低频脉冲电场对胰岛素介导的细胞增殖影响及其作用机理研究

THE EFFECTS OF EXTREMELY LOW FREQUENCY PULSED ELECTRICAL FIELD ON PROLIFERATION OF HUMAN LIVER CELL MEDIATED BY INSULIN*

本文以胞外信号分子胰岛素为研究对象,从细胞信号系统与电磁场相互耦合角度,通过MTT比色法和间接免疫荧光技术研究脉冲电场(f=50Hz,τ=20μS,Epp=1V/m)对人肝细胞(L-02细胞株)的增殖能力以及胰岛素与细胞表面受体的专一结合特性的影响。MTT比色分析结果发现在脉冲电场对细胞增殖的直接作用中,处理5、10、20分钟对细胞增殖的抑制百分率分别为10.13％、18.10％和11.85％;脉冲电场对细胞增殖的间接作用中,胰岛素经脉冲电场处理20、40分钟,对细胞增殖的抑制百分率分别为10.31％和14.12％;流式细胞术检测结果表明细胞悬液和胰岛素共同经脉冲电场处理20分钟组其平均荧光强度降低22.74％,仅胰岛素受电场处理组和仅细胞受电场处理组的平均荧光强度分别下降12.96％和10.51％。本实验结果显示 脉冲电场对人肝细胞的作用体现在两个方面:一是电场直接作用于细胞膜上的胰岛素受体及其他膜成分;二是脉冲电场还可通过改变培养基中的胰岛素分子的构像,影响胰岛素与受体的结合能力,进而影响细胞的增殖等活动。

The effects of extremely low frequency pulsed electrical field (f = 50Hz, t = 20piS, Epp = 1 V/m) on liver cells mediated by insulin were examined by two independent methods: 1) measurement of cell proliferation using MTT colorimetric assay; 2) monitoring of insulin-insulin receptor binding properties by flow cytometry. The MTT test showed that an inhibition in cell proliferation to 10,13% ,18. 10% and 11.85% when both cell and insulin were exposed to electrical field for 5 min, 10 min and 20min in direct effects, respectively; In indirect effects, cell proliferation inhibited by 10.31% and 14.12% when only insulin was exposed to electrical field for 20 min and 40min, respectively; Flow Cytometry analysis of FITC staining demonstrated that the binding capacity of live cells decreased by 22.74% when both cells and insulin were exposed to electrical field for 20min, the binding capacity decreased by 12.96% when only insulin was exposed to electrical field for 20min and the binding capacity decreased by 10.51 % when only cells were exposed to electrical fields for 20min. These results indicated that the effects of extremely low frequency pulsed electrical field on liver cells have two aspects; 1). The extremely low frequency pulsed electrical field could affect insulin receptor and other membrane components; 2). The extremely low frequency pulsed electrical field could change the conformation of insulin which added to culture medium, and influence the binding capacity of insulin-insulin receptor, so as to influence cell proliferation and other metabolic activities.