Molecular factors resulting in tooth agenesis and contemporary approaches for regeneration: a review.
Trauma, genetic disorders, and malformations have created the need for functional, mechanical, and aesthetic replacement of missing teeth. Current methods of tooth replacement include dental implants, tooth transplantation, and prosthodontics; however, these options are not considered permanent and significant complications exist with these methods [Logeart-Avramoglou et al., 2005; Mastrogiacomo et al., 2005]. Ideally, the altered signaling cascade that interupts tooth development could be treated, or an autogenous tooth replacement developed. This paper discusses the genetic regulation and signaling networks involved in epithelial-mesenchymal interactions during tooth development and the signaling pathway alterations that result in hypodontia. Tooth development includes reciprocal interactions between the epithelium and mesenchyme, which is governed by the expression of sonic hedgehog (Shh), fibroblast growth factors (FGFs), bone morphogenetic proteins (BMPs), and wingless (Wnt) signaling families [Thesleff and Sharpe, 1997; Yen and Sharpe, 2008]. The aim of this paper is to dissect these complex interactions and related mutations so we may better understand and develop new dental regeneration and growth factor supplementation procedures.
In the discussion section, generalisations have been made regarding the location of signaling factors, and these specifically refer to tooth development unless otherwise specified. For example, if a factor is said to only be expressed in the mesenchyme, it is referring specifically to the epithelial-mesenchymal interactions during tooth development.
[FIGURE 1 OMITTED]
Tooth development In utero, ectomesenchyme from the neural crest cells migrates to the site of tooth development and forms a complex signaling network with the dental epithelium from the first branchial arch. The dental epithelium thickens and buds into the underlying dental mesenchyme, and the epithelium condenses and forms knots at the site of the future cusps [Jernvall et al., 1998]. By the cap stage, these knots are fully developed and have begun to serve as central regulators of tooth development by secreting signaling factors from the BMP, FGF, Shh, and Wnt families [Thesleff and Sharpe, 1997].
Throughout tooth development, the epithelium and mesenchyme synergistically activate each other. As the epithelium thickens, it will activate the mesenchyme, which, during the cap stage, will induce adjacent epithelium to differentiate into the enamel organ [Kollar and Baird 1970]. Once the mesenchyme is activated, it is responsible for inducing the specific morphology of the tooth and cusps, while still depending on signaling feedback from the epithelium [Hu et al., 2006; Takahashi et al., 2010]. In the absence of signaling factors from the epithelium or mesenchyme, tooth genesis may be suspended.
Signaling factors Epithelial-mesenchymal interactions are extremely intricate and require numerous signaling cascades (Figure 1). The following factors have been implicated as major contributors to tooth development:
Sonic hedgehog (shh) Shh is believed to effect the formation, proliferation, and temporal development of the enamel knot [Tucker and Sharpe, 1999; Zhang et al., 2005]. Shh expression occurs downstream of Fgf8 and Fgf9, is regulated by Bmp4, and is responsible for regulating Bmp2 [Zhao et al., 2000; Thesleff et al., 2001]. Shh is locally expressed at the tip of the thickening epithelium and is responsible for activating patterning and proliferation factors in the mesenchyme, such as the Ptc gene [Chuong et al., 2000]. Excess Bmp4 inhibits Shh, which inhibits Bmp2. This has no effect on knot formation, suggesting Shh may not have an effect on cusp development as was once believed [Zhao et al., 2000].
Interestingly, all epithelial appendages, such as teeth, hair, eyes, tongue papillae, and lungs, have a conserved genetic pathway that has been re-arranged to result in novel organ morphologies [Chuong et al., 2000]. Therefore, mutations in Shh have systemic effects. For example, PTC is an inhibitory receptor for Shh, and mutations in Ptc result in deregulation and over-expression of Shh causing a malignancy of epithelial basal cells [Tojo et al., 1999]. This is associated with basal cell nevus syndrome (Gorlin syndrome/nevoid basal cell syndrome) with symptoms such as numerous basal cell carcinomas, tumour formation, odontogenic keratocysts, and small abnormally shaped teeth and alveolar bone [Chuong et al., 2000; Ohki et al., 2004; Cobourne et al., 2009].
