黄芪多糖诱导SOCS3表达对鸡巨噬细胞炎症反应的抑制作用  

Inhibitory Effect of Astragalus Polysaccharide on Inflammatory Response of Chicken Macrophages by Inducing SOCS3 Expression

作　　者：陶新磊 刘丹华 田旭 郑世民[1] 高雪丽[1] 吕晓萍[1] 刘超男[1] TAO Xinlei;LIU Danhua;TIAN Xu;ZHENG Shimin;GAO Xueli;LYU Xiaoping;LIU Chaonan(Heilongjiang Key Laboratory for Laboratory Animals and Comparative Medicine,College of Veterinary Medicine, Northeast Agricultural University,Harbin 150030,China;CTI Biotechnology(Suzhou)Co.,Ltd.,Suzhou 215300,China)

机构地区：[1]东北农业大学动物医学学院,黑龙江省实验动物与比较医学重点实验室,哈尔滨150030 [2]苏州华测技术有限公司,苏州215300

出　　处：《中国畜牧兽医》2021年第11期4284-4291,共8页China Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(31472169);黑龙江省教育厅科学技术研究项目基金(12511029)。

摘　　要：试验旨在探究黄芪多糖(Astragalus polysaccharide,APS)对LPS诱导的鸡巨噬细胞(HD11)炎症模型的抗炎效果和作用机制。用不同浓度的LPS刺激鸡巨噬细胞(HD11),通过CCK8法检测细胞活力,实时荧光定量PCR检测白细胞介素-1β(IL-1β)mRNA表达的变化,以确定构建细胞炎症模型的LPS最适添加浓度。将HD11分为对照组(C)、脂多糖(LPS)组(L)、黄芪多糖(APS)组(A)和黄芪多糖抑制脂多糖组(A+L),在LPS刺激后的2、6、12、24、48 h用实时荧光定量PCR法检测IL-1β、肿瘤坏死因子-α(TNF-α)、核转录因子(NF-κBp65)、p38丝裂原活化蛋白激酶(p38MAPK)和细胞因子信号转导抑制因子3(SOCS3)mRNA表达的变化,Western blotting法检测NF-κBp65、p38MAPK和SOCS3蛋白含量变化,ELISA检测IL-1β和TNF-α蛋白含量变化。细胞活力和IL-1β检测结果表明,构建细胞炎症模型的LPS最适添加浓度为0.5μg/mL。实时荧光定量PCR结果表明,与对照组相比,L和A组IL-1β、TNF-α、NF-κBp65、p38MAPK和SOCS3 mRNA表达均显著升高(P<0.05);与L组相比,A+L组在LPS刺激后的IL-1β、TNF-α、NF-κBp65和p38MAPK mRNA表达含量均显著降低(P<0.05),SOCS3 mRNA表达显著增加(P<0.05)。Western blotting结果表明,与对照组相比,A组P-NF-κBp65/NF-κBp65、P-p38MAPK/p38MAPK和SOCS3/α-tubulin比值均显著增加(P<0.05);与L组相比,A+L组P-NF-κBp65/NF-κBp65和P-p38MAPK/p38MAPK比值均显著降低(P<0.05);A+L组SOCS3/α-tubulin比值显著升高(P<0.05)。ELISA结果表明,与对照组相比,L组IL-1β和TNF-α蛋白含量在2~48 h均显著增加(P<0.05);与L组相比,A+L组IL-1β和TNF-α的蛋白含量在LPS刺激后12~48 h均显著降低(P<0.05)。以上结果表明,在LPS诱导的HD11细胞炎症模型中,APS通过促进SOCS3的表达抑制NF-κBp65和p38MAPK信号通路的过度活化,从而发挥抗炎作用。The aim of this study was to investigate the anti-inflammatory effect and action mechanism of Astragalus polysaccharide(APS)on LPS-induced chicken macrophage(HD11)inflammation model.HD11 cells were stimulated with different concentrations of LPS.Cell viability was detected by CCK8 assay and interleukin-1β(IL-1β)mRNA expression was detected by Real-time quantitative PCR in order to determine the optimal concentration of LPS for the construction of cellular inflammation model.HD11 cells were divided into control group(C),lipopolysaccharide(LPS)group(L),APS group(A)and APS inhibited LPS group(A+L).The mRNA expressions of IL-1β,tumor necrosis factor-α(TNF-α),NF-κBp65,p38MAPK and suppresser of cytokine signaling 3(SOCS3)were detected by Real-time quantitative PCR,the protein contents of NF-κBp65,p38MAPK and SOCS3 were detected by Western blotting,and the protein contents of IL-1βand TNF-αwere detected by ELISA at 2,6,12,24 and 48 h after LPS stimulation.The results of cell viability and IL-1βdetection showed that the optimal addition concentration of LPS for construction of cellular inflammation model was 0.5μg/mL LPS.Real-time quantitative PCR results showed that compared with control group,the mRNA expressions of IL-1β,TNF-α,NF-κBp65,p38MAPK and SOCS3 in L and A groups were significantly increased(P<0.05).Compared with L group,the mRNA expression levels of IL-1β,TNF-α,NF-κBp65 and p38MAPK in A+L group were significantly decreased(P<0.05),and the mRNA expression level of SOCS3 was significantly increased(P<0.05).Western blotting results showed that compared with control group,the ratios of P-NF-κBp65/NF-κBp65,P-p38MAPK/p38MAPK and SOCS3/α-tubulin in A group were significantly increased(P<0.05).Compared with L group,the ratios of P-NF-κBp65/NF-κBp65 and P-p38MAPK/p38MAPK in A+L group were significantly decreased(P<0.05),and SOCS3/α-tubulin ratio in A+L group was significantly increased(P<0.05).ELISA results showed that compared with control group,the protein contents of IL-1βand TNF-αin L grou

关 键 词：黄芪多糖 抗炎 细胞因子信号转导抑制因子3(SOCS3) 

分 类 号：S831.7[农业科学—畜牧学]