Leo Tjaderhane and Susanna Paju
2.1 Introduction
Although dentin and pulp are fundamentally different in that dentin is a mineralized tissue and the pulp is a soft tissue, they are developmentally interdependent and remain anatomically and functionally closely integrated throughout the life of the tooth. Thus, the two tissues are often referred to as the dentin-pulp complex.
Dentin and pulp develop from embryonic connective ectomesenchymal cells from the cranial neural crest during the bell stage of the tooth development (Figure 2.1). The inner dental epithelium of the “bell” encases the condensed mesenchyme. The epithelial-ectomesenchymal interactions initiate the differentiation of the first odontoblasts at the periphery of the dental papilla, and the rest of the mesenchyme will form into future pulp. The differentiating odontoblasts start the secretion of dentin proteins and initiate enamel matrix secretion by ameloblasts [138].
When root formation initiates after crown morphogenesis, Hertwig's epithelial root sheath (HERS) develops from the epithelium at the cuff of the enamel organ. When the HERS grows apically, the adjacent dental papilla cells differentiate into odontoblasts to form root dentin. HERS is critical for root dentin formation: if HERS is disrupted, the dental papilla cells fail to differentiate. On the other hand, cross talk between differentiating odontoblasts and HERS is also necessary for appropriate root formation. The HERS fragmentation allows dental follicle cells to contact the root dentin surface and to differentiate into cementoblasts to form cementum. Also, some of the HERS cells may undergo transition to become cementoblasts. Dental follicle cells secrete collagen fibers that are embedded into the cementum matrix and form the periodontal ligament. Parts of HERS remain in the pulp and in the periodontal connective tissue (Figure 2.2) as epithelial cell rests of Malassez [88, 112, 221]. The formation of lateral root canals and an apical delta of accessory canals rather than a single apical foramen may be a normal variant or it may be due to disturbances of HERS.
The soft tissue of the dental pulp communicates directly with the periodontal ligament (PDL) through the apical foramen or foramina. Sometimes the apical area consists of a delta of accessory canals with several communications between the pulp and the PDL. The rest of the PDL is separated from the pulp-dentin organ by the cementum. Periodontal ligament fibers are embedded in the cementum and alveolar bone as Sharpey's fibers, and attach the teeth to the alveolar bone (Figure 2.3).
2.2 Dentin
Dentin is the largest structural component of human tooth. Dentin provides support to enamel, preventing enamel fractures during occlusal loading. It also protects the pulp from potentially harmful stimuli and participates in the overall protection of the continuum of the hard and soft tissue often referred as the dentin-pulp complex. Dentin in different locations of a tooth may qualitatively differ from each other, which enables it to meet the requirements in that specific location.
Figure 2.1 Initiation of dentinogenesis at the bell stage of tooth development (DP, dental papilla; O, odontoblasts; PD, predentin; IE, inner dental epithelium).
Figure 2.2 Section through the periodontal ligament (PDL) showing Malassez's epithelial rests (M) adjacent to cementum (C) (AB, alveolar bone).
Figure 2.3 The apical portion of a tooth showing alveolar bone (AB), periodontal ligament (PDL), cementum (C), dentin (D), Volkmann's canal (VC).
Dentin is mineralized connective tissue, nanocrystalline-reinforced collagen biocomposite, with unique properties that provide teeth with mechanical strength under heavy occlusal forces. About 70 w-% (55 vol-%) is minerals and 20 w-% (30 vol-%) organic components, the rest being water. However, since the structure of dentin varies within a tooth, these values are only average [208]. About 90% of dentinal organic matrix is highly crosslinked type I collagen, the rest being non-collagenous proteins such as proteoglycans and other proteins, growth factors and enzymes, and small amount of lipids. The mineral is hydroxyapatite (Ca10(PO4)6(OH)2), but contains impurities (CO3, Mg, Na, K, Cl) and fluoride, and should thus be called biological apatite [208].
The major part of dentin is intertubular, formed by the dentin-forming odontoblasts at the dentin-pulp border. Almost the entire dentin organic component is located in inter- tubular dentin. While forming dentin, odontoblasts leave behind dentinal tubules, in which peritubular dentin is later formed.
Peritubular dentin formation leads to a slow occlusion of tubules. Since peritubular dentin is highly mineralized, the mineral-organic matrix ratio increases from the dentin-pup border towards the dentin-enamel junction, and with age [208, 209].
2.2.1 Dentin Formation
2.2.1.1 Odontoblasts, Predentin and Mineralization Front
Odontoblasts are the outermost cells of the pulp, separated from the rest of the pulp tissue (pulp proper) by a cell-poor layer of Weil. During and immediately after the differentiation the odontoblasts organize into a distinguished odontoblast cell layer, and the mineralization of organic matrix completes the mantle dentin formation [138] (Figure 2.4). In the coronal part of the tooth, the morphological features and cell membrane polarization are unique among collagen- synthesizing cells [39, 205]. Odontoblasts are terminally differentiated post-mitotic cells, meaning that they have withdrawn from the cell cycle and cannot be replaced by cell division [39, 209]. Coronal odontoblasts are highly polarized both morphologically [39] and by cell membrane polarity [205] and organized in a pseudostratified palisade, while in root they form a single cell layer [39]. The cell body is located on a pulpal wall of dentin and odontoblast processes inserted into dentinal tubules (Figure 2.5). The cell body is 20-40 µm tall, depending on dentinogenic activity, and contains a large nucleus at the basal portion of the cell, Golgi apparatus, rough endoplastic reticulum, several mitochondria and other intracellular structures [39]. Adjacent odontoblasts are attached with extensive tight junctions forming a stable barrier between cell bodies but may be disrupted e.g. as a response to trauma or caries [23, 40, 209]. The cytoplasmic odontoblast process penetrates into mineralized dentin tubules. It has a 0.5-1 µm main trunk and thinner lateral branches through which the processes may be connected with each other [27, 39, 209] (Figure 2.6). The odontoblast processes are suggested to detect the integrity of the region, acting as a receptor field. Any stimulation is transmitted to the cell body, inducing responses that aim to maintain the tooth integrity. At the same time, the processes withdraw, leaving the tubules empty [127], which in ground section is seen as so-called dead tracts. The extent of odontoblast processes into dentinal tubules is still a matter of debate due to the conflicting results obtained with different research methods and by the possible species differences. In rat molars, odontoblast processes extend all the way to the DEJ [127]. In human teeth, most studies indicate that the odontoblast cell processes would not extend far from the dentin-pulp border (200-700 µm) [27, 209].
Figure 2.4 Tall, columnar odontoblasts (O), the relatively cell-free zone (CF) and the relatively cell- rich zone (CR) in the dental pulp ((BV), blood vessels).
The 10-30 µm layer of unmineralized predentin is located between odontoblasts and mineralized dentin (Figure 2.5). This is where the dentin organic matrix is organized [14] before the controlled mineralization at the mineralization front to form intertubular dentin. The backbone of the organic matrix is type I collagen, whereas non-collagenous proteins - glycoproteins, proteoglycans and enzymes - control the matrix maturation and mineralization (Figure 2.5). The mineralization of dentinal collagen happens via proteoglycan- collagen interaction in the collagen gap zone (intrafibrillar mineralization) [43, 209]. Interestingly, matrix vesicles that are responsible for immature bone and calcifying cartilage mineralization are involved in mantle dentin and reparative dentin but not in primary or secondary dentin mineralization [193]. The mineralization front is often considered to be linear, but actually mineralized globular protrusions called calcospherites are common [209, 213].
Figure 2.5 Schematic description of odontoblast function in dentin formation. Factors involved in dentin matrix formation, maturation, and mineralization. (a) Ca2+ ion transport and handling in the odontoblasts to provide calcium for mineralization and to maintain physiological cytosolic Ca2+ concentration. Intravesicular Ca-ATPase-dependent accumulation of Ca2+ is needed for the controlled transport in the mineralization front. Some calcium may also enter via an intercellular route despite the presence of tight junctions at least in cases of odontoblast cell layer integrity disruption (arrow with question marks). (b) Dentin organic matrix components are processed by odontoblasts and secreted into predentin at precise locations (e.g. dentin phosphoprotein, DPP, directly at or close to the mineralization front). Differential presence of Dentin matrix protein-1 (DMP1), proteoglycans (PGs, e.g. decorin [Dec]) or their side-chains (keratin sulfate [KS], chondroitin sulfate [CS]) indicate enzymatic modifications of the proteins in the predentin for controlled mineralization. Enzymes such as matrix metalloproteinases (e.g. MMP-2 and -3) participate in protein processing during predentin maturation. Cell membrane is highly polarized into basolateral process (blue dotted line) and apical cell body membrane (red dashed line), divided by tight junctions [205]. (c) Odontoblasts also have a transport system that excludes the unwanted proteins and degradation products from predentin. (d) Odontoblasts are also responsible for peritubular dentin formation. Modified from [209].
Figure 2.6 Odontoblasts processes (OP) with presence of intercellular connections (empty arrow) filling the the dentinal tubules enveloped by intertubular dentinal matrix (DM) (original magnification (a) 10,000x; (b) 2500x; inset 20,000x). (Reproduced from [27] with permission from Tissue and Cell.)
2.2.2 Dentin Structure
Dentin can also be divided according to the time of formation into: dentin-enamel junction (DEJ); mantle dentin; primary and secondary dentin; and tertiary dentin, which is further divided into reactionary or reparative dentin, according to the structure and the cells responsible for formation.
2.2.2.1 DEJ and Mantle Dentin
In humans, DEJ is a 7-15 µm wide wavy, scalloped structure [59, 131, 169, 213] that is different from both enamel and dentin [59].
