ZEA和DON及其联合作用对小鼠T淋巴细胞Fas/FasL凋亡信号通路的影响  

Effects of ZEA,DON and their combination on apoptosis of mouse T lymphocytes via Fas/FasL signaling pathway in vitro

作　　者：夏苏干 孙凯[1] 蔡国栋[1] 邹辉[1] 顾建红[1] 袁燕[1] 刘学忠[1] 刘宗平[1,2] 卞建春 XIA Su-gan;SUN Kai;CAI Guo-dong;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 Coinnovation Centerfor Prevention and Control of Important Animal Infectious Diseases and Zoonoses , Yangzhou 225009, China)

机构地区：[1]扬州大学兽医学院,江苏扬州225009 [2]江苏省动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《中国兽医科学》2019年第6期781-788,共8页Chinese Veterinary Science

基　　金：国家重点研发计划支持项目(2016YFD0501208);江苏高校优势学科建设工程资助项目(PAPD)

摘　　要：为揭示玉米赤霉烯酮(ZEA)、脱氧雪腐镰刀菌烯醇(DON)及其联合作用免疫毒性的机理,试验研究了ZEA、DON及联合作用对刀豆蛋白A(Con A)刺激的小鼠离体T淋巴细胞细胞凋亡和Fas/FasL凋亡信号通路的影响。从BALB/c小鼠脾分离原代T淋巴细胞,分别设细胞空白组、刀豆蛋白A对照组(Con A组)、Con A+不同浓度ZEA染毒组、Con A+不同浓度DON染毒组、Con A+不同浓度ZEA与DON联合染毒组,染毒时间为24 h,利用流式细胞术检测细胞凋亡率,Western-blot检测Fas、FasL、Cleaved-caspase8、Bax、Bcl-2等关键蛋白表达情况。结果显示,与细胞空白组相比,Con A组细胞凋亡率上升但差异不显著(P>0.05);与Con A组相比,各染毒组细胞凋亡率均上升,多组出现显著或极显著差异(P<0.05或P<0.01);联合处理组较单一毒素处理组细胞凋亡率升高,经CompuSyn软件拟合,ZEA和DON以20∶1联合作用时协同效应最为明显。Con A组与细胞空白组相比,Bax/Bal-2值显著降低(P<0.05)、Fas、FasL、Cleaved-caspase8等蛋白的表达量无明显变化(P>0.05);与Con A对照组相比,各染毒组Bax/Bal-2、Fas、FasL、Cleaved-caspase8等蛋白的表达量均上升,且呈剂量依赖性,多组出现显著或极显著差异(P<0.05或P<0.01);联合染毒组较单一染毒组各蛋白表达量均升高。结果表明, ZEA、DON均可通过Fas/FasL凋亡信号通路诱导T细胞的凋亡,ZEA与DON联合可发挥协同效应。In order to reveal the immunotoxic mechanism of zearalenone(ZEA),deoxynivalenol(DON)and their combination,we studied the effects of ZEA,DON and their combination on the apoptosis of mouse T lymphocytes stimulated by concanavalin A(Con A) in vitro and Fas/Fas L apoptotic signaling pathway. T lymphocytes were isolated from the spleen of BALB/c mice. Cell control group(cell only),Con A group,Con A group with different concentrations of DON or ZEA,and Con A group with different concentrations of ZEA and DON were set up. After being exposed for 24 h,the apoptosis rate was detected by flow cytometry. The expression of key protein,Fas,Fas L,Cleaved-caspase8,Bax and Bcl-2 were detected by Western-blot.In result,compared with the cell control group,the apoptosis rate of the Con A group was increased,but the difference was not significant(P>0.05).Compared with the Con A group,the apoptotic rate of each toxin group increased. Many of them showed significant or extremely significant differences(P<0.05 or P<0.01).The apoptotic rate of the combined treatment group was higher than the single toxin treatment groups. By the Compu Syn software fitting,the synergistic effect of ZEA and DON combined with 20 ∶ 1 ratio was most obvious. Compared with the cell control group,the ratio of Bax/Bal-2 was significantly decreased in the Con A group(P<0.05).While the expression of Fas,Fas L,and Cleaved-caspase8 was not significantly changed(P>0.05).Compared with the Con A group,the expression of Bax/Bal-2,Fas,Fas L,and Cleaved-caspase8 increased in a dose-dependent manner in the toxin groups. Significant or extremely significant differences were found in many of them(P<0.05 or P<0.01).The expression of each protein in the combined exposure group was higher than that in the single exposure group. In conclusion, Both ZEA and DON can induce T cell apoptosis through Fas/Fas L apoptosis signaling pathway, and the combination of ZEA and DON could induce a synergistic effect.

关 键 词：玉米赤霉烯酮 脱氧雪腐镰刀菌烯醇 T细胞 凋亡 Fas/FasL信号通路 

分 类 号：S859.8[农业科学—临床兽医学]