Effects of emulsified isoflurane on apoptosis in human neuroblastoma cells: the role of c-Jun N-terminal kinase

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 table: control 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], EI2 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). 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). 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. Key words: Isoflurane; Fat emulsions, intravenous; Neurons; Apoptosis; JNK mitogen-activated protein kinases