DDK1阻断Wnt/β-catenin信号通路抑制百草枯诱导的肺成纤维细胞转分化  

Blockade of Wnt/β-catenin signaling pathway by DKK1 inhibits paraquat-induced transition of lung fi broblasts

作　　者：王蒙蒙 杨志洲 任艺 李亮 曹丽萍 孙兆瑞 Wang Mengmeng;Yang Zhizhou;Ren Yi;Li Liang;Cao Liping;Sun Zhaorui(Department of Emergency Medicine,Jinling Hospital,Medical School of Nanjing University(General Hospital of Eastern Theater Command,PLA),Nanjing 210002,China)

机构地区：[1]南京大学医学院附属金陵医院(东部战区总医院)急诊医学科,南京210002

出　　处：《中华急诊医学杂志》2022年第3期310-314,共5页Chinese Journal of Emergency Medicine

基　　金：国家自然科学基金(82102311,81701894),江苏省自然科学基金(BK20190247)。

摘　　要：目的探讨抑制Wnt/β-catenin信号通路对百草枯(paraquat,PQ)诱导的人胚肺成纤维细胞(MRC-5)转分化的影响及相关分子机制。方法将MRC-5细胞分为三组,对照组(Control组):不加药物处理;PQ组:以50μmol/L的PQ刺激MRC-5细胞72 h诱导建立转分化模型;PQ+DKK1组:以50μmol/L的PQ和10 ng/mL的DKK1同时处理细胞72 h。收集细胞,采用细胞免疫荧光和Western Blot分别检测三组细胞转分化标记物α-SMA、Vimentin和Collagen I的表达水平;同时,应用Western Blot、细胞免疫荧光和实时荧光定量PCR(RT-PCR)技术检测转分化过程中Wnt通路相关信号分子β-catenin、Cyclin D1和WISP1的表达变化情况;进一步采用Wnt/β-catenin通路抑制剂Dickkopf-1(DKK1)阻断该信号途径,Western Blot检测β-catenin、Cyclin D1和WISP1表达变化情况,以验证干预效果并分析干扰Wnt/β-catenin信号传递对转分化标记分子α-SMA、Vimentin和Collagen I表达的影响。结果处理72 h后,与对照组细胞相比,PQ组MRC-5细胞α-SMA、Vimentin和Collagen I的表达水平均显著增加(P<0.05);同时,在PQ诱导MRC-5转分化过程中β-catenin、Cyclin D1和WISP1表达水平显著上调(P<0.05);采用DKK1干扰Wnt/β-catenin信号通路能够抑制PQ诱导MRC-5细胞转分化过程中α-SMA、Vimentin和Collagen I的高表达(P<0.05)。结论DKK1能够通过干扰Wnt/β-catenin信号的激活抑制PQ诱导的成纤维细胞转分化,有望进一步抑制PQ中毒引起的肺纤维化的发生。Objective To investigate the effects of inhibition of Wnt/β-catenin signaling pathway on paraquat(PQ)-induced transition of human lung fibroblast cell line MRC-5 and related molecular mechanisms.Methods The MRC-5 cells were divided into three groups.Control group:without drug treatment;PQ group:the cells were treated by PQ(50μmol/L)for 72 hours to establish cell transition model;PQ+DKK1 group:the cells were treated with PQ(50μmol/L)and DKK1(10 ng/mL)for 72 hours.Then,the cells were collected,and the transition related signatures includingα-SMA,Vimentin and Collagen I were detected by immunofl uorescence and Western blot(WB);Meanwhile,the expression levels of Wnt pathway-related molecules includingβ-catenin,Cyclin D1 and WISP1 were determined by WB,immunofl uorescence and real-time fl uorescence quantitative PCR(RT-PCR)during the transition;In addition,the inhibitor of Wnt/β-catenin pathway Dickkopf-1(DKK1)was applied to block the signaling.Then the expression changes ofβ-catenin,cyclin D1 and WISP1 were detected by WB to verify the inhibitory effect,and the transition marker molecules includingα-SMA,Vimentin and Collagen I were also determined by WB.Results After 72 hours,compared with the Control group,the expression levels ofα-SMA,Vimentin and Collagen I of MRC-5 cells in PQ group were increased signifi cantly(P<0.05);The expression levels ofβ-catenin,Cyclin D1 and WISP1 of MRC-5 cells in PQ group were signifi cantly up-regulated(P<0.05);DKK1 could inhibit the high expression ofα-SMA,Vimentin and Collagen I of MRC-5 cells during PQ-induced transition(P<0.05).Conclusions DKK1 could inhibit PQ-induced transition of lung fi broblasts by interference with Wnt/β-catenin signaling,which was expected to further inhibit pulmonary fi brosis caused by PQ.

关 键 词：百草枯 成纤维细胞 转分化 WNT/Β-CATENIN信号通路 抑制剂 

分 类 号：R595.4[医药卫生—内科学]