Kerstin M. Galler
8.1 Principles of Regeneration and Repair
Root canal treatment can prevent tooth loss by eliminating pulpal and periradicular disease; however, it is not a biology-based approach. Advances in pulp biology and tissue engineering challenge the traditional concept of replacing lost pulp with inert synthetic materials. Regenerative strategies aim at the production of new vital tissue that resembles the original in architecture, structure and function. As for the dental pulp, it has been known for decades that this tissue possesses regenerative capacity, and dentistry has been pioneering regenerative approaches with the use of agents such as calcium hydrox- ide to promote healing after pulp-capping procedures [41]. Such therapeutic measures are mainly empirical, but as we unravel the biological basis and start to comprehend the mechanisms underlying regeneration and repair, regenerative medicine offers great scope also for the field of endodontics.
The term ’regeneration’ refers to the recreation of the original architecture, form, and function of a specific tissue type; ’repair’ denotes healing by formation of a tissue that has partially lost the biological function of the original tissue. The distinction between repair and regeneration is often difficult in the clinical setting. Moreover, as biological processes, the two overlap in most situations. A complex fracture of the long bones will typically heal by virtually complete regeneration of the bone itself, but the surface wound will heal by fibrous tissue formation and a skin scar.
8.1.1 Pulp and Periapical Regeneration Processes
Periradicular bone possesses the capability to regenerate fully after removal of the inflammatory trigger. Healing of bony lesions is a major goal of endodontic therapy and it occurs predictably if the clinician succeeds in eliminating or sufficiently reducing bacterial activity within the root canal system. Thus, the connective tissues of the periapical bony structures have an innate capacity, with an adequate supply of multipotent cells, for complete regeneration of the periodontal ligament and the alveolar bone in the right circumstance.
Pulp tissue can also regenerate; however, it remains a challenge to elucidate exactly under which circumstances this may take place. It is important to identify the cell sources that may be activated for regeneration. Remaining islands of vital pulp can form the point of origin for regenerated pulpal tissue [77]. Stem cells of the apical papilla in immature teeth is another source of cells that can proliferate, differentiate, and form odontoblasts and pulp tissue [44]. These cells may cause regeneration of the pulp after revitalization procedures. In teeth with incomplete root formation and pulp necro- sis, revitalization can be performed as an alternative treatment to apexification, where provocation of bleeding into the canal flushes in the respective cells from the apical papilla [54]. Even in teeth with complete root formation, the periradicular tissues comprise mes- enchymal stem cells that can be delivered into the canal [14]; however, it is uncertain whether formation of a pulp-like tissue is biologically possible after pulp necrosis in mature teeth. The tissue formed inside the root canal after revitalization procedures contains elements of pulp tissue (fibroblasts, connective tissue, blood vessels, collagen), but other cell types are missing, among them notably odontoblasts, whereas non-targeted cell types or tissue such as osteoblasts and cementum may be present [56, 97]. Thus revitalization procedures may not so much give healing by regeneration but rather healing by repair.
8.2 Vital Pulp Therapy
Measures such as indirect and direct pulp capping or pulpotomy aim at maintaining tooth vitality and supporting the pulp's inherent capability of regeneration and repair. In either case, the pulp will respond with the formation of tertiary dentine. The generation of tertiary dentine is an active defense mechanism to create a mineralized barrier that separates the tissue from the site of injury; as such, it is a measurable parameter of healing. Tertiary dentine can be either reactionary or reparative. During tooth development, odontoblasts secrete primary dentine at a rate of 4-8 pm per day [49, 57], but they adopt a resting state after the com- pletion of root formation where secretion is reduced to about 0.5 pm per day [18, 79]. Mild stimulation induces an upregulation of odontoblast activity, where primary odonto- blasts increase their secretory activity to the original level, leading to rapid deposition of reactionary dentine, which displays a tubular structure [16, 81]. After therapeutic intervention, reversible inflammatory processes within the pulp tissue are expected to heal by regeneration. With increasing intensity of the stimulus and delayed intervention, healing will more likely take place as repair. If the original odontoblast layer is lost, e.g. after intense stimulation or pulp exposure, it can be replaced after differentiation of stem cells into secondary odontoblasts [16, 22]. These cells deposit reparative dentine, a mineralized matrix that may not exhibit the characteristic tubular structure, but resemble bony tissue and often display cellular inclusions [33]. It remains unclear whether osteodentine formation is particular to the cell source, which is not comprised of original odontoblasts, or to the intensity of stimulation, which leads to hasty deposition of a mineralized tissue that is less organized. Furthermore, tertiary dentine is also characterized by increased peritubular dentine deposition [81], a vital process which has to be differentiated from intra-tubular-calcifications due to physicochemical precipitation of mineral crystals [5]. Thus, the structure of tertiary dentine is highly variable.
