芪珍胶囊抗结肠癌的作用机制研究  

Study on mechanism of Qizhen Capsule against colon cancer

作　　者：郭丹丹 郭铖洁 桑婷婷 王玉洁 吴凯凯 郭翠玲 方柳 王兴亚 GUO Dan-dan;GUO Cheng-jie;SANG Ting-ting;WANG Yu-jie;WU Kai-kai;GUO Cui-ling;FANG Liu;WANG Xing-ya(Zhejiang Chinese Medical University,Hangzhou 310053,China)

机构地区：[1]浙江中医药大学,浙江杭州310053

出　　处：《中草药》2021年第10期2967-2977,共11页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金面上项目(81973521);浙江省科技厅一流学科药学学科基金资助项目(2018-2020)。

摘　　要：目的探究芪珍胶囊对人结肠癌细胞HCT116及裸鼠移植瘤模型的作用及机制。方法HCT116细胞给予芪珍胶囊进行干预,采用MTT法和流式细胞术检测细胞存活率和凋亡情况;采用Western blotting法考察芪珍胶囊对聚腺苷二磷酸核糖聚合酶(poly ADP-ribose polymerase,PARP)、B淋巴细胞瘤-2(B-cell lymphoma-2,Bcl-2)、半胱氨酸蛋白酶-3(Caspase-3)、Caspase-9、剪切型Caspase-9(cleaved Caspase-9)、哺乳动物雷帕霉素靶蛋白(mammalian target of rapamycin,m TOR)、磷酸化m TOR(p-mTOR)、腺苷酸活化蛋白激酶(adenosine monophosphate-activated protein kinase,AMPK)、p-AMPK、细胞外信号调节蛋白激酶(extraeellular signal regulated kinase,ERK)、p-ERK、p38和p-p38蛋白表达的影响;采用qRT-PCR法检测芪珍胶囊对程序性死亡分子配体-1(programmed death ligand 1,PD-L1)m RNA表达的影响。建立结肠癌裸鼠移植瘤模型,考察芪珍胶囊对裸鼠体质量、瘤体积和瘤质量的影响;采用ELISA法检测各组裸鼠血清中肿瘤坏死因子-α(tumor necrosis factor alpha,TNF-α)、白细胞介素-6(interleukin-6,IL-6)和IL-1β水平;采用qRT-PCR法检测各组裸鼠肿瘤组织核因子-κB(nuclear factor-κB,NF-κB)、TNF-α、IL-6、CD68、单核细胞趋化蛋白-1(monocyte chemoattractant protein-1,MCP-1)、诱导型一氧化氮合酶(inducible nitric oxide synthase,iNOS)和PD-L1 mRNA表达;采用Western blotting法考察各组裸鼠肿瘤组织PD-L1蛋白表达。以γ-干扰素(interferon-γ,IFN-γ)诱导HCT116细胞,考察芪珍胶囊对细胞存活率及PD-L1蛋白表达的影响。结果芪珍胶囊显著抑制HCT116细胞存活率(P<0.01、0.001),显著促进细胞凋亡(P<0.01、0.001),显著下调细胞中Caspase-9、Caspase-3和Bcl-2蛋白表达水平(P<0.05、0.01),显著上调细胞中cleaved Caspase-9、cleaved PARP、p-mTOR/mTOR、p-AMPK/AMPK、p-ERK/ERK和p-p38/p38蛋白表达水平(P<0.05、0.01、0.001),显著下调细胞中PD-L1m RNA表达水平(P<0.05)。结肠癌裸鼠移植瘤模型Objective To explore effect and mechanism of Qizhen Capsule(芪珍胶囊)on human colon cancer cell HCT116 and transplanted tumor models in nude mice.Methods HCT116 cells were treated with Qizhen Capsules for intervention.MTT and flow cytometry were used to detect cell survival and apoptosis;Western blotting was used to investigate the effects of Qizhen Capsules on poly ADP-ribose polymerase(PARP),B-cell lymphoma-2(Bcl-2),Caspase-3,Caspase-9,cleaved Caspase-9,mTOR,phosphorylated m TOR(p-mTOR),adenosine monophosphate-activated protein kinase(AMPK),p-AMPK,extraeellular signal-regulated protein kinase(ERK),p-ERK,p38 and p-p38 expressions;qRT-PCR was used to detect the effect of Qizhen Capsule on programmed death ligand 1(PD-L1)mRNA level.Transplanted tumor model of colon cancer in nude mice was established,effects of Qizhen Capsule on body weight,tumor volume and tumor weight of nude mice were investigated;ELISA was used to detect tumor necrosis factor alpha(TNF-α),interleukin-6(IL-6)and IL-1βlevels in serum of nude mice;qRT-PCR was used to detect nuclear factor-κB(NF-κB),TNF-α,IL-6,CD68,monocyte chemoattractant protein-1(MCP-1),inducible nitric oxide synthase(i NOS)and PD-L1 mRNA levels in tumor;Western blotting was used to investigate PD-L1 expression in tumor tissues of nude mice in each group.IFN-γwas used to induce HCT116 cells to investigate the effect of Qizhen Capsule on cell survival rate and PD-L1 expression.Results Qizhen Capsule significantly inhibited the survival rate of HCT116 cells(P<0.01,0.001),significantly promoted cell apoptosis(P<0.01,0.001),significantly down-regulated expressions of Caspase-9,Caspase-3 and Bcl-2 in cells(P<0.05,0.01),significantly up-regulated expressions of cleaved Caspase-9,cleaved PARP,p-mTOR/mTOR,p-AMPK/AMPK,p-ERK/ERK and p-p38/p38 in cells(P<0.05,0.01,0.001),significantly down-regulated PD-L1 mRNA level in cells(P<0.05).In transplanted tumor model of colon cancer in nude mice,Qizhen Capsules significantly inhibited the growth of transplanted tumors(P<0.05,0.01),redu

关 键 词：芪珍胶囊 结肠癌 增殖 凋亡 炎性反应 程序性死亡分子配体-1 

分 类 号：R285.5[医药卫生—中药学]