TGF-β1调控Cdk5表达在人肾系膜细胞外基质沉积中的作用  

Role of TGF-β1 in regulating Cdk5 expressionin human mesangial extracellular matrix deposition

作　　者：周毅[1] 杨亚坤 康平 柳丹 夏顺杰 张悦 刘巍[2] ZHOU Yi;YANG Ya-kun;KANG Ping;LIU Dan;XIA Shun-jie;ZHANG Yue;LIU Wei(The Second Hospital,Hebei Medical University,Shijiazhuang 050017,China;Dept of Pathology,Hebei Medical University,Shijiazhuang 050017,China;Dept of Clinical Diagnostics,Hebei Medical University,Shijiazhuang 050017,China)

机构地区：[1]河北医科大学第二医院,河北石家庄050017 [2]河北医科大学病理学教研室,河北石家庄050017 [3]河北医科大学诊断学教研室,河北石家庄050017

出　　处：《中国药理学通报》2021年第12期1688-1693,共6页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 81770720);河北省自然科学基金杰出青年科学基金(No H2019206550);河北省自然科学基金面上项目(No H2019206549);河北省科技厅重点研发计划项目(No 19277784D)。

摘　　要：目的研究TGF-β1调控Cdk5表达在人肾系膜细胞外基质沉积中的作用。方法10μg·L^(-1)的TGF-β1刺激体外培养的人肾小球系膜细胞(HMC),检测其对系膜细胞Cdk5、p35/p25表达的影响;转染Cdk5过表达质粒,Western blot、免疫荧光检测胶原蛋白FN、CollagenⅣ表达的变化;应用Cdk5激酶活性抑制剂Roscovitine或转染Cdk5干扰质粒,Western blot、Real time-PCR、免疫荧光检测抑制Cdk5对TGF-β1刺激HMC后FN、CollagenⅣ蛋白及mRNA表达的影响。结果TGF-β1作用于HMC后,Cdk5、p35/p25蛋白及mRNA表达均呈时间依赖性增高;与正常对照组相比,转染Cdk5过表达质粒,FN、CollagenⅣ蛋白表达增加,FN(绿色荧光)、CollagenⅣ(红色荧光)增强;与TGF-β1刺激组相比,加用10μmol·L^(-1)的Roscovitine或转染Cdk5干扰质粒,FN、CollagenⅣ蛋白及mRNA表达均减少,FN(绿色荧光)、CollagenⅣ(红色荧光)减弱,而空质粒转染组没有明显变化。结论一定浓度的TGF-β1可诱导系膜细胞内Cdk5表达增加,抑制Cdk5可降低系膜细胞外基质沉积。Aim To investigate the role of TGF-β1 in regulating Cdk5 expression in human mesangial extracellular matrix deposition.Methods TGF-β1(10μg·L^(-1))stimulated human glomerular mesangial cells(HMC)in vitro to detect the expression of Cdk5 and p35/p25.Cdk5 overexpression plasmid was transfected,and the expressions of FN and collagen IV were detected by Western blot and immunofluorescence.Using Roscovitine,an inhibitor of Cdk5 kinase,or transfecting Cdk5 interfering plasmid,the effects of Cdk5 inhibition on the expression of FN and collagen IV in HMC stimulated by TGF-β1 were assessed by Western blot,Real time-PCR and immunofluorescence.Results The protein and mRNA expression of Cdk5 and p35/p25 increased in a time-dependent manner after HMC was stimulated by TGF-β1.Compared with normal control group,the expression of FN and collagen IV increased after transfection with Cdk5 overexpression plasmid.FN(green fluorescence)and collagen IV(red fluorescence)were enhanced.Compared with TGF-β1 stimulation group,the protein and mRNA expression of FN and collagenⅣdecreased when Roscovitine(10μmol·L^(-1))was added or Cdk5 interference plasmid was transfected.And FN(green fluorescence)and collagenⅣ(red fluorescence)were reduced.However,there was no significant change in the empty plasmid transfection group.Conclusions A certain concentration of TGF-β1 can increase the expression of Cdk5,and the inhibition of Cdk5 can reduce the deposition of extracellular matrix in mesangial cells.

关 键 词：TGF-Β1 CDK5 系膜细胞 肾纤维化 基质沉积 ROSCOVITINE 

分 类 号：R322.61[医药卫生—人体解剖和组织胚胎学] R329.24[医药卫生—基础医学]