Dental Regeneration Research Articles

Ultrasonic techniques to obtain dental pulp from impacted third molars.

BackgroundIn the dental clinic impacted teeth are frequent findings, especially upper and lower third molars, leading to their exodontia. Among surgical techniques piezosurgery is advantageous for delicate structures in the oral cavity. Extracted teeth, usually discarded, have been revalued as biological material, providing living tissues with possible applications in regenerative dentistry. The aim was to compare cross-section methods of upper included third molars by ultrasonic piezoelectric technique to obtain dental pulp, with diamond-coated tip (DT) against titanium nitride-coated tip (TN), according to the pulp tissue cell viability and the section surface characteristics. Material and MethodsPatients attending dental consultation were evaluated. Upper third molars (n= 24) were avulsed from 15 patients with exodontia indication, age 18-26 years old, who agreed to participate of the study. Third molars were cross-sectioned at amelocemental junction level with piezoelectric device using DT or TN inserts. Pulps were mechanic and enzymatically treated, and tissue viability determined by Trypan Blue test. Sectioned teeth were visualized using Scanning Electron Microscope (SEM). Ethical principles of biomedical research were respected; all patients gave their informed consent. ResultsViability of pulp tissue was 84.71% not associated to sex (p= 0.141) nor to teeth position, upper right third molar or upper left third molar (p= 0.580). According to the insert used, pulp tissue viability was 85.21% with TN, similar to 84.00% with DT (p= 0.611). By SEM, cut performed by TN insert showed smooth and uniform surfaces, while DT insert surfaces were irregular, porous, with fissures. ConclusionsPiezosurgery applied to cross-section upper third molars with both types of inserts showed differences in the cut surfaces but similar effectiveness regarding preservation of pulp tissue with high viability, thus, they could be allocated for further cellular developments. Key words:Impacted teeth, third molars, piezosurgery, regenerative dentistry.

The Beneficial Role of Filipendula ulmaria Extract in Prevention of Prodepressant Effect and Cognitive Impairment Induced by Nanoparticles of Calcium Phosphates in Rats.

Mineral components of dental composites are used in many medical and dental applications, including preventive, restorative, and regenerative dentistry. To evaluate the behavioural alterations induced by nanosized particles of novel dental composites, by means of depressive level and cognitive functions, experimental groups of rats were chronically administered with nanosized hydroxyapatite (HA), tricalcium phosphate (TCP), and amorphous calcium phosphate (ACP) with or without simultaneous application of Filipendula ulmaria L. (FU) methanolic extract. The significant prodepressant action was observed in groups solely treated with HA and ACP. Besides, prolonged treatment with ACP also resulted in a significant decline in cognitive functions estimated in the novel object recognition test. The adverse impact of calcium phosphates on estimated behavioural functions was accompanied by increased oxidative damage and apoptotic markers in the prefrontal cortex, as well as diminished specific neurotrophin (BDNF) and gabaergic expression. The results of our investigation showed that simultaneous antioxidant supplementation with FU extract prevented calcium phosphate-induced behavioural disturbances, as well as prooxidative and apoptotic actions, with the simultaneous restoration of BDNF and GABA-A receptors in the prefrontal cortex. These findings suggest that FU may be useful in the prevention of prodepressant impact and cognitive decline as early as the manifestation of calcium phosphate-induced neurotoxicity.

Proteomic Profiling of the First Human Dental Pulp Mesenchymal Stem/Stromal Cells from Carbonic Anhydrase II Deficiency Osteopetrosis Patients.

Osteopetrosis is a hereditary disorder characterized by sclerotic, thick, weak, and brittle bone. The biological behavior of mesenchymal cells obtained from osteopetrosis patients has not been well-studied. Isolated mesenchymal stem/stromal cells from dental pulp (DP-MSSCs) of recently extracted deciduous teeth from osteopetrosis (OP) patients and healthy controls (HCs) were compared. We evaluated whether the dental pulp of OP patients has a population of MSSCs with similar multilineage differentiation capability to DP-MSSCs of healthy subjects. Stem/progenitor cells were characterized using immunohistochemistry, flow cytometry, and proteomics. Our DP-MSSCs were strongly positive for CD44, CD73, CD105, and CD90. DP-MSSCs obtained from HC subjects and OP patients showed similar patterns of proliferation and differentiation as well as gene expression. Proteomic analysis identified 1499 unique proteins with 94.3% similarity in global protein fingerprints of HCs and OP patients. Interestingly, we observed subtle differences in expressed proteins of osteopetrosis disease-related in pathways, including MAPK, ERK 1/2, PI3K, and integrin, rather than in the stem cell signaling network. Our findings of similar protein expression signatures in DP-MSSCs of HC and OP patients are of paramount interest, and further in vivo validation study is needed. There is the possibility that OP patients could have their exfoliating deciduous teeth banked for future use in regenerative dentistry.

