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PubMed1995

Ultrastructural relation between nerve terminals and dentine bridge formation after pulpotomy in human teeth.

Close association between nerve terminals and preodontoblasts, odontoblasts and predentine was observed during healing after pulpotomy. The nerve terminals frequently contained large numbers of synaptic vesicles. Terminals with many vesicles tended to be fewer in the predentine than in the odontoblastic layer. The distribution of terminals was more dense at the stage before the regenerated odontoblasts became arranged regularly beneath the predentine. It is suggested that these terminals have some efferent role(s), especially during collagen synthesis at the early stage of dentinogenesis. The nerves may release their abundant synaptic vesicles, in addition to serving a sensory role for monitoring the increased sensitivity in the injured areas.

Dentin, Secondary↗
PubMed1995

Providing an environment for reparative dentine induction in amputated rat molar pulp by high molecular-weight hyaluronic acid.

To study provision of this environment, wound healing was examined by light and electron microscopy following pulp amputation and direct capping with hyaluronic acid. Molar pulps of female Sprague-Dawley rats were mechanically exposed and directly capped; the cavities were then restored with glass-ionomer cement. As an experimental control. Calvital (a commercial preparation of calcium hydroxide paste) was used as the direct pulp-capping agent in other rats, and its effect on pulp healing was compared with that of hyaluronic acid. In hyaluronic acid-treated pulps, at 2 days after amputation, wound surfaces were covered with blood and fibrin clots and inflammatory cells such as neutrophils and macrophages. At 1 week, differentiation of fibroblastic and odontoblast-like cells was observed beneath the wound layer; and odontoblast-like cells produced globular calcified nodules along the existing dentine walls. At 2 weeks, a layer of reparative dentine had been formed by odontoblast-like cells over the dentine walls. Between 30 and 60 days, the formation of reparative dentine had extended throughout the pulp chamber. The healing observed after direct capping with hyaluronic acid was, except for the formation of dentine bridges, similar to that seen after Calvital capping. These results suggest that high molecular-weight hyaluronic acid can provide an environment suitable for reparative dentine formation through mesenchymal cell differentiation during healing of the amputated dental pulp.

Animals↗
PubMed2006

Isolation and partial sequencing of cDNA clones from rat incisor after Nd:YAG laser irradiation in root canal.

OBJECTIVE: The purpose of this study was to identify the genes expressed in odontoblast-like cells and related to osteodentin production of pulp cells in vivo during the repair process. METHODS: A pulsed Nd:YAG laser was irradiated in mandibular incisors of rats at 2 W and 20 pulses/s for 5 s. We constructed a unidirectional cDNA library from the non-calcified portion of irradiated incisors, sequenced cDNA clones and classified their sequences by homology search through the GenBank database. RESULTS: Of the sequences determined, 26% represented sequences of new genes that were not related to any previously reported gene. 65.5% of the clones strongly matched genes and proteins in the databases. Among them, sequences found more than once were amelogenin, ameloblastin, amelin, collagen alpha1 type 1, osteonectin, nestin, enamelin and osteocalcin. Sequences detected only once were transforming growth factor beta-1 (TGF-beta1), growth hormone receptor (Ghr), glypican, enamelysin (Mmp20), amelin 2, integral membrane protein 2B, amelogenin precursor (Ame1), splicing factor 3b, insuline-like growth factor binding protein 5. CONCLUSION: This library reflects the gene expression during the dentin repair process in vivo including the odontoblast-like cell specific gene expression and osteodentin-production. Using this library, we indicate the candidate genes which might be related to odontoblast-differentiation and formation of osteodentin.

Animals↗
PubMed2006

Targeted expression of csCSF-1 in op/op mice ameliorates tooth defects.

