Poster abstracts

Poster number 25 submitted by Sverre Aune

Sivelestat does not reduce the burst of superoxide at reperfusion in isolated rat hearts

Sverre E. Aune (Emergency Medicine, Tzagournis Medical Research Facility), Mark G. Angelos (Emergency Medicine, Tzagournis Medical Research Facility)

Abstract:
Introduction: Sivelestat, a neutrophil-elastase inhibitor, has been shown to preserve post-ischemic cardiac function in the absence of neutrophils (1). It is unclear if a reduction in the burst of superoxide is a mechanism of Sivelestat’s protection. Aim: To test the hypothesis that Sivelestat reduces the superoxide burst at full reperfusion when administered during low flow following global ischemia in the rat heart. Methods: Hearts isolated from male Sprague-Dawley rats (n=6/group) were perfused at 75 mmHg and 37.4º C with Krebs-Henseleit buffer. Following 25 min of global ischemia, hearts were given: a) immediate full reperfusion at 75 mmHg, b) 3 min of low flow at 20% of baseline coronary flow prior to full reperfusion, or c) 3 min of 20% low flow with Sivelestat (100 µg•mL-1) prior to full reperfusion. Hearts were infused continuously for the first two minutes and then at 3 min and 10 min of full reperfusion with the superoxide radical spin trap 5,5-dimethyl-1-pyrroline-N-oxide (DMPO, 40 mM). Coronary effluent samples were collected into liquid nitrogen to test for the presence of DMPO-OH, the stable EPR-active product of the reaction of DMPO with superoxide. Frozen effluent samples were thawed and transferred to a quartz flat-cell inside an EPR-300 X-band (9.7 GHz) spectrometer. Quantification of superoxide was performed by comparing the double integral of the observed signal with that of a known aqueous solution of 4-hydroxy tempol. Significance was set at p=0.05. Results: A characteristic increase in superoxide radical formation in the heart was observed in all groups upon full reperfusion. No significant differences in superoxide radical concentration were observed between groups at any time points. Conclusions: A reduction in the superoxide burst upon full reperfusion is not a mechanism of Sivelestat’s dramatic preservation of post-ischemic cardiac function in rats. It is unclear whether other types of ROS are involved.

References:
1. Kambe M, et al. Sivelestat reduces myocardial ischemia and reperfusion injury in rat hearts even when administered after onset of myocardial ischemia. ICVTS 2009 (8):629-634.

Keywords: ischemia-reperfusion, heart, EPR