Cell-biological and molecular biological analyzes of reverse cholesterol transport as a protective system against atherosclerosis

1 .胆固醇酯转移蛋白（CETP）缺乏症中脂蛋白功能异常分析CETP缺乏症中脂蛋白异常的特征是存在大量且富含胆固醇酯（CE）的HDL和小的多分散LDL。纯合子cetp缺失组获得的大且富含ce的hdl2对降低脂质巨噬细胞胆固醇含量的效果不如对照组。患者的小多分散LDL对正常人成纤维细胞LDL受体的亲和力降低。这些由CETP缺乏引起的血浆脂蛋白体外功能异常可能与动脉粥样硬化有关。ii . CETP缺乏的分子基础我们发现，在日本普通人群中，CETP基因中内含子14的5'剪接供体条G-to-A突变的频率约为1%。我们还在明显高脂蛋白血症患者的第15外显子（D442； G）上发现了一个新的错义突变。这一新的突变与日本的内含子14剪接缺陷一样普遍。iii . CETP缺陷的动脉粥样硬化性冠心病（CHD）和青少年角膜混浊的高脂蛋白血症患者（Atherosclerosis 53; 207-212, 1984）被鉴定为外显子15错义突变的纯合。我们还发现，低HTGL的cetp缺乏患者可能易患冠心病。我们发现了一个独特的区域，在那里由内含子14剪接缺陷引起的显著高脂蛋白血症（HALP）明显累积。在这一区域，老年存活者的HALP和内含子14 CETP基因突变的频率低于年轻存活者。这些结果提示CETP缺乏可能不是一种长寿综合征。iv .原发性胆汁性肝硬化（PBC）脂蛋白异常已知PBC常与HALP和黄瘤相关。我们发现，与CETP缺乏相比，PBC高脂蛋白血症患者血浆CETP水平显著升高，HTGL活性降低。v . HDL与肝细胞的相互作用，反向胆固醇转运的终端我们发现，HDL被人肝癌细胞系Mahlavu吸收并降解，而在肝外周细胞中，HDL不发生降解。结果表明，高密度脂蛋白相关胆固醇可能通过不同于低密度脂蛋白代谢的途径加工。少

I.Analysis of abnormal function of lipoproteins in cholesteryl ester transfer protein (CETP) deficiencyLipoprotein abnormalities in CETP deficiency were characterized by the presence of large and cholesteryl-ester(CE)-rich HDL and small polydisperse LDL.Large and CE-rich HDL_2 obtained from homozygous CETP-deficient subjects was less effective for reducing cholesterol content in lipid-laden macrophages than that from control subjects. The small polydisperse LDL of patients had reduced affinity for the LDL receptor of normal human fibroblasts. These abnormal in vitro function of plasma lipoproteins from CETP deficiency may be associated with athrosclerosis.II.Molecular basis of CETP deficiencyWe found that the frequency of the G-to-A mutation at the 5' splice donor slite of intron 14 in the CETP gene was approximately 1% in the Japanese general population. We also found a novel missense mutation in exon 15(D442 ; G) in markedly hyperalphalipoproteinemic patients. This novel mutation was … More revealed to be as common as the intron 14 splicing defect in Japan.III.Atherogenicity in CETP deficiencyThe hyperalphalipoproteinemic subjects with coronary heart disease (CHD) and juvenile corneal opacification (Atherosclerosis 53 ; 207-212, 1984), were identified to be homozygous for the exon 15 missense mutation. We also found that CETP-deficient patients with low HTGL may be susceptible to CHD.We found a unique area where a marked hyperalphalipoproteinemia (HALP) caused by the intron 14 splicing defect was markedly accumulated. In this area, a marked HALP and the intron 14 CETP gene mutation were less frequentry observed in the eiderly survived subjects than in the younger subjects. These results suggested that CETP deficiency may not be a longevity syndrome.IV.Lipoprotein abnormalities in primary biliary cirrhosis (PBC)PBC was known to be often associated with HALP and xanthomas.We found that plasma levels of CETP were markedly increased and HTGL activities were reduced in hyperalphalipoproteinemic patients with PBC, in contrast to CETP deficincy.V.Interaction between HDL and hepatocytes, a terminal of reverce cholesterol transportWe found that HDL is taken up and degraded by a human hepatoma cell line, Mahlavu, in contrast to extrahepatic peripheral cells, in which the degradation of HDL does not occur. The results indicated that HDL-associated cholesterol may be processed via a pathway different from that of LDL metabolism. Less