86 Fundamentals of Oral Histology and Physiology

Circumpulpal dentin
Enamel The bulk of dentin is constituted of circumpulpal dentin exclu-
sively produced by the odontoblasts (Fig.  5.5a). Odontoblasts
initially form predentin, which undergoes mineralization and
Primary
Mantle becomes dentin. The daily production of dentin decreases from
dentin
dentin 10 μm/day at the onset of dentin formation to 4 μm/day. This
generates a rhythmic formation, and at each 4 μm a daily incre-
Secondary
dentin mental (von Ebner’s) line appears. This circadian rhythm is
Pulp
interacting with another longer-period rhythm forming Owen’s
Predentin
lines, appearing each 16–20 μm, due to unknown mechanisms.
The continual centripetal formation of primary dentin results in
Cementum
gradual reduction of the pulp volume (Figs. 5.2 and 5.3).
As each odontoblast deposits dentin matrix and retreats
toward the central pulp, it extends an elongated process from the
distal end of the cell body. Continual deposition and mineraliza-
tion of dentin matrix around the odontoblast process creates a
tubule within the dentin, from near the dentinoenamel junction
to the pulpal surface of the dentin. Dentin contains about 20,000
dentinal tubules per mm2, with variations between the outer
Figure 5.1 General organization of a tooth. Beneath the coronal and inner parts of the dentin layer. Due to space restriction,
enamel, the different layers of dentin are found. Mantle dentin is the more tubules are present in the inner third than in the outer
first dentin deposited; circumpulpal or primary dentin constitutes the third (between 18,000 and 21,000/mm2). The tubules are curved,
bulk of the tooth. Secondary dentin is deposited at a slow rate after displaying a gradual S shape from the dentinoenamel junction to
primary dentin formation is complete. A layer of predentin separates
the pulp. The primary dentin of the crown and most of the root
the dentin from the pulp. The root is covered by cementum.

Dentin location, types and structures

Enamel DEJ
Primary dentin Metadentin
Neonatal line Secondary
dentin Odontoblast
processes pulp
Hoehl' s cells

Odontoblast
cell bodies
predentin
Circumpulpal dentin

Mantle dentin
Hopewell-Smith & Tomes' granular layer

Figure 5.2 Schematic distribution of the different dentin layers located between the dentinoenamel junction (DEJ) and the dental pulp. Two
atubular peripheral layers are located beneath the enamel or cementum: the mantle dentin in the crown and the peripheral layers in the root,
including the atubular hyaline Hopewell-Smith layer and/or the Tomes’ granular layer. Primary dentin includes the atubular peripheral layers and
the tubular circumpulpal dentin of the crown and root. The neonatal line separates dentin formed before birth from dentin formed after birth.
Secondary dentin is formed continuously after the completion of primary dentin formation. The mineralization front, or metadentin, is the border
where dentin mineralization is initiated. Odontoblast processes cross the predentin (15-20 μm in width) and enter the dentinal tubules. Hoehl’s cells
may differentiate into odontoblasts, the dental pulp being located beneath.
Chapter 5 Dentin, Pulp, and Tooth Pain 87

(a) (b)

cp pd

e a
odontoblasts
pd

cp
d
odontoblasts

Hoehl' s
cells pulp

pulp
10 μm

Figure 5.3 Light micrographs of odontoblasts and forming dentin. (a) One-μm-thick section of a rat incisor. On the right, enamel (e) is formed by
secretory ameloblasts (a) The mineralization front (arrows) located at the edge of the dentin layer (d) is covered by silver grains (radioautography
using (3H) serine as phosphoprotein precursor). Odontoblast processes (arrowheads) extend through the predentin (pd). Dentin extracellular
matrix molecules are synthesized in odontoblast cell bodies. Hoehl’s cells may differentiate into odontoblasts. The odontoblast and Hoehl’s cell
layers include endothelial cells forming capillaries (cp). The dental pulp is located more centrally. Bar = 10 μm. (b) Hematoxylin- and eosin-stained
section of a human molar. Labels as in 5.3(a). Arrowheads indicate odontoblast processes, and arrows indicate Hoehl’s cells.

