Enamel:

Composition, Formation &

Structure

PROF. DR. KARTHIKEYAN RAMALINGAM

 ENAMEL
 It is the hardest calcified matrix in the body.
 Ameloblasts are the cells responsible for enamel
formation.
 These cells are lost as the tooth erupts into the
oral cavity and hence, enamel cannot renew
itself.
 Enamel has a complex structure and high
degree of mineralization when compared to
other mineralized tissues like cementum, dentin
and bone.
 Can withstand mechanical forces during tooth
function.
 Physical characteristics

Fully formed enamel has

approximately
95% of inorganic mineral
Crystalline calcium phosphate
(hydroxyapatite)
Can be substituted with ions like
carbonate, strontium, magnesium, lead
and fluoride, if present during enamel
formation. These ions affect the
susceptibility to dissolution.
4% of organic material/proteins
1% of water
 Physical characteristics
 Enamel is translucent
 Varies in thickness – 2.5mm over cusp tips
to feather edge in cervical line
 Color varies from light yellow to gray
white. Underlying yellow dentin could be
seen in thinner regions.
 Extremely hard
 Brittle in nature. Underlying resilient dentin
is necessary to maintain enamel integrity.
 Loss of dentin by caries/cavity
preparation leads to unsupported enamel
which fractures easily.
 Structure of enamel
Structure is difficult to study
Inter-
Demineralized sections rod
show empty space
occupied by mature
enamel.
Fundamental organizational Ro
units are the rods (prisms) d

and inter-rod enamel (inter-

prismatic substance).
 Structure of enamel
Crystals
 Enamel rod was first described as
hexagonal and prism-like in cross
section. But these rods do not have
regular geometry and are not
prismatic.
 Enamel has closely packed, long,
ribbon-like carbonatoapatite
crystals (60-70nm in width, 25-30nm
in thickness).
 Structure of enamel

Calcium phosphate unit has

a hexagonal symmetry.
Fully mature crystals are
irregular in outline as they
are pressed against each
other during final growth.
Rod directions can be
studied under electron
microscopy.
 Rod
Structure of enamel sheath

 Enamel rod is shaped like a

cylinder with crystals running
along the long axis
 Inter-rod region surrounds each
rod and crystals are oriented in
a different direction.
 The narrow space containing Inter-
organic material between rod Ro rod
and inter-rod enamel is called d
rod sheath.
 Ro
d Ro
Structure of enamel Rod
sheath d

Inter-
 In some areas, there is no space or rod

rod sheath between rod & inter-rod

enamel. Ro
Ro
d
d
 The rod crystals are confluent with
inter-rod enamel. This is compared Inter-
with the shape of a key-hole. rod

 Hence, basic pattern in mammalian

enamel are cylindrical rods Ro
embedded in the inter-rod enamel. d
 Amelogenesis

 Amelogeneis or enamel formation is a two-step process.

 When enamel forms, it is only partially mineralized (30%)
 Subsequently, the organic material is broken down and
removed.
 The crystals grow wider and thicker to reach the >96% mineral
content.
 Ameloblasts secrete the matrix proteins that favor mineral
deposition.
Light Microscopy of Amelogenesis

Short columnar cells of the IEE show

reversal of polarity towards the stratum
intermedium.
Induces differentiation of ectomesenchymal
cells to odontoblasts.
Dentin formation initiated.
Induces the ameloblasts to lay down the
initial layer of enamel protein without rods.
Ameloblasts move away from dentin
surface.
Reciprocal
Induction

