Homeostasis and its pathology of cellular lipid

本研究利用三种模型系统研究了载脂蛋白-细胞相互作用介导的HDL组装系统。1）小鼠白血病细胞系RAW 264中载脂蛋白相互作用位点的研究。必须激活一种激酶，使该细胞表达与载脂蛋白的结合位点，并使HDL与细胞脂质结合。完全表达需要几个小时，其被各种蛋白质合成的代谢抑制剂抑制。因此，提出了膜蛋白的贡献。有趣的是，ABC 1的表达没有明显差异，ABC 1已被证明是家族性HDL缺乏症中的突变位点。2）在人白血病细胞系THP-1中与HDL组装系统相关的细胞内胆固醇运输的研究。THP-1细胞在分化前仅通过载脂蛋白生成细胞磷脂和HDL，而PMA诱导分化后HDL富含胆固醇。ABC 1和小窝蛋白-1都被PMA诱导，并且反义DNA选择性地降低所产生的HDL中的胆固醇含量。因此，小窝蛋白-1负责将胆固醇掺入HDL中。3）通过使用载脂蛋白-细胞相互作用的抑制剂普罗布考在小鼠模型中研究HDL组装系统的生理相关性。普罗布考可迅速而显著地降低小鼠HDL，而HDL相关蛋白（包括ABC 1）的含量和HDL清除率无显著变化。普罗布考可显著降低载脂蛋白和腹腔巨噬细胞的HDL组装，表明这是小鼠血浆HDL的主要来源。

We studied the HDL assembly system mediated by apolipoprotein-cell interaction by using three model systems.1) Study on the apolipoprotein interaction site in mouse leukemic cell line cells RAW 264. A kinase must be activated for this cell to express the binding site with apolipoprotein and to gegerate HDL with cellular lipid. It takes several hours for the full expression which is suppressed by various metabolic inhibitors for protein synthesis. Therefore, contribution of a membrane protein(s) is suggested. Interestingly, there was no apparent difference in expression of ABC1 which has been shown as a mutation site in familial HDL deficiencies.2) Study on intracellular cholesterol trafficking linked to the HDL assembly system in human leukemic cell line cells THP-1. While THP-1 cells gegerate HDL only with cellular phospholipid by apolipoprotein before differentiation, HDL is enriched with cholesterol after PMA induces differentiation. Both ABC1 and caveolin-1 are induced by PMA, and the antisense DNA selectively reduces cholesterol content in the HDL generated. Therefore, caveolin-1 is responsible for incorporation of cholesterol into the HDL.3) Study on physiological relevance of the HDL assembly system in a mouse model by using an inhibitor of the apolipoprotein-cell interaction, probucol. Mouse HDL is rapidly and remarkably decreased by probucol, while no significant change is demonstrated in the massages of HDL-relating proteins including ABC1 and in the HDL clearance. The HDL assembly by apolipoprotein and the peritoneal macrophage is markedly decreased by probucol, showing that this is a major source of plasma HDL in mice.