Summary
The effects of plasmin treatment upon washed human platelets were studied in an attempt
to elucidate the mechanisms underlying thrombin-induced platelet aggregation. At calcium
concentrations of 10–20 μM, plasmin (0.2 CTA U/ml) inhibited thrombin-induced aggregation
almost completely, but did not diminish the thrombin-induced release of adenine nucleotides,
5-hydroxytryptamine, or calcium. Increasing the calcium concentration partially antagonized
plasmin’s inhibition of aggregation.
Studies utilizing calcium chelators and the Kunitz soybean trypsin inhibitor (SBTI)
as a plasmin inhibitor indicated that in order to achieve maximal block of aggregation,
plasmin must act upon a substrate made fully available only after an initial thrombin-platelet
interaction has taken place. Moreover, the time course of this inhibition parallels
the time course of the thrombin-induced release reaction.
Plasmin inhibition of aggregation could not be mimicked by exposing the platelets
to proteolytic digests of fibrinogen at concentrations as high as 17% total platelet
protein. Nor could inhibitory activity be recovered from supernatants of plasmin -treated
platelets, upon centrifugation and treatment with SBTI.
With the use of a “cold initiation” technique, the release by thrombin of 46.7 ± 6-7
(mean ± SEM) μg of fibrinogen immunological equivalents per mg platelet protein could
be demonstrated. Platelets in which thrombin-induced aggregation was abolished by
plasmin treatment (and the plasmin subsequently inactivated by SBTI) aggregated normally
upon addition of as little as 10 μg human plasma fibrinogen per mg platelet protein.
It is concluded that plasmin inhibition of aggregation most likely results from its
attack upon a protein that is released or becomes fully available subsequent to interaction
of thrombin with a platelet receptor mediating release. The results of this study
are consistent with a cofactor role for fibrinogen in the aggregation of human platelets
by thrombin.
