HIF1α/HIF2α经EGF调控胶质瘤细胞化疗抵抗机制研究  

Study on the mechanism of HIF1α/HIF2αregulating glioma cell chemotherapy resistance via EGF

作　　者：杜军 汪攀 龚胜 赵璐 郭海燕 廖彬 DU Jun;WANG Pan;GONG Sheng;ZHAO Lu;GUO Haiyan;LIAO Bin(Department of Neurosurgery,Chongqing General Hospital,Chongqing 401147,China;Department of Neurosurgery,Qianjiang Central Hospital,Chongqing 409000,China)

机构地区：[1]重庆市人民医院神经外科,401147 [2]重庆市黔江中心医院神经外科,409000

出　　处：《重庆医学》2022年第8期1261-1265,共5页Chongqing medicine

基　　金：重庆市渝中区基础研究与前沿探索项目(20200133);重庆市人民医院医学科技创新基金(Y2020MSXM03)。

摘　　要：目的通过体外实验探讨低氧诱导因子1α(HIF1α)和HIF2α经表皮生长因子(EGF)调控胶质瘤细胞化疗抵抗的机制。方法采用CRISPR-CAS9系统单独或同时敲除U87细胞HIF1α和HIF2α表达(分为HIF1α单独敲除组、HIF2α单独敲除组、HIF1α/HIF2α同时敲出组),另设未转染的U87细胞为对照组。缺氧或常氧培养各组细胞并给予替莫唑胺(TMZ)处理,检测各组细胞毒性率和凋亡率。缺氧培养HIF1α/HIF2α同时敲出组的U87细胞,RT-qPCR检测EGF mRNA表达水平,进一步在培养基中添加EGF后予以TMZ处理,检测细胞毒性率和凋亡率。结果Western blot提示U87细胞HIF1α和HIF2α敲除成功。缺氧条件下,HIF1α和HIF2α单独敲除组及HIF1α/HIF2α同时敲除组EGF表达水平较对照组显著降低(P<0.05),HIF1α/HIF2α同时敲除组EGF表达水平也低于HIF1α、HIF2α单独敲除组(P<0.05)。缺氧条件下,TMZ处理的HIF1α、HIF2α单独敲除组的细胞毒性率与对照组比较差异无统计学意义(P>0.05),HIF1α/HIF2α同时敲除组细胞毒性率高于对照组及HIF1α、HIF2α单独敲除组(P<0.05);TMZ处理的HIF1α、HIF2α单独敲除组及HIF1α/HIF2α同时敲除组细胞凋亡率均高于对照组(P<0.05),HIF1α/HIF2α同时敲除组细胞凋亡率高于HIF1α、HIF2α单独敲除组。缺氧条件下,HIF1α/HIF2α同时敲除组添加EGF后,TMZ处理的细胞毒性率和细胞凋亡率均低于未添加EGF的HIF1α/HIF2α同时敲除组(P<0.05)。结论HIF1α和HIF2α经EGF促进胶质瘤细胞化疗抵抗。Objective To investigate the mechanism of hypoxia-inducible factor 1α(HIF1α)and HIF2αvia epidermal growth factor(EGF)on the chemotherapy resistance of glioma cells through in vitro experiments.Methods CRISPR-CAS9 system was used to knock out HIF1αand HIF2αin U87 cells alone or simultaneously(the HIF1αknockout alone group,the HIF2αknockout alone group,the HIF1α/HIF2αsimultaneously knockout group),and additional untransfected U87 cells were set as control group.Temozolomide(TMZ)was administered to detect the cytotoxicity and apoptosis rates in each group.After U87 cells of the HIF1αand HIF2αsimultaneously knockout group were cultured under hypoxia,EGF mRNA expression level was detected by RT-qPCR.TMZ was treated after EGF was added,then the cytotoxicity and apoptosis of cells of the HIF1α/HIF2αsimultaneously knockout group were detected.Results The results of Western blot showed that HIF1αand HIF2αwere knocked out successfully in U87 cells.Under hypoxia,the expression level of EGF in HIF1αand HIF2αknockout alone groups and the HIF1α/HIF2αsimultaneously knockout group were lower than that in the control group(P<0.05),the expression level of EGF in HIF1α/HIF2αknockout simultaneously group were lower than that in the HIF1αand HIF2αknockout alone groups(P<0.05).Under hypoxia,the cytotoxicity rate of TMZ to HIF1αand HIF2αknockout alones group was no different from that of the control group(P>0.05).The cytotoxicity rat of TMZ to HIF1α/HIF2αknockout simultaneously group was higher than that of the control group,the HIF1αand HIF2αknockout alone groups(P<0.05).The apoptosis of HIF1αand HIF2αknockout alone and simultaneously groups were higher than that of control group(P<0.05),while the apoptosis rate of HIF1α/HIF2αsimultaneously knockout group was higher than that of HIF1αand HIF2αknockout alone groups.Under hypoxia,the cytotoxicity and apoptosis of HIF1α/HIF2αknockout simultaneously group supplemented with EGF were lower than those without EGF(P<0.05).Conclusion HIF1αand HIF2αcan pro

关 键 词：胶质瘤 低氧诱导因子1Α 低氧诱导因子2α 表皮生长因子 缺氧 化疗抵抗 

分 类 号：R739.41[医药卫生—肿瘤]