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目的:以对缺氧敏感的转录调控因子——缺氧诱导因子(HIF)-1α作为评价指标,比较物理方法与化学方法建立大鼠心肌微血管内皮细胞缺氧模型效果。方法:1植块法培养2周龄雄性SD大鼠心肌微血管内皮细胞,鉴定并传代培养。2在不同时间点分别以缺氧发生装置和氧耗剂Na2S2O4对心肌微血管内皮细胞进行缺氧干预,选择最佳干预时间。3免疫印迹和细胞免疫荧光法检测两种方法建立心肌微血管内皮细胞缺氧模型后,HIF-1α的表达情况。结果:1植块法培养原代心肌微血管内皮细胞纯度较高,可见典型铺路石样形态和管腔样结构;26hHIF-1α表达较其他时间点高,故选择6h作为最佳缺氧干预时间;3物理缺氧法干预心肌微血管内皮细胞后HIF-1α表达较化学缺氧法显著增加(P<0.05)。结论:对构建体外心肌微血管内皮细胞缺氧模型,物理法效果优于化学法。
OBJECTIVE: To hypoxia-inducible factor Hypoxia-inducible factor 1 (HIF-1α), an indicator of hypoxia, was used to evaluate the effects of HIF-1α in rat model of myocardial microvascular endothelial cell hypoxia by comparing physical and chemical methods. Methods: 1, 2-week-old male Sprague-Dawley rat myocardial microvascular endothelial cells were cultured, identified and subcultured. 2 at different time points were hypoxia device and oxygen consumption agent Na2S2O4 myocardial microvascular endothelial cells were hypoxic intervention, select the best intervention time. 3 Western blot and immunofluorescence detection of two methods to establish myocardial microvascular endothelial cell hypoxia model, the expression of HIF-1α. Results: 1 The culture of primary cultured myocardial microvascular endothelial cells with high purity showed typical paving stone-like morphology and lumen-like structure; 26h expression of HIF-1α was higher than other time points, so 6h was chosen as the best time of hypoxic intervention; Compared with the chemical hypoxia method, HIF-1α expression was significantly increased after physical hypoxia intervention in myocardial microvascular endothelial cells (P <0.05). CONCLUSION: Physical method is better than chemical method in establishing model of myocardial microvascular endothelial cell hypoxia in vitro.