A low concentration of boron (B) accelerates the proliferation and differentiation of mammalian osteoblasts. The aim of this study was to investigate the effects of 0.1 mM of B on the membrane function of osteoblastic cells in vitro. Genes involved in cell activity were investigated using gene expression microarray analyses. The Ca influx and efflux were evaluated to demonstrate the activation of L-type Ca channel for the Ca influx, and that of Na/K-ATPase for the Ca efflux. A real-time PCR analysis revealed that the messenger RNA (mRNA) expression of four mineralization-related genes was clearly increased after 3 days of culture with a B-supplemented culture medium. Using microarray analyses, five genes involved in cell proliferation and differentiation were upregulated compared to the control group. Regarding the Ca influx, in the nifedipine-pretreated group, the relative fluorescence intensity for 1 min after adding B solution did not increase compared with that for 1 min before addition. In the control group, the relative fluorescence intensity was significantly increased compared with the experimental group (P < 0.05). Regarding the Ca efflux, in the experimental group cultured in 0.1 mM of B-supplemented medium, the relative fluorescence intensity for 10 min after ouabain treatment revealed a significantly lower slope value compared with the control group (P < 0.01). This is the first study to demonstrate the acceleration of Ca flux by B supplementation in osteoblastic cells. Cell membrane stability is related to the mechanism by which a very low concentration of B promotes the proliferation and differentiation of mammalian osteoblastic cells in vitro.
Tilapia type I atelocollagen (TAC) is a strong candidate for clinical application as its biological scaffold due to a high degeneration temperature and biologically safe properties. The aim of this study was to confirm the biological effects of TAC in vitro on osteoblastic cells, simulating its clinical application. The proliferation and differentiation of typical preosteoblasts, MC3T3-E1 cells, were investigated using a microarray analysis, staining assay for mineralization, and real-time PCR analysis of the expression of mineralization-related genes. The mRNA expression of 10 genes involved in proliferation and differentiation increased after 3-day culture on an TAC gel, with an average balanced score ratio exceeding 1.5 compared to the control. After two weeks of culture, all three experimental groups showed stronger alkaline phosphatase staining than after one week. The genes expression of alkaline phosphatase, osteocalcin, and bone sialoprotein increased under the experimental conditions. The gene expression of osteopontin did not increase, and no statistical differences were noted among the three experimental groups. The present and previous findings suggest that TAC is not only a suitable alternative to collagen products originating from mammals but also a novel biomaterial with cell differentiation ability for regenerative medicine.
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