摘要
目的:探讨整合素连接激酶(ILK)基因沉默后及中药肺岩宁方对相关下游靶基因表达的影响。方法:构建载体转染人肺腺癌细胞A549后,随机分为阴性对照病毒感染细胞(NC)组、RNAi靶点病毒感染细胞(KD)组、10%肺岩宁含药血清干预的NC组与KD组、20%肺岩宁含药血清干预的NC组与KD组,以及阳性对照10%DDP干预的KD组,应用Real time-PCR及Western Blot方法检测下游靶基因蛋白激酶B(Akt/PKB)、糖原合成酶激酶-3β(GSK-3β)、转录因子-1(AP-1)、细胞周期蛋白D1(cyclinD1)、基质金属蛋白酶-9(MMP-9)、血管内皮生长因子(VEGF)、β-链蛋白(β-catenin)的mRNA及蛋白表达水平。结果:与NC组比较,KD组GSK-3β、VEGF mRNA和蛋白表达显著下调(P<0.05,P<0.01),cyclinD1、MMP-9 mRNA和蛋白表达显著上调(P<0.05,P<0.01),Akt表达无变化,AP-1与β-catenin在mRNA水平表达升高(P<0.01)、蛋白水平表达则下降;与KD组比较,KD+10%肺岩宁含药血清和KD+10%DDP组具有下调VEGF、cyclinD1、MMP-9和β-catenin蛋白表达、上调GSK-3β蛋白表达,以及下调AP-1、cyclinD1 mRNA表达的作用(P<0.01);与KD组相比,KD+20%肺岩宁含药血清具有下调AP-1、cyclinD1和MMP-9蛋白表达的作用。结论:沉默ILK基因后可以明显降低GSK-3β、VEGF的表达,10%的中药肺岩宁方含药血清可能通过下调靶基因AP-1、cyclinD1、MMP-9、VEGF和β-catenin蛋白表达的作用而发挥其抗肿瘤的作用。
Objective：To discuss the effect of＂Feiyanning Decoction＂on the expression of target gene after the ILK gene silencing.Methods：The transfected A549 cells were constructed and randomized into negative control（ NC） group,Knock Down（ KD） group and treated group by 10% and 20% FYN drug serum and KD group treated by 10% DDP.The mRNA and protein expression Protein kinase B（ Akt）,Glycogen synthase kinase 3（ GSK-3beta）,transcription factor-1（ AP-1）,cell cycle protein 1（ CyclinD1）,Matrix metalloproteinase-9（ MMP-9）,Vascular endothelial growth factor（ VEGF） andβ-catenin genes were tested by the realtime-PCR and western blot.Results：Compared with the NC group,the mRNA and protein expression of GSK-3β and VEGF genes were significantly down-regulated（ P 0.05 or P 0.01）; the expression of cyclinD1 and MMP-9 genes were significantly up-regulated（ P 0.05 or P 0.01）; expression of AKT gene wasn＇t regulated.The mRNA expression of AP-1 and β-catenin was significantly up-regulated（ P 0.01）; the protein expression was downregulated; compared with the KD group,the protein expression of VEGF,cyclinD1,MMP-9 andβ-catenin were downregulated in the group of KD treated with 10% FYN drug serum and 10% DDP.The protein expression of GSK-3β was upregulated.The mRNA expression of AP-1 and cyclinD1 were significantly down-regulated（ P 0.01）; the protein expression of AP-1,cyclinD1 and MMP-9 were down-regulated.Conclusion：The expression of GSK-3β and VEGF genes were downregulated after the ILK silencing,the anti-tumor effects of＂Feiyanning Decoction＂can be played through the way by downregulating the protein expression of AP-1,cyclinD1,MMP,VEGF and β-catenin.
出处
《上海中医药大学学报》
CAS
2014年第1期51-55,共5页
Academic Journal of Shanghai University of Traditional Chinese Medicine
基金
国家自然科学基金资助项目(81072739)
上海中医药大学杏林学者资助项目(2209-13-03)
关键词
肺岩宁方
整合素连接激酶
肺癌
基因表达
＂Feiyanning Decoction＂
integrin-linked kinase（ILK）
lung cancer
gene expression
