi formation (Greer et al., 2019). Additionally, fibrinogen activity can
be measured by the Clauss-clot based method or by immunoassays and nephelometric methods
(Keohane et al., 2020).

The disorder exhibits an autosomal recessive pattern of inheritance, wherein many of the
reported cases are due to consanguineous relationships between two carriers, producing an

2
offspring with the disorder. It is manifested at birth and thus diagnosed due to the presence of
severe and uncontrollable bleeding from the umbilical cord (NORD, 2018).

A similar case reported by Malaquin et al. (2016) showed that sudden onset of bleeding
either spontaneously or after surgery or trauma as well as thrombo-embolic complications are
reported in patients with congenital afibrinogenemia. The report describes a 19-year-old female
patient with congenital afibrinogenemia who developed similar symptoms as she was admitted to
the hospital, namely abdominal pain and nausea, and was also treated with similar medications.
The same diagnosis of hemoperitoneum which complicated the rupture of an ovarian cyst was
given to the patient in the case report. Low levels of hemoglobin were observed as well, along
with later signs of increased heart rate, abdominal pain, and low blood pressure after the patient
was admitted. In the case report by Malaquin et al., the patient was suffering from spontaneous
hepatic hematoma with numerous splenic ruptures. The clinical manifestations which are not
attributed to congenital afibrinogenemia in the case, such as tachycardia and hypotension, may
be due to the aforementioned severe bleeding episode. This occurrence does not frequently occur
among individuals with congenital afibrinogenemia, and may happen after a traumatic event or
as a result of the progression of hepatic disease.

Individuals who experience severe spontaneous bleeding exhibit tachycardia, or an

increase in heart rate, as the body compensates for the lack of blood by preserving oxygen
delivery to 2 organs: the brain and the heart. Abdominal pain may have been due to the
exacerbation of an intrahepatic hemorrhage, similar to the aforementioned case report.
Hypotension is also one of the manifestations of the loss of a large volume of blood, which
suggests that the patient may also be experiencing hemorrhagic shock. The decrease in blood
pressure occurs due to a severe decrease in blood volume; the decrease in blood volume is also
aligned with the decrease in hemoglobin level seen in the patient (Hooper & Armstrong, 2013).

2. Give the etiology and pathophysiology of the disorder. Indicate how it is related
to hemostasis as well.

Congenital Afibrinogenemia is a rare homozygous autosomal recessive bleeding disorder

with the gene located on chromosome 4. It is characterized by absence of fibrinogen that is less
than 10 mg/dL of fibrinogen in the plasma (Ciesla, 2018 p.275). Mutations in one of three genes,
known as the fibrinogen alpha-chain (FGA), beta-chain (FGB), or gamma-chain (FGG), cause the
absence of fibrinogen in the blood. Some mutations make it difficult to produce fibrinogen by
preventing the DNA from being read correctly. Other mutations impact the fibrinogen protein itself,
affecting its production, secretion, or fusion of the protein's component subunits. The end outcome
is the same in every case: fibrinogen is not present in the blood. (NORD, 2018).

3
 The conversion of fibrinogen to fibrin is one of the most important steps in the coagulation
cascade, its absence inhibits clot formation and can result in episodes of prolonged bleeding.
(NORD, 2018) It also serves as a support for platelet aggregation and a substrate for fibrin clot
formation in both primary and secondary hemostasis. Due to the absence of fibrinogen,
Secondary hemostasis cannot function well as it is defined as the synthesis of fibrin in the
coagulation cascade. (Van Herrewegen et al., 2012) In addition, fibrinogen is involved in a number
of important biological processes, including angiogenesis, tissue healing, and the immunological
response. (Casini et al., 2020). Specifically, fibrinogen is essential in the inflammatory process as
it interacts with leukocytes through the surface receptors, serves as a ligand for intercellular
adhesion molecule-1 and facilitates cell-to-cell communication (Malaquin et al., 2016)

Some features of this disorder are poor wound healing and spontaneous abortion. Infants
with this disorder exhibit bleeding from the umbilical stump, along with hematomas or intracranial
hemorrhage due to birth trauma. This disorder is similar to hemophilia wherein spontaneous
hemorrhage, and excessive post-traumatic and postsurgical bleeding is observed. This can result
in hemarthroses, excessive ecchymoses, intracranial hemorrhage, and gastrointestinal bleeding.
(Ciesla, 2018 p.275) (Rodak et al., p. 727) An example of spontaneous hepatic hematoma is
splenic rupture, where patients may experience hypotension or shock, acute abdomen, and
abdominal pain. (Malaquin et al., 2016).

