Increased macrophage glutathione content reduces cell-mediated oxidation of LDL and atherosclerosis in apolipoprotein E-deficient mice

We used the apolipoprotein E deficient (apo e−/−) mice to analyze the role of macrophage reduced glutathione (GSH) content in cell-mediated oxidation of LDL and in atherosclerotic lesion development. Apo e−/− mice were supplemented with -2-oxo-4-thiazolidin carboxylate (OTC, which supplies cysteine residues, 500 mg/kg/day), or with buthionine sulfoximine (BSO, a specific inhibitor of GSH synthesis, 400 mg/kg/day) for 6 weeks. Then mouse peritoneal macrophages (MPM) and the mice aortas were collected. MPM from apo e−/− mice contained decreased GSH levels (by 58%), and a four-fold increased lipid peroxides content compared to control macrophages from C57BL6 mice. These MPM demonstrated increased capability to release superoxide anions and to oxidize LDL in comparison to control MPM. OTC supplementation resulted in a 26% increase in macrophage GSH, paralleled by a 25% reduction in cellular lipid peroxides content. Decrement by 30% in superoxide anion release and LDL oxidation by MPM, and also in the atherosclerotic lesion size by 25%, was found in the OTC-treated mice, compared to placebo-treated apo e−/− mice. In contrast, in BSO-treated mice MPM a further depletion of cellular GSH by 22% was found, paralleled by a two-fold increase in lipid peroxides content, and a 41% increased superoxide anion release and cell-mediated LDL oxidation, compared to placebo-treated apo e−/− mice MPM. Most important, BSO supplementation to apo e−/− mice caused a 59% increase in the atherosclerotic lesion area. An additional way to increase cellular GSH content was the use of dietary antioxidants. Vitamin E (40 mg/kg/day) or the isoflavan glabridin (25 μg/kg/day) administration for 2 months to apo e−/− mice resulted in the accumulation of these antioxidants in their MPM, and increased MPM GSH content by 24 and 80%, respectively. MPM lipid peroxides content was reduced by 31 or 60% upon vitamin E or glabridin supplementation, paralleled by a 30 or 60% decrease in cell-mediated oxidation of LDL, respectively. Finally, a significant inverse correlation (R=0.83) was found between macrophage GSH content and cell-mediated oxidation of LDL. We conclude that enrichment in vivo of macrophages with GSH, significantly decreases cellular oxidative stress, leading to reduced capability of the macrophages to oxidize LDL, and hence GSH may attenuate the development of atherosclerosis.