高糖刺激的巨噬细胞源外泌体通过激活NF-κB/TGF-β1信号通路促进肾小球纤维化  

Macrophage-derived exosomes from high-glucose stimulated cells promote glomerular fibrosis via NF-κB/TGF-β1 signaling pathway

作　　者：宋惠杰 李阳[1] 谭燚[1] 何旭梅[1] SONG Hui-jie;LI Yang;TAN Yi;HE Xu-mei(Department of Endocrinology,Wuhan First Hospital,Wuhan 430022,Hubei,China)

机构地区：[1]武汉市第一医院内分泌科,湖北武汉430022

出　　处：《广东医学》2024年第11期1391-1396,共6页Guangdong Medical Journal

基　　金：湖北省卫生健康委员会科研项目(20210845)。

摘　　要：目的探讨高糖(HG)处理的巨噬细胞源性外泌体(Exos)是否能激活肾小球系膜细胞和产生促纤维化细胞因子,并探讨其作用机制是否与激活核因子-κB(NF-κB)/转化生长因子-β1(TGF-β1)信号通路有关。方法将高糖处理Raw264.7细胞的外泌体(HG-Exos)和SV40MES13细胞体外孵育24 h。通过EdU和TUNEL实验评估Exos对SV40MES13细胞增殖、凋亡的影响,ELISA法测定上清液中细胞外基质(ECM)的主要成分[Ⅳ型胶原(ColⅣ)和纤维连接蛋白(FN)]。采用定量RT-PCR分析细胞分泌的TGF-β1 mRNA表达,采用Western blot法检测细胞中NF-κB/TGF-β1/Smad3信号通路蛋白表达。结果与NG-Exos相比,HG-Exos处理的SV40MES13细胞分泌ColⅣ、FN蛋白和TGF-β1 mRNA显著增加(P<0.05)。Western blot分析证实,经HG-Exos处理的SV40MES13细胞内TGF-β1、Smad3、p-Smad3、NF-κB p65和p-NF-κB p65蛋白表达较NG-Exos处理显著升高(P<0.05)。当用PDTC(NF-κB特异性抑制剂)与HG-Exos联合处理SV40MES13细胞24 h后,细胞内ColⅣ、FN、TGF-β1、Smad3和p-Smad3蛋白表达较单用HG-Exos明显降低。结论高糖处理的巨噬细胞源性Exos可能通过NF-κB/TGF-β1信号通路激活肾小球系膜细胞,加速肾纤维化。Objective To investigate whether exosomes(Exos)derived from macrophages under high glucose(HG)conditions can activate glomerular mesangial cells and promote the production of pro-fibrotic cytokines,and to explore whether this effect is related to the activation of the nuclear factor kappa-B(NF-κB)/transforming growth factor-β1(TGF-β1)signaling pathway.Methods Exosomes from high glucose-treated Raw264.7 cells(HG-Exos)were incubated with SV40MES13 cells for 24 hours.The effects of Exos on SV40MES13 cell proliferation and apoptosis were assessed using EdU and TUNEL assays.ELISA was used to measure the levels of extracellular matrix(ECM)components[Collagen TypeⅣ(ColⅣ)and fibronectin(FN)]in the supernatant.Quantitative RT-PCR was used to analyze the expression of TGF-β1 mRNA,and Western blotting was performed to detect the protein levels of NF-κB/TGF-β1/Smad3 signaling pathway components.Results Compared to NG-Exos,HG-Exos significantly increased the secretion of Col IV,FN,and TGF-β1 mRNA in SV40MES13 cells(P<0.05).Western blot analysis confirmed that TGF-β1,Smad3,p-Smad3,NF-κB p65,and p-NF-κB p65 protein levels were significantly higher in SV40MES13 cells treated with HG-Exos compared to NG-Exos(P<0.05).Co-treatment with PDTC(an NF-κB specific inhibitor)and HG-Exos for 24 hours significantly reduced the levels of Col IV,FN,TGF-β1,Smad3,and p-Smad3 proteins compared to HG-Exos treatment alone.Conclusion High glucose-treated macrophage-derived exosomes may promote glomerular fibrosis by activating glomerular mesangial cells through the NF-κB/TGF-β1 signaling pathway.

关 键 词：高糖 巨噬细胞 外泌体 肾脏纤维化 NF-κB/TGF-β1信号 

分 类 号：R58[医药卫生—内分泌] R34[医药卫生—内科学]