Fibroblast growth factors (FGFs) FGFs are a highly conserved family of epithelial signaling factors that are crucial for tooth development, craniofacial patterning, growth regulation, skeleton genesis, and synchondrosis regulation [Nie et al., 2006a]. Fgf3 is expressed solely in the mesenchyme, while Fgf4, Fgf8, and Fgf10 are expressed by the epithelium. FGF3 stimulates cell proliferation in the mesenchyme. Fgf4 is activated by LEF1 from the Wnt pathway, is expressed only in the knot, and is responsible for enamel epithelium and dental papilla proliferation [Miletich and Sharpe, 2003; Zhang et al., 2005]. Fgf8 is expressed prior to the bud stage and responsible for tooth patterning and positioning. Fgf10 stimulates cell proliferation in the epithelium [Thesleff and Sharpe, 1997; Zhang et al., 2005; Nie et al., 2006a].
FGF3 and FGF10 have some form of functional redundancy, as Fgf3-/- or Fgf10-/- mice develop teeth normally, but mice that lack the FgfR2b receptor (the receptor for Fgf3 and Fgf10) exhibit teeth that arrest at the bud stage [Zhang et al., 2005].
Cbfa1/Runx2 protein is a critical transcriptional regulator of osteoblast differentiation that is down-regulated by Msx1 most likely via FGF3 [Zhang et al., 2005]. If Cbfa1/Runx2 is reduced, Fgf3 and Fgf10 are down-regulated, resulting in a systemic disorder called cleido-cranial dysplasia with a systemic presentation of supernumerary teeth that fail to erupt, defective development of cranial bones, and the partial or complete absence of clavicles [Zhang et al., 2005; Wang and Fang 2011].
Lhx6/7 and Gsc develop the oral-aboral polarity of the mandible [Caton and Tucker, 2009; Denaxa et al., 2009]. FGF8 induces Lhx6/7 which are responsible for activating mesenchymal homeobox genes to coordinate odontogenesis. In molar patterning, Lhx6/7 maintain expression of Msx1 , and Msx1 stimulates Bmp4 [Zhang et al., 2005; Denaxa et al., 2009]. In incisor patterning, Lhx6/7 maintain agenesis of the diastema between the rat incisor and 1st molar [Zhang et al., 2005; Denaxa et al., 2009].
It is not completely understood to what extent Fgf8 regulates the odontogenic potential of the dental epithelium, as Fgf8-/mice lack molars and have developed incisors; it is possible FGF9 rescues the incisor development when FGF8 is not present [Zhang et al., 2005; Pani, 2011]. While FGF expression is limited to the enamel knot, the knot lacks FGF receptors and the cells are unable to proliferate. FGF release activates the surrounding epithelium, which proliferates around the enamel knot and induces the transition from the bud to the cap stage [Thesleff et al., 2001]. Additionally, Fgf4 regulates factors that limit ossification (Fgf3) and other factors that control osteoblast differentiation and function (Runx2). Another important role of the FGF family (specifically Fgf2, Fgf4, Fgf8, and Fgf9) is to up-regulate Msx1 expression for odontogenesis, as Msx/- mice have reduced expression of Bmp4, Fgf3, Lef1, Dlx2, Ptc, Syndecan-1, and tenascin [Zhang et al., 2005].
Bone morphogenetic proteins (BMPs) BMPs are an important family of redundant activators for incisor tooth development. During early development, Bmp2, Bmp4, and Bmp7 are expressed in high amounts from the enamel knot and play a role in cell proliferation and apoptosis, position and number of cusps, and tooth patterning [Zhang et al., 2005].
Bmp4 interacts with multiple growth factors and is critical in downstream signaling events that are required for the formation of the enamel knot [Kapadia et al., 2007]. Prior to laminar thickening, Pax9 expression is upregulated until the end of the cap stage, when it is reduced. Bmp4 inhibits Pax9, while Fgf8 activates Pax9.
