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PubMed1992

A light and electron microscopic study on pulpal nerve fibers in the lower incisor of the mouse.

Light and electron microscopic studies were made on pulpal nerve fibers in mouse lower incisors, typical continuously growing teeth. Serial sections, from the apex of the odontogenic sheath to the incisal edge of the apical foramen, were examined by light microscopy to identify myelinated fibers passing through the apical foramen. The fine structure of the pulpal nerves was examined by electron microscopy at three sites: 1) the level at the incisal edge of the apical foramen; 2) a level 5 mm incisal from the apex of the odontogenic sheath; and 3) the level where the incisor comes out of the alveolar bone. No myelinated fibers were found passing through the apical foramen; they were also lacking at the three levels of the pulp. At level 2, unmyelinated axons were seen in close contact with smooth muscle fibers of arterioles. At level 3, nerve fibers were difficult to distinguish from processes of fibroblasts and odontoblasts. Degenerating axons were present in Schwann cells, and fine unmyelinated axons running through the odontoblast cell layer were seen. Various types of unmyelinated axons were observed in the apical region (level 1). These axons were classified into 6 types on the basis of their fine structures: Type I, bundles of unmyelinated axons completely or partly ensheathed by Schwann cell cytoplasm (mature type); Type II, bundles of unmyelinated axons in a space formed by a Schwann cell membrane (regenerating type); Type III, bundles of unmyelinated axons ensheathed not by a Schwann cell, but merely by a basal lamina (regenerating type); Type IV, single axons in direct contact with the basal lamina (regenerating or terminal type); Type V, naked, electron-dense axons with many vesicles and mitochondria (growth cone-like type); and Type VI, electron opaque axons, due to loss of axonal organellae (degenerating type). The significance of these structures is discussed in relation to the continuous growth of the rodent incisor.

Animals↗
PubMed1998

Immunohistochemical localizations of class II antigens and nerve fibers in human carious teeth: HLA-DR immunoreactivity in Schwann cells.

Nerve fibers and class II major histocompatibility complex (MHC) antigen-expressing dendritic cells have been known to gather in the dental pulp beneath carious lesions. Significant functional interactions presumably occur between the neural and immune elements. The present study analyzed the morphological relationship between class II-expressing cells and nerve fibers in fuman carious teeth, visualized by a HLA-DR monoclonal antibody and a protein gene product 9.5 (PGP 9.5) polyclonal antibody; a confocal laser scanning microscope (CLSM) and an electron microscope were used. In pulps affected by early caries, HLA-DR-positive dendritic cells aggregated mainly in the cell-free zone associated with bundles of PGP 9.5-immuno-reactive nerve fibers. In pulps affected by advanced caries, the accumulated HLA-DR-positive cells and PGP 9.5-immunoreactive nerve fibers showed close association with each other especially beneath the odontoblast layer: the cells even embraced the nerve fibers. Intriguingly, class II molecules were recognized not only in dendritic cells but also in the Schwann cells of non-myelinated nerves in the pulp. Using immuno-electron microscopy, class II molecules were localized on the surface of the non-myelinating Schwann cells and also within some vesicles, whereas myelinating Schwann cells lacked this immunoreactivity. PGP 9.5-immunoreactive nerve fibers were also observed densely in the odontoblast layer, and CLSM revealed an intimate association of the nerve fibers and dendritic cells. The immunoreactivity for HLA-DR in Schwann cells depended upon the severity of the carious lesion. Class II-expressing Schwann cells are suggested to function as antigen-presenting cells in addition to dendritic cells.

Adult↗
PubMed1978

Alkaline phosphatase in developing teeth and bone of man and macaque monkey.

