title: Thiolprotease inhibitor, EST, can inhibit thrombin-induced platelet activation.
abstract: Participation of thiolprotease in platelet activation was investigated. When platelets were treated with EST (L-trans-epoxysuccinyl-leucylamide (3-methyl)butane-ethyl ester, a membrane-permeable thiolprotease inhibitor) for 30 min, thrombin-induced aggregation and secretion were inhibited, and remained so even when the platelets were washed and resuspended in EST-free medium after the pretreatment. The inhibitory action of EST on thrombin-induced platelet aggregation and secretion was both dose-dependent and incubation-time-dependent. The inhibitory action of EST on platelet aggregation and secretion was shown not only in response to thrombin but also to platelet activating factor (PAF). Pretreatment of platelets with 1 mM EST for 30 min inhibited the 65% of calpain (thiolprotease) activity in platelets. The phosphorylation of 40 kDa and 20 kDa proteins of platelets caused by stimulation with thrombin was blocked by the pretreatment with 1 mM EST. Phosphatidylinositol hydrolysis and inositol-1-phosphate production, which appear after stimulation of platelets with thrombin, were also inhibited by the pretreatment with 1 mM EST. The results suggest that EST was incorporated to inside of platelets, and inhibited activation of platelet through inhibition of thiolprotease.
authors.affiliation: Department of Clinical Biochemistry, Faculty of Pharmaceutical Sciences, Teikyo University, Kanagawa, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Ishii
authors.forename: M
authors.initials: M
authors.lastname: Kuboki
authors.forename: J
authors.initials: J
authors.lastname: Fujii
authors.forename: S
authors.initials: S
authors.lastname: Hiraishi
authors.forename: M
authors.initials: M
authors.lastname: Kazama
mesh_terms: D000328:Adult; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D002154:Calpain / Q000037:antagonists & inhibitors; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D007295:Inositol Phosphates / Q000378:metabolism; D007930:Leucine / Q000031:analogs & derivatives* / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000378:metabolism; D010766:Phosphorylation; D010972:Platelet Activating Factor / Q000037:antagonists & inhibitors; D015539:Platelet Activation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D012701:Serotonin / Q000378:metabolism; D013917:Thrombin / Q000037:antagonists & inhibitors; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D015853:Cysteine Proteinase Inhibitors; D007295:Inositol Phosphates; D010716:Phosphatidylinositols; D010972:Platelet Activating Factor; D010975:Platelet Aggregation Inhibitors; C002647:inositol 1-phosphate; D012701:Serotonin; D010738:Type C Phospholipases; D013917:Thrombin; D002154:Calpain; D007930:Leucine; C108192:aloxistatin