黄芪多糖抑制肺腺癌A549/DDP细胞移植瘤裸鼠EMT改善顺铂耐药的机制  

Astragalus Polysaccharides Improve Cisplatin Resistance by Inhibiting EMT of Lung Adenocarcinoma A549/DDP Cells Transplanted into Nude Mice

作　　者：张颖[1] 王淳[2] 于丹[2] 高原[2] 井欢[2] 王莹[2] 刘春英[2] ZHANG Ying;WANG Chun;YU Dan;GAO Yuan;JING Huan;WANG Ying;LIU Chun-ying(Jining Medical University,Jining 272000,China;Liaoning University of Traditional Chinese Medicine,Shenyang 110032,China)

机构地区：[1]济宁医学院,山东济宁272000 [2]辽宁中医药大学,沈阳110032

出　　处：《中国实验方剂学杂志》2022年第6期79-85,共7页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金面上项目(81973735,81774184);济医院字[2019]18号中西医结合治疗脾胃病创新团队。

摘　　要：目的:探讨黄芪多糖(APS)抑制转化生长因子-β1(TGF-β1)诱导的肺腺癌耐顺铂细胞株A549/顺铂细胞移植瘤裸鼠上皮-间质转化(EMT)进程和改善顺铂耐药的分子机制。方法:BALB/c裸鼠随机分为空载组、模型组、顺铂组、联合组(顺铂与黄芪多糖联合应用)。将TGF-β1基因过表达慢病毒载体及阴性对照组感染A549/顺铂细胞,感染成功的细胞进行裸鼠皮下接种。除TGF-β1组A549/顺铂细胞接种至空载组外,其他组均接种TGF-β1基因稳定过表达的A549/顺铂细胞。细胞接种8 d后,开始进行药物干预。联合组灌胃APS(0.3 g·kg^(-1)·d^(-1)),腹腔注射顺铂(0.003 5g·kg^(-1));顺铂组腹腔注射顺铂(0.003 5 g·kg^(-1))。细胞接种32 d后,处死裸鼠,取肿瘤组织、肺。称量瘤质量,计算抑瘤率;显微镜下计数整张玻片肺肿瘤转移结节数;免疫组化、蛋白免疫印迹法(Western blot)、实时荧光定量聚合酶链式反应(Real-time PCR)检测肿瘤EMT标志物α-连环蛋白(α-catenin)、N-钙黏蛋白(N-cadherin)和肿瘤耐药标志物人肺耐药蛋白(LRP)、多药耐药相关蛋白(MRP)、P-糖蛋白(Pgp)蛋白和mRNA表达情况。结果:与空载组比较,模型组瘤质量增加,肺肿瘤转移结节增多(P<0.05),α-catenin蛋白和mRNA表达降低(P<0.05),N-cadherin、LRP、MRP、P-gp蛋白和mRNA表达升高(P<0.05)。与模型组、顺铂组比较,联合组瘤质量减轻,肺肿瘤转移结节减少(P<0.05);α-catenin蛋白和mRNA表达升高(P<0.05),N-cadherin、LRP、MRP、P-gp蛋白和mRNA表达降低(P<0.05)。结论:黄芪多糖可抑制肺腺癌移植瘤生长和转移,改善顺铂耐药,这一作用可能与其抑制肿瘤细胞EMT进程相关。Objective: To investigate the inhibitory effect of Astragalus polysaccharide(APS)on epithelial-mesenchymal transition(EMT) induced by transforming growth factor-β1(TGF-β1)in cisplatin(DDP)-resistant lung adenocarcinoma cell line A549/DDP cells transplanted into nude mice and the molecular mechanism in improving DDP resistance. Method: BALB/c nude mice were randomly divided into a blank group,a model group,a DDP group,and a combination group(APS combined with DDP). A549/DDP cells were infected with TGF-β1-overexpressed lentiviral vector and the negative control. The infected cells were inoculated subcutaneously in nude mice. The A549/DDP cells with TGF-β1 gene overexpression were inoculated into all groups except the control group with negative TGF-β1 gene overexpression. The drug intervention was performed eight days after cell inoculation. The mice in the combination group received intragastric administration of APS(0.3 g·kg^(-1)·d^(-1))and intraperitoneal injection of cisplatin(0.003 5 g·kg^(-1)),and those in the cisplatin group received intraperitoneal injection of cisplatin(0.003 5 g·kg^(-1)). After 32 days of cell inoculation,the nude mice were killed and the tumor tissues and lungs were collected. The tumor weight was recorded and the inhibition rate was calculated. The number of metastatic nodules of the lung tumor on the whole slide was counted under the microscope. Immunohistochemistry,Western blot,and real-time fluorescence-based quantitative polymerase chain reaction(Real-time PCR)were used to detect the protein and gene expression of EMT molecular markers α-catenin and N-cadherin,and tumor drug resistance markers human lung resistance protein(LRP),multidrug resistance-associated protein(MRP),and P-glycoprotein(P-gp)in the transplanted tumor. Result: Compared with the blank group,the model group showed increased tumor weight and metastatic nodules of the lung tumor(P<0.05),decreased protein and mRNA expression of α-catenin(P<0.05),and elevated protein and mRNA expression of N-cadherin,LRP,MR

关 键 词：黄芪多糖 上皮-间质转化(EMT) 顺铂耐药 肺腺癌 

分 类 号：R22[医药卫生—中医基础理论] R242[医药卫生—中医学]