Biomedical subjects

Shinichiro Oida

Publications and source records attributed to Shinichiro Oida.

10 recordsLinked to original sources

PubMed2007

Twist negatively regulates osteoblastic differentiation in human periodontal ligament cells.

Periodontal ligament (PDL) is a thin fibrous connective tissue located between two mineralized tissues, alveolar bone and cementum, which maintains a constant width physiologically. The mechanisms by which PDL resists mineralization are not well understood. Twist is a basic helix loop helix protein that plays a central role in regulation of early osteogenesis. We investigated the localization of Twist in PDL and compared the expression of Twist and osteoblast-related genes in PDL cells with those in osteoblast-like cells in the presence or absence of recombinant human bone morphogenetic protein (BMP)-2. Histochemical analysis showed that Twist was expressed along alveolar bone surface in PDL. PDL cells constitutively expressed Twist gene and the expression level was higher than that in osteoblast-like cells. In osteoblast-like cell culture, BMP-2 enhanced osteoblast-related gene expression, while Twist expression was slightly decreased. In contrast, BMP-2 increased runt-related transcription factor (Runx)-2, but failed to enhance alkaline phosphatase (ALP) and osteocalcin (OCN) gene expression in PDL cells. Interestingly, unlike in osteoblast-like cells, Twist expression was upregulated by BMP-2 in PDL cells. We transiently knocked down Twist gene in PDL cells using a short interference RNA expression vector (siTwist) and found that ALP, osteopontin (OPN), bone sialoprotein (BSP) genes expression and basal level of ALP activity were slightly increased, whereas Runx2 and OCN genes were not affected. Collectively, these results suggest that Twist may act as a negative regulator of osteoblastic differentiation in PDL cells.

Cell Differentiation↗

PubMed2006

Porcine sheath proteins show periodontal ligament regeneration activity.

The purpose of this study was to identify the periodontal regeneration factors of enamel protein extracts that induce cementum and bone regeneration in vivo. Cementum regeneration, one aspect of periodontal ligament regeneration, was examined using a buccal dehiscence model of dogs. Enamel matrix protein fractions were prepared from developing porcine incisors. Cementum-regeneration activity was found to reside in a protein aggregate composed of amelogenins and sheath proteins extracted from newly formed secretory enamel. Cementum-regeneration activity was not observed in protein fractions containing only amelogenin or its derivatives. The sheath proteins were purified to homogeneity and tested for alkaline phosphatase (ALP)-inducing activity on human periodontal ligament (HPDL) cells. The induction of ALP was observed following application of the 17-kDa sheath protein but not of the lower-molecular-weight sheath proteins. Although transforming growth factor-beta1 also shows ALP-inducing activity, contamination with growth factors was excluded because synthetic peptides (based on the sheath protein's sequence) also showed ALP-inducing activity. The 17-kDa sheath protein showed both cytodifferentiation and cementum-regeneration activity, but it is unclear whether its cementum-regeneration activity is derived from its ALP-inducing activity on HPDL cells.

Alkaline Phosphatase↗

PubMed2006

Porcine enamel protein fractions contain transforming growth factor-beta1.

BACKGROUND: Enamel extracts are biologically active and capable of inducing osteogenesis and cementogenesis, but the specific molecules carrying these activities have not been ascertained. The purpose of this study was to identify osteogenic factors in porcine enamel extracts. METHODS: Enamel proteins were separated by size-exclusion chromatography into four fractions, which were tested for their osteogenic activity on osteoblast-like cells (ST2) and human periodontal ligament (HPDL) cells. RESULTS: Fraction 3 (Fr.3) and a transforming growth factor-beta 1 (TGF-beta1) control reduced alkaline phosphatase (ALP) activity in ST2 but enhanced ALP activity in HPDL cells. The enhanced ALP activity was blocked by anti-TGF-beta antibodies. Furthermore, using a dual-luciferase reporter assay, we demonstrated that Fr.3 can induce the promoter activity of the plasminogen activator inhibitor type 1 (PAI-1) gene. CONCLUSION: These results show that the osteoinductive activity of enamel extracts on HPDL cells is mediated by TGF-beta1.

Alkaline Phosphatase↗

PubMed2005

Gene expression and localization of insulin-like growth factors and their receptors throughout amelogenesis in rat incisors.