8.2.1 Bioactive Materials
Bioactive materials have long been used to induce tertiary dentine formation and to support healing of the dentine pulp complex. A material is termed bioactive if it exerts a positive influence on vital tissues and elicits a desirable biological response at the interface [39]. The response can be indirect, through antibacterial activity, or direct, by interaction with adjacent cells, e.g. stimulation of proliferation, differentiation, and/or biominerali- zation. Calcium hydroxide was introduced in root canal therapy nearly a hundred years ago [40], and studies with this material in contact with the pulp have demonstrated that it enables the tissue to remain vital and to form a mineralized barrier [41]. Calcium hydroxide has been extensively used in endodontics and dental traumatology, and it has been the material of choice for vital pulp therapy for several decades. Due to its high alkalinity, it not only exerts antibacterial and antifungal activity [80], but induces necrosis of adjacent cell layers and an inflammatory reaction in the underlying tissue. Proinflammatory cytokines and chemokines recruit immune cells, which promote healing by clearance of the injury site, and stem cells, which differentiate into secondary odonto- blasts. Furthermore, calcium hydroxide releases growth and differentiation factors that are bound in the dentin matrix [34, 94], which affect and modulate cellular behavior [94, 99]. The use of calcium hydroxide for direct pulp capping has been shown to result in the formation of a mineralized barrier [2, 29], its thickness increasing with longer postoperative periods [29]. Histological analysis after pulp capping shows a superficial layer of tissue debris beneath the calcium hydroxide [29] and an adjacent, mineralized barrier. This barrier may display a tubular structure continuous with the original dentin, only with fewer and more curved tubules [29, 67], but more often it presents as an amorphous and atubular calcified tissue with cellular inclusions [33, 67, 71].
Application of calcium hydroxide thus has several effects that enable healing by regeneration. Negative properties, however, include high solubility and low mechanical stability. Moreover, the mineralized barrier formed may be porous and exhibit tunnel defects [19, 60], and calcium hydroxide appeared to be inferior regarding the thickness of newly formed mineralized tissue compared to Portland-cement-based mate- rials such as mineral trioxide aggregate (MTA) [2, 95]. Due to these drawbacks, calcium hydroxide is progressively displaced by such materials, collectively referred to as hydraulic calcium silicate cements. These cements form a stable calcium-silicate-hydrate matrix; calcium hydroxide and calcium ions are side products of this reaction and create the bioactive effects [9, 10]. Similar to calcium hydroxide, MTA solubilizes dentin matrix proteins [91], which is likely to contribute to the material's bioactivity. Besides increased stability of these materials compared to calcium hydroxide, hydraulic calcium silicate cements offer a wide range of applications in endodontics due to their capability to seal and disinfect and to induce dentinogenic or osteogenic mineralization [21]. Several studies provide evidence that the use of hydraulic calcium silicate cements results in a less distinct necrotic zone, hyperemia, and inflammatory reaction compared to calcium hydroxide, to a more homogenous and solid layer of tertiary den- tin, and to a superior clinical performance regarding failure rates and pulp vitality [1, 42, 64, 89].
8.3 Cell Types Involved in Pulp Healing
Among the cells that constitute the pulp tissue, the odontoblasts are the first target for external stimuli due to their peripheral localization in the dental pulp and their extension into dentine. Odontoblasts are post-mitotic cells, i.e. they are not replaced during the life of the organism under physiological conditions. As such, they share certain features with neurons and myocardiocytes as static cell populations [70, 92]. Stimuli include thermal variations and biomechanical forces, but also molecular products derived from microorganisms.