OBJECTIVE: The aim of this study was to characterize the tooth phenotype of CSF-1-deficient op/op mice and determine whether expression of csCSF-1 in these mice has a role in primary tooth matrix formation. DESIGN: Ameloblasts and odontoblasts, isolated from wt/wt frozen sections using laser capture microdissection, were analysed for csCSF-1, sCSF-1 and CSF-1R mRNA by RT-PCR. Mandibles, excised from 8 days op/op and wt/wt littermates, were examined for tooth morphology as well as amelogenin and DMP1 expression using in situ hybridisation. op/opCS transgenic mice, expressing csCSF-1 in teeth and bone using the osteocalcin promoter, were generated. Skeletal X-rays and histomorphometry were performed; teeth were analysed for morphology and matrix proteins. RESULTS: Normal dental cells in vivo express both CSF-1 isoforms and CSF-1R. Compared to wt/wt, op/op teeth prior to eruption showed altered dental cell morphology and dramatic reduction in DMP1 transcripts. op/opCS mice showed marked resolution of osteopetrosis, tooth eruption and teeth that resembled amelogenesis imperfecta-like phenotype. At 3 weeks, op/op teeth showed severe enamel and dentin defects and barely detectable amelogenin and DMP1. In op/opCS mice, DMP1 in odontoblasts increased to near normal and dentin morphology was restored; amelogenin also increased. Enamel integrity improved in op/opCS, although it was thinner than wt enamel. CONCLUSIONS: Results demonstrate that ameloblasts and odontoblasts are a source and potential target of CSF-1 isoforms in vivo. Expression of csCSF-1 within the tooth microenvironment is essential for normal tooth morphogenesis and may provide a mechanism for coordinating the process of tooth eruption with endogenous matrix formation.

Ameloblasts↗
PubMed2004

Differential expression patterns of the dentin matrix proteins during mineralized tissue formation.

Sequential and reciprocal interactions between the oral ectoderm and neural crest-derived mesenchyme are responsible for tooth development. During dentin formation, there are three components that are necessary for proper mineralization, namely, collagen which forms a scaffold, noncollagenous proteins that can specifically bind to the collagen template and function as a mineral nucleator and crystalline calcium phosphate deposited in an ordered manner. It is well established that noncollagenous proteins play an important role during mineralized tissue formation. Here we demonstrate by in situ hybridization techniques that the noncollagenous dentin matrix proteins 1, 2 (DMP1, 2) and dentin sialoprotein (DSP) have characteristic temporal and spatial expression patterns within odontogenic tissues during dentin mineralization. DMP1, DMP2 and DSP mRNA are expressed in the odontoblasts at specific and overlapping time points and are thus presumably used for different functions during dentin formation. In developing rat incisors and molars, high levels of expression of DMP2 mRNA were seen in polarized odontoblasts and preameloblasts, while DSP mRNA was expressed at significantly lower levels and was expressed by highly differentiated odontoblasts. However, their expression was continuously maintained during the mineralization of the organic matrix. In the adult rats, DMP2 and DSP mRNA was also detected in the osteoblasts. The expression of DMP1 mRNA was found to coincide with the start of the mineral nucleation process and gradually decreased during the maturation of the mineralized matrix during odontogenesis. In this study, we have also correlated the expression of these proteins relative to the presence of type I collagen and calcium phosphate crystals. Thus, the temporal and spatial differences between DMP1, DMP2 and DSP might implicate a direct demonstration of the functional difference between these three genes during calcified tissue formation.

Animals↗
PubMed2005

In vitro cytotoxicity of a remineralizing resin-based calcium phosphate cement.

UNLABELLED: Recently, a resin-based calcium phosphate cement (RCPC) has been reported as a remineralizing pulp-capping or lining cement. RCPC consists mainly of tetracalcium and dicalcium phosphates, ethoxylated bisphenol A dimethacrylate and pyromellitic glycerol dimethacrylate monomers and photo- and chemical initiators. OBJECTIVES: Here, the cytotoxic effects of RCPC were evaluated. The hypothesis was that RCPC induced only minor cytotoxic response in immortalized murine odontoblast and pulp cells, comparable to that produced by similar dimethacrylates due to unpolymerized dimethacrylate monomer present after curing. METHODS: Cytotoxicity was determined following the changes in cell succinate dehydrogenase activity after 24 h exposure to the cement components and after a 24 h recovery period. A fourfold range of concentrations was tested of the monomers, the eluate of cured RCPC leached in Dulbecco's modified Eagle's medium, and crushed cured cement in dimethyl sulfoxide. RESULTS: The monomers themselves had cytotoxicities similar to those reported for other dimethacrylates, although they are significantly less toxic than Bis-GMA. Differential cell sensitivity was demonstrated, with the pulp cells having greater sensitivity to the unpolymerized monomer than the odontoblast-like cells. The leached components have cytotoxicity similar to that of the free monomers. The crushed material demonstrated no apparent cytotoxicity at the dilutions tested. SIGNIFICANCE: These data demonstrate that RCPC has an in vitro cytotoxicity that is comparable to other materials containing dimethacrylate monomers and suggest that the material may be suitable for use in dental restorations. The data also indicate that the pulp cells appear more sensitive to dimethacrylates than the odontoblasts.

