米贝地尔对罗哌卡因诱导SH-SY5Y细胞凋亡的抑制作用研究

Inhibitory effect of mibefradil on ropivacaine-induced apoptosis in SH-SY5Y cell

目的:观察T型钙通道拮抗剂米贝地尔对罗哌卡因致SH-SY5Y细胞凋亡的影响,探讨罗哌卡因诱发神经细胞毒性是否与T型钙通道相关。方法:将SH-SY5Y细胞随机分为4组:SH-SY5Y细胞正常培养组(A组);SH-SY5Y细胞5μmol/L米贝地尔培养组(B组);SH-SY5Y细胞3 mmol/L罗哌卡因培养组(C组);SH-SY5Y细胞5μmol/L米贝地尔+3 mmol/L罗哌卡因培养组(D组)。各组细胞分别在有或无3 mmol/L罗哌卡因处理开始时(T0)、处理后1 h(T1)、6 h(T2)、12 h(T3)、24 h(T4),MTT法检测细胞活力、流式细胞术和Hoechst33258染色法检测细胞凋亡率。结果:与A组相比,C组、D组在T1、T2、T3、T4时点细胞活力明显降低(P<0.05);但C组降低幅度更明显,C组与D组比较,两组在T1、T2、T3、T4时点差异有统计学意义(P<0.05)。C组随时间的递增其细胞凋亡率逐渐增加,在T4时凋亡率达到最高;D组在T1、T2、T3、T4时点的细胞凋亡率明显比C组降低,两组差异有统计学意义(P<0.05)。结论:罗哌卡因对SH-SY5Y细胞有毒性作用,米贝地尔可减轻罗哌卡因诱发的神经细胞损伤,提示罗哌卡因诱发神经毒性可能与T型钙通道有关。

Objective To observe the effects of T type calcium channel blocker mibefradil on ropivacaine-induced SH-SY5Y cell apoptosis,and investigate whether ropivacaine-induced neurotoxicity is related to T type calcium channel.Methods SH-SY5Y cells were randomly divided into four groups： SH-SY5Y cells were cultured in normal conditions（group A）,SH-SY5Y cells were cultured with 5 mol/L mibefradil（group B）,SH-SY5Y cells were cultured with 3 mmol/L ropivacaine（group C）;SH-SY5Y cells were cultured with 5 mol/L mibefradil and 3 mmol/L ropivacaine（group D）.Following SH-SY5Y cells treated or untreated with 3 mmol/L ropivacaine for 0 h（T0）,1 h（T1）,6 h（T2）,12 h（T3） and 24 h（T4）,cell viability was quantitatively determined by the MTT colorimetry assay.Flow cytometry and Hoechst33258 staining were used to analyse the apoptosis rates.Results Compared with Group A,cell viability decreased significantly in group C and D at T1、T2、T3、T4（P 0.05）;Compared with group D,a greater decrease of cell viability was observed in group C.The difference between group C and D at T1、T2、T3、T4 was significant（P 0.05）.With the increasement of time,the apoptosis rates of group C increased gradually,and was highest at T4;the apoptosis rates of group D at T1、T2、T3、T4 were significantly lower than those of group C（P 0.05）.Conclusion 5 mol/L mibefradil can reduce ropivacaine-induced apoptosis SH-SY5Y cells,which indicates that ropivacaine-induced neurotoxicity is probably related to T type calcium channel.