低氧和缺氧诱导因子-1α对人白血病细胞株K562细胞血管相关生成因子的影响  

Effect of hypoxia and hypoxia-inducible factor-1α on angiogenesisrelated factors in human leukemia K562 cells

作　　者：解友邦[1] 李建平[1] 沈括[1] 孟芳[1] 王莉[1] 韩国雄[1] 艾国[1] 蒋白丽 赵强强[1] 候艳 杨红艳[1] 李文倩[1] XIE Youbang;LI Jianping;SHEN Kuo;MENG Fang;WANG Li;HAN Guoxiong;AI Guo;JIANG Baili;ZHAO Qiangqiang;HOU Yan;YANG Hongyan;LI Wenqian(Department of Hematology and Rheumatology,Qinghai Provincial People's Hospital,Xining,Qinghai 810007,China)

机构地区：[1]青海省人民医院血液风湿科,西宁810007

出　　处：《重庆医学》2019年第3期364-369,共6页Chongqing medicine

基　　金：青海省卫生和计划生育委员会2016年科研指导项目(青卫科[2016]22号2016-wjzdx-17);青海省国家重点专科项目(卫办医政函[2011]873号);2014年度青海省人才小高地项目(青海人才字[2014]12号);青海省高层次卫生人才(骨干人才和领军人才)计划

摘　　要：目的观察低氧和缺氧诱导因子-1α(HIF-1α)对K562细胞分泌血管相关生成因子的影响。方法选用携带和干扰HIF-1α基因的慢病毒转染K562细胞,以转染携带和干扰HIF-1α基因的K562细胞分别作为过表达组、干扰组,同时以空载病毒转染K562细胞作为对照组,3组在低氧状态下培养72h后收集细胞。通过荧光显微镜观察转染效率,采用RT-PCR法检测HIF-1α和血管相关生成因子mRNA表达水平;ELISA法检测培养上清液中血管相关生成因子蛋白水平。结果慢病毒转染K562细胞最佳感染复数(MOI)为10,转染效率约50%,经过嘌呤霉筛选后转染阳性细胞达90%以上。干扰组人血管生成素(ANG)-Ⅱ和血管内皮生长因子(VEGF)mRNA表达均低于过表达组,而转化生长因子(TGF)-βmRNA表达高于对照组和过表达组,过表达组ANG-ⅡmRNA表达高于对照组,差异有统计学意义(P<0.05)。培养上清液中TGF-α未检测到,过表达组ANG-Ⅱ水平低于对照组(P<0.05);过表达组VEGF和TNF-α水平高于对照组,干扰组TGF-β和VEGF水平低于对照组(P<0.05)。干扰组TGF-β、VEGF和TNF-α水平低于过表达组,差异有统计学意义(P<0.05)。结论低氧和HIF-1α可影响慢性白血病K562细胞血管生成相关因子的表达和自分泌,但在mRNA表达和蛋白分泌水平上存在差异。Objective To investigate the effect of hypoxia and hypoxia inducible factor 1α(HIF-1α)on human leukemia cell line K562 secreting angiogenesis-related factors.Methods The lentivirus carrying and interfering HIF-1α was transfected into K562 cells.The K562 cells transfection carrying and interfering HIF-1α gene served as the overexpression group and interference group respectively.Meanwhile K562 cells with empty virus transfection served as the control group.The cells in 3 groups were harvested after 72h of incubation under hypoxia.The transfection efficiency was observed by using the fluorescence microscope.The expression levels of HIF-1αand angiogenesis-related factors mRNA were detected by RT-PCR.The level of angiogenesis-related factor protein in the culture supernatant was detected by ELISA.Results The optimal multiplicity of infection(MOI)of transfected K562 cells with lentivirus was 10 and the transfection efficiency was about 50%.The transfection positive cells after screening by puromycin reached more than 90%.The expressions of ANG-Ⅱand VEGF mRNA in the interference group were all lower than those in the overexpression group,while the TGF-β mRNA expression was higher than that in the control group and overexpression group(P<0.05);the ANG-ⅡmRNA expression level in the overexpression group was higher than that in the control group,the difference was statistically significant(P<0.05).TGF-α in the culture supernatant fluid was not detected.The ANG-Ⅱ level in the overexpression group was lower than that in the control group;the VEGF and TNF-αlevels in the overexpression group were higher than those in the control group,the TGF-β and VEGF levels in the interference group were lower than those in the control group(P<0.05).The levels of TGF-β,VEGF and TNF-α in the interference group were lower than those in the overexpression group.The above differences were statistically significant(P<0.05).Conclusion Hypoxia and HIF-1α could affect the expression and autocrine of angiogenesis-related factors in

关 键 词：白血病 K562细胞 缺氧诱导因子-1Α 低氧 血管相关生成因子 

分 类 号：R557.3[医药卫生—血液循环系统疾病]