白花蛇舌草提取物通过抑制有氧糖酵解和促进氧化磷酸化抑制肝癌细胞增殖  

Hedyotis diffusa extract inhibits aerobic glycolysis and promotes oxidativephosphorylation to suppress the proliferation of liver cancer cells

作　　者：贺红艳 晁满香 翟翠 张晴 李海燕 HE Hongyan;CHAO Manxiang;ZHAI Cui;ZHANG Qing;LI Haiyan(Department of Medical Technology,Xi’an Medical College,Xi’an 710021;Department of Neurology Air Force 986th Hospital,Air Force Military Medical University,Xi’an 710000;Shaanxi Key Laboratory of Brain Disorders,Institute of Basic and Translational Medicine,Xi’an Medical College,Xi’an 710021,China)

机构地区：[1]西安医学院医学技术学院,陕西西安710021 [2]空军军医大学空军第九八六医院神经内科,陕西西安710000 [3]西安医学院基础与转化医学研究所,陕西省脑疾病防治重点实验室,陕西西安710021

出　　处：《西安交通大学学报（医学版）》2024年第4期656-662,共7页Journal of Xi’an Jiaotong University（Medical Sciences）

基　　金：国家自然科学基金资助项目(No.81700546);陕西省科技厅自然科学基础研究计划项目(No.2023-JC-QN-0863)。

摘　　要：目的 探讨白花蛇舌草提取物(HDE)对肝癌细胞增殖的影响及其与有氧糖酵解和氧化磷酸化的关系,并分析其可能机制。方法 采用CCK-8实验和细胞增殖实验(5-ethynyl-2-deoxyuridine, EDU)检测不同质量浓度(20、40、80 mg/mL)HDE对肝癌细胞SNU-368增殖的影响;采用酶标仪检测乳酸脱氢酶活性、葡萄糖摄取、乳酸生成、线粒体呼吸链复合体活性,采用pH计检测细胞外液pH值,采用细胞氧耗仪检测细胞氧耗,分析HDE对SNU-368细胞有氧糖酵解和氧化磷酸化的影响;采用qRT-PCR实验检测各组SNU-368细胞中GLUT1、GLUT4、HK2、GPI、PFKL、ALDOA和HIF-1α的mRNA表达,采用Western blotting检测HIF-1α的蛋白表达。用过表达HIF-1α的慢病毒转染肝癌细胞SNU-368构建过表达HIF-1α稳转细胞株,并进行HDE相应干预后,采用qRT-PCR和Western blotting检测HIF-1α的mRNA及蛋白表达。结果 CCK-8实验显示,HDE呈一定的浓度依赖效应抑制肝癌细胞增殖(均P<0.05);糖代谢相关检测结果显示,HDE可以抑制葡萄糖摄取和乳酸生成,降低乳酸脱氢酶活性,升高细胞外培养液pH值,增加细胞氧耗和线粒体呼吸链复合体Ⅰ、Ⅱ、Ⅲ、Ⅳ活性(均P<0.05);qRT-PCR结果显示,HDE抑制了GLUT1、HK2、GPI、ALDOA mRNA的表达(均P<0.05)。qRT-PCR和Western blotting实验显示,与对照组相比,HDE组HIF-1α mRNA及蛋白的表达明显减少;而与HDE组相比,HDE干预过表达HIF-1α的HIF-1α-LV组中,HIF-1α mRNA及蛋白表达再次增加(P<0.05)。结论 HDE通过抑制肝癌细胞有氧糖酵解、促进氧化磷酸化而抑制肝癌细胞增殖,作用机制可能与抑制HIF-1α的表达有关。Objective To investigate the effect of Hedyotis diffusa extract(HDE)on the proliferation of liver cancer cells and its relationship with sugar metabolism reprogramming and oxidative phosphorylation and analyze its possible mechanisms.Methods CCK-8 and EDU experiments were used to determine the effect of different concentrations(20,40,80 mg/mL)of HDE on the growth of liver cancer cell line SNU-368.Lactate dehydrogenase activity,glucose uptake,lactate production,extracellular pH,mitochondrial respiratory chain complex activity,and cellular oxygen consumption were measured to analyze the effect of HDE on aerobic glycolysis and oxidative phosphorylation in liver cancer cells.qRT-PCR experiments were used to detect the mRNA expressions of GLUT1,GLUT4,HK2,GPI,PFKL,ALDOA and HIF-1αin SNU-368 cells of different groups.Western blotting experiments were used to detect the protein expression of HIF-1α.A stable cell line overexpressing HIF-1αwas constructed by lentivirus transfection of liver cancer cells SNU-368 and then intervened with HDE;the expression of HIF-1αmRNA and protein was detected with qRT-PCR and Western blotting.Results CCK-8 results showed that the HDE exhibited a concentration-dependent inhibitory effect on the proliferation of liver cancer cells(all P<0.05).Results from glucose metabolism-related tests indicated that the HDE could inhibit glucose uptake and lactate production,decrease lactate dehydrogenase activity,increase extracellular pH value,enhance cellular oxygen consumption,and elevate activities of mitochondrial respiratory chain complexesⅠ,Ⅱ,ⅢandⅣ(all P<0.05).qRT-PCR results revealed that the HDE suppressed the mRNA expressions of GLUT1,HK2,GPI,and ALDOA(all P<0.05).qRT-PCR and Western blotting experiments showed that compared to the control group,the expression of HIF-1αmRNA and protein in the HDE group was significantly reduced.However,when HIF-1αwas overexpressed and HDE was added in the HIF-1α-LV group,the expression of HIF-1αmRNA and protein increased again compared to the HD

关 键 词：白花蛇舌草提取物 肝癌细胞 糖酵解 氧化磷酸化 增殖 

分 类 号：R285[医药卫生—中药学]