8.3.1 Tertiary Dentine
An essential feature of pulpal defense is the formation of tertiary dentine. As discussed above, reactionary dentine has to be distin- guished from reparative dentine, as they arise from two different populations of cells and thus their genesis and nature are distinct. Reactionary dentinogenesis refers to the secretion of a tertiary dentine matrix by sur- viving post-mitotic odontoblasts, which increase their secretory activity in response to an appropriate stimulus. Implantation of dentin extracellular matrix components into unexposed cavities in ferret teeth leads to a localized stimulation of reactionary dentine [85]. This can mainly be attributed to the presence of transforming growth factor p-1 (TGF p-1), the most abundant growth factor in the dentine matrix, which is known to markedly upregulate odontoblast secretory activity [81]. Additionally, secretion of matrix at the mineralization front and along the odontoblast process leads to a progressive increase in thickness of the peritubular den- tin, and the gradual occlusion of the dentinal tubules by centripetal deposition of calcium phosphate crystals leads to sclerosis and thus a decreased permeability of dentine [91]. In carious lesions, the demineralization of den- tine induced by bacterial acids and the subsequent solubilization of bioactive molecules, in particular TGFp-1, is considered responsible for initiating the stimulatory effect on the odontoblasts and thus the main cause of reactionary dentine formation. In contrast, reparative dentinogenesis is a more complex biological process. Stronger stimuli will lead to the death of the odontoblasts, but if conditions are favorable, a new generation of odontoblast-like cells may differentiate from stem or precursor cells within the pulp.
8.3.2 Stem Cells: Sources and Activation
A major cell source for regeneration or repair is the pool of stem cells that is present in the dental pulp, papilla and periapical tissues. Stem cells have been isolated from the pulp of permanent [35] as well as deciduous teeth [59] and furthermore from the apical papilla of teeth with incomplete root formation [44, 87]. Most commonly, stem cells are separated from the population of ex vivo cultured pri- mary cells based on presence or absence of specific glycoproteins on the cells' surface. Thus, patterns characteristic for mesenchy- mal stem cells can be recognized, and cells can be sorted by use of specific antibodies.
By definition, stem cells are characterized by their capacity for self-renewal and their ability to differentiate into different cell types. In general, stem cells are involved in regular tissue turnover to replace aged cells, and during repair and regeneration. Located in the perivascular niche [78], where they are kept in an undifferentiated stage, stem cells in the dental pulp remain quiescent until an insult occurs. Stem cells undergo asymmetric division, meaning that one cell gives rise to an identical cell to keep the pool of stem cells constant, whereas the second daughter cell enters the path of differentiation. Chemotactic signaling recruits stem cells to the site of injury, they leave their niche, migrate, and differentiate into secondary odontoblasts [22], cells that can produce a mineralized barrier at the interface of soft and mineralized tissue. Thus, stem cells ful-fil an essential role during regeneration and repair of the dentine pulp complex. See Figures 8.1 and 8.2.
8.3.3 Neurovascular Components in Regeneration
Pulpal inflammation and subsequent healing are also influenced by a complex neurovas- cular relationship and the interplay between the inflammatory process and sensory nerve fibers. Besides nociception, nerve fiber functions in normal and injured tissues include vasodilatation and neurogenic inflammation. Noxious stimuli trigger the release of neuropeptides, in particular substance P (SP), which induce vasodilation, increase pulpal blood flow and vascular permeability, and activate immune cells. The expression of SP in dental pulp is significantly upregulated in carious teeth [74]. Neuropeptides may exert stimu- latory effects on odontoblast-like cells by increasing their expression of bone morphogenetic protein 2 (BMP-2), thus inducing tertiary dentin formation [12]. Furthermore, dental sensory fibers react to damage and inflammation by sprouting of their terminal branches into the surviving pulp beneath the injured site [51]. Increased neuropep- tide levels from sprouting fibers can enhance inflammation through the recruitment of immune cells and the increase of vascular permeability [7, 37]. Denervated teeth show less survival of vascular pulp and accelerated loss of pulp tissue in comparison with innervated pulps [7]. Thus, if the sensory innervation is removed before injury, the pulp is less able to defend itself, to trigger neurogenic inflammation and to heal after pulpal exposure.
Figure 8.1 Perivascular niche and multipotency of mesenchymal stem cells. (a) Mesenchymal stem cells (MSC) reside in perivascular niches where they undergo self-renewal and maintain the surrounding cells or tissue. Under specific signaling conditions, MSC can undergo differentiation into different lineages. From [65]. (b) Immunolocalization of the CD146 antigen, an endothelial surface marker, to blood vessel walls in human dental pulp tissue. (c) Co-localization of blood vessels and mesenchymal stem cells in dental pulp. Dual immunofluorescence staining showing reactivity of an antibody to the mesenchymal stem cell marker STRO-1 labeled with Texas red to a blood vessel to the endothelial marker CD 146 labeled with fluorescein isothiocyanate. From [78].
