Cross-talking between lymphocytes and platelets and its regulation by nitric oxide and peroxynitrite in physiological condition and endotoxemia

Cross-talk between platelets and lymphocytes may play a role in different pathological conditions like sepsis. This study aimed to investigate the effect of lymphocytes on platelet aggregation in lipopolysaccharide (LPS)-stimulated and non-stimulated cells. Lymphocytes and platelet-rich plasma (PRP) were obtained from rat arterial blood. Platelets (1.2×108platelets/ml) were incubated with lymphocytes (0.8×106cells/ml) in the presence or not of LPS (100μg/ml), after which ADP (5μM)-induced platelet aggregation was carried out. Lymphocytes inhibited by 51% the platelet aggregation, which was significantly prevented by the non-selective NO inhibitor l-NAME (300μM) or the selective iNOS inhibitor 1400W (100μM), as well as by the soluble guanylyl cyclase (sGC) inhibitor ODQ (10μM). The platelet inhibition by lymphocytes was accompanied by 2-fold increase of intraplatelet cGMP levels. Next, lymphocytes and platelets were co-incubated with LPS for 6h. In LPS-treated cells, lymphocytes produced a larger inhibition of platelet aggregation (62%), despite the same elevation of cGMP levels (2.2-fold increase). This inhibitory effect was prevented by l-NAME and 1400W, but rather unaffected by ODQ. The peroxynitrite (ONOO−) scavenger −(−)epigallocatechin gallate (ECG, 100μM) abolished the inhibition by lymphocytes on platelet aggregation in LPS-treated cells, but not in non-treated cells. Our results show that lymphocytes act to inhibit platelet aggregation via iNOS-derived NO release and cGMP generation. In presence of LPS, ONOO− production accounts for the platelet inhibition.