Summary
Acquired platelet storage pool disease has been shown to be associated with reduced
platelet thromboxane synthesis. However, the mechanisms for this dysfunction are incompletely
understood. The present experiments were designed to evaluate some of the possible
defects which may account for impaired thromboxane formation in human platelets previously
exposed to thrombin in vitro. Washed platelets pretreated with 0.5 U/ml thrombin for
20 sec and subsequently recovered as single degranulated platelets were incapable
of forming normal amounts of thromboxane upon a second stimulation with thrombin (as
compared to Tyrode-pretreated control platelets). In contrast, thrombin-degranulated
platelets released additional amounts of thromboxane in response to arachidonate,
or collagen, indicating that short-time exposure to thrombin does not irreversibly
inactivate platelet cyclooxygenase or thromboxane synthetase. Upon incubation of the
thrombin-pretreated platelets in autologous plasma in the presence of 14C-arachidonate, the label became associated with the platelets to the same extent
as with control platelets. However, the platelets did not recover their ability to
synthesize normal amounts of thromboxane upon restimulation with thrombin, and only
about half of the label was lost from the thrombin-pretreated platelets as compared
to control platelets in response to thrombin. The ability of collagen to cause loss
of 14C-arachidonate and formation of thromboxane was the same regardless of whether or
not the platelets had been pretreated with thrombin. Incubation of platelets in plasma
in the presence of added arachidonate for 90 min resulted in complete refractoriness
to a second stimulation with thrombin but not with collagen. However, the control
platelets also lost most of their ability to synthesize thromboxane when incubated
with arachidonate for 90 min and thereafter stimulated with thrombin. Thus, the presence
of added arachidonate affects the thrombin-inducible thromboxane synthesis after prolonged
incubation of human platelets in plasma. Our observations suggest that depletion of
endogenous arachidonate is not a major cause for the defective thrombin-induced thromboxane
synthesis in thrombin-pretreated platelets. It is more likely that impaired mobilization
of endogenous arachidonic acid explains this dysfunction. Defective mobilization of
arachidonate in thrombin-degranulated platelets may be due to agonists-specific receptor
desensitization, but the responsible mechanism has not been identified.
Keywords
Degranulated platelets - Thrombin - Collagen - Thromboxane synthesis - Arachidonic
acid metabolism - Receptor desensitization