Figure 8.2 Stem cell types relevant for regenerative processes in the pulp and periapex. BMSC: bone marrow stem cells; iPAPC: inflamed periapical progenitor cells; DPSC: dental pulp stem cells; SHED: stem cells from human exfoliated deciduous teeth; SCAP: stem cells of the apical papilla. Modified from [38].
The role of pulp fibroblasts in the process of defense and regeneration has also been explored. Pulp fibroblasts produce all com- ponents of the complement system and can form the membrane attack complex (MAC), which is effective against cariogenic bacteria [48]. Furthermore, stimulation with bacterial toxins enables pulp fibroblasts to guide nerve sprouting during pulp regeneration through complement system activation and the production of neurotrophic factors [8, 13].
Immune cells are naturally involved in regeneration and repair of the pulp tissue, which will be discussed in more detail below.
8.4 The Role of Inflammation
As carious lesions progress, bacterial toxins and eventually bacteria themselves enter the pulp space via the dentinal tubules [53]. Odontoblasts sense bacterial toxins (such as cell wall components of bacteria) through receptors of the Toll-like (TLR) and nod-like (NLR) families [88], and contribute substantially to the pulpal immune response.
The dental pulp reacts to an infection with an inflammatory response as part of the - initially protective - host response. The most common cause for an invasion of microbes towards the pulp is tooth decay. Owing to its tubular structure, dentine is a penetrable barrier, where permeability increases due to an increase in both tubule density and diameter with decreasing dis- tance from the pulp tissue. Thus, bacterial toxins or components and eventually intact bacteria reach the pulp via the dentinal tubules, and their penetration accelerates with increasing depth. The initial invasion of microbial components and toxins activates innate immunity, which in general is not antigen-specific, but recognizes molecular patterns that are specific to bacteria and ini- tiates their attack and elimination by phagocytotic killing. Due to the unique anatomical location of bacteria in carious lesions, phagocytosis will not take place until the carious front has reached the pulp. However, the innate response of the dentine-pulp complex mobilizes a variety of protective measures, including the reactions of odontoblasts, the production of neuropeptides, the activation of immune cells, and the production of chemokines and cytokines [36].
Generally, the outward flow of dentine liquor due to the positive intrapulpal pressure prevents sporadic migration of oral bacteria from exposed dentine surfaces into the pulp. Thus, the rate of bacterial invasion is significantly lower in vital compared to non-vital teeth [61]. As an initial protective mechanism against caries progression, the rate of outward flow can be increased by vasodila- tation after the neuropeptide release of intra- dental sensory afferent nerves [55, 58].
8.4.1 Pulp Responses to Bacterial Challenges
The odontoblasts are anatomically located in the periphery of the pulp, and their cellular processes extend well into the dentinal tubules. Therefore, they are the first cells to encounter bacterial antigens. In order to sense potential threats, odontoblasts consti- tutively express so-called pattern recognition receptors (PRRs), which recognize and can bind to various bacterial components. Receptor binding leads to intracellular activation of signal transducers and the secretion of interleukins, pro-inflammatory cytokines, and chemokines. These messengers stimu- late the diapedesis of leucocytes and recruit immature dendritic cells [25, 43], which process and present antigens and thus act as messengers between the innate and adaptive immune systems.
The production of the acute-phase, lipopolysaccharide-binding protein (LBP) by odontoblasts contributes to the protective mechanisms. LBPs neutralize bacterial cell wall components and can attenuate the immune response by inhibiting the production of pro-inflammatory cytokines [26]. Challenged with bacterial toxins, odontoblasts also produce vascular endothelial growth factor (VEGF), a potent inducer of vascular permeability and vasculogenesis [90]. Furthermore, antibacterial peptides called defensins are secreted [23]; these exert a broad spectrum of antimicrobial activity as they generate channels or micropores in the cell membrane of microorganisms. Moreover, odontoblasts secrete TGF-p1, which stimulates dentine matrix secretion as well as acting as an anti-inflammatory mediator; its expression increases in irreversible pulpitis [68]. Dental pulp cells may also produce anti-inflammatory signaling molecules, which attenuate the immune response to restrict tissue damage, while simultaneously stimulating odonto- blast differentiation and formation of new dentin [26].