Animals↗
PubMedDate not supplied

Effects of a functional agar surface on in vitro dentinogenesis induced in proteolytically isolated, agar-coated dental papillae in rat mandibular incisors.

In an attempt to study the effects of a three-dimensional agar surface on in vitro dentinogenesis both in the growing end and in incisally cross-cut pulp, the possible expression of odontoblast phenotype was investigated morphologically, autoradiographically and immunohistochemically. Explants were incubated for 8 days. In the growing end, during the last 4 days, mitotic cells differentiated into [3H]-thymidine-labelled, tubular matrix-forming cells. In cross-cut pulp, however, during the first 4 days, mitotic cells differentiated into [3H]-thymidine-labelled, tubular matrix-forming cells. Electron microscopy demonstrated that, in both regions, tubular matrix-forming cells had characteristics similar to those of primary odontoblasts. When agar was incubated alone, exogenous fibronectin was deposited on it rapidly. After 12 h, endogenous fibronectin appeared on explant peripheral cells. Collagen and materials reacting positively to periodic acid-Schiff (PAS) were first interposed between agar and explant after 4 days. After 8 days, an inner immunonegative layer corresponding to materials reacting positively to PAS or toluidine blue and an outer immunopositive layer of fibronectin or collagen were visible adjacent to the rows of elongated columnar cells. In the presence of Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP), a competitive inhibitor of attachment of cells to fibronectin, explants became detached from the agar surface, and no dentinogenesis occurred. These results indicate that, when in contact with an agar surface that becomes modified by fibronectin and/or by a complex of fibronectin with deposited matrix, dental mesenchymal cells progressively differentiate into tubular matrix-forming cells. Possibly the functional agar surface has the important role of providing a foothold for cell attachment, which is the first step towards in vitro odontoblast differentiation. This system of inducing tubular matrix-forming cells constitutes a useful model for the study of in vitro dentinogenesis.

Agar↗
PubMed1998

Effects of recombinant basic fibroblast growth factor, insulin-like growth factor-II and transforming growth factor-beta 1 on dog dental pulp cells in vivo.

The effects of recombinant basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF)-II and transforming growth factor (TGF)-beta 1 on dental pulp cells were investigated by light and transmission electron microscopy after their implantation for 1 and 3 weeks at central sites of mechanically exposed pulps in dog molar and canine teeth. The implants were Millipore filters that have been soaked with solutions containing 100 or 500 ng/ml of bFGF or IGF-II or 100 ng/ml of TGF-beta 1. Control filters were soaked with dog albumin. No changes in cell organization or matrix synthesis were seen after implantation of control filters. Groups of columnar, polarized cells with numerous mitochondria and Golgi elements or elongated cells unassociated with any matrix deposition were demonstrated after 1 or 3 weeks, respectively, in close proximity to the filters that had been soaked with bFGF solution; at a distance from these implants enhanced formation of an osteotypic matrix was seen beneath the exposure site. No particular response was found in close proximity to the filters that had been soaked with IGF-II solution after 1 or 3 weeks implantation but thick zones of osteodentine were found beneath the exposure site and at adjacent circumferential dentine sites. Numerous elongated, polarized cells with long cytoplasmic extensions invading the filter pores were consistently seen after 1 week in close proximity to the filters that had been soaked with TGF-beta 1 solution. After 3 weeks implantation of these filters, deposition of a tubular matrix surrounding the implants was seen in association with the highly elongated odontoblast-like cells, while enhancement of circumferential dentine formation was also found at adjacent peripheral sites. These experiments demonstrate that TGF-beta 1 when implanted for short term periods at central pulp sites exerted dentine-specific effects, inducing differentiation of odontoblast-like cells and stimulating primary odontoblasts. Implantation of bFGF and IGF-II did not result in reparative dentine formation, but did stimulate osteotypical matrix deposition at a distance from the implants.

