人脐带间充质干细胞共培养减轻高糖诱导的足细胞固有免疫反应  

Human umbilical cord mesenchymal stem cells co- culture ameliorates the innate immunity of podocytes induced by high glucose

作　　者：王英惠 吕莎莎[1] 宋燕[1] 程静[1] 刘珊珊 张亚萍 赵龙[1] 王学龄 孙丽娜 柳刚[1] 

机构地区：[1]山东大学第二医院肾内科山东大学肾脏病研究所,济南250033

出　　处：《中华肾脏病杂志》2017年第11期849-855,共7页Chinese Journal of Nephrology

基　　金：国家自然科学基金（81570653）;山东省自然科学基金（ZR2014HM045、2016ZRA13030、ZR2015PH021）;济南市临床医学科技创新计划（201503005）

摘　　要：目的探讨人脐带间充质干细胞（HUC-MSCs）对高糖状态下Toll样受体（TLR）介导的足细胞固有免疫反应的影响。方法将小鼠足细胞分为4组：正常糖浓度组（NG）、甘露醇组（NG＋MA）、高糖浓度组（HG）、高糖浓度下足细胞与HUC—MSCs共培养组（HG＋MSC）。刺激72h后，ELISA方法检测各组培养基上清中炎性因子白细胞介素6（IL-6）、肿瘤坏死因子仅（TNF-α）、TLR配体热休克蛋白70（HSP70）、高迁移率族蛋白1（HMGB1）的表达，实时定量PCR法检测各组细胞表面及细胞提取物中TLR2、TLR4的mRNA表达，Western印迹检测TLR2、TLR4、髓样分化因子88（MyD-88）、磷酸化P65（p-P65）的蛋白表达，免疫荧光染色观察P—P65在足细胞内的定位。结果与NG组比较，HG组炎性因子IL-6、TNF-α、TLR配体HSP70、HMGBl的蛋白表达增多（均P〈0．05），TLR2、TLR4的mRNA和蛋白水平增加（均P〈0．05），MyD-88及P—P65蛋白表达增多（均P〈0．05），免疫荧光染色可见P．P65核内转移。与HG组比较，HG＋MSC组炎性因子IL-6、TNF—α，TLR配体HSP70、HMGB1蛋白表达减少（均P〈0．05），TLR2、TLR4mRNA和蛋白水平减少（均P〈0．05），MyD-88、P—P65的蛋白表达减少（均P〈0．05），免疫荧光染色示p-P65核内转移不明显。结论高糖刺激可使TLR介导的足细胞固有免疫反应增强，HUC．MSCs共培养可通过抑制TLR通路减轻免疫反应。Objective To explore the effects of human umbilical cord mesenchymal stem cells （HUC- MSCs） on the innate immunity of podoeytes mediated by Toll- like receptor （TLR）signaling pathway under high glucose （HG） condition. Methods Podoeytes were divided into four groups according to the treatment： normal glucose group （NG）, mannitol control group （NG＋MA）, high glucose group （HG） and HUC-MSCs co-culture group （HG＋MSC）. After 72 hours treatment, the protein levels of interleukin- 6 （IL- 6）, tumor necrosis factor- α （TNF- α）, heat shock protein 70 （HSP70）, highmobility group box-1 （HMGB1）in culture medium were measured by ELISA. Real-time PCR was used to detect the mRNA expressions of TLR2 and TLR4. Western blotting was used to detect the protein expressions of TLR2, TLR4, myeloid differentiation factor 88 （MyD- 88） and phospho- P65 （p-P65）. Immunofluorescence staining was used to study the localization of p- P65 in podoeytes. Results High glucose induced the inflammation of podoeytes by activating the TLR signaling, which increased the secretion of IL-6, TNF-α, HSP70, HMGB1, the mRNA level of TLR2, TLR4 and the protein level of TLR2, TLR4, MyD-88 and p-P65 （all P 〈 0.05）. High glucose also activated NF-κB and induced its nuclear translocation. HUC- MSCs co- cuhure decreased the inflammation and restrained the TLR signaling. Conclusions HUC-MSCs co-culture decreases the inflammation and innate immunity of podocytes induced by HG.

关 键 词：糖尿病肾病 足细胞 干细胞 间充质 固有免疫 TOLL样受体 

分 类 号：R587.2[医药卫生—内分泌] R692.9[医药卫生—内科学]