Mitsuro Tanaka

Distribution of 14C-bisphenol A in pregnant and newborn mice

OBJECTIVES The purpose of the present investigation was to trace the fate of bisphenol A injected into pregnant mice, focusing on its potential accumulation in the fetus and the brain, critical targets of hormonal chemicals, using whole-body autoradiography. METHODS Pregnant mice were injected intraperitoneally with 0.46MBq of (14)C-BPA and then killed at 1h or 1, 3, or 5 days after injection. Sections for autoradiography were prepared in a cryomicrotome and the exposed imaging plate was processed using a fluorescent/radioisotope image analyzer. RESULTS Intraperitoneally injected (14)C-BPA was distributed throughout the body, including the fetus and the brain, within 1h. Radioactivity faded gradually from the whole body by the fifth day, and no accumulation in any specific organ was found. However, although (14)C was detected in the fetuses immediately after injection, the transfer of BPA from mother to newborn was not observed. SIGNIFICANCE The routes of rapid BPA discharge were confirmed, and BPA neither accumulated in the body nor was it transferred to newborn mice. No evidence was observed to suggest the existence of a blood-placenta or blood-brain barrier for BPA. This information should be taken into consideration when assessing the risks of using dental materials that contain BPA.

Differential effects of TGF-β1 and FGF-2 on SDF-1α expression in human periodontal ligament cells derived from deciduous teeth in vitro

Stromal cell-derived factor (SDF)-1α has been reported to play a crucial role in stem cell homing and recruitment to injured sites. However, no information is available about its role in periodontal tissues. The aim of this in vitro study was to investigate the effects of basic fibroblast growth factor (FGF-2) and transforming growth factor (TGF)-β1 on SDF-1α expression in immortalized periodontal ligament (PDL) cells derived from deciduous teeth (SH9 cells). Real-time PCR and western blot analyses showed that SDF-1α mRNA expression in SH9 cells was markedly inhibited by FGF-2 treatment for 48 h. SU5402, which directly interacts with the catalytic domain of the FGF receptor 1 (FGFR1) and suppresses its phosphorylation, inhibited the FGF-2-related decrease in SDF-1α expression. These results suggest that FGF-2 signaling via the FGFR1 pathway inhibits SDF-1α expression. Conversely, SDF-1α expression in SH9 cells was increased by TGF-β1 treatment for 12 h. Western blot analysis showed that this treatment induced Smad2/3 phosphorylation. A time-course experiment showed that SDF-1α expression levels reached a maximum 12 h after the TGF-β1 treatment and returned to basal levels by 48 h. Real-time PCR analysis showed that Smad7 mRNA expression peaked by 6 h after TGF-β1 treatment. Since Smad7 siRNA downregulated Smad7 expression by approximately 2.5-fold compared with the negative control siRNA, the induction of SDF-1α expression was prolonged. Furthermore, treatment of SH9 cells with TGF-β1 for 12 h induced transwell migration of UE7T-13 cells, which are mesenchymal stem cells derived from human bone marrow. Therefore, SDF-1α may play an important role in stem and progenitor cell recruitment and homing to injured sites in the periodontal ligament, and regulation of SDF-1α expression may be a useful tool in cell-based therapy for periodontal tissue regeneration.

Fibroblast growth factor 2 inhibits the expression of stromal cell-derived factor 1α in periodontal ligament cells derived from human permanent teeth in vitro

Although cells derived from periodontal ligament (PDL) tissue are reported to have stem cell-like activity and are speculated to play a crucial role for tissue healing and regeneration after injury or orthodontic treatment, mechanisms regulating their recruitment and activation remain unknown. Recently, stromal cell-derived factor 1α (SDF-1α) has been reported to be important for stem cell homing and recruitment to injured sites. The aim of this study was to evaluate whether fibroblast growth factor 2 (FGF-2) affects the expression of SDF-1α in PDL cells derived from human permanent teeth in vitro. Using real-time PCR, the expression of SDF-1α mRNA in PDL cells was inhibited by treatment with 10 ng/ml FGF-2. When PDL cells were treated with SU5402 (an inhibitor of FGF receptor 1) in combination with FGF-2, the FGF-2-reduced expression of SDF-1α was inhibited. In the presence of the JNK inhibitor SP600125, SDF-1α mRNA in PDL cells was not suppressed by the FGF-2 treatment. Western blot analysis also showed that SDF-1α production was suppressed by treatment with FGF-2, but it recovered with treatment by FGF-2 + SU5402. These findings suggest that SDF-1α from PDL cells plays an important role in the regeneration and homeostasis of periodontal tissues via the recruitment of stem cells.