Animals↗
PubMed1999

Neurokinin-1 receptor expression in the mature dental pulp of rats.

Substance P induces inflammatory reactions in peripheral tissues including the dental pulp, but its regulatory effects in target tissues are dependent on receptor signalling. Here the expression of the substance-P receptor neurokinin-1 (NK1) in the mature molar pulp of the rat was examined in order to localize the main target areas for substance P. A polyclonal antibody directed against the C-terminal of the receptor was used, and immunohistochemistry was performed by the avidin-biotin peroxidase complex method. The results showed that the NK1 receptor was intensely expressed along vessel-like structures in the odontoblast and subodontoblast layer. A granulated and diffusely distributed NK1-receptor labelling was found along larger blood vessels in the root pulp and pulp proper. NK1 receptor-positive cells were frequently observed in the cell-rich zone beneath the odontoblast layer. The results indicate that, in the mature rat molar pulp, the main targets for substance P acting through the NK1 receptors are tissues related to blood vessels in the odontoblast and subodontoblast area. Furthermore, the expression of NK1 receptors on cells located in the subodontoblast area could indicate that substance P also affects cell functions in this area.

Animals↗
PubMed2003

Notch2 protein distribution in human teeth under normal and pathological conditions.

Notch signaling is essential for the appropriate differentiation of many cell types during development and, furthermore, is implicated in a variety of human diseases. Previous studies have shown that although the Notch1, -2, and -3 receptors are expressed in developing and injured rodent teeth, Notch2 expression was predominant after a lesion. To pursue the role of the Notch pathway in tooth development and disease, we have analyzed the expression of the Notch2 protein in embryonic and adult wounded human teeth. During the earlier stages of tooth development, the Notch2 protein was expressed in the epithelium, but was absent from proliferating cells of the inner enamel epithelium. At more advanced stages, Notch2 was expressed in the enamel-producing ameloblasts, while it was absent in mesenchyme-derived odontoblasts that synthesize the dentin matrix. Although Notch2 was not expressed in the pulp of adult intact teeth, it was reexpressed during dentin repair processes in odontoblasts and subodontoblastic cells. Transforming growth factor beta-1, which stimulates odontoblast differentiation and hard tissue formation after dental injury, downregulated Notch2 expression in cultured human dental slices, in vitro. These observations are consistent with the notion that Notch signaling is an important element in dental physiological and pathogenic conditions.

Adult↗
PubMed2002

Different expressions of connexin 43 and 32 in the fibroblasts of human dental pulp.

The expression and localization of gap junctional proteins connexin (Cx) 26, 32, and 43 was examined in human dental pulp. Dental pulp tissues were obtained from human third molars immediately after extraction. Some pulp tissues were used for cell culture, and the rest for histological observations. Immunostaining for cultured dental pulp fibroblasts (DPFs) showed that Cx32 and 43 were expressed in human DPFs, and proteins corresponding to 27 (Cx32) and 43kDa (Cx43) were identified by Western blot analysis. Immunostaining for tissue sections showed that the expression of Cx32 and 43 was observed in the entire region of the pulp and further strong expression of Cx32 was established beneath the cell-rich zone. Considering the close relationship between Cx types and cell functions, the results indicate that DPFs beneath the cell-rich zone may have specific, Cx32-related functions. The cell rich zone is thought to contain progenitor odontoblasts that can be induced to differentiate into mature odontoblasts in response to wounding. Therefore, it may be hypothesized that DPFs just beneath the cell-rich zone produce proteins and induce odontoblast differentiation from the cells in the cell-rich zone.

Adult↗
PubMed1993

Inductive effect of native dentin on the dentinogenic potential of adult dog teeth.