 At the late bell stage, the IEE differentiates into ameloblasts and as it
differentiates, it induces the undifferentiated ectomesenchyme to form
odontoblasts.
 Odontoblasts forms the initial dentin which will in turn induce ameloblasts to
secrete enamel.
 IEE have been shown to express and secrete several growth factors – TGF β1,
BMP2, IGF, Enamel proteins
 Light microscopy of
amelogenesis
 Can be seen in late bell stage of tooth
formation.
 The low columnar cells of inner enamel
epithelium gradually become taller and
columnar as it is traced coronally.
 Nuclei become aligned to proximal ends
adjacent to stratum intermedium
 Ameloblasts lay down partially-mineralized,
initial enamel without any rods.
 After this first increment, ameloblasts move
away from dentin surface.
 Enamel can be seen as a deep-staining layer.
 Light microscopy of
amelogenesis
 Tome‘s process is the cytoplasmic extension
of ameloblasts giving a picket-fence or saw-
toothed appearance.
 Blood vessels invaginated deep into the
enamel organ to form a convoluted
structure called papillary layer.
 Ameloblasts and papillary layer regress after
enamel maturation.
 Reduced enamel epithelium is formed that
remains till the tooth erupts and interacts with
oral epithelium to form junctional epithelium
in cervical region.
 Life cycle of ameloblast
1. Morphogenetic stage
2. Histo-differentiation stage
3. Formative stage
 Initial secretory stage (no tome’s
process)
 Secretory stage (tomes process)
4. Maturative stage
 Ruffle ended ameloblast
 Smooth ended ameloblast
5. Protective stage
6. Desmolytic stage
Morphogenetic Stage
 Cells are short columnar with large oval
nuclei in the center.
 Golgi apparatus and centrioles are
located in the proximal end of the cell.
 Mitochondria are evenly dispersed through
out the cell.
 During differentiation, mitochondria
migrate to the proximal part of the cell.
 IEE is separated from the dental papilla by
a delicate basement membrane.
Organizing stage

 IEE differentiate into ameloblasts.

 Ameloblast induces differentiation of dental papilla
into odontoblast.
 Ameloblast elongate, nucleus shift proximally
toward the stratum intermedium (Reversal of
polarity).
 Basement membrane fragmented by cytoplasmic
projection and mantle dentin formation.
Organizing stage

 Golgi complex migrates distally from its proximal

position.
 RER increases significantly.
 Mitochondria cluster in the proximal region.
 Distal junctional complex develops compartmentalizing
the ameloblast, into a body and distal extension called
Tomes process.
 During the terminal phase of the organizing stage, mantle
predentin is formed by odontoblast.
Organizing stage

Reversal of the nutrition stream occurs due to the

deposition of mineralized dentin below the
ameloblast.
Proliferation of capillaries of the dental sac along
with the deduction and gradual disappearance of
the stellate reticulum occurs.
Thus, the distance between the capillaries and the
stratum intermedium and the ameloblast layer is
shortened.
 Importance of Junctional Complex
Alignment of adjacent ameloblasts are
maintained by JC.
These complexes encircle the cells at their
distal and proximal extremities.
Fine actin containing filaments radiate from
the JC into the cytoplasm of the ameloblast.
These JC play a major role in amelogenesis
by determining at different times, what may
or may not pass between ameloblasts (to
enter or leave the enamel).
Formative stage
 After the first layer of predentin has been formed,
ameloblast enter into this stage.
 Golgi complex is extensive,
 Numerous cisternae of RER occupy a large part of
the distal region of the cell.
 The mRNA for enamel protein is translated by
ribosomes on the membrane of RER and the
synthesized protein is translocated into the ER.
Formative stage

Enamel proteins progress through the golgi complex

for continued post translational modification and are
packaged in the membrane bound secretory granules.
These granules migrate into distal extremity of the
cell - into Tomes Process.
Secretory granules are released against the mantle
dentin along the surface of the process to form an
initial layer of enamel.
 Maturation Stage:
Enamel maturation occurs after most
of the thickness of enamel matrix has
been formed in the occlusal or incisal
area.
In the cervical part, enamel matrix
formation is still progressing.
Reduction in height of the
ameloblasts
Decrease in the volume and organelle
content.
 Maturation Stage:
Ameloblasts undergo programmed cell death
(apoptosis).
 25% of the cells die during transitional phase and another 25% die as
enamel maturation proceeds.

