Incorporation of Lipids Labeled with Various Fatty Acids into the Cytoskeleton of Aggregating Platelets

Earlier studies showed that during the first 20 to 25 seconds of aggregation induced by thrombin (0.1 U/mL) or adenosine diphosphate (ADP) (2 microM) of rabbit or human platelets prelabeled with [3H]palmitic acid, labeled lipid became associated with the cytoskeleton (isolated after lysis with 1% Triton X-100, 5 mM EGTA [ethylene glycolbis-(beta-aminoethyl ether(N,N,N',N'-tetraacetic acid] in the presence of 0.5 mM leupeptin and 50 mM benzamidine). In comparison with labeled lipid in intact platelets, the labeled lipid that was associated with the cytoskeleton was enriched in phospholipids and ceramide. To determine whether these effects were specific for lipids labeled with palmitic acid, we studied rabbit platelets in which lipids had been labeled by incubation of the platelets with pairs of 14C- or 3H-labeled palmitic, stearic, arachidonic, and linoleic acids. Examination of the distribution of label among the lipid classes of intact platelets showed that phospholipids contained most of the label. Under the conditions of limited, thrombin-induced aggregation used, labeled lipids were not lost from the platelets and the distribution of label among the lipid classes was essentially unchanged. There were major differences in the incorporation of labeled lipids into the cytoskeleton. The greatest incorporation (2.1 to 2.8% of the label in the platelets) was observed with palmitic acid-labeled lipids; by direct comparison, only 44% as much of the label of stearic acid-labeled lipids, 21% as much of the label of linoleic acid-labeled lipids, and only 6% as much of the label of arachidonic acid-labeled lipids was incorporated into the cytoskeleton.(ABSTRACT TRUNCATED AT 250 WORDS)