The scalloped form of the interface is believed to improve the mechanical attachment of enamel to dentin. Mantle dentin is 5-30 µm thick layer of the outermost dentin. The matrix is formed during and immediately after the odontoblast terminal differentiation, contains organic remnants of dental papilla, and the mechanisms of mineralization is different from that at the mineralization front [193, 209]. Instead of large tubules, small ramifications of each tubule are present in mantle dentin (Figure 2.7). Unlike the rest of dentin, mantle dentin contains type III collagen (so-called von Korff fibers).
Figure 2.7 (a) Intensive branching of human tooth dentinal tubules close to DEJ (arrows). (b) Intensive branching of dentinal tubules in the middle part of dentin (bars: 20 pm). (Reproduced from [101] with permission from Anatomy and Embryology.)
There appears to be a gradual change of the mineralization rate from the mantle dentin towards the pulp [197], which may create up to 500 pm “resilience zone” necessary to prevent fractures under high occlusal forces [197, 208, 231, 232].
2.2.2.2 Primary and Secondary Dentin
Primary dentin formation (primary dentinogenesis) occurs during the formation and growth of the bulk of the crown and root, forming the main portion of dentin. After it, dentin formation continues as secondary dentin at approximately 1/10 of the rate [208]. The exact time for the “end” of primary dentinogenesis is vague, and actually primary dentin formation slows down gradually [100]. The difference between primary and secondary dentin even in histological or electron microscopy images is often difficult, and has no clinical relevance. Secondary dentin formation continues throughout life, leading to gradual obliteration of the pulp chamber and root canals [208].
2.2.2.3 Dentinal Tubules and Peritubular Dentin
Dentin tubularity contributes e.g. to the mechanical properties [11-13, 129] and behavior in dentin bonding [202]. Generally speaking, the tubules extend from the DEJ at right angles and run smoothly S-shaped course to the dentin-pulp border, but the direction may be different immediately beneath enamel [232]. Tubule orientation may also be different between the dental arches [232], which may affect the mechanical response to loading of teeth in occlusion [208, 232]. The density of the tubules varies depending on the location in the tooth, but is always highest in the dentin-pulp border and reduces towards the DEJ [142] (Figure 2.8). The number of tubules slowly decreases towards the apex, and in the root dentin and especially in the apical area, extensive branching occurs [77, 129, 142, 143]. In coronal area, it is highest and the direction is straighter under the cusps, where also the odontoblast processes [212, 229] and dense nerve innervation [23] penetrate deeper into the tubules. This may relate to the sensing of external irritation and contribute to the regulation of dentin-pulp complex defensive reactions.
Peritubular (intratubular) dentin forms in a regular circular manner on the walls of the dentinal tubules (Figure 2.9). This highly mineralized structure results with an age- related reduction in tubular lumen diameter, even complete occlusion of the tubule.
Figure 2.8 Dense tubularity close to the dentin-pulp border (a) and much sparser close to the DEJ (b). Toluidine blue staining of a human third molar from a young patient. OB: odontoblasts; PD: predentin; D: dentin.
Figure 2.9 Fractured dentin showing intertubular dentin (ID), peritubular dentin (PD) and dentinal tubules (DT): (a) newly erupted tooth; (b) obturated dentinal tubules (OT) in a tooth from an old individual.
2.2.2.4 Tertiary Dentin
Tertiary dentin is formed as a response to external irritation, including physiological and pathological wear and erosion, trauma, caries (in case of which both the lesion size and activity may affect [16]) and cavity preparation, and chemical irritation. The growth factors and other bioactive molecules fluid and its components to pass across the peritubular dentin.
This is called dentin sclerosis. The tubules may also be occluded by mineral crystals due to reprecipitation or from the mineral ions from the dentinal fluid in cases of extensive wear or caries. Often this phenomenon is also called dentin sclerosis, although “reactive (dentin) sclerosis” would be a more appropriate term [208]. Peritubular dentin is often heterogeneous, and it is perforated by tubular branches and several small fenestrations [70], which allow dentinal present in mineralized dentin and liberated during caries or wear are believed to initiate and control the tertiary dentin formation and structure [187]. Tertiary dentin increases the mineralized barrier thickness between oral microbes and other irritants and pulp tissue, aiming to retain the pulp tissue vital and non-infected. The form and regularity of tertiary dentin depends on the intensity and duration of the stimulus. There are two kinds of tertiary dentin: reactionary dentin, formed by original odontoblasts, and reparative den- tin, formed by newly differentiated replacement odontoblasts [16, 138, 171, 173, 209] (Figure 2.10). Reactionary dentin is tubular and relatively similar to secondary dentin in structure, while reparative dentin (also called fibrodentin or even “calcified scar tissue” [16, 138, 171, 209]) is usually atubular or poorly tubularized and may present in variable forms (Figure 2.11). Reparative dentin is believed to be relatively impermeable, forming a barrier between tubular dentin and pulp tissue.
2.2.2.5 Root Dentin
Root dentin bears some distinct differences to coronal dentin. Right under cement, the granular layer of Tomes represents coronal mantle dentin with thin canaliculi and poorly fused globules. The granules contain uncalcified or poorly calcified collagen fiber bundles, and has been suggested to function as a “resilience zone” similar to mantle dentin [102]. As mentioned above, tubular density in root dentin is lower than in coronal dentin, especially in the most apical part [77, 129, 142, 143] (Figure 2.12). Age-related root tubular sclerosis starts from the apical region and advances coronally [149, 216], influencing root dentin permeability [164, 198]. Also, other regional differences occur, as buccal and lingual root canal dentin has patent tubules, while the mesial and distal dentin can be completely occluded [164, 198] (Figure 2.13). These tubular patency/occlusion patterns may correspond to stress distributions under occlusal loading [208], and affect both the bacterial penetration and disinfection [164, 198].
The apical part has also relatively large number of accessory root canals and apical branching (apical delta) and cementum-like lining the apical root canal wall [208] (Figure 2.14). The percentage of apical delta varies between 5.7% (maxillary anterior teeth) to 16.5% (mandibular molars), with the average number of canals being 4 (range 3-18) and about 87% having vertical extension of 3 mm or less [60]. The traditional form of single narrow apical constrict was questioned in a recent micro-CT study, identifying long (> 1 mm) parallel form as the most common, and also a tapered form with no clear constrict as relatively frequent in all types of teeth [179] (Figure 2.15).
Figure 2.10 Intensive irritation (e.g. deep caries lesion) induces local odontoblast destruction and apoptosis and differentiation of replacement odontoblasts forming reparative dentin (RD). Normal wear or other mild irritation induces reactionary dentin formation by primary odontoblasts. Crowding of the odontoblasts causes apoptosis of selected cells (black cells). Modified from [209].
Figure 2.11 (a) A radiograph of a lower molar tooth with deep occlusal caries. (b) Tertiary dentin, consisting of both reactionary and reparative type. A limited area of necrosis is present in the mesial pulp horn (hematoxylin-eosin, original magnification 16x). (c) A section close to that in B (Taylor's modified Brown and Brenn, original magnification 16x). (d) A detailed view of the local microabscess. Bacteria surrounded by inflammatory PMNs on the right, fibroblasts on the left (original magnification 100x; inset 400x). (Reproduced from [172] with permission from Journal of Endodontics.)
Figure 2.12 Longitudinal view of dentinal tubules: (a) in the crown; (b) in the root. The tubules are further apart in the root than in the crown and numerous fine branches are found in the root. Hematoxylin and eosin stained sections.
Figure 2.13 Cross-section of a tooth showing the typical bucco-lingual dye penetration (a). (b-d) Backscattered electron micrograph of areas with (b) and without (c, d) dye penetration. Patent dentinal tubules in dye- penetrated area and marked tubular sclerosis in approximal, non-dyed areas (c, d). Original magnifications: (a) 16x; (b), (c) 1000x; (d) 3000x. (Reproduced from [164] with permission from Journal of Endodontics.)
Figure 2.14 Human canine root tip with the major foramen (arrow) and four accessory foramina (arrowheads) large enough to easily fit ISO 10 instrument.
2.2.3 Dentinal Fluid
The space between the odontoblast process and tubule wall is filled with dentinal fluid. The odontoblast cell layer forms a functional barrier which mostly restrains the passage of fluid, ions and other molecules along the extracellular pathway, and at least in teeth without tissue damage (e.g. caries, cavity preparation, abrasion), dentinal fluid content is believed to be strictly under odontoblast control [209] (Figure 2.5). Dentin also contains several serum proteins, at least albumin, IgG, transferrin, fetuin-A and superoxide dismutase 3 (SOD3) [135], believed to be present mainly in dentinal tubules. With the exception of transferrin [160] they are not expressed by the odontoblasts. Therefore, serum proteins have a passage to dentinal fluid, even in intact teeth. However, the presence of SOD3 [165] and even 100 to 200-fold higher con- centration of fetuin-A in dentin compared to serum [200] strongly indicate active transport systems by the odontoblasts [209].
Figure 2.15 Cross-sectional apical root canal areas with different forms of the apical constriction, and the distribution (percentage with 95% confidence intervals) in different tooth groups. Each point represents a canal cross-section. X-axis: distance to apical foramen in mm, Y-axis: canal area in mm2, mirrored at the zero axis. Modified from [179].
Some evidence exists that physiological dentinal fluid flow may be controlled by endocrine system. A factor called parotid hormone is suggested to affect dentinal fluid flow rate. This hormone, secreted under hypothalamus control has been isolated from bovine, rat and porcine parotid glands and is present in plasma [175, 209]. A synthetic parotid hormone has equal biological activity to respective parotid gland-purified hormone in enhancing intradentinal fluid movement [233].
