香叶木素通过抑制Wnt/β-catenin途径抑制A549细胞增殖和肿瘤球形成  

Diosmetin Inhibits Proliferation and Tumor Sphere Formation of A549 Cells by Inhibiting Wnt/β-catenin Pathway

作　　者：崔东[1] 冯雨 栾加强 侯曼曼 赵悦[2] 钱如林[1] Cui Dong;Feng Yu;Luan Jiaqiang(Department of Thoracic Surgery,Henan Provincial Chest Hos pital,Zhengzhou 450003,China)

机构地区：[1]河南省胸科医院胸外科,郑州450003 [2]河南省胸科医院医学检验科,郑州450003

出　　处：《华中科技大学学报（医学版）》2023年第3期334-339,350,共7页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：河南省医学科技攻关计划项目(No.LHGJ20190740)。

摘　　要：目的探究香叶木素对A549细胞增殖、干细胞特性的影响及其机制。方法MTT法检测不同浓度香叶木素对细胞活性的作用,随后将A549细胞分为对照组、香叶木素低、中、高剂量组和顺铂处理组。集落形成实验检测A549细胞增殖;成球实验检测成球直径和成球数量;流式细胞术检测肿瘤干细胞标记物CD133的表达,定量逆转录聚合酶链反应(qRT-PCR)和Westernbolt分别检测CD44、SOX2、乙醛脱氢酶1(ALDH1)、OCT4、Wnt、β-catenin和GSK-3β的mRNA和蛋白相对表达量。建立异种移植瘤模型,免疫组化检测CD44和SOX2的表达,Westernblot检测β-catenin蛋白表达量。结果与对照组相比,中、高剂量香叶木素处理后,A549细胞活力和克隆形成率降低,干细胞成球直径和成球数量降低,CD133阳性细胞率减少,CD44、SOX2、ALDH1、OCT4、Wnt和β-catenin的mRNA和蛋白相对表达量降低,GSK-3β的mRNA和蛋白相对表达量升高,GSK-3β蛋白磷酸化水平降低。在体内,香叶木素处理组的肿瘤重量减轻,肿瘤组织中CD44和SOX2阳性细胞数均减少,β-catenin蛋白表达量下调。结论香叶木素可能通过抑制Wnt/β-catenin途径抑制A549细胞增殖及干细胞活性。Objective To explore the effects of diosmetin on the prolifera tion,stem cell characteristics of A549 cells and its mechanism.Methods MTT method was used to detect the effect of different concentrations of diosmetin on cell activity,and then A549 cells were divided into control group,low-dose,medium-dose and high-dose diosmetin groups and cisplatin treatment group.The proliferation of A549 cells was detected by colony formation assay.The diameter and number of spheres were ana-lyzed by sphere formation assay.The expression of tumor stem cell marker CD133 were detected by flow cytometry.The relative mRNA and protein expression levels of CD44,SOX2,aldehyde dehydrogenase 1(ALDH1),OCT4,Wnt,B-catenin and GSK--3βwere detected by quantitative real-time polymerase chain reaction(qRT-PCR).The expression of CD44,SOX2,ALDH1,OCT4,Wnt,β-catenin,p-GSK-3βand GSK-3βproteins was detected by W estern blotting.A xenograft tumor model was establi shed,the expression of CD44 and SOX2 was detected by immunohistochemistry,and the expression ofβ-catenin protein was detected by Western blot ting.Results Compared with the control group,the cell viability and clone formation rate of A549 cell were de-creased,the spheres diameters and spheres number were decreased,and the positive cell rate of CD133 was decreased.The rela-tive mRNA and protein expression levels of CD44,SOX2,ALDH1,OCT4,Wnt andβ-catenin were decreased,the mRNA and protein relative expression level of GSK-3βwas increased,and the phosphorylation level of GSK-3βprotein was decreased after medium and high doses of diosmetin treatment.In vivo,the tumor weight was reduced,the number of CD44 and SOX2 positive cells in tumor tissue was reduced,and the expression level ofβ-catenin protein was down-regulated in diosmetin treatment group.Conclusion Diosmetin may inhibit the proliferation and stem cell activity of A549 cells by inhibiting Wnt/β-catenin path-way.

关 键 词：肺癌 香叶木素 增殖 肿瘤球形成 Wnt/β-catenin途径 

分 类 号：R734.2[医药卫生—肿瘤]