环孢素引起肾小管上皮细胞培养液中肾损伤分子-1水平升高的机制

摘要

Objective To investigate the mechanism of increasing of the level of kidney injury molecule-1(KIM-1)in culture supernatant of human kidney cells(HKC)induced by cyclosporine A(CsA),and to clarify the relationships between the expression levels of KIM-1 and p38 MAPK pathway and ERK1/2MAPK pathway in HKC. Methods The HKC at logarithmic growth phase were randomly divided into control group, CsA control group, CsA + p38 kinase inhibitor group, p38 kinase inhibitor group, CsA + ERK1/2 inhibitor group and ERK1/2 kinase inhibitor group.The inhibitory rates of proliferation of HKC in various groups were detected by MTT assay, and the expression levels of KIM-1 in HKC supernatant in various groups were detected by ELISA;the survival rates,apopototic rates and necrotic rates of the HKC in various groups were detected by flow cytometry. Results Compared with control group,the expression level of KIM-1 protein in the supernatant of HKC in CsA control group was significantly increased (P<0.05),and the survival rate was significantly decreased (P<0.05), while the apoptotic rate and the necrotic rate were significantly increased (P<0.05 ). Compared with control group,the survival rates, the apoptotic rates and the necrosis rates of cells in p38 kinase inhibitor group and ERK1/2 kinase inhibitor group had no significant differences(P>0.05).Compared with CsA control group,the expression levels of KIM-1 protein in CsA+ p38 kinase inhibitor group and CsA+ ERK1/2 kinase inhibitor group were significantly decreased (P<0.05),and the survival rate was significantly increased (P<0.05),while the apoptotic rate and the necrotic rate were significantly decreased (P<0.05).Conclusion p38 MAPK pathway and ERK1/2MAPK pathway are involved in the process of up-regulation of the KIM-1 level in HKC culture supernatant induced by CsA,and the expression of KIM-1 may become the biochemical marker of clinical monitoring of CsA nephrotoxicity.%目的：探讨环孢素（CsA）引起人肾小管上皮细胞（HKC）培养液中肾损伤分子-1（KIM-1）水平升高的作用机制，阐明KIM-1表达与 p38 MAPK通路和 EKR1/2 MAPK通路的关系。方法：将处于对数生长期的 HKC分为空白组、CsA损伤组、CsA与 p38激酶抑制剂合用组、p38激酶抑制剂组、CsA与 ERK1/2激酶抑制剂合用组和 ERK1/2激酶抑制剂组。MTT法检测各组 HKC增殖抑制率，ELISA法测定各组 HKC上清液中 KIM-1水平，流式细胞术检测各组细胞存活率、细胞凋亡率和细胞坏死率。结果：与空白组比较， CsA损伤组细胞上清液中KIM-1水平显著升高（P＜0.05），细胞存活率显著降低（P＜0.05），细胞凋亡率和细胞坏死率显著升高（P＜0.05）；p38激酶抑制剂组和 ERK1/2激酶抑制剂组中细胞存活率、细胞凋亡率和细胞坏死率比较差异无统计学意义（P＞0.05）。与CsA损伤组比较，CsA与 p38激酶抑制剂合用组、CsA与 ERK1/2激酶抑制剂合用组细胞上清液中KIM-1水平显著降低（P＜0.05），细胞存活率显著升高（P＜0.05），细胞凋亡率和细胞坏死率显著降低（P＜0.05）。结论：p38 MAPK通路和ERK1/2 MAPK通路参与CsA素引起肾小管上皮细胞培养液中KIM-1水平升高的过程，KIM-1的表达可能成为临床监测CsA肾毒性的生化指标。