泡球蚴蛋白通过Wnt/β-Catenin信号通路调控小鼠肝星状细胞二肽基肽酶-4的表达研究  

Regulation of Dipeptidyl Peptidase 4 expression in mouse hepatic stellate cells by Echinococcus multilocularis protein via Wnt/β-Catenin signaling pathway

作　　者：木克西娜·木拉提 努尔拜提·库苏曼 毕晓娟 杨宁 楚瑨 刘辉 房彬彬 孙立 何小龙 高瑾 林仁勇 Mukexina Mulati;Nuerbaiti Kusuman;BI Xiaojuan;YANG Ning;CHU Jin;LIU Hui;FANG Bingbing;SUN Li;HE Xiaolong;GAO Jin;LIN Renyong(Department of Biochemistry and Molecular Biology College of Basic Medicine.Xinjiang Medical University,Urumqi 830011,China;China State Key Laboratory of Pathogenesis,Prevention and Treatment of High Incidence Diseases in Central Asia,Xinjiang Medical University;Clinical Medical Research Institute,the First Affiliated Hospital of Xinjiang Medical University;College of Veterinary Medicine,Xinjiang Agricultural University)

机构地区：[1]新疆医科大学基础医学院生物化学与分子生物学教研室,新疆乌鲁木齐830011 [2]省部共建中亚高发病成因与防治国家重点实验室,新疆医科大学 [3]新疆医科大学第一附属医院临床医学研究院 [4]新疆农业大学动物医学学院

出　　处：《中国病原生物学杂志》2024年第3期275-279,284,共6页Journal of Pathogen Biology

基　　金：国家自然科学基金项目(No.81860361,82060371,32060223);新疆维吾尔自治区自然科学基金项目(No.2022D01D59,2022D01E67);新疆维吾尔自治区“天山英才”培养计划(No.2022TSYCLJ0032);新疆维吾尔自治区重点实验室开放课题项目(No.2020D04028);省部共建中亚高发病成因与防治国家重点实验室开放课题(No.SKL-HIDCA-2021-31)。

摘　　要：目的探讨泡球蚴蛋白对肝星状细胞中纤维化关键基因二肽基肽酶-4(Dipeptidyl Peptidase 4,DPP4)表达的调控机制。方法免疫组织化学法检测二肽基肽酶-4在泡型棘球蚴病患者肝脏病灶近端与远端的表达情况。体外培养小鼠肝星状细胞系JS1,以60μg/mL浓度的泡球蚴蛋白(Echinococcus multilocularis Protein,EmP)刺激JS1细胞,检测细胞活化及Wnt通路的激活情况,Wnt通路抑制剂(IWP-2)和激动剂(LY2090314)靶向干预检测肝星状细胞活化标志物COL1A1、α-SMA与PPARγ,Wnt通路关键分子Wnt5a、β-catenin、GSK3β以及DPP4的表达水平。结果免疫组化检测结果显示泡型棘球蚴病患者肝脏病灶近端DPP4的表达(5927±987.1)显著高于病灶远端(2478±696.9),差异有统计学意义(t=9.026,P<0.05);EmP可显著促进JS1细胞活化并促进DPP4的表达,差异有统计学意义(F_(DPP4)=10.65,P<0.05);与对照组相比,EmP刺激组Wnt通路激活,Wnt5a、β-catenin、p-GSK3β的表达水平上升,差异有统计学意义(WB:F_(Wnt5a)=18.28,Fβ-Catenin=14.98,FP-GSK3β=28.52;RT-qPCR:F_(Wnt5a)=27.29,F_(CTNNB1)=21.24,F_(GSK3β)=7.974,P<0.05);Wnt通路抑制剂IWP-2干预可抑制EmP蛋白对α-SMA、DPP4的表达的促进作用,上调PPARγ的表达,差异有统计学意义(WB:F DPP4=26.27,Fα-SMA=126.4,F_(PPAR-γ)=8.187;RT-qPCR:F_(DPP4)=41.23,F_(ACTA2)=185.2,F_(PPARG)=136.2,P<0.05);与EmP刺激组相比,Wnt通路激动剂可促进JS1细胞的活化及DPP4的表达,差异有统计学意义(WB:F_(DPP4)=151.0;RT-qPCR:F_(DPP4)=29.71,P<0.05)。结论EmP蛋白可通过激活Wnt/β-Catenin通路促进纤维化关键基因DPP4的表达。Objective To determine the regulatory mechanism of Echinococcus multilocularis protein on the expression of Dipeptidyl Peptidase 4(DPP4),a key gene for fibrosis in hepatic stellate cells.Methods Mouse hepatic stellate cell line JS1 was cultured in vitro,and JS1 cells were stimulated with E.multilocularis protein(EmP)concentration of 60μg/mL to detect cell activation and activation of Wnt pathway,the expression of Dipeptidyl Peptidase4 in the closed and distant liver lesions of alveolar echinococcosis patients was detected by immunohistochemistry.Wnt pathway was targeted by Wnt pathway inhibitor(IWP-2)and agonist(LY2090314),and the expression levels of hepatic stellate cell activation markers COL1A1,α-SMA and PPARγ,Wnt pathway key molecules Wnt5a,β-catenin,GSK3βand DPP4 were detected Results EmP could significantly promote the activation of JS1 cells and the expression of DPP4,with statistical significance(F_(DPP4)=10.65,P<0.05),The expression of DPP4 in the closer to the liver lesions(5927±987.1)in alveolar echinococcosis patients was significantly higher than that in the distal end of liver lesions,with statistical significance(2478±696.9),(t=9.026,P<0.05 Compared with the control group,the Wnt pathway was activated in EmP stimulation group,and the expression levels of Wnt5a,β-catenin and P-GSK3βwere increased,with statistical significance(WB:F_(Wnt5a)=18.28,Fβ-Catenin=14.98,FP-GSK3β=28.52;RT-qPCR:F_(Wnt5a)=27.29,F_(CTNNB1)=21.24,F_(GSK3β)=7.974,P<0.05).Wnt pathway inhibitor IWP-2 could inhibit the promoting effect of EmP protein on the expression ofα-SMA and DPP4,and upregulate the expression of PPARγ,with statistical significance(WB:F DPP4=26.27,Fα-SMA=126.4,F_(PPAR-γ)=8.187;RTqPCR:F_(DPP4)=41.23,F_(ACTA2)=185.2,F_(PPARG)=136.2,P<0.05).Compared with EmP stimulation group,Wnt pathway agonists could further promote JS1 cell activation and DPP4 expression,and the difference was statistically significant(WB:F_(DPP4)=151.0;RT-qPCR:F_(DPP4)=29.71,P<0.05).Conclusion EmP can promote the expression of D

关 键 词：泡球蚴 肝纤维化 二肽基肽酶-4 WNT/Β-CATENIN通路 

分 类 号：R383[医药卫生—医学寄生虫学]