Pax9 physically interacts with Msx1 in a feedback loop to regulate Bmp4 expression in the mesenchyme [Vieira et al., 2004; De Coster et al., 2009; Zhou et al., 2011]. Msx1 and Pax9 alleles are over-transmitted together suggesting both genes interact in tooth agenesis [Vieira et al., 2004; Zhou et al., 2011]. If Msx1 or Pax9 are not expressed, the mesenchyme will not be activated to release Bmp4, which in turn causes Lef1, Dlx2, Plx2, Ptc, and Fgf3, to not be expressed, and the tooth bud will not develop to the cap stage [Zhao et al., 2000; Matalova et al., 2008; De Coster et al., 2009].
Lef1 works in association with Bmps to activate Alx4 to signal mesenchymal condensation and is required to activate Fgf4 expression for the Wnt cascade [Hudson et al., 1998; Nie et al., 2006b]. Dlx1, 2-/- mice may only lack maxillary molars due to compensation by other members of the Dlx family (possibly Dlx5,6) to restore mandibular molar and all incisor development [Caton and Tucker, 2009; Pani, 2011]).
If the mesenchyme is not activated to release Bmp4 and exogenous Bmp4 supplements are provided, molars will develop to the cap stage. Down-regulation of Msx1 causes a decrease in Fgf8; Fgf8 does not signal Fgf3; Fgf3 does not signal Syndecan-1; the absence of Syndecan-1 causes cessation of tooth development during the cap stage [Zhao et al., 2000]. Osr2 is also activated by Pax9 and restricts Msx1 and Pax9 activation of BMP4 via stable protein-protein complexes [Zhou et al., 2011]. In Osr2-/- mice, Msx1 expression will activate excessive Bmp4 synthesis, resulting in supernumerary lingually displaced molars, similar to non-mammalian multi-rowed dentitions [Mikkola, 2009; Zhou et al., 2011]. In Msx1-/- or Pax9-/- mice, Osr2 will limit Bmp4 activation by Pax9 or Msx1, respectively [Zhou et al., 2011]. However, if the mouse is Msx-/- and Osr2-/-, 1st molar (not the 2nd molar) morphogenesis will be rescued as Pax9 occurs upstream of Msx1 and is capable of activating basal levels of Bmp4 [Zhou et al., 2011]. Additionally, it has been hypothesised Msx2, a close relative and mesenchymal redundancy of Msx1, may become active when Msx1 expression is absent and amplify Bmp4 via a positive Bmp4-Msx2 feedback loop [Mikkola, 2009]. However, other authors believe Msx1 occurs downstream of Bmp4, and they activate each other in a positive feedback loop which accounts for the downregulation of Lef1 in mutant Msx1 cases [Miletich and Sharpe, 2003]). It is hypothesised "the Bmp4-Msx1 pathway is a driving force in the activator-inhibitor network regulating sequential tooth initiation, while Msx1 and Osr2 act antagonistically to pattern the tooth morphogenic field by controlling the expression and spatial distribution of mesenchymal odontogenic signals along the buccolingual axis" [Zhang et al., 2009]. Pax9-/- mice have a downregulation of Msx1, Bmp4, and Lef1, resulting in missing molars, while Msx1-/- mice lack 2nd premolars and 3rd molars [Miletich and Sharpe, 2003; Zhang et al., 2005].
Bmp4 is important during the bud-cap stage transition, as it activates Msx1 and Msx2 in the mesenchyme and is necessary for molar development and odontoblast differentiation [Nie et al., 2006b]. Once the mesenchyme is activated, it will begin to produce Bmp4 as well, which will reciprocally activate the enamel knot [Tnesleff et al., 2001; Nie et al., 2006b]. As the odontoblasts express BMP, it induces ameloblasts to differentiate. At the same time, follicular activin induces epithelial follistatin, which inhibits odontoblast-derived BMPs. Differential downregulation of follistatin leads to asymmetric distribution of enamel, which is responsible for individual tooth morphology [Caton and Tucker, 2009].
