Hoff H F, Clevidence B A
Exp Mol Pathol. 1987 Jun;46(3):331-44. doi: 10.1016/0014-4800(87)90054-2.
We have previously shown that a lipoprotein fraction consisting of large cholesteryl ester-rich particles can be isolated from homogenates of human aortic plaques by gel exclusion chromatography. This fraction was recognized by a high-affinity binding site on mouse peritoneal macrophages (MPM) resulting in unregulated uptake, stimulation of cholesterol esterification, and massive accumulation of cholesteryl esters. In this report we have further characterized such a fraction, designated lipid-protein complex (LP), which can be isolated from the void volume fraction of a Bio-Gel A-150m column following chromatography of plaque extracts. LP possessed a mean cholesterol-to-protein ratio of 2.3; it was heterogeneous in size and structure as observed by electron microscopy after negative staining, and it stimulated cholesterol esterification in MPM in a linear fashion over a 48-hr time interval, suggesting that the binding site on MPM recognizing LP was not down-regulated by intracellular cholesterol content. This uptake resulted in the presence of oil red O-positive intracellular droplets and numerous vacuoles containing electron-dense structures, whereas MPM incubated without lipoprotein showed few vacuoles or lipid droplets. Using SDS-PAGE and immunoblot and dot-blot techniques, we found that the major proteins associated with LP were albumin and fibronectin, whereas apoB and apoE were present in lower amounts. These proteins may be responsible for opsonization of LP, making it recognizable to receptors on MPM and facilitating LP uptake by MPM. LP isolated from tissue extracts without homogenization had the same structural and functional characteristics, suggesting that homogenization per se was not responsible for creating a particle that was recognized by MPM. However, homogenization yielded two to three times more LP. MPM uptake of LP derived from lysed foam cells may represent one of the mechanisms by which fatty streak lesions may grow to larger atherosclerotic lesions.
我们之前已经表明，通过凝胶排阻色谱法可从人主动脉斑块匀浆中分离出一种由富含胆固醇酯的大颗粒组成的脂蛋白组分。该组分可被小鼠腹膜巨噬细胞（MPM）上的高亲和力结合位点识别，从而导致不受调控的摄取、胆固醇酯化的刺激以及胆固醇酯的大量积累。在本报告中，我们进一步对这样一种组分进行了表征，其被命名为脂蛋白复合物（LP），可在对斑块提取物进行色谱分析后从Bio-Gel A-150m柱的空体积组分中分离得到。LP的平均胆固醇与蛋白质之比为2.3；经负染色后通过电子显微镜观察，其大小和结构具有异质性，并且在48小时的时间间隔内以线性方式刺激MPM中的胆固醇酯化，这表明MPM上识别LP的结合位点不会因细胞内胆固醇含量而下调。这种摄取导致出现油红O阳性的细胞内液滴以及许多含有电子致密结构的液泡，而未与脂蛋白一起孵育的MPM几乎没有液泡或脂滴。使用SDS-PAGE、免疫印迹和斑点印迹技术，我们发现与LP相关的主要蛋白质是白蛋白和纤连蛋白，而载脂蛋白B和载脂蛋白E的含量较低。这些蛋白质可能负责LP的调理作用，使其能够被MPM上的受体识别并促进MPM对LP的摄取。从未经匀浆的组织提取物中分离得到的LP具有相同的结构和功能特征，这表明匀浆本身并非产生被MPM识别的颗粒的原因。然而，匀浆产生的LP要多两到三倍。MPM对源自裂解泡沫细胞的LP的摄取可能代表了脂肪条纹病变发展为更大动脉粥样硬化病变的机制之一。
