Abstract: Objective　 To investigate the efficacy of tacrolimus（FK506）on the proliferation，apoptosis and expression of inflammatory factors of high glucose-induced human renal tubular epithelial（HK-2）cells.Methods　From October 2014 to May 2015,HK-2 cells cultured in vitro were randomly divided into a blank control group，high glucose induction group（30 mmol/L high glucose），mannitol induction group（24.5 mmol/L mannitol），20 μmol/L FK506 control group，different concentration FK506 inhibition groups（30 mmol/L high glucose+1，5，10 and 20 μmol/L FK506），and NF-κB signaling inhibitor group（30 μmol/L Bay11-7082+30 mmol/L high glucose）.After 24 hours of intervention，the expression level of NF-κB mRNA was determined by RT-PCR；those of interleukin-6（IL-6）and tumor necrosis factor-α（TNF-α）in supernatant were detected by ELISA；cell proliferation was measured by CCK-8；cell apoptosis was detected by flow cytometry.Results　The difference in the expression level of NF-κB mRNA among the groups showed a statistical significance（P<0.001）.Compared with the high glucose induction group，the expression level of NF-κB mRNA was lower in the blank control group（P<0.05）；and it was also lower in 1，5，10，20 μmol/L FK506 inhibition groups in a concentration dependent manner（P<0.05）.NF-κB signaling inhibitor group had lower expression level of NF-κB mRNA than the blank control group，high glucose induction group，and 1，5，10 μmol/L FK506 inhibition groups（P<0.05）.The expression levels of both IL-6 and TNF-α were statistically significant among all the groups（P<0.001）.Compared with the blank control group，the expression levels of both IL-6 and TNF-α were higher in the high glucose induction group（P<0.05）.The expression levels of both IL-6 and TNF-α were higher in the 1，5，10，20 μmol/L FK506 inhibition groups than those in the blank control group,but lower than those in the high glucose induction group in a concentration dependent manner（P<0.05）.Compared with the NF-κB signaling inhibitor group，the expression levels of both IL-6 and TNF-α were lower in the blank group and higher in the high glucose induction group and 1，5，10 μmol/L FK506 inhibition groups（P<0.05）.CCK-8 method showed that processing method and time exerted interaction effect on the proliferation of HK-2 cells；processing method led to main effect on the proliferation of HK-2 cells；processing time had main effect on the proliferation of HK-2 cells（Finteraction effect=16.315，Pinteraction effect<0.001；Fgroup=24.618，Pgroup<0.001；Ftime=16.315，Ptime<0.001）.The results of flow cytometry analysis showed that the difference in 24 h apoptosis rate of HK-2 cells among all the groups exhibited statistical significance（P<0.001）.Compared with the high glucose induction group，the 24 h apoptosis rate of HK-2 cells was lower in the bland control group；and it was also lower in 1，5，10，20 μmol/L FK506 inhibition groups and the NF-κB signaling inhibitor group（P<0.05）.ConclusionFK506 showed cytotoxicity for human normal HK-2 cells，while the HK-2 cells under high glucose FK506 is manifested protection.High glucose mediated kidney inflammation through NF-κB signaling，and FK506 alleviated the progression of diabetic nephropathy，which may be related to NF-κB signaling.

Key words: Diabetic nephropathies, HK-2 cells, Tacrolimus, NF-kappa B

摘要： 目的探讨他克莫司（FK506）对高糖诱导人肾小管上皮细胞（HK-2）增殖、凋亡及其表达相关炎性因子的影响。方法　于2014年10月—2015年5月，以人HK-2为研究对象，分为空白对照组、高糖诱导组（30 mmol/L高糖）、甘露醇组（24.5 mmol/L甘露醇）、20 μmol/L FK506对照组、不同浓度FK506干预组（30 mmol/L高糖+1、5、10、20 μmol/L FK506）及核因子（NF）-κB信号通路抑制剂组（30 μmol/L Bay11-7082+30 mmol/L高糖），各组均培养24 h。应用反转录聚合酶链式反应（RT-PCR）检测各组HK-2 NF-κB mRNA的表达；酶联免疫吸附试验（ELISA）检测白介素6（IL-6）和肿瘤坏死因子α（TNF-α）的表达；CCK-8法测定细胞增殖；流式细胞技术检测细胞凋亡。结果　各组NF-κB mRNA表达水平比较，差异有统计学意义（P<0.001）；其中高糖诱导组NF-κB mRNA表达水平高于空白对照组；不同浓度FK506干预组NF-κB mRNA表达水平低于高糖诱导组，且呈浓度依赖性；NF-κB信号通路抑制剂组NF-κB mRNA表达水平低于空白对照组、高糖诱导组及1、5、10 μmol/L FK506干预组（P<0.05）。各组IL-6、TNF-α表达水平比较，差异均有统计学意义（P<0.001）；其中高糖诱导组IL-6、TNF-α表达水平高于空白对照组；不同浓度FK506干预组IL-6、TNF-α表达水平高于空白对照组，低于高糖诱导组，且呈浓度依赖性；NF-κB信号通路抑制剂组IL-6、TNF-α表达水平高于空白对照组，低于高糖诱导组和1、5、10 μmol/L FK506干预组（P<0.05）。处理方法与时间对HK-2增殖存在交互作用（F交互=16.315，P交互<0.001）；处理方法对HK-2增殖主效应显著（F组间=24.618，P组间<0.001）；时间对HK-2增殖主效应显著（F时间=16.315，P时间<0.001）。各组24 h凋亡率比较，差异有统计学意义（P<0.001）；其中高糖诱导组24 h凋亡率高于空白对照组；不同浓度FK506干预组和NF-κB信号通路抑制剂组24 h凋亡率低于高糖诱导组（P<0.05）。结论　FK506对正常人HK-2表现为毒性作用，而对高糖环境下的HK-2则表现为保护作用。高糖可通过NF-κB信号通路介导肾脏炎症，而FK506缓解糖尿病肾病进展的机制可能与NF-κB信号通路有关。

关键词: 糖尿病肾病, 肾小管上皮细胞, 他罗利姆, NF-κB