Dentinal fluid has a distinct role on the stress-strain distribution within the bulk dentin, increasing resilience (the capacity to absorb energy elastically upon loading) and toughness (the ability to resist fracture) [109]. In carious teeth, dentinal fluid is considered a protective factor through the occlusion of dentinal tubules (especially in slowly progressing, chronic lesions) [141] and as part of the innate response of the dentin-pulp complex with the deposition of intratubular immunoglobulins [73]. Both the quality and the quantity of immuno- globulins seem to vary according to caries depth and intensity even in uninfected tubules [73]. However, it is important to realize that inward flow also occurs all the way to the pulp [23, 117] even through enamel, at least in young teeth [23]. The study with different size microspheres demonstrated the size-dependence of penetration, the larger ones (0.2-1 ^m, the size of small microbes) in the inner third of dentin and the smallest (0.02-0.04 µm) even in the pulp [117]. Thus, outward fluid flow is not capable of “washing out” the noxious stimuli from the tubules. Dentinal fluid also affects the success of adhesive restorative procedures. Increased dentinal wetness, due to increased size of dentinal tubules and fluid flow, makes successful bonding in deep cavities (close to pulp) more difficult than to superficial dentin [166]. Dentinal fluid may cause degradation of hydrophilic adhesives, but also increase the collagen degradation rate in the hybrid layer, both leading to decrease in bond strength durability [202].
2.3 Pulp Tissue and its Homeostasis
The pulp tissue - sometimes called pulp proper - is loose connective tissue with type I and III collagens. Cells and structures are embedded in a gelatinous ground substance, containing mainly of chondroitin sulfates, hyaluronates and proteoglycans and interstitial fluid. The cells depend on the interstitial fluid as a mean for nutrient and oxygen transportation and elimination of metabolic waste products (Figure 2.16). Nerves and blood vessels enter the pulp through the apical foramen or foramina (Figure 2.17). They run close together until the main branching takes place in the coronal pulp and final, profuse branching in the odontoblast/ sub-odontoblast region (Figure 2.18).
2.3.1 Pulp Cells
The main cell population in the pulp tissue are fibroblasts. Immediately under the odontoblast layer there is relatively cell-free layer (of Weil) rich in tenascin and fibronectin but low amounts of type III collagen [134]. Below that is the cell-rich layer with dense population of fibroblasts (Figure 2.4). The distribution of the fibroblasts in the rest of the pulp is less dense and relatively uniform. The pulp also contains mesenchymal stem cell-like dental stem cells with self-renewal capacity and multidifferentiation potential [18]. Pericytes are perivascular stellate cells forming a discontinuous layer in close contact with the endothelial cells sur- rounding capillaries and a continuous layer around microvessels [176]. They are classically considered as regulators of angiogenesis and blood pressure. Nowadays, pericytes (or their precursors) are recognized to have mesenchymal stem cell characteristics, including multipotentiality. They are selectively capable of differentiating into adipocytes and hard-tissue-forming cells osteoblasts and chondrocytes [95, 176] also in dental pulp [162, 230], possibly along with other mesenchymal stem cells of a nonpericyte origin [55].
Figure 2.16 Schematic diagram illustrating capillary, cells and interstitial fluid. Blood is brought to the capillaries by bulk flow, and diffusion links plasma and interstitial fluid. The cells are surrounded by interstitial fluid acting as an extension of the plasma.
Figure 2.17 Vessels (V) entering/leaving the pulp through numerous apical foramina (F) in a dog tooth. The number of foramina is not representative for human teeth. (Reproduced from [192] with permission from Journal of Endodontics.)
2.3.2 Blood and Lymph Vessels
Like in any tissues, blood flow is required in the pulp to bring oxygen and nutrients to the cells, and to remove carbon dioxide and metabolic waste products. The pulp circulation is supplied by the maxillary artery, dividing into dental arteries and further arterioles that enter the teeth via apical foramina and through lateral canals. Arterioles are centrally located, and some of them pass directly to the coronal pulp while others supply the root pulp. The blood drains into venules, which largely follow the same course as the arterioles and a triad of arteriole, venule and nerve is often found in central pulp (Figure 2.19). The vasculature differs between the crown and the root. In the root, blood vessels penetrate the apical area of the pulp and form tiny branches. In crown area, capillaries form a subodontoblastic plexus of successive individual glomerular structures that each supply 100-150 qm of subodontoblastic and odontoblastic areas [90]. In young teeth with rapid dentinogenesis, capillaries enter the odontoblast cell layer to ensure their nutrition. Pulp capillaries are relatively thin-walled, may be dis- continuous and fenestrated [28, 52, 90]. Pericytes are embedded within the capillary basement membrane, where they may migrate and undergo transition to a fibroblastic phenotype [28], modulate inflammatory events (e.g. leakage of plasma proteins) and may be involved with calcification of blood vessels [186] and thus be related to pulp stone formation. The blood vessels of pulp are innervated by sensory and by sympathetic nerve fibers (Figure 2.20) [23, 81].
Figure 2.18 Blood vessels in the pulp. Note terminal capillary network (CN) subjacent to the predentin. (Reproduced from [192] with permission from Journal of Endodontics.)
Figure 2.19 Area from the central part of the pulp showing the triad arteriole (A), venule (V) and nerve (N).
The pulp tissue interstitial fluid has lower colloidal osmotic pressure than blood plasma, favoring capillary absorption. This helps to retain low tissue pressure, which is essential for the proper function of the blood vessels in the dentin-encased low-compliance environment. Surprisingly, the presence of lymphatics in dental pulp still remains controversial. The earlier studies indicating pulp lymphatic capillaries have lately been disputed especially in studies using specific lymphatic markers [64, 119, 133, 218].
Figure 2.20 Serial cross-sections of vessels (V) from cat canine pulp. Network of sensory nerve fibers containing the neuropeptides CGRP (a), substance P (b) and neuropeptide Y (c) in the vessel walls. (Reproduced from [81] with permission from Acta Odontologica Scandinavica.)
2.3.3 Nerves
Both myelinated and unmyelinated nerves are present in the pulp (Figure 2.21), majority of them being sensory. The sensory innervation of the pulp is very effective, terminating mostly in the odontoblast layer, predentin, and inner 0.1 mm of mineralized coronal dentin, where they run in close proximity of the odontoblast processes [27, 41]
(Figure 2.22). There are at least six dental sensory nerve fiber types with specific distribution to focus on particular regions of blood vessels, coronal pulp, and dentin (Table 2.1). The sensory fibers are especially dense near the pulp horn tips, where sensitivity is also greatest, and gradually decrease towards the dentin-enamel junction (DEJ). Only few nerve endings are present in root pulp and dentin-pulp border [23]. Pulp innervation is closely related to the microvasculature [90] where the blood vessels are innervated by sensory and by sympathetic fibers, and a few other sympathetic fibers are located in cervical pulp [23, 90]. Nociceptive nerve fibers alert of damage and cause reflex withdrawal that limits the intensity of the initial injury. They also facilitate repair via amplifications of inflammatory, immune and healing mechanisms. Pulpal peripheral nerve fibers secrete a variety neuropeptides that activate receptors on the plasma membrane on target cells and affect tissue homeostasis, blood flow, immune cell function, inflammation, and healing. This phenomenon is called neuro- genic inflammation, and occurs in the absence of direct chemical, thermal or microbial irritation [19, 23, 30]. Nerve fibers also adjust their own functions, cytochemistry, and structure to fit the tissue conditions [23, 65]. On the other hand, adrenergic agonists - better known for their vasoconstriction effect - may directly inhibit of dental nociceptor afferents [31, 76] and environmental conditions, such as pH, can regulate the nociceptive afferent activity, and may be significant in the clinical development and amelioration of dental pain [69]. These examples demonstrate bi-directional tissue-nerve interactions and neuroplasticity especially prominent in nociceptive sensory fibers, the major component of dental innervation (Figure 2.20) [23].
Figure 2.21 Electron micrograph illustrating details of a nerve from the central part of a pulp with myelinated (M) and unmyelineated (U) nerve fibers. (Reproduced from [41] with permission from Acta Odontologica Scandinavica.)
Figure 2.22 Nerve fibers in the periodontoblastic space (a) in the predentin (PD) and (b) in dentin and close contact with the odontoblast processes (OP). (Reproduced from [41] with permission from Acta Odontologica Scandinavica).
Table 2.1 The types, roles, mechanisms of activation, and sites of terminal endings of pulpal nerves.
|
Fiber type |
Sensation |
Activation |
Terminal sites |
|
Sensory A-beta |
“Prepain" sharp pain |
Mechanical: vibration, dentin fluid movement Electric (low voltage) Chemical: mustard oil, serotonin |
Primary: Predentin, OBs, Secondary: dentin, pulp |
|
A-delta fast |
“Prepain" sharp pain |
Intense cold Mechanical: dentin fluid movement Electric (mid-voltage) Chemical: mustard oil, serotonin |
Primary: dentin, predentin, OBs Secondary: pulp |
|
A-delta slow |
Ache |
Intense cold Electric (high voltage) Pulp damage (ATP) Chemical: capsaisin |
Primary: pulp Secondary: blood vessels |
|
C-fiber - polymodal |
Ache |
Intense heat Electric (high voltage) Pulp damage (ATP) |
Primary: pulp Secondary: blood vessels |
|
C-fiber - silent |
Ache |
Chemical: capsaisin, histamine, bradykinin |
Primary: pulp Secondary: blood vessels |
|
C-fibers |
Ache |
Electric (high voltage) Tissue damage (?) |
Primary: OBs, pulp, blood vessels |
|
Sympathetic |
|||
|
C-fiber |
Sympathetic activation, inflammatory mediators |
Primary: blood vessels, pulp |
2.3.4 Pulp Stones
Pulp stones are discrete or diffuse pulp calcifications. One tooth may contain one or several pulp stones with varying size in coronal or in radicular pulp. The exact cause of pulp calcifications remains largely unknown.
