Phenotypic properties and characteristics of superoxide production by mouse coronary microvascular endothelial cells
Phenotypic properties and characteristics of superoxide production by mouse coronary microvascular endothelial cells
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DOI：
10.1006/jmcc.2001.1372
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发表时间：
2001-06-01
影响因子：
5
通讯作者：
Shah, AM
中科院分区：
文献类型：
--
作者：
Li, JM;Mullen, AM;Shah, AM
冠状动脉微血管内皮细胞对心肌细胞的功能发挥（病理）生理作用，这可以使用纯细胞群进行实验研究。作为一个必要的先决条件，从基因修饰小鼠的细胞的调查，我们研究了冠状动脉微血管内皮细胞从正常小鼠分离的表型特性，和生化特征的超氧化物的生产这些细胞。用胶原酶、胰蛋白酶和DNA酶连续消化后，从失活的小鼠心室组织中分离微血管内皮细胞。采用差速离心、平板培养等方法从心肌细胞和其他细胞中分离出冠状动脉微血管内皮细胞。小鼠冠状动脉微血管内皮细胞在汇合时显示出不规则的“鹅卵石”形态，通过FACS分析对⑶ 31呈>98%阳性，并且通过共聚焦显微镜对VE-钙粘蛋白和内皮型一氧化氮合酶（eNOS）也呈阳性。细胞摄取荧光标记的乙酰化低密度脂蛋白，但对α-平滑肌肌动蛋白、结蛋白和细胞角蛋白呈阴性。与人内皮细胞不同，小鼠冠状动脉微血管内皮细胞仅弱表达血管性血友病因子。免疫印迹显示小鼠细胞表达吞噬细胞型NADPH氧化酶的组分。它们表现出NADPH依赖的O-2（-）-生成活性，这是由血管紧张素II增加，但完全抑制diphenyleniodonium。因此，小鼠冠状动脉微血管内皮细胞表达eNOS和NADPH氧化酶，它们之间的相互作用可能在内皮细胞病理生理学中发挥作用。(C)北京：科学出版社.
Coronary microvascular endothelial cells exert (patho)physiological effects on the function of cardiac myocytes, which may be studied experimentally using pure cell populations. As an essential pre-requisite to the investigation of cells from gene-modified mice, we studied the phenotypic properties of coronary microvascular endothelial cells isolated from normal mice, and biochemically characterized the superoxide production by these cells. Microvascular endothelial cells were isolated from devitalized mouse ventricular tissue after sequential digestion with collagenase, trypsin and DNase. Coronary microvascular endothelial cells were separated from cardiac myocytes and other cells by differential centrifugation, plating and culture. Mouse coronary microvascular endothelial cells showed an irregular "cobblestone" morphology at confluence, were >98% positive for CD31 by FACS analysis, and were also positive for VE-cadherin and endothelial-type nitric oxide synthase (eNOS) by confocal microscopy, The cells took up fluorescently labelled, acetylated low-density lipoprotein, but were negative for a alpha -smooth muscle actin, desmin and cytokeratin. Unlike human endothelial cells, mouse coronary microvascular endothelial cells only weakly expressed von Willebrand factor. immunoblotting showed that the mouse cells expressed components of a phagocyte-type NADPH oxidase. They exhibited NADPH-dependent O-2(-)-generating activity, which was increased by angiotensin II but completely inhibited by diphenyleneiodonium. Thus, mouse coronary microvascular endothelial cells express both eNOS and NADPH oxidase, interactions between which may play a role in endothelial cell pathophysiology. (C) 2001 Academic Press.