Study for the mechanism of LDL modification search for its inhibitor.

动脉粥样硬化早期病变的特征性事件之一是脂质泡沫细胞的积累。最近的研究表明，大多数泡沫细胞来源于单核巨噬细胞。这些巨噬细胞在其细胞质中积累脂质，其中大部分来自低密度脂蛋白（LDL）。由于这些细胞表达的经典LDL受体数量很少，因此LDL掺入巨噬细胞的机制一直是广泛研究的焦点。Goldstein等人报道了通过乙酰化化学修饰的LDL可以被巨噬细胞上的受体识别和结合，并将其命名为乙酰化LDL受体。然而，LDL的这种乙酰化似乎不太可能在体内发生。因此，一些研究者一直在研究体内LDL的修饰形式。内皮细胞修饰的LDL、氧化的LDL和香烟烟雾修饰的LDL现在被认为是自然发生的修饰LDL的候选者。最近有很多证据表明内皮细胞修饰的LDL和氧化的LDL是一样的。parthasarathi在体外观察到，通过Cu^<++>或Fe^<++>化学氧化将LDL转化为带负电荷的形式，将巨噬细胞转化为泡沫细胞，并提示巨噬细胞对这种氧化LDL的摄取机制是由乙酰-LDL受体介导的。然而在我们之前的研究中，我们发现乙酰-LDL对巨噬细胞降解[^<125>I]氧化LDL的抑制作用仅为55%。这一结果使我们推测这两种脂蛋白的摄取机制可能是不同的。我们目前的研究是通过比较氧化LDL、香烟烟雾修饰LDL和乙酰-LDL与巨噬细胞的结合特性来检验上述假设。为此，我们对巨噬细胞的结合和降解进行了竞争和交叉竞争。结果清楚地表明，修饰LDL的巨噬细胞受体至少由三种不同的受体组成，它们具有不同的特异性。其中两种对这三种低密度脂蛋白是特异性的，其余的是氧化低密度脂蛋白和香烟烟雾修饰低密度脂蛋白以及乙酰低密度脂蛋白的共同受体。在用Cu^<++>、Fe^<++>或香烟提取物修饰LDL的过程中，我们发现超氧化物阴离子（O_2）在修饰LDL过程中起重要作用，而超氧化物歧化酶（SOD）抑制了这些反应。但过氧化氢酶不能阻止LDL的修饰。因此，我们筛选出了几种抗氧化剂，如普罗布考、维生素E或维生素C衍生物，而不是S.O.D。到目前为止，普罗布考是唯一一种可以阻止LDL修饰的药物。其他药物不能阻止这些反应因为它们不能被纳入LDL颗粒中。接下来，我们用whhl -家兔进行了体内实验，以了解普罗布考作为抗氧化剂的抗动脉粥样硬化活性。从2月龄开始，连续17个月饲喂1%普罗布考饲料，主动脉动脉粥样硬化病变面积为23%，而对照组为93%。在兔8月龄病变面积为54%时，即使饲喂1%普罗布考日粮，6个月后病变面积平均约为38%。这些数据表明，氧化LDL是体内修饰LDL的候选之一，普罗布考具有很强的抗动脉粥样硬化作用。少

One of the characteristic events in the early lesions of atherosclerosis is the accumulation of lipid-laden foam cells. Recent studies have demonstrated that most of the foam cells are derived from monocute-macrophages. These macrophages accumulate lipids in their cytoplasms, most of which are derived from low density lipoproteins (LDL). Because these cells express very few number of classical LDL receptor, the mechanism of LDL incorporation into macrophages has been a focus of extensive investigation.Goldstein et al. reported that LDL modified chemically by acetylation can be recognized and incorporated by a receptor on macrophages and they named it acetyl-LDL receptor. However, it seems unlikely that such acetylation of LDL would occur in vivo. Therefore, several investigators have been studying what are modified forms of LDL in vivo. Endothelial cell modified LDL, oxidized LDL and cigarette smoke modified LDL are now proposed as a candidate for naturally occurring modified LDL. And … More recently there are lots of evidences that endothelial cell modified LDL is as same as oxidized LDL. Parthasarathy have observed in vitro that LDL converted by chemical oxidation with Cu^<++> or Fe^<++> into a negatively-charged form transforms macrophages into foam cells and suggested that the uptake mechanism of this oxidized LDL by macrophages is mediated by the acetyl-LDL receptor. However in our previous study, we found that acetyl-LDL inhibited the degradation of [^<125>I]oxidized LDL by macrophages only by 55 %. This result has led us to postulate that the uptake mechanisms of the two lipoproteins may be different. 0ur current study was undertaken to examine the above hypothesis by comparing the binding properties of oxidized LDL, cigarette-smoke modified LDL and acetyl-LDL to macrophages. To this, we have performed competition and cross-competition in binding and degradation in macrophages. The results clearly showed that the macrophage receptors for modified LDL are composed of at least three distinct receptors with different specificities. Two of them are specific for those three LDL and the rest is a common receptor for both oxidized and cigarette smoke modified LDL, and acetyl-LDL. In the process of modifying LDL by Cu^<++>, Fe^<++> or cigarette extract, we found that superoxide anion (O_2) played an important role for modification and super-oxide dismutase (SOD) inhibited those reactions. However catalase could not prevent the modification of LDL. Therefore, in stead of S.O.D., we screened out several agents as an antioxidant, such as probucol, vitamin E or vitamin C derivatives.Probucol, so far, is an only agents which could prevent the modification of LDL. Other agents could not prevent those reaction because those could not be included into LDL particles. We next performed in vivo experiment to ask antiatherogenic activity of probucol as an antioxidant using WHHL-rabbits. When rabbits were fed 1 % probucol diet for 17 months since they were 2 month of age, the surface area of atheromatous lesion in aorta was 23 % in comparison with that of 93 % in control WHHL-rabbit. Even when rabbits were fed 1 % probucol diet at 8 month of age when their surface area of lesion was 54 %, 6 month later their lesion became about 38 % on an average. Those date indicated that oxidized LDL is one of the candidate modified LDL in vivo and probucol has a strong antiatherogenic effect. Less