肾小管上皮细胞中可溶性表氧化物水解酶对小鼠巨噬细胞极化的调控作用  

Role of soluble epoxide hydrolase in renal tubular epithelial cells in the regulation of macrophage polarization in mice

作　　者：王倩[1] 赵向娅[1] 赵倩茹[1] 杨轶[1] 杨胜楠[1] 李冰[1] 李馨 田蕊[1] WANG Qian;ZHAO Xiang-ya;ZHAO Qian-ru;YANG Yi;YANG Sheng-nan;LI Bing;LI Xin;TIAN Rui(Department of Geriatric Medicine,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 45000,Herman Province,China)

机构地区：[1]郑州大学第一附属医院老年综合二科,河南郑州450052

出　　处：《新乡医学院学报》2018年第5期355-360,共6页Journal of Xinxiang Medical University

基　　金：国家自然科学基金资助项目(编号:81300614);河南省教育厅科技计划项目(编号:142300410376)

摘　　要：目的探讨肾小管上皮细胞中可溶性表氧化物水解酶(sEH)在小鼠巨噬细胞极化中的作用。方法以人近端肾小管上皮细胞系HK-2细胞及小鼠巨噬细胞系RAW264.7细胞为研究对象。将HK-2细胞分为正常对照组、sEH抑制剂组、尿蛋白组及sEH抑制剂联合尿蛋白组;正常对照组细胞不给予任何干预处理;sEH抑制剂组细胞给予1μmol·L^(-1)sEH抑制剂;尿蛋白组细胞给予10 g·L^(-1)尿蛋白;sEH抑制剂联合尿蛋白组细胞给予10 g·L^(-1)尿蛋白和1μmol·L^(-1)sEH抑制剂;各组细胞均培养24 h。将RAW264.7细胞分为A、B、C、D组及干扰素-γ(IFN-γ)阳性对照组、白细胞介素(IL)-4阳性对照组。A、B、C、D组细胞分别加入正常对照组、sEH抑制剂组、尿蛋白组、sEH抑制剂联合尿蛋白组培养24 h的HK-2细胞培养基孵育24 h;IFN-γ阳性对照组和IL-4阳性对照组细胞分别加入M1型巨噬细胞诱导剂IFN-γ及M2型巨噬细胞诱导剂IL-4。采用Western blot法检测HK-2细胞中sEH蛋白表达,实时荧光定量聚合酶链反应检测HK-2细胞中单核细胞趋化蛋白-1(MCP-1)、IL-6、集落刺激因子-1(CSF-1)、肿瘤坏死因子-α(TNF-α)mRNA表达及RAW264.7细胞中诱导型氮氧化物合酶(iNOS)、IL-6、精氨酸酶-1(Arg^(-1))及IL-10 mRNA表达。酶联免疫吸附试验检测各组HK-2细胞培养上清液中14,15-环氧二十碳三烯酸(14,15-EET)和14,15-脱氧二十碳三烯酸(14,15-DHET)水平,计算14,15-EET/14,15-DHET比值。结果正常对照组和sEH抑制剂组HK-2细胞中sEH蛋白表达及MCP-1、IL-6、CSF-1、TNF-αmRNA表达、细胞培养上清液中14,15-EET/14,15-DHET比较差异均无统计学意义(P>0.05)。与正常对照组比较,蛋白尿组HK-2细胞中sEH蛋白及MCP-1、IL-6、CSF-1、TNF-αmRNA表达均显著增加(P<0.05),细胞培养上清液中14,15-EET/14,15-DHET显著降低(P<0.05)。与尿蛋白组比较,sEH抑制剂联合尿蛋白组HK-2细胞中MCP-1、IL-6、CSF-1及TNF-αmRNA表达显著下降(P<0.05),培养上Objective To investigate the role of soluble epoxide hydrolase（ sEH） in renal tubular epithelial cells in the regulation of macrophage polarization in mice. Methods Human proximal tubular epithelial cell line（ HK-2） and mouse macrophage cell line（ RAW264. 7） were studied in the present study. HK-2 cells were divided into normal control group,sEH inhibitor group,urine protein group and sEH inhibitor combined urine protein group. The cells in the normal control group wasn’t given any intervention. The cells in the sEH inhibitor group and the urine protein group were given 1 μmol·L-1 sEH inhibitor and 10 g · L-1 urine protein,respectively; while the cells in the sEH inhibitor combined urine protein group were given 1 μmol·L-1 sEH inhibitor and 10 g·L-1 urine protein. All the cells in each group were cultured for 24 hours. RAW264. 7 cells were divided into group A,group B,group C,group D,interferon gamma（ IFN-γ） positive control group and interleukin（ IL）-4 positive control group. The cells of group A,group B,group C and group D were respectively added to the HK-2 cell cultured medium of the normal control group,sEH inhibitor group,urine protein group and sEH inhibitor combined urine protein group; then the cells were cultured for 24 h. IFN-γ and IL-4 were respectively added in IFN-γ positive control group and IL-4 positive control group. The expression of sEH protein in HK-2 cells was measured by Western blot. The expression of monocyte chemotactic protein-1（ MCP-1）,IL-6,colony stimulating factor-1（ CSF-1）,tumor necrosis factor-α（ TNF-α） mRNA in the HK-2 cells and the expression of inducible nitric oxide synthase（ iNOS）,arginase（ Arg-1）,IL-10 mRNA in the RAW264. 7 cells were measured by real-time polymerase chain reaction. The levels of 14,15-epoxyeicosatrienoic acids（ 14,15 EET） and 14,15-dihydroxyeicosatrienoic acid（ 14,15-DHET） in the supernate of HK-2 cell medium in each group were detected by enzyme linked immunosorbent assay,and the 14,15-EET/14,15-D

关 键 词：可溶性表氧化物水解酶 肾小管上皮细胞 巨噬细胞极化 

分 类 号：R589[医药卫生—内分泌]