Establishment of Clonal Periodontal Ligament Cell Line Derived from Deciduous Tooth Immortalized by Human Telomerase Reverse Transcriptase (hTERT) Gene Transfer

Recently, many studies have shown that mesenchymal stem cells (MSCs) exist in dental pulp and periodontal ligament (PDL) of human deciduous teeth. MSCs derived from extracted teeth have been considered an appropriate candidate for not only dental tissue regeneration but also for the treatment of general diseases. However, there was no immortal PDL cell line derived from human deciduous teeth for cell-based regeneration study. In this study, we established tree PDL cell lines derived from human deciduous tooth immortalized by the transfection with human telomerase reverse transcriptase (hTERT) gene. The transfected cells were named SH, Single cell from Human periodontal ligament. All SH clones expressed hTERT and scleraxis, periostin, cementum-derived protein 23 and tenomodulin. Though all SH clones expressed osteoblastic marker genes, only the clone, SH 9 cells, differentiated mature osteoblasts. These clones could be useful for the research in regenerative medicine.

Dental pulp cells derived from permanent teeth express higher levels of R-cadherin than do deciduous teeth: implications of the correlation between R-cadherin expression and restriction of multipotency in mesenchymal stem cells.

OBJECTIVES The aim of this study was to characterize the expression status of cadherins in dental pulp-derived mesenchymal progenitor/stem cells from deciduous and permanent teeth, and to determine how cadherins affect the multipotency of the progenitor/stem cells. MATERIALS AND METHODS We evaluated and compared the expression status of cadherins in dental pulp-derived cells from deciduous teeth and in cells from permanent teeth by using an array of primers for amplification of RNA encoding human cell adhesion molecules and a real time PCR system. In order to elucidate how cadherins (which are differentially expressed in deciduous and permanent teeth) affect the multipotency of the dental pulp-derived progenitor/stem cells, the ability of the dental pulp cells to differentiate into adipocytes and osteoblasts was evaluated. RESULTS R-cadherin was found to be vigorously expressed in the dental pulp cells derived from permanent teeth but not in the dental pulp cells derived from deciduous teeth. N-cadherin was found to be expressed essentially equally in both types of cells. The ability of the dental pulp cells of deciduous teeth to differentiate into adipocytes and osteoblasts was found to be much higher than that of cells obtained from permanent teeth. CONCLUSION R-cadherin may be a key molecule for providing control over the multipotency of the dental pulp-derived mesenchymal stem cells.

Dominant prevalence of Porphyromonas gingivalis fimA types I and IV in healthy Japanese children

Background/purpose Porphyromonas gingivalis is a major causative agent of chronic periodontitis, whilst circumstances for acquisition of the bacterium remain to be elucidated. To examine prevalence of the bacterium harboring distinct fimA types in dental plaque of children, we established PCR procedures that are applicable for specimens with limited amounts. By this method, all six fimA types including type I and Ib were directly identified, and prevalence of fimA types and their frequency of guardian-child transmission in Japanese children were assessed. Materials and methods Genomic DNA was purified from dental plaque specimens of 132 periodontally healthy children (2–12 years old, 4.8 ± 0.2 years) and 19 mothers of resultant P. gingivalis-positive child subjects. PCR-based fimA genotyping was performed, and untypeable strains in the first PCR analysis were determined by a nested PCR. Results P. gingivalis was found in 15.2% of the subjects (2–10 years old, 5.1 ± 0.6 years), and the most prevalent types were I and IV (37.0% each), followed by Ib and III (11.1% each), and then II (7.4%). Seven (35.0%) of the 20 P. gingivalis-positive subjects had combined colonization of type I with other fimA types. In most cases, bacterial prevalence and fimA types in the children were distinct from those of their mothers, indicating that its maternal transmission was not significant. Conclusion These results suggest that colonization of non-disease-associated fimA types I and IV P. gingivalis to the oral cavity initiates from early childhood without showing any periodontal inflammation.

Maternal Transmission of Mutans and Other Oral Streptococcal Species

In this study, the distribution of mutans and non-mutans streptococci in the plaque samples from 320 children was investigated, and then the maternal transmission in the 16 mother-and-child pairs was assessed. In the children’s plaque, 14 oral streptococci including mutans streptococci (MS) were identified, although the frequencies of the species were quite different among species. The relatively higher number of species was detected in the MS-positive children than the MS-negative one. In the mother-and-child pairs, the genotypic fingerprinting analysis using pulse-field gel electrophoresis (PFGE) revealed a high frequency in matching genotypes of MS. Furthermore, the distribution as well as the quantitative level of non-mutans streptococci was similar to each other. Thus, the present results strongly suggested that the maternal transmission does occur in not only mutans streptococci but also non-mutans streptococci.

An intruded immature permanent incisor splinted with a mouthguard: A case report

An intruded upper left immature permanent central incisor of a 6-year-and-10-month-old girl was splinted with a mouthguard-type splint because the adjacent teeth could not be used for fixation. She visited our clinic about 1 h after she fell from an iron bar at school and injured her tooth. After repositioning, the incisor was splinted for 4 weeks with a mouthguard fabricated using the pressure molding technique. Although the incisor erupted in the normal position, a radiograph 1 year after the injury showed partial obliteration of the pulp cavity, and total obliteration was observed at 1 year and 10 months. The apical shape of the root was round and the root was shorter than that of the right central incisor. The incisor showed vitality 9 months after the injury using an electric pulp tester and the vitality equaled that of the right incisor at 1 year and 10 months. A mouthguard-type splint is less likely to cause ankylosis compared to rigid fixture and is useful for the fixation of an erupting immature incisor.