Guyard-Dangremont V, Lagrost L, Gambert P
Laboratoire de Biochimie des Lipoprotéines, INSERM CJF 93-10, Faculté de Médecine, Dijon, France.
J Lipid Res. 1994 Jun;35(6):982-92.
The aim of the present study was to determine in vitro the effects of various purified apolipoproteins (apo) on the activity of the cholesteryl ester transfer protein (CETP). It appeared that the ability of apoA-I, A-II, and A-IV to modulate the CETP-mediated transfer of radiolabeled cholesteryl esters between low density lipoproteins (LDL) and high density lipoproteins (HDL) was markedly influenced by the final apolipoprotein:lipoprotein ratio in incubation mixtures. At low apolipoprotein:lipoprotein ratio, the rate of radiolabeled cholesteryl esters transferred from HDL3 to LDL was significantly increased in the presence of apoA-I and apoA-IV. Under similar conditions, the rate of radiolabeled cholesteryl esters transferred from LDL to HDL3 was increased in the presence of apoA-I while apoA-IV had no significant effects. At high apolipoprotein:lipoprotein ratio, the ability of apoA-I and apoA-IV to enhance the rate of radiolabeled cholesteryl esters transferred either from HDL3 to LDL or from LDL to HDL3 was considerably reduced. At the highest apolipoprotein:lipoprotein ratio studied, apoA-I and A-IV became inhibitors of the CETP-mediated transfer reaction. Interestingly, apoA-II differed markedly from other apolipoproteins as, even at a low apolipoprotein:lipoprotein ratio, it significantly inhibited CETP activity as measured either from HDL3 to LDL or from LDL to HDL3. The inhibition by apoA-II was concentration-dependent and, at the highest apolipoprotein:lipoprotein ratio studied, cholesteryl ester transfer activity was totally suppressed. The possibility of a direct interaction between CETP and the two major HDL apolipoproteins, apoA-I and apoA-II, was further investigated by combining crosslinking and immunoblotting techniques. Whereas CETP alone had an apparent molecular mass of 76,000 +/- 3,100 Da, crosslinking reactions in incubation mixtures containing CETP and either apoA-I or apoA-II revealed the appearance of additional protein bands with apparent molecular masses of 99,600 +/- 6,100 and 86,900 +/- 4,500 Da, respectively. These complexes corresponded to the association of one molecule of CETP with one molecule of apoA-I or apoA-II. Interestingly, the mass concentrations of apoA-II needed to produce visible CETP-apolipoprotein complexes appeared to be about ten times higher as compared with apoA-I, suggesting that CETP may have a lower affinity for apoA-II than for apoA-I. In conclusion, data from the present study indicate that apolipoproteins A-I, A-II, and A-IV could be potent modulators of the CETP-mediated transfer of cholesteryl esters between HDL and LDL fractions.(ABSTRACT TRUNCATED AT 400 WORDS)
本研究的目的是在体外确定各种纯化的载脂蛋白（apo）对胆固醇酯转运蛋白（CETP）活性的影响。结果显示，apoA-I、A-II和A-IV调节CETP介导的放射性标记胆固醇酯在低密度脂蛋白（LDL）和高密度脂蛋白（HDL）之间转运的能力，受到孵育混合物中最终载脂蛋白与脂蛋白比例的显著影响。在低载脂蛋白与脂蛋白比例时，在apoA-I和apoA-IV存在的情况下，从HDL3转运至LDL的放射性标记胆固醇酯的速率显著增加。在类似条件下，在apoA-I存在时，从LDL转运至HDL3的放射性标记胆固醇酯的速率增加，而apoA-IV无显著影响。在高载脂蛋白与脂蛋白比例时，apoA-I和apoA-IV增强从HDL3转运至LDL或从LDL转运至HDL3的放射性标记胆固醇酯速率的能力大幅降低。在研究的最高载脂蛋白与脂蛋白比例时，apoA-I和A-IV成为CETP介导的转运反应的抑制剂。有趣的是，apoA-II与其他载脂蛋白明显不同，即使在低载脂蛋白与脂蛋白比例时，它也能显著抑制从HDL3到LDL或从LDL到HDL3的CETP活性。apoA-II的抑制作用呈浓度依赖性，在研究的最高载脂蛋白与脂蛋白比例时，胆固醇酯转运活性被完全抑制。通过结合交联和免疫印迹技术，进一步研究了CETP与两种主要HDL载脂蛋白apoA-I和apoA-II之间直接相互作用的可能性。单独的CETP的表观分子量为76,000±3,100 Da，而在含有CETP和apoA-I或apoA-II的孵育混合物中的交联反应显示，出现了表观分子量分别为99,600±6,100和86,900±4,500 Da的额外蛋白条带。这些复合物对应于一个CETP分子与一个apoA-I或apoA-II分子的结合。有趣的是，产生可见的CETP-载脂蛋白复合物所需的apoA-II的质量浓度似乎比apoA-I高约十倍，这表明CETP对apoA-II的亲和力可能低于对apoA-I的亲和力。总之，本研究的数据表明，载脂蛋白A-I、A-II和A-IV可能是CETP介导的HDL和LDL组分之间胆固醇酯转运的有效调节剂。（摘要截短为400字）
