IMMUNOLOGY

CREATEB BY: MOATH FATEH

 Lecture 1-3: Introduction to immune system and cells
Jenner à Cowpox-vaccinated milkmaids did
not get smallpox. Ehrlich à Specific antigen
elicited the production of a specific antibody.

Germ theory and Koch postulates: If you

isolate the microorganism causing the disease
in organism, culture it, and reintroduced it into
healthy organisms, they'll get the disease.

Bone marrow (BM) is place of synthesis of

immune and non-immune blood cells (hematopoiesis).

Immune system recognize pathogens, restore homeostasis and remember the pathogen
(memory cells).

Cells: 4 Granulocytes, 3 main lymphocytes, 3 phagocytes, 3 APCs.

Neutrophils : PMN = segmented nucleus (3-5 connected lobules), short lifespan,

circulate for 6 hours. G-CSF & IL-6 stimulate their production from BM. If there are
microbes in tissues, neutrophils migrate to them, function for a few hours and then die.
NETs are neutrophils DNA that bind pathogens. Has two types of granules : Azurophilic
(lysosOmes) + specific granules (Enzymes: lysozYme, collagenase, and elastase). If
low à neutropenic fever.

Once monocytes enter tissues, they mature and become resident macrophages with
different names according to tissue: microglia (CNS) , Kupffer (liver) , osteoclasts (bone)
Roles: secrete cytokines , display antigens , scavengers, repair of damaged tissue
(angiogenesis & fibrosis). Both neutrophils and macrophages form phagolysosome to
digest microbes.

Eosinophils : deficiency of eosinophils is not important clinically. They migrate and reside
in thymus or GI tract. IL-5 and eotaxin chemokines recruit them into tissues.

Mast cells reside in skin and mucosal epithelium. Basophils only in blood and < 1%. Both
play role in allergic symptoms. Their surfaces coated with IgE & IgG. They can be triggered
by antigen binding to the IgE, releasing histamine stored in granules.

Antigen-presenting cells (APC): APCs links the innate and adaptive immune systems.
Dendritic cells, Macrophages, B cells. Present extracellular proteins on MHC-II.

Types of Dendritic cells :

A. Conventional Dendritic cells (cDC) (Major initiator of naive T cells - Migratory)

B. Plasmacytoid (pDC) (interferon α/ß production; viral infections)
C. Follicular : not BM-derived but from fetal Hematopoietic Stem Cells, always lie in
germinal centers of lymph nodes (LN) & spleen. They trap antigens complexed to
antibodies or complement and display them for B cells.

cDC have membranous projections that facilitate phagocytosis. After doing so, they migrate
to LN and present antigens to T cells. Macrophages present antigens to T helper in the tissue
itself. B cells present antigens to helper T cells in LN and spleen.

Natural killer cells perform their killing function without clonal expansion & di^erentiation.
Fight against intracellular viruses and bacteria. They have receptors that when recognize
MHC-I on host cells, get inhibited. If no MHC-I is there, NK destroy that cell.

65% of lymphocytes are in LN & spleen. Each clone of lymphocytes expresses antigen
receptors with single specificity, produced by recombination of DNA segments during the
maturation of cells. B and T produced from BM. Early maturation of B cells occurs in BM
while for T cells in thymus. B cells à humoral (circulating antibodies). T cells à cell
mediated immunity. CD3 are present on both T helper and cytotoxic T lymphocytes (CTL).
CD19 on B cells.

• Helper T cells activate macrophages to kill the phagocytosed microbes.

• CTL recognize abnormal proteins on MHC-I and destroy the infected host cells by
perforins and granzymes.
TCR binds MHC:peptide complex. TCR of CD4 + cells binds MHC-II (4*2=8) and CD8+
cells binds MHC-I (8*1=8)
• Regulatory T cells suppress immunity. γδ lymphocytes are innate immunity like NK.
 Lecture 4: Tissues of immune system
Primary central lymphoid organs : BM, Thymus. Secondary : spleen, LN (capsulated)
MALT (non-encapsulated).

