摘要
背景与目的:Wnt/β-catenin信号通路在人类肿瘤尤其在大肠癌的发生发展中起着重要作用。本研究分析了香加皮杠柳苷(periplocin extracted from cortex periplocae,CPP)对人结肠癌细胞SW480增殖的抑制作用,及对Wnt/β-catenin信号通路的调控作用。方法:MTT法检测CPP对SW480细胞增殖的影响,流式细胞技术检测细胞周期的变化和凋亡。Western blot法检测CPP处理组与对照组细胞总蛋白、细胞浆蛋白及细胞核蛋白中β-catenin表达变化,电泳迁移率改变法分析CPP作用后SW480细胞核TCF复合物与其特异性DNA结合序列结合能力变化。半定量RT-PCR法检测CPP作用后细胞中β-catenin、survivin、c-myc和cyclin D1 mRNA的表达。结果:CPP明显抑制SW480细胞增殖(P<0.01),并呈时间和浓度依赖性;0.5μg/mLCPP可将SW480细胞阻滞于G0/G1期,并诱导细胞凋亡(P<0.05)。CPP作用后SW480细胞总蛋白、胞浆蛋白及细胞核蛋白中的β-catenin表达均明显降低(P<0.01),细胞核中TCF复合物与其特异性DNA结合序列结合能力受到抑制,其下游靶基因mRNA表达水平下降(P<0.01),而β-catenin mRNA表达未见明显改变。结论:CPP可明显抑制SW480细胞增殖并诱导细胞凋亡,其作用机制与抑制细胞Wnt/β-catenin信号转导通路有关。
Background and Objective： The Wnt/β-catenin signaling pathway plays an important role in the development and progression of human cancers, especially in colorectal carcinomas. This study was to analyze the inhibition effect of periplocin extracted from cortex periplocae （CPP） on proliferation of human colon carcinoma cell line SW480 and the underlying mechanism. Methods： Cell proliferation of SW480 cells was measured by M-FF assay. Cell apoptosis and cell cycle were analyzed by flow cytometry. Protein expression of β-catenin in total cell lysates, cytosolic extracts, and nuclear extracts were detected by Western blot. Binding activity of the T cell factor （TCF） complex in nucleus to its specific DNA binding site was measured by electrophoretic mobility shift assay （EMSA）. Expressions of β-catenin, survivin, c-myc and cyclin D1 mRNA in cells after the treatment with CPP were detected by semi-quantitative RT-PCR. Results： CPP significantly inhibited the proliferation of SW480 cells in a time- and dose- dependent manner （P〈0.01）. CPP （0.5 μg/mL） also caused Go/G1 cell cycle arrest of SW480 cells and induced cell apoptosis （P〈0.05）. Compared to untreated control cells, after the treatment with CPP, the protein levels of β-catenin in total cell lysates, cytosolic extracts, and nuclear extracts were reduced （P〈0.01）; the binding activity of the TCF complex in nucleus to its specific DNA binding site was suppressed; mRNAs of the downstream target genes survivin, c-myc and cyclin D1 were decreased （P〈0.01） while β-catenin mRNA remained unchanged. Conclusion： CPP could significantly inhibit the proliferation of SW480 cells, which may be through down- regulating the Wnt/β-catenin signaling pathway.
出处
《癌症》
SCIE
CAS
CSCD
北大核心
2009年第5期456-460,共5页
Chinese Journal of Cancer
基金
国家自然科学基金资助项目(No.30772752)
河北省自然科学基金项目(No.C2008000952)~~