External irritation (caries, attrition) certainly may induce pulpal calcifications, but pulp stones also appear in teeth with no apparent cause (e.g. impacted third molars). There seems to be an increase in prevalence with age, especially with the cumulative effect of external irritation [67]. The age-related pulp calcifications have been related to the blood vessels and nerve fibers. Structurally, there are “true” pulp stones, lined with odontoblasts (or rather odontoblast-like cells) and containing dentinal tubules; and “false” pulp stones, which are more or less atubular calcifications, also described as dystrophic calcification [171]. The distinction between the “true” and “false” pulp stones may be artificial, as both tubular and atubular dentin can be present in a single pulp stone (Figure 2.23). Large pulp stones in pulp chamber may obstruct the canal orifices, and in root canal they may complicate access to the apical canal [67, 208]. Apart from creating problems with endodontic procedures, pulp stones do not seem to have any other significance [67].
2.4 Pulp Inflammation
The encasement of the pulp within dentin and enamel creates a low-compliance environment that is unique in human body in terms of inflammatory tissue response. As in any other tissue, external irritation regardless of its nature (chemical, mechanical or thermal) induces a local inflammatory reaction characterized by the dilation of the vessels and decrease in the blood flow resistance (Figure 2.24). Vasodilation and the early recruitment of immune cells are mainly regulated by the sensory nerves via the release of the vasoactive neuropeptides (Table 2.1; Figure 2.25) [30]. The pertinent role of sensory nerves was demonstrated in studies where denervation caused a significant reduction of immunocompetent cells [57] and dramatically advanced pulp necrosis [24] after pulp exposure. Vasodilation together with lower resistance cause an increase in intravascular pressure and capillary blood flow, leading to leukocyte extravasation and filtration of the serum proteins and fluid into the tissue, mainly in the subodontoblastic area [201]. The increase in vascular permeability and accumulation of the proteins can happen quite rapidly, and it is clearly observable already four hours after the cavity preparation [201]. The vascular reactions aim to provide inflammatory cells and to eliminate microbial toxins and metabolic waste products from the area. However, if the external irritation exceed certain threshold level (e.g. continues and intensifying microbial stimuli from advancing caries lesion), it is possible that the pulpal reaction does not limit to the restricted area. Because of the protein and fluid filtration and the increase in cell content, the tissue becomes edematous and tissue pressure increases. In almost all other tissues this would lead to swelling, but in a pulpal low-compliance environment the tissue pressure may increase to the level that exceeds venular pressure, causing the compression of the venules (Figure 2.24). This is followed by increased flow resistance and concomitant decrease in blood flow, because the venous drainage is impeded. The slower blood flow causes aggregation of red and other blood cells and local elevation of the blood viscosity, which further reduces blood flow. The following local hypoxia, increase in metabolic waste products and carbon dioxide, and decrease in pH lead to vasodilation of the adjacent vascular structures, thus leading to the spreading of inflammation (Figure 2.24). The local inflammatory reaction will lead to local necrosis (local pulpal abscess) sometimes called necrobiosis, where part of the pulp necrotic and infected and the rest is irreversibly inflamed [2, 115] (Figure 2.11). Matrix metalloproteinases (MMPs), the enzymes degrading collagen and other extracellular matrix proteins and produced by odontoblasts [206, 207, 211] and especially by the inflammatory cells (PMN- leukocytes, macrophages and plasma cells), aim to confine the spreading of infection. Both chemical [204] and genetic [136, 220] impairment of MMP function leads to increased apical pathogenesis. However, uncontrolled enzyme activity may also lead to increased tissue destruction [4, 72, 183, 211, 219]. If this vicious circle continues, it will slowly spread and lead into total necrosis (Figure 2.24).
Figure 2.23 Approximately 3-mm human dental pulp stone, stained with hematoxylin-eosin (HE) (a, c) and Toluidine blue (b, d-f) staining. (a, b) Toluidine blue demonstrates heterogeneous structure compared to the solid appearance with HE staining. (c) Higher magnification of the lower left corner of (a). Odontoblast-like cells line the lower border of the pulp stone, while the left side is devoid of cells. (d) Same area stained with Toluidine blue demonstrates tubular structure at the site of the odontoblast-like cells, while the area without cells is free of tubules. (e) Higher magnification of (d). Well-formed longitudinally cut tubules are on the left side, while on the right side less organized tubules cut across the tubule direction. (f) In another part of the pulp stone, tubules appear sparse and with numerous fine branches and microbranches. (Reproduced from [208] with permission from Endodontic Topics.)
Figure 2.24 Localized inflammatory reactions in the pulp by e.g. caries, while the adjacent tissue remains intact. At the site of inflammation, inflammatory mediators lead to changes in vascular size and blood flow. If the reaction continues, the vicious circle leads first to a local and finally to total necrosis. (Modified from [201].)
Figure 2.25 Neuropeptide release and their role in neurogenic inflammation. Different irritations and inflammatory mediators such as bradykinin (BK) and prostaglandins (PG), lipopolysaccharides (LPS) and capsaicin result in release of calcitonin gene-related peptide (CGRP) and tachykinins substance P (SP), neurokinin A and B (NKA and NKB, respectively) from C and A-delta sensory fibers. In addition to vasodilation, they activate immune cells and odontoblasts. Vasoactive intestinal polypeptide (VIP) is released from parasympathetic fibers stimulated by nitric oxide (NO), LPS and cytokines, generating vasodilation and exerting immunomodulatory effects on different immune cells. Sympathetic fibers release neuropeptide Y (NPY), resulting with vasoconstriction and reduced fluid tissue pressure. NPY also inhibits neuronal activity in normal conditions. Adapted from [30].
Pulp vasculature is equipped with means to control the spread of inflammatory reaction and necrosis. Arterial U-loops and arterio-venous anastomoses (AVAs) may redirect some of the arterial blood flow away from the inflamed area (Figure 2.26). There are also arterial branching sites with constrictions or sphincters that can control the blood flow into the arterioles leading to the inflamed area. The controlled blood flow may control the level of tissue pressure in a manner that will allow sufficient function of the vasculature in and around the inflammatory area [201]. Neuropeptide Y (NPY), released from sympathetic fibers, generates vasoconstriction, which also will lead to reduced fluid tissue pressure [30].
Figure 2.26 Pulp vascular structures that regulate the blood flow. U-loop and arteriovenous anastomosis (AVA) can be opened to redirect the blood flow away from the area of inflammation to reduce the interstitial tissue pressure. Smooth muscle constrictions in the branching of the arterioles can restrict or close the diameter of the arteriole. Modified from [201].
Pulpal healing and regeneration processes require angiogenesis. Pro- and anti-angiogenic factors liberated from dentin or pulp cells, neuropeptides and hypoxia control pulp vascularization [3, 10, 71, 174, 218], and inflammation leads to angiogenic capillary sprouting.
2.4.1 Immune Cells in Pulp
Human dental pulp contains resident immunocompetent cells that participate in the maintenance of tissue homeostasis and are able to mount innate adaptive immune responses against approaching infection [73, 74]. Inflammatory mediators initiating these responses are released from carious dentin or resident pulp cells [38, 187, 189]. The control of the resolution or advance of the initial inflammation is further regulated by a complex network of inflammatory chemokines, directing the trafficking of immune cells, and cytokines that regulate immune and inflammatory responses [73, 74, 82]. They are produced either by the odontoblasts [73, 74, 85, 116, 159], pulp cells [38, 73, 74], or (mostly) by the immune cells attracted to the site of inflammation. The effects of mediators are temporal con- text dependent: if inflammation is resolved, low levels of proinflammatory mediators may then promote tissue repair, whereas in chronic inflammation repair mechanisms become inhibited [38].
Immature dendritic cells (DCs) and T cells are important in immunosurveillance as part of the innate response to caries. Class II major histocompatibility complex (MHC)- positive immature DCs are located especially in the subodontoblastic but also in the odontoblast-predentin area [40, 52, 153], some of them extending their cytoplasmic processes into dentinal tubules [153, 154]. They detect microbial antigens, initiate maturation, and then migrate to regional lymph nodes to present them to naïve T cells. Streptococcus mutans can rapidly transform monocytes into mature DCs within 24 hours in vitro [75], which may contribute to the local maturation of DCs in inflamed pulps. As a reaction to caries, subodontoblastic DCs infiltrate the odontoblast layer and invade reactionary dentin [40] attracted by TGF-beta liberated from dentin or secreted by the odontoblasts [54]. During maturation, DCs produce pro-inflammatory cytokines and chemokines that recruit circulating immature DCs, DC precursors and T cells to inflamed tissues [114].
The predominant T-cell type in healthy pulp is the memory CD8+ T cell [54, 61, 94], which have higher migratory capacity across endothelial cells than CD4+T cells. Its functions in the normal pulp remain undefined, but in general an immunosurveillance role of CD8+T cells has been proposed [110]. Natural killer (NK) cells are found in the bloodstream and can respond to inflammatory chemokines by extravasating into inflammatory sites. A very small population of NK cells may be present in healthy pulp, where also they may participate in immuno- surveillance [61]. Regulatory T cells (T-reg) are absent [20] or present only in very low amounts [61] in healthy human pulp.
Neutrophils and macrophages are professional phagocytes in innate immune responses. Tissue macrophages are generally derived from circulating monocytes and show a high degree of heterogeneity, which is influenced by their microenvironment [73]. Small number of macrophages [93] and even smaller number of neutrophils [61] may be present in the pulp of an intact tooth. The quantity of macrophages increases reasonably early under caries lesion, when caries is limited to the DEJ or outer dentin [93, 94]. Macrophages may be activated in the early stage of pulpitis to protect the dental pulp by increasing vascular permeability, and to remove foreign antigens and damaged tissue caused by proteases [73, 94]. Activated macrophages are effective in eliminating pathogens in both innate and adaptive immune responses, important in tissue homeostasis through the clearance of senescent cells, and in remodeling and repair of tissue after inflammation. Neutrophils may not be important in early and reversible pulpitis, as they are few in pulpal tissues under shallow caries, but the numbers increase when caries approaches the pulp [94].