At the bell stage, the primary knot will disappear in consequence of apoptotic events regulated by Bmp4 apoptosis and secondary knots will form at each cusp of premolar and molar teeth [Thesleff and Jernvall, 1997]. The secondary knots, which are located over the cusp tips of multi-cusp teeth, have slightly different signaling. For example, Bmp2 is upregulated in the primary knot, but not present in the secondary knots. It is hypothesized that the stage-specific signaling differences between the primary and secondary knot is responsible for altering odontoblast differentiation, patterning, and morphogenesis in order to develop the appropriate dental anatomy for each tooth [Thesleff et al., 2001]. Due to BMPs integral involvement in epithelial-mesenchymal interactions, it is used extensively in the study of tooth regeneration.
Wingless signaling (Wnt) The Wnt signaling pathway is active during the entire tooth development process. Wnt induces [beta]-catenin production, which results in cell proliferation. Intracellular levels of [beta]-catenin are regulated by the APC complex via proteolysis, and the Wnt/[beta]-catenin complex may be antagonised by Osr2 [Chen et al., 2009].
During the initiation stage, Wnt/[beta]-catenin signaling is localised in the epithelium and may play a positive role in initiating laminar thickening. During the bud stage, Wnt/[beta]-catenin signal is necessary for proper epithelial-mesenchymal crosstalk. In the bud stage, two main factors may inhibit proper Wnt signaling. First, inadequate Bmp4 leads to suppression of Msx1 and Msx2, causing the up-regulation of Dkk1 and the arrest of tooth development at the bud stage. Unregulated Dkk1 is significant as it is a potent and specific inhibitor of Wnt signaling by binding and inhibiting LPR co-receptors, which are required for the Wnt signaling pathway activation [Miletich and Sharpe, 2003; Nie et al., 2006b; Chen et al., 2009; Liu and Millar, 2010]. Second, [beta]-catenin may not be synthesised, resulting in inactive Fgf3 and Lef1 and developmental arrest prior to primary enamel knot formation [Nie et al., 2006b; Chen et al., 2009; Liu and Millar, 2010]. Lef1 is important for reciprocal epithelial mesenchymal interactions because it induces Fgf4 expression in the knot, which leads to Fgf3 expression in the mesenchyme and Shh in the epithelium [Kratochwil et al., 2002; Zhang et al., 2005]. During the cap stage, Wnt/[beta]-catenin is upregulated in the knot.
Excess [beta]-catenin is known to induce new enamel knots to bud off the existing epithelium, resulting in simply shaped, lingually displaced supernumerary teeth [Jarvinem et al., 2006; Chen et al., 2009]. A defective APC complex, mutations that prevent [beta]-catenin degradation, or excess Bmp4 promote Wnt activity and have been linked to excess [beta]-catenin, which suggests Wnt is active in regulating knot differentiation. It has been hypothesised that excess BMP4 upregulates the Wnt cascade, resulting in supernumerary teeth. [Jarvinem et al., 2006; Liu, 2008; Mikkola, 2009].
While it is not well understood, Barx1 is an antagonist to Wnt and is regulated by an antagonistic Fgf8 (upregulates Barx1)Bmp4 (downregulates Barx1) interaction. This antagonistic relationship between Wnt and Barx1 is important to differentiate molars from incisors and is required for the transition from bud to cap stage [Nie et al., 2006a; Song et al., 2006; Caton and Tucker, 2009]. Barx1 is restricted to the posterior dentition mesenchyme and important for premolar and molar specification [Miletich et al., 2005; Nie et al., 2006b; Caton and Tucker, 2009]. The Fgf8-Bmp4 interaction is initially activated by Shh and restricts Pax9 to the presumptive dental mesenchyme and Pitx2 to the presumptive dental epithelium [Caton and Tucker, 2009; Nie et al., 2006b]. Pitx2 activates Dlx2, is involved in a positive feedback loop with FGF8, and is required for the regulation and orientation of tooth development [Zhang et al., 2005; Dressler et al., 2010].