Autogenous dentinal matrix was exposed to the pulp cells of adult dogs in order to determine whether the mature pulp cells possess the ability to differentiate into odontoblast-like cells as a direct response to known inductive influences. The pulps of molars, premolars, and canines of three dogs (2 to 4 yr old) were mechanically exposed through buccal class V cavities. Pieces of demineralized or native dentin and predentin were implanted in the pulp sites for periods of 2 to 6 wk. The reactions were analyzed by light microscopy. Induction of dentin formation was observed only after native dentin implantation; either as early response to exposure of predentinal surfaces or around mineralized dentin after 3 postoperative wk. Encapsulation by fibrous connective tissue or matrix degradation was seen around demineralized dentin implants. A characteristic enhancement of circumferential pulpal dentin deposition around the implantation site was demonstrated after native dentin exposure to light and scanning electron microscopic examination. These data indicate that specific inductive influences given by the native but not the acid-conditioned dentin, when it is exposed to the pulp environment of adult teeth, are able to direct differentiation of odontoblast-like cells and to enhance the biosynthetic activity of primary odontoblasts.

Animals↗
PubMed1996

Electron microscopic study on interodontoblastic collagen fibrils in amputated canine dental pulp.

The purpose of this research was to study the presence and the ultrastructural features of interodontoblastic collagen fibrils and their contribution to the formation of reparative dentin in dog incisors and premolars for a period of 30 days following experimental pulpotomy. On the seventy day after pulp exposure and capping with calcium hydroxide, short cylindrical-shaped cells collected at the coronal end of the vital pulp tissue. Many collagen fibrils were synthesized in the intercellular spaces and in the direction of the long axis of these cells. On the fourteenth day, large bundles of collagen fibrils were observed in the intercellular spaces of young odontoblasts. These bundles spread out in a fan-shaped arrangement. On the thirtieth day, the odontoblasts formed a tubular dentin matrix. Small numbers of twisted collagen bundles crossed the distal junctional complex of the odontoblast cell bodies and entered the dentin matrix.

Animals↗
PubMed1986

Localization of type V collagen with monoclonal antibodies in developing dental and peridental tissues of the rat and hamster.

The distribution of collagen type V in developing dental and peridental tissues was investigated with the indirect immunofluorescence technique using unfixed, frozen sections of jaws from 1-2 day old neonatal rats and hamsters. Immunostaining for type V collagens was found both intracellularly and extracellularly in dental tissues of mesenchymal origin. In non-dental tissues, weak immunostaining was observed in the mesenchymally derived stroma surrounding the developing molar tooth germs but was more pronounced in larger cells, probably young osteoblasts in close vicinity to alveolar bone, and in some cells within developing salivary glands. Decalcification revealed a strong immunostaining in the extracellular bone matrices. In the dental tissues, the mesenchymally derived cells of the papilla exhibited an intracellular staining for type V collagen. In odontoblasts, increased immunostaining over that of other pulpal cells was observed just prior to or coinciding with the onset of predentin secretion and reactivity remained high in fully differentiated odontoblasts. A weak staining was observed in predentin but only after the onset of mineralization. As was the case for bone, after demineralization the dentin matrix stained intensely for type V collagen. The results demonstrate that type V collagen is actively synthesized by mesenchymal cells of developing hard tissues and that this type of collagen is an intrinsic component of hard connective tissue matrices. The data suggest that in developing tooth germs type V collagen is not involved in the differentiation process of either odontoblasts or ameloblasts.

Animals↗
PubMed2001

Transdentinal stimulation of reactionary dentinogenesis in ferrets by dentine matrix components.

OBJECTIVES: Trans-dentinal stimulation of reactionary dentinogenesis may be mediated through cellular signalling by bio-active components released from the dentine matrix during injury. Understanding of these processes will be important to guide dentinal repair activity following restorative surgery. The purpose of this study was to investigate the effects of implanting isolated dentine matrix proteins within cavity preparations on dentinal repair activity and odontoblast survival using a controlled experimental animal model. METHODS: Forty-five ferret canine teeth each had a standardised non-exposed cylindrical Class V cavity cut into the buccal dentine. Ten cavities were restored with zinc oxide eugenol, as a control. Two different lyophilised preparations of dentine matrix components were implanted on the axial floor of the remaining 35 cavities prior to filling with zinc oxide eugenol. After post-operative periods of 2, 7, 14, 28, and 90 days, the teeth were extracted and examined histomorphometrically and the data analysed statistically by analysis of variance tests. RESULTS: The odontoblasts beneath the restored cavities responded to the presence of the two dentine matrix preparations by increasing the mean area of reactionary dentine secreted by 433 and 578%, and the numbers of odontoblasts remained stable. CONCLUSION: Dentine matrix components can stimulate reactionary dentinogenesis in non-exposed cavity preparations. It will now be important to identify how this may be harnessed as a part of routine restorative surgery to optimise treatment outcomes with a biological basis.