3. What are the clinical signs and symptoms unique to the disorder that led you to
that diagnosis?

Individuals who are affected by afibrinogenemia may be prone to severe bleeding episodes,
especially during infancy and childhood. Bleeding problems may vary from mild to severe in
patients with this disease. In many cases, bleeding from the umbilical cord is an early sign of
concern. Other signs and symptoms that a patient with this disease may experience include the
difficulty in stopping nose bleeds or epistaxis, mucus membranes and joint bleeding, bruising
easily, gastrointestinal bleeding, postpartum hemorrhage and menorrhagia, which is prolonged
menstrual bleeding, excessive bleeding after an accident or surgery, spontaneous splenic rupture,
miscarriage, and even intracrtanial hemorrhage, which is rarely observed in some patients.
(Genetic and Rare Diseases Information Center, 2021, February).

According to Malaquin, et al. (2016) in their study about Congenital Afibrinogenemia, the
most prevalent clinical symptoms include mucocutaneous, soft-tissue, joint, and genito-urinary
bleeding that occurs spontaneously, as well as traumatic or surgical bleeding, while for
thrombosis, poor wound healing, and splenic rupture are uncommon symptoms. In addition,
evidence of gastrointestinal bleeding when a patient has acute abdominal pain and nausea, as

4
well as massive bleeding, which was suffered by the patient several years ago are some of the
clinical signs and symptoms that led to the diagnosis of congenital afibrinogenemia. (Greer et al.,
2019).

4. What treatment is given to patients with this disorder?

Patients with afibrinogenemia have a high rate of hemorrhagic symptoms, and bleeding is
the leading cause of mortality in around one-third of them. This disorder has no known prevention
or cure. Fibrinogen concentrates, cryoprecipitate, and fresh frozen plasma (FFP) are all options
for treating extensive hemorrhages in individuals with this condition, but fibrinogen concentrates
are the most prominent. They are virally quiescent and can be administered in small amounts with
a lower risk of allergic reaction. Some patients, however, acquire antibodies to fibrinogen,
rendering this treatment ineffective (Keohane et al., 2020, p. 753). When fibrinogen concentrates
are unavailable in an emergency, fresh frozen plasma and cryoprecipitate are administered.

FFP involves thawing and ABO compatibility blood matching prior administration and has
a relatively low fibrinogen concentration (varying amounts of other coagulation components),
which inhibits precise dosing. Substitution of fibrinogen with FFP needs a substantial transfusion
amount that leads to volume excess. Moreover, FFP frequently does not undergo pathogen
inactivation, causing a threat of pathogen infection, and carries antigens and antibodies that may
cause severe immunological or allergic reactivity, such as the risk of transfusion-related acute
lung injury. Cryoprecipitate, on the other hand, is a human plasma by-product that needs to be
cross matched and thawed before use, and it has a more prominent and considerably less
variable fibrinogen content than FFP. Cryoprecipitate is derived from several donors and, similar
to FFP, is associated with safety risks.

Due to the constraints of FFP and cryoprecipitate, human fibrinogen concentrate (HFC) has
emerged as the primary alternative for fibrinogen substitution in situations of congenital fibrinogen
deficiency and is the ideal replacement treatment in individuals with afibrinogenemia. Faster
preparation (wherein there is no thawing necessary and no requirement for blood matching), more
rapid administration (such as low infusion amount), and improved purity are advantages of HFC
over FFP and cryoprecipitate. Furthermore, HFC has a more constant fibrinogen concentration
that can be reliably detected, enabling regulated dosing. With no threat of volume overload and a
lower risk of pathogen transmission, HFC is a safer alternative to FFP and cryoprecipitate
(Lissitchkov, et. al., 2020).

