benzyloxycarbonylvalyl-alanyl-aspartyl-fluoromethyl-ketone and Arteriosclerosis

The mechanisms of thrombosis on plaque erosion are poorly understood. We examined the potential role of endothelial apoptosis in endothelial erosion and vessel thrombosis.. Segments of New Zealand White rabbit femoral arteries were temporarily isolated in vivo. One artery was incubated with staurosporin for 30 minutes, whereas the contralateral artery was incubated with saline and served as control. Three days later, thrombosis was evaluated angiographically and histologically. TUNEL score in the endothelial layer was significantly increased in staurosporin-treated arteries compared with controls (2.43+/-0.30 versus 0.93+/-0.44, respectively; P=0.001). Large areas of endothelial denudation were detectable in staurosporin-treated vessels, whereas endothelium integrity was almost preserved in the saline group. Vessel thrombosis occurred in 58% of staurosporin-treated arteries (7 of 12) but in only 8% of saline-treated segments (P<0.01). Immunoreactivities for tissue factor, platelets, and fibrin were detectable within the thrombus. Addition of ZVAD-fmk (0.1 mmol/L) significantly reduced the occurrence of thrombosis (1 of 7 arteries or 14%, P=0.04). These results were confirmed in balloon-injured atheromatous arteries.. In vivo induction of endothelial apoptosis leads to both vessel thrombosis and endothelial denudation. Endothelial apoptosis may be a critical step in the transition from a stable endothelialized plaque to plaque erosion and thrombosis.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Arteriosclerosis; Catheterization; Cysteine Proteinase Inhibitors; Endothelial Cells; Endothelium, Vascular; Femoral Artery; Fibrin; In Situ Nick-End Labeling; Platelet Count; Rabbits; Staurosporine; Thromboplastin; Thrombosis; Tunica Intima

In vitro studies of macrophage death in response to oxidized LDL (oxLDL) were undertaken as a model for the formation of the necrotic core of atherosclerotic plaque.. Thioglycollate-elicited mouse peritoneal macrophages avidly incorporated both oxLDL and acetylated LDL (acLDL) to become foam cells. oxLDL-treated macrophages, but not acLDL-treated macrophages, showed nearly 100% death, with characteristics consistent with apoptosis, including cell surface phosphatidylserine exposure, intracellular caspase-3 activity, cleavage of caspase-3 substrates, and DNA fragmentation, as shown by TUNEL assay. The activated form of caspase-3 (p17 cleaved form) was present in attached, viable macrophages before exposure to oxLDL. This p17 form was also found in apparently viable as well as in TUNEL-positive cells within atherosclerotic lesions of chow-fed apolipoprotein E-deficient (ApoE-/-) mice. The amount of p17 caspase-3 was reduced by in vitro blockade of FasL with an FasL-blocking antibody and was absent in macrophages from lpr/lpr mice, which lack functional Fas. Moreover, lpr/lpr macrophages resisted oxLDL cytotoxicity.. The naturally occurring Fas-FasL induction of caspase-3 cleavage after macrophage attachment may represent an important physiologic mechanism that primes for cytotoxicity by oxLDL and possibly, other death-inducing molecules.

Topics: Amino Acid Chloromethyl Ketones; Animals; Apoptosis; Arteriosclerosis; Caspase 3; Caspase Inhibitors; Caspases; Cysteine Proteinase Inhibitors; DNA Fragmentation; Enzyme Activation; Enzyme Inhibitors; Fas Ligand Protein; fas Receptor; Foam Cells; In Situ Nick-End Labeling; Lipoproteins, LDL; Macrophages, Peritoneal; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Oligopeptides