Heart microvessels and aortic endothelial cells express the 15 kDa heart-type fatty acid-binding proteins
Heart microvessels and aortic endothelial cells express the 15 kDa heart-type fatty acid-binding proteins
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DOI：
10.1016/s0171-9335(98)80022-8
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发表时间：
1998-06-01
影响因子：
6.6
通讯作者：
Simionescu, M
中科院分区：
文献类型：
--
作者：
Antohe, F;Popov, D;Simionescu, M
由于它们的疏水性质，游离脂肪酸需要载体来跨细胞和在细胞内运输。内皮层是脂肪酸从血浆到下层细胞和组织的第一道屏障。我们试图找出是否胞质脂肪酸结合蛋白（FABP）存在于大血管内皮细胞（主动脉内皮细胞）和小血管（心肌毛细血管）使用以下实验方法：（i）分别用[C-14]棕榈酸酯或[C-14]油酸酯加载脱脂的主动脉内皮细胞（EC）匀浆和心脏胞质蛋白和膜蛋白，随后进行放射自显影检测电泳分离的条带;（ii）使用针对牛H-FABP产生的亲和纯化抗体通过免疫沉淀检测心脏型FABP（H-FABP）（抗H-FABP），和（iii）通过应用于培养的EC和小鼠心脏的厚和薄冷冻切片的间接免疫荧光和金免疫细胞化学定位FABP，结果显示：（i）在EC匀浆中，表达对[C-14]棕榈酸酯的亲和力的蛋白质具有15000和40000-45000的表观M-r，其分别对应于胞质和膜FABPs的分子量。通过与[C-14]油酸酯孵育发现类似的亲和力，其结合心脏胞质溶胶中的M-r 15000蛋白和膜部分中的40-45 kDa蛋白：（ii）抗H-FABP免疫沉淀特异性的胞质15 kDa肽（H-FABP）;（iii）通过间接免疫荧光，胞质H-FABP定位于心脏微血管和心肌细胞，也定位于培养的主动脉EC，其中存在胞浆抗原特征性的强烈斑点荧光;（iv）通过免疫细胞化学，在EC胞质中检测到H-FABP，并且在质膜和囊泡膜的胞质方面非常接近。这些数据共同证明了主动脉和心脏微血管内皮中存在15 kDa的心脏型FABP。
Due to their hydrophobic nature, free fatty acids require carriers for transport across and within the cells. The endothelial layer is the first barrier to be traversed by the fatty acids, from the plasma to the underlying cells and tissues. We tried to find out whether cytosolic fatty acid-binding proteins (FABPs) are present in the endothelium of large vessels (aortic endothelial cells) and small vessels (myocardial capillaries) using the following experimental approaches: (i) loading the delipidated aortic endothelial cell (EC) homogenate and the heart cytosolic proteins and membrane proteins with [C-14]palmitate or [C-14]oleate, respectively, followed by autoradiographic detection of electrophoretically separated bands; (ii) detection by immunoprecipitation of heart-type FABP (H-FABP) using an affinity-purified antibody raised against bovine H-FABP (anti-H-FABP), and (iii) localization of FABP by indirect immunofluorescence and gold-immune cyto chemistry applied to cultured EC and to thick and thin frozen sections of mouse heart, The results showed that: (i) within the EC homogenate proteins that express affinity for [C-14]palmitate have an apparent M-r of 15000, and 40000-45000, that correspond as molecular mass to cytosolic and membrane FABPs, respectively. Similar affinity was found by incubation with [C-14]oleate, that binds to a protein of M-r 15000 in the heart cytosol, and to a 40-45 kDa protein in the membrane fraction; (ii) anti-H-FABP immunoprecipitated specifically a cytosolic 15 kDa peptide (H-FABP); (iii) by indirect immunofluorescence, cytosolic H-FABP was localized on heart microvessels and myocytes and also in cultured aortic EC where intense spotted fluorescence characteristic for cytosolic antigens was present; (iv) by immunocytochemistry, H-FABP was detected in the EC cytoplasm, and in close proximity to the cytoplasmic aspect of plasmalemma and vesicle membranes. Together the data attest the presence of the 15 kDa, heart-type FABP in the endothelium of aorta and heart microvessels.