[Genic and non-genic regulation of low frequence pulsed electromagnetic fields on osteoblasts differentiation].

Objective: To study the genic and non-genic regulation of 50 Hz 0.6 mT pulsed electromagnenic fields (PEMF) on rat calvarial osteoblasts (ROB) differentiation. Methods: ROBs were achieved by enzyme digestion, and treated with 50 Hz 0.6 mT PEMFs for 1.5 hours after subculture. The alkaline phosphatase (ALP) activity, mRNA transcription of ALP, Runx2 and OSX and protein expression of Runx2 and OSX were detected at 0, 3, 6, 9 and 12 hours after PEMF treatment. Results: The ALP activity at 3 hours after treatment was significantly higher than that in the control(P<0.01), while the mRNA transcription of ALP began to increase at 6 hours after treatment. The mRNA transcription of Runx2 increased immediately after treatment and regressed at 6 hours, then increased again. The protein expression of it corresponded but with a little lag. The mRNA transcription of OSX also raised instantly after treatment, then returned to the level of control at 6 hours, and lower than control at 12 hours significantly. The protein expression of it also corresponded but with a bit delay. Conclusions: There are genic regulation for the protein expression of Runx2 and OSX, and non-genic regulation for the ALP activity on the process of 50 Hz 0.6 mT PEMFs prompts ROBs differentiation.

Low-intensity laser irradiation stimulates mineralization via increased BMPs in MC3T3-E1 cells.

BACKGROUND: Previously, we reported that low-intensity laser irradiation accelerated bone formation, and that this mechanism deeply involved insulin-like growth factor I expression. However, as bone formation is supported by many local factors, the mechanism involved in laser irradiation remains incompletely understood. Therefore, the purpose of this study was to determine the effects of laser irradiation on the osteogenic response in vitro. METHODS: Mouse osteoblast-like cells, MC3T3-E1, were cultured and were irradiated for 5-20 minutes (0.96-3.82 J/cm(2)) at the subconfluent stage using a low-intensity Ga-Al-As diode laser apparatus. After laser irradiation, expression of bone morphogenetic proteins (BMPs), transcription factors (Runx2, Osterix, Dlx5, Msx2), and phosphorylation of Smad1 were determined, and calcium content of cell cultures was also determined. RESULTS: Irradiation at 1.91 J/cm(2) significantly increased the expression of BMPs and Runx2, Osterix, Dlx5, Msx2, and the phosphorylation of Smad1. Noggin, a BMP receptor blocker, inhibited the laser-induced Runx2 expression and phosphorylation of Smad1. Moreover, laser irradiation significantly increased the calcium content of cell cultures, and noggin inhibited this increase. CONCLUSION: These results suggest that low-intensity laser irradiation stimulates in vitro mineralization via increased expression of BMPs and transcription factors associated with osteoblast differentiation.

BMP-7 and CBFA1 in allograft bone in vivo bone formation and the influence of gamma-irradiation.

An initial study showed that morselized human bone grafts were osteoconductive and osteoinductive when implanted in nude rat tibial window defects, and 25 kGy of gamma-irradiation significantly reduced those properties. The mechanism of the osteoinductivity and the influence of gamma-irradiation required further investigation. In this study we assessed the paraffin sections of seven morselized human bone grafts implanted into rat tibial defects for 3 weeks after being treated with 0, 15, or 25 kGy gamma-radiation respectively. Osteoclast-like cell counting and protein expressions of bone morphogenetic protein-7 (BMP-7), core binding factor alpha1 (CBFA1), and proliferating cell nuclear antigen (PCNA) were investigated and the positive signals were quantitatively analyzed. More new bone formation was observed in the 0 and 15 kGy groups compared with 25 kGy groups. The newly formed bones were found mainly from the intact cortex into the defects bridged by the implanted grafts. A dense staining of BMP-7 and CBFA1 was noted in the osteoblast-like cells in those areas. The BMP-7 and CBFA1 staining was also seen in the cells surrounding the implanted grafts in the centre areas of the defects in distance from the intact cortex. Quantitative analysis of immunohistochemical staining of the centre areas of the defects showed that gamma-irradiation (15 and 25 kGy) significantly reduced the expression of CBFA1 and BMP-7. In conclusion, morselized human bone grafts may contain some factors, which induced osteoblast lineage differentiation and bone formation and gamma-irradiation damages those bone inducing factors.