Rozenberg Orit, Rosenblat Mira, Coleman Raymond, Shih Diana M, Aviram Michael
Lipid Research Laboratory, Department of Anatomy and Cell Biology, the Technion Faculty of Medicine, The Rappaport Family Institute for Research in the Medical Sciences and Rambam Medical Center, Haifa, Israel.
Free Radic Biol Med. 2003 Mar 15;34(6):774-84. doi: 10.1016/s0891-5849(02)01429-6.
Human serum paraoxonase (PON1), an HDL-associated esterase, protects lipoproteins against oxidation, probably by hydrolyzing specific lipid peroxides. As arterial macrophages play a key role in oxidative stress in early atherogenesis, the aim of the present study was to examine the effect of PON1 on macrophage oxidative stress. For this purpose we used mouse arterial and peritoneal macrophages (MPM) that were harvested from two populations of PON1 knockout (KO) mice: one on the genetic background of C57BL/6J (PON1(0)) and the other one on the genetic background of apolipoproteinE KO (PON1(0)/E(0)). Serum and LDL, but not HDL, lipids peroxidation was increased in PON1(0), compared to C57BL/6J mice, by 84% and by 220%, respectively. Increased oxidative stress was shown in peritoneal and in arterial macrophages derived from either PON1(0) or PON1(0)/E(0) mice, compared to their appropriate controls. Macrophage oxidative stress was expressed by increased lipid peroxides content in MPM from PON1(0) and from PON1(0)/E(0) mice by 48% and by 80%, respectively, and by decreased reduced glutathione (GSH) content, compared to the appropriate controls. Furthermore, increased capacity of MPM from PON1(0) and PON1(0)/E(0) mice to oxidize LDL (by 40% and by 19%, respectively) and to release superoxide anions was observed. In accordance with these results, PON1(0) mice MPM exhibited 130% increased translocation of the cytosolic p47phox component of NADPH-oxidase to the macrophage plasma membrane, suggesting increased activation of macrophage NADPH-oxidase in PON1(0) mice, compared to control mice MPM. The increase in oxidative stress in PON1-deficient mice was observed despite the presence of the two other members of the PON gene family. PON2 and PON3 activities and mRNA expression were both found to be present in PON1-deficient mice MPM. Upon incubation of PON1(0)/E(0) derived macrophages with human PON1 (7.5 arylesterase units/ml), cellular peroxides content was decreased by 18%, macrophage superoxide anion release was decreased by 33%, and macrophage-mediated oxidation of LDL was reduced by 22%. Finally, a 42% increase in the atherosclerotic lesion area was observed in PON1(0)/E(0) mice, in comparison to E(0) mice under regular chow diet. We thus concluded that PON1 can directly reduce oxidative stress in macrophages and in serum, and that PON1-deficiency results in increased oxidative stress not only in serum, but also in macrophages, a phenomenon that can contribute to the accelerated atherosclerosis shown in PON1-deficient mice.
人血清对氧磷酶（PON1）是一种与高密度脂蛋白（HDL）相关的酯酶，可能通过水解特定的脂质过氧化物来保护脂蛋白免受氧化。由于动脉巨噬细胞在早期动脉粥样硬化的氧化应激中起关键作用，本研究的目的是检测PON1对巨噬细胞氧化应激的影响。为此，我们使用了从小鼠动脉和腹腔巨噬细胞（MPM）中分离出的两种PON1基因敲除（KO）小鼠：一种是C57BL/6J遗传背景（PON1(0)），另一种是载脂蛋白E基因敲除的遗传背景（PON1(0)/E(0)）。与C57BL/6J小鼠相比，PON1(0)小鼠的血清和低密度脂蛋白（LDL）（而非HDL）的脂质过氧化分别增加了84%和220%。与相应对照组相比，PON1(0)或PON1(0)/E(0)小鼠来源的腹腔和动脉巨噬细胞均表现出氧化应激增加。PON1(0)和PON1(0)/E(0)小鼠的MPM中脂质过氧化物含量分别比相应对照组增加了48%和80%，还原型谷胱甘肽（GSH）含量降低，这表明巨噬细胞氧化应激增强。此外，观察到PON1(0)和PON1(0)/E(0)小鼠的MPM氧化LDL的能力（分别增加40%和19%）以及释放超氧阴离子的能力增强。与这些结果一致，PON1(0)小鼠的MPM中NADPH氧化酶的胞质p47phox成分向巨噬细胞质膜的转位增加了130%，这表明与对照小鼠的MPM相比，PON1(0)小鼠的巨噬细胞NADPH氧化酶激活增加。尽管存在PON基因家族的另外两个成员，但仍观察到PON1缺陷小鼠的氧化应激增加。PON1缺陷小鼠的MPM中同时存在PON2和PON３的活性及mRNA表达。用人类PON1（7.5芳基酯酶单位/毫升）孵育PON1(0)/E(0)来源的巨噬细胞后，细胞过氧化物含量降低了18%，巨噬细胞超氧阴离子释放减少了33%，巨噬细胞介导的LDL氧化减少了22%。最后，与正常饮食的E(0)小鼠相比，PON1(0)/E(0)小鼠的动脉粥样硬化病变面积增加了42%。因此，我们得出结论，PON1可以直接降低巨噬细胞和血清中的氧化应激，PON1缺乏不仅会导致血清中氧化应激增加，还会导致巨噬细胞中氧化应激增加，这种现象可能导致PON1缺陷小鼠动脉粥样硬化加速。