The remaining dentine thickness appears to be critical. Studies assessing the immune response of pulp tissue due to caries show the full spectrum of the cellular host response if the layer of dentine falls below 0.5 mm [15]. Apparently, the progression rate is also of importance, as slowly progressing caries is characterized by a different bacterial flora, color, and consistency compared to rapidly progressing caries [6]. Slow caries progression allows for defensive measures as described above as well as for peritubular mineral depo- sition leading to dentine sclerosis in order to shut the gates for invading microbes.
8.4.2 The Link between Inflammation and Regeneration
Thus the innate immune response offers a repertoire of protective mechanisms that aim at the restoration of pulp homeostasis. Therapeutic intervention and removal of caries and other sources of infection can thereby result in resolution of inflammation and healing. The initial inflammatory response, however, promotes regeneration, and the general importance of this close link was established with the observation that healing after myocardial infarction was compromised by corticosteroids in animals [52]. A prerequi- site for healing is the elimination of bacterial noxa as well as the production of pro-inflammatory mediators by the host. The immune system produces both pro-inflammatory as well as anti-inflammatory signaling molecules. Whereas pro-inflammatory mediators initiate the immune response, anti-inflammatory molecules prohibit an excessive response and lead back to tissue homeostasis. Pathogen removal results in attenuation of the response and remaining toxins will eventually be neutralized [26]. Undoubtedly, if the damage to the dentine pulp complex is severe enough, it cannot be resolved by healing. Increasing numbers of immigrating immune cells induce extensive collateral tissue damage. They produce proteases, which on the one hand enable passage of immune cells through the tissue, but on the other lead to tissue dissolution. Immune cells furthermore secrete reactive oxygen species and enzymes, which not only damage bacteria but also local cells. Signals derived from necrotic cells promote the inflammatory process further and lead to exacerbation. Prolonged stimuli evoke chronic inflammation characterized by moderate immune cell infiltrates, collagen fibrosis, and premature aging of the tissue with reduced host response, and/or it may lead to necrosis and spread of microorganisms into the root canal system and eventually the periapical region.
In contrast to a slow invasion of microor- ganisms towards the pulp in tooth decay, the pulp can be exposed to microbial contamination and infection rapidly after dental trauma and crown fractures. In that case, a healthy tissue with a fully functional immune response is confronted directly with microorganisms. A classic study in monkeys found that the zone of inflammation one week after pulp exposure by grinding or fracture did not extend beyond a depth of some 2 mm [20] without significant differences compared to the penetration depth after 48 hours. The healthy pulp thus has a remarkable resistance to bacterial invasion.
8.5 Signaling Molecules in Dentine
During tooth development, neural crest- derived cells of the dental papilla undergo terminal differentiation into dentine-forming odontoblasts. Upon completion of cytodifferentiation, odontoblasts start their secretory phase and produce an organic template of collagenous and non-collagenous proteins, which later mineralizes with hydroxyapatite crystals to form calcified dentine. During this synthesis process, the odontoblasts not only lay down the predentine, but also express a variety of bioactive molecules, which are secreted into the extracellular space [28, 73, 100]. During mineralization, these bioactive factors become embedded and immobilized in the dentine matrix. Whereas proteins and growth factors in an active form have a short half-life, binding to extracellular matrix components may serve to maintain their bio- activity by protecting them from proteolytic degradation and thus prolonging their life span. Growth factor-binding compounds include mainly proteoglycans [72, 96], but also specific binding proteins [3], glycoproteins such as fibronectin [69], and different types of collagen [66, 86].