Ameloblasts show modulation, the cyclic creation,

loss and recreation of a
highly invaginated, ruffle ended, apical or
a smooth surface.
The cells of the stratum intermedium lose their
cuboidal shape and regular arrangement to assume
a spindle shape.
Protective Stage:
 Ameloblast cease to be arranged in a well defined
layer and can no longer be differentiated from the
cells of the stratum intermedium and outer enamel
epithelium.
 These cell layers then form a stratified epithelial
covering of the enamel, so called Reduced Enamel
Epithelium (A+SI+OEE).
 REE protects the enamel by separating it from the
connective tissue until the tooth erupts.
 Desmolytic Stage:
 REE proliferates and induce atrophy of the
connective tissue separating it from the oral
epithelium,
 There is fusion of REE with oral epithelium.
 Enzymes that are elaborated by the epithelial
cells may destroy connective tissue fibers.
 Premature degeneration of REE may prevent
the eruption of tooth.
Lifecycle of Ameloblast
 Electron microscopy of amelogenesis
Ameloblasts have multiple activities throughout their
life cycle.
It is described as 6 phases, with 3 main functional
stages,
Pre-secretory – differentiating ameloblasts show polarity
change, develop protein synthetic apparatus and prepare
to secrete organic matrix.
Secretory or formative – ameloblasts secrete and organize
the entire enamel thickness
Maturation – ameloblasts modulate and transport specific
ions for mineral deposition.
Electron Microscopy of Amelogenesis
 Pre-secretory stage
 Morphogenetic phase
 Differentiation phase
 Secretory stage
 Maturation stage
 Transitional phase
 Maturation proper
 Protective stage
 Mineral pathway
 Secretory stage
Secretory granules are stored at the cytoplasm.
Tomes process initially has only a proximal portion.
Content of secretory granules is released against the
mantle dentin along the surface of the process to lay
down initial layer of the enamel which lacks enamel rods.
Ameloblasts migrate away from the dentin and develop
the distal portion of Tomes process
 Once the distal portion of Tomes’ process is
developed, the secretion of enamel proteins is
confined to two sites.
 Secretion from the proximal part of the process
that is close to JC, around the periphery of the cell
along with that from adjoining ameloblasts, results
in the formation of Inter-rod enamel.
 Secretion from one face of the distal portion of
the Tomes’ process results in the formation of Rod
enamel.
Distal portion of Tomes process increases in
length but becomes thinner as the rod growing in
diameter press it against the wall of inter-rod
cavity.
Distal process finally disappears creating a narrow
space between rod and inter-rod enamel that fills
with organic material forming the rod sheath.
The final layer of enamel protein is formed
without the distal extension of Tomes process and
lacks the rod and inter-rod configuration.
 Maturation Stage

Transitional Phase:
Maturation Proper:
 Modulation of Ameloblasts.
 Cells alternate between ruffle border
or a smooth border.
 Ruffle border with
 Proximal Junctions towards stratum
intermedium - leaky
 Distal Junctions towards enamel - tight.
Transitional phase

 Once full thickness of immature enamel is formed, the

ameloblast changes in morphology to prepare for next phase –
enamel maturation.
 It involves reduction of height, volume and organelle content of
ameloblasts.
 Around 25% of ameloblasts die during transitional phase and
25% die during enamel maturation – cell death by apoptosis &
necrosis.
 Apoptosis also occurs in enamel knot and helps in
morphogenesis.
 Bcl-2 family of apoptosis regulatory proteins play a key role.
Maturation stage

Cells alternate with a

Ruffled border – introduction of
inorganic material
Smooth border - removal of organic
material
Water and organic material are
selectively removed from the enamel
Additional inorganic material is
introduced by alternate bursts of activity.
Maturation stage Ruffle border ameloblasts contain
lysosomes
calcium binding proteins
membrane associated calcium ATPases - promote the
pumping calcium ions into maturing enamel.
Smooth border ameloblasts
secrete enzymes that degrade various matrix proteins
extracellularly into small polypeptide fragments.
These small fragments leave the enamel layer through
leaky distal junctions.
 As the ameloblasts begin their modulation cycles, they deposit a basal lamina at their
flattened apex.
 Basal lamina are rich in glyco-conjugates.
 These glycosylates molecules help regulate the movement of material into and out
of the enamel layer.
 Ameloblasts attach to it by hemi-desmosomes.
Mineral pathway