Insulin-like growth factors (IGFs) are expressed in many tissues and control cell differentiation, proliferation, and apoptosis. In teeth, the temporo-spatial pattern of expression IGFs and their receptors has not been fully characterized. The purpose of this study was to obtain a comprehensive profile of their expression throughout the life cycle of ameloblasts, using the continuously erupting rat incisor model. Upper incisors of young male rats were fixed by perfusion, decalcified, and embedded in paraffin. Sections were processed for in situ hybridization and immunohistochemistry. mRNA and protein expression profiles IGF-I, IGF-II, IGF-IR, and IGF-IIR mRNA were essentially identical. At the apical loop of the incisor, very strong signals were seen in the outer enamel epithelium while the inner enamel epithelium showed a moderate reaction. In the region of ameloblasts facing pulp, inner enamel epithelium cells were still moderately reactive while signals over the outer enamel epithelium were slightly reduced. In the region of ameloblasts facing dentin and the initial portion of the secretory zone, signals in ameloblasts were weak while those over the outer enamel epithelium were strong. In the region of postsecretory transition, signals in both ameloblasts and papillary layer cells gradually increased. In maturation proper, signals in ameloblasts appeared as alternating bands of strong and weak reactivities, which corresponded to the regions of ruffle-ended and smooth-ended ameloblasts, respectively. Papillary layer cells also showed alternations in signal intensity that matched those in ameloblasts. These results suggest that the IGF family may act as an autocrine/paracrine system that influences not only cell differentiation but also the physiological activity of ameloblasts.

Amelogenesis↗

PubMed2005

Analysis of amelogenin gene (AMGX, AMGY) expression in ameloblastoma.

Although the amelogenin gene is expressed in ameloblastoma, the precise expression pattern of X and Y amelogenin genes (AMGX, AMGY) in this tumor has not yet been identified. In this study, we analyzed amelogenin gene expression in 19 samples (9 male, 10 female) of oral ameloblastomas by RT-PCR and detect the chromosomal origin of amelogenin mRNA by restriction enzyme digestion of the RT-PCR product. All tumor samples expressed amelogenin mRNA. We could detect increased level of AMGY expression in all male samples, higher than that of AMEX. It is an interesting finding as in normal male tooth development, the expression of AMGY is very much lower than that of AMGX. We postulate that epigenetic change of sex chromosomes may have some correlations with tumorigenesis of ameloblastoma. We also discuss the other possible mechanisms and points for future studies on this change in expression pattern.

Ameloblastoma↗

PubMed2004

Effect of heat treatment on bioactivities of enamel matrix derivatives in human periodontal ligament (HPDL) cells.

OBJECTIVES: It has been shown that Emdogain (EMD) containing enamel matrix derivative has cementum- and osteo-promotive activities in vivo and in vitro. Nevertheless, the commercial sale of EMD was halted because it has some possible risk for infectiosity. At present, Emdogain Gel (Emd-Gel) containing the EMD heated to avoid the infectiosity is commercially available. The purpose of this study was to compare the in vitro bioactivities of Emd-Gel and EMD. MATERIAL AND METHODS: Healthy human periodontal ligament (HPDL) cells were used to study the effect of Emd-Gel and EMD on cell differentiation. The HPDL cells exposed to Emd-Gel and EMD were evaluated for the following effects: (i) alkaline phosphatase (ALP) activity; (ii) mitogenic (MTT) assay; (iii) biomineralization activity; (iv) gene expressions using reverse transcription-polymerase chain reaction (RT-PCR). The effect of EMD with or without heat treatment was examined for ALP activity on the cell differentiation. RESULTS: The effect of Emd-Gel on ALP activity was greater than that of EMD. It was confirmed from the effect of EMD with heat treatment at 60 degrees C, 80 degrees C and 100 degrees C on the ALP activity. The effect of Emd-Gel on the biomineralization activity was also greater than that of the EMD. The Emd-Gel has a stronger effect for the expression of osteoblast-like phenotype than the EMD. CONCLUSION: The results indicate that the Emd-Gel has greater bioactivities than the EMD in vitro.

Alginates↗

PubMed2003

FGF-4 and titanium implants: a pilot study in rabbit bone.

AIM: To evaluate the effect of a local single injection of amino-terminally truncated recombinant human fibroblast growth factor-4 (rhFGF-4s) on titanium implant incorporation in a rabbit bone. MATERIAL AND METHODS: Thirty-six threaded titanium implants were inserted into the femur and tibia of six rabbits. Three weeks prior to implant insertion 10 microg of FGF-4 in an altelocollagen carrier or the carrier alone was injected into the intended implant sites. Biomechanical evaluation by (i) resonance frequency analysis and (ii) peak removal torque measurements was performed after 6 weeks. The implants with surrounding tissue were processed to undecalcified ground sections followed by light-microscopic quantifications of the bone in threaded area and the apical parts of the implants. RESULTS: A general trend, however not statistically significant, with higher mean values obtained in the above-mentioned tests was found. The FGF-4-treated implants revealed a significantly increased bone area in the apical part of the implants compared to the corresponding control implants. CONCLUSION: A local single injection of rhFGF-4 may stimulate bone formation around titanium implants in bone. However, further research is required to confirm these results, understand the mechanisms of FGF, and establish optimal dose and delivery methods.

