Nitro-fatty acids possess anti-atherogenic properties, but their effects on macrophage oxidative status and lipid metabolism that play important roles in atherosclerosis development are unclear. This study compared the effects of nitro-oleic acid (OLA-NO2) with those of native oleic acid (OLA) on intracellular reactive oxygen species (ROS) generation, anti-oxidants and metabolism of triglycerides and cholesterol in J774A.1 macrophages. Upon incubating the cells with physiological concentrations of OLA-NO2 (0-1 µM) or with equivalent levels of OLA, ROS levels measured by 2, 7-dichlorofluorescein diacetate, decreased dose-dependently, but the anti-oxidative effects of OLA-NO2 were significantly augmented. Copper ion addition increased ROS generation in OLA treated macrophages without affecting OLA-NO2 treated cells. These effects could be attributed to elevated glutathione levels and to increased activity and expression of paraoxonase2 that were observed in OLA-NO2 vs OLA treated cells. Beneficial effects on triglyceride metabolism were noted in OLA-NO2 vs OLA treated macrophages in which cellular triglycerides were reduced due to attenuated biosynthesis and accelerated hydrolysis of triglycerides. Accordingly, OLA-NO2 treated cells demonstrated down-regulation of diacylglycerol acyltransferase1, the key enzyme in triglyceride biosynthesis, and increased expression of hormone-sensitive lipase and adipose triglyceride lipase that regulate triglyceride hydrolysis. Finally, OLA-NO2 vs OLA treatment resulted in modest but significant beneficial effects on macrophage cholesterol metabolism, reducing cholesterol biosynthesis rate and low density lipoprotein influx into the cells, while increasing high density lipoprotein-mediated cholesterol efflux from the macrophages. Collectively, compared with OLA, OLA-NO2 modestly but significantly reduces macrophage oxidative status and cellular triglyceride content via modulation of cellular anti-oxidants and triglyceride metabolizing enzymes.
Keywords: Antioxidants; Atherosclerosis; Cholesterol metabolism; Fatty acid modifications; Free radicals; Triglyceride metabolism.