The activity of nonspecific alkaline phosphatase (E.C. 3.1.3.1) in developing teeth and bone of human fetuses and young macaque monkeys has been studied by means of histochemistry. The incubations for alkaline phosphatase were performed at pH 8.2 using naphthol-AS-MX-phosphate as substrate and Fast Blue RR salt or Fast Red Violet LB salt as couplers. By means of pretreatment with heat (56 degrees C), or addition of sodium metavanadate, ortho- or pyrophosphate, two alkaline phosphatases were demonstrated in the developing teeth. Prior to hard tissue formation all alkaline phosphatase activity was inhibited by the addition of vanadate, phosphate, or by pretreatment with heat. Pretreatment with heat or addition of vanadate or phosphate also inhibited alkaline phosphatase activity in the odontoblasts and in the pulpal connective tissue, whereas the activity in the subodontoblastic cell layer, stratum intermedium, outer enamel epithelium, and the the outer cells of the reduced enamel epithelium were much less affected. A week resistant activity was also noted in odontoblasts and pulpal connective tissue.

Alkaline Phosphatase↗
PubMed1998

Cloned 3T6 cell line from CD-1 mouse fetal molar dental papillae.

Only primary pulpal cell cultures and one virally transformed mouse cell culture have been formally reported in the literature to synthesize proteins such as phosphophoryn which are unique to dentin matrix. In the present study, a mixed culture was derived from dental papilla cells of 18-19 fetal day CD-1 mouse mandibular first molars, maintained on a 3T6 plating regimen, and subsequently cloned after 28 passages. This cloned cell line (MDPC-23) exhibited several unique features, some of which were characteristic of odontoblasts in vivo. The features of this cell line included (1) epithelioid morphology of all cells with multiple cell membrane processes, (2) high alkaline phosphatase activity in all cells, (3) formation of multilayered nodules and multilayered cultures when maintained in ascorbic acid and beta-glycerophosphate, and (4) expression of two markers for odontoblast differentiation, i.e. dentin phosphoprotein and dentin sialoprotein.

Animals↗
PubMed1998

Expression of dentin sialoprotein (DSP) and other molecular determinants by a new cell line from dental papillae, MDPC-23.

The purpose of this study was to characterize the molecular expression of a spontaneously immortalized and cloned cell line (MDPC-23) derived from 18-19 day CD-I fetal mouse molar dental papillae to determine if these cells were odontoblast-like. Western blots showed that a protein band, at approximately 105 kDa, reacting positively with anti-DSP antibodies and co-migrating with mouse DSP, was present in lysates of cells from passages 7, 37 and 77, in serum-free conditioned medium from passage 37 cells, and in mouse dentin extract. A minor band at 55 kDa was also apparent in cell lysates. Using a cDNA probe for a 486bp mouse DSP coding sequence, DSP or DSP-PP mRNA expression was detected by Northern analysis as well as Southern analysis after RT-PCR in all three passages. It was also shown that in these cells 1,25 (OH)2 vitamin D3 upregulated both osteopontin and osteocalcin mRNA, and dexamethasone downregulated alkaline phosphatase and alpha2(I) collagen mRNA. Thus, MDPC-23 cells express proteins which are common to mineralizing tissue. The expression of DSP and DSP-PP strongly suggests that this cell line is from the odontoblast lineage.

Animals↗
PubMed1998

Mechanisms of mineralization in the enameloid of elasmobranchs and teleosts.

Ultrastructural and cytochemical studies on the mineralization of enameloid were performed using Heterodontus japonicus, an elasmobranch, and Tilapia buttikoferi, a teleost as materials. The mineralization of the enameloid in the Heterodontus was divided into the following two steps: (1) initial crystallization in the tubular vesicles that originated from the odontoblasts, and (2) crystal growth that was accompanied by the degeneration and removal of the organic matrix around the crystals. In the Tilapia, the mineralization of the cap enameloid followed three steps: (1) initial crystallization at the matrix vesicles, (2) aggregation of fine slender crystals along collagen fibrils, and (3) crystal growth with the degeneration and removal of the organic matrix. The pattern of early mineralization and the composition of organic matrix in enameloid were considerably different between the two species examined, while in both species the odontoblasts were mainly involved in the formation of the organic matrix of enameloid and in the initial mineralization. In the next step, remarkable crystal growth associated with the degeneration and removal of the organic matrix occurred in both the elasmobranch and the teleost species. The absorptive functions of the dental epithelial cells in the later stages of enameloid formation is probably similar in the two types of enameloid, and is essential for the production of well-mineralized enameloid.

