硫酸吲哚酚通过上调微小RNA-21表达参与心肌成纤维细胞的旁分泌  

MiroRNA- 21 mediates myocardial fibroblasts paracrine stimulated by indoxyl sulfate in vitro

作　　者：龚建光[1] 李一文[1] 金娟[1] 赵黎[1] 李一乔[1] 何强[1] 

机构地区：[1]浙江省人民医院杭州医学院附属人民医院肾脏病科,杭州310014

出　　处：《中华肾脏病杂志》2017年第12期935-939,共5页Chinese Journal of Nephrology

基　　金：浙江省中医药科技计划项目（2016ZA023、2017ZA008）;浙江省自然基金面上项目（Y16H050024）

摘　　要：目的探讨硫酸吲哚酚（Is）对心肌成纤维细胞微小RNA-21（miRNA-21）表达的影响，并探讨miRNA-21对心肌成纤维细胞旁分泌的作用。方法小鼠心肌成纤维细胞分成空白对照组和IS组培养48h，实时荧光定量PCR法检测miRNA．21的表达。采用RNA干扰技术沉默miRNA-21表达，将小鼠心肌成纤维细胞分成：血肌酐（Scr）组（Scr处理48h）、Scr＋IS组（Scr＋50μmol／LIS处理48h），Scr＋miRNA-21抑制物组（miRNA．21抑制物处理24h后加Scr处理48h）和Scr＋miRNA-21抑制物＋IS组（miRNA-21抑制物处理24h后加Ser和IS处理48h）。采用ELISA及Western印迹分别检测细胞外白细胞介素1（IL-1）、白细胞介素6（IL-6）、肿瘤坏死因子仪（TNF—α）的表达及细胞内转化生长因子β（TGF-β）、基质金属蛋白酶2（MMP2）、基质金属蛋白酶9（MMP9）、基质金属蛋白酶1的组织抑制因子（TIMPl）的蛋白表达。结果与空白对照组比较，IS组心肌成纤维细胞miRNA-21的表达增加（P〈0．01）。与Scr组比较，Scr＋IS组IL-1、IL-6、TNF—α、TGF-β、MMP2、MMP9表达均升高（均P〈0．05），TIMPl表达降低（P〈0．05）；与Scr＋IS组比较，Scr＋miRNA-21抑制物＋IS组IL-1、IL-6、TNF-α、TGF—β、MMP2表达均降低，TIMP1表达也进一步下降（均P〈0．05）。结论尿毒症毒素IS可以促进心肌成纤维细胞miRNA-21的表达上调，miRNA-21参与调节心肌成纤维细胞炎性因子、促纤维化因子等的分泌。Objective To explore the expression of miroRNA-21 （miRNA- 21） in myocardial fibroblasts stimulated by indoxyl sulfate （IS） and its role on paracrine factors of myocardial fibroblasts. Methods Myocardial fibroblasts which derived from C57BL/6J mice were divided into control group and IS group, and their expressions of miRNA-21 were detected by real time PCR after 48 h. MiRNA-21 inhibitor transfection was applied to silence miRNA-21 expression. Myocardial fibroblasts were divided into creatinine （Scr） group （Set treated for 48 h）, Scr＋IS group （Ser and 50μmol/L IS treated for 48 h）, Scr＋miRNA-21 inhibitor group （miRNA-21 inhibitor treated for 24 h and then Scr treated for 48 h） and Scr＋miRNA-21 inhibitor＋IS group （miRNA-21 inhibitor pretreated for 24 h and then Set and IS treated for 48 h）. Enzyme- linked immunosorbent assay was performed to evaluate the expressions of interleukin-1 （IL-1）, interleukin-6 （IL-6） and tumor necrosis factor-α （TNF-α）. Western blotting was applied to detect the expressions of transforming growth factor-β（TGF-β）, matrix metalloproteinases 2 （MMP2）, matrix metalloproteinases 9 （MMP9） and tissue inhibitor of metalloproteinases 1 （TIMP1）. Results The expression of miRNA-21 was obviously increased in IS group than that in control group （P 〈 0.01）. Compared with those in Scr group, the expressions of IL-1, IL-6, TNF-α, TGF-β, MMP2 and MMP9 significantly increased （all P 〈 0.05）, while the expression of TIMP1 decreased （P 〈 0.05）. When the expression of miRNA-21 was inhibited, the expressions of IL-1, IL-6, TNF-α, TGF-β and MMP2 in Scr＋miRNA-2l inhibitor＋IS group significantly decreased than those in Scr＋IS group （all P 〈 0.05）, and the expression of TIMPI decreased further （P 〈 0.05）. Conclusions IS can promote the expression of miRNA- 21 in myocardial fibroblasts. MiRNA- 21 plays an important role in regulating inflammatory factors and pro-fibrogenic cytokines in myocardial fibroblast

关 键 词：尿毒症 微RNAS 旁分泌细胞细胞间通讯 靛甙 

分 类 号：R692.5[医药卫生—泌尿科学]