摘要
为探究内质网释放的Ca^2+是否参与ZEA诱导TM4细胞自噬,本研究以小鼠睾丸支持细胞细胞株(TM4细胞)为材料,用不同浓度ZEA(0、5、10、20μmol/L)及BAPTA-AM(Ca^2+螯合剂)、2-APB(内质网Ca^2+通道IP3R受体拮抗剂)单独或联合作用TM4细胞,染毒时间24 h。流式细胞术检测胞浆Ca^2+浓度变化,Western-blot检测内质网应激及自噬相关蛋白表达水平。结果显示,随着ZEA染毒浓度的增加,TM4细胞胞浆内Ca^2+浓度呈剂量依赖性升高;与ZEA处理组相比,BAPTA-AM与ZEA共处理组胞浆Ca^2+浓度极显著下降(P<0.01),内质网应激相关蛋白BIP、ATF6、ATF4表达量显著或极显著降低(P<0.05或P<0.01),自噬相关蛋白LC3Ⅱ和P62表达量分别呈显著性上升和极显著降低(P<0.05或P<0.01);2-APB与ZEA共处理组胞浆Ca^2+浓度显著下降(P<0.05),内质网应激相关蛋白BIP、ATF6、ATF4、CHOP表达量显著或极显著降低(P<0.05或P<0.01),自噬相关蛋白LC3Ⅱ和P62表达量分别极显著升高和极显著下降(P<0.05或P<0.01)。结果表明,ZEA可以引起TM4细胞内Ca^2+浓度上升,且部分胞内Ca^2+来源于内质网且参与了ZEA诱导TM4细胞内质网应激与自噬。
The study aims to research whether endoplasmic reticulum IP3R calcium channel participates in the process of TM4 cell autophagy induced by zearalenone(ZEA). In this study,the experiment material is mouse sertoli cell line(TM4 cells),which was exposed to different concentrations of ZEA(0,5,10 and 20 μmol/L),BAPTA-AM(Calcium chelator)+ZEA,2-APB(endoplasmic reticulum Ca^2+channel IP3R receptor antagonist)+ZEA,alone and in combination,for 24 hours,cytoplasmic Ca^2+concentration was detected by flow cytometry;and the expression levels of endoplasmic reticulum stress and autophagy associated proteins were detected by Western-blot. In result,the concentration of cytoplasmic Ca^2+in TM4 cells increased with the increase of ZEA concentration. Compared to the ZEA treatment group,in the BAPTA-AM and ZEA co-treatment group,the concentration of Ca^2+in cytoplasm decreased significantly(P <0.01),the expression of endoplasmic reticulum stress associated proteins BIP,ATF6 and ATF4 decreased significantly or extremely significantly(P<0.05 or P<0.01),and the expression of autophagy associated protein LC3 Ⅱ and P62 increased significantly and decreased significantly respectively(P <0.05 or P <0.01);in the 2-APB and ZEA co-treatment group,the concentration of Ca^2+in cytoplasm decreased dramatically(P<0.05),the expression of endoplasmic reticulum stress associated proteins BIP,ATF6,ATF4 and CHOP decreased significantly(P<0.05 or P<0.01),and the expression of autophagy associated protein LC3 Ⅱ and P62 increased significantly and decreased significantly respectively(P <0.05 or P <0.01).The results indicated that ZEA could increase the concentration of Ca^2+in TM4 cells,and some of the intracellular Ca^2+originated from endoplasmic reticulum and participated in ZEA-induced endoplasmic reticulum stress and autophagy in TM4 cells.
作者
郑豪义
冯楠楠
邹辉
顾建红
袁燕
刘学忠
刘宗平
卞建春
ZHENG Hao;FENG Nan-nan;ZOU Hui;GU Jian-hong;YUAN Yan;LIU Xue-zhong;LIU Zong-ping;BIAN Jian-chun(College of Veterinary Medicine.Yangzhou University,Yangzhou 225009,China;Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and,Zoonoses,Yangzhou 225009,China)
出处
《中国兽医科学》
CAS
CSCD
北大核心
2019年第12期1594-1601,共8页
Chinese Veterinary Science
基金
国家重点研发计划项目(2016YFD0501208)
江苏高校优势学科建设工程资助项目(PAPD)
作者简介
郑豪(1995-),男,安徽天长人,硕士生,研究方向为动物营养代谢病与中毒病,E-mail:305858082@99.com;通信作者:卞建春(1965-),男,教授,博士,主要从事动物营养代谢病与中毒病的研究,Tel:0514-87979042,E-mail:jcbian@yzu.edu.cn。
