硬脂酸通过CD36影响酮病奶牛CD4^(+)T细胞IL-17的表达  

Stearic acid affects the expression of IL-17 in CD4^(+)T cells from ketosis cows through CD36

作　　者：季子崴 李思瑶 张海鑫 李紫薇 张上明珠 杨威[2] 徐闯 张冰冰[1] JI Ziwei;LI Siyao;ZHANG Haixin;LI Ziwei;ZHANG Shangmingzhu;YANG Wei;XU Chuang;ZHANG Bingbing(College of Life Science and Technology,Heilongjiang Bayi Agricultural University,Daqing,Heilongjiang 163319,China;College of Animal Science and Veterinary Medicine,Heilongjiang Bayi Agricultural University,Daqing,Heilongjiang 163319,China;College of Veterinary Medicine,China Agricultural University,Beijing 10019l,China)

机构地区：[1]黑龙江八一农垦大学生命科学技术学院,黑龙江大庆163319 [2]黑龙江八一农垦大学动物科技学院,黑龙江大庆163319 [3]中国农业大学动物医学院,北京100191

出　　处：《中国兽医学报》2025年第3期602-610,共9页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(32172926)。

摘　　要：采集健康、酮病奶牛外周血液并分离出CD4^(+)T细胞,Western blot检测脂质合成相关蛋白脂肪酸合成酶(fatty acid synthase,FASN)、乙酰辅酶A羧化酶1(acetyl coenzyme A carboxylase 1,ACC1)、白细胞分化抗原36(cluster of differentiation 36,CD36)及钙库操纵性钙离子内流(store-operated calcium entry,SOCE)相关蛋白ORAIl、ORAI2、ORAI3、STIM1、STIM2表达;流式细胞术检测IL-17表达。从1日龄犊牛脾脏分离CD4^(+)T细胞用于体外培养,细胞处理分为阴性对照siRNA处理组(Ctrl组)、沉默CD36(siCD36)组、硬脂酸(stearic acid,SA)组、SA+siCD36组。使用75 pmol/L阴性对照siRNA转染Ctrl组和SA组细胞48 h,然后用200μmol/L SA刺激SA组细胞24 h;使用75 pmol/L CD36 siRNA转染siCD36组和SA+siCD36组细胞48 h,然后用200μmol/L SA刺激SA+siCD36组细胞24 h。Western blot检测FASN、ACC1、CD36、钙释放激活钙调节因子1(calucium release-activated calcium channel protein 1,CRCM1,也称ORAI1),ORAI2、ORAI3、基质相互作用分子1(slromal interaction molecule 1,STIM1)、STIM2蛋白表达;流式细胞术检测IL-17表达。结果显示,与健康奶牛相比,酮病奶牛外周血CD4^(+)T细胞中IL-17表达显著升高(P<0.01);并上调了FASN、CD36、STIM1(P<0.05)及ACC1、ORAI2、ORAI3、STIM2的蛋白水平(P<0.01)。体外试验结果显示,相比于Ctrl组,SA组上调了CD36、ACC1、ORAI3(P<0.05)以及FASN、STIM1的蛋白表达(P<0.01),IL-17表达显著升高(P<0.05);相比于SA组,SA+siCD36组下调了STIM1、ORAI1(P<0.05)及CD36、ACC1、FASN、ORAI2的蛋白表达(P<0.01),IL-17表达降低(P<0.05)。结果表明,SA通过CD36促进酮病奶牛CD4^(+)T细胞脂质合成并激活SOCE通道,促进IL-17表达。The peripheral blood of healthy or ketosis dairy cows was collected,and CD4^(+)T cells were isolated.The expressions of lipid synthesis related proteins fatty acid synthase(FASN),acetyl coenzyme A carboxylase 1(ACC1),cluster of differentiation 36(CD36)and store-operated calcium entry(SOCE)related proteins ORAI1,ORAI2,ORAI3,STIM1,STIM2 were detected by Western blot.IL-17 cells were detected by flow cytometry.CD4^(+)T cells were isolated from the spleen of 1-day-old calves and cultured in vitro.Cells were treated and divided into control(Ctrl)group,silenced CD36(siCD36)group,stearic acid(SA)group,and SA+siCD36 group.Cells in the Ctrl and SA groups were transfected with 75 pmol/L negative control siRNA for 48 h,and then stimulated with 200μmol/L SA for 24 h;Cells in the siCD36 group and SA+siCD36 group were transfected with 75 pmol/L CD36 siRNA for 48 h,and then stimulated with 200μmol/L SA for 24 h in the SA+siCD36 group.The protein expression of FASN,CD36,ACC1,ORAI1,ORAI2,ORAI3,STIM1 and STIM2 was detected by Western blot,and IL-17 cells were detected by flow cytometry.The results showed that the expression of IL-17 in peripheral blood CD4^(+)T cells of ketosis dairy cows was significantly increased compared to that of healthy cows(P<0.01).Additionally,the protein level of FASN,CD36,STIM1(P<0.05),and ACC1,ORAI2,ORAI3,STIM2(P<0.01)were up-regulated.Compared with the Ctrl group,the protein expression levels of CD36,ACC1 and ORAI3(P<0.05)were up-regulated in the SA group,as well as the protein expression of FASN and STIM1(P<0.01).Additionally,the expression of IL-17 was significantly increased(P<0.05).Compared with the SA group,there was a decrease in the protein expression of STIM1,ORAI1(P<0.05)and CD36,ACC1,FASN,ORAI2(P<0.01)in the siCD36+SA group,as well as IL-17(P<0.05).These results suggest that SA can promote the expression of IL-17 in CD4^(+)T cells in ketosis cows by regulating fatty acid synthesis and activating SOCE channels through CD36.

关 键 词：硬脂酸 CD36 CD4^(+)T细胞 IL-17 酮病 奶牛 

分 类 号：S852.3[农业科学—基础兽医学] S856[农业科学—兽医学]