Thymic medullary epithelial cells present self-antigens to developing T cells and causing
their deletion. Maturation in thymus begins in the cortex, then they migrate toward the
medulla which contains mostly mature T cells. DiGeorge à heart & T cell deficiency

Unlike dendritic cells which initiate adaptive immunity to microbes in tissues, spleen initiate
adaptive immune responses to blood-borne antigens. Spleen's red pulp has macrophages
that filters out immune complexes and opsonized microbes from blood. White pulp is
lymphocyte rich & lie around central arterioles, separated from red pulp by marginal zone.
Having no spleen ↑ risk of infections with encapsulated bacteria.

LN Anatomy : Capsule, subcapsular sinus, cortex made of B cell zones or follicles with
germinal centers that have Follicular DC, T cell zone and medulla.

Mucosa-associated lymphoid tissue à immune responses to ingested (GI) and inhaled

(bronchial) antigens / microbes. Examples of MALT are tonsils and Peyer's patches
(aggregates of lymphoid follicles in small bowel).

Lecture 5: Pattern recognition molecules

Both innate and adaptive immunity recognize antigens; the antigens that stimulate
innate immunity are limited in number and of 2 types:
1. PAMPs (e.g. LPS in G-ve bacteria, lipoteichoic acid or peptidoglycan in G+ve bacteria
mannose-rich oligosaccharides, bacterial flagellin protein, viral dsRNA).
2. DAMPs (released from damaged host cells by means other than apoptosis like by
infection, trauma, burns, chemicals, ischemia (decreased blood supply)) e.g. Heat shock
protein HSP.

In order to produce cytokines (IL, chemokines, interferons, TNF), PAMPs and DAMPs bind to
pattern recognition receptors (PRR) found on/in innate immune cells. PRR can be cell
bound (to plasma membrane or endosomes or in cytosol) or soluble (in blood, ECF).

PRR types (most of these are cell bound):

1- C-type lectin family (mannose and dectin receptors) found soluble in blood / ECF or
bound on macrophages and dendritic cells.
2- Scavenger proteins mainly on macrophages.
 3- N-Formyl met-leu-phe receptors binds N-formylmethionine residue on bacterial
proteins and direct the movement of neutrophils and macrophages toward bacteria
(chemoattractants).
4- Toll Like Receptors TLRs (10 di^erent subtypes 1 to 10, cell bound). TLR4 (cell surface)
binds bacterial LPS. TLR3 (endosomes inside the cell) binds viral dsRNA. When TLR
bind to a ligand, 2 TLRs come together and form a dimer.

MD2 and CD14 (Co-signaling molecules), are proteins needed for TLR signaling. MD2
recognizes and binds lipid A component of LPS before binding to TLR4. Subsequently,
TLR4 or other TLRs activate MyD88 (then NF-kB induce expression of IL and
chemokines). Only TLR3 activate TRIF after binding viral dsRNA to synthesize type 1
interferon.

5- NOD and RIG like receptors (free in cytoplasm; considered cell bound).
NOD1 and NOD2 like receptors respond to bacterial cell wall peptidoglycans. NLRP3
produce inflammasomes, which generate active forms of IL-18 & IL-1b (produced as
pro IL-1b by TLR and NOD mediated NF-KB pathways, cleaved by NLRP3
inflammasome).
RLRs can recognize double-stranded and single-stranded viral RNA.

Acute inflammatory mediators:

1. TNF produced by T-cells & macrophages

2. IL-1: macrophages, mast cells, keratinocytes, epithelial and endothelial cells.
3. IL-6: macrophages, mast cells
TNF can have pathological e^ects like low cardiac output and insulin resistance. IL-6
stimulate di^erentiation of IL-17–producing helper T cells. IL-1 & IL-6 induces liver
synthesis of acute phase proteins. IL-1 and TNF together increase capillary
permeability (tissue swelling). All 3 produce fever and induce leukocyte production in
bone marrow. (1,6 liver // 6à17)
4. Type I interferons (3 types α,ß (anti-viral) & γ)
- cause sequestration of lymphocytes in lymph nodes
- increase the cytotoxicity of NK cells and CD8+ CTLs
- Upregulate expression of class I MHC molecules to be killed by CD8+ CTLs.
- Interferons induce expression of enzymes that block viral replication within
cells and degrade its RNA.
 Lecture 6: Complement system and other soluble PRRs
The soluble e^ector molecules (part of innate immunity) function in 2 major ways:

