Thrombin increases cardiomyocyte acute cell death after ischemia and reperfusion

Thrombin exerts multiple actions on cardiomyocytes leading to increased intracellular Na+ and Ca2+ concentrations, and to activation of a Ca2+-independent PLA2, and has been proposed to favor the genesis of arrhythmias and ischemic injury in acute coronary syndromes. However, the influence of thrombin on cardiomyocyte cell death during ischemia-reperfusion has not been studied. A beneficial influence of low thrombin concentrations has been described in other cell types. HL-1 cardiomyocytes were subjected to simulated ischemia (SI) and reperfusion (SR) and cell death was assessed by means of LDH release to the incubation media. Thrombin dose-dependently increased cell death in normoxic cells, in cells subjected to SI, and in cells subjected to SR (by 20 ± 8%, 95 ± 32% and 35 ± 9%, respectively, at 100 U/ml). The effects of thrombin were associated to increased cytosolic Ca2+ overload, mimicked by 100 μM PAR-1 agonist peptide SFLLRNPNDKYEPF, and reversed by the direct thrombin inhibitor lepirudin (IC50 = 1.3 ± 0.2 μg/ml). The presence of thrombin during simulated ischemia-reperfusion increases cardiomyocyte cell death by a mechanism that involves activation of PAR-1 receptors and can be prevented by the direct thrombin inhibitor lepirudin.