In addition to fibrinogen concentrates, the patient was said to be treated with tranexamic
acid (during menstrual periods), and oral contraception monthly. Tranexamic acid suppresses
fibrinolysis by adhering to plasmin and could manage to treat mucosal bleeding. It can be used

5
orally, intravenously, or by using mouthwash. It may, however, increase the likelihood of
thrombosis and should be used with caution in individuals who have had thrombosis previously.
It should also be avoided by pregnant women, individuals who underwent surgery or are
immobilized, and those who have hematuria. In individuals with end-stage renal disease,
tranexamic acid should be provided at a low dosage. On the other hand, hemoperitoneum caused
by follicular rupture appears to be preventable with oral contraceptives. Regardless, it may
increase the risk of thrombosis (Tziomalos et al., 2009).

The Food and Drug Administration (FDA) has authorized some orphan drugs for the
treatment of this disorder. FIBRYGA is a human fibrinogen concentrate that is used to help
alleviate bleeding episodes in adults and adolescents with congenital fibrinogen deficiency, such
as afibrinogenemia and hypofibrinogenemia. Utilizing two virus inactivation or elimination phases
ensures high purity and pathogen safety. Human fibrinogen concentration that has been
pasteurized could potentially be used. An example of this would be RiaSTAP® and
Haemocomplettan® P.

IV. Reference/s:

Casini, A., Neerman-Arbez, M., & Moerloose, P. de. (2020, December 26). Heterogeneity of
congenital afibrinogenemia, from epidemiology to clinical consequences and management.
Blood Reviews. Retrieved March 14, 2022, from
https://www.sciencedirect.com/science/article/pii/S0268960X20301430

Ciesla, B. [V. N. V. (2018). Hematology in practice. F.A. Davis Company.

Congenital afibrinogenemia. NORD (National Organization for Rare Disorders). (2018). Retrieved
from https://rarediseases.org/rare-diseases/afibrinogenemia-congenital/

Genetic and Rare Diseases Information Center (2021). Afibrinogenemia. Retrieved from
https://rarediseases.info.nih.gov/diseases/5761/afibrinogenemia.

Greer, J. P., Arber, D. A., Glader, B., List, A. F., Means, R. T., Paraskevas, F., & Rodgers, G. M.
(2019). Wintrobe's clinical hematology (13th ed.). Wolters Kluwer.

Hillyer C., Silberstein L., Ness P., Anderson., & Roback J., (2006) Blood Banking and Transfusion
Medicine Basic Principles and Practice (2nd ed.)

Hooper, N. (2021, July 13). Hemorrhagic shock. StatPearls [Internet]. Retrieved March 15, 2022,
from https://www.ncbi.nlm.nih.gov/books/NBK470382/

Keohane, E. M., Walenga, J. M., & Otto, C. N. (2020). Rodak's hematology: Clinical principles
and applications (6th ed.). Elsevier.

6
Lissitchkov, T., et al. (2020). Fibrinogen concentrate for treatment of bleeding and surgical
prophylaxis in congenital fibrinogen deficiency patients. Journal of thrombosis and
haemostasis : JTH. Retrieved from
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7187153/

Malaquin, S., Rebibo, L., Chivot, C., Badoux, L., Mahjoub, Y., & Dupont, H. (2016, July).
Congenital afibrinogenemia: A case report of a spontaneous hepatic hematoma. Medicine.
Retrieved from https://www.ncbi.nlm.nih.gov/pmc/articles/

Rodak B. F., Fritsma G. A., & Keohane E. M. Hematology: Clinical Principles and Applications
(4th ed.). Elsevier

Tziomalos, K., Vakalopoulou, S., Perifanis, V., & Garipidou, V. (2009). Treatment of congenital
fibrinogen deficiency: Overview and recent findings. Vascular health and risk management.
Retrieved from https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2762433/

Van Herrewegen, F., Meijers, J. C. M., Peters, M., & van Ommen, C. H. (2012, February). Clinical
practice: The bleeding child. part II: Disorders of secondary hemostasis and fibrinolysis.
European journal of pediatrics. Retrieved March 16, 2022, from
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3258398/