Analysis of Variance↗
PubMed2001

Inositol hexasulphate, a casein kinase inhibitor, alters the distribution of dentin matrix protein 1 in cultured embryonic mouse tooth germs.

Immunohistochemical studies using a polyclonal antibody, raised against the recombinant form of dentin matrix protein 1 (DMP1), show that DMP1 was detected mainly in odontoblasts in cultured mouse embryonic tooth germs. However, in restricted areas, DMP1 staining was also observed in secretory ameloblasts, in the stratum intermedium and stellate reticulum, but only when the odontoblasts located in front of them were unstained. When the embryonic tooth germs were cultured in the presence of inositol hexasulfate, a casein kinase I and II inhibitor, staining of odontoblasts was weak or nil, whereas, in contrast, ameloblasts and enamel organ were strongly immunolabelled, suggesting an enhanced translocation of DMP1 after secretion to the secretory ameloblasts and/or stratum intermedium and stellate reticulum. Moreover, DMP1--was shown to be a good substrate for gelatinase A (MMP-2), but not to gelatinase B (MMP- 9). We hypothesized that DMP1--or the sub-fractions cleaved by the MMP--could behave as diffusible signaling molecule (s) rather than as a true dentin extracellular matrix component.

Ameloblasts↗
PubMed2003

Remaining dentine thickness and human pulp responses.

AIM: To evaluate pulp responses as a function of remaining dentine thickness (RDT) of 98 class V cavity preparations in 49 teeth of 31 patients aged 10-16 years. METHODOLOGY: Shallow cavities were restored with amalgam, deeper cavities or pulp exposures were restored with amalgam lined with calcium hydroxide or with zinc oxide eugenol. Teeth were extracted after 3-89 days for orthodontic reasons. Following processing for light microscope analysis, the number of odontoblasts, pulp inflammation, and repair was recorded. RESULTS: In comparison with independent odontoblasts, the numbers of odontoblasts were reduced by 13.6% beneath a RDT of 2.5-0.5 mm, 33.7% beneath a RDT of 0.5-0.01 mm and 99.0% beneath pulp-exposed cavities. Reparative dentine was observed following pulp exposure and reactionary dentine was observed with a mean RDT of 0.77 mm (2.5-0.01 mm). Reactionary dentine secretion was influenced by RDT and restorative materials. Pulp inflammation was not influenced by RDT in the present study. CONCLUSIONS: Cavity RDT mediates a powerful influence on underlying pulp tissue vitality but it has little effect on reactionary dentine secretion and inflammatory activity. Gross tissue injury explains the poor pulp capping prognosis following exposure and underlies the need to avoid this type of injury. Following restoration, a RDT of 0.5 mm or greater is necessary to avoid evidence of pulp injury.

Adolescent↗
PubMed2002

Age-related odontometric changes of human teeth.

OBJECTIVE: The number of older patients requiring restorative treatment are likely to increase due to improvements in oral health and increased longevity. However, aging odontometric data are lacking. The aim of this study was to determine possible changes in pulp cell density, pulp area, and dentinal thickness with age. STUDY DESIGN: Incisors (50), canines (39), premolars (51), and molars (7) extracted from 60 patients aged between 10 and 59 years, were analyzed histomorphometrically for cell density (odontoblasts, subodontoblasts, and pulp core fibroblasts) and dentinal thickness. RESULTS: With increasing patient age, in both crown and root aspects of teeth, dentinal thickness increased (P <.001), while the density of odontoblasts (P <.001), subodontoblasts (P = 0.001), and pulp fibroblasts (crown, P <.011; root, P =.0015) decreased. The degree of age-related changes in teeth appeared to be asymmetrical, with decreases in the root being greater than in the crown. At all ages pulp cell densities, including odontoblasts, within the crown were greater than in the root (P <.001), even though the calculated rate of dentinal deposition was greatest in the root. CONCLUSION: Decreases in pulp cell density may reduce pulp repair activity after restorative treatments, although increases in dentinal thickness may aid pulp protection. An understanding of these age-related changes will influence the provision of restorative and endodontic care and benefit older patients.

Adolescent↗