反义ClC-3寡核苷酸促进thapsigargin诱导的PC12细胞凋亡

目的　研究反义ClC 3寡核苷酸对thapsigargin诱导的细胞凋亡的影响。方法　蛋白免疫印迹法检测ClC 3蛋白的表达 ;MTT法检测反义ClC 3寡核苷酸对TG诱导的细胞生长的影响 ;形态学方法、流式细胞仪和DNA琼脂糖凝胶电泳观察和分析PC12细胞在转染ClC 3反义寡核苷酸后 ,TG诱导的细胞形态、DNA含量的变化和DNA断裂的情况。结果　与对照组相比 ,反义ClC 3寡核苷酸呈浓度和时间依赖性地抑制ClC 3蛋白的表达 ;使TG诱导的PC12细胞存活率降低 ,凋亡程度加强 ,差异有显著性 ,而正义及随义ClC 3寡核苷酸对此没有影响。结论　反义ClC 3寡核苷酸对TG诱导的PC12细胞凋亡有促进作用。

The effect of ClC-3 antisense oligonucleotide on apoptosis induced by thapsigargin in PC12 cells

AIM To investigate the effect of ClC-3 antisense oligonucleotide on apoptosis induced by thapsigargin in PC12 cells. METHODS Western-blot was performed to detect the protein expression of ClC-3 in PC12 cells. MTT assay was used to measure the effect of ClC-3 antisense oligonucleotide on growth inhibition induced by thapsigargin. The effect of ClC-3 antisense oligonucleotide on apoptosis was studied with the fluorescent microscopy, DNA agarose gel electrophoresis, flow cytometry analysis. RESULTS Compared with control group, transient transfection of PC12 cells with antisense oligonucleotide specific to ClC-3 caused an inhibitory effect on expression of ClC-3 protein in a time-and concentration-dependent manner,whereas the thapsigargin-induced reductions of viability of PC12 cells and apoptosis were markedly enhanced (P<0.05 vs control group). Sense and missense oligonucleotide had no effects. CONCLUSION The results suggest that transient transfection of PC12 cells with antisense oligonucleotide specific to ClC-3 causes cells is more susceptible to apoptosis induced by thapsigargin. Thus, ClC-3 chloride channel may be involved in the regulation of apoptosis.