1. Opsonization which facilitates phagocytosis. Opsonins include antibodies and

complement proteins. The Fab portion of antibody binds to the antigen and Fc
portion binds to Fc receptors on macrophage/ neutrophil to facilitates phagocytosis
of microbe.
2. Direct killing of microbes

There are 5 soluble PRRs:

1. Natural antibodies; produced by B cells without the need for exposure to foreign
antigens. Mainly pentameric IgM. Bind carbs or lipid molecules on bacterial and
apoptotic cell membranes.
2. Collectins:
Made up of 3 or 6 subunits, each of which contains a collagen-like tail
connected to a calcium-dependent (C-type) lectin head. Example:

Mannose-binding lectins MBL

Found in plasma
Bind to carbs with terminal
mannose and fructose
activates lectin pathway
3. Ficolins are plasma proteins similar to collectins, but have a Fibrinogen-type
carbohydrate recognition domain instead of C-type lectin head.
4. Pentraxins: Both CRP and SAP bind to a few PAMPs and DAMPs, and can bind C1q
and initiate the classical pathway. These are acute phase proteins (IL-1b & IL-6
stimulated their synthesis in liver)
5. Complement system does 3 functions:
Opsonization è C3b , C4b
Draw in leukocytes and inflammation è C3a , C5a {a: anaphylaxis}
Pore in bacterial cell wall and lysis è MAC / C5b – C9

Classical pathway starts with C1q, Lectin pathway starts with binding of
Ficolins and MBL. In these 2 pathways, C3 convertase is made from C2bC4b.
Alternate pathway starts with C3 convertase which is made by C3bFB

Regulators of the complement system: CD46 (inhibits C3 convertase formation), CD55

(accelerate Convertase delay), CD59 (prevent MAC/TCC formation), C1-INH (inhibits
classical pathway)
 Lecture 7: Epithelial barriers and leukocyte migration
1- Tight junctions of skin and mucosal surfaces
2- Mucus which contains: lysozYmes (cleaves peptidoglycans), IgA, glycoproteins
such as lactoferrin and mucins (antimicrobials) + inorganic salts + its viscosity.
Mucus facilitates ciliary and peristaltic removal of microbials. [altered consistency
of mucus in cystic fibrosis, immobile cilia in primary ciliary dyskinesia both lead
to recurrent chest infections]
3- Antimicrobial peptides possessing a net +ve charge are attracted and incorporated
into -ve charged bacterial membranes thus disturbing them. E.g. Defensins
(epithelial cells, neutrophils, NK, CD8+), Cathelicidins (epithelial cells, neutrophils)
4- Intraepithelial T lymphocytes: γδ receptor, non CD4+ non CD8+, do not depend on
MHC, recognize limited peptide and nonpeptide antigens (innate immunity).

Leukocyte recruitment from the blood and lymph nodes into tissues releasing
chemokines signals depends first on adhesion of the leukocytes to endothelium.
Leukocyte migration has 4 steps:

1. Rolling / tethering : Ligands (complex sialylated carbohydrate) on leukocytes bind to E-

selectin and P-selectin on endothelial cells. This low a^inity binding results in
leukocyte rolling (Slowing down leukocytes).
2. Integrin activation : Leukocytes' integrin molecules change from low to high a^inity
in response to chemokines. (integrin is receptor)
3. Firm adhesion : ICAM-1 ligand on endothelial cells bind to LFA-1 integrin on
leukocytes resulting in firm adhesion.
4. Extravasation : paracellular transmigration in between endothelial cells.

IL-8 is a CXC chemokine that induces chemotaxis in neutrophils & stimulates phagocytosis
once neutrophils have arrived. CXC means 2 cysteine (CC) residues are separated by 1 amino acid (X).

Leukocyte adhesion deficiency is immunodeficiency disorder where patient cannot

form pus because leukocytes cannot migrate to infection site. Eventually recurrent
bacterial infections.
 Lecture 8: Antibodies
Always first antibody is IgM. In primary response, lag phase is 1 week, steady state in 3 wks.
Secondary response: short lag time, larger amount of antibodies which decline later.

A^inity: strength of binding of single site of antibody to 1 epitope.

Avidity: binding of 1 antibody to multiple epitopes (1 IgM molecule has 10 binding sites; so
more avidly bind to antigen than 1 IgG binding 2 epitopes than 1 IgG binding 1 epitope).

