DIDS对SIN-1诱导大鼠海马神经元凋亡及PARP-1/AIF表达的影响

Effects of DIDS on cultured hippocampal neuronal apoptosis and the expression of PARP-1/AIF induced by SIN-1 in rats

目的观察氯通道阻断剂4,4'-二异硫氰基芪-2,2'-二磺酸(DIDS)对NO诱导的大鼠离体海马神经元凋亡的保护作用。方法离体培养12 d的SD大鼠海马神经元,随机分为正常对照组、3-吗啡斯德酮亚胺(3-morpholinosyndnomine,SIN-1)处理组、SIN-1+DIDS组。对各组神经元分别在相应的时间点用MTT法测定细胞生存率、Hoechst 33258测定凋亡百分数、免疫化学荧光分析检测凋亡信号蛋白PARP-1/AIF的变化、Western blot分析凋亡蛋白Caspase-3的变化。结果 DIDS呈剂量依赖性地抑制SIN-1诱导的神经元损伤,减少凋亡发生数目,并能抑制损伤所引起的Caspase-3的激活、削弱PARP-1/AIF的表达。结论氯通道可能参与了NO诱导的海马神经元损伤,氯通道阻断剂DIDS的保护作用可能与削弱PARP-1/AIF的表达有关。

Aim To observe the protective effects of DIDS,one kind of chloride channel blockers,on hippocampal neuronal damage induced by NO in culture.Methods The cultures were randomly assigned into control group,3-morpholinosydnonimine（SIN-1 0.5 mmol·L-1 for 18 h） treatment group,SIN-1 ＋4,4′-disothiocyanatostilbene-2,2′disulfonic acid（DIDS,0.1 mmol ·L-1） group.The neuronal viabilities were detected with the methods of MTT and the neuronal apoptosis was assyed by Hoechst 33258 stainning and activated caspase-3 by Western blot,the both expressions of PARP-1 and AIF were also assayed with immunofluorscent staining.Results There was a significant protective effect of DIDS on neuronal damage in a dose-dependent manner.DIDS inhibited the activation of Caspase-3 and the expression of PARP-1/AIF.Conclusions Chloride channel activities may be involved in neuronal injury induced by NO,and its mechanism underlying the protection of chloride channel blocker against neuronal injury may relate to the down regulation of PARP-1/AIF expression.