学术论文

      缺氧诱导因子-1α改善缺氧血管内皮细胞线粒体损伤的实验研究

      Hypoxia-inducible factor-1α relieves mitochondrial damage in hypoxic vascular endothelial cells

      摘要:
      目的 探讨缺氧诱导因子-1 α(HIF-1 α)对缺氧状态下血管内皮细胞的保护机制. 方法 ①建立小鼠失血性休克模型,选取休克组与假手术组(n=3)血管内皮细胞,进行转录组测序,分析主要差异基因.②检测对照组(正常培养)与缺氧组(缺氧培养6h)(n=3)中人脐静脉内皮细胞(HUVECs)的HIF-1α及葡萄糖转运蛋白1(GLUT1)的表达变化,以及线粒体膜电位变化.③干扰组HUVECs转染HIF-1α的siRNA 48 h,干扰HIF-1α表达,对照组转染对照siRNA(n=3),检测HIF-1α的表达以确定干扰效果.干扰成功后缺氧培养6h后检测干扰组和对照组(n=3)GLUT1的表达情况,荧光探针法检测线粒体膜电位变化. 结果 ①休克组与假手术组血管内皮细胞存在明显差异的基因有25条.假手术组和休克组HIF-1α表达量分别为53.49±3.26、111.70±15.97,差异有统计学意义(P=0.023).②缺氧组HUVECs中HIF-1α和GLUT1的表达均升高,与对照组比较差异均有统计学意义(P<0.05).缺氧组和对照组线粒体膜电位分别为0.781±0.023、1.177±0.062,差异有统计学意义(P<0.05).③干扰组和对照组线粒体膜电位分别为1.011±0.076、1.151±0.031,差异无统计学意义(P>0.05);但是予缺氧培养6h后,干扰组和对照组线粒体膜电位分别为0.514±0.018、0.769±0.044,差异有统计学意义(P<0.05). 结论 HIF-1 α在失血性休克状态下表达明显升高,在缺氧状态下HUVECs中HIF-1α可上调GLUT1的表达,促进葡萄糖转运,改善线粒体损伤,保护血管内皮细胞.靶向调控HIF-1α可能有助于失血性休克的治疗.
      Abstract:
      Objective To investigate the protective mechanism of hypoxia-inducible factor-1α (HIF-1α) in vascular endothelial cells under hypoxia.Methods 1.After a hemorrhagic shock model was established in mice,the vascular endothelial cells were sorted in a shock group (n =3) and a sham operation group (n =3) for RNA-sequencing to analyze the main differential molecules.2.The expression of HIF-1α and glucose transporter-1 (GLUT1) was measured in human umbilical vein endothelial cells (HUVECs) and the mitochondrial membrane potentials were detected in a control group (normal culture,n =3) and a hypoxia group (hypoxia culture for 6 h,n =3).3.The HUVECs cells were transfected with HIF-1α siRNA for 48 h to interfere with HIF-1 α expression,and the control group(n =3) was transfected with control siRNA.The expression of HIF-1α was detected to determine the interference effect.The mRNA and protein expression of GLUT1 was detected in the interference and the control groups after 6 h of hypoxia culture.The mitochondrial membrane potential was detected by fluorescent probe method.Results 1.The transcriptome sequencing in the vascular endothelial cells in the shock and sham operation groups indicated 25 genes with significant differences.The HIF-1 α expression was significantly increased in the shock group (111.70 ± 15.97) than in the sham operation group (53.49 ± 3.26) (P =0.023).2.The expression of HIF-1 α and GLUT1 in the HUVECs cells was significantly increased in the hypoxia group compared with the control group (P < 0.05).The mitochondrial membrane potential was significantly lower in the hypoxia group (0.781 ± 0.023) than in the control group (1.177 ± 0.062) (P < 0.05),indicating aggravated injury.3.There was no significant difference in the mitochondrial membrane potential between the interference group (1.011 ± 0.076) and the control group (1.151 ± 0.031) (P > 0.05).However,after hypoxia culture for 6 h,there was a significant difference in the mitochondrial membrane potential between the interference group (0.514 ± 0.018) and the control group (0.769 ± 0.044) (P < 0.05),also indicating aggravated damage.Conclusions The expression of HIF-1 α may be significantly increased in hemorrhagic shock.In HUVECs under hypoxia,HIF-1 α may up-regulate the expression of GLUT1,promote glucose transport,improve mitochondrial damage and protect vascular endothelial cells.Thus,targeting HIF-1 α may contribute to the treatment of hemorrhagic shock.
      作者: 陈实 [1] 范金柱 [2] 李忠 [3] 谢建刚 [4]
      Author: Chen Shi [1] Fan Jinzhu [2] Li Zhong [3] Xie Jiangang [4]
      作者单位: 西安交通大学附属红会医院急诊科 710054 西安交通大学附属红会医院骨显微修复外科 710054 西安交通大学附属红会医院创伤骨科 710054 空军军医大学附属西京医院急诊科,西安,710032
      刊 名: 中华创伤骨科杂志 ISTICPKU
      年,卷(期): 2019, 21(3)
      在线出版日期: 2019年5月9日
      基金项目: 陕西省重点研发计划一般项目-社会发展领域(S2017-ZDYF-YBXM-SF-0442)Key Research and Development Programs of Science and Technology in Shaanxi Province-Projects in Social Development