2.4.2 Odontoblasts as Immunological Cells
In dentin-pulp complex, odontoblasts represent the first line of cellular defense against external irritants, and odontoblasts have several means to participate in the initiation and development of dentin-pulp complex inflammatory and/or immune responses [53, 68, 116, 217]. Odontoblasts express Toll-like receptors (TLRs), which are a group of trans- membrane glycoproteins that recognize microbial and viral particles, fungal proteins, and viral and bacterial RNAs and DNAs [194]. TLRs initiate the early activation of innate immune responses such as the recruitment of inflammatory cells, production of antimicrobial peptides, and maturation of dendritic cells [53], and may participate in reactionary dentin formation [33]. Out of the 10 TLRs identified in humans, TLR1-9 genes are expressed in mature human odontoblasts or human cultured odontoblast-like cells [48, 53, 158], with TLR2, -4 and -8 (recognizing lipoteichoic acid [LTA] from Gram-positive bacteria, lipopolysaccharide [LPS] from Gram-positive bacteria, and viral RNA, respectively) being most studied [53, 98, 147, 158, 217] (Figure 2.27). In principle, odontoblast-expressed TLRs would be capable of recognizing practically all essential microbial components.
Defensins are a group of small (3-5 kDa) peptides with broad-spectrum antimicrobial, immune cell chemotactic and bacterial toxin inactivation activities [97]. Seventeen defensins in two subfamilies, a- and the β-defensins, are found in humans [97]. Human β-defensins (hBD) -1 and -2 are expressed by the odontoblasts both in vivo and in vitro [46, 47] and suggested to participate in the innate host defense of human dental pulp [46]. hBD-2 is a host-derived ligand for TLR4 [15], and the inhibition of central TLR signaling [80] blocks the S. mutans-induced increase in IL-6 and -8 gene expression in odontoblast-like cells in vitro [45]. Odontoblast-derived hBDs may thus also participate in the autoor intracrine regulation of odontoblast immunodefense.
Figure 2.27 Immunohistochemical staining of TLR8 in human third molar. (a) An intensive staining is present in the dentin-pulp border in all parts of the tooth, with varying staining patterns between different areas. (b) Occlusal dentin-pulp border (higher magnification of the area marked with red square in (a)). (c) Approximal wall of the pulp chamber (higher magnification of the area marked with cyan square in (a)). (Reproduced from [158] with permission from International Endodontic Journal.)
Proteinase-activated receptors (PARs) are G protein-coupled receptors that undergo irreversible proteolytic activation by proteases. They participate in controlling a wide range of biological processes, such as inflammation, hemostasis, thrombosis, and embryonic development [156], and in skeletal growth and bone repair [62]. PAR-2 is present in dental pulp fibroblast-like cells [121, 145], and PAR-1 and -2 are present in human odontoblasts [9]. The expression is significantly increased in response to caries both in the odontoblasts [9] and in pulp tissue [121, 145], indicating a regulatory role in reparative dentin formation and/or in pulp inflammation.
Even though odontoblasts appear to have a distinct role in the regulation of the initiation and progress of the dentin-pulp complex inflammatory response, more research is needed before any clinical treatment approaches are justified. For example, the most potential reparative dentinogenesis-inducing dentinal growth factor TGF-p affects inflammatory interleukin production in mature human odontoblasts [159], inhibit TLR2 and TLR4 expression, and decreases odontoblast-like cell responses to caries pathogens in vitro [86]. Much of the data come from in vitro studies, which differ significantly from the clinical reality. The complexity of the dentin-pulp complex calls for detailed in vivo or in situ studies, with approaches as close as possible to clinical reality.
2.5 Pulp Nociception and Hypersensitivity
Psychological studies of humans show only three main perceived sensations from teeth [23]. There is an initial poorly defined low- stimulus “pre-pain” sensation based on fast- conducting A-beta and A-delta fibers. That sensation shifts to sharp pain at higher stimulation intensities. In addition, there are dull ache sensations related to C-fiber and probably slow A-delta signaling. Dental nociceptive neurons may also have proper- ties unique from other tissues [36]. An unusual feature of dental innervation is the sensitivity of all types of nerve fibers to intense cooling of the tooth [23]. Several differences distinguish pain in teeth from other tissues. Especially in pulpitis, slight thermal or air stimuli that elsewhere are felt as cold, hot or light “breeze” can easily evoke pain in teeth. The low-compliance environment of the pulp and nerve sprouting and changes in neuropeptide expression of dental afferent neurons may lead to increased pain sensitivity [181]. Thus, stimulations of tooth by any type of stimulus result in a painful sensation, unlike in other tissues in the body [23, 36].
Although the sensory nerves even in completely healthy pulp may deliver pain sensation, inflammation intensifies those sensations. Nociceptor sprouting has been characterized as an early neural reaction to dentin injury, related to reactionary but not reparative dentinogenesis [23, 40]. Analyses of single nerve fibers in animals show expansion of dentinal receptive fields for A-fiber activation [150] as well as central plasticity [181] under those conditions.
Dentinal hypersensitivity is due to the activation of nociceptive trigeminal ganglion neurons that innervate dental pulp (i.e., dental primary afferents). The exact mechanism of this activation is not perfectly clear. The hydrodynamic mechanisms, in which tubular fluid movements are mechanically detected by nerve endings near the dentin- pulp border, has been widely accepted and dominating theory behind dental nociception. However, accumulative evidence of other potential mechanisms has led to other possible theories: neural theory, with nerve endings in dentinal tubules directly responding to external stimuli; and odontoblast transducer theory, where odontoblasts them- selves may act as pain transducers [36, 126] (Figure 2.28).
Instead of mere mechanical stretching, mechanoreceptors of nociceptive afferents in dentinal tubules offer a potential mechanism in the pain delivery mechanism in hydro- dynamic theory (Figure 2.28). Even normal chewing forces may create sufficient fluid flow to excite putative mechanoreceptors [163], especially in horizontal loading [191]. Especially transient receptor potential vanilloid 1 and 2 (TRPV1, TRPV2), transient receptor potential ankyrin 1 (TRPA1), and transient receptor potential melastatin 8 (TRPM8), expressed in dental afferents [36, 105, 106], are good candidates. The significant increase of TRPA1-positive axons in the painful irreversible pulpitis [106] and the role of TRPA1 in perception of cold [103] further indicates the importance of this receptor in dental hypersensitivity.
Figure 2.28 Molecular mechanisms of hydrodynamic, odontoblast transducer and neural theories of dental nociception. Fluid movement initiated by diverse external stimuli eventually activates mechanoreceptors in dental primary afferents. In neural theory, nerve afferents in the dentinal tubule are directly activated by external stimuli. Candidates of mechano- and thermosensitive molecules are listed. Adapted from [36].
The expression of voltage-gated Ca2+, Na+ and K+ channels, Ca2+-activated K+ channels, store-operated calcium channels and Na+/Ca2+ exchanger [7, 8, 26, 89, 120, 125, 126, 182, 222] indicate the excitability of odontoblasts. Indeed, action potentials can be evoked in odontoblasts by electrical stimulation in vitro [8, 91, 126]. Human odontoblasts also have electrical coupling, allowing information transmission between large number of neighboring cells after electric stimulation. This way the receptive field to external irritation may be significantly larger that the area of irritation [91]. Human odontoblasts express at least TRPV1, TRPV4, TRPA1, TRPM8, mechanosensitive TWIK-Related K+ channel (TREK-1), pH- sensing epithelian Na+ (EnaC) channels and cannabinoid receptor CB1 [35, 49, 50, 106, 125, 168, 188, 195]. Since synaptic structures between odontoblasts and nerve afferents do not exist, ATP [37, 49, 118, 182] and glutamate [35, 107, 152] have been proposed as signal mediators between odontoblasts and neural afferents. Purinergic P2X ATP receptors [6, 99, 105] and mGluR5 glutamate receptors [107, 152] are present in nerve fibers in the pulp, subodontoblastic plexus of Raschkow, odontoblast layer and even in dentinal tubules.
Another possible mechanism for sensory function for the odontoblasts is primary cilia. Primary cilia are single non-motile flagellar organelles present on nearly all vertebrate cells, where they serve a diverse set of signaling functions [63]. Primary cilia also function as flow sensors in osteoblasts, osteocytes, and chondrocytes [128, 223]. In bone cells, cilia are responsible for changes in cellular activity [128], suggesting a role in bone remodeling. Odontoblasts express primary cilia [39, 123, 199], where they may participate in odontoblast polarization and terminal differentiation and in signals that influence cell movement toward the pulp [39, 63, 123]. The cilia may respond to mechanical stimulus from dentinal fluid movements [124] or other signals from either dentin or pulp extracellular milieu or both. In odontoblasts, primary cilia locate in apical pole of cells in vivo [39, 199], as is the case also with bone cells [128]. The putative sensor function, the intimate relationship between odontoblast primary cilia and nerve fibers (Figure 2.29), and the capacity to generate action potentials indicate the possible role of odontoblast primary cilia in tooth pain transmission [199].
The neural theory is based on the dental afferent expression of receptors that take part in the transduction of a specific stimulus to electrical impulses [36]. In coronal dentin, especially in the pulp horn area, afferent nerves penetrate into dentinal tubules to approximately 100 ^m depth [23, 27]. The presence of nociceptive and thermosensitive TRP receptors in dental afferent neurons would facilitate the direct pain sensation without need for hydrodynamic mechanical or odontoblast-transduced stimulus (Figure 2.28).