Mineralization spans both secretory and maturation phases of

enamel formation.
Massive influx of mineral in a short period of time during
maturation phase.
Ameloblasts control the influx of Calcium into mineralized
enamel.
Calcium moves from blood vessels thru the enamel organ to
enamel.
Intercellularly in secretory phase
Transcellularly thru the ruffle ended ameloblast during maturation.
Mineralization of enamel

No vesicles in enamel formation.

Immediate formation of
crystallites in newly secreted
enamel proteins.
No equivalent of pre-dentin /
pre-osteoid.
 Mineralization of enamel
In four stages,
Primary mineralization
Secondary mineralization
Tertiary mineralization
Quaternary mineralization
Mineralization of enamel

Primary – partially mineralized enamel matrix – 30%

mineralization (inner most 8µm next to DEJ, is heavily
mineralized – enamelin)
Secondary – starts at the surface, sweeps rapidly into deeper
layers until the 8µm layer.
Tertiary – increase in mineral rebounding from the innermost
layer out toward the enamel surface. Surface layer is 15µm
wide and mineralized more slowly.
Quaternary – outer layer mineralizes rapidly and becomes the
most mineralized part of the enamel.
 Mineralization of enamel

Thus enamel is
Highly mineralized at its surface
Degree of mineralization decreasing towards the DEJ until
the innermost layer is reached,
8µm layer has increased mineralization.
Ameloblast secretory products
Name of protein Features
Amelogenin (AMELX, • Main protein present in forming enamel
AMELY) • Expression stops when enamel has reached full thickness
• Is a low molecular weight protein that inhibits lateral growth of hydroxyapatite
crystals
Ameloblastin (AMBN) • Smaller amounts than amelogenin but larger molecular weight.
• Seen in newly formed enamel and on outer surface than near DEJ.
• Assist ameloblasts in adherence to forming enamel surface.
• Amelin/Sheathlin is the older term
Enamelin (ENAM) • Largest and least abundant
• Binds strongly to mineral and modulates mineralization and promote crystal
elongation.
Odontogenic • Secreted by ameloblasts and seen throughout maturation in basal lamina
ameloblast-associated
(ODAM)
and
Amelotin (AMTN)
Enamelysin (MMP20) • Calcium dependant metalloproteinase found in newly formed enamel.
• Cleaves amelogenin, ameloblastin and enamelin.
Enamel matrix serine • Serine protease belonging to kallikrein subfamily.
protease (KLK4) • Secreted when enamel has reached full thickness
 Morphogenetic Inner Enamel Epithelium
Phase
Differentiates into

Differentiation
Ameloblasts
Phase

Induces the dental papilla

•Nucleus, mitochondria - Proximal
•Golgi ER – Distal,
•Distal JC is formed
•Polarized Ameloblast Odontoblasts

PreDentin
…contd
…contd Secretory
Induced Ameloblasts Phase
 Extensive Golgi
No rod and
Complex and RER.
inter rod Initial Enamel Layer
• Protein is synthesized
configuration
by translation of mRNA
• Secretory proteins
Migration of Ameloblast migrate to the proximal
away from the part of Tomes’ Process.
Dentin surface

Tomes’ Process

Proximal Process Distal Process

…contd

Proximal Process Distal Process

Inter rod substance Enamel Enamel rod

Distal process Narrow space

goes out of filled with org material
existence called Rod Sheath

Ameloblast become
shorter

Final enamel Without rod and

inter rod partitions
formation
 …contd
Reduction in organelle
Transitional Phase Content and height of
Ameloblasts.

Inorganic material
Maturation Stage Replaces water and
Organic material.

