Abstract
Inflammatory activity and hypoxia in atherosclerotic plaques are associated with plaque instability and thrombotic complications. Recent studies show that vascular cell metabolism affects atherogenesis and thrombogenicity. This study aimed to identify the metabolites in macrophage-rich unstable plaques that modulate atherogenesis and serve as potential markers of plaque instability. Atherosclerotic plaques were induced by balloon injury in the iliofemoral arteries of rabbits fed on a conventional or 0.5% cholesterol diet. At 3 months post-balloon injury, the arteries and cardiac tissues were subjected to histological, quantitative real-time polymerase chain reaction, and metabolomic analyses. The identified metabolite-related proteins were immunohistochemically analyzed in stable and unstable plaques from human coronary arteries. The factors modulating the identified metabolites were examined in macrophages derived from human peripheral blood mononuclear cells. Metabolomic analysis revealed that choline and guanine levels in macrophage-rich arteries were upregulated compared with those in non-injured arteries and cardiac tissues. Vascular choline levels, but not guanine levels, were positively correlated with the areas immunopositive for macrophages and tumor necrosis factor (TNF)-α and matrix metalloproteinase (MMP) 9 mRNA levels in injured arteries. In human coronary arteries, choline transporter-like protein (CTL) 1 was mainly localized to macrophages within plaques. The area that was immunopositive for CTL1 in unstable plaques was significantly higher than that in stable plaques. Intracellular choline levels were upregulated upon stimulation with TNF-α but were downregulated under hypoxia in cultured macrophages. Administration of choline upregulated the expression of TNF-α and CTL1 mRNA in cultured macrophages. The transfection of CTL1 small interfering RNA decreased CTL1, TNF-α, and MMP9 mRNA levels in cultured macrophages. These results suggest that choline metabolism is altered in macrophage-rich atherosclerotic lesions and unstable plaques. Thus, CTL1 may be potential markers of plaque instability.