穿心莲内酯在镉致IEC6细胞线粒体自噬中的作用  

Role of andrographolide in mitochondrial autophagy in IEC6 cells induced by cadmium

作　　者：王涛[1] 蔡瑶 曹彬彬 袁燕[1] 张泉[1] 刘宗平[1] WANG Tao;CAI Yao;CAO Bin-bin;YUAN Yan;ZHANG Quan;LIU Zong-ping(College of Veterinary Medicine,Yangzhou University,Yangzhou 225009,China)

机构地区：[1]扬州大学兽医学院,江苏扬州225009

出　　处：《中国兽医科学》2022年第4期524-530,共7页Chinese Veterinary Science

基　　金：江苏高校优势学科建设工程资助项目(PAPD);国家自然科学基金青年基金项目(31802260);江苏省自然科学基金青年基金项目(BK20180917);江苏省高等学校自然科学研究面上项目(18KJB230009)。

摘　　要：为探讨穿心莲内酯(Ad)在镉诱导肠上皮细胞线粒体自噬中的作用,体外培养IEC6细胞,以5μmol/L镉处理不同时间(0、6、12 h),利用荧光显微镜观察细胞中外源性GFP-RFP-LC3聚点的变化。用7μmol/L Ad预处理IEC6细胞16 h后,5μmol/L镉处理细胞12 h,利用Western-blot方法检测线粒体自噬相关蛋白p62、LC3、PINK1、Parkin的表达水平;利用荧光显微镜观察GFP-RFP-LC3和Bcl-XL在细胞中分布的变化。结果,5μmol/L镉处理IEC6细胞不同时间后,红色LC3聚点数量增多,与6 h组相比,镉处理细胞12 h后,LC3红色聚点数量下降;Western-blot结果显示,与镉单独处理组相比,Ad与镉共处理组Parkin蛋白表达水平升高,p62、LC3-II、PINK1表达水平降低;荧光观察结果显示,与镉单独处理组相比,Ad与镉共处理组LC3、Bcl-XL红色信号增强,绿色信号减弱。上述结果表明,镉能够引起IEC6细胞自噬流阻滞,Ad可以通过缓解镉诱导的自噬流阻滞,促进镉致IEC6细胞线粒体自噬的发生。To investigate the effect of andrographolide on mitophagy induced by cadmium(Cd)in intestinal epithelial cells,IEC6 cells were cultured with 5μmol/L Cd for different time(0,6,12 h),the change of exogenous GFP-RFP-LC3 puncta was observed by using fluorescence microscope.IEC6 cells were pretreated with 7μmol/L Ad for 16 h and then treated with 5μmol/L Cd for 12 h,the expression of p62,LC3,PINK1 and Parkin proteins which was involoved in the regulation of mitophagy was detected by Western-blot,and the distribution of GFP-RFP-LC3 and Bcl-XL in cells was observed by using fluorescence microscope.In result,after IEC6 cells treated with 5μmol/L Cd for different time,the number of red LC3 puncta increased.The number of LC3 red puncta decreased in IEC6 cells with Cd treatment for 12 h compared with the 6 h group.The result of Western-blot showed that the expression of parkin increased while the expression level of P62,LC3-II and PINK1 decreased in cells treated with Cd in the presence of Ad compared with the Cd group.Fluorescence result showed that the red signal of LC3 and Bcl-XL was enhanced while the green signal was degraded in cells treated with Cd in the presence of Ad compared with the Cd group.In conclusion,Cd triggers the blockage of autophagic flux.Ad promoted the occurrence of mitophagy via alleviating the blockage of autophagic flux induced by Cd in IEC6 cells.

关 键 词：穿心莲内酯 线粒体自噬 镉 IEC6细胞 

分 类 号：S853.74[农业科学—临床兽医学]