Mitochondrial Oxidative Stress Mediates Macrophage Pro-inflammatory Metabolic Switch in Atherosclerotic Vascular Disease in Aging

Abstract

Aging elevates cardiovascular disease risk, including atherosclerosis. Macrophages play crucial role in vascular aging by promoting inflammation and atherosclerosis progression. Age-related increase in NOX4 NADPH oxidase expression correlates with mitochondrial dysfunction, inflammation, and atherosclerosis severity. We hypothesized that NOX4-dependent mitochondrial oxidative stress induces macrophage metabolic dysfunction and an inflammatory phenotype in aging-associated atherosclerotic disease. Aortic and brachiocephalic artery lesion areas were comparable in 5-month-old (young) Nox4-/-/Apoe-/- and Apoe-/- mice, increased significantly in 16-month-old (aged) mice, but were significantly lower in Nox4-/-/Apoe-/- versus Apoe-/- mice. In aged Nox4-/-/Apoe-/- mice, atherosclerotic lesions had reduced CD11b+ area, lower expression of CCL2, IL1β, and IL6, and fewer classically activated pro-inflammatory macrophages (CD38+CD80+). Notably, there was also an increased proportion of alternatively activated pro-resolving macrophages (CD163+CD206+). Spectral flow cytometry and t-SNE analysis revealed a significantly lower proportion of activated inflammatory macrophages and macrophage-like cells in atherosclerotic lesions of aged Nox4-/-/Apoe-/- compared to Apoe-/- mice. Macrophages from aged Apoe-/- mice had altered metabolic function. In contrast, macrophages from Nox4-/-/Apoe-/- mice were less glycolytic, more aerobic, and had preserved basal and maximal respiration and mitochondrial ATP production. Nox4-/-/Apoe-/- macrophages had lower mitochondrial ROS and reduced IL1β secretion, compared with Apoe-/- mice. In aged Apoe-/- mice, inhibition of NOX4 using GKT137831 significantly reduced macrophage ROS and improved mitochondrial function. This resulted in a decreased CD68+CD80+ and increased CD163+CD206+ lesion macrophages and attenuated atherosclerosis. Our results imply that NOX4-dependent mitochondrial oxidative stress in aging contributes to macrophage mitochondrial dysfunction, glycolytic metabolic switch, and pro-inflammatory phenotype, advancing atherosclerosis. Inhibition of NOX4 could alleviate vascular inflammation and atherosclerosis by improving mitochondrial function in macrophages.