Mutations in Pitx2 may result in Axenfeld-Rieger syndrome with variable malformations to the eyes, teeth, maxilla, heart, ears, and brain [Miletich et al., 2005]. If Pitx1 is downregulated, Fgf8 is reduced and Bmp4 is increased, the result is failure to develop molars [Caton and Tucker, 2009]. In cases of extreme [beta]-catenin expression, systemic effects may be present in the form of familial colorectal polyposis/Gardner syndrome (familial adenomatous polyposis, numerous osteomas, thyroid cancer, fibromas, epidermoid cysts, and supernumerary teeth) [Wang et al., 2011]. Wnt initiates the formation of the secondary enamel knots and has been implicated in regulating tooth size and occlusal morphology [Sarkar and Sharpe, 2000].
Genetic defects resulting in altered and absent dentition
Any alteration of the epithelial-mesenchymal interactions can have deleterious effects on tooth development and may have systemic effects. Numerous mutations that result in tooth agenesis were mentioned above and four genes that are frequently affected will be discussed in more detail: MSX1, Pax9, Axin2, and Eda (Table 1).
MSX1 (OMIM 142983: 4p16.3-p16.1) is a homeobox gene with mutations that effect second premolars and third molars with an association for cleft lip and/or palate and Witkop syndrome (Zhang et al., 2005; Fleischmannova et al., 2008; Kavitha et al., 2010; Pani, 2011). The majority of MSX1 mutations are autosomal dominant and result in hypodontia and oligodontia, however an autosomal recessive mutation has been identified [Chishti et al., 2006; Nieminen, 2009; Pani, 2011]. Msx1 is strongly expressed in the mesenchyme and interacts with Bmp4 and Pax9 to determine the shape and position of teeth [Zhang et al., 2005; Matalova et al., 2008]. To date, five distinct mutations have been identified in MSX1 that preferentially induce posterior dentition agenesis (second premolars and third molars), while anterior teeth may be affected in severe cases. The documented mutations significantly alter the structure of MSX1 which impedes its ability to bind DNA [Chishti et al., 2006; Matalova et al., 2008]. For example, a missense mutation (R196P) in the homeodomain causes the protein to have reduced thermo-stability, which severely impairs its biochemical activity and eliminates its ability to interact with other proteins and transcription factors [Vastardis et al., 1996]. MSX1 has a clear link to tooth agenesis and explains a reduced percentage of tooth agenesis cases [Vieira, 2008].
PAX9 (OMIM 167416: 14q12-q13) is a transcription factor that binds downstream enhancers in order to modify transcription activity and is necessary at multiple stages of development. During the bud stage, Pax9 induces the mesenchyme to express BMP4, Msx1, and Lef1 [Neubuser et al., 1997]. Autosomal dominant nonsense, missense, insertions, and deletion mutations in PAX9 have been documented to be responsible for a high incidence of absent permanent molars and in more severe cases missing second premolars and mandibular central incisors [Matalova et al., 2008].
PAX9 mutations are associated with oligodontia, microdontia, benign hereditary chorea, cleft of secondary palate, and absent thymus and parathyroid glands [Kavitha et al., 2010; Pani, 2011]. Interestingly, the gene dosage of Pax9 decreases from anterior to posterior teeth in the dental arch, which parallels cases of PAX9 deficiencies where posterior teeth are absent more frequently than anterior teeth [Kist et al., 2005]. Mutations in PAX9 lead to haploinsuficiency that halts tooth development in an autosomal dominant manner by not activating BMP4.
AXIN2 (OMIM 604025: 17q23-q24) is an important regulator of the stability of [beta]-catenin in the Wnt/[beta]-catenin pathway and is found in high concentrations in the mesenchyme and enamel knot during tooth development. Mutated AXIN2 results in anodontia and accumulation of intracellular [beta]-catenin in cancerous cells in the colon [Lammi et al., 2004]. It is hypothesised AXIN2 acts in a negative feedback loop with the Wnt cascade, and a loss of this gene reduces Wnt signaling [Caton and Tucker, 2009]. An intricate control of Wnt signal activity is necessary for normal tooth development, since point change and frameshift mutations may inhibit or stimulate Wnt signaling and may result in severe oligodontia, sporadic incisor agenesis, and colorectal cancer [Lammi et al., 2004; Pani 2011].
