| BackgroundThe involvement of an inflammatory response in the pathophysiology of myocardial ischemia/reperfusion (MI/R) has been already recognized. Adhesion of polymorphonuclear leukocytes (PMN) to endothelial cells (EC) is an early and requisite event in the inflammatory response. Therefore, the PMN/EC adherence during reperfusion represents one such important target for therapeutic intervention because of the pivotal role played by PMN in deleterious events following reflow.Insulin infusion has been shown to protect endothelium in the critically ill patients, but the underlying mechanisms are still unclear. Recent results from our laboratory as well as others'have revealed that insulin causes a suppression of several proinflammatory transcription factors including nuclear factor-κB (NF-κB),early growth response-1 (Egr-1) and tumor necrosis factor-α(TNF-α),and acts as an anti-inflammatory agent. Meanwhile, amounting evidence has shown that insulin can protect the MI/R-injured heart, and insulin-induced anti-apoptosis and coronary artery protection may be one of the mechanisms. However, whether insulin could exert an anti-adhesive effect on the coronary endothelium and thus inhibit MI/R-induced injury remains largely elusive.AimsTherefore, the aim of the present study was to investigate whether insulin inhibits PMN/EC adherence and reduces MI/R-induced injury.MethodsPMNs were isolated from arterious blood of anesthetized rabbits before MI/R using standard dextran sedimentation and gradient separation on Lympholyte-Rabbit (density = 1.0965 g/mL). PMNs labeled with PKH26 red fluorescent cell linker were prepared. Anesthetized rabbits were subjected to 45 min MI followed by R for 4 h and were randomly received one of the following treatments: saline (0.9% NaCl), GIK (glucose: 250 g/L, insulin: 60 U/L and KCl: 80 mmol/L), or GK (n = 10 in each group) at 2 ml·kg?1·h?1, beginning 15 min after MI and continuing throughout the 4 h reperfusion.Blood glucose, electrolytes, arterial blood pressure and cardiac function were monitored throughout the experiment. Plasma creatine kinase (CK) activity was measured spectrophotometrically. Myocardial infarction and PMN accumulation (MPO activity) were determined in a blinded manner. In addition, P-selectin and ICAM-1 were immunohistochemically detected after 20 min and 24 h resperctively.Left circumflex coronary artery was isolated after 4 h reperfusion and was cut into segments to measure adherence of fluorescently labeled meutrophils by epifluorescence microscopy.ResultsAdministration of GIK, but not GK, significantly attenuated myocardial injury as evidenced by reduced infarct size (35.7±2.7% vs. 48.6±3.2% in vehicle, P<0.01, n=10) and plasma creatine kinase activity,and improved recovery of cardiac systolic/diastolic function as evidenced by increased +LVdP/dtmax and ?LVdP/dtmax at the end of reperfusion in rabbits subjected to MI/R.GIK markedly decreased the expressions of coronary endothelial P-selectin and intercellular adhesion molecule-1 (ICAM-1) (10.9±2.7% vs. 24.8±3.5% and 32.4±2.2% vs. 58.6±3.7% in vehicle, P<0.01) at 20 min and 24 h following reperfusion respectively. Sequentially,insulin inhibited leukocyte-endothelium adherence (108±8 vs. 156±10 PMNs/mm2 in vehicle, P<0.01, n=5), and thus attenuated PMN accumulation as manifested by myeloperoxidase (MPO) activity (0.62±0.08 vs. 1.18±0.11 U/100g tissue, P<0.01) in ischemic/reperfused hearts compared with saline group. While GK infusion didn't show these benefits in MI/R-injured rabbits suggesting that insulin may play a pivotal role in the anti-inflammatory effect of GIK on I/R myocardium.ConclusionIn the present study, we demonstrate firstly that insulin reduces coronary endothelial P-selectin and ICAM-1 expressions, inhibits PMN/EC adherence, and consequently attenuates PMN accumulation in ischemic/reperfused rabbit hearts. The anti-adhesive property of insulin may contribute to its cardioprotection on I/R myocardium.
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