P155	Clearance of apoptotic cells by macrophages is impaired in human and experimental atherosclerosis.
1D.M.Schrijvers, 1G.R.Y.De Meyer, 2M.M.Kockx, 1A.G.Herman, 1W.Martinet
1University of Antwerp, Division of Pharmacology, Wilrijk, BE; 2AZ Middelheim, Department of Pathology, Antwerp, BE.

Objective - Apoptotic cell death is a prominent feature of human atherosclerotic plaques. Apoptotic cells (AC) have atherothrombogenic potential, due to phosphatidylserine (PS) exposure on the cell surface. Moreover, it is known that plaque-derived AC account for the high tissue factor activity in plaques, which is a key element in the initiation of the coagulation cascade. Rapid recognition and clearance of AC by phagocytes is essential for normal development and prevention of inflammation and disease. When the apoptotic remains of a cell are not efficiently removed, inevitable leaking of cell contents occurs, inciting an inflammatory reaction. Since human atherosclerotic plaques show signs of both apoptosis and inflammation, we questioned whether cells induced to undergo apoptotic cell death, are efficiently removed. Methods and Results - Human endarterectomy specimens and human tonsils were co-stained for CD68 (macrophages) and TUNEL (apoptosis). Free and phagocytized AC were counted in both tissues. The ratio of free versus phagocytized AC was 19 times higher in human atherosclerotic plaques as compared to human tonsils, indicating a severe defect in clearance of AC. Moreover, in tonsils macrophages engulfed several AC at the same time to ensure effective and swift clearance, whereas in atherosclerotic plaques, the majority of phagocytic macrophages engulfed one single apoptotic cell. Examination of plaques from cholesterol-fed rabbits not only confirmed our findings in human plaques but also showed that impaired clearance of AC persisted during plaque progression. In vitro experiments with J774 or peritoneal mouse macrophages showed that several factors caused impaired phagocytosis of AC including cytoplasmic overload of macrophages with indigestible material (beads), free radical attack and competitive inhibition among oxidized red blood cells, oxLDL and AC for the same receptor(s) on the macrophage. Foam cell formation induced by aggregated lipoproteins or platelets did not alter phagocytosis efficiency. Conclusion - Our data demonstrate that phagocytosis of AC is impaired in atherosclerotic plaques, which is at least partly attributed to oxidative stress and cytoplasmic saturation with indigestible material. This could have severe consequences regarding further accumulation of necrotic debris, plaque thrombogenicity and stability of human atherosclerotic plaques.

Copyright © 2005 S. Karger AG, Basel. Any further use of this abstract requires written permission from the publisher.