【作者】 朱俭勋； 钟石； 李有贵； 计东风；

【Author】 ZHU Jian-xun;ZHONG Shi;LI You-gui;JI Dong-feng;Sericultural Research Institute,Zhejiang Academy of Agricultural Science;

【机构】 浙江省农业科学院蚕桑研究所；

【摘要】 通过MTT、流式细胞计数、RT-PCR等方法,研究桑黄、蛹虫草子实体多糖及两者复合多糖对肺癌A549细胞的抑制作用,并探讨其作用机制。结果发现,桑黄、蛹虫草多糖(0.8mg/ml)对肺癌细胞A549增殖具有一定的抑制作用,两者复合多糖(1:1,0.8mg/ml)则表现出协同增效抑制细胞A549增殖作用,抑制率可达54.7%;蛹虫草多糖(0.8mg/ml)可影响A549细胞周期各时相的分布,桑黄多糖(0.8mg/ml)无明显作用,但复合多糖(0.8mg/ml)对A549细胞周期各时相分布影响显著加强,G0/G1期细胞比例从(75.39±2.49)%降低至(64.43±2.12)%(P<0.01),S期细胞比例从(16.61±0.87)%升高至(27.97±2.82)%,(P<0.01);凋亡结果显示,桑黄+蛹虫草复合多糖(0.8mg/ml)作用48h后,晚期凋亡率从正常组(2.46±0.30)%提高至(20.75±0.24)%;坏死细胞率由正常组(1.88±0.43)%提高至(9.64±0.42)%,且表现出明显的协同增效诱导A549细胞凋亡作用;RT-PCR结果显示,桑黄、蛹虫草多糖处理后细胞内周期调控相关蛋白Cyclin A、CDK4 mRNA水平显著下调,Cyclin D1、CDK2表达显著提高;凋亡相关基因Bid、Bak、细胞色素C(Cyt-C)、Caspase3基因的表达水平也显著升高。结果表明,桑黄、蛹虫草多糖具有协同增效抑制肺癌A549细胞增殖,通过调控Cyclin A、CDK4、Bid、Bak的表达,导致Cyt-C释放、启动caspase级联反应,诱导细胞凋亡。更多还原

【Abstract】 This paper is aim to study the inhibitive effect and its underlying mechanism(s)of polysaccharides from Phellinus spp and Cordyceps sinensis on human lung adenocarcinoma A549 cells by MTT,flow cytometry,and RT-PCR methods.The results showed that polysaccharides from Phellinus linteus and Cordyceps sinensis exhibited a significant dose-dependent inhibitive effect on A549 cells proliferation.Moreover,the mixture polysaccharides(1:1,0.8 mg/ml)from Phellinus linteus and Cordyceps sinensis could increase the inhibitive effect on A549 cells,and the inhibitive ratio was increased to 54.41%.Flow cytometric analysis(FACS)results showed that cell cycle phase(G0/G1,S,G2/M)of A549 cells were affected after polysaccharides from Cordyceps sinensis treatment for 48 h,but the proportion of A549 cells in different phase was not significantly changed in polysaccharides from Phellinus spp treated group.But,the mixture polysaccharides(CP+PP,1:1)could cooperative induce S phase arrest on A549 cells,the number of cells in G0/G1 phase was significandy decreased from(75.39±2.49)%to(64.43±2.12)%,and S phase was increased from(16.61±0.87)%to(27.97±2.82)%evidently.Apoptosis results showed that A549 cells treated with mixture polysaccharides(CP+PP,1:1,0.8 mg/ml)for 48 h,the late apoptosis rate of A549 cell was significandy increased from(2.46±0.30)%to(20.75±0.24)%,while the necrosis ratio was increased from(1.88±0.43)%to(9.64±0.42)%,suggested that the polysaccharides from Phellinus spp and Cordyceps sinensis could cooperative induce A549 cell apoptosis.RT-PCR results showed diat the mRNA levels of cell cycle regulatory protein Cyclin A.CDK4 were significantly reduced,but the expression of Cyclin D1.CDK2 were evidently increased in A549 cells with polysaccharides from Phellinus linteus and Cordyceps sinensis treatment for 48 h.Furthermore,the mRNA levels of pro-apoptotic protein gene Bid,Bak,and apoptosis related gene cytochrome C(Cyt-C),Caspase3 were also significantly increased in different polysaccharides treated group.These results indicated that the mixture polysaccharides(1:l,0.8 mg/ml)from Phellinus linteus and Cordyceps sinensis could cooperative inhibit A549 cells proliferation and induced the apoptosis by regulating the expression of Cyclin A.CDK4,Bid,Bak,which resulted in the release of Cyt-C,activated die caspase cascade reaction.更多还原