Analysis of different mechanisms through hydrogen sulphide (H2S) would inhibit platelet aggregation.

BERMEJO, EMILSE; SAENZ, DANIEL; ALBERTO, FABIANA; ROSENSTEIN, RUTH

Amsterdam, Netherland

Congreso; XXIV Congress of the International Society on Thrombosis and Haemostasis (ISTH); 2013

International Society on Thrombosis and Haemostasis

Background Hydrogen sulphide (H2S) is the most recent of the gasotransmitters to be identified and investigated. H2S is an important endogenous modulator, which exhibits beneficial effects on the cardiovascular system. However, the mechanisms involved in the different effects on platelets pathways are still unclear. Objective: In order to investigate H2S inhibitory pathways on platelet function we performed several tests: the opening of KATP-channels, the cGMP and cAMP accumulation, fibrinogen binding (B-Fg) and Ca2+ influx. Methods: Tests were performed using platelet rich plasma (PRP)(300 × 109 plat/L). Sodium hydrogen sulfide (NaHS) was used as H2S donor and 100 μM glibenclamide (Glib) as a selective K+-channel blocker. Samples were preincubated with buffer, 100 mM glib (during 30 min) at 37°C , then with or without NaHS (1, 5 and 10 mM) for 2 min before adding 2 μg/mL collagen as agonist. The expression of fibrinogen binding (B-Fg) and Ca2+ influx were assessed by flow cytometry using mean fluorescence intensities. cGMP and cAMP accumulation by RIA were measured as pmol/ 106plat. Student?s t-test control vs. NaHS. : * = P< 0.05, ** = P < 0.01. Results: H2S significantly inhibited platelet aggregation (P< 0.01) induced by 2 μg/ml Col. Glib failed to block the inhibition induced by NaHS. The accumulation of cGMP and cAMP in presence of NaHS have no statistical difference 10 ±2 vs 9±1 and 5±1 vs. 4±2 respectively. Levels of B-Fg (126±12 vs. 15±5)** were inhibited in the presence of H2S, and Ca2+ influx was unaffected. Conclusion: Platelet aggregation induced by collagen was significantly inhibited in a concentration-dependent by NaHS. The test with Glib ruled out the role of KATP channel in H2S-induced antiaggregation. The accumulation of cGMP and cAMP confirmed that these are not the mediators of the inhibitory effect of NaHS. This result, together with a significant blocked of B-Fg levels, suggest the inhibition on platelet function would be through another signaling pathway, such as CalDAG-GEFI/Rap1.Thus, H2S may constitute a novel target for antithrombotic drug development.