Objective: To investigate the effect of low-frequency pulsed electromagnetic fields (PEMF) on the maturation and mineralization of rat cranial osteoblasts in vitro and its relation to IGF-1R/NO signaling pathway.
Methods: The rat osteoblasts were isolated and cultured in vitro and randomly divided into blank control group, PEMF group, GSK group (IGF-1R blocker) and PEMF+GSK group. The cells were treated with 50 Hz 0.6 mT PEMF for 1.5 h/d. After 3 d of PEMF treatment, the expressions of protein kinase (AKT), inducible nitric oxide synthase (iNOS) and cGMP-dependent protein kinase (PKG) were detected by Western blotting; on 6 d of PEMF treatment alkaline phosphatase (ALP) activity was determined; on 12 d of PEMF treatment the calcification nodule formation was demonstrated by Alizarin red staining.
Results: NO level was significantly increased in rat osteoblasts treated with 50 Hz 0.6 mT PEMF for 1.5 h/d. Western blot analysis showed that the expressions of AKT, iNOS and PKG protein in PEMF group were higher than those in the control group (all P<0.01); the ALP activity was increased(P<0.05), and the PEMF group had the largest area of Alizarin red staining (P<0.01). The expressions of AKT, iNOS and PKG protein in GSK group were lower than those in the control group; the ALP activity was decreased (P<0.05), and the GSK group had the least area of Alizarin red staining (P<0.01). The expressions of AKT, iNOS, PKG protein, the ALP activity and the area of Alizarin red staining in PEMF+GSK group were between PEMF group and GSK group.
Conclusions: PEMF may enhance the maturation and mineralization of rat cranial osteoblasts in vitro through IGF-1R/NO signaling pathway.
目的: 观察低频脉冲电磁场（PEMF）是否通过胰岛素样生长因子1受体（IGF-1R）/一氧化氮（NO）信号通路促进大鼠颅骨成骨细胞成熟及矿化。
方法: 体外分离培养乳鼠颅骨成骨细胞，传代后随机分成9组，每天用50 Hz 0.6 mT PEMF处理不同的时间，通过检测成骨细胞中NO含量确定PEMF最佳处理时长。将传代后的成骨细胞随机分成空白对照组、PEMF组、GSK（IGF-1R阻断剂）组、PEMF+GSK组，PEMF处理成骨细胞3 d后，采用蛋白质印迹法检测蛋白激酶（AKT）、诱导型一氧化氮合酶（iNOS）和cGMP依赖性蛋白激酶（PKG）蛋白表达水平；PEMF处理第6天时测定碱性磷酸酶（ALP）活性；PEMF处理第12天时采用茜红素染色法观察钙化结节形成情况。
结果: 经1.5 h电磁场处理后的成骨细胞中NO含量显著提高（ P < 0.01），遂后续采用PEMF处理1.5 h。与空白对照组比较，PEMF组AKT、iNOS、PKG蛋白表达量增加（均 P < 0.01），ALP活性升高（ P < 0.05），且茜红素染色面积增大（ P < 0.01）；GSK组AKT、iNOS、PKG蛋白表达量减少，ALP活性降低（ P < 0.05），茜红素染色面积最小（ P < 0.01）；PEMF+GSK组上述实验结果均高于GSK组但低于PEMF组（ P < 0.05或 P < 0.01）。
结论: PEMF可能通过IGF-1R/NO信号通路促进大鼠颅骨成骨细胞成熟与矿化。