Multivalent antigen has more than 1 identical epitope.

Antigen à specific BCR à phagocytosis à MHC-II à activate T-cells à cytokines à

proliferation into clonal B-cells à di^erentiation into memory & plasma (with no BCR).

T-cells à cytokines à Genetic rearrangement in B cells à change constant regions à class

switching : IgM à IgG/ IgA/ IgE with same antigen specificity (same V regions).

Repertoire: total collection of antibodies with di^erent specificities. Random

recombination of genes encoding variable regions results in diversity (ability to bind large
number of antigens). Aiinity maturation occurs when somatic mutations result in new V
regions with higher a^inity and the selection of B cells producing them.

Antibodies get rid of microbial antigens through 4 methods: neutralization (prevent binding
of toxins or microbes), opsonization (Fc binds to its receptor on phagocytes), antibody
dependent cellular cytotoxicity (NK) & complement activation (Fc binds to complement).

J chain present in IgM / IgA. Each antibody is made of 4 polypeptides. Most of the variability
between di^erent antibodies are confined to 3 hypervariable regions (CDRs) which are
present in heavy and light chain V region. (note: single Ab have a 12 HVR)

The following 5 isotypes diier in heavy chain C region.

IgM: Pentameric with many binding sites = agglutination, activates complement, 10 binding
sites, 60 HVR.

IgG (G is mother, g is newborn) : the most abundant immunoglobulin in serum , long-lived

antibody response , cross the placental barrier (passive immunity to newborn). Formed in
large amounts during the secondary response to antigenic stimulus, and usually follows
production of IgM in the course of a viral or bacterial infection, has a 12 HVR.

IgA (A Apple ‫)ﺗﻔﺎﺣﺔ‬: provides local immunity by preventing colonization and infection.
Secreted by plasma cells in mucosal tissues then each 2 IgA molecules combine (dimer)
with secretory piece (secretory IgA (sIgA)).
IgE (ellergy) : immunity against helminths + mediate type I hypersensitivity (asthma, allergic
rhinitis and atopic dermatitis). IgD : receptor on immature B-cell (also secreted in small
amount).

Lecture 9: B-cell activation

How B cell activated via binding to T cells ( T- dependent) in the lymph node
Chemokine receptor & its ligands Function
CCR7 CCL19 and CCL21 B cells express CCR7 to move away
from follicle toward T cell zone
CXCR5 CXCL13 B cells move toward the follicle.
When B / T cells start expressing T cells express CXCR5 to move toward
one receptor, the other
decreases. the follicle.
Survival of naïve B cells depends on : Signals from 1- BCR 2- BAFF

- Follicular B cells are not static but recirculating between LNs searching antigens. Naïve
follicular B cells have IgM and IgD receptors.
- B-1 B cells (secrets natural antibodies): spontaneously secrete IgM react with
microbial polysaccharides and lipids. Rapid antibody production against microbes in
peritoneum. At mucosal sites, as many as half the IgA-secreting cells derived from B-1
cells.
- Marginal zone B cells (spleen; respond to polysaccharide antigens and generate
natural antibodies.
How Antigens reach B-cells (points 1-2)? How B-cells get activated (points 3-5)?
1) Follicular B cells recirculate between LNs searching antigens.
2) Subcapsular sinus of LN contain macrophages that capture large microbes and
antigen-antibody complexes and deliver them to follicles. Medium sized antigens à
Dendritic cells deliver them to follicles. In both cases, Antigens reach B cells
unchanged, unlike when these cells present them to T cells.
3) The activation of antigen-specific B lymphocytes is initiated by the binding of antigen to
membrane IgM/ IgD molecules (BCR), which, in conjunction with the associated Igα and
Igβ proteins, make up the antigen receptor complex. For full responses to be induced,
other stimuli cooperate with BCR engagement, including complement proteins (binding
to CR2), pattern recognition receptors (like TLR), and, in the case of protein antigens,
helper T cells.
4) After antigen binding, B cells activate and engulf the antigen to present it on MHC-II for
CD4 +ve T cells. Cells enter G1 phase & produces bcl-2, an anti-apoptotic protein.
5) TCR of T helper cells (CD4) bind to MHC-II : peptide complex & produce cytokines that
activate B cells.