Alkaline Phosphatase↗
PubMed1996

In situ hybridization evidence for a paracrine/autocrine role for insulin-like growth factor-I in tooth development.

Insulin-like growth factor-I(IGF-I) has both metabolic and growth-promoting activities in many cell and tissue types. Although IGF-I is present in serum, it is also thought to have important autocrine and paracrine functions. Immunohistochemistry for IGF-I and its receptor have shown that IGF-I is synthesised locally by the tooth forming cells which exhibit both the IGF-I and the growth hormone receptors. This concept required to be tested by in situ hybridization. Using a digoxigenin-labelled synthetic oligodeoxyribonucleotide probe for IGF-I, we investigated the distribution of IGF-I mRNA in the continuously erupting rat incisor by in situ hybridization. The distribution and intensity of the hybridization signal varied with the developmental stage of the rat incisor. The cells of the apical loop expressed a positive hybridization signal, but the earliest polarised odontoblasts and pre-ameloblasts did not show any positive signal. The onset of enamel secretion was accompanied by a strong hybridization signal in the secretory ameloblasts as well as the odontoblasts. Maturation ameloblasts also demonstrated IGF-I message in their cytoplasm as well as their nuclei. The cells of the pulp and the dental follicle were consistently negative. However, in the adjacent alveolar bone, the signal was high in the osteoblasts and osteoclasts. These findings support the notion of paracrine or autocrine function for IGF-I in tooth development.

Alkaline Phosphatase↗
PubMed1998

Basic fibroblast growth factor (FGF-2) in mouse tooth morphogenesis.

This study describes the spatio-temporal expression of basic Fibroblast growth factor (FGF-2) during odontogenesis of mouse as revealed by immunohistology. Parasagittal sections of mouse embryo head (13-18 day of gestation) containing various stages of developing tooth were incubated with a polyclonal anti-FGF-2 antibody and positive binding was evidentiated by using Streptavidin-Biotin complex-HRP system and AEC staining. We observed no FGF-2 staining at the dental lamina stage. At the bud stage slight staining is seen, limited to some epithelial cells. The intensity of the staining increases at the cap stage. In the bell stage, the stellate reticulum cells stain intensely. Later, odontoblasts and the dentin matrix stain deeply; but the epithelial cells stain faint. The extra cellular matrix of the dentin and dental papilla stain very intense but the enamel matrix is found negative. These results indicate the participation of FGF-2 in differentiation rather than in proliferation of tooth-forming cells. In particular, it appears that FGF-2 participates in odontoblast differentiation and in dentin matrix deposition. The spatio-temporally specific distribution pattern of FGF-2 in developing mouse tooth reported here emphasizes the importance of FGF-2 in mammalian odontogenesis.

Animals↗
PubMed1999

Ultrastructural study of fossil human tooth tissue.

The preservation of fossil human soft tooth tissue from extinct populations which inhabited the northeast of Argentina (fourteenth and fifteenth centuries) is described. Studies were performed using both Scanning Electron Microscopy (SEM) and Transmission Electron Microscopy (TEM). The preservation of the surface structure in Tomes fibrils and in odontoblastic processes was determined by SEM; whereas by TEM we could observe the dentin ducts occupied by acellular material, being the inner structure of the odontoblastic processes poorly preserved. It is suggested that such preservation is due to an "instantaneous phosphatization" occurring immediately after death as a result of the presence of calcium phosphate deposits.

Archaeology↗
PubMed2000

Actions of bovine plasma fibronectin on cultured human dental pulp cells.

