Vila Andrew, Korytowski Witold, Girotti Albert W
Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
Biochemistry. 2002 Nov 19;41(46):13705-16. doi: 10.1021/bi026467z.
Under oxidative pressure in the vascular circulation, erythrocytes and phagocytic cells may accumulate membrane lipid hydroperoxides (LOOHs), including cholesterol- and phospholipid-derived species (ChOOHs, PLOOHs). LOOH translocation from cells to low-density lipoprotein (LDL) might sensitize the latter to free radical-mediated oxidative modification, an early event associated with atherogenesis. To test this, we examined the spontaneous transfer kinetics of various ChOOH species (5 alpha-OOH, 6 alpha-OOH, 6 beta-OOH, 7 alpha/7 beta-OOH) and various PLOOH groups (PCOOH, PEOOH, PSOOH, SMOOH) using photoperoxidized erythrocyte ghosts as model donors and freshly prepared LDL as an acceptor. LOOH departure or uptake was monitored by reverse-phase HPLC with reductive electrochemical detection. Mildly peroxidized ghost membranes transferred overall ChOOH and PLOOH to LDL with apparent first-order rate constants approximately 60 and approximately 35 times greater than those of the respective parent lipids. Individual ChOOH rate constants decreased in the following order: 7 alpha/7 beta-OOH > 5 alpha-OOH > 6 alpha-OOH > 6 beta-OOH. Kinetics for reverse transfer from LDL to ghosts followed the same trend, but rates were significantly higher for all species and their combined activation energy was lower (41 vs 85 kJ/mol). PLOOH transfer rate constants ranged from 4- to 15-fold lower than the composite ChOOH constant, their order being as follows: PCOOH approximately PEOOH approximately PSOOH > SMOOH. Similar PLOOH transfer kinetics were observed when LDL acceptor was replaced by unilamellar liposomes, consistent with desorption from the donor membrane being the rate-limiting step. The susceptibility of transfer LOOH-enriched LDL to Cu2+-induced chain peroxidative damage was assessed by monitoring the accumulation of conjugated dienes and products of free radical-mediated cholesterol oxidation. In both cases, transfer-acquired LOOHs significantly reduced the lag time for chain initiation relative to that observed using nonperoxidized ghosts. These findings are consistent with the idea that LDL can acquire significant amounts of "seeding" LOOHs via translocation from various donors in the circulation.
在血管循环中的氧化压力下，红细胞和吞噬细胞可能会积累膜脂质氢过氧化物（LOOHs），包括胆固醇衍生和磷脂衍生的种类（ChOOHs、PLOOHs）。LOOH从细胞转移到低密度脂蛋白（LDL）可能会使后者对自由基介导的氧化修饰敏感，这是与动脉粥样硬化发生相关的早期事件。为了验证这一点，我们使用光过氧化红细胞血影作为模型供体，新鲜制备的LDL作为受体，研究了各种ChOOH种类（5α - OOH、6α - OOH、6β - OOH、7α/7β - OOH）和各种PLOOH基团（PCOOH、PEOOH、PSOOH、SMOOH）的自发转移动力学。通过反相高效液相色谱结合还原电化学检测来监测LOOH的释放或摄取。轻度过氧化的血影膜将总的ChOOH和PLOOH转移到LDL上，其表观一级速率常数分别比各自的母体脂质大约高60倍和大约35倍。各个ChOOH的速率常数按以下顺序降低：7α/7β - OOH > 5α - OOH > 6α - OOH > 6β - OOH。从LDL到血影的逆向转移动力学遵循相同趋势，但所有种类的速率都显著更高，并且它们的综合活化能更低（41对85 kJ/mol）。PLOOH的转移速率常数比复合ChOOH常数低4至15倍，其顺序如下：PCOOH ≈ PEOOH ≈ PSOOH > SMOOH。当LDL受体被单层脂质体取代时，观察到类似的PLOOH转移动力学，这与从供体膜上解吸是限速步骤一致。通过监测共轭二烯的积累和自由基介导的胆固醇氧化产物，评估了转移获得的富含LOOH的LDL对Cu2 +诱导的链式过氧化损伤的敏感性。在这两种情况下，相对于使用未过氧化血影观察到的情况，转移获得的LOOH显著缩短了链引发的滞后时间。这些发现与以下观点一致，即LDL可以通过从循环中的各种供体进行易位而获得大量的“引发”LOOH。
