酸感受离子通道亚基1a敲减后的细胞抗酸性研究

Acid resistance research on acid-sensing ion channel 1a knock-down in cells

目的:通过酸感受离子通道(ASIC)1 a表达抑制的细胞模型,探讨在酸性条件下ASIC1 a对细胞生长增殖的影响。方法:以大鼠神经胶质瘤(C6)为模型细胞,利用RNA i技术建立ASIC1 a表达抑制的细胞模型。通过绘制细胞生长曲线、集落形成试验、MTT试验和乳酸脱氢酶(LDH)释放试验来观察不同pH条件下,ASIC1 a表达抑制细胞和阴性对照细胞生长增殖能力的变化。结果:RNA i技术可明显抑制C6细胞ASIC1 a蛋白的表达。在正常pH条件下,ASIC1 a表达缺失对C6细胞的生长增殖没有显著影响。但在不同酸性条件下,ASIC1 a表达抑制细胞与阴性对照细胞相比对酸的耐受性明显提高,表现为细胞损伤减轻,存活数目明显增多。结论:C6细胞中有ASIC1 a,ASIC1b,ASIC2 a,ASIC2b和ASIC3亚基的表达。抑制ASIC1 a的表达不影响正常pH条件下细胞的生长增殖,但能明显提高细胞对酸的耐受性,提示ASIC1 a可能是潜在的抗酸靶分子。

Objective： To study the possible effects of acid-sensing ion channel 1 a subunit on the growth of rat neuroglioma cells（C6） under the conditions of various pH values. Methods： RNAi technique was used to silence the expression of ASIC1 a subunit mRNA and protein in C6 cells. The growth and reproduction of ASIC1 a knock-down cells were observed by making out the cell growth curve and measuring the rate of plating efficiency. The acidosis-induced lesions of ASIC1a knock-down cells and the control group cells were observed by performing the experiments of methyl thiazolyl tetrazolium （MTT） assays and LDH release assays. Results：The expression of ASIC1a, 1b,2a,2b and 3 subunits was found in C6 cells. ASIC1a knock-down cells built by RNAi technique presented remarkable suppression on the expression of ASIC1a protein. Under the normal pH condition, there was no difference on the growth and reproduction of the ASIC1a knock-down cells and the control group cells. ASIC1a knock-down cells displayed a stronger resistance to acid injury compared with the control cells. Conclusions：Suppression of ASIC1 a expression has no effect on the growth and reproduction of cells under the normal pH condition. The livability of cells is evidently enhanced by inhibiting the expression of ASIC1a under acidic conditions, suggesting that ASIC1a may be a potential anti-acid target.