As there is no turnover in dentine extracellular matrix, bioactive regulatory molecules can be reactivated much later in life upon release from their bond. During caries, bacterial acids, such as lactic acid, expose the organic component of dentine and release bioactive factors, which may modify the immune response, cell recruitment and differentiation (Figure 8.3) (Table 8.1) [24]. Application of dental materials onto dentine, e.g. calcium hydroxide or hydraulic calcium silicate cements, but also self-etching dental adhesives, release bioactive factors [27, 34, 93]. Organic acids or chelating agents such as ethylenediaminetetraacetic acid (EDTA), which is commonly used in endodontic treatment, are also suitable for dentine deminer- alization. A multitude of bioactive molecules have been identified in dentine extracellular matrix (for review see [84]). These include non-collagenous proteins that regulate mineralization, as well as growth and differ- entiation factors, cytokines, chemokines, and neurotrophic factors. Growth factors present in human dentine extracellular matrix include transforming growth factor pi (TGF-pi), basic fibroblast growth factor (bFGF), bone morphogenetic protein 2 (BMP-2), platelet- derived growth factor (PDGF), placenta growth factor (PIGF), and epidermal growth factor (EGF), in addition to angiogenic factors such as vascular endothelial growth factor (VEGF) [11, 28, 73]. These molecules are effective at very low concentrations and elicit cellular responses even at the picogram level. They modify immune reactions, stimulate angiogenesis, exert chemotactic effects to recruit cells to the site of injury, and promote proliferation, differentiation and mineralization [4, 83, 98]. Dentine matrix proteins and their roles in healing and regeneration are under continuous study, and their bioactive properties are explored for tissue engineering approaches.
Figure 8.3 Simplified schematic drawing of odontoblast responses to carious lesions. Odontoblasts express TLRs (Toll-like receptors) to recognize various bacterial products (red). Receptor binding activates intracellular signaling pathways. Odontoblasts produce antibacterial peptides such as beta-defensin 2, cytokines and chemokines to attract immune cells, growth factors such as vascular endothelial growth factor to increase vascular permeability and transforming growth factor β-1 to induce mineralization. A phagocytotic activity of odontoblasts has been suggested. The response is likely to be modulated by growth factors released from the dentine matrix by demineralization caused by bacterial acids. Based on data in [36].
Table 8.1 Bioactive dentine matrix components and their putative functions, a non-exhaustive overview.
|
Growth Factors and Cytokines |
Function |
|
Transforming growth factor beta 1 (TGF-P1) (Isoforms 1, 2, 3) |
Anti-inflammatory; induction of odontoblast differentiation and matrix secretion; chemotaxis |
|
Basic fibroblast growth factor (bFGF) |
Induction of cell proliferation; chemotaxis; angiogenesis |
|
Vascular endothelial growth factor (VEGF) |
Angiogenesis |
|
Bone morphogenetic proteins (BMPs) (BMP 2, 4 and 7) |
Differentiation; mineralization |
|
Adrenomedullin (ADM) |
Anti-inflammatory; odontoblastic differentiation |
|
Insulin-like growth factor (IGF-1 and -2) |
Proliferation; differentiation |
|
Platelet-derived growth factor (PDGF) |
Regulation of cell growth and division; angiogenesis |
|
Placenta growth factor (PGF) |
Angiogenesis; chemotaxis; modulation of odontoblastic differentiation |
|
Hepatocyte growth factor (HGF) |
Proliferation; migration; cell survival |
|
Epidermal growth factor (EGF) |
Cell growth and neurogenic differentiation |
|
Interleukins (IL-8, IL-10) |
Anti-inflammatory |
|
Neuropeptides and neurotrophic factors |
Function |
|
Calcitonin (CT) |
Ca-metabolism |
|
Calcitonin gene-related peptide |
Vasodilation,; nociception |
|
Neuropeptide Y (NPY) |
Neurotransmitter |
|
Substance P (SP) |
Neurotransmitter; neuromodulator |
Note that bioactive molecules are often multifunctional. From [82, 84].
8.6 Tissue Engineering Approaches to Dental Pulp Regeneration
Tissue engineering is an interdisciplinary field of research, which involves the use of materials and cells with the goal of under- standing tissue function and eventually enabling specific tissues to be made de novo. According to the classical tissue engineering paradigm, stem cells are seeded into a suitable scaffold laden with growth factors, which, after transplantation into a host, induce stem cell differentiation and tissue formation (Figure 8.4). The isolation of pulp-derived stem cells has opened new avenues for tissue engineering approaches in dentistry. Stem cells have been isolated from permanent [35] and deciduous teeth [59], and from the apical papilla of teeth with incomplete root formation [44].
