Gerry Andrew B, Satchell Leanne, Leake David S
Cardiovascular Research Group, Biomolecular Sciences Section, School of Biological Sciences, University of Reading, Whiteknights, PO Box 228, Reading, Berkshire RG6 6AJ, UK.
Atherosclerosis. 2008 Apr;197(2):579-87. doi: 10.1016/j.atherosclerosis.2007.08.026. Epub 2007 Oct 22.
LDL oxidation may be important in atherosclerosis. Extensive oxidation of LDL by copper induces increased uptake by macrophages, but results in decomposition of hydroperoxides, making it more difficult to investigate the effects of hydroperoxides in oxidised LDL on cell function. We describe here a simple method of oxidising LDL by dialysis against copper ions at 4 degrees C, which inhibits the decomposition of hydroperoxides, and allows the production of LDL rich in hydroperoxides (626+/-98 nmol/mg LDL protein) but low in oxysterols (3+/-1 nmol 7-ketocholesterol/mg LDL protein), whilst allowing sufficient modification (2.6+/-0.5 relative electrophoretic mobility) for rapid uptake by macrophages (5.49+/-0.75 microg (125)I-labelled hydroperoxide-rich LDL vs. 0.46+/-0.04 microg protein/mg cell protein in 18 h for native LDL). By dialysing under the same conditions, but at 37 degrees C, the hydroperoxides are decomposed extensively and the LDL becomes rich in oxysterols. This novel method of oxidising LDL with high yield to either a hydroperoxide- or oxysterol-rich form by simply altering the temperature of dialysis may provide a useful tool for determining the effects of these different oxidation products on cell function.
低密度脂蛋白（LDL）氧化在动脉粥样硬化中可能起重要作用。铜对LDL的广泛氧化会诱导巨噬细胞摄取增加，但会导致氢过氧化物分解，使得研究氧化LDL中的氢过氧化物对细胞功能的影响变得更加困难。我们在此描述一种简单的方法，即在4℃下通过与铜离子透析来氧化LDL，这种方法可抑制氢过氧化物的分解，并能产生富含氢过氧化物（626±98 nmol/mg LDL蛋白）但氧化甾醇含量低（3±1 nmol 7-酮胆固醇/mg LDL蛋白）的LDL，同时允许进行足够的修饰（相对电泳迁移率为2.6±0.5）以便巨噬细胞快速摄取（18小时内，5.49±0.75 μg富含氢过氧化物的（125）I标记LDL，而天然LDL为0.46±0.04 μg蛋白/mg细胞蛋白）。在相同条件下于37℃透析时，氢过氧化物会大量分解，LDL会富含氧化甾醇。这种通过简单改变透析温度就能将LDL高产率地氧化为富含氢过氧化物或氧化甾醇形式的新方法，可能为确定这些不同氧化产物对细胞功能的影响提供一个有用的工具。