EDA (OMIM 300451: Xq12-q13.1) is in the TNF family and is an important gene for enamel knot formation, tooth morphogenesis, and cusp patterning [Kolenc-Fuse, 2004; Matalova et al., 2008]. Increased Eda expression produces more secondary enamel knots, resulting in more cusps, changes in cusp shape and position, the formation of longitudinal crests, and increased number of teeth [Kangas et al., 2004]. Eda expression is regulated by Wnt during the formation of secondary enamel knots [Caton and Tucker, 2009]. If the Wnt complex, Eda, or the Eda receptor is mutated, the size and shape of the enamel knot is altered [Caton and Tucker, 2009]. Rather than forming a visible cluster of cells at the enamel knot, Eda mutants form a flat, elongated sheet called an enamel rope. While the signaling factors, such as Shh, BMP4, FGF4, Wnt, and Eda may still be expressed at high levels, their spatial patterning is altered, which results in small malformed molars with few short, flattened cusps and conical incisors [Tucker et al., 2000; Caton and Tucker, 2009]. Translocations, point mutations, and frame shifts may alter Eda expression that result in misshapen conical teeth or varying degrees of tooth agenesis. Additionally, Eda is another conserved signaling gene that has many systemic effects. Ectodermal dysplasia (ED) develops due to Eda inactivation and results in abnormal teeth, hair, sweat gland, skin, and nail development.
Molecular approach for regeneration of missing teeth
During tooth development, altered gene expression has widespread effects on other signaling pathways and may result in the cessation of tooth development. Many different approaches are being undertaken to treat different aetiologies of tooth agenesis. In cases where growth factors are absent, post-natal supplements have been proposed, while in cases of tooth avulsion or the lack of development, the growth of complete bioengineered teeth is being attempted.
In one of the first studies of its kind, foetal mice with ED were treated with a recombinant Eda protein [Gaide and Schneider, 2003]. Eda was attached to the Fc portion of human IgG. The Fc-Eda recombinant complex was stable, aggregated, and capable of transplacental delivery. It was injected intravenously into the pregnant mother at set intervals. While the mother appeared unaffected during pregnancy, all of her offspring reverted to the wild-type-like phenotype. Though the Fc-Eda restored dental anatomy, third molars were still frequently absent.
In a similar study, canine dogs with ED were treated intravenously with Fc-Eda after birth [Casal et al., 2007]. Most of the dogs that received an extended treatment of Fc-Eda responded well, developed a normal dentition, and eliminated or reduced most other ED symptoms.
In a population of mice with inactivated Pax9, the number and severity of missing teeth was dosage dependent; the concentration of Pax9 in the posterior arch was substantially less than the anterior arch [Kist et al., 2005]. As a result, first molars developed to a more advanced stage than second molars (though they still did not always form completely), which developed to a further stage than third molars. It has been hypothesised that by providing post-natal doses of Pax9, dental deficits that are a result of inadequate Pax9 may be treated and partial dentition may be restored in a manner similar to the Fc-Eda treated canines.
Another area of investigation has examined the feasibility of growing bioengineered teeth. Currently, there are two main approaches to complete tooth generation. The first method is to develop a biodegradable scaffold that cells will be seeded onto. In one study, a tooth bud from a rat was removed, the cells were isolated, single-cell isolates were seeded onto a polymer scaffold which was grown in a rat omentum, and the teeth were removed after 12 weeks [Duailibi et al., 2004]. The resulting teeth exhibited mineralisation and proper colour, shape, and size. Despite the promise of this scaffold study, the growth patterning was not fully understood and application of this technology would require tooth bud isolation during early development. Another significant complication is the development of a biocompatible, resorbable, and drug delivering scaffold.