OBJECTIVE: Fibronectin (FN) has been shown to play a crucial role in odontoblast differentiation, and it is involved in the repair of dentine and pulp. The responsiveness of human dental pulp (HDP) cells to bovine plasma FN was investigated. MATERIALS AND METHODS: After isolation and selection noncarious freshly extracted third molars in vitro, HDP cells were incubated for 72 hours one of four concentrations (10, 20, 40, 80 micrograms/ml) of bovine plasma FN with Dulbecco's modified eagle medium (DMEM) containing 1% fetal calf serum (FCS). We examined cell proliferation (using an MTT assay), DNA synthesis (using incorporation of [3H]-thymidine), the cell area, and alkaline phosphatase (ALPase) activity of the culture supernatant. The expression of FN, types I and III collagens, binding of lectins concanavalin(ConA), or wheat germ agglutinin (WGA) in vitro was examined by means of immunocytochemistry. The expression degree and the cell area were analyzed semi-quantitatively with an image processing and analysis system. RESULTS: After 72 hours, HDP cells appeared contracted and smaller in the absence of FN when compared to cells incubated with FN. HDP cells were in spread state with all four concentrations of FN. FN (20, 40 micrograms/ml) stimulated cell proliferation, promoted incorporation of [3H]-thymidine, and increased the levels of ALPase activity in supernatant. The binding of ConA, the expression of type I collagen, and FN were significantly increased, while the binding of WGA, and the expression of type III collagen were significantly decreased in the FN (40, 80 micrograms/ml) groups when compared to the FN (10, 20 micrograms/ml) groups. CONCLUSION: These findings suggest that HDP cells utilize FN for attachment and subsequent spreading. FN might have mitogenic activity and some role in the regulation of differentiation of pulp cells into odontoblasts.

Alkaline Phosphatase↗
PubMed1999

[Experimental study on the effect of restorative dentin formation with human dentin phosphoprotein in immature permanent teeth of minipig].

OBJECTIVE: To investigate the function of human dentin phosphoprotein(DPP) on dentine mineralization in vivo. METHODS: Pulp capping with human DPP was made in the permanent teeth of minipigs with calcium hydroxide and zinc oxide eugenol cement as controls. The minipigs were sacrificed at intervals of 14, 30, 90 days respectively after operation. RESULTS: The histopathological results revealed that pulp cells had developed into odontoblastoid cells around the major restorative dentin body after two weeks. There was complete restorative dentin bridge a month later. The bridge became thick and dense, and mainly was tubular dentin in 3 month. There were well developed odontoblasts under the bridge. In the calcium hydroxide capping group, inflammatory cytonecrosis zone was found in the surface of dental pulp and only a little calcial bodies formed in two weeks. Complete restorative dentin bridge was formed in 1 to 3 months, the speed of dentin bridge formation was not as quick as that of DPP group. CONCLUSION: DPP can induce pulp cells to develop into odontoblasts and formation of dentin bridge with little stimulation to the dental pulp. This study suggests that DPP plays an important role in the biomineralization of dentin development and restoration.

Animals↗
PubMed2001

[Lasers in dentistry. Part B--Interaction with biological tissues and the effect on the soft tissues of the oral cavity, the hard tissues of the tooth and the dental pulp].

The interaction of laser energy with target tissue is mainly determined by two non operator-dependent factors: the specific wavelength of the laser and the optical properties of the target tissues. Power density, energy density, pulse repetition rate, pulse duration and the mode of energy transferring to the tissue are dictated by the clinician. Combination of these factors enables to control optimal response for the clinical application. Four responses are described when the laser beam hits the target tissue: reflection, absorption, transmission and scattering. Three main mechanisms of interaction between the laser and the biological tissues exist: photothermic, photoacoustic and photochemical. The effect of lasers on the soft tissues of the oral cavity is based on transformation of light energy into thermal energy which, in turn heats the target tissue to produce the desirable effect. In comparison to the scalpel used in surgical procedures, the laser beam is characterized by tissue natural sterility and by minimum bleeding during the surgical procedures due to blood vessels welding. The various effects achieved by the temperature elevation during the laser application on the soft tissue are: I. coagulation and hemostasis II. tissue sterilization III. tissue welding IV. incision and excision V. ablation and vaporization Ablation and melting are the two basic modalities by which the effect of lasers on the hard tissues of the tooth is produced. When discussing the effect of laser on dental hard tissues, the energy absorption in the hydroxyapatite plays a major role in addition to its absorption in water. When laser energy is absorbed in the water of the hard tissues, a rapid volume expansion of the evaporating water occurs as a result of a substantial temperature elevation in the interaction site. Microexplosions are produced causing hard tissue disintegration. If pulp temperatures are raised beyond 5 degrees C level, damage to the dental pulp is irreversible. Histologically, after laser ablation, presence of odontoblastic nuclei is important. Consistency and composition of the intracellular tissue is another factor influencing cell viability. If heat is intensive and exists for an extended time, the consistency of the intracellular ground substance may not be preserved. Accordingly, the application of excessive energy densities has been shown to result in significant damage to pulp tissue and in particular to odontoblasts. Studies showed that the use of Er:YAG laser to treat dental hard tissues is both safe and effective for caries removal, cavity preparation and enamel etching.

