原花青素下调wnt/β-catenin通路拮抗砷诱导的人肝细胞L-02增殖  

Effects of proanthocyanidins down-regulating wnt/β-catenin signaling pathway to antagonize arsenic-induced proliferation of human hepatocyte L-02

作　　者：任庆新 张卫兵[1] 黄建萍[1] 安娜[1] 柴婧 REN Qing-xin;ZHANG Wei-bing;HUANG Jian-ping;AN Na;CHAI Jing(Nantong Center for Disease Control and Prevention,Nantong,Jiangsu 226007,China;不详)

机构地区：[1]南通市疾病预防控制中心,江苏南通226007 [2]乌鲁木齐市疾病预防控制中心

出　　处：《现代预防医学》2024年第8期1486-1492,1529,共8页Modern Preventive Medicine

基　　金：国家自然科学基金项目(81760584)。

摘　　要：目的 本次研究使用原花青素(Proanthocyanidins, PC)和亚砷酸钠(Sodium arsenite, NaAsO2)对人肝细胞L-02进行干预,探究砷致细胞增殖的效应,PC对砷作用的拮抗以及与wnt/β-catenin信号通路的关系。方法 以不同剂量的PC单独或与NaAsO2联合干预细胞,利用细胞计数试剂盒-8(Cell Counting Kit-8, CCK-8)法检测细胞增殖活力,使用细胞划痕实验、Transwell实验检测细胞的迁移和侵袭能力,使用实时聚合酶链反应(Real-time Polymerase Chain Reaction, Real-time PCR)法和免疫印迹(Western Blotting, WB)法检测相关因子的信使RNA(message RNA, mRNA)和蛋白表达水平。结果 NaAsO2致细胞增殖活性(1.740±0.105)升高,迁移(556.000±3.606)、侵袭(439.667±8.622)能力增强,上调原癌基因c-myc(0.739±0.012)、细胞周期蛋白D1(cyclinD1)(0.943±0.002)、血管内皮生长因子(Vascular endothelial growth factor, VEGF)(1.020±0.019)、凋亡抑制基因survivin(0.571±0.033)和基质金属蛋白酶-7(Matrix metalloproteinase-7, MMP-7)(0.695±0.059)的蛋白表达水平,上调wnt3a和β-连环蛋白(β-catenin)的蛋白(分别为1.044±0.010, 0.958±0.037)和mRNA(分别为1.789±0.165, 1.596±0.217)水平。当PC与NaAsO2联合处理后,相较于NaAsO2单独干预,c-myc(0.438±0.046)等增殖相关蛋白及信号通路核心分子wnt3a(0.822±0.015)、β-catenin(0.832±0.064)表达水平下降,通路抑制因子糖原合成酶激酶-3β(Glycogen synthase kinase-3β, GSK-3β)(0.885±0.058)和轴蛋白(Axin)(0.749±0.016)表达升高。结论 NaAsO2诱导L-02细胞增殖,这可能是由其激活了wnt/β-catenin信号通路导致的;PC可以下调wnt/β-catenin通路,进而拮抗NaAsO2所致的细胞增殖。Objective To explore the effect of arsenic on cell proliferation,the antagonism of PC against arsenic and the relationship associated to wnt/β-catenin signaling pathway by using Proanthocyanidins(PC)and sodium arsenite(NaAsO2)to intervene human hepatocyte L-02.Methods We treated cells with different doses of PC alone or in combination with NaAsO2.The cell activities were detected by Cell Counting Kit-8(CCK-8)method.Cell scratch experiment and Transwell experiment were used to detect the ability of cell migration and invasion.Real-time Polymerase Chain Reaction(Real-time PCR)method and western blotting(WB)method were used to detect the expression levels of message RNA(mRNA)and protein.Results NaAsO2 increased the activity of cell proliferation(1.740±0.105),enhanced the ability of migration(556.000±3.606)and invasion(439.667±8.622).NaAsO2 up-regulated the protein expression of proto-oncogene c-myc(0.739±0.012),cyclinD1(0.943±0.002),Vascular Endothelial Growth Factor(VEGF)(1.020±0.019),apoptosis inhibitor gene survivin(0.571±0.033)and Matrix Metalloproteinase-7(MMP-7)(0.695±0.059).NaAsO2 could also up-regulate the protein and mRNA levels of pathway core moleculars wnt3a(protein:1.044±0.010,mRNA:1.789±0.165)andβ-catenin(protein:0.958±0.037,mRNA:1.596±0.217).Compared with NaAsO2 group,when PC intervention was introduced,the levels of proliferation related proteins such as c-myc(0.438±0.046)decreased.The levels of mRNA and protein of wnt3a(mRNA:1.181±0.018,protein:0.822±0.015)andβ-catenin(mRNA:0.965±0.078,protein:0.832±0.064)decreased.The expression of pathway inhibitors Glycogen synthase kinase-3β(GSK-3β)(0.885±0.058)and Axin(0.749±0.016)increased.Conclusion NaAsO2 induce L-02 cell proliferation,which may be due to its activation of wnt/β-catenin signaling pathway.PC can down-regulate wnt/β-catenin pathway,thereby antagonizing the cell proliferation induced by NaAsO2.

关 键 词：砷 原花青素 增殖 WNT/Β-CATENIN信号通路 

分 类 号：R113[医药卫生—公共卫生与预防医学]