The second method for tooth regeneration is to isolate and transplant stem cells to establish epithelial-mesenchymal interactions and reconstruct the tooth germ. Five major sources of stem cells have been proposed: dental pulp stem cells (DPSCs); stem cells from human exfoliated teeth (SHEDs); periodontal ligament stem cells (PDLSCs); stem cells from the apical root papilla (SCAPs); dental follicle stem cells (DFSCs) (Table 2, Figure 2). DPSCs from extracted third molars have been shown to contain pluri-potent dental pulp stem cells that have the ability to expand into odontoblasts, osteoblasts, chondrocytes, and adipocytes [Peng et al., 2009; Sloan et al., 2009; Volponi et al., 2010; Kawashima, 2012]. In vitro, studies have demonstrated the ability for DPSCs to form calicific nodules with functional dental tissue with enamel, dentine, and pulp-like complexes [Volponi et al., 2010; Kawashima, 2012]. SHEDs have increased population doublings, osteo-inductive capacity, and higher population rates than DPSCs [Peng et al., 2009; Sloan et al., 2009; Volponi et al., 2010; Kawashima, 2012]. PDLSCs may differentiate along mesenchymal cell lineages to form cementoblast-like cells, adipocytes, and collagen [Peng et al., 2009; Volponi et al., 2010]. SCAPs are isolated prior to tooth eruption (such as the 3rd molar); these cells can differentiate into odontoblasts and adipocytes and show higher rates of proliferation than DPSCs. When SCAPs and PDLSCs were implanted into an alveolar socket of a pig, dentine and a periodontal ligament were found, suggesting the possible development of a natural tooth root implant with a PDL rather than a titanium screw implant [Volponi et al., 2010]. DFSCs are isolated from the epithelium and mesenchyme prior to eruption and may form cementoblast-like cells, PDL, and osteoblasts. One study isolated the epithelially activated mesenchyme, reorganised the cells, recombined the cells with epithelium, and placed the epithelial and mesenchymal cells in a mouse renal capsule [Yamamoto et al., 2003]. After allowing time to develop, the teeth were removed and shown to contain enamel, dentine, and pulp tissue (not cementum). The clinical application to transform these stem cells into teeth appears promising; however, more research is needed to study the effects of combining multiple lineages of stem cells to facilitate the accelerated development of an entire tooth.
[FIGURE 2 OMITTED]
Epithelial-mesenchymal interactions are a complex and loosely understood network of reciprocal cascades that are necessary for proper tooth development. During initial tooth development, the Shh, FGF, BMP, and Wnt signaling pathways are required for epithelial thickening, bud development, and knot formation. Also, numerous genetic mutations have been identified in MSX1, PAX9, EDA, and AXIN2 that are directly responsible for oligodontia. Multiple approaches are being studied to resolve genetic and syndromic tooth agenesis. Preliminary trials of signaling factor supplementation with animals that have ectodermal dysplasia appear promising, while treatment options for the other genetic disorders still must be elucidated. Other studies are developing methods to grow new teeth with different dental stem cell populations. Future studies will need to examine the effect of growth factor supplementation on tooth development and experiment with stem cell population combinations to grow functional teeth.
This manuscript is based on a paper submitted as part of the requirements of the University of Pittsburgh School of Dental Medicine course ORBIOL 5174 Craniofacial Genetics. We thank Sarah Visnki for revising grammar and style. This work is dedicated to Dr. Nicholas Piesco for his teachings of tooth development and a career dedicated to integrating basic scientific concepts into clinical training.
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S.M. Cudney, A.R. Vieira
Department of Oral Biology, University of Pittsburgh School of Dental Medicine, Pittsburgh, USA.
Postal address: Dr A.R. Vieira, Department of Oral Biology, University of Pittsburgh School of Dental Medicine, 614 Salk Hall, 3501 Terrace Street, Pittsburgh, PA 15261, USA.