Absorption↗
PubMed2001

[Expression changes of nerve growth factors (NGF) in pulps of injury teeth].

OBJECTIVE: The aim of this study is to investigate the expression and functions of nerve growth factors during teeth injury. METHODS: Twenty male Wistar rats were given mechanical irritation in upper first molar and their molecular and biological changes were examined in different periods. Four of them were killed immediately after the teeth were irritated, the other four were killed 3 days, 5 days, and 9 days respectively after the teeth were stimulated. The perfusion was performed through the left ventricle with 4% paraformaldehyde and 0.2% picric acid in 0.1% phosphate buffer. The maxilla was decalcified and embedded. Sections of 15 microns in thickness were cut with freezing macrodome in the sagittal planes. Then the sections were stained with immunohistochemical method to observe changes of NGF in the injured dental pulp, comparing with that in the normal dental pulp. In the end, we observed the NGF distribution in the pulp under optical microscope, and analyzed the expression level of NGF with the image pattern analysis instrument. The results were statistically analyzed. RESULTS: In the normal group, the NGF expression was not observed. In the experimental group of zero hour, the NGF expression could be observed, but was very low. In the experimental group of three days, the NGF expression increased greatly, and the positive sites mainly distributed in the odontoblastic and the sub-odontoblastic layer. They also distributed in dentinal tubules. But in the group of 5 days, the level of NGF decreased. In the group of 9 days, it was the same as that of the normal group. Using the image pattern analysis instruments and statistical analysis, the results clearly showed that the expression level were lower in the group of three days than that of the other groups, and the expression levels of the groups 0 and 5 days were lower than that of the normal control group, however, that of 9-day group was the same as the normal group. CONCLUSION: NGF may play a role in the reaction and nerve restoration after teeth injury and, the expression change in different stages.

Animals↗
PubMed2003

[Effects of dentin sialophosphoprotein antisense oligodeoxynucleotide on ultrastructure of mouse tooth germ].

OBJECTIVE: To investigate more deeply the function and mechanism of DSPP during tooth development. METHODS: Explants of tooth germs from embryonic 17th day mice were divided into two groups. In the control experiment, explants were cultured in agarose semi-solid medium under serum-free and chemically defined conditions, while explants in the other group were cultured with 30 mumol/L, 15 bp antisense oligodeoxynucleotide targeted to DSPP mRNA. After 10 ds, the explants were examined by transmission electron microscope. The width of dentin matrix at the tip of the cusps were then measured and statistically analyzed with Student t-test. RESULTS: Ultrastructure analyses showed that large cisternae of the rough endoplasmic reticulum (RER) existed in the odontoblasts at the tip of the cusps of antisense-treated explants and the average thickness of dentin matrix (2.5 microns) was thinner compared to the control ones (3 microns, P < 0.001). In addition, the collagen fibers in extracellular matrix were disorganized. CONCLUSION: These findings indicated that DSPP played an important role in keeping tooth normal development, as well as in dentin mineralization by maintaining odontoblasts' secreting ability and controlling fiber structure and orientation.

Animals↗
PubMed2003

[Expression of homeobox gene Msx-1, Msx-2 and Dlx-2 during murine mandibular first molar development].