2.6 Age-related Changes in Dentin-pulp Complex
Even though the capacity for dentin formation remains throughout life, even odontoblasts show signs of aging. The first signs occur during the transition between primary and secondary dentinogenesis, when dramatic changes occur in the odontoblast phenotype [39] (Figure 2.30), related to differential transcriptional activity [185] and increase of odontoblast apoptosis due crowding [56, 137]. Mature human odontoblasts also develop an age-related autophagic-lysosomal system with autophagic vacuoles to facilitate the turnover and degradation of cellular components [39]. Autophagy is a housekeeping process with an “anti-aging” function active in most long-lived cells, consisting of self- digestive pathways mediated by lysosomes to maintain cellular homeostasis. In odontoblasts, autophagic activity has been proposed as essential survival mechanisms. Autophagy could also constitute an alternative mechanism of cell death, named “autophagic type II programmed cell death” (Type II PCD) [39].
Figure 2.29 Primary cilium in human odontoblasts in vivo, confocal laser microscopy (a, b, c, e) and TEM analysis (d). (a) Arrow indicates odontoblast (od) primary cilium axoneme (bar: 10 pm). (b) Green color indicates rootlet in cilium basal body and pinkish-red the cilium; arrow shows nerve fiber; p: pulp core, *: dentin (bar: 10 pm). (c) Close relationship between primary cilium axoneme ("tail") (c) and a nerve fiber (arrow); green indicates cilium basal body at the odontoblast cell membrane (bar: 1.25 pm). (d) TEM image of a primary cilium stemming via the basal body from the odontoblast in close contact with a nerve-like structure (arrow) (bar: 0.20 pm). (e) Intimate contact between a ciliary structure (red; arrowhead) and a nerve fiber (green; arrow) (bar: 0.65 pm). (Reproduced from [199] with permission from Journal of Dental Research.)
As the pulp ages and reparative dentin gradually accumulates, pulp tissue tends to become more fibrous and the number of the odontoblasts and fibroblasts decreases [42, 87, 146]. Reduction of both the odontoblasts and fibroblasts is more pronounced in root than in coronal pulp [42, 146]. The capillary endothelium shows morphological and cytoskeletal changes, including increased trans- end othelial transport [52]. The innervation decreases and shifts its cytochemistry [23]. A comprehensive comparison of gene expression between young and old teeth indicated reduced expression of the functional gene groups involved with cell and tissue development, cell growth and proliferation, and hematological and immune system development and function in aged teeth. Conversely, the expression of genes involved with apoptosis were increased [210]. Overall, these changes may reflect a reduced ability of regeneration and repair in case of tissue destruction.
Figure 2.30 Upper panel: Schematic presentation of the age effects on odontoblast morphology. (a-c) Light microscope images of the coronal odontoblastic layer from 15-year (a), 25-year (b) and 75-year (c) patients' teeth. Odontoblasts in young and young-adult individuals are columnar cells with large autophagic vacuoles (a, b, arrows), while in older teeth odontoblasts become shorter and flattened with dense deposits accumulated within autophagic vacuoles (c, arrows). (d-f) TEM images of the primary cilium (PC) with the respective ages, showing that PC is a persistent structure in odontoblasts. C, centrosome; D, dentin; GC, Golgi complex; PD, predentin. Scale bars: 50 pm (a-c) and 0.5 pm (d-f). (Reproduced from [39] with permission from Journal of Dental Research.)
2.6.1 Age-related Changes in Dentin
In terms of clinical endodontology, the most important age-related change in dentin-pulp complex is the gradual obliteration of the pulp chamber and root canals even in intact teeth [5, 130, 146, 167] because of the slow secondary dentin formation (Figure 2.31). Relative increase in dentin thickness is higher in the root than in coronal area [146]. External irritation by caries, restorative procedures etc. may naturally speed up the process significantly. In incisors, canines, and premolars the obliteration initiates from the coronal direction, but in molars also the pulp chamber floor dentin frequently grows towards the roof. The obliteration may make the location of the root canals difficult [170] and the initial root canal preparation and creation of the glide path more challenging.
The effects of aging on dentin mechanical properties have been debated for decades, but the more recent studies strongly indicate that aging induces changes in the strength and resilience of mineralized dentin. While dentin tensile strength may increase with the occlusion of tubules [129], it occurs with the cost of reduced flexural strength [184]. The most important aspect is the increased mineral- to-collagen ratio due to the peritubular dentin occlusion of dentinal tubules [11, 144, 184] in aged dentin, which increases the hardness, especially in outer dentin [144]. As a result, the fatigue crack growth exponent is about 40% lower [13], the endurance strength about 48% lower [12], and the fatigue crack propagation over 100 times faster [13] in old than in young dentin. Dentin flexure strength reduces approximately 20 MPa/decade, and correlates well with tubular occlusion [11, 184] (Figure 2.32). The tubules in root dentin are similarly occluded [198] (Figure 2.33).
Figure 2.31 Reduction of the pulp space with age. PHr is the pulp volume percentage of the total and hard tissue volume. Data adapted from [5] and [167].
Age
Figure 2.32 The change in dentin tubular lumen dimensions and the influence of age on the dentin strength in human third molars. "Cuff" indicates peritubular dentin. Data adapted from [11].
Age
Figure 2.33 Relative mean dye penetration (in percentage of complete dentin area) after extensive incubation of Methylene blue in instrumented root canals. Data adapted from [198].
Human root dentin may have higher flexural strength than coronal dentin [51, 184], but is similarly reduced with age [184, 228] along with increase in mineral content and hardness [226].
Age-related changes in the organic components seem also to contribute to the dentin mechanical properties [12, 184, 228]. Aged dentin is more cross-linked [228] and has high levels of pentosidine cross-links of collagen [184]. This non-enzymatic advanced glycosylation end-product (AGE) - cross- linking between individual collagen molecules stiffens the collagen matrix, making it more fragile, and participates in low bone strength in osteoporosis patients [177, 215]. Also in human dentin, high AGE levels significantly reduce dentin flexural strength and contributes to the lower mechanical strength of aged dentin both experimentally [139] and in vivo [184].
Other changes in dentin organic matrix with age may also occur: decrease and loss of matrix-degrading enzymes has already been demonstrated [132, 151, 196], which may implicate also changes in their substrates, namely collagen and non-collagenous proteins.
2.6.2 Caries-affected Dentin
Caries damage develops and increases with age and has clinical significance in relation to tooth restoration. Minimally invasive restorative dentistry aims to avoid unnecessary removal of tooth structure, and cavity preparation is limited to the removal of caries- infected dentin, leaving the restoration to be adhesive-bonded to caries-affected dentin. Caries-affected dentin has lower mineral content and altered structure and composition of dentin organic components, including collagen. These changes reduce dentin hardness, stiffness, tensile strength, modulus of elasticity and shrinkage during drying [203]. As a result, dentin in and under the composite-dentin interface is more susceptible to cohesive failures due to the polymerization stress and occlusal forces [202]. Even short exposure of dentin to lactic acid (the most important acid produced by cariogenic bacteria [34]) significantly reduces dentin fatigue strength, increases the crack extension rate, and reduces the fatigue crack growth resistance [44, 155]. Since fatigue crack is a precursor to unstable fracture, lactic acid exposure increases the probability of restored tooth fracture at lower occlusal forces [155]. Especially the endodontically treated teeth are more prone to fractures because of the weaker structure due to loss of tissue, but also because the incremental crack extension occurs with significantly lower cyclic stresses in deep compared to superficial dentin [92]. To avoid catastrophic, unrepairable tooth fractures, restorative procedures should be performed in a manner to protect and preserve the remaining tooth structure, especially after endodontic treatment.
2.7 The Periodontium
The periodontium includes the gingiva, the periodontal ligament (PDL), the alveolar bone, and the dental cementum (Figure 2.34). Its main function is to provide an attachment for the teeth to the alveolar bone. Periodontium is actually a fibrous joint of the gomphosis type, where a conical process (root) is inserted into a socket via a fiber ligament (i.e. PDL). In humans, periodontium is the only gomphosis-type joint and allows minor adjustments in the position of the teeth. Thus it is a resilient suspensory apparatus that provides optimal conditions for masticatory functions. The apical periodontium, including the PDL, cementum and the alveolar bone, is of prime importance for endodontology and these parts of the periodontium will be described in some detail.
2.8 The Periodontal Ligament (PDL)
The PDL is a dense connective tissue with islands of interstitial loose connective tissue interspersed between the dense bundles of collagen fibers (Figure 2.2, Figure 2.34). The principal fibers of the PDL extend from the cementum to the alveolar bone. However, each fiber does not reach the entire distance between cementum and bone, and collectively they constitute an intricate branching and reuniting pattern of fibers. They are embedded deep into the two mineralized tissues as Sharpey's fibers (Figures 2.34 and 2.35). The fibers inserting into cementum are smaller and more numerous than those entering into alveolar bone.
Four groups of principal fibers are present in the PDL. The alveolar crest fibers pass downward from the cementum in the cervical region of the tooth to the alveolar crest; the horizontal fibers comprises the cervical third of the PDL; the oblique fibers run from the alveolar bone somewhat apically to the cementum; and the apical fibers radiate from the cementum towards the alveolar bone in all directions.
Figure 2.34 Radiograph (a) and microradiographs (b, c) of alveolar bone (AB) in undermineralized (a, c) and demineralized sections (b). Note islands of blood vessels and loose connective tissue (CT) among the fibers of the periodontal ligament and radiolucent Sharpey's fibers that attach to alveolar bone (AP, alveolar process of the mandible).
Figure 2.35 Electron micrograph showing fine collagen fibers inserting into a cellular cementum as Sharpey's fibers. (Reproduced from [58] with permission from Acta Odontologica Scandinavica.)
