Nonanticoagulant heparin reduces myocyte Na+ and Ca2+ loading during simulated ischemia and decreases reperfusion injury

Barry WH, Zhang XQ, Halkos ME, Vinten-Johansen J, Saegusa N, Spitzer KW, Matsuoka N, Sheets M, Rao NV, Kennedy TP. Nonanticoagulant heparin reduces myocyte Na+ and Ca2+ loading during simulated ischemia and decreases reperfusion injury. Am J Physiol Heart Circ Physiol 298: H102-H111, 2010. First published October 23, 2009; doi:10.1152/ajpheart.00316.2009.-Heparin desulfated at the 2-O and 3-O positions (ODSH) decreases canine myocardial reperfusion injury. We hypothesized that this occurs from effects on ion channels rather than solely from anti-inflammatory activities, as previously proposed. We studied closed-chest pigs with balloon left anterior descending coronary artery occlusion (75-min) and reperfusion (3-h). ODSH effects on [Na+](i) (Na Green) and [Ca2+](i) (Fluo-3) were measured by flow cytometry in rabbit ventricular myocytes after 45-min of simulated ischemia [metabolic inhibition with 2 mM cyanide, 0 glucose, 37 degrees C, pacing at 0.5 Hz; i.e., pacing-metabolic inhibition (PMI)]. Na+/Ca2+ exchange (NCX) activity and Na+ channel function were assessed by voltage clamping. ODSH (15 mg/kg) 5 min before reperfusion significantly decreased myocardial necrosis, but neutrophil influx into reperfused myocardium was not consistently reduced. ODSH (100 mu g/ml) reduced [Na+](i) and [Ca2+](i) during PMI. The NCX inhibitor KB-R7943 (10 mu M) or the late Na+ current (INa-L) inhibitor ranolazine (10 mu M) reduced [Ca2+](i) during PMI and prevented effects of ODSH on Ca2+ loading. ODSH also reduced the increase in Na+ loading in paced myocytes caused by 10 nM sea anemone toxin II, a selective activator of INa-L. ODSH directly stimulated NCX and reduced INa-L. These results suggest that in the intact heart ODSH reduces Na+ influx during early reperfusion, when INa-L is activated by a burst of reactive oxygen production. This reduces Na+ overload and thus Ca2+ influx via NCX. Stimulation of Ca2+ extrusion via NCX later after reperfusion may also reduce myocyte Ca2+ loading and decrease infarct size.