IL-6经NF-κB信号通路上调人胎盘MSC SPP1表达促进M2型巨噬细胞极化的作用机制研究  

IL-6 upregulates the expression of human placental MSC SPP1 through the NF-κB signaling pathway and promotes M2 macrophage polarization

作　　者：潘琳 蔡睿志 陶金 李莉 马淑琴 叶鹏 朱永朝 PAN Lin;CAI Ruizhi;TAO Jin;LI Li;MA Shuqin;YE Peng;ZHU Yongzhao(Medical Experiment Center,General Hospital of Ningxia Medical University,Yinchuan 750004,China)

机构地区：[1]宁夏医科大学总医院医学实验中心,宁夏银川750004

出　　处：《细胞与分子免疫学杂志》2024年第11期961-967,共7页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(82060020);宁夏回族自治区重点研发项目(2022BEG03088,2021BEG02032);宁夏自然科学基金(2021AAC03335,2022AAC03539)。

摘　　要：目的探究白细胞介素6(IL-6)调控人胎盘来源间充质干细胞(MSC)分泌型磷酸蛋白1(SPP1)表达,影响巨噬细胞极化的分子机制。方法利用酶消化法及成分明确的无血清培养基制备出人胎盘来源间充质干细胞(hfPMSC),形态学观察,流式细胞仪检测MSC表面标志分子CD14、CD34、CD45、CD73、CD90、CD105、人类白细胞抗原DR(HLA-DR)的表达;利用终剂量100 ng/mL的IL-6处理hfPMSC 24 h,ELISA、Western blot法和实时定量PCR在蛋白和mRNA表达水平上检测SPP1的表达;将IL-6处理感染SPP1干扰慢病毒的hfPMSC与80 ng/mL佛波酯(PMA)联合100 ng/mL脂多糖(LPS)诱导活化的THP-1巨噬细胞共培养,流式细胞术检测THP-1细胞CD11c和CD206的阳性细胞比例;利用IL-6和/或核因子κB p65(NF-κB p65)特异性抑制剂SC75741处理hfPMSC,Western blot法检测NF-κB信号通路与SPP1的表达。结果IL-6显著上调hfPMSC SPP1蛋白及mRNA的表达水平;干扰SPP1显著下调hfPMSC SPP1的表达,显著降低共培养巨噬细胞CD206阳性的细胞比率;IL-6显著上调hfPMSC p-p65的表达,激活NF-κB信号通路;SC75741显著下调经IL-6处理hfPMSC中p-p65和SPP1的蛋白表达水平。结论IL-6经NF-κB信号通路上调hfPMSC SPP1的表达,增强其诱导巨噬细胞向M2型极化的作用。Objective To investigate the molecular mechanism of IL-6 regulating the expression of secretory phosphoprotein 1(SPP1)in human placental-derived mesenchymal stem cells(hfPMSCs)and influencing the polarization of macrophages.Methods hfPMSCs were prepared by enzymatic digestion and cultured in a defined,serum-free medium.The morphology of hfPMSCs was observed under a microscope,and the expression of surface markers CD14,CD34,CD45,CD73,CD90,CD105,and human leukocyte antigen DR(HLA-DR)was detected by flow cytometry.hfPMSCs were treated with IL-6 at the final dosage of 100 ng/mL for 24 hours.ELISA,Western blot,and real-time quantitative PCR were used to detect the expression of SPP1 at the protein and mRNA levels;after being treated with IL-6,hfPMSCs infected with SPP1 interference lentivirus were co-cultured with activated macrophage THP-1 cells induced by 100 ng/mL lipopolysaccharide(LPS)and 80 ng/mL phorbol 12-myristate 13-acetate(PMA),and flow cytometry was used to detect the proportion of CD11c and CD206 positive cells in THP-1 cells;hfPMSCs were treated with IL-6 and/or specific inhibitors of nuclear factor kappa B p65(NF-κB p65),SC75741,and Western blot was used to detect the expression of genes in the NF-κB signaling pathway and SPP1.Results IL-6 significantly upregulates the expression of SPP1 in protein and mRNA levels in hfPMSCs;interference of SPP1 significantly downregulates the expression of SPP1 in hfPMSCs and significantly reduces the positive cell ratio of CD206 in co-cultured macrophages;IL-6 significantly upregulates the expression of p-p65 in hfPMSCs,activating the NF-κB signaling pathway;SC75741 significantly downregulates the expression of p-p65 and SPP1 in hfPMSCs treated with IL-6.Conclusion IL-6 upregulates the expression of SPP1 through the NF-κB signaling pathway in hfPMSCs,which enhances the capability to induce macrophages to polarize towards the M2 phenotype.

关 键 词：人胎盘来源间充质干细胞(hfPMSC) IL-6 SPP1 巨噬细胞极化 

分 类 号：R392[医药卫生—免疫学] R364.5[医药卫生—基础医学] R392.12R392.11