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目的探讨50 Hz 工频磁场对人羊膜成纤维细胞微丝装配的影响,并分析磁场对肌动蛋白和表皮生长因子受体蛋白表达的影响。方法将人羊膜成纤维细胞分别用0.1、0.2、0.3、0.4、0.5 mT,50 Hz 工频磁场辐照30 min,以异硫氰四甲基罗丹明-鬼笔环肽标记细胞微丝并在显微镜下观察分析微丝形态的变化。用荧光光谱及紫外光谱检测法测定细胞内微丝总含量,用激光共聚焦显微镜逐层扫描的方法观测细胞微丝骨架的高度,用扫描电镜观测细胞外部形态等方法进一步分析磁场对细胞微丝骨架及细胞形态的影响。用 Western blotting 检测去垢剂不可溶的蛋白的表达量以分析磁场作用的可能机制。结果人羊膜成纤维细胞经不同强度工频磁场辐照30 min 后,只有0.2 mT 工频磁场辐照显著导致细胞中平行排列的微丝应力纤维减少,诱导细胞周边出现致密微丝外周带并伸出丝状伪足,停止辐照2 h 后微丝形态恢复,伪足消失。光谱检测发现胞内微丝总含量无显著变化。激光共聚焦显微镜逐层扫描观测到0.2 mT 磁场辐照使细胞微丝骨架平均高度由(12.37±1.28)μm降低到(9.97±0.38)μm(t=6.96,P>0.05)。扫描电镜图像显示磁场辐照使细胞更为扁平,有足状伪足出现。Western blotting 检测到磁场辐照后细胞骨架中去垢剂不可溶部分的肌动蛋白和表皮生长因子受体含量与对照组相比,分别升高(16.8±2.3)%(t=16.68,P<0.05)和(31.2±4.1)%(t=17.10,P<0.05)。结论 0.2 mT 工频磁场短时辐照影响了人羊膜成纤维细胞微丝骨架的形态,此效应可随磁场撤销而解除,且可能与磁场诱导了细胞膜上表皮生长因子受体聚簇有关。
Objective To investigate the effect of magnetic field of 50 Hz on assembly of human amnion fibroblasts microfilaments and analyze the effect of magnetic field on the expression of actin and epidermal growth factor receptor protein. Methods Human amnion fibroblasts were irradiated with 0.1,0.2,0.3,0.4,0.5 mT and 50 Hz frequency magnetic field for 30 min respectively, and labeled with isothiocyanate tetramethylrhodamine-phalloidin Microscopic observation and analysis of changes in the morphology of the filaments. The total content of microfilaments in the cells was determined by fluorescence spectroscopy and UV spectrophotometry. The height of the microfilament skeleton was observed by confocal laser scanning microscope. The morphology of the microfilaments was observed by scanning electron microscopy. Skeleton and cell morphology. Western blotting was used to detect the amount of insoluble protein in the detergent to analyze the possible mechanism of magnetic field. Results After human amniotic fibroblasts were exposed to a power frequency magnetic field of different intensities for 30 min, only the 0.2 mT power frequency magnetic field irradiation significantly reduced the parallel arrangement of microfilament stress fibers in the cells, inducing the appearance of dense microfilament peripheral zone and extending A filopodia, stop irradiation 2 h after the recovery of the microfilament, pseudopods disappear. Spectrum detection found no significant changes in the total content of intracellular filaments. Laser scanning confocal microscopy showed that the average microfilament skeletal height decreased from (12.37 ± 1.28) μm to (9.97 ± 0.38) μm (t = 6.96, P> 0.05) after 0.2 mT magnetic field irradiation. Scanning electron microscopy images showed that the magnetic field irradiation made the cells flatter with foot-like pseudopods. Compared with the control group, the content of actin and epidermal growth factor receptor in the insoluble part of the cytoskeleton in the cytoskeleton increased by (16.8 ± 2.3)% (t = 16.68, P < 0.05) and (31.2 ± 4.1)% (t = 17.10, P <0.05). Conclusion Short-time exposure of 0.2 mT power frequency magnetic field affects the morphology of human amniotic fibroblasts microfilament, which can be removed by the magnetic field withdrawal, and may be related to the magnetic field induced epidermal growth factor receptor clustering on the cell membrane.