The effect of phospholipase C on human blood platelets.
The effect of phospholipase C (EC 3.1.4.3) on human blood platelets has been studied. Phospholipase C from Bacillus cereus was purified to homogeneity as judged by analytical and sodium dodecyl sulphate disc gel electrophoresis and by immunoelectrophoresis. Human platelets isolated from platelet-rich plasma by gel filtration or by centrifugation and washing were incubated with phospholipase C. A loss of 20-45% of the total platelet phospholipid was observed, whereas 88% was hydrolyzed when platelet homogenates were submitted to identical enzyme treatment. Intact platelets lost 50-75% phosphatidylethanolamine, 20-50% phosphatidylcholine, and 20-25% phosphatidylserine. Sphingomyelin was not a substrate for the enzyme under the conditions used. The platelets contained no detectable endogenous phospholipase C activity. The loss of phospholipid was not accompanied by aggregation of the platelets, nor did the platelets lose their ability to aggregate with ADP or thrombin. Total platelet factor 3 releasable by freezing and thawing was reduced. Measurements of releasable platelet factor 4 and the efflux of serotonin showed that no release reaction was triggered even when up to 45% of the total phospholipid in the platelets was hydrolyzed. When sphingomyelinase was added together with, before, or after phospholipase C, aggregation occurred. Sphingomyelinase alone gave no aggregation. The gel-filtered platelets also aggregated upon addition of purified phospholipase C from Clostridium perfringens. The distribution of phospholipids in the platelet membrane is discussed.
D001409:Bacillus cereus / Q000201:enzymology; D001792:Blood Platelets / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D010713:Phosphatidylcholines / Q000378:metabolism; D010714:Phosphatidylethanolamines / Q000378:metabolism; D010740:Phospholipases / Q000302:isolation & purification / Q000494:pharmacology*; D010743:Phospholipids / Q000378:metabolism; D010974:Platelet Aggregation / Q000187:drug effects; D010978:Platelet Factor 4; D012701:Serotonin / Q000378:metabolism; D013109:Sphingomyelins / Q000378:metabolism
D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010743:Phospholipids; D013109:Sphingomyelins; D012701:Serotonin; D010978:Platelet Factor 4; D010740:Phospholipases