Increased expression of procoagulant activity on the surface of human platelets exposed to heavy-metal compounds

Abstract

One of the essential roles for platelets in haemostasis is in the potentiation of blood clotting due to the contribution of anionic phospholipid from the surface of the cells, as an essential cofactor to the proteolytic reactions of coagulation (platelet procoagulant activity). Only a limited number of agonists are known to initiate platelet procoagulant activity. In this study the rate of thrombin formation on the platelet surface was observed to increase in a dose-dependent manner upon treatment of washed platelets with heavy-metal compounds. Unlike the immediate increase observed upon treatment of platelets with calcium ionophore, A23187, the change due to these agents was progressive, approaching a maximum after 10 min. The maximum-fold acceleration of the rate of thrombin formation compared with control platelets was calculated for HgCl2 (56-fold), AgNO3 (42-fold) phenylmercuriacetate (24-fold) and thimerosal (14-fold), compared with 70-fold observed for calcium ionophore. The increase in procoagulant activity due to HgCl2 coincided with a large increase in intracellular calcium and phosphorylation of 22 and 45 kDa proteins. It is considered that the mechanism responsible for the increase in procoagulant activity is exposure of anionic phospholipids. This was detected by a 2-fold increase in the binding of 125I-annexin V upon addition of HgCl2, compared with resting platelets (3-fold on treatment of platelets with calcium ionophore). In contrast to the generation of activity by A23187 and other known agonists of this reaction, heavy-metal compounds appeared to cause little or no release of microparticles from the platelet surface. Since HgCl2 did not cause aggregation of platelets or significant release of serotinin, these findings may give further support to the need for exposure and ligation of glycoprotein IIb:IIIa for vesiculization to occur. Treatment of platelets with heavy metals may constitute a new approach to investigating the early changes in the cell membrane which lead to increased expression of anionic phospholipid.

Platelet Agonists Enhance the Import of Phosphatidylethanolamine into Human Platelets

“…Assessment of Platelet Procoagulant Activity-Procoagulant activity in the presence of activated platelets was determined essentially as described previously (19). Platelets (5 ϫ 10 6 /0.5 ml) were incubated without or with the indicated concentrations of thrombin and collagen plus either LDL or phospholipid vesicles for 5 min at 37°C in a buffer composed of (in mM): 145 NaCl, 10 Hepes, 5 KCl, 1 MgCl 2 , 5 glucose (pH 7.4, "coagulation buffer").…”

The organomercurial 4-aminophenylmercuric acetate, independent of matrix metalloproteinases, induces dose-dependent activation/ inhibition of platelet aggregation

“…Several mercurial compounds have the ability to penetratep latelets, bind to plasma membrane receptorsand block sulfhydryls, mobilize intracellular calcium, increase expression of procoagulant activity and releaseserotonin,activate phospholipase A2,increase arachidonic acid metabolism, phosphorylate pleckstrin and myosin light chain, and modulate plateletaggregation. Independent stimulatorye ffectsa tl ow dose and inhibitorye ffectso np lateletr esponses( aggregation and dense granule release) to agonists at high dose have beenreported with mercurials (25)(26)(27). Basedon studiesofthiomersal (25), Hg 2+ mayact as asulfhydryl reagent and an oxidizing agentorthe reaction with asulfhydryl group of the type R-SH mayr epresent an exchange reaction.…”

HAuCl4, Putative General Aquaporins Blocker, Reduces Platelet Spreading, Filopodia Formation, Procoagulant Response, and Thrombus Formation Under Flow

“…The most extensively used nonspecific AQPs’ inhibitor – HgCl 2 , is highly toxic and its general inhibitory properties has been questioned since activation of AQP6 by HgCl 2 has also been observed ( Lee et al, 2012 ). What is more, mercurials (including HgCl 2 ) have been found to notably increase human platelet procoagulant activity at concentrations even lower than those routinely used to block AQPs ( Goodwin et al, 1995 ), which makes them unsuitable tools to study antiplatelet effect of AQPs inhibition. Studies performed on various cell types suggest that gold-containing compounds are promising candidates for general inhibitors of AQPs, with comparable inhibitory potential to HgCl 2 but without side-effects and direct toxicity toward platelets ( Niemietz and Tyerman, 2002 ; Yang et al, 2006 ; Martins et al, 2013 ; Misztal et al, 2018 ).…”