健脾解毒方通过Wnt/β-catenin信号通路下调幽门螺杆菌诱导的胃癌血管生成因子的研究  

Jianpi Jiedu Recipe down-regulates angiogenic factors in Helicobacter pylori-induced gastric cancer cells by inhibiting Wnt/β-catenin signaling pathway

作　　者：刘宁宁[1] 周利红[1] 孙筱婷 李春浦 李琦[1] 

机构地区：[1]上海中医药大学附属曙光医院肿瘤科上海中医药大学附属曙光医院中西医结合肿瘤研究所,上海201203

出　　处：《上海中医药杂志》2017年第10期84-88,共5页Shanghai Journal of Traditional Chinese Medicine

基　　金：国家自然科学基金资助项目(81202663;81273958);上海市自然科学基金资助项目(12ZR1449300);上海市卫生局资助项目(20134309);上海市领军人才;上海市医学领军人才;上海市杏林新星计划(ZY3-RCPY-2-2006);上海市浦东新区名中医工作室建设项目(PDZYXK-3-2014001;PDZYXK-6-2014017)

摘　　要：目的研究健脾解毒方对幽门螺杆菌(HP)感染胃癌细胞诱导的血管生成因子(VEGF)、血管生成素-2(Ang-2)、成纤维细胞生长因子(b FGF)的影响,并探讨其可能的机制。方法采用HP标准株NCTC11637与人胃癌MKN45细胞共培养的方法感染MKN45细胞,分为对照组、HP感染组、健脾解毒方醇提物低中高剂量组(低中高剂量分别为0.5 g/L、1.0 g/L和2.5 g/L),采用ELISA法检测健脾解毒方对HP感染诱导的人胃癌MKN45细胞VEGF、b FGF、Ang-2表达的影响;使用Wnt/β-catenin特异性抑制剂FH-535(20μmol/L)抑制Wnt/β-catenin活性后,再进行HP感染,ELISA法检测人胃癌MKN45细胞VEGF、b FGF、Ang-2蛋白表达的变化。Western blotting检测健脾解毒方对HP感染激活的人胃癌MKN45细胞Wnt/β-catenin信号通路活性的调控作用。结果 HP感染MKN45细胞12 h后,VEGF、b FGF、Ang-2蛋白的表达明显上调(P<0.01)。健脾解毒方醇提物低中高剂量组均可下调HP诱导的人胃癌MKN45细胞VEGF、b FGF、Ang-2蛋白的表达,并呈一定的剂量依赖效应。采用Wnt/β-catenin特异性抑制剂FH-535抑制Wnt/β-catenin活性后可明显抑制HP诱导的VEGF、b FGF、Ang-2蛋白的表达(P<0.01)。HP感染胃癌MKN45细胞12 h后,胞浆和胞核β-catenin蛋白表达明显上调(P<0.01),健脾解毒方可下调胞浆和胞核β-catenin蛋白表达,并呈一定的剂量依赖效应。结论健脾解毒方可下调HP诱导的人胃癌细胞VEGF、b FGF、Ang-2表达,抑制Wnt/β-catenin信号通路是其机制之一,这可能是该方抗胃癌作用的重要机制。Objective To study the influence of Jianpi Jiedu Recipe on the expression of vascular endothelial growth factor （VEGF）, angiopoietin-2 （Ang-2） and basic fibroblast growth factor（bFGF） in Helicobacter pylori （HP） -infected gastric cancer ceils, and discuss its possible mechanism. Methods Human gastric cancer MKN45 cells were infected by HP standard strain NCTC11637 by co-culture. The cells were divided into the control group, HP infection group, alcohol extract of Jianpi Jiedu Recipe（JPJD） groups with low-, medium- and high-dose（ as 0.5 g/L, 1.0 g/L and 2.5 g/L respectively）. The effects of Jianpi Jiedu Recipe on the expression of VEGF,bFGF and Ang-2 in HP infected MKN45 cells were detected by ELISA. After the inhibition on Wnt/13-catenin activity by the specific inhibitor FH-535 at concentration of 20 μmol/L, the cells were infected by HP once again, and the expression changes of VEGF, bFGF and Aug-2 in MKN45 ceils were detected by ELISA. The regulation of Jianpi Jiedu Recipe on the activity of Wnt/β-catenin signaling pathway in HP infected MKN45 ceils were determined by Western blotting. Results After 12 hours of HP infection, the expression of VEGF, bFGF and Aug-2 in MKN45 cells were obviously up-regulated （ P 〈 0.01 ）. The JPJD alcohol extract with low-, medium- and high-dose could all down-regulate the expressions of VEGF, bFGF and Ang-2 in HP infected MKN45 cells, with a certain dose-dependent manner. After the intervention of FH-535, the activity of Wnt/β-catenin was suppressed, the expressions of VEGF, bFGF and Ang-2 were obviously inhibited in HP infected MKN45 cells （P 〈0. 01 ）. After 12 hours of HP infection, the protein expression of β-catenin in cytoplasm and nucleus of MKN45 ceils was obviously up-regulated（ P 〈0.01 ）. Jianpi Jiedu Recipe could down-regulate the protein expression of β-catenin in cytoplasm and nucleus,with a certain dose-dependent manner. Conclusion Jianpi Jiedu Recipe could down-regulate the expressions of VEGF,bFGF and Ang-2 in HP-infe

关 键 词：幽门螺杆菌 健脾解毒方 胃癌 WNT/Β-CATENIN 血管新生 

分 类 号：R285[医药卫生—中药学]