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目的 研究 5 0Hz正弦磁场对细胞连接蛋白Cx4 3移位的影响 ,以探索极低频磁场 (ELFMF)抑制细胞间隙连接通讯 (GJIC)的机制。方法 用 5 0Hz、0 .8mT的正弦磁场和 (或 )十四酰基咐哌醇酯 (TPA ,5ng/ml)对培养的中国仓鼠肺成纤维细胞 (CHL)处理 2 4h(其中TPA处理 1h)。采用间接免疫荧光细胞化学法和激光共聚焦显微镜进行连接蛋白Cx4 3定位的测定 ;采用Westernblot方法检测胞浆和核内Cx4 3蛋白的含量。结果 正常组细胞间连接处有明亮的点状标记 ,连接成线 ;TPA处理组、0 .8mT正弦磁场单独作用组及 0 .8mT正弦磁场联合TPA处理组细胞的连接处标记斑点均较正常组减少 ,大量Cx4 3蛋白标记斑点出现在胞浆内 ,且在核附近聚集。各组核内Cx4 3蛋白条带很淡 ,而胞浆内Cx4 3蛋白含量ELF组 (2 .0 3± 0 .89)和TPA组 (2 .4 3± 0 .82 )明显增加 ,与正常组 (1.0 4± 0 .17)相比 ,差异有显著性 (P <0 .0 1)。结论 5 0Hz正弦磁场直接和 (或 )协同TPA抑制体外培养细胞GJIC功能与细胞连接蛋白Cx4 3内移至胞浆有关
Objective To investigate the effect of 50 Hz sinusoidal magnetic field on the translocation of Cx4 3 in order to explore the mechanism of ELFMF inhibiting GJIC. Methods Cultured Chinese hamster lung fibroblasts (CHL) were treated with sinusoidal magnetic field of 50Hz and 0.8mT and / or tetradecanoyl piperidyl ester (TPA, 5ng / ml) for 24 hours (TPA treatment for 1 hour) . Indirect immunofluorescence cytochemistry and confocal laser scanning microscopy were used to determine the location of connexin Cx4 3. The content of Cx4 3 protein in cytoplasm and nucleus was detected by Western blot. Results In the normal group, there was a bright dot-like mark at the junction of the cells. The TPA-treated group, 0.8mT sinusoidal magnetic field alone group and 0.8mT sinusoidal magnetic field combined with TPA-treated group had more marked spots on the junction than the normal group Reduce, a large number of Cx4 3 protein mark spots appear in the cytoplasm, and gathered in the vicinity of the nucleus. The Cx4 3 protein bands in each group were very weak, while the intracellular Cx4 3 protein levels in ELF group (2.03 ± 0.89) and TPA group (2.43 ± 0.82) were significantly increased compared with normal Group (1.0 4 ± 0.17), the difference was significant (P <0.01). CONCLUSIONS: 5 0 Hz sinusoidal magnetic field directly and / or in combination with TPA inhibits the function of GJIC in cultured cells in vitro and correlates with the intracellular localization of Cx43