7
 CASE 1: HEMOPHILIA A o Hemophilia A is more common in males. It
• Which part of the secondary hemostasis is mainly is rare in females
affected? o Carrier female: only has 1 abnormal X
o Coagulation chromosome
• Factor that lacks in Hemophilia A o Males cannot be carriers. They are
o Factor VIII (Antihemophilic factor) automatically affected if they have the
• Alternative name: Classic Hemophilia abnormal X chromosome
• Common clinical manifestations • Can Hemophilia A be acquired?
o Prolonged bleeding o When it becomes an autoimmune
o Bruising disorder - There is autoimmune antibody
• What are the key points that made you realize that attacks the Factor VIII
that it is hemophilia A (3 points) o Diagnosis: Bethesda Assay – quantitative
o History test used to determine the concentration
o Reduced Factor VIII levels of the inhibitor in the patient
o Triggering event → Bleeding after the cut sample
• Is the disorder described as mild, moderate or • Inherited Hemophilia A/ Classic Hemophilia A Lab
severe? results:
o Moderate because of 4% Factor VIII o PT – normal (it assesses extrinsic and
activity or because of triggering event common pathway but Factor VIII is
• 3 levels of severity → factor VIII activity and intrinsic)
clinical manifestation o PTT – prolonged
o Severe: o BT – normal (it affects primary
▪ <1% factor VIII activity hemostasis)
▪ Occurs in neonatal period o Platelet aggregation – normal
▪ Clinical manifestation: Neonates o Fibrinogen – normal
experience bleeding, easy o vWF – normal
bruising, bleeding from other o Thrombin time – normal
parts of the body, hematuria, o Mixing studies:
umbilical stump neurologic ▪ Aged serum: NC
symptoms (intracranial bleeding, ▪ Adsorbed plasma: C
memory loss, coma) ▪ Aged plasma: NC
o Moderate ▪ Fresh plasma: C
▪ 1-5% factor VIII activity • Treatment:
▪ Clinical manifestation: Non-stop o Is there a standard cure? NO cure. There
bleeding after cut or triggering is only management
event o Replacement therapy – alternative blood
▪ Commonly occurs in early products for Factor VIII concentrate are
childhood cryoprecipitate (fibrinogen, factor VIII,
o Mild XIII, and vWF) and fresh frozen plasma
▪ 6-20% factor VIII activity (has all coagulation factors)
▪ Manifests in triggering event or o Drugs: Anti-fibrinolytics
trauma, surgery or dental ▪ Trenaxemic acid
extraction ▪ Epsilon-aminocaproic acid
▪ Do not manifest immediately • Hemophilia B is also known as Christmas Disease
• In this case, it is inherited: o Factor IX deficiency
o Looking at the pedigree of inheritance o X-linked recessive
pattern, the mother is a carrier • Hemophilia C aka Rosenthal Syndrome
o X-linked recessive pattern → 1 X gene only o Factor XI deficiency
in males but 2 X genes in females o Autosomal dominant

