内质网应激预适应对LPS诱导的山羊子宫内膜上皮细胞炎性反应的保护作用  

Protective Effect of Endoplasmic Reticulum Stress Preadaptation on LPS-Induced Inflammatory Response in Goat Endometrial Epithelial Cells

作　　者：郜康康 扆妍妍 赵一腾 林鹏飞[1,2] 陈华涛 靳亚平[1,2] GAO Kangkang;YI Yanyan;ZHAO Yiteng;LIN Pengfei;CHEN Huatao;JIN Yaping(College of Veterinary Medicine,Northwest A&F University,Yangling,Shaanxi 712100,China;Key Laboratory of Animal Biotechnology of the Ministry of Agriculture and Rural Affairs,Yangling,Shaanxi 712100,China)

机构地区：[1]西北农林科技大学动物医学院,杨凌712100 [2]西北农林科技大学农业农村部动物生物技术重点实验室,杨凌712100

出　　处：《畜牧兽医学报》2023年第8期3546-3556,共11页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：宁夏回族自治区科技厅科技项目(201802060004);科技部高端外国专家引进计划(G0222172033L)。

摘　　要：探究内质网应激(endoplasmic reticulum stress,ERS)在脂多糖(lipopolysaccharide,LPS)诱导的山羊子宫内膜上皮细胞(goat endometrial epithelial cells,gEECs)炎性反应中的作用,为阐明ERS在反刍动物子宫内膜炎中的作用机制提供前期基础。本研究使用ERS激活剂衣霉素(tunicamycin,TM)和抑制剂4-苯基丁酸(4-phenylbutyric acid,4-PBA)分别单独处理gEECs激活或抑制ERS,应用RT-qPCR和Western blot技术检测ERS对炎症相关基因表达的影响;接着,使用TM和4-PBA分别预处理gEECs,再利用大肠杆菌LPS处理诱导gEECs炎性反应,通过RT-qPCR和Western blot检测ERS对LPS诱导的gEECs炎性反应的影响及作用机制。结果显示,TM处理激活ERS显著抑制gEECs中炎性细胞因子IL-6和TNF-αmRNA的表达(P<0.05),显著抑制经典炎症通路相关基因TLR4、NF-κB P65和NLRP3 mRNA的表达(P<0.01);同时,显著抑制NF-κB P65蛋白的磷酸化以及NLRP3蛋白的表达(P<0.05)。4-PBA处理则显著促进gEECs中炎症相关基因(IL-6、TNF-α、TLR4、NF-κB P65和NLRP3)的表达(P<0.05)。进一步研究发现,TM预处理gEECs显著抑制LPS诱导的炎性细胞因子IL-6 mRNA的表达(P<0.05);显著抑制LPS诱导的TLR4、NF-κB P65和NLRP3 mRNA的表达以及NF-κB P65蛋白的磷酸化和NLRP3蛋白的表达(P<0.05)。4-PBA预处理gEECs则显著促进LPS诱导的gEECs炎性反应(P<0.05)。综上表明,ERS通过抑制NF-κB通路和NLRP3炎性小体通路的激活减轻LPS诱导的gEECs炎性反应,为进一步阐明山羊子宫内膜炎的发生发展提供了前期理论基础。We attempted to investigate the role of endoplasmic reticulum stress(ERS)in lipopolysaccharide(LPS)-induced inflammatory responses in goat endometrial epithelial cells(gEECs)and provide a preliminary basis for elucidating the mechanism of ERS in ruminant endometritis.In this study,gEECs were treated with ERS activator tunicamycin(TM)and inhibitor 4-phenylbutyric acid(4-PBA)separately to activate and inhibit ERS,and RT-qPCR and Western blot techniques were used to detect the effect of ERS on inflammation-related genes expression.Next,gEECs were pretreated with TM and 4-PBA,respectively,and then treated with E.coli LPS to induce inflammatory responses in gEECs,the effects and mechanisms of ERS on the inflammatory responses induced by LPS in gEECs were detected by RT-qPCR and Western blot.(Results)The results showed that activation of ERS by TM treatment significantly inhibited the expression of inflammatory cytokines IL-6 and TNF-αmRNA in gEECs(P<0.05).The mRNA expression levels of classical inflammatory pathway-related genes TLR4,NF-κB P65,and NLRP3 was also significantly suppressed(P<0.01).Meanwhile,it also significantly inhibited the phosphorylation of NF-κB P65 protein and the expression of NLRP3 protein(P<0.05).However,4-PBA treatment significantly promoted the expression of inflammation-related genes(IL-6,TNF-α,TLR4,NF-κB P 65,and NLRP3)in gEECs(P<0.05).Further study revealed that TM pretreatment of gEECs significantly inhibited the expression of LPS-induced inflammatory cytokine IL-6 mRNA(P<0.05).It also significantly inhibited LPS-induced TLR4,NF-κB P 65,and NLRP 3 mRNA expression as well as phosphorylated NF-κB P65 protein and NLRP3 protein(P<0.05).4-PBA pretreatment of gEECs then significantly promoted the LPS-induced inflammatory response in gEECs(P<0.05).(Conclusion)The above results suggest that ERS alleviates LPS-induced inflammatory response in gEECs by inhibiting the activation of NF-κB pathway and NLRP3 inflammatory vesicle pathway.These results provide a pre-theoretical basis for further e

关 键 词：内质网应激 子宫内膜炎 脂多糖 NF-ΚB通路 NLRP3炎性小体 

分 类 号：S857.2[农业科学—临床兽医学]