T-dependent = protein antigens. T-independent = polysaccharides (haptens)

Polysaccharides display multiple identical epitopes on each molecule or on a cell surface.

Therefore, such multivalent antigens e^ectively cross-link many B cell antigen receptors and
initiate responses (secretes IgM) even though they are not recognized by T helper cells.

Hapten-carrier eiect: Haptens are small chemicals that can be bound by specific
antibodies but are not immunogenic by themselves. If, however, haptens are coupled to
proteins, which serve as carriers, the conjugates are able to induce antibody responses
against the haptens.

!This can be used in the production of conjugate vaccines. A conjugate vaccine consists
of a polysaccharide antigen that is conjugated to a carrier molecule.

Hapten need Help from other proteins to initiate an immune response

Lecture 10: B-cell activation 2

On activation (After binding of B and T cells), helper T cells express CD40 ligand (CD40L),
which engages its receptor, CD40, on antigen-stimulated B cells
Initial response (Extra follicular) VS. late response (Follecular):

Initillay: extrafollicular activation of B cells into short lived plasma cells, which secretes IgM
(mainly), without memory B cells, and by the help of Th extrafollicular cells

Late: in the germinal center, the plasma cells and memory cells got released into the blood
stream, with high a^inity and diversity.

Plasma cells will resides into the bone marrow while memory B cells will be recirculated

what happens within the germinal center?

aiinity maturation (to have antibodies with better a^inities), isotype switching (switching
from IgM to IgG, IgE or IgA), generation of memory B cells, and long-lived plasma cell
diierentiation.

Isotype switching:
- Occures both in the extrafollicular and follicular B cell activation
- Depends mainly on the cytokines release by T cells, which activated by specific
antigen
 - Polysaccharides and lipids, T-independent, stimulate IgM
- Viruses and many bacteria, stimulate IFN-γ.
- Helminths and parasites, stimulate IL-4, which stimulate IgE release

A1ininty maturation:
- Occures only in the germinal center
- It means the increased a^inity of antibodies to the antigen, by something known as
somatic mutation in the specific regions, known as Hypervariable region (
complementary determining region)
- Not happened without the help of CD40L:CD40 interaction (need Th cell)

In the follicles, only B cells that binds to antigen presnted on the FDC will be released in the
blood, and any cell that doesn’t bind will die by IL-21 secrted by Tfh cells.

There are 2 types of plasma cells (activated B cells that secretes antibodies):

1- Short-lived plasma cells are generated during T-independent responses and early
during T cell– dependent responses in extrafollicular B cell foci.
2- long-lived plasma cells are generated in T-dependent germinal center responses to
protein antigens. Signals from the B cell antigen receptor and IL-21 cooperate in the
generation of plasma cells, acquire the ability to home to the bone marrow, where
they are maintained by cytokines of the BAFF family.

Major site of production of antibodis is BONE MARROW, where plasma cells live and do not
resirculate (VS. memory cells which circulates), they need a Bcl-2 gene to prevent apoptosis
and to live for a long time (until the host dies).
Antibody feedback (regulation of antibodies secretion);

- once B cells secrete an antibodies, some of them binds to the inhibitory FcγRIIB by
using phpsphate group, which binds to Ig-alpha and beta, thus inhibiting the
antibody secretion.

‫ﺗﻢ ﺑﺤﻤﺪ ﷲ‬
‫ وإن أﺳﺄﻧﺎ ﻓﻤﻦ أﻧﻔﺴﻨﺎ وﻣﻦ اﻟﺸﯿﻄﺎن‬..‫إن أﺣﺴﻨﺎ ﻓﻤﻦ ﷲ‬
‫ وﻟﻛن ﺗؤﺧذ اﻟدﻧﯾﺎ ﻏﻼﺑﺎ‬.. ‫وﻣﺎ ﻧﯾل اﻟﻣطﺎﻟب ﺑﺎﻟﺗﻣﻧﻲ‬

‫ وﻟﮫ اﻟﺣﻣد ﻓﻲ اﻟﺳﻣﺎوات‬،‫ اﻟﺗوﻓﯾق واﻟﻣﻧﺔ‬9‫ھذا وﺑﺎ‬

‫ وﻟﮫ اﻟﺣﻛم وإﻟﯾﮫ ﺗرﺟﻌون‬،‫واﻷرض‬