Antibiotics: A changing scenario in Regenerative Endodontics

The main aim of Endodontics is to either prevent the endodontic disease or to treat it. This is achieved by maintaining the pulp health in cases of pulp inflammation. If there is pulp necrosis, pulp health can be maintained by regenerating the healthy pulp tissue. Hence various researches and the clinical studies tried to regenerate the healthy functioning pulp-dentin complex. Regenerative dentistry aims to biologically replace the dental tissues along with their supporting structures. Regenerative Endodontics can be defined as “ biologically based procedures designed to replace damaged structures such as dentin, root structures, and cells of the pulp-dentin complex.” Root canal anatomy is complex, hence even after mechanical instrumentation, there exists some portions of the root canal that remain untouched. Only instrumentation cannot completely eliminate the microbes existing into the root canal. Hence there is need to additionally irrigate the canals and also using medicaments is required to disinfect the root canal and promote healing. This article aims at focussing on various antibiotics used and role of these materials to disinfect the canals during revascularization treatment.

Gingival-Derived Mesenchymal Stem Cell from Rabbit (Oryctolagus cuniculus): Isolation, Culture, and Characterization.

Objective This study aims to confirm whether the GDMSCs isolated from rabbit’s ( Oryctolagus cuniculus ) gingiva are mesenchymal stem cells (MSCs). Materials and Methods This study design was partly quasi-experimental with an observational design. GDMSCs were isolated from the gingiva of healthy male rabbits ( O. cuniculus ) ( n = 2), 6 months old, and 3 to 5 kg of body weight. The specific cell surface markers of MSCs; clusters of differentiation (CD), namely, CD44, CD73, CD90, CD105, and CD200 expressions; and hematopoietic stem cell surface markers CD34 and CD45 were examined using flow cytometry and immunohistochemistry with immunofluorescence. The osteogenic differentiation of isolated GDMSCs was examined using alizarin red staining. Results GDMSCs in the fourth passage showed a spindle-like formation and fibroblast-like cells that attached to the base of the culture plate. GDMSCs were MSCs that positively expressed CD44, CD73, CD90, CD105, and CD200 but did not express CD34 and CD45 when examined using flow cytometry and immunohistochemical analysis. GDMSCs had osteogenic differentiation confirmed by calcified deposits in vitro with a red–violet and brownish color after alizarin red staining. Conclusion GDMSCs isolated from the rabbits ( O. cuniculus ) were confirmed as MSCs in vitro documented using immunohistochemistry and flow cytometry. GDMSCs can differentiate into osteogenic lineage in vitro that may be suitable for regenerative dentistry.

Current and Advanced Nanomaterials in Dentistry as Regeneration Agents: An Update.

In modern dentistry, nanomaterials have strengthened their foothold among tissue engineering strategies for treating bone and dental defects due to a variety of reasons, including trauma and tumors. Besides their finest physiochemical features, the biomimetic characteristics of nanomaterials promote cell growth and stimulate tissue regeneration. The single units of these chemical substances are small-sized particles, usually between 1 to 100 nm, in an unbound state. This unbound state allows particles to constitute aggregates with one or more external dimensions and provide a high surface area. Nanomaterials have brought advances in regenerative dentistry from the laboratory to clinical practice. They are particularly used for creating novel biomimetic nanostructures for cell regeneration, targeted treatment, diagnostics, imaging, and the production of dental materials. In regenerative dentistry, nanostructured matrices and scaffolds help control cell differentiation better. Nanomaterials recapitulate the natural dental architecture and structure and form functional tissues better compared to the conventional autologous and allogenic tissues or alloplastic materials. The reason is that novel nanostructures provide an improved platform for supporting and regulating cell proliferation, differentiation, and migration. In restorative dentistry, nanomaterials are widely used in constructing nanocomposite resins, bonding agents, endodontic sealants, coating materials, and bioceramics. They are also used for making daily dental hygiene products such as mouth rinses. The present article classifies nanostructures and nanocarriers in addition to reviewing their design and applications for bone and dental regeneration.