Table 5.1 Schematic distribution of the successive layers and types of sound dentin in longitudinal section between the dentinoenamel (DEJ)
and the pulp

Location Dentin Type Structure

Peripheral dentin Crown: Atubular dentin, 30–150 µm thick.

Circumpulpal dentin
Peripheral dentin
Circumpulpal dentin
● Mantle dentin Initial dentin formed in the crown.
Non-phosphorylated proteins.
Resilient border.
Root: Initial atubular dentin.
● Hopewell-Smith layer: 8–15 µm Calcospherites or globular structures and interglobular spaces.
and/or A few bent minute tubules.
● Tomes’ Granular layer: 8–15 µm
Primary dentin The dentin formed between the creation of initial peripheral
dentin and early tooth functioning. The tubules display a
longitudinal S-shaped curvature. The formation of primary dentin
is limited by the outer DEJ and the initiation of secondary dentin
deposition.
Secondary dentin Formed normally and continuously after completion of the crown
and root. Results from the daily 4-µm dentin formation. The S-shape
of the tubules is more accentuated, and the tubules are less
numerous, due to space restriction.
Atubular dentin ● Mantle dentin
● Hopewell-Smith & Tomes’ granular layers
● Fibrodentin formation in the mesial and distal surfaces of
the root canal
Tubular dentin Intertubular dentin: a collagen-rich dentin associated with non-
collagenous proteins along and between the collagen fibrils. The
hydroxyapatite crystallites are 3–5 nm thick and 60 nm long.
Peritubular dentin: 25–30 nm crystallites, and an amorphous
network without collagen fibrils lining the lumina of dental
tubules.
88 Fundamentals of Oral Histology and Physiology

Table 5.2 Pathologic dentin

Causative Event Response Structural Alteration

Genetic alterations: Gene deletion of collagen I, or phosphorylated Abnormal dentin properties and tooth structure
● dentinogenesis imperfecta proteins (SIBLINGs) (DSPP, DSP, DMP1)
● dentin dysplasia
● X-linked hypophosphatemia
Environmental effects: Odontoblast gene expression changes Hyper/hypomineralization, structural alterations
● fluorosis
● dioxin
● other toxic agents
Loss of tooth structure: Reactionary dentin Ortho- or osteodentin formed beneath a
● caries calciotraumatic line
● abrasion
● restorative dental material
Pulp exposure Reparative dentin Dentinal bridge, partial or total pulp mineralization

(a) (b)

IG

DEJ MD CD GT
E
E

10 μm CA 10 μm

Figure 5.4 Electron micrographs of a rat molar. (a) Atubular mantle dentin (MD) in the coronal part of the forming molar. DEJ: dentinoenamel
junction. E: enamel. CD: circumpulpal dentin. (b) Beneath the forming enamel (E) and cervical ameloblasts (CA), calcospherites contribute to the
formation of the superficial Tomes’ granular layer (GT). An interglobular space (IG) is located between the granules. Bar = 10 μm.

