包虫抗原B通过抑制TAZ促进RANKL/NF-κB/TAK1介导的破骨细胞发生  

Hydatid Antigen B Promote RANKL/NF-κB/TAK1-mediated Osteoclastogenesis via Inhibition of TAZ

作　　者：吾路汗·马汗 谢增如[1] Wuuhan Mahan;XIE Zengru(Department of Trauma and Orthopedics,First Affiliated Hospital of Xinjiang Medical University,China)

机构地区：[1]新疆医科大学第一附属医院创伤骨科,乌鲁木齐市830011

出　　处：《医学分子生物学杂志》2025年第1期8-15,共8页Journal of Medical Molecular Biology

基　　金：新疆维吾尔自治区自然科学基金(No.2021D01D19)。

摘　　要：目的探讨包虫抗原B(hydatid antigen-B,Hyd-B)促进破骨细胞发生的分子机制。方法细胞实验分组-1:BMSCs细胞分为Control组、MCSF+RANKL组和MCSF+RANKL+Hyd-B组;使用巨噬细胞集落刺激因子(macrophage colony-stimulating factor,MCSF)联合可溶性核因子κB受体激活配体(receptor activator of nuclear factor-κb ligand,RANKL),外加/不加Hyd-B联合诱导骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)向破骨细胞分化。细胞实验分组-2:BMSCs细胞分为Ctrl组和Hyd-B处理组(Treat组)。免疫共沉淀(co-IP)法测定Hyd-B对TAZ(transcriptional coactivator with PDZ-binding motif)和TAK1(transforming growth factorβ-activated kinase 1)的直接相互作用的影响,使用抗TAK1抗体进行IP、IB检测TAZ的表达。细胞实验分组-3:BMSCs细胞分为Control组、MCSF+RANKL组、MCSF+RANKL+Hyd-B组、MCSF+RANKL+Hyd-B+TAZ-OE组和MCSF+RANKL+Hyd-B+OE-vector组。BMSCs转染腺病毒-TAZ OE或腺病毒-OE vector介导过表达TAZ。qPCR法检测破骨细胞分化标志物TRAP和Cathepsin K mRNA相对表达水平。蛋白质印迹检测细胞核p-P65、细胞质P65、细胞核NFATc1、p-AKT、AKT、p-ERK 1/2、ERK 1/2、p-TAZ、TAZ和TAK1的表达水平。结果与Control组比较,MCSF+RANKL组和MCSF+RANKL+Hyd-B组细胞TRAP和Cathepsin K mRNA水平升高(P<0.05);p-P65、p-AKT、p-ERK 1/2水平升高(P<0.05);细胞核内p-P65和NFATc1的水平升高(P<0.05)。与MCSF+RANKL组相比较,MCSF+RANKL+Hyd-B组细胞上述指标表达水平进一步升高(P<0.05)。与MCSF+RANKL+Hyd-B组相比较,MCSF+RANKL+Hyd-B+TAZ OE组细胞上述指标表达水平降低(P<0.05)。Co-IP结果,与Ctrl组相比较,Treat组中TAZ与TAK1的相互作用增加(P<0.05)。与Ctrl组相比较,Treat组中细胞质p-TAZ磷酸化水平增加(P<0.05),TAZ表达水平降低(P<0.05)。结论Hyd-B通过抑制TAZ上调RANKL/NF-κB/TAK1介导的破骨细胞发生。Objective To investigate the molecular mechanisms of hydatid antigen-B(Hyd-B)in osteoclastogenesis.Methods Cell experiment group-1:Bone marrow mesenchymal stem cells(BMSCs)were divided into 3 groups(Control group,MCSF+RANKL group,MCSF+RANKL+Hyd-B group),cells in the MCSF+RANKL group and MCSF+RANKL+Hyd-B group were induced to differentiate into osteoclasts by using macrophage colony-stimulating factor(MCSF)combined with soluble receptor activator of nuclear factor-κb ligand(RANKL),and then treated with or without Hyd-B respectively.Cell experiment group-2:BMSCs were divided into Ctrl group and Hyd-B treatment group(Treat group),the effect of Hyd-B on the direct interaction between TAZ and TAK1 was determined by co-immunoprecipitation(co-IP).IP was performed by using anti-TAK1 antibody,and TAZ expression level was detected by IB.Cell experiment group-3:BMSCs were divided into 5 groups:Control group,MCSF+RANKL group,MCSF+RANKL+Hyd-B group,MCSF+RANKL+Hyd-B+TAZ-OE group and MCSF+RANKL+Hyd-B+OE-vector group.TAZ overexpression plasmid or vector plasmid was transfected to the BMSCs in the MCSF+RANKL+Hyd-B+TAZ-OE group and MCSF+RANKL+Hyd-B+OE-vector group respectively.qPCR was used to detect the mRNA expression levels of osteoclast differentiation markers of TRAP and Cathepsin K.Western blotting was used to detect the expression levels of nuclear p-P65,cytoplasmic P65,nuclear NFATc1,p-AKT,AKT,p-ERK 1/2,ERK 1/2,p-TAZ,TAZ and TAK1.Results Compared with those in the Control group,the TRAP and Cathepsin K mRNA expression levels in the MCSF+RANKL and MCSF+RANKL+Hyd-B groups were increased(P<0.05);the expression levels of p-P65,p-AKT and p-ERK 1/2 were increased(P<0.05);the expression levels of p-P65 and NFATc1 in nucleus were increased(P<0.05).The expression levels of the above indicators were further increased in the MCSF+RANKL+Hyd-B group when compared with those in the MCSF+RANKL group(P<0.05).Compared with those in the MCSF+RANKL+Hyd-B group,the expression levels of the above indicators in MCSF+RANKL+Hyd-B+TAZ-OE group we

关 键 词：骨包虫病 包虫抗原B RANKL/NF-κB/TAK1 TAZ 破骨细胞分化 

分 类 号：R532.32[医药卫生—内科学]