The principal fibers of the PDL have a functional arrangement in that groups of fibers act against different types of forces, including resistance to rotation of the teeth. Although collagen is inelastic, slight movement of the teeth is possible due to the wavy course of the fibers, allowing them to stretch during stress. Changes in the blood flow and blood pressure of the PDL induce minor movement of the teeth [1]. The blood and tissue fluid also act as a hydrodynamic system that absorbs occlusal forces.
2.8.1 Cells of the PDL
Fibroblasts are the prevailing cells of the PDL. Those that are located between the principal fibers are long, slender cells, but those in the interstitial tissue are irregular or stellate-shaped. The function of the fibro- blasts is to maintain the collagen fibers and the glycosaminoglycans and glycoproteins that constitute the ground substance during the normal turnover processes and during repair. Macrophages and mast cells, including associated cytokines, are also found in the normal PDL and they increase markedly during inflammation. Cementoblasts locate on the cementum side and osteoblasts on the alveolar bone side of the PDL. Osteoclasts and odontoclasts allow bone and tooth resorption. They play important roles in the turnover of the PDL, during periodontal, including apical, disease and during orthodontic movement of teeth. They also play an essential role in the development of apical periodontitis. Odontoclasts and osteoclasts also play a central role in shedding of primary teeth and during eruption of the permanent teeth.
Sympathectomy increases osteoclast- mediated bone resorption [113] and root dentin resorption [78], indicating that sympathetic nerves have an inhibitory effect on osteo- and odontoclasts. Sympathetic nerves in the PDL are also important for the recruitment of granulocytes as demonstrated by experimental tooth movement [78]. In experimentally induced periapical lesions, sympathetic nerves have an inhibitory effect on the size of the lesion, the number of osteoclasts lining the lesion and the amount of IL-1a within the lesion [79].
2.8.2 Epithelial Cell Remnants
Both the developing and mature teeth are surrounded by a continuous, net-like pattern of epithelial cells originating from the Hertwig's root sheath. These so-called Malassez's epithelial rests (Figure 2.2) are located close to the root cementum in its entire length, and persist within the periodontal membrane throughout the life of the tooth [225]. The reason why the epithelial rests of Malassez persist in the PDL is unknown, but it has been proposed that these cells are important to prevent ankylosis and hinder bone ingrowth [225]. Stimulation of these epithelial cells may induce cell proliferation [66] that may form cyst linings at the periphery of lesions. These epithelial cells undergo apoptosis, which, together with proliferation, may play a role in the decrease and/or turnover of the epithelial cells of Malassez's rests in the periodontium [32].
2.8 The Periodontal Ligament (PDL)
Malassez's cells may function as targets for developing periodontal nerves. Ruffini-like receptors and free nerve endings relate close to these cells (Figure 2.36) [81]. Furthermore, immunohistochemical studies have shown that Malassez's cells contain neuropeptides such as CGRP and SP and may thus have functional endocrine roles [81]. Malassez's epithelial rests may also play a role in the formation of cementicles by undergoing mineralization. They may be attached or free and may contain cell rests much like pulp stones which may be found in the apical area of the teeth (Figure 2.37).
2.8.3 Turnover
It is important to note that bone is a more dynamic tissue than cementum. The normal turnover of bone also affects the alveolar processes. Cementum is usually covered by a thin, unmineralized precementum, and slow cementogenesis compensates for wear of the teeth. Bone, on the other hand, is covered by osteoid only during bone formation. Unmineralized matrix tends to resist odon- toclastic activity [190]. Thus, osteoclastic activity seems to predominate over odonto- clastic or cementoclastic activity in the PDL and Howship's lacunae in bone defects develop more commonly than tooth resorption. Both resorptive processes may take place simultaneously. Reparative cementum formation by cellular cementum may fill in resorption defects.
The difference in resorptive activity between bone and cementum in the PDL is the basis for orthodontic tooth movement. Moderate forces applied to a tooth will result in bone resorption on the pressure side and bone formation on the tension side without resorption of the cementum. Thus, the tooth will move in the direction of the force applied. The use of excessive force may also result in resorption of the cementum and may reach clinically significant proportions.
Figure 2.36 (a) Apical periodontal ligament of cat incisor richly supplied with protein gene product (PGP) immunoreactive nerves (arrows) from apical bone (B). (b) Enlargement of framed area showing nerve fibers supplying the blood vessels (V) and extensive branching of nerves towards the root cementum (C) and Malassez's epithelial cells (M) (D, dentin). (Reproduced from [81] with permission from Acta Odontologica Scandinavica.)
2.8.4 Circulation in the PDL
The blood supply to the PDL is complex. Although tendinous, the PDL is highly vascularized. The total vascular volume has been calculated to be approximately 20% of the tissue, compared to only 3-4% in most other tissues. PDL receives its blood via vessels from the alveolar bone, periosteum, gingiva, and pulp. The main vessel supply originates from the intraosseous arteries. As the arterial supply arises from different sources and the venules and veins drain both into the bone marrow and gingiva, the vascular bed of the PDL should not be regarded as an isolated functional unit. This implies that inflammatory and pathological changes in blood flow, tissue fluid pressure, or blood pressure in the surrounding adjacent tissues, will also influence the periodontal circulation. Inflammatory vasodilation in parallel- coupled vessels in the alveolar bone or gingiva may cause reduced blood flow in the PDL due to a fall in pressure in the arterioles feeding the PDL.
The main vessels of the ligament run parallel to the long axis of the tooth in compartments of loose connective tissue between the fibers. The arterioles branch to form capillaries arranged in a flat, basketlike network that surrounds the root surface. The capillary network is located closer to the alveolar bone than to the cementum, and the vessels perforating the alveolar walls are most abundant in the apical third.
Blood flow in the PDL seems to be con- trolled mainly by sympathetic fibers causing vasoconstriction and by sensory fibers for vasodilation [104]. However, due to the lack of suitable methods to measure blood flow in a tissue with such a tiny volume and a position secluded between bone and tooth as the PDL, reliable quantitative measurements of blood flow in the PDL are lacking.
Figure 2.37 (a) Attached (AC) and (b) free cementicles (FC) in the periodontal ligament (C, cementum).
Qualitative measurements of PDL blood have shown that sympathetic vasoconstrictor fibers take part in regulation of PDL blood flow. The constrictor effect is greatly reduced, but never abolished, by alpha-adrenoceptor antagonists, indicating that some of the sympathetic fibers innervating the PDL contain NPY, which has been con- firmed by immunohistochemical studies [81, 157] (Figure 2.36). Sympathetic induced vasodilations, due to activation of beta receptors that are most probably located in postcapillary resistance vessels, have also been found in the PDL [1].
Changes in PDL blood flow affect the tooth position, and external forces applied to the tooth crown may greatly influence PDL blood flow [29, 161]. Such changes in the normal conditions may possibly be related to changes in PDL tissue fluid pressure [111]. This pressure has been recorded as relatively high compared to most other tissues [111, 161]. It is claimed to affect tooth position, blood flow, the eruptive force of teeth, and probably pain sensation. In common with the pulp, also the PDL is enclosed in a rigid low-compliance environment between alveolar bone and tooth. Changes in blood volume induced by venous stasis or cardiac arrest are thus rapidly transmitted to the PDL tissue fluid pressure [111].
Unlike the pulp, where the main sensation is pain, the PDL also recognizes touch, pressure, movements, and position of teeth, in addition to pain. A variety of Ruffini-like mechanoreceptor structures are found in the PDL [22]. Some of the fully encapsulated mechanoreceptor structures locate in the interstitial loose connective tissue next to blood vessels. Thus, increased PDL blood flow causing increased tissue pressure would most probably cause excitation of mechanoreceptors. An increased blood volume in the PDL raises the tissue pressure and causes tooth extrusion, whereas decreased blood volume causes tooth intrusion and reduces the tissue pressure [1, 111]. Another aspect of the low compliance in the PDL is that changes in tissue pressure most likely will affect pain sensation, i.e., increased tissue fluid pressure causes increased activity in sensory A-delta and C fibers, much the same way as in the pulp.
2.8.5 Innervation of PDL
Although less innervated than the dental pulp, the PDL is richly supplied with nerves entering both from the apical and lateral alveolar bone (Figures 2.36 and 2.38). The apical part of the ligament is most heavily innervated and the major part seems to be of sensory origin, whereas sympathetic, NPY- carrying fibers are rarely found. However, some larger vessels in the mandibular canal are densely innervated by NPY fibers in their walls [81]. Sprouting of sympathetic NPY- containing nerves may be observed in the inflamed PDL after pulp exposure [79]. The significance of this sprouting is not clearly understood, but it may affect blood flow and immunomodulation. Since peripheral endogenous sympathetic neurotransmitters are important regulators of vascular growth factors [30], it might suggest that sympathetic nerves play a role in revascularization during repair and healing processes in the inflamed PDL.
Figure 2.38 Section through the periodontal ligament from apical third of cat canine showing numerous nerve fibers (arrows) approaching the ligament from alveolar bone (AB) (D, dentin; C, cementum). (Reproduced from [81] with permission from Acta Odontologica Scandinavica.)