is formed before eruption. The curvature is more accentuated in
tubules formed after the eruption of the tooth.
Secondary dentin formation begins after completion of pri-
mary dentin formation. The structure and composition of sec-
ondary dentin are similar to those of primary dentin. A slight
change in the direction of the dentinal tubules occurs when sec-
ondary dentin formation begins. The formation of secondary
dentin continues at a slow rate throughout life, but is gradually
reduced in later years (Fig. 5.2 and Table 5.1).
Between the tubules, intertubular dentin is found (Figs. 5.5a,
5.6 and Table 5.1). This dentin specifically results from the con-
tinuous apposition of predentin components at the mineraliza-
tion front (Fig.  5.3). It is a collagen-rich structure, containing
fibrils about 80 to 100 nm in diameter (Fig. 5.6). Non-collagenous
proteins are associated either with the surface of collagen fibrils
and holes due to the quarter-stagger transversal bands of colla-
gen fibrils, or they fill the spaces between collagen fibrils
(Fig. 5.7). The dentin organic matrix plays a crucial role in pro-
moting crystal nucleation and allowing limited growth to
hydroxyapatite crystallites. The needle-like crystallites are 3 nm
in thickness and 60 nm in length. Each crystallite is formed by
the association of two plates (each 1 nm thick) separated by a
thin electron-lucent layer (Fig. 5.8a).
The lumen of the dentinal tubules contains non-mineralized
collagen fibrils and some amorphous proteins forming a perio-
dontoblastic structure around the odontoblast process. Peritubular
dentin forms a highly mineralized ring around the tubules but
never completely fills the lumen. Isodiametric crystals about
25 nm in width and 9 to 10 nm in length resist abrasive forces and
occlusal pressures (Fig. 5.5b and Table 5.1). These crystallites have
high magnesium and carbonate content. They display high solu-
bility in acid and chelating solutions. Peritubular dentin formation
begins within the tubules, a few micrometers from the predentin-
dentin junction. Present mostly in the inner two-thirds of the den-
tin, it is reduced in the outer one-third and almost lacking in the
mantle dentin. Peritubular dentin does not contain collagen fibrils,
but is comprised of non-collagenous proteins, namely phospholip-
ids or lipoproteins and proteoglycans (Fig. 5.5b).
 (a) (b)

Lu

100 nm 100 nm

Figure 5.5 Electron micrographs of unstained ultrathin sections of human dentin. (a) Intertubular dentin. Crystallites are located along and
between collagen fibrils. The periodic banding of collagen is apparent. Barâ•›=â•›100â•›nm. (b) Peritubular dentin displays isodiametric crystallites about
35â•›nm in diameter. Lumen of dentinal tubule (Lu). Barâ•›=â•›100â•›nm.

co

100 nm

Figure 5.7 Electron micrograph of phosphorylated extracellular

0.5 µm
Figure 5.6 Electron micrograph of type I collagen fibrils (co) in
intertubular dentin. Demineralized section. Barâ•›=â•›0.5â•›μm.
matrix€proteins stained with phosphotungstic acid–chromic acid (PTA)
in intertubular dentin. The electron-dense staining is located along and
between the collagen fibrils. Barâ•›=â•›100â•›nm.

(a) (b)

