Myocardial infarction, stroke and other diseases related to thrombosis and atherosclerosis are still intransigent in their resistance to prophylaxis and therapy. More fundamental knowledge is needed. In this application we propose studies on platelets, endothelial cells and blood vessels to elucidate their role in thrombosis and the underlying atherosclerotic lesions in blood vessels. The focus shall be on three of the modulating elements or sub-processes involved in the induction and progression of thrombosis, atherosclerosis and inflammation.
The aims i nclude the following: 1) Studies on platelet-activating factor (PAF), a potent inducer of platelet aggregation and involved in inflammatory processes - these comprise the determination of the structure and biosynthesis of PAF formed by human platelets and endothelial cells obtained from blood vessels of adult humans and the metabolism of PAF by these cells. 2) Studies on the metabolism of arachidonic acid by endothelial cells derived from adult human blood vessels with differing susceptibility to atherosclerosis. 3) Studies on the influence of diabetes or sex hormones on platelet and blood vessel prostaglandin synthesis. The methods employed include the formation of PAF in response to stimuli, its extraction and separation by high performance liquid chromatography and its estimation by its ability to induce platelet aggregation. These studies also include measurement of acetyl transferase activity and determination of the 1-0-alkyl moiety of PAF by gas liquid chromatography of a derivative of its 1-0-alkyl glycerol. The metabolism of arachidonic acid by endothelial cells will be followed by stimulating the cultured cells with radio-labelled arachidonic acid in the presence or absence of inhibitors of certain enzyme activities, extracting the metabolites and separating them by thin layer chromatograhy and measuring the radioactivity in the zones corresponding to authentic standards. In some studies the cultured cells will be incubated with physiological stimuli such as bradykinin, histamine or angiotensin II and the prostacyclin formed will be measured by radioimmunoassay of 6-keto-PGF-1 alpha and HETE by spectrophotometry at 235 nm. Platelet and blood vessel prostaglandin synthesis will be studied in male rabbits with experimentally induced diabetes and in female and pregnant female rabbits and compared to our previous findings in normal male rabbits. Specific radioimmunoassays for 6-keto-PGF-1 alpha and thromboxane B-2 will be employed.
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