P2589Platelets serve as carriers of cardioprotective factors after remote ischemic conditioning in humans

Abstract

Abstract Background/Introduction Brief episodes of ischemia/reperfusion (I/R) in a tissue or organ remote from the heart reduce myocardial infarct size after sustained severe myocardial I/R in all species tested so far, including humans. Remote ischemic conditioning can be induced before (pre-), during (per-) or following (post-) myocardial ischemia. Signal transfer from the remote tissue/organ to the heart is both, neuronal and humoral. Humoral signal transfer has been evidenced by the transfer of cardioprotection via plasma or plasma derivatives from one individual to another individual's heart, even across species. Circulating blood cells have been considered as targets for cardioprotection, but so far not as carriers of cardioprotective signals. Purpose To investigate the role of platelets as potential carriers of cardioprotection by remote ischemic preconditioning (RIPC). Methods Peripheral venous blood samples were collected from healthy volunteers (5 male/5 female, mean age 26±5 years) before and 60 min after RIPC (3×5 min blood pressure cuff inflation at 200 mmHg on the left upper arm/5 min deflation) or placebo (PLA) protocol (blood pressure cuff uninflated). RIPC and PLA protocols, respectively, were performed in randomized sequence at an interval of one week. Blood (80 mL) was drawn into tubes containing sodium citrate, apyrase and prostaglandin E1. Blood cells were counted using a hematology analyzer. Blood was centrifuged (100 g, 15 min, at room temperature) to obtain platelet-rich plasma (PRP). PRP was washed twice with buffer (pH 6.5), and the pellet was re-suspended in suspension buffer (pH 7.35); the platelet amount was adjusted to 2.5x103 platelets/μL. The platelet suspension was supplemented with 1 mol/L CaCl2 and centrifuged (14,000 g) at 4°C for degranulation. The supernatant, i.e. the platelet releasate, was retrieved. Male Lewis rats were sacrificed, their hearts isolated and perfused at constant pressure. Diluted platelet releasate (1:10) was infused into the isolated perfused rat hearts for 8 min followed by 2 min washout before 30/120 min global I/R. Infarct size (percentage of ventricular mass) was demarcated with staining by triphenyltetrazolium chloride. Data are presented as mean±SD. Results The platelet count was increased after RIPC, but unchanged after PLA (RIPC: from 204±19x103 platelets/μL to 247±16x103 platelets/μL; PLA: from 230±16x103 cells/μL to 222±18x103 platelets/μL; PLA vs. RIPC p<0.01, RIPC before vs. after p<0.01, two-way ANOVA for repeated measures with Fisher's least significant differences post-hoc tests), whereas all other blood cell counts remained unchanged. Infarct size was less with infusion of platelet releasate after RIPC in comparison to platelet releasate before RIPC and to platelet releasate before and after PLA, respectively (see Figure). Conclusion In response to RIPC in healthy volunteers, platelets carry soluble cardioprotective factor(s). Their precise nature must still be identified.