缺氧微环境下HIF-1α通过PD-L1调控肝癌恶性生物学行为和上皮间质转化的机制研究  

HIF-1αregulates malignant biological behavior and epithelial interstitial transformation of hepatocellular carcinoma through PD-L1 in hypoxia microenvironment

作　　者：李昂 查文章 轩福明 王宏伟 周勇[2] LI Ang;ZHA Wenzhang;XUAN Fuming;WANG Hongwei;ZHOU Yong(Yancheng Clinical College of Xuzhou Medical University,Yancheng,Jiangsu 224005,China;Yancheng Clinical Medical College of Jiangsu University,Yancheng,Jiangsu 224005,China)

机构地区：[1]徐州医科大学盐城临床学院,江苏盐城224005 [2]江苏大学盐城临床医学院,江苏盐城224005

出　　处：《安徽医药》2025年第1期189-195,F0003,共8页Anhui Medical and Pharmaceutical Journal

基　　金：江苏省卫生健康委员会科研项目面上项目(M2020100);江苏大学医教协同创新基金项目(JDYY2023106)。

摘　　要：目的探讨缺氧微环境下,缺氧诱导因子-1α(HIF-1α)通过程序性死亡配体1(PD-L1)调控肝癌细胞增殖、迁移、侵袭和上皮间质转化的机制。方法选取2016年9月至2018年9月在徐州医科大学附属盐城临床学院确诊治疗的57例肝癌病人作为研究对象。免疫组织化学检测HIF-1α、PD-L1的表达水平,统计学分析HIF-1α、PD-L1与肝癌病人临床病理及总生存期之间的关系。通过氯化钴建立肝癌细胞缺氧模型,用细胞计数试剂盒(cell counting kit-8,CCK-8)法检测不同浓度氯化钴作用下细胞存活率;划痕实验、Transwell实验检测缺氧细胞及亲本细胞迁移、侵袭能力的变化;蛋白质印迹法检测缺氧细胞及亲本细胞HIF-1α、PD-L1蛋白表达水平。设计针对HIF-1α的siRNA(小干扰RNA,small interfering RNA)及PD-L1质粒,脂质体转染肝癌细胞氯化钴缺氧细胞,划痕实验、Transwell侵袭实验检测转染前后细胞迁移、侵袭能力的变化;蛋白质印迹法检测转染前后细胞HIF-1α、PD-L1、上皮间质转化(EMT)相关蛋白表达水平。结果与癌旁组织相比,HIF-1α、PD-L1在肝癌组织中高表达(P<0.05)。肝细胞癌中HIF-1α高表达组与低表达组在肿瘤长径、肿瘤分化程度和CNLC分期差异有统计学意义(P<0.05);PD-L1高表达组与低表达组在肿瘤数目、AFP表达水平和CNLC分期差异有统计学意义(P<0.05);且log-rank检验方法显示:HIF-1α、PD-L1高表达组病人的总生存期较低表达组明显降低(P<0.05)。与常氧细胞相比,缺氧细胞穿透小室的细胞数显著增加[(429.33±12.01)个比(244.3±8.14)个,t=−22.08,P<0.001],氯化钴缺氧肝癌细胞增殖、迁移及侵袭能力增强,HIF-1α、PD-L1表达水平明显增加(P<0.05)。与亲本细胞相比,HIF-1αsiRNA作用后肝癌缺氧细胞增殖、迁移及侵袭能力明显降低(P<0.05),且PD-L1、波形蛋白(Vimentin)及扭曲相关蛋白1(Twist1)蛋白表达明显降低,而上皮钙黏素(E-cadherin)蛋白表达明显增�Objective To investigate the mechanism of HIF-1α regulating hepatocellular carcinoma(HCC)cell proliferation,migration,invasion and epithelial mesenchymal transformation through programmed death ligand 1(PD-L1)in hypoxia microenvironment.Methods Fifty-seven patients diagnosed with liver cancer and treated at the Yancheng Clinical College of Xuzhou Medical University from September 2016 and September 2018 were selected as the study objects.The expression of HIF-1α and PD-L1 were detected by immunohistochemistry,and the relationship between HIF-1α,PD-L1 and clinicopathological and overall survival was statistically analyzed.The hypoxia model of HCC was established by Cobalt Chloride(CoCl2),and the survival rate of HCC under different concentrations of CoCl2 was detected by Cell Counting Kit-8(CCK-8)method.The changes of migration and invasion ability of hypoxic cells and parental cells were detected by scratch test and transwell invasion test.The expressions of HIF-1α and PD-L1 in hypoxic cells and parental cells were detected by Western blot.Small interfering RNA(siRNA)targeting HIF-1α and PD-L1 plasmid were designed to transfect HCC cells with liposome into hypoxic cells.The changes of cell migration and invasion ability before and after transfection were detected by scratch assay and Transwell(migration/invasion)invasion assay.The expression levels of HIF-1α,PD-L1 and Epithelial mesenchymal transformation(EMT)-related proteins before and after transfection were detected by Western blotting.Results HIF-1α and PD-L1 were highly expressed in HCC tissues compared with paracancellular tissues(P<0.05).There were significant differences in tumor size,tumor differentiation degree and China liver cancer staging(CNLC)stage between high and low PD-L1 expression groups(P<0.05).There were significant differences in tumor number,AFP expression level,and CNLC stage between high and low PD-L1 expression groups(P<0.05).Kaplan-Meier analysis showed that the survival rate of patients in HIF-1αand PD-L1 high expression group

关 键 词：癌 肝细胞 缺氧微环境 缺氧诱导因子-1Α 程序性死亡受体-配体1 上皮间质转化 

分 类 号：R735.7[医药卫生—肿瘤]