Wang W Q, Moses A S, Francis G A
CIHR Group on Molecular and Cell Biology of Lipids and Department of Medicine, University of Alberta, Edmonton, Alberta, Canada.
Biochemistry. 2001 Mar 27;40(12):3666-73. doi: 10.1021/bi002141j.
Despite very low plasma levels of HDL, carriers of the apolipoprotein AI Arg173 --> Cys mutation apoAI(Milano) (AIM) have no apparent increase in risk for atherosclerotic vascular disease. HDL apolipoprotein species in AIM carriers include apoAI-AII heterodimers, previously found to confer the enhanced ability of tyrosyl radical-oxidized HDL to mobilize cholesterol for removal from cultured cells. To determine whether enhanced mobilization of cholesterol by apoprotein species in AIM explains a cardioprotective action of this mutation, we examined the ability of lipid-free and lipid-bound AIM and AIM-AII heterodimers to deplete cholesterol from cultured cells. Free AIM and AIM-AII heterodimers showed a decreased capacity to act as acceptors of cholesterol from cholesterol-loaded human fibroblasts compared with native apoAI but similar capacities to deplete fibroblasts of the pool of cholesterol available for esterification by acyl-CoA:cholesterol acyltransferase (ACAT). Discoidal reconstituted HDL (rHDL) containing apoAI depleted both of these cholesterol pools more readily than AIM-containing rHDL when compared at equivalent rHDL protein levels, but similar abilities of these rHDL to deplete cell cholesterol were seen when compared at equivalent phospholipid levels. Spherical rHDL generated using the whole lipid fraction of HDL and apoAI or AIM showed similar capacities to deplete total and ACAT-accessible cell cholesterol when compared at similar protein levels, but an increased capacity of AIM-containing particles was seen when compared at equivalent phospholipid levels. Unlike the apoAI-AII heterodimer in tyrosylated HDL, AIM-AII heterodimer-containing spherical rHDL showed no increased capacity to deplete either of these pools of cholesterol. These results suggest a similar or better capacity of native apoAI in lipid-free or lipid-bound form in discoidal rHDL to enhance the mobilization of cellular cholesterol when compared to AIM in its free or lipid-bound forms. Any increase in depletion of cellular cholesterol by lipid-bound AIM in spherical rHDL appears related to altered phospholipid-binding rather than intrinsic cholesterol-mobilizing characteristics of this protein compared to native apoAI. The lack of major differences in these studies in cholesterol mobilization by native apoAI and AIM, or by apoAIM-AII heterodimers, suggests that any protection against atherosclerosis conferred by this mutation is likely related to other beneficial vascular effects of AIM.
尽管载脂蛋白AI的精氨酸173突变为半胱氨酸（apoAI[米兰]，AIM）携带者的血浆高密度脂蛋白（HDL）水平极低，但他们患动脉粥样硬化性血管疾病的风险并未明显增加。AIM携带者的HDL载脂蛋白种类包括apoAI-AII异源二聚体，此前发现这种异源二聚体可增强酪氨酰自由基氧化的HDL从培养细胞中转运胆固醇以供清除的能力。为了确定AIM中载脂蛋白种类增强的胆固醇转运能力是否解释了这种突变的心脏保护作用，我们检测了无脂和结合脂质的AIM以及AIM-AII异源二聚体从培养细胞中清除胆固醇的能力。与天然apoAI相比，游离的AIM和AIM-AII异源二聚体作为从胆固醇负载的人成纤维细胞中接受胆固醇的受体的能力降低，但在耗尽成纤维细胞中可被酰基辅酶A：胆固醇酰基转移酶（ACAT）酯化的胆固醇池方面能力相似。在相同的重组高密度脂蛋白（rHDL）蛋白水平下比较时，含有apoAI的盘状重组HDL比含有AIM的rHDL更容易耗尽这两个胆固醇池，但在相同的磷脂水平下比较时，这些rHDL清除细胞胆固醇的能力相似。使用HDL的全脂质部分和apoAI或AIM生成的球形rHDL在相似的蛋白水平下比较时，在耗尽总胆固醇和ACAT可及的细胞胆固醇方面能力相似，但在相同的磷脂水平下比较时，含有AIM的颗粒的能力有所增强。与酪氨酰化HDL中的apoAI-AII异源二聚体不同，含有AIM-AII异源二聚体的球形rHDL在耗尽这两个胆固醇池方面没有增强的能力。这些结果表明，与游离或结合脂质形式的AIM相比，盘状rHDL中无脂或结合脂质形式的天然apoAI在增强细胞胆固醇转运方面具有相似或更好的能力。球形rHDL中结合脂质的AIM在清除细胞胆固醇方面的任何增加似乎与磷脂结合的改变有关，而不是与该蛋白相对于天然apoAI的内在胆固醇转运特性有关。在这些研究中，天然apoAI和AIM或apoAIM-AII异源二聚体在胆固醇转运方面缺乏主要差异，这表明这种突变赋予的对动脉粥样硬化的任何保护作用可能与AIM的其他有益血管效应有关。