OBJECTIVE: To observe the expression of homeobox gene Msx-1, Msx-2 and Dlx-2 during murine mandibular first molar development. METHODS: The murine heads or mandibles on embryonic days 11-18 (E11-18) and postnatal day 1-3 (P1-3) were removed, fixed and embedded, 5 micro m serial sections were cut in the coronal plane. Msx-1, Msx-2 and Dlx-2 RNA probes were synthesized by in vitro transcription and labeled with digoxigenin. Msx-1, Msx-2 and Dlx-2 mRNA expression was observed after in situ hybridization. RESULTS: During molar development Msx-1 transcripts appeared only in mesenchymal cells, not in epithelial cells. Msx-2 and Dlx-2 both expressed in the epithelial and mesenchymal cells. At the initiation stage of the molar development Msx-2 and Dlx-2 had similar expression. At the bud stage (E13-14) Msx-2 mRNA signaling was intensive in the enamel organ and slight in the dental mesenchyme; Dlx-2 signaling was stronger in the dental papilla. At cap stage (E15-16) Msx-2 showed prominent mRNA signaling in enamel knot and Dlx-2 was maximal in the dental papilla. At the late bell stage (P2-3) Msx-2 transcripts were observed in odontoblasts but not labeled in ameloblasts, and Dlx-2 transcripts appeared in ameloblasts but no labeling was seen in odontoblasts. CONCLUSIONS: Msx-1, Msx-2 and Dlx-2 are expressed in various patterns during murine mandibular first molar development, suggesting they possibly play a role in the interaction between the epithelium and mesenchyme during the molar development.

Animals↗
PubMed2004

[Dynamics in the content of neuromediators in amine-containing structures during development of tooth hard tissues].

Using luminescent-histochemical methods the contents of neurotransmitters (catecholamines, serotonin, histamine) was determined during the development of dental hard tissues in man. In the dental papilla the luminescent blood vessels were initially demonstrated, while single nerve fibers branching around the odontoblasts appeared later. It was found that enamel organ differentiation at the early stages took place without the participation of granular luminescent cells and mast cells. These cells appeared starting at week 22 of fetal development. Their appearance coincided with the process of enameloblast and odontoblast differentiation. The contents of neurotransmitters in the dental structures studied was found to increase with the fetal age.

Adrenergic Fibers↗
PubMed1992

Effects of inflammation on dental sensory nerves and vice versa.

A brief review of the normal sensory innervation of teeth is presented, especially concerning the fibers that are immunoreactive (IR) for calcitonin gene-related peptide (CGRP). Numerous CGRP-IR fibers innervate coronal dentin at sites populated by primary odontoblasts and associated pulp cells that synthesize nerve growth factor (NGF). If the primary odontoblasts and adjacent pulp cells are lost as a consequence of injury, CGRP-IR dentinal innervation is greatly reduced. The responses of CGRP-IR nerve fibers to pulpal injury are reviewed. Those reactions show that pulpal sensory nerve fibers can alter the size, shape and immunoreactivity of their terminal branches in response to different stages of inflammation and healing.

Animals↗
PubMed2005

[Reconstruction of dentin-pulp complex structure by tissue engineering technology].

OBJECTIVE: To investigate the possibility of reconstruction of dentin-pulp complex by tissue engineering technology. METHODS: Rat dental pulp stem cells were seeded into HA-TCP scaffold and incubated for 20 hours in vitro. Then the cell-scaffold complex was implanted subcutaneously into the dorsal side of nude mice. 8 weeks postimplantation, the samples were extracted for histological and immunohistochemical examinations. RESULTS: Three strata of tissue were observed in the hole of HA-TCP scaffold. They were dentin-like tissue, predentin-like tissue and pulp-like tissue respectively from the inner surface of the pore to the center. Dentin tubules were obvious in predentin-like and dentin-like tissue lining from the pulp-like tissue through predentin-like tissue and dentin-like tissue. Cells localized along the edge of pulp-like tissue were dense and polarized, resembling odontoblasts. Immunohistochemical study demonstrated DSP and DMP1 expression in these odontoblast-like cells and in the area of predentin-like tissue. CONCLUSIONS: Tissue-engineered rat dentin-pulp complex was reconstructed by seeding HA-TCP scaffold with rat dental pulp stem cells.

Animals↗