【摘要】 目的应用衣霉素诱导内质网应激(ERS),观察Roscovtine对ERS导致足细胞损伤的保护作用。方法体外培养永生化小鼠足细胞,取37℃分化成熟细胞随机分组:(1)正常对照组、DMSO组及衣霉素Tunicamycin(1.0μmol/L,TM)组,各实验组分别刺激3、6、12 h。(2)正常对照组、Tunicamycin(1.0μmol/L,TM)组及Tunicamycin+Rosco-vitine(20、40μmol/L,TM+ROS)组,各实验组分别刺激12 h。应用流式细胞术及TUNEL法检测足细胞凋亡情况;应用Western blot检测细胞周期素依赖性蛋白激酶5(Cdk5)及ERS标志蛋白GRP78、Caspase-12、CHOP的表达变化。结果 (1)与正常对照组和DMSO组相比,Tunicamycin刺激3、6及12 h后,TM组足细胞凋亡率及Cdk5、GRP78、Caspase-12和CHOP蛋白的表达水平均呈明显的时间依赖性增高(P<0.05);(2)加入Roscovitine干预后,与TM组相比,TM+ROS(20、40μmol/L)组足细胞凋亡率及GRP78、Caspase-12和CHOP蛋白的表达水平均显著降低(P<0.05),其干预作用呈现明显的剂量依赖性。结论 Cdk5抑制剂Roscovitine能显著抑制衣霉素诱导的足细胞凋亡,从而发挥细胞保护作用。Roscovitine的足细胞保护作用可能有助于糖尿病肾病的治疗。更多还原

【Abstract】 Objective To observe the protective effects of roscovitine on the podocyte injury induced by endoplasmicreticulum stress(ERS) caused by tunicamycin. Methods The differentiated podocytes cultured at 37 ℃ were randomly di-vided into:(1) Control group,DMSO group and tunicamycin group(TM,1.0 μmol/L). The treatment was given for 3,6 and 12 hours in three groups.(2) For control group,tunicamycin group,tunicamycin+roscovitine group(20,40 μmol/L,TM+ROS),the treatment was given for 12 hours. The podocyte apoptosis was detected by flow cytometry and TUNEL method.The ex-pressions of Cdk5,GRP78,Caspase-12 and CHOP were detected by Western blot assay. Results(1) Compared with con-trol group and DMSO group,the podocyte apoptosis was increased significantly in a time dependent manner after tunicamy-cin treatment in TM group; the protein expressions of Cdk5,GRP78,Caspase-12 and CHOP were also up-regulated signifi-cantly in TM group(P < 0.05).(2) Flow cytometry and TUNEL analysis showed that tunicamycin induced apoptosis in podo-cytes,which was significantly inhibited by roscovitine in a concentration dependent manner in TM+ROS group as comparedto that of TM group(P < 0.05). The protein expressions of GRP78,Caspase-12 and CHOP were also significantly decreasedin a concentration dependent manner in TM+ROS group compared to those of TM group(P < 0.05). Conclusion Roscovi-tine,the inhibitor of Cdk5,can reduce the podocyte apoptosis induced by tunicamycin. The protective effects of roscovitineon podocytes can be a novel approach of treating diabetic nephropathy.更多还原