The periodontal ligament contains myelinated and unmyelinated nerve fibers. CGRP fibers appear more frequently than SP fibers at all levels in the PDL [81]. Most nerves in the PDL localize in the apical third, closely associated with blood vessels. They are frequently observed on the exact border between PDL and cellular cementum, where some form round, coiled, nerve-like endings. Others are closely associated with periodontal epithelial rests of Malassez's cells (Figure 2.36) [81, 122]. In the apical part of cat PDL, immunoreactive cells forming a net-like pattern are regularly displayed in Malassez's epithelial rests surrounded by numerous nerve fibers. Some of the cells located in Malassez's rests contain CGRP and SP. Thus, in common with specialized epithelial cells from other locations [148], the Malassez's epithelium may comprise cells that could be classified as endocrine cells due to their content of neuropeptides. Periapical inflammation and orthodontic tooth movement induce a transient periapical sprouting of sensory axons [25, 108, 214]. Larger axons often form specialized terminals, predominantly in the apical area, described as Ruffini-like endings. Thin axons usually terminate as free nerve endings. Both electrophysiological and histological data suggest that the Ruffini-like terminals as well as free nerve endings may function as mechanoreceptors [122]. Cementum appears not to be innervated [81], but extensive branching of sensory nerves is often found adjacent to the apical cellular cementum, where few blood vessels are located (Figure 2.36).
2.9 Cementum
Cementum is avascular, mineralized connective tissue that covers the root of the tooth. The main function of cementum is to attach the principal fibers of PDL to the tooth. Generally, the coronal half of the cementum is acellular while the apical part is cellular and has cementocytes embedded in its matrix. The acellular cementum is thin, about 50-200 pm thick [227]. The cellular cementum is thicker and usually multilayered, with individual layers of 10-100 pm thickness [227]. Acellular cementum is important for tooth support while cellular cementum has a role in adjustment of tooth position after eruption.
Cementum-dentin junction (CDJ), a region where cementum attaches to dentin, is a 100-200 pm thick interspace with a 10-50 pm hygroscopic proteoglycan-rich layer [83, 227]. The tight attachment of cementum to dentin at CDJ is mediated by bridges of continuous collagen fibers [83, 208] (Figure 2.39). The hygroscopic CDJ has also been suggested to be a gomphosis, a fibrous joint between cementum and root dentin that is capable of accommodating functional loads in a way similar to that between cementum and alveolar bone [83, 84].
Age-related changes occur in physico-chemical properties of cementum, suggesting cementum as adaptive in nature [96]. Cementum hardness increases with age, but the width of the CDJ decreases, as measured from the cementum-enamel junction (CEJ) to the tooth apex [96]. The thickness of the cellular cementum increases with age. Compensatory cementum deposition occurs in the apical area to counterbalance occlusal attrition. Occasionally cementum formation may exceed this physiologic limit and result in hypercementosis, which may affect a single tooth or all the teeth. Local abnormal thickening of the cementum may be found in connection with chronic periapical inflammation. A more generalized hypercementosis may be associated with certain systemic disorders. Cementomas, cement-producing tumors, have also been described. If a root fractures, cementum may form between the root fragments and at the peripheral site of the fracture. If root resorption occurs, repair of the defects by cellular cementum formation may take place.
Figure 2.39 FE-SEM images of human cement- dentinal junction (CDJ). (a) 10 to 15 pm cementum layer in intimate contact with dentin. Magnification = 500x; bar = 10 pm. (b,c) Higher magnification demonstrates the mineralized collagen fiber continuity from cementum to underlying dentin. Magnifications = 2,500x (b) and 5,000x (c); Bars = 10 pm (b) and 1 pm (c). (Reproduced from [208] with permission from Endodontic Topics.)
Cementicles are mineralized structures that may be found freely residing in the PDL or attached to the root surface (Figure 2.37). They may be formed by mineralization of degenerating epithelial rests or from thrombosed vessels. Cementicles contain bone sialoprotein (BSP) and osteopontin, two non- collagenous matrix proteins typically found in bone and cementum [17]. When present, cementicles are often found on most teeth.
2.9.1 Cement Structure and Formation
Collagen fibers are major organic components of cementum. Two types of collagen fibers are present in cementum, Sharpey's fibers and matrix fibers. Fine Sharpey's fibers (or extrinsic fibers) represent the termination of the principal fibers of the PDL and penetrate the cementum (Figure 2.35). The matrix fibers are oriented parallel to the root surface and they are interwoven with the Sharpey's fibers. The Sharpey's fibers are formed by the fibroblasts in the PDL while the matrix fibers are formed by the cementoblasts. When the position of the tooth is altered, e.g., during tooth eruption or during orthodontic tooth movement, new attachment of periodontal fibers will take place and the fibers in the cementum orient at different angles to the surface. In cellular cementum, the layers of cellular intrinsic fiber cementum form an alternating lamellar pattern of two types of lamellae, transversely and longitudinally arranged, and this arrangement may be controlled by cementoblasts [227].
The cellular components of cementum include cementoblasts and cementocytes. The cementoblasts, which line the root surface, have all the ultrastructural characteristics of cells capable of synthesizing collagen and protein-polysaccharide complexes. During the formation of cellular cementum at the apical portion of the root, some cementoblasts progress to become cementocytes. The cementocytes are harbored in lacunae, which are embedded in the matrix of the cellular cementum. The functions of cementocytes are not known, but they have recently been compared to osteocytes; cementocytes and osteocytes harbor several commonalities in transition, communication, and specific markers, suggesting that cementocyte may have similar dynamic functions to osteocyte [234].
The ground substance of interfibrillar matrix contains proteoglycans and glycoproteins similarly with other periodontal tissues. Two major constituents are BSP and osteopontin. Collagens have been characterized better than other components in cementum, but there are new developments in the composition of extracellular matrix. Recent mice experiments revealed a novel structural component, Fibulin-4, expressed in dental cementum [180]. Proteomic analysis from mouse dental cementum identified a protein- protein interaction network that includes indicators of metabolic function, possibly reflecting the activity of cementocytes [178].
The precementum is a thin unmineralized layer, which covers the cementum. It pre- vents, up to a certain point, the root from resorption. If it mineralizes, root resorption may occur.
2.10 Alveolar Bone
The alveolar bone is that part of the alveolar processes of the mandible and maxilla that lines the alveoli for the teeth (Figure 2.40). It has all the features of cortical bone but is further characterized by harboring the Sharpey's fibers from the PDL. Alveolar bone has numerous channels for blood and lymph vessels (Volksmann's canals) from the cancellous bone to the PDL and is therefore often referred to as the cribriform plate. Despite all the channels passing, it appears as a radiopaque line on clinical radiographs (Figure 2.41a), which gives it the name lamina dura. It is an important diagnostic landmark. A breach in its continuity on radiographs may be a sign of resorption, often associated with pulp infection or periodontal disease. However, the degree of mineralization of the alveolar bone is no different from that of the rest of the cortical bone in the alveolar process (Figure 2.41b). The tangential superimposition of the alveolar bone on radiographs gives the lamina dura its characteristic radiodensity.
The buccal and lingual laminae of the alveolar processes of the mandible and maxilla vary depending on the location within the jaw. The buccal (vestibular) lamina is usually thinner than the lingual (palatal) lamina, except in the mandibular molar region, where the lingual lamina is thinner. This relationship is important to keep in mind when teeth are extracted and for draining abscesses from periapical areas since they tend to follow the path of least resistance. The close relationship between the alveoli and the maxillary sinus (Figure 2.42a) is also of importance during surgical extractions, implant placement, and endodontic procedures. (Figure 2.42b)
Figure 2.40 Alveolus with Volkmann's canals (VC) perforating the alveolar bone (AP, alveolar process of the mandible). (Reproduced from [140] with permission from Munksgaard.)
2.10.1 Alveolar Bone Structure
The alveolar bone has all the basic characteristics of bone tissue, including osteoblasts, osteocytes, and osteoclasts (Figure 2.43). Its development is closely associated with the presence of teeth. If teeth are lost, the alveolar bone undergoes resorption. If teeth do not erupt, alveolar bone will not develop.
Figure 2.41 (a) Radiograph of a molar tooth showing the radio-dense alveolar bone (arrows) which gives it the name lamina dura. (b) Microradiograph of a ground section showing dentin (D), cementum (C), periodontal ligament space (PDL) and alveolar bone (AB) (VC, Volkmann's canal).
Figure 2.42 (a) A split maxilla at the level of the hard palate showing apices of the roots of the first permanent molar (arrows) extending into the maxillary sinus. (Reproduced from [140] with permission from Munksgaard.) (b) A dissected specimen from a series of experimental endodontic procedures on monkeys showing perforation into the maxillary sinus by gutta-percha points. (Courtesy of Drs D. 0rstavik and I.A. Mjor.)
Osteoblasts are matrix-producing cells, with a well-developed Golgi apparatus, granular endoplasmic reticulum, and mitochondria. Osteoblasts that become embedded in the bone matrix are referred to as osteocytes. They lose many of their organelles, but they maintain cytoplasmic contact with neighboring osteocytes. Osteocytes are significant in controlling responses to mechanical forces and therefore may be central to tooth eruption, physiological and pathological tooth movement, and in orthodontics [21]. Collagen formation may take place in the periosteocytic space between the osteocytes and the wall of the lacunae. Mineralization of the lacunae may occur, resulting in “plugged lacunae" Osteolysis may also occur in the lacunae and it represents a part of the mineral metabolism of bone tissue [224]. However, the main part of the turnover associated with bone remodeling involves osteoclastic activity (Figure 2.43) and new bone formation by osteoblasts.
Figure 2.43 Demineralized section showing human alveolar crest and adjacent tissues (PDL, periodontal ligament; OB, osteoblasts; SF, Sharpey's fibers; LP, lamina propria of attached gingiva; BL, bone lacunae with osteocytes; OC osteoclast; IL incremental lines; RL, reversal lines). (Courtesy of Dr K. Reitan.)
The normal turnover rate results in bone tissue of different ages in the alveolar processes. Osteons with different degrees of mineralization are present at any given time. Bone lamella, which are the incremental growth lines of bone tissue, are discernible. Osteoid is present anywhere bone formation takes place, but it does not cover fully formed bone. The unmineralized core of the relatively thick Sharpey's fibers inserting into the alveolar bone gives it a striated appearance in microradiographs (Figure 2.34c).
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