Abstract
BackgroundDirect interaction between Red blood cells (RBCs) and platelets is known for a long time. The bleeding time is prolonged in anemic patients independent of their platelet count and could be corrected by transfusion of RBCs, which indicates that RBCs play an important role in hemostasis and platelet activation. However, in the last few years, opposing mechanisms of platelet inhibition by RBCs derived nitric oxide (NO) were proposed. The aim of our study was to identify whether RBCs could produce NO and activate soluble guanylate cyclase (sGC) in platelets.MethodsTo test whether RBCs could activate sGC under different conditions (whole blood, under hypoxia, or even loaded with NO), we used our well-established and highly sensitive models of NO-dependent sGC activation in platelets and activation of purified sGC. The activation of sGC was monitored by detecting the phosphorylation of Vasodilator Stimulated Phosphoprotein (VASPS239) by flow cytometry and Western blot. ANOVA followed by Bonferroni’s test and Student’s t-test were used as appropriate.ResultsWe show that in the whole blood, RBCs prevent NO-mediated inhibition of ADP and TRAP6-induced platelet activation. Likewise, coincubation of RBCs with platelets results in strong inhibition of NO-induced sGC activation. Under hypoxic conditions, incubation of RBCs with NO donor leads to Hb-NO formation which inhibits sGC activation in platelets. Similarly, RBCs inhibit activation of purified sGC, even under conditions optimal for RBC-mediated generation of NO from nitrite.ConclusionsAll our experiments demonstrate that RBCs act as strong NO scavengers and prevent NO-mediated inhibition of activated platelets. In all tested conditions, RBCs were not able to activate platelet or purified sGC.
Highlights
Direct interaction between Red blood cells (RBCs) and platelets is known for a long time
Studies propose that RBCs, under certain conditions, can produce nitric oxide (NO) and inhibit platelet activation
The activation of platelet soluble guanylate cyclase (sGC)/cGMP/Protein Kinase G (PKG) by NO released from RBCs was not directly proven
Summary
Direct interaction between Red blood cells (RBCs) and platelets is known for a long time. The bleeding time is prolonged in patients with anemia independent on their platelet count and could be corrected by RBCs transfusion, and bleeding defects are directly connected with platelet function but not with the blood coagulation system [1,2,3]. RBCs transfusion could increase platelet activation and might be dangerous in treatment of coronary artery diseases [4] These clinical observations clearly pointed out that RBCs, by still unidentified mechanism(s), play some important role in platelet activation. Recently in several papers [5,6,7,8,9] platelet inhibition by RBCs-derived NO was described. In none of these papers direct activation of platelet soluble Guanylate Cyclase (sGC) by RBCs-derived NO was shown. The connection between RBCs, eNOS and hemoglobin (Hb) to platelet sGC activation and platelet inhibition cannot be proven
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