Autologous Materials in Regenerative Dentistry: Harvested Bone, Platelet Concentrates and Dentin Derivates.

The jawbone is a peculiar type of bone tissue, unique for its histological, anatomical and physiological characteristics. Therefore, a defect in the maxilla or in the mandible, because of pathological sequelae is difficult to prevent and to restore. Several biomaterials have been and are currently being developed to respond to the demands of regenerative medicine. A specific group of biomaterials used in regenerative dentistry is represented by the autologous materials. Platelet concentrates harvested bone and dentin derivates are indeed used in an attempt to minimise the alveolar resorption or in vertical ridge augmentation procedures or in sinus lift interventions. The aim of this review is to examine the properties of the above-listed materials, to compare them and to indicate eventual clinical applications.

Integration of Regenerative Dentistry Into the Dental Undergraduate Curriculum

Integration of Regenerative Dentistry Into the Dental Undergraduate Curriculum

2,3,5,4′-tetrahydroxystilbene-2-O-b-D-glucoside triggers the pluripotent-like possibility of dental pulp stem cells by activating the JAK2/STAT3 axis: Preliminary observations

Background/PurposeAlthough 2,3,5,4′-Tetrahydroxystilbene-2-O-beta-glucoside (THSG) reportedly has anti-inflammatory properties, its role in inducing the dedifferentiation of human dental pulp stem cells (DPSC) into pluripotent-like stem cells remains to be determined. The purpose of this study is to evaluate the effects of THSG on the pluripotent-like possibility and mechanism of DPSC.Materials and methodsDPSCs were treated with THSG, and cell viability was determined using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTS) assay. Real-time polymerase chain reaction was used to analyze the mRNA expression levels of pluripotency-associated genes and oncogenes and to detect telomerase activity in the cells. Embryoid body formation assay was conducted, and pluripotency-related proteins were identified using Western blotting. Data were analyzed using one-way analysis of variance.ResultsCell viability, telomerase activity, and embryoid body formation were enhanced in THSG-treated DPSCs. The mRNA expression levels of pluripotent-like genes (including Nanog homeobox [NANOG], SRY-box 2 [SOX2], and POU class 5 homeobox 1 [POU5F1/OCT4]) significantly increased after THSG treatment. The expression levels of pluripotency-related genes (Janus kinase-signal transducer 2 [JAK2] and signal transducer and activator of transcription 3 [STAT3]) increased, whereas those of oncogenes (Ras, SRC, HER2, and C-sis) decreased. Furthermore, the expression levels of the phosphorylated JAK2 and STAT3 proteins significantly increased after THSG treatment.ConclusionTHSG treatment may enhance the pluripotent-like possibility of DPSC through the JAK2/STAT3 axis. Hence, it may be used as an alternative cell-based therapeutic strategy in regenerative dentistry.

Dental pulp stem cells response on the nanotopography of scaffold to regenerate dentin-pulp complex tissue.

The study of regenerative dentistry receives a fast growing interest. The potential ability of the dentin-pulp complex to regenerate is both promising and perplexing. To answer the challenging nature of the dental environment, scientists have developed various combinations of biomaterial scaffolds, stem cells, and incorporation of several growth factors. One of the crucial elements of this tissue engineering plan is the selection and fabrication of scaffolds. However, further findings suggest that cell behavior hugely depends on mechanical signaling. Nanotopography modifies scaffolds to alter cell migration and differentiation. However, to the best of the author's knowledge, there are very few studies addressing the correlation between nanotopography and dentin-pulp complex regeneration. Therefore, this article presents a comprehensive review of these studies and suggests a direction for future developments, particularly in the incorporation of nanotopography design for dentin-pulp complex regeneration.

