脂筏在工频磁场诱导受体聚簇效应中的作用及机制初探

目的:研究50 Hz工频磁场诱导细胞膜表面表皮生长因子受体(EGFR)聚簇与脂筏(lipid rafts)结构以及酸性鞘磷脂酶(acid sphingomyelinase,ASM)之间的关系,探索工频磁场诱导细胞膜受体聚簇及其可能的相关机制。方法:将人羊膜上皮细胞(FL)暴露于50 Hz、0.4 mT工频磁场15 min,设立假辐照组和EGF阳性对照组,同时各组增加制霉菌素(nystatin)预处理组,细胞经处理后采用间接免疫荧光法分别用相应抗体标记EGFR和ASM,用FITC标记的霍乱毒素B亚基标记脂筏结构。制片后通过激光共聚焦显微镜进行观察分析。结果:阳性对照组和50 Hz磁场处理组均能观察到EGFR的聚簇现象;制霉菌素预处理1 h后,细胞膜上脂筏结构被破坏,阳性对照组和磁场处理组中受体聚簇现象基本消失。在ASM转位分析中,阳性对照组和磁场处理组Cy3标记的ASM和FITC标记的脂筏结构产生共定位现象,并且在胞膜表面形成特定的富集区域,而在假辐照组中基本处于散在状态。结论:工频磁场诱导FL细胞膜表面EGFR发生聚簇与脂筏结构密切相关,ASM有可能参与了工频磁场诱导的受体聚簇及/或信号转导过程。

Preliminary study on role of lipid rafts in receptor clustering induced by 50 Hz magnetic fields and its mechanism

Objective: To investigate the relationship among a 50 Hz magnetic field(MF)-induced epidermal growth factor receptor(EGFR) clustering,lipid rafts and acid sphingomyelinase(ASM),and to explore its possible mechanism.Methods: Human amnion FL cells were exposed to 50 Hz,0.4 mT MF for 15 min.EGF treatment was used as positive control.Nystatin was employed to study lipid rafts since it could disrupt lipid rafts structure.The EGF receptors,ASM and lipid rafts were labeled with polyclonal anti-EGFR antibody,anti-ASM antibody and FITC-Cholera toxin B,respectively.The images were observed by laser confocal scanning microscope.Results: Both EGF treatment and 50 Hz MF exposure could induce EGFR clustering;however,nystatin pretreatment disrupted this effect.MF exposure turned ASM(labeled with Cy3) from a diffused state in the sham exposure group to a concentrated state on the cell membrane,which co-localized with lipid rafts(labeled with FITC).Conclusion: The results suggest that the EGFR clustering induced by 50 Hz MF depends on intact lipid rafts on cellular membrane,and the ASM might participate in the process of EGFR clustering.