HSP70 mRNA expression by cells of the epithelial rest of Malassez due to mechanical forces in vitro.

BACKGROUND: The purpose of the present study was to examine the in vitro responses of ERM cells under the combination of centrifugal and compression forces, in terms of their expression of HSP70 mRNA. METHODS: The ERM cells were positive for CK19 indicating that they were derived from the odontogenic epithelium. Cultured ERM cells were applied centrifugal force and compressing force at one to three times as mechanical forces. After addition of forces, cells were observed using scanning electron microscope (SEM) and were measured expression of HSP70 mRNA by RT-PCR. RESULTS: SEM observations showed the cells were flattened immediately after the application of mechanical force, but nuclear protrusions recovered the same as the control 3 h later. A significantly higher expression of HSP70 mRNA was observed in ERM cells under mechanical force compared with the control, but it gradually decreased with time. No accumulation of HSP70 mRNA expression occurred with intermittent force. However, the expression of HSP70 mRNA with intermittent force repeated 3 times was significantly higher compared with intermittent force applied only once or twice. CONCLUSIONS: These findings suggest that ERM cells express HSP70 mRNA in response to mechanical force, and that intermittent force maintains the level of HSP70 mRNA expression.

Effect of EDTA-treated dentin on the differentiation of mouse iPS cells into osteogenic/odontogenic lineages in vitro and in vivo.

To investigate the effect of EDTA-treated dentin on the differentiation of mouse induced pluripotent stem (iPS) cells. Dentin discs were prepared from bovine incisors and treated with 17% EDTA. Embryoid bodies (EBs) formed from mouse iPS cells were seeded on the dentin discs for the experiment. The roughness of the EDTA-treated dentin surface, Sa and Sdr, was higher and collagen fibrillike structures were observed by the scanning electron microscopy (SEM) in vitro. In RT-PCR, the mRNA levels of the osteoblast markers Bsp and Ocn were significantly higher in the experimental group. Expression of the DMP1, DSP, and BSP proteins were more notable in the experimental group by immunofluorescence (ICF) study. In vivo study, cartilage and bone-like tissue were observed adjacent to the EDTA-treated dentin. The study demonstrates that the dentin treated with 17% EDTA induces mouse iPS cells to differentiate into the osteo/odontogenesis.

Topography enhances Runx2 expression in outgrowing cells from iPS cell-derived embryoid bodies.

The effect of differing polystyrene substrate topographies on the osteogenic potential of the outgrowing cells (OGCs) formed from mouse-induced pluripotent stem (iPS) cells (miPSCs)-derived embryoid bodies (EBs) was investigated. Polystyrene substrates were sandblasted with 25, 50, and 150 µm aluminum oxide particles to obtain topographies with average Sa values of 0.6, 1.1, and 1.8 µm, respectively. 3D-SEM was used to evaluate substrate's topographies. Examination was done by scanning electron microscopy (SEM), by immunocytofluorescence (ICF) analysis for vinculin, Runx2 and collagen type I, and by quantitative RT-PCR (qRT-PCR) analysis for Runx2 and collagen type I. SEM and ICF analyses revealed that surface roughness caused cells elongation (2, 6, 8, 10 times for the NT, 0.6 µm, 1.1 µm, and 1.8 µm, respectively). Vinculin staining demonstrated how the Sa value affected cellular attachment to the substrate. FA points were randomly distributed on flat surfaces, but rough surfaces resulted in more concentrated FA points on the podia of the cells (11.7, 25.2, 26.7, 16.6 vinculin spots per 20 µm2 for the NT, 0.6 µm, 1.1 µm, and 1.8 µm, respectively). qRT-PCR revealed that Runx2 expression was highest on day 16 on surfaces with Sa of 0.6 µm and 1.1 µm. Collagen type I expression increased from day 0 to day 16, no significance was found among the groups. In conclusion, surface topography affects cell shape and expression of early osteogenic potential in OGC, particularly surfaces with Sa values of 0.6 µm and 1.1 µm which showed the highest concentration of FA points on podia. These findings could be utilized in the development of inner surface topographies of scaffolds used with iPSCs. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 2288-2296, 2019.

Effects of CO2 Lasers on Dental Pulp Biology in Rats.

OBJECTIVE: The purpose of this study was to investigate the effects of CO2 lasers on the proliferation and differentiation of dental pulp cells, and their latent self-recovery in connection with their stemness using reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry. MATERIALS AND METHODS: The first molars from male Sprague-Dawley rats, each weighing ∼150-200 g, were used for this study. The upper first molars were irradiated with a 10,600 nm wavelength CO2 laser under identical parameters (2 W CO2 laser, energy 4J, energy density 203.84 J/cm(2) for 8.8 sec) through the dentin of the occlusal surface. The molars were extracted immediately, or at 1, 3 or 5 days after the laser irradiation. RT-PCR analysis using primers specific for heat shock protein 70 (Hsp70), adenosine triphosphate (ATP)-binding cassette transporter G2 (ABCG2), dentin sialophosphoprotein (DSPP), and dentin matrix protein 1 (DMP1), and immunohistochemistry using antibodies specific for proliferating cell nuclear antigen (PCNA), ABCG2, CD34, and CD44 were performed. RESULTS: RT-PCR analysis revealed that Hsp70 mRNA expression in the immediate group and ABCG2 mRNA expression at day 1 were the highest. DSPP and DMP1 mRNA expression in the laser-irradiated groups increased gradually, reaching its peak on the 5th day of the experiment, although no significant difference found among groups with regard to DMP1 expression. Immunohistochemically, PCNA-positive cells were observed at all times after the laser irradiation; however, they were most evident on day 3. CD44-positive cells were observed strongly on day 1 and day 3, while ABCG2-positive cells were the most evident on day 3. CONCLUSIONS: These results demonstrate that CO2 laser irradiation induces degeneration in the pulp tissue, which is then repaired by newly formed odontoblast-like cells.