Modulation
Ruffle border Smooth border

• Lysozome, calcium binding • Interstitial fluid leak to neutralize

proteins, calcium ATPase pump the enamel fluid.
calcium ions into the enamel. • Matrix proteins digested by bulk
• Some degraded proteins can be degrading enzymes into
absorbed. polypeptide fragments to leave
the enamel through leaky distal
junction.

Calcification
…contd
Calcification

Ameloblasts decrease
Protective Stage in height and deposit a
Basal lamina at their
Flattened apex

Ameloblasts fuse with

Reduced Enamel the stratum inter
Epithelium medium and outer
Epithelium.
Life cycle of Ameloblast
LM of Enamel
Striae of Retzius
Hunter-Schreger Bands
Enamel lamellae
Enamel tufts
Enamel spindle
Gnarled enamel
DEJ
CEJ
Enamel cuticle
Enamel surface features
Striae of retzius
 Incremental growth lines.
 Successive apposition of layer of
enamel during formation of enamel.
 Brown bands in ground section.
 Striae of retzius
 In longitudinal section – surround the tip
of dentin.
 In cervical part – run obliquely
 DEJ to enamel surface-deviate occlusally.
 In Transverse sections –appear as
concentric circles which are compared to
growth rings in a tree.
 Striae of retzius
 Striae of retzius occurs as a result
of temporary constriction of tomes
processes, associated with a
corresponding increase in secretory
face forming inter-rod enamel.
 Lines of retzius represent change in
enamel growth- accentuated by
disease or change in nutrition.
 Neonatal line

 Nutrition changes prenatally and

after birth.
 Enamel formed after birth shows
accentuated lines of retzius-
dividing the prenatal enamel from
that produced after birth –
NEONATAL LINE.
Hunter Schreger Bands
Series of alternating dark & light bands
seen in L.S section of enamel, when
viewed by reflected light .

Change in direction of rods is responsible

for their appearance.

Originate from DEJ & run perpendicular

or oblique to striae of retzius
 Hunter Schreger Bands

Careful decalcification & staining -

alternate zones having different
permeability & content of organic
material.
Diazones - Bands in which rods are
sectioned more transversely, appear
dark,
Parazones - in which rods are sectioned
more longitudinally, appear light.
It is not just an optical phenomenon.
 Enamel Lamellae

 Thin leaf like structures – extending from enamel

surface towards DEJ.
 May develop in plane of tension & consist of
organic material.
 Lamellae was thought to act as foci of caries -
creating a hypomineralized area containing cellular
debris + other particles from oral cavity.
 Distinguished from artifactual cracks by
decalcification - cracks disappear & lamellae
persist.
Enamel lamellae
Three types of lamellae:
Type a: lamellae composed of poorly calcified rod
segment
Type b: lamellae consisting of degenerated cells.
Type c: lamellae arising in erupted teeth where cracks are
filled with organic matter (saliva).

Type a – within enamel

Type b & c: reach dentin.
 Enamel Tufts
 Arises from DEJ –reaches into enamel to about
1/5th -1/3rd of its thickness.
 Appeared to be branched & contain greater
concentration of enamel proteins.
 Formed as rows with same orientation as the
lamellae.
 Hence seen in abundance in horizontal than the
L .S .
Enamel Tufts
 Tufts consists of hypocalcified enamel
rods & interprismatic substance.

 Occur developmentally because of

abrupt changes in direction of rods that
arises from different region of the
scalloped DEJ.

 Tufts & lamellae – no clinical

significance & do not appear to be sites
of increased vulnerability to caries
attack.
 Enamel Spindles
 The slender projection that transverse the DEJ from
underlying odontoblast.
 Project at right angles to DEJ & thereby form an
oblique angle to the direction of enamel rods.
 Theory 1: Elongated odontoblastic processes that have
insinuated between ameloblast during formative period
of enamel production.
 Theory 2: Another theory it has been noted that the
processes of ameloblast may project well into the
dentin matrix Enamel matrix is deposited in the
territory of dentin & will, therefore surround the
processes of ameloblasts.
 Gnarled Enamel
When direction of enamel rod in
cuspal region of crown become
irregular & twisted – GNARLED .
Seen as series of ridges that probably
increases the adherence between
enamel and dentin.
This configuration provides strength
& resistance to the crushing &
shearing stress of mastication.
Dentino-Enamel Junction (DEJ)
 Surface of dentin at DEJ is pitted.
 Into the shallow depression of the dentin,
fits the rounded projections of the enamel –
Tight grip of enamel
 Appears not as a straight but scalloped line
–convexities of the scallop directed towards
dentin.
Cemento-enamel junction

