Bilirubin Prevents Atherosclerotic Lesion Formation in Low‐Density Lipoprotein Receptor‐Deficient Mice by Inhibiting Endothelial VCAM‐1 and ICAM‐1 Signaling

BACKGROUND: Numerous epidemiological studies support an inverse association between serum bilirubin levels and the incidence of cardiovascular disease; however, the mechanism(s) by which bilirubin may protect against atherosclerosis is undefined. The goals of the present investigations were to assess the ability of bilirubin to prevent atherosclerotic plaque formation in low‐density lipoprotein receptor‐deficient (Ldlr (−/−)) mice and elucidate the molecular processes underlying this effect. METHODS AND RESULTS: Bilirubin, at physiological concentrations (≤20 μmol/L), dose‐dependently inhibits THP‐1 monocyte migration across tumor necrosis factor α–activated human umbilical vein endothelial cell monolayers without altering leukocyte binding or cytokine production. A potent antioxidant, bilirubin effectively blocks the generation of cellular reactive oxygen species induced by the cross‐linking of endothelial vascular cell adhesion molecule 1 (VCAM‐1) or intercellular adhesion molecule 1 (ICAM‐1). These findings were validated by treating cells with blocking antibodies or with specific inhibitors of VCAM‐1 and ICAM‐1 signaling. When administered to Ldlr (−/−) mice on a Western diet, bilirubin (30 mg/kg intraperitoneally) prevents atherosclerotic plaque formation, but does not alter circulating cholesterol or chemokine levels. Aortic roots from bilirubin‐treated animals exhibit reduced lipid and collagen deposition, decreased infiltration of monocytes and lymphocytes, fewer smooth muscle cells, and diminished levels of chlorotyrosine and nitrotyrosine, without changes in VCAM‐1 or ICAM‐1 expression. CONCLUSIONS: Bilirubin suppresses atherosclerotic plaque formation in Ldlr (−/−) mice by disrupting endothelial VCAM‐1‐ and ICAM‐1‐mediated leukocyte migration through the scavenging of reactive oxygen species signaling intermediaries. These findings suggest a potential mechanism for the apparent cardioprotective effects of bilirubin.