Platelet phagocytosis and processing of beta-amyloid precursor protein as a mechanism of macrophage activation in atherosclerosis.

In human occluded saphenous vein grafts, we previously demonstrated cytotoxic foam cells, presumably derived from macrophages engulfing platelets. In the present study, we investigated whether platelet phagocytosis occurs in human atherosclerotic plaques, whether this activates macrophages, and whether the platelet constituent, amyloid precursor protein (APP), was involved. Immunohistochemistry documented the presence of APP, beta-amyloid peptide (Abeta, cleaved from APP), and platelets (CD9), along with inducible NO synthase (iNOS) and cyclooxygenase-2, two markers of macrophage activation, around microvessels in advanced human carotid artery plaques (n=18). Abeta colocalized with iNOS-expressing macrophages that were often surrounded by platelets. In vitro, murine J774 and human THP-1 macrophages were incubated with or without washed human platelets. Coincubation of macrophages and platelets led to platelet phagocytosis (electron and confocal microscopy) and formation of lipid-, APP-, and Abeta-containing foam cells. These expressed iNOS mRNA (reverse transcription-polymerase chain reaction) and protein and produced nitrite and tumor necrosis factor-alpha (ELISA). Macrophage pretreatment with 4-(2-aminoethyl)benzenesulfonyl fluoride, a protease inhibitor, reduced APP processing and inhibited NO biosynthesis induced by platelet phagocytosis but not by lipopolysaccharides. Human atherosclerotic plaques and J774 and THP-1 macrophages contained mRNA of the APP-cleaving enzyme beta-secretase. This is the first demonstration of Abeta, a peptide extensively studied in Alzheimer's disease, in human atherosclerotic plaques. It was present in activated iNOS-expressing perivascular macrophages that had phagocytized platelets. In vitro studies indicate that platelet phagocytosis leads to macrophage activation and suggest that platelet-derived APP is proteolytically processed to Abeta, resulting in iNOS induction. This represents a novel mechanism for macrophage activation in atherosclerosis.

Nitric oxide donor molsidomine favors features of atherosclerotic plaque stability during cholesterol lowering in rabbits.

Rupture-prone atherosclerotic plaques are characterized by a thin fibrous cap containing numerous macrophage-derived foam cells and few smooth muscle cells (SMC). Decreasing the ratio between macrophages and SMC might favor plaque stabilization. Macrophages expressing inducible nitric oxide (NO) synthase become hypersensitive to killing by exogenous NO donors. Therefore, we investigated in cholesterol-fed rabbits (20 weeks 0.3% cholesterol) the effect of 4 weeks cholesterol withdrawal alone and in combination with the NO donor molsidomine on plaque size, cell composition, superoxide production and extracellular superoxide dismutase (ecSOD) mRNA expression in the atherosclerotic plaques in the aorta. Cholesterol withdrawal alone did not alter atherosclerotic plaque size, the increased superoxide production or the decreased ecSOD mRNA, but led to the formation of a thin subendothelial macrophage-free layer and reduced both vascular cell adhesion molecule-1 expression and cell replication in the luminal part of the plaques. Treatment with molsidomine (1 mg/kg/day) during cholesterol withdrawal did not affect plaque size but increased the thickness of the subendothelial macrophage-free layer consisting of SMC, and normalized both superoxide production and ecSOD mRNA expression. The latter findings demonstrate that molsidomine, when combined with cholesterol lowering, decreases signs of oxidative stress and increases features of stable atherosclerotic plaques.