Effects of emulsified isoflurane on apoptosis in human neuroblastoma cells : the role of c-Jun N-terminal kinase / 中华麻醉学杂志

ABSTRACT

Objective To investigate the effects of emulsified isoflurane on apoptosis in human neuroblastoma cells and the role of c-Jun N-terminal kinase (JNK) in it.Methods The human neuroblastoma SHSY-5Y cells were seeded in 96-well plates or dishes and then randomly divided into 8 groups using a random number tablecontrol group(group C,n=24),different concentrations of lipid emulsion groups(LE1 groups [n =24],LE2 group [n =24] and LE3 group [n =72]),different concentrations of emulsified isoflurane groups (EI1 group [n=24],El2 group [n=24] and EI3 group [n=72]),and emulsified isoflurane + JNK inhibitor SP600125 group (group EI-SP,n =24).At 24 h after the cells were plated,in LE1-3 groups,30％ lipid emulsion was added to the culture medium with the final concentrations of 0.395 6,0.791 2 and 1.582 4 μl/ml,respectively;in EI1-3 groups,8％ emulsified isoflurane was added with the final concentrations of 0.56,1.12 and 2.24 mmol/L,respectively;in group EI-SP,emulsified isoflurane was added with the final concentration of 1.12 mmol/L,and SP600125 was added at 1 h before addition of emulsified isoflurane with the final concentration of 10 μmol/L;the cells were cultured normally in group C.At 6,12 and 24 h of incubation in EI3 and LE3 groups,and at 24 h of incubation or culture in the other groups,the morphology of cells was detected,the cell viability was measured using methyl thiazolyl tetrazolium assay,and the expression of JNK,phosphorylated JNK (p-JNK) and cytochrome c (Cyt c) was detected by Western blot.Results Compared with group C,the cell viability was significantly decreased,and the expression of p-JNK and Cyt c was significantly up-regulated at 24 h of incubation in group EI2 and at 12 and 24 h of incubation in group EI3,the cell viability was significantly decreased (P＜0.05),and no significant change was found in the expression of p-JNK and Cyt c in group EI-SP,and no significant change was found in the cell viability and expression of p-JNK and Cyt c in LE1-3 and EI1 groups (P＞0.05).Compared with group EI1,the cell viability was significantly decreased,and the expression of p-JNK and Cyt c was significantly up-regulated at 24 h of incubation in EI2.3 groups (P＜0.05).Compared with group EI2,the cell viability was significantly decreased,and the expression of p-JNK and Cyt c was significantly up-regulated at 24 h of incubation in group EI3,and the cell viability was significantly increased,and the expression of p-JNK and Cyt c was significantly down-regulated in group EI-SP (P＜0.05).There was no significant difference in JNK expression between the eight groups (P＞0.05).Conclusion High concentrations of emulsified isoflurane can induce apoptosis in neurons only when applied for a long time,while low concentrations do not have the effect when applied for a short time.The mechanism by which emulsified isoflurane induces neuronal apoptosis is related to activation of JNK pathway.