黄蜀葵花对碘普罗胺诱导肾小管上皮细胞损伤的作用及其机制研究  

Effect of Abelmoschus manihot on iopromide-induced renal tubular epithelial cell damage and its underlying mechanisms

作　　者：徐中驰 刘春玲[2] 林欣[2] 高坤[3] 牛茹歌 Zhong-chi Xu;Chun-ling Liu;Xin Lin;Kun Gao;Ru-ge Niu(The First Clinical Medical College,Nanjing University of Chinese Medicine,Nanjing,Jiangsu 210023,China;Department of Cardiology,Jiangsu Provincial Hospital of Traditional Chinese Medicine,Nanjing,Jiangsu 210004,China;Department of Nephrology,Jiangsu Provincial Hospital of Traditional Chinese Medicine,Nanjing,Jiangsu 210004,China)

机构地区：[1]南京中医药大学第一临床医学院,江苏南京210023 [2]江苏省中医院心内科,江苏南京210004 [3]江苏省中医院肾内科,江苏南京210004

出　　处：《中国现代医学杂志》2022年第8期20-27,共8页China Journal of Modern Medicine

基　　金：国家自然科学基金(No:81973762);江苏省中医院夏桂成创新发展基金(No:Y19019)。

摘　　要：目的研究黄蜀葵花防治碘普罗胺诱导的人近端肾小管上皮细胞(HK2)损伤的作用和机制。方法利用碘普罗胺孵育HK2细胞诱导的体外细胞损伤模型,采用黄蜀葵花制剂(TFA)干预碘普罗胺处理的HK2细胞,分为空白组、模型组(111 mgI/mL碘普罗胺)、TFA组(0.6 mg/mL TFA)、TFA+模型组(111 mgI/mL碘普罗胺+0.6 mg/mL TFA)、N-乙酰半胱氨酸(NAC)组(10 mmol/L NAC)、NAC+模型组(111 mgI/mL碘普罗胺+10 mmol/L NAC)。采用CCK-8法检测细胞活性,活性氧(ROS)检测试剂盒检测ROS,Annexin V-FITC及TUNEL染色检测细胞凋亡,免疫荧光法检测p-ASK1荧光,Western blotting检测通路凋亡蛋白表达。结果碘普罗胺呈浓度依赖性诱导HK2细胞死亡,促进HK2细胞ROS的产生。空白组、TFA组与NAC组HK2细胞活性比较,差异无统计学意义(P>0.05);TFA+模型组、NAC+模型组HK2细胞活性高于模型组(P<0.05)。Annexin V-FITC、TUNEL染色及流式细胞术结果显示,与空白组比较,模型组荧光强度明显增加;TFA组、NAC组与空白组比较无明显差异;与模型组比较,TFA及+模型组和NAC+模型组细胞荧光强度减弱。免疫荧光法结果表明,与空白组比较,模型组p-ASK1荧光表达明显增强;TFA组、NAC组与空白组比较,荧光表达无明显差异;TFA+模型组、NAC+模型组的p-ASK1荧光强度较模型组减弱。Western blotting检测结果表明,与空白组比较,模型组Bcl-2/Bax蛋白表达降低(P<0.05),Cleaved caspase-3/Caspase-3、pP38/P38、pJNK/JNK蛋白表达升高(P<0.05);与模型组比较,TFA+模型组和NAC+模型组Bcl-2/Bax蛋白表达升高(P<0.05),Cleaved caspase-3/Caspase-3、pP38/P38、pJNK/JNK蛋白表达降低(P<0.05)。结论TFA可以减少碘普罗胺诱导的HK2细胞凋亡,其机制可能与TFA抑制ROS-ASK1-MAPKs信号通路激活有关。Objective To study the effect of Abelmoschus manihot on iopromide-induced renal tubular epithelial cell damage and its underlying mechanisms.Methods Human kidney-2 cells(HK2)were incubated with ioproamine in vitro to establish renal tubular epithelial cell damage models,which were followed by the treatment with total flavone of Abelmoschus manihot(TFA)or not.The cells were divided into blank group(untreated),model group(111 mgI/mL of iopromide),TFA group(0.6 mg/mL of TFA)group,TFA-model group(111 mgI/mL of iopromide plus 0.6 mg/mL of TFA),NAC group(10 mmol/L of NAC),and NAC-model group(111 mgI/mL of iopromide plus 10 mmol/L of NAC).The cell viability was detected via CCK-8 assay,and the reactive oxygen species were detected via commercial kits.The cell apoptosis was analyzed by Annexin V-FITC apoptosis staining and TUNEL staining.The expression of p-ASK1 was analyzed by immunofluorescence staining,while the levels of apoptosis-associated proteins were measured via Western blotting.Results Iopromide induced HK2 cell death in a concentration-dependent manner and promoted the production of ROS in HK2 cells.There was no difference in the cell viability among the blank group,TFA group and NAC group(P>0.05),while the viability of HK2 cells in the TFA-model group and NAC-model was higher than that in the model group(P<0.05).As shown in Annexin V FITC and TUNEL staining as well as the flow cytometry,the fluorescence intensity of p-ASK1 was higher in the model group relative to that in the blank group,and was lower in the TFA-model group and NAC-model group compared with the model group.However,there was no difference in the fluorescence intensity of p-ASK1 between the TFA group and NAC group.The Western blotting exhibited that the Bcl-2/Bax expression ratio was decreased,but the cleaved Caspase-3/Caspase-3,pP38/P38 and p-JNK/JNK expression ratios were increased in the model group compared with the blank group(P<0.05).In comparison with the model group,the Bcl-2/Bax expression ratio was increased,but the cleaved Caspase-3

关 键 词：造影剂肾病 碘普罗胺 黄蜀葵花总黄酮 活性氧 ASK1 

分 类 号：R692[医药卫生—泌尿科学]