MARIANO, D. 1
 CASE 2: von Willebrand Disease Type II M to as Autosomal
• vWF in adhesion Hemophilia (?)
• Protects factor VIII from proteolysis o Type 3: Quantitative (most severe;
• Helps adhere collagen complete absence)
• 1. EC: Weibel-palade bodies ▪ Autosomal recessive (all are
• 2. Megakaryocytes: Alpha granules in platelets dominant except for this)
• Factor VIII • LAB FINDINGS:
• Platelet GP Ib/IX/V, collagen o Types 1 & 3 = Similar findings
• Platelet GP IIb/IIIa o Type 1 < 3 (Extent)
• Is the vWF acquired or inherited? o BT – prolonged
o Both o Plt aggregation w/ ristocetin – Decreased
▪ 2b: increased ristocetin
• Autosomal dominant
o Plt count – normal EXCEPT 2B (moderate
• Genetic defect is mutation in the gene located in
or mild thrombocytopenia)
chromosome 12
o PT – Normal
o Defe
o aPTT – Prolonged
• What hemostatic step is impaired?
• 3 tests:
o Platelet adhesion
o vWF antigen assay – decreased (Types 1
• Prolonged bleeding or mucocutaneous bleeding
and 3)
• Mutation – VWF Defects – Impaired platelet
o vWF activity – decreased
adhesion – Bleeding
o Factor VIII assay – decreased (Type 1, 3
• Aside from vWF abnormalities, what other and 2N)
abnormalities may cause that bleeding
• Multimeric analysis – to differentiate subtypes 2A
o Factor VIII decreases as well
and 2B
• Bleeding is caused by: vWF and Factor VIII
• Activity to antigen ratio
deficiency
o <0.5 → Type 2
• What levels of Factor VIII can cause severe o >0.5 → Type 1
bleeding: <30%
• Similar with Bernard-Soulier Syndrome in platelet
• Types: aggregation
o Type 1: Quantitative (most common;
partial deficiency))
CASE 3: Vitamin K Deficiency
o Type 2: Qualitative (problem is the
structure of vWF) • Other name: Hemorrhagic Disease of the
▪ Contains small, intermediate, Newborn
and HMW multimers • Importance of Vitamin K in coagulation:
• 2A: no high and • In the absence of Vitamin K, what is being
intermediate produced?
• 2B: no high but with o PIVKA = Protein induced by vitamin K
intermediate Antagonism
▪ Problems with binding site o PIVKA – no gamma-carboxylation
(platelet, collagen factor VIII) o Normal prothrombin group – gamma
carboxylation allow prothrombin to bind
• 2B: platelet (increased
to phospholipids or calcium since calcium
affinity); 2 specific results
→ platelet count is low, is a cofactor. Coagulation cascade will not
increased ristocetin proceed w/o calcium
induced plt aggregation • Newborns: Umbilical cord removal (bleeding) ?
• 2M: platelet (decreased • Adults: Inadequate intake of Vitamin K (biliary
platelet binding) obstruction diseases), malnutrition, prolonged IV
or parenteral nutrition
• 2N: Factor VIII binding
site; normal vWF is • Medications that can cause Vit K deficiency:
normal but factor VIII is o Broad spectrum antibiotics – destroys
abnormal; also referred intestinal flora
o Coumadin and warfarin anticoagulants
MARIANO, D. 2
 o Accidental rat poisoning o Leukocytes
• Why are breastfed infants more prone? • Formation of thrombi:
o Low Vit K in breastmilk o Thrombin - activates hemostatic system
o Decreased normal flora in the intestine of o Thrombi partially occlude blood vessels
the baby (impede blood flow)
• 3 types: • 3 parts:
o Early – 0-24 hrs due to maternal intake of o Clot formation
certain drugs ▪ Thrombin – primary culprit
o Classical - within a week because it activates platelets and
o Late – common in infants who are coagulation proteins and
breastfed consumes all of these materials
• Drugs that can cause early Vit K deficiency: o Fibrinolysis
o Anti-TB drugs → low Vit K of mother ▪ Fibrinolytic system is activated
• IS Vit. K deficiency common in adults? but during these process
o Rare because it is only due to dietary ▪ In DIC, endothelial cells become
reasons damaged and there is free
• Anemia – Excessive bleeding plasmin
• Lab tests: ▪ Coagulation factors affected by
o BT – normal free plasmin → V, VIII, IX, XI
o Plt count – normal o Monocytes
o PT- abnormal ▪ Cause the secretion of tissue
o PTT – either factor (III) → release of cytokines
o TT – normal because of the inflammation that
• Prevention for the newborn: is also happening
o Vitamin K administration as prophylaxis • 2 types:
• Treatment: o Acute – hemostatic components are
o Severe: FFP transfusion deficient (Uncompensated DIC)
o Vitamin K administration o Chronic – body is able to compensate (CF
can be normal or abnormal)
CASE 4: Disseminated Intravascular Coagulation • Fibrin monomers – in DIC, it is soluble. They do not
polymerize
• Disseminated – found in all over the body
o Coat the platelets and coagulation
• Intravascular – inside the blood vessels
proteins → anticoagulant effect →
• Coagulation – mainly coagulation but as it
bleeding
progresses, there is also fibrinolysis
• Plasmin – digests all form of fibrinogen forming
• Fibrinolysis → anemia
the fibrin degradation products
• Excessive thrombin formation is connected with
• Platelets are decreased because they are
too much clotting
consumed
• Consumption Coagulopathy – other name
• Etiology:
o Platelets and clotting factors are
o DIC is secondary to systemic diseases
consumed
o Bacterial infection
o Primary and secondary hemostasis are
o Malignancies – pancreatic, prostatic,
consumed
ovarian, lung,
• Both coagulation and bleeding
o Cause of acute DIC – burns, sepsis, injury,
• Very complicated condition
intravascular hemolysis, viremia, acute
• General description of DIC – generalized activation inflammation, crush injuries, obstetric…
of hemostasis
• Lab test:
• Involved in the hemostatic systems:
o Blood vessels
o Platelets and coagulation factors
o Coagulation control pathways and
fibrinolysis

