低氧条件下沉默HIF-1α与TNF-α、IL-6的相关性研究及人真皮微血管内皮细胞生物学行为的变化  

The Relationship among HIF-1α, TNF-α and IL-6 and the Change of the Biological Behavior of Human Dermal Microvascular Endothelial Cells after Silencing HIF-1α Gene under Hypoxia

作　　者：罗鸿 桂福强 刘洪[1] 张矛[1] 赵渝[1] 

机构地区：[1]重庆医科大学附属第一医院血管外科,重庆400016

出　　处：《中国细胞生物学学报》2018年第1期62-70,共9页Chinese Journal of Cell Biology

基　　金：重庆市卫生计划生育委员会医学科研项目(批准号:20141002)资助的课题~~

摘　　要：该文探讨了低氧条件下沉默缺氧诱导因子-1α(hypoxia inducible factor-1α,HIF-1α)与肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白介素-6(interleukin-6,IL-6)的相关性及人真皮微血管内皮细胞生物学行为的变化。采用三气培养箱培养人真皮微血管内皮细胞,根据时间和氧浓度分组,确定最适培养时间及缺氧浓度。同时,构建靶向HIF-1α的腺病毒并感染人真皮微血管内皮细胞,分为对照组、空载组和沉默组。应用RT-PCR和Western blot检测HIF-1α、TNF-α和IL-6的mRNA和蛋白质水平。Transwell检测细胞迁移能力,流式细胞技术观察细胞凋亡及细胞周期变化。结果显示,缺氧条件下,HIF-1α、TNF-α和IL-6的表达随缺氧时间延长而增加,随氧浓度降低而增加。与对照组相比,沉默组的细胞迁移能力(P<0.01)及周期(P<0.01)均明显抑制,细胞凋亡明显增加(P<0.01)。与对照组相比,沉默组TNF-α和IL-6的mRNA和蛋白质水平均减少(P<0.01)。由此可见,缺氧条件下,HIF-1α、TNF-α和IL-6的表达呈时间依赖性和氧浓度递减依赖性。人真皮微血管内皮细胞HIF-1α的表达与炎性因子TNF-α、IL-6具有明显的正相关。沉默HIF-1α后,能明显抑制细胞增殖,抑制细胞迁移,促进细胞凋亡。This work is aimed to study the relationship among hypoxia inducible factor-1α（HIF-1α）, tumor necrosis factor-α（TNF-α） and interleukin-6（IL-6） and observe the change of the biological behavior of human dermal microvascular endothelial cells（HDMECs） after silencing HIF-1α gene under hypoxia. A three-gas incubator was used to cultured the HDMECs, which were grouped by the time and oxygen concentration to determine the optimum culture time and hypoxia concentration. Meanwhile, adenovirus targeting HIF-1α was constructed and infected HDMECs. The HDMECs were divided into control group, non-HIF-1α-siRNA group and HIF-1α-siRNA group. RT-PCR and Western blot were then used to detect the mRNA and protein levels of HIF-1α, TNF-α and IL-6, respectively. The cell migration ability was detected by Transwell, and apoptosis and cycle changes were observed by flowcytometry. Under hypoxia, the gene expressions of HIF-1α, TNF-α and IL-6 increased with the increase of hypoxia time, and increased with the decrease of oxygen concentration. Compared with the control group, the cell migration ability（P〈0.01） and the cycle（P〈0.01） of the HIF-1α-siRNA group were significantly inhibited, and the apoptosis was significantly increased（P〈0.01）. Compared with the control group, the mRNA and protein levels of TNF-α and IL-6 genes in HIF-1α-siRNA group were decreased（P〈0.01）. So, under hypoxia, the gene expressions of HIF-1α, TNF-αand IL-6 were in time dependent and oxygen concentration decreasing dependent manner. The expression of HIF-1α in HDMECs was positively related to inflammatory factors TNF-α and IL-6. Silencing HIF-1α can obviously inhibit cell proliferation, inhibit cell migration and promote apoptosis.

关 键 词：缺氧诱导因子 人真皮微血管内皮细胞 静脉性溃疡 生物学行为 炎性因子 

分 类 号：R543.6[医药卫生—心血管疾病]