Dental pulp tissue can also be engineered by the transplantation, in experimental animals, of dental pulp stem cells in a suitable carrier system. Tooth slices, dentine cylinders, and whole tooth roots laden with scaffolds and dental stem cells can be transplanted subcutaneously in the back of mice, and vascularized tissue similar to dental pulp forms after few weeks in situ with cells adjacent to the dentine differentiating to deposit tubular dentine [17, 31, 45, 76]. Other animal models can more closely mimic clinical regenerative endodontic treatment procedures: following pulpotomies and pulpectomies in dogs' canines, filling the void with a scaffold material containing stem cells and recombinant growth factor resulted in the formation of vital, pulp-like tissue [46, 47, 62]. There is also evidence that functional recovery of pulp tissue after transplantation of autolo- gous dental pulp stem cells into the root canals of humans is possible [63]. These cell- based experiments follow the tissue engi- neering concept described above, which involves the delivery of stem cells and recombinant growth factors in a carrier material. However, transplantation of stem cells has several problems, including limited availability of cell sources, technical issues of cell harvest and expansion, logistics, high cost, nd regulatory hurdles for approval and clinical translation.
Figure 8.4 Illustration of selected steps of a cell-homing approach for pulp tissue engineering. (a) After conditioning with EDTA, endogenous growth factors are released into saline by ultrasonic activation. The scaffold is laden with isolated growth factors and injected in the canal space; the tooth is restored with bioactive cement and a tight coronal seal. (b) Schematic illustrating potential actions initiated by exposed or supplemented endogenous growth factors in a cell-homing approach in mature teeth. Bioactive molecules may induce chemotaxis of resident stem cells (DPSC if remnant pulp tissue is present, PDLSCs or iPAPCs), their proliferation and attachment as well as odontoblast-like cell differentiation. From [32].
Alternatively, cell-free approaches to dental pulp tissue engineering have been proposed [30, 50, 75]. Following the principle of cell- homing, resident stem cells from remnant pulp or periapical tissues can be recruited into spe- cifically designed biomaterials via recombinant or endogenous, dentine-derived growth factors. Evidence is accumulating that a regenerative endodontic procedure following the principle of cell homing might be a feasible and affordable alternative to cell transplantation [32, 50, 75].
General principles of regenerative mechanisms and tissue engineering approaches in treatment modalities for pulpitis or pulp necrosis are listed in Table 8.2.
Table 8.2 Current and envisioned treatment modalities for pulpitis or pulp necrosis.
|
Current treatment modalities |
|||||
|
Type of assault |
Type of tooth |
Treatment |
Cell type activated/recruited |
Healing |
Proposed Mechanism |
|
Infection - mild stimulus (Reversible pulpitis) |
Mature tooth |
Indirect pulp capping Restoration |
Primary odontoblast |
Regeneration or repair |
Upregulation of odontoblast secretory activity |
|
Infection - intense stimulus (Irreversible pulpitis) |
Mature tooth |
Direct pulp capping or pulpotomy Restoration |
Dental pulp stem cells |
Repair |
Differentiation of stem cells into odontoblast-like or mineralizing cells |
|
Pulp necrosis/apical periodontitis |
Immature tooth |
Revitalization |
Stem cells of the apical papilla or Ectopic cells (bone, PDL) |
Regeneration, more likely repair |
Differentiation of stem cells into odontoblasts, completion of root formation or Apposition of cementum, ingrowth of bone |
|
Immature tooth |
Apexification |
Bone cells/periodontal ligament cells |
Regeneration of bone Repair in contact with a synthetic material |
Induction of a mineralized barrier by a bioactive material |
|
|
Mature tooth |
Root canal treatment |
Regeneration of bone Repair in contact with synthetic material |
Healing in contact with a biocompatible material |
||
|
Proposed tissue engineering-based treatment modalities |
|||||
|
Type of assault |
Type of tooth |
Treatment |
Cell type activated/recruited |
Healing |
Proposed Mechanism |
|
Irreversible pulpitis or pulp necrosis/apical periodontitis |
Mature and immature |
Tissue engineering by cell transplantation |
Transplanted cells (e.g. dental pulp stem cells) |
Regeneration |
Differentiation of transplanted stem cells Ingrowth of vasculature |
|
Immature tooth |
Tissue engineering by cell-homing |
Dental pulp stem cells and/or Stem cells of the apical papilla |
Regeneration |
Differentiation of resident, recruited stem cells |
|
|
Mature tooth |
Tissue engineering by cell-homing |
Dental pulp stem cells or mesenchymal stem cells from the periapical region |
Regeneration |
Differentiation of resident, recruited stem cells |
|
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