Relation between cementum &

enamel at cervical region is
variable.
30% of teeth cementum meet
cervical end of enamel.
 Cemento-enamel junction
10% of teeth cementum & enamel
don`t meet.

Occurs when enamel epithelium in

cervical portion of root is delayed its
separation from dentin.
Cemento-enamel junction

 60% - cementum overlaps the cervical

end of enamel.

 Occurs when enamel epithelium

degenerates at its cervical termination,
permitting connective tissue to come in
contact with enamel surface.
 Cross -striation
Human enamel is formed at a rate of
approximate - 4µ a day.
Along the length of rod ,cross striation
–observed at an interval of 4µ -rod a
segmented appearance.
Represent transverse zone or area of
organic content.
Crystallite orientation or localized
width difference -creates optical effect
of striated marking.
 Enamel surface features

Perikymata- striae of retzius often extends

from DEJ to outer surface of enamel ending
in shallow furrows .
Run in circumferentially horizontal lines,
across the face of crown.
 Pellicle
 It is a precipitate of salivary proteins.
 Reforms after an hour –mechanically cleaned.
 Becomes colonized by microorganism to form plaque.
Mammelons
Developmental structures present in newly erupted incisors.
Seen as three prominences or scallop along incisal edges.
Worn off early in life.
Cracks
appear as jagged lines in various regions of tooth surface.
Fissures & grooves
 Seen on tooth surface of crown.
Clinical implications
• Fluoridation
• Acid etching
• Enamel hypoplasia
• Age changes
Fluoridation

 Fluoride ion into hydroxyapatite crystal - More resilient to acid

dissolution.
 Prevents demineralization and in caries prevention.
 If fluoride is present during enamel formation –resistant to acid
dissolution.
Fluoridation

 Semi permeable nature of young enamel

 Enables topical fluorides, fluoridated tooth paste& fluoridated drinking
water to provide an increased higher concentration in enamel surface
of erupted teeth.
 Enhances chemical reaction that precipitate calcium phosphate.
 ACID ETCHING

 Acid etch or enamel conditioning has become an important technique in clinical practice .
 Use of tissue sealants, in bonding of restorative material & cementing of orthodontic bands
to tooth surface.
 Effect is achieved by:
removal of plaque & other debris
increasing the porosity of exposed surfaces through selective dissolutions of crystals for
better bonding surface.
 Etching pattern in enamel
• Etching pattern depends
upon crystal orientation.
• Ultrastructural studies of
crystal dissolution indicate-
crystal dissolve more at
ends than on sides.
• Crystals lying perpendicular
to enamel surface are more
vulnerable.
Age changes
 Enamel is a non vital tissue incapable of
regeneration.
 Masticatory attrition.
 Older patient wear facets & portion of enamel &
dentin erodes.
 Discoloration: teeth darken with age.
addition of organic material from
environment
deepening of dentin color,
visible through the translucent enamel.
 Permeability decreases with age.
 Amelogenesis Imperfecta

Hereditary enamel defect

It can be classified according to the
stages in which enamel is formed:
Hypoplastic
Hypocalcified
Hypomaturative
 Environmental Enamel Hypoplasia

Defects in enamel can be caused by febrile diseases.

 Characterized by distinctive bands of malformed enamel.
Defects can be formed by tetracycline induced disturbances
in teeth.
 Band of brown pigmentation or even total pigmentation.
Defects in enamel can be caused by interference of Fluoride
ion with amelogenesis.
 White patches of hypo mineralized and altered enamel.