MARIANO, D. 3
 o Non-specific – not associated with
bleeding or hemorrhae (more aligned
with thrombosis)
• Systemic Lupus Erythematosus
o Butterfly rash
o More common in female (8x) because
they have estrogen which is an
immunoenhancing hormone. → more
prone to autoimmune
• Lupus anticoagulant → antiphospholipid
anticoagulant
• Not all SLE patients are positive with lupus
anticoagulant
• The patient is more prone to thrombosis than
hemorrhage
o Plt count – decreased
• How does lupus anticoagulant cause thrombosis?
o PT – increased
o No definite reason
o PTT – increased
o In general, since LA are antiphospholipid
o D-dimer – increased
antibodies, they will attack the cells and
o Fibrinogen – either increase or decrease
tissues with these
(cannot be used to diagnose)
• Why is it called anticoagulant if thrombosis is its
main problem?
o Lupus anticoagulant has 2 mechanisms:
▪ In vitro – outside the body,
prolongs the clotting time (based
on test)
▪ In vivo – effect inside the body
• 4 criteria to diagnose:
o 2009 INTERNATIONAL SOCIETY ON
o Peripheral blood smear – anemia with THROMBOSIS AND HEMOSTASIS (ISTH):
schistocytes (due to hemolysis; plasmin Guidelines for LA Detection
triggers the complement system → • 2 diagnostic tests:
hemolysis) o Dilute Russell Viper Venom Test
o Protein C, protein S, and AT activity o Silica-based PTT or aPTT – prolonged
assays – decreased (<50%) ▪ INTRINSIC TENASE COMPLEX
o TT – prolonged ▪ EXTRINSIC TENASE COMPLEX
o Reptilase time - prolonged ▪ PROTHROMBINASE COMPLEX
• Treatment: ▪ Phospholipid surfaces is where
o Acute DIC – slow down the clotting these 3 will bind
process, replace the loss of ______ ▪ When plasma is with Lupus
through transfusion anticoagulant, they inhibit these
o Chronic DIC – surgery, anti-inflammatory complexes from attaching to
agents, obstetric procedures these complexes
• Peripheral blood smear: presence of schistocytes
CASE 5: Circulating Anticoagulants (Lupus (autoimmune disorder so it attacks its own rbcs)
Anticoagulant) • Antinuclear Antibody Test – not diagnostic
• What are circulating anticoagulants? o SLE does not have a definite antibody
o Inhibitors of clotting proteins o Negative – you can eliminate SLE in the
• 2 types: possible diagnosis
o Specific – can exhibit hemorrhagic
episodes

MARIANO, D. 4
 CASE 6: Congenital Afibrinogenemia
• Fibrinogen – protein responsible for the formation
of clots
• Why in afibrinogenemia, bleeding time is
prolonged because fibrinogen is responsible for
platelet aggregation and adhesion.
• Platelet count – normal
o There is impaired plt function due to lack
of fibrinogen
• Factor I is part of common time
• Clotting time – prolonged (no stable clot)
• Fibrinogen NV: 200-400 mg/dL
o 220-498 mg/dL
• Fibrinogen is involved in:
o Primary hemostasis: binds to the
GpIIB/IIIa receptor
o Secondary hemostasis: part of the
coagulation cascade
• Autosomal recessive
o ¼ chance
o 25% affected; 50% unaffected / carrier;
25% unaffected / not carrier
• Distinct clinical manifestation:
o Epistaxis
o Gingival bleeding
o Joint swelling
o Spontaneous abortion
• Fibrinogen assay:
• In afibrinogenemia, lab tests are prolonged
compared to hemophilia A
• Treatment:
o Fibrinogen concentrate
o Medications approved by FDA:
▪ FIBRYGA
▪ RIASTAP
• Can produce against fibrinogen → if this happens,
these drugs are not effective

MARIANO, D. 5