PD PD

MD

D 0.5 µm 0.1µm
D

Figure 5.8 Electron micrographs of rat molar dentin. (a) Unstained section. Needle-like crystallites located at the mineralization front between the
predentin (PD) and dentin (D) are seen along the collagen fibrils. Barâ•›=â•›0.5â•›μm. (b) The distal border (metadentin: MD) located between the predentin
(PD) and intertubular dentin (D) is densely stained by the phosphotungstic acid–chromic acid mixture. Barâ•›=â•›0.1â•›μm.
90
In the sclerotic zone of carious decay, or after pathologic abra-
sion, or as a consequence of the aging process, the tubules may
be filled by intratubular mineralization. Such heterogeneous
mineralization results from the occurrence of pathologic pro-
cesses that favor the precipitation of calcium and phosphate ions
into non-apatitic forms within the lumen of the tubules.
The reparative process occurs in arrested carious lesions, or in
response to the preparation of cavities. It also may be due to the
effects of the release of resin monomers by restorative dental
materials. These events lead to the formation of tertiary dentin,
classified as either reactionary dentin or reparative dentin.
Hoehl’s cells (Fig.  5.3) and surviving secretory odontoblasts
form reactionary dentin, which appears either as orthodentin or
osteodentin (Table  5.2). If both the odontoblastic and Hoehl’s
cell layers are irreversibly altered, pulp progenitor cells contrib-
ute to the formation of reparative dentin.
Dentin includes atubular peripheral dentin (the mantle
dentin in the crown and Hopewell-Smith and/or Tomes’
granular layers in the root) and tubular circumpulpal den-
tin. After the formation of primary dentin, secondary dentin
is formed during the entire life of the functioning tooth.
Between the tubules, there is a network of intertubular
dentin, whereas around the lumen of the tubules peritubu-
lar dentin increases resistance to occlusal pressures. After
a mild lesion (caries, abrasion, and/or restorative material),
odontoblasts and Hoehl’s cells contribute to the formation
of reactionary dentin (also named tertiary dentin) beneath
a calciotraumatic line. After irreversible alteration of these
cells, or a pulp exposure, pulp cells contribute to the forma-
tion of reparative dentin.
Dentin composition
Dentin consists of an organic matrix composed largely of Type I
collagen, within which hydroxyapatite mineral crystals are
deposited. Mature dentin contains more organic substance than
enamel, but less mineral (Table 5.3). In addition to Type I colla-
gen, small amounts of Type III and Type V collagen are present.
Non-collagenous components, including phosphorylated and
non-phosphorylated proteins, proteoglycans, lipids, growth
factors, and enzymes, account for about 10% of the organic
matrix (Table 5.4). While many of the non-collagenous compo-
nents also are present in bone and other tissue, products of the
dentin sialophosphoprotein gene are specific to odontoblasts and
Table 5.3 Summary of the global composition of dentin.
Component By Weight By Volume
Mineral content 68–70% 50%
Organic content 21% 30%
Water (free and bound) 11% 20%
Note: Substantial differences occur between the different types of dentin
described in the text.
Fundamentals of Oral Histology and Physiology
dentin. These non-collagenous components have important
functions in mineral nucleation and crystal growth.
Dentin formation
Pre-odontoblast differentiation and tooth
morphogenesis
As described in Chapter 3, at early stages of tooth formation, neural
crest-derived mesenchymal cells migrate toward the first branchial
arch. Interaction between these cells (condensing mesenchyme)
and epithelial cells located specifically at placode sites leads to the
proliferation of the epithelial cells and formation of a dental lamina.
The two tissues form the rudiment of the embryonic tooth, epithe-
lial cells forming the enamel organ and mesenchymal cells forming
the dental papilla, or embryonic pulp (Fig. 5.9).
Crown morphogenesis starts with the formation of dental buds,
followed by the cap stage, and later by early and late bell stages. The
lateral proliferation of epithelial cells occurring at the periphery con-
tributes to increase the diameter of the enamel organ. Four epithelial
layers form the enamel organ: the outer enamel epithelium, the stel-
late reticulum, the stratum intermedium, and the inner enamel
epithelium, the latter eventually differentiating to ameloblasts
(Figs. 5.9–5.12). The cells undergo a series of divisions, leading to the
formation of folds, the onset of shaping grooves and cusps. In the
mouse molar, cell division in the embryonic mesenchymal pulp stops
before it does in the enamel organ, leading to a larger surface of inner
enamel epithelium compared with pre-odontoblasts. Folds begin to
form, leading to cusp formation. In the central area of the enamel
organ, the enamel knot regulates the morphogenetic process, influ-
encing the process through cell proliferation and apoptosis.
Epithelial-mesenchymal interactions are instrumental in the
transition from mesenchymal embryonic pulp cells to a pre-
odontoblastic stage. The cells at the periphery of the embryonic
pulp undergo asymmetric division. After the last mitosis, the
larger cells in contact with the basement membrane (BM)
(basal lamina) become early pre-polarized odontoblasts
(Figs.  5.10 and 5.11). These cells ultimately differentiate into
polarized secretory odontoblasts (Figs. 5.13a and b). The cells
some distance away from the BM are smaller and retain their
potential as stem cells. They are grouped in the sub-odontoblas-
tic cell layer of Hoehl (Figs. 5.2 and 5.3), which may differentiate
ultimately toward the odontoblastic lineage. It is crucial to high-
light that odontoblasts are post-mitotic cells. In case of carious
decay or odontoblast injury, differentiation of Hoehl’s cells is
reactivated and polarized secretory odontoblasts are implicated
in reactionary dentin formation (Figs. 5.14–5.16b and 5.16c). If
both the odontoblastic and Hoehl’s cell layers are irreversibly
Early tooth development involves reciprocal interactions
between neural crest-derived mesenchymal cells and oral
ectoderm-derived cells of the enamel organ. The peripheral
mesenchymal cells of the dental papilla proliferate and
become pre-polarized odontoblasts, subsequently differen-
tiating into odontoblasts. Cells in the sub-odontoblastic
layer retain their potential as stem cells.