Table 1. Gene mutations and phenotypes. This table summarises the inheritance and expression of mutations in Msx1, Pax9, Axin2, and Eda. Gene Location Inheritance Msx1 4p16.3-16.1 Autosomal dominant (AD) and autosomal recessive (AR) Pax9 14q12-q13 AD Axin2 17q23-q24 AD Eda Xq12-q13.1 AD, AR, and X-linked recessive Gene Phenotype and associated conditions Msx1 Absent second premolars third molars, and in severe cases anterior teeth; CUP; Witkop syndrome Pax9 Absent molars and in severe cases second premolars and mandibular central incisors; peg laterals; microdontia; benign cleft of secondary palate; hereditary chorea; absent thymus and parathyroid glands Axin2 Absent molars, premolars, mandibular incisors, and maxillary lateral incisors, colorectal cancer Eda Varying degree of tooth agenesis, conical teeth, and ectodermal dysplasia Gene Molecular consequence Msx1 Reduced expression of Bmp4 and Fgf3; altered mesenchymal signaling causing failure of enamel knot development Pax9 Reduced expression of Bmp4, Msx1, and Lef1; altered mesenchymal signaling causing failure of enamel knot development Axin2 Accumulation of [beta]-catenin and reduced Wnt Eda Altered epithelial and enamel knot signaling Gene Reference Msx1 Zhang et al., 2005; Chishti et al., 2006; De Coster et al., 2009; Nieminen, 2009; Pani, 2011 Pax9 Nieminen, 2009; Kavitha et al., 2010; Pani, 2011; Matalova et al., 2008. Axin2 Lammi et al., 2004; Nieminen, 2009; Pani, 2011 Eda Tucker et al., 2000; Nieminen, 2009; Pani, 2011 Table 2. Proposed stem cells for tooth regeneration. This table summarises the stem cells that have been proposed for tooth regeneration, their harvest site, and potential to differentiate. Cell Type Harvest site Dental pulp Pulp of extracted stem cells (DPSC) third molars Stem cells from human Pulp of primary teeth exfoliated teeth (SHED) Periodontal ligament Periodontal ligament space stem cells (PDLSC) Stem cells from the Developing root prior to apical root tooth eruption papilla (SCAP) (i.e. third molars) Dental follicle Epithelium and mesenchyme stem cells (DFSC) of tooth germ Cell Type Differentiate into Dental pulp Osteoblasts, chondrocytes, stem cells (DPSC) adipocytes, and neurons Stem cells from human Osteoblasts, neurons, exfoliated teeth (SHED) adipocytes, and atypical odontoblasts Periodontal ligament Cementoblast-like cells, stem cells (PDLSC) adipocytes, odontoblasts, and collagen Stem cells from the Odontoblasts, adipocytes, apical root and osteocyte-like cells papilla (SCAP) Dental follicle Osteoblasts, cementoblasts, stem cells (DFSC) adipocytes, and neurons Cell Type Product/Characteristics Dental pulp Sporadic calcific nodules stem cells (DPSC) with enamel, dentine, and pulp Stem cells from human Dentine-like tissues; exfoliated teeth (SHED) increased population doublings, osteoinductive capacity, and higher population rates than DPSCs Periodontal ligament Formation of cementum-like stem cells (PDLSC) calcifications with PDL attachments Stem cells from the Formation of cementum with apical root PD complex; formation of papilla (SCAP) dentine-pulp like tissues; higher proliferation rate than DPSCs Dental follicle Formation of mineralised stem cells (DFSC) tissues and a PDL Cell Type Reference Dental pulp Volponi et al., 2010; stem cells (DPSC) Kawashima, 2012 Stem cells from human Peng et al., 2009; exfoliated teeth (SHED) Sloan et al., 2009; Volponi et al., 2010; Kawashima, 2012 Periodontal ligament Peng et al., 2009; stem cells (PDLSC) Volponi et al., 2010 Stem cells from the Volponi et al., 2010; apical root Kawashima, 2012 papilla (SCAP) Dental follicle Peng et al., 2009; stem cells (DFSC) Volponi et al., 2010 before eruption
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|Author:||Cudney, S.M.; Vieira, A.R.|
|Publication:||European Archives of Paediatric Dentistry|
|Date:||Dec 1, 2012|
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