Animals↗

PubMed2003

Characterization of porcine dentin sialoprotein (DSP) and dentin sialophosphoprotein (DSPP) cDNA clones.

Dentin sialophosphoprotein (DSPP) is a chimeric glycoprotein with dentin sialoprotein (DSP) on its N-terminus and dentin phosphoprotein (DPP) on its C-terminus. We have constructed and screened a unidirectional cDNA library derived from the pulp organ of developing pig teeth, and isolated cDNA clones encoding DSP-only, as well as two DSPP clones with alternative sequences in their 3' coding regions. The DSP-only transcript has an open reading frame of 386 codons, and is generated through the use of a polyadenylation signal within intron 4, immediately following the DSP coding region. the use of this polyadenylation signal deletes the DPP coding region and places a TGA translation termination signal as the fourth codon following the exon 4-encoded segment. The DSPP cDNAs contain open reading frames of 593 and 600 codons. Northern blots hybridized to radiolabeled DSP probes showed bands at 1.4, 2.5, 4.4, and 4.8 kb. Cloning and characterization of reverse transcriptase polymerase chain reaction products confirmed the existence of mRNA encoding pDSP386, pDSPP593, and pDSPP600in vivo, but also suggested that DNA sequence redundancies in the DSPP coding region make it prone to cloning artifacts.

Amino Acid Sequence↗

PubMed2002

Porcine N-acetylgalactosamine 6-sulfatase (GALNS) cDNA sequence and expression in developing teeth.

Mucopolysaccharidosis type IVA (Morquio A syndrome, MPS IVA) is a rare, autosomal recessive disorder with a prevalence of 1 in 170,000 live births. It is caused by a deficiency of N-acetylgalactosamine 6-sulfatase (GALNS), a lysosomal hydrolase encoded by a gene on human chromosome 16q24.3. Mucopolysaccharidosis type IVA is the only known MPS that is associated with structural defects in dental enamel. GALNS cleaves the sulfate group from N-acetylgalactosamine 6-sulfate and galactose 6-sulfate, which are specifically found in keratan sulfate and chondroitin 6-sulfate. A pathologic absence of GALNS activity results in the accumulation of these glycosaminoaglycans in the urine and in the lysosomes of tissues that turn them over. There is currently no animal model for MPS IVA. To learn more about how a GALNS deficit could lead to enamel defects, we have cloned and characterized a full-length pig GALNS cDNA. GALNS mRNA was localized in developing teeth by in situ hybridization, Northern blot, and reverse-transcription polymerase chain reaction analyses, while GALNS substrates were localized using immunohistochemistry. We report that secretory ameloblasts were positive for GALNS mRNA, as well as for keratan sulfate and chondroitin 6-sulfate. We conclude that enamel defects associated with the loss of GALNS activity in persons with MPS IVA are likely to result from the pathological accumulation of keratan sulfate and chondroitin 6-sulfate in the lysosomes of secretory stage ameloblasts.

Amino Acid Sequence↗

PubMed2002

The effect of swimming on cartilage formation.

Swimming is a non-weight-bearing exercise. Therefore it has the advantage of maintaining skeletal integrity in aged persons with weakened skeletal structures. Unlike other weight-bearing aerobic exercises, however, it does not appear to exert sufficient stimulus on bone-remodeling activities because the local load-bearing on bone tissues is mild. The purpose of this study was to investigate the effect of swimming on bone remodeling, especially with the use of implanted pellets containing bone morphogenetic protein (BMP) and demineralized bone matrix during the initial stages of the differentiation of mesenchymal cells to cartilage cells. Six-week-old female rats were divided into the swimming group and a control, nonswimming group. Test animals were forced to swim in a water bath for 30 min daily for 2 wk. After the swimming protocol, pellets were implanted and harvested. Messenger RNA isolated from pellets was quantified by means of a reverse transcription-polymerase chain reaction. The expression of RNAs for bone sialoprotein and BMP-6 in pellets from the swimming group was apparently enhanced at 7 d after implantation. These results suggested that systemic hormonal and/or metabolic changes that promote cartilage formation might have occurred after swimming because the effect was observed after the swimming protocol had ended and the pellets were implanted at a non-weight-bearing site.

Animals↗