Stem Cells in Regenerative Dentistry: An Overview

The study of stem cells in regenerative dentistry is the most precious in-progress research. Stem cells arethe cells that have the self-renewal capacity and can differentiate to mature cells. Stemcells also have theprospective to form dentin matrix, bone and neuronal cellsrapidly. Stem cell-based therapeutic approachesprovide a better and attractive alternative that the classical dental treatments because of their questionableefficacy and durability. The most promising alternative for tooth loss is the utilization of stem cells andinduced pluripotent for the regeneration of the whole tooth. Therefore helping with the progress in inducingbone rejuvenation, treating damaged teeth, stem cell-based therapy plays a vital role. So, this article willhighlight the role and importance of stem cells in regeneration in Dentistry

The role of nanohydroxyapatite on the morphological, physical, and biological properties of chitosan nanofibers

This study aimed to evaluate the effects of nanohydroxyapatite (nHAp) particles on the morphological, chemical, physical, and biological properties of chitosan electrospun nanofibers. nHAp particles with a 1.67 Ca/P ratio were synthesized via the aqueous precipitation method, incorporated into chitosan polymer solution (0.5 wt%), and electrospun into nHAp-loaded fibers (ChHa fibers). Neat chitosan fibers (nHAp-free, Ch fibers) were used as the control. The electrospun fiber mats were characterized using morphological, topographical, chemical, thermal, and a range of biological (antibacterial, antibiofilm, cell viability, and alkaline phosphatase [ALP] activity) analyses. Data were analyzed using ANOVA and Tukey's test (α = 0.05). ChHa fibers demonstrated a bead-like morphology, with thinner (331 ± 110 nm) and smoother (Ra = 2.9 ± 0.3 μm) distribution as compared to the control fibers. Despite showing similar cell viability and ALP activity to Ch fibers, the ChHa fibers demonstrated greater antibacterial potential against most tested bacteria (except for P. intermedia), and higher antibiofilm activity against P. gingivalis biofilm. The incorporation of nHAp particles did not jeopardize the overall morphology, topography, physical, and biological characteristics of the chitosan nanofibers. The combination of nHAp particles with chitosan can be used to engineer bioactive, electrospun composite nanofibers with potential applications in regenerative dentistry.

Marine Based Biomaterials in Dental Regeneration

The novel approach of tissue engineering aims at regenerating the functional alveolar or periodontal tissues through a series of key events that is modulated by the use of scaffolds, cells and signalling molecules. Many synthetic and natural polymers have been used as tissue engineering constructs so far with varying results in regeneration. Developing a biomaterial to replace the damaged tissue is of paramount importance for effective regeneration. Due to its rich biodiversity, marine environment yields structures with immense potential for biomedical application. These bio molecules offer many applications in cartilage and bone tissue engineering, dental tissue regeneration, wound healing and local drug delivery system. These substances are usually nontoxic, bio compatible and well tolerated by the body, which boost their efficacy for tissue engineering application. In this article, we are trying to brief the various marine based biomaterials used in dental regeneration, their possible sources and clinical applications.

Recent Advances of Chitosan-Based Injectable Hydrogels for Bone and Dental Tissue Regeneration.

Traditional strategies of bone repair include autografts, allografts and surgical reconstructions, but they may bring about potential hazard of donor site morbidity, rejection, risk of disease transmission and repetitive surgery. Bone tissue engineering (BTE) is a multidisciplinary field that offers promising substitutes in biopharmaceutical applications, and chitosan (CS)-based bone reconstructions can be a potential candidate in regenerative tissue fields owing to its low immunogenicity, biodegradability, bioresorbable features, low-cost and economic nature. Formulations of CS-based injectable hydrogels with thermo/pH-response are advantageous in terms of their high-water imbibing capability, minimal invasiveness, porous networks, and ability to mold perfectly into an irregular defect. Additionally, CS combined with other naturally-derived or synthetic polymers and bioactive agents has proven to be an effective alternative to autologous bone and dental grafts. In this review, we will highlight the current progress in the development of preparation methods, physicochemical properties and applications of CS-based injectable hydrogels and their perspectives in bone and dental regeneration. We believe this review is intended as starting point and inspiration for future research effort to develop the next generation of tissue-engineering scaffold materials.

