Nitric oxide mediates the effects of pulsed electromagnetic field stimulation on the osteoblast proliferation and differentiation

The purpose of this research was to investigate whether the effects of pulsed electromagnetic field (PEMF) stimulation on the osteoblast proliferation and differentiation are mediated by the increase in the nitric oxide (NO, nitrogen monoxide) synthesis. The osteoblasts (MC3T3-E1 cell line) were cultured in the absence (-NMMA group) or in the presence (+NMMA group) of the NO synthase inhibitor L-NMMA. First, osteoblasts were subjected to PEMF stimulation (15 Hz and 0.6 mT) up to 15 days. The DNA content and the NO concentration in the conditioned medium were determined on the 3rd, 7th, and 15th days of culture. Following, osteoblasts were stimulated in the proliferation (P-NMMA and P+NMMA groups) or in the differentiation (D-NMMA and D+NMMA groups) stages of maturation, and the alkaline phosphatase (AlPase) activity was determined on the 15th day of culture for all groups. PEMF stimulation increased significantly the nitrite concentration in the -NMMA group on the 3rd, 7th, and 15th days of culture. However, this effect was partially blocked in the +NMMA group. The DNA content in the -NMMA group, but not in the +NMMA group, increased significantly on the 3rd and 7th days of culture. The AlPase activity in the P-NMMA and D-NMMA groups, but not in the P+NMMA and D+NMMA groups, also increased significantly. In conclusion, the PEMF stimulatory effects on the osteoblasts proliferation and differentiation were mediated by the increase in the NO synthesis.