摘要
目的:缺氧是造成胃癌化学治疗(以下简称"化疗")耐药的重要原因,但目前对单纯缺氧环境下胃癌的生长情况了解甚少,本研究通过观察缺氧对胃癌细胞生长的影响,探讨缺氧环境中胃癌细胞对化疗药物5-氟尿嘧啶(5-fluorouracil,5-FU)的反应.方法:设置缺氧(1%氧气)及常规空气环境,将胃癌细胞MKN45分别置于上述环境中培养,同时设立化疗药物5-FU干预组.采用细胞计数试剂盒-8(cell counting kit-8,CCK-8)法、JC-1线粒体膜电位法及Annexin-V/PI双染法检测不同氧含量环境中、不同干预组胃癌细胞的增殖及凋亡情况,流式细胞术检测细胞周期的改变,在电镜下观察细胞线粒体变化情况.结果:不加5-FU干预的情况下,缺氧组与常氧组相比,JC-1线粒体膜电位法、Annexin-V/PI双染法及CCK-8检测结果显示胃癌细胞早期凋亡、晚期凋亡率更高,细胞死亡率更高;流式细胞术检测结果显示细胞周期被阻止在G0/G1期,不进展;电镜结果显示线粒体破坏更严重.而在加入5-FU干预的情况下,缺氧组与常氧组相比,凋亡率低,细胞周期更易进展,线粒体破坏更少.结论:缺氧环境促进胃癌细胞凋亡甚至死亡,但是缺氧环境对化疗药物5-FU的起效产生反作用,可能是造成5-FU化疗耐药的因素.
Abstract
Objective:Hypoxia is an important cause of chemotherapy resistance in gastric cancer.However,little is known about the growth of gastric cancer under purely hypoxia conditions.This study aims to study the effect of hypoxia on the growth patterns of gastric cancer cells and explore the response of gastric cancer cells to the chemotherapeutic drug 5-fluorouracil(5-FU)in a hypoxic environment.Methods:Gastric cancer cells MKN45 were cultured under 1%oxygen hypoxia and conventional air conditions.An intervention group with the addition of the chemotherapeutic drug 5-FU was also established.The proliferation and apoptosis of gastric cancer cells under different oxygen conditions and intervention groups were detected using the cell counting kit-8(CCK-8)method,JC-1 mitochondrial membrane potential assay,and Annexin-V/PI double staining method.Cell cycle changes were detected by flow cytometry,and mitochondrial changes were detected using electron microscopy.Results:In the absence of 5-FU intervention,compared with the normoxia group,the hypoxia group showed higher rates of early and late apoptosis and higher cell death rates as indicated by the JC-1 mitochondrial membrane potential assay,Annexin-V/PI double staining,and CCK-8 results.Flow cytometry results showed that the cell cycle was arrested in the G0/G1 phase without progression.Electron microscopy revealed more severe mitochondrial destruction.However,with 5-FU intervention,the hypoxia group showed lower apoptosis rates,more cell cycle progression,and less mitochondrial destruction compared with the normoxia group.Conclusion:Hypoxic environments promote apoptosis and even death in gastric cancer cells,but hypoxia counteracts the efficacy of the chemotherapeutic drug 5-FU,which may contribute to 5-FU chemotherapy resistance.
基金项目
兰州大学第二医院萃英学子科研培育计划(CYXZ2022-08)
维普
万方数据