Magnesium incorporation into primary dental enamel and its effect on mechanical properties

Cross-sectional study of sound primary dental enamel revealed hardness zonation and, in parallel, significant change in the Mg content below the prismless layer. Mg content is known to play an important role in enamel apatite biomineralization, therefore, Mg ion exchange experiments were carried out on the outer surface of sound primary molars and on reference abiogenic Ca-phosphates using MgCl2 solution. Effects of Mg incorporation on crystal/particle size, ionic ratio and morphology were compared and the observed changes were explained by parallel diffusion and dissolution/reprecipitation processes.Based on depth profile analysis and high resolution electron microscopy of the Mg-exchanged dental enamel, a poorly ordered surface layer of approximately 10–15 nanometer thickness was identified. This thin layer is strongly enriched in Mg and has non-apatitic structure. Below the surface layer, the Mg content increased only moderately (up to ~3 at%) and the apatite crystal structure of enamel was preserved. As a common effect of the Mg exchanged volume, primary dental enamel exhibited about 20% increase of nanohardness, which is intrepreted by strengthening of both the thin surface layer and the region below due to the decreased crystallite size and the effect of incorporated Mg, respectively. Statement of SignificanceDental enamel is the most durable mineralized tissue in the human body, which, in spite to be exposed to extreme conditions like mastication and acidic dissolution, is able to fulfill its biological function during lifetime. In this study we show that minor component magnesium can affect hardness properties of human primary dental enamel. Then, through Mg incorporation experiments we provide an additional proof for the poorly ordered Mg-containing intergranular phase which has been recently observed. Also, we report that the hardness of dental enamel can be increased by ca. 20% by Mg incorporation. These results contribute to a deeper understanding of sound primary dental enamel structure and may inspire new pathways for assisted remineralization of enamel and regenerative dentistry.

Prospects of Advanced Therapy Medicinal Products–Based Therapies in Regenerative Dentistry: Current Status, Comparison with Global Trends in Medicine, and Future Perspectives

IntroductionRegenerative medicine offers innovative approaches to restore damaged tissues on the basis of tissue engineering (TE). Although research on advanced therapy medicinal products (ATMPs) has been very active in recent years, the number of licensed products remains surprisingly low and restricted to the treatment of severe, incurable diseases. MethodsThis paper provides a critical review of current literature on the regulatory, clinical, and commercial status of ATMP-based therapies in the EU and worldwide and the hurdles to overcome for their broader application in Regenerative Dentistry. ResultsCompetent authorities have focused on developing regulatory pathways to address unmet patient needs. Oncology represents the dominating field, followed by cardiovascular, musculoskeletal, neurodegenerative, immunologic, and inherited diseases. Yet, the status remains in early development, and scientific, regulatory, and cost-effectiveness issues impose considerable hurdles toward marketing authorization, technology adoption, and patient accessibility. In this context, although regenerative dentistry has achieved breakthrough innovations in TE of several dental/oral tissues in preclinical models, it has hardly harnessed research progress to integrate innovative regenerative treatments into clinical practice. ConclusionGlobal demographic changes, which demonstrate a steady increase of the aging population, highlight the societal need for the application of ATMP-based therapies in the treatment of noncommunicable diseases (NCDs). Although oral diseases, as an integral part of NCDs, are not life-threatening and largely preventable, they sustain high prevalence, with severe burden on economy and quality of life. In this perspective, the urgent request to ultimately translate draining research in dental TE conducted during the last decades into innovative treatments brought safely and cost-effectively into society at large still holds the stage. This review provides an overview of the regulatory, clinical, and commercial status of ATMP-based therapies in the European Union and worldwide and the hurdles to overcome for their broader application in regenerative dentistry.

Scaffold-based and Scaffold-free Strategies in Dental Pulp Regeneration

Regenerative dentistry has come a long way from pulp capping to pulp regeneration research, which aims to regenerate the pulp-dentin complex and restore its functions compromised by pulp injury and/or inflammation. Because of unique anatomic limitations of the tooth structure, engineering a suitable microenvironment that facilitates angio/vasculogenesis and innervation is a challenging task. Cell-based tissue engineering approaches have shown great potential in achieving this goal. Biomedical approaches in creating a regenerative microenvironment are mainly represented by either scaffold-based or scaffold-free strategies. The scaffold-based strategy mainly relies on the use of biomaterials to create a structural base that supports cells throughout the process of tissue formation. The scaffold could be a classic 3-dimensional construct with interconnected pores, a hydrogel with cells embedded in it, or a combination of these 2. The scaffold-free approach has been considered a bottom-up strategy that uses cell sheets, spheroids, or tissue strands as building blocks. The outcome of this strategy relies on the capacity of these building blocks to secrete a favorable extracellular matrix and to fuse into larger tissue constructs. Both the scaffold-free and scaffold-based systems are required as complementary, rather than competing, approaches for pulp regeneration. A combined synergetic strategy, through which multicellular building blocks could be integrated with robust 3-dimensional scaffolds, might represent an optimal solution to circumvent some of the major drawbacks of the current methods in pulp regeneration while concurrently fostering their advantages.

