缺氧对体外培养动脉内皮细胞中诱生型一氧化氮合酶表达和细胞形态的影响

Effect of hypoxia on the expression of inducible nitric oxide synthase and morphology of arterial endothelial cells cultured in vitro

目的观察体外培养的动脉内皮细胞在缺氧条件下诱生型一氧化氮合酶的表达和细胞形态的变化情况。方法实验于2004-09/11在哈尔滨医科大学组织学与胚胎学教研室实验室完成。原代培养牛动脉内皮细胞,将培养的内皮细胞随机分为5组,即常氧组、缺氧0.5,1,2,3h组。每组切片选取12个400倍视野进行观察。诱生型一氧化氮合酶的表达采用免疫组织化学法测定。用计算机图文分析系统对各组切片内皮细胞诱生型一氧化氮合酶阳性表达的平均积分吸光度和缺氧后的细胞形态进行分析。结果①缺氧1h,2h,3h组的平均积分吸光度显著高于常氧组[0.1008±0.019,0.1179±0.037,0.1964±0.023,0.0735±0.032(t=-10.364～2.133,P<0.05～0.01)]。②缺氧后细胞的最大直径和最小直径均变小,核浆比例增大,细胞数量减少。结论缺氧后动脉内皮细胞的生长数量和形态发生明显变化,使其屏障作用降低;同时诱生型一氧化氮合酶表达的增加提示诱生型一氧化氮合酶在缺氧后内皮细胞的损伤中发挥重要作用。

AIM： To observe the changes of the expression of inducible nitric oxide synthase （iNOS） and morphology of pulmonary artery endothelial cells （PAEC） under the hypoxia cultured in vitro. METHODS： The experiment was conducted at the Staff Room of Histology and Embryology, Harbin Medical University from September to November 2004. Bovine artery endothelial cells were done the primary cuhure and divided randomly into 5 groups： normal group and hypoxic 0.5, 1, 2 and 3 hours groups. Twelve sections from every＇ group were selected to observe with 400-power field. The expression of the iNOS was detected with immunohistochemical method. The mean integral absorbance （A） and the morphology of cells after hypoxia of positive expression of iNOS were analyzed by computer image analysis system. RESULTS： （1） The mean integral A in the hypoxia 1, 2 and 3 hours groups was higher significantly than that in the normal group [0.100 8±0.019, 0.117 9±0.037,0.196 4±0.023,0.073 5±0.032 （t=-10.364 to 2.133.P 〈 0.05-0.01 ）]. （2） The maximal diameter and the minimal diameter of hypoxie cells became smaller with inereased karyoplasms and decreased number of cells. CONCLUSION： After hypoxia the number of artery endothelial cells and the morphology, change significantly, which make the barrier effect decrease. Meanwhile, the increase of the expression of iNOS indicates that the iNOS plays an important role in hypoxic endothelial cells injury.