间充质干细胞外泌体通过TGF-β信号通路抑制成骨分化

Mesenchymal stem cell-derived exosomes inhibit osteogenic differentiation through TGF-βsignaling pathway

目的 探究间充质干细胞(mesenchymal stem cell,MSC)来源外泌体对MSC成骨分化的影响及其可能的机制。方法 超速离心获得未被地塞米松干预和用地塞米松(10μmol/L)干预的MSC外泌体:Exo-1和Exo-2。将MSC分为3组:对照组、Exo-1组和Exo-2组,成骨诱导分化培养7 d。生化检测碱性磷酸酶(alkaline phosphatase,ALP)活性。流式检测细胞凋亡率。qRT-PCR检测炎症因子及TGF-β信号通路相关基因表达。Western blot检测TGF-β信号通路相关蛋白表达。结果 用地塞米松干预的MSC来源外泌体能够降低MSC细胞中ALP活性,细胞凋亡率升高,下调NF-κB mRNA表达、Smad3和RUNX2 mRNA和蛋白表达(P<0.05),上调IL-1β和TNF-αmRNA表达、TGF-βmRNA和蛋白表达(P<0.05)。结论 用地塞米松(10μmol/L)干预的MSC来源外泌体能通过TGF-β信号通路抑制MSC成骨分化,促进细胞凋亡和炎症反应。

Objective To explore the possible mechanism of the effect of mesenchymal stem cell-derived exosomes on osteogenic differentiation of MSC.Methods Exo-1 and EXO-2 of MSC-derived exosomes without or with dexamethasone(10μmol/L)intervention were obtained by supercentrifugation.MSC was divided into three groups:control group,Exo-1 group,and Exo-2 group.Osteogenic differentiation was induced and cultured for 7 days.Alkaline phosphatase(ALP)activity was determined.The apoptosis rate was detected with flow cytometry.qRT-PCR was used to detect the expression of inflammatory factors and TGF-βsignaling pathway related genes.Western blotting was used to detect the expression of TGF-βsignaling pathway related proteins.Results MSC-derived exosomes treated with dexamethasone reduced ALP activity and increased apoptosis rate by MSC cells.Dexamethasone-treated MSC-derived exosomes down-regulated the expressions of NF-κB mRNA,Smad3,RUNX2 mRNA and protein(P<0.05),and up-regulated the expression of IL-1βand TNF-αmRNA,TGF-βmRNA and protein(P<0.05).Conclusion Dexamethasone-treated(10μmol/L)MSC-derived exosomes inhibit osteogenic differentiation of MSC through TGF-βsignaling pathway and promote apoptosis and inflammation.