The Emerging Role of Stem Cells in Regenerative Dentistry.

Progress of modern dentistry is accelerating at a spectacular speed in the scientific, technological and clinical areas. Practical examples are the advancement in the digital field, which has guaranteed an average level of prosthetic practices for all patients, as well as other scientific developments, including research on stem cell biology. Given their plasticity, defined as the ability to differentiate into specific cell lineages with a capacity of almost unlimited self-renewal and release of trophic/immunomodulatory factors, stem cells have gained significant scientific and commercial interest in the last 15 years. Stem cells that can be isolated from various tissues of the oral cavity have emerged as attractive sources for bone and dental regeneration, mainly due to their ease of accessibility. This review will present the current understanding of emerging conceptual and technological issues of the use of stem cells to treat bone and dental loss defects. In particular, we will focus on the clinical application of stem cells, either directly isolated from oral sources or in vitro reprogrammed from somatic cells (induced pluripotent stem cells). Research aimed at further unraveling stem cell plasticity will allow to identify optimal stem cell sources and characteristics, to develop novel regenerative tools in dentistry.

Resveratrol and Celastrol Loaded Collagen Dental Implants Regulate Periodontal Ligament Fibroblast Growth and Osteoclastogenesis of Bone Marrow Macrophages.

Collagen is widely used for dental therapy in several ways such as films, 3D matrix, and composites, besides traditional Chinese medicine (TCM), has been used in tissue regeneration and wound healing application for centuries. Hence, the present study was targeted for the first time to fabricate collagen film with TCM such as resveratrol and celastrol in order to investigate the human periodontal ligament fibroblasts (HPLF) growth and bone marrow macrophages (BMM) derived osteoclastogenesis. Further, the physicochemical, mechanical and biological activities of collagen-TCM films crosslinked by glycerol and EDC-NHS (1-ethyl-3-(3-dimethylaminopropyl)carbodiimide-N-hydroxysulfosuccinimide) were investigated. Collagen film characterization was significantly regulated by the nature of plasticizers like hydrophobic and degree of polarity. Interestingly, the collagen film's denaturation temperature was increased by EDC-NHS than glycerol. FT-IR data confirmed the functional group changes due to chemical interaction of collagen with TCM. Morphological changes of HPLF cells cultured in control and collagen films were observed by SEM. Importantly, the addition of resveratrol upregulated the proliferation of HPLF cells, while osteoclastogenesis of BMM cells treated with mCSF-RANKL was significantly downregulated by celastrol. Accordingly, the collagen-TCM film could be an interesting material for dental regeneration, and especially it is a therapeutic target to restrain the elevated bone resorption during osteoporosis.

The Dental Lamina: An Essential Structure for Perpetual Tooth Regeneration in Sharks

In recent years, nonclassical models have emerged as mainstays for studies of evolutionary, developmental, and regenerative biology. Genomic advances have promoted the use of alternative taxa for the study of developmental biology, and the shark is one such emerging model vertebrate. Our research utilizes the embryonic shark (Scyliorhinus canicula) to characterize key developmental and regenerative processes that have been overlooked or not possible to study with more classic developmental models. Tooth development is a major event in the construction of the vertebrate body plan, linked in part with the emergence of jaws. Early development of the teeth and morphogenesis is well known from the murine model, but the process of tooth redevelopment and regeneration is less well known. Here we explore the role of the dental lamina in the development of a highly regenerative dentition in sharks. The shark represents a polyphyodont vertebrate with continuously repeated whole tooth regeneration. This is presented as a major developmental shift from the more derived renewal process that the murine model offers, where incisors exhibit continuous renewal and growth of the same tooth. Not only does the shark offer a study system for whole unit dental regeneration, it also represents an important model for understanding the evolutionary context of vertebrate tooth regeneration.