Abstract 379: Insulin-induced Cardioprotection against Ischemia-Reperfusion Damage: Role for NO and Upregulation of the Dual-specificity Phosphatase MKP-1
Insulin-induced MKP-1 protein expression in cardiomyocytes has been shown to require ERKp44/p42, JNK and PI3-K/Akt pathways. Insulin also stimulates eNOS activation and NO production. We propose that NO contributes to the cardioprotective actions of insulin via upregulation of MKP-1.
Methods: Isolated perfused working hearts (preload 15cm H2O; afterload 100cm H2O) were subjected to 35 min low flow ischemia (LFI: 0.2ml/min), in the presence or absence of insulin (100nM). During reperfusion mechanical recovery was assessed and hearts freeze-clamped at different time-points. MKP-1 expression, p38MAPK and PKB/Akt activation were determined by Western blot (n=6 hearts/series).
Results: Aortic output (AO), cardiac output and work performance during reperfusion after 35min LFI were significantly higher in insulin-treated hearts (AO before LFI: 37±0.8ml/min; After LFI: untreated: 6±4ml/min; insulin: 21.5±4ml/min, p<0.05 vs untreated). This was associated with a significant 1.45±0.11 fold upregulation of MKP-1(p<0.05 vs untreated), reduced activation of p38MAPK (fold increase vs untreated; 0.54±0.17, p<0.05) and increased Akt activation ((fold increase vs untreated; 1.52±0.1 p<0.05) at 30 min reperfusion. Pretreatment with the NOS inhibitor L-NAME (30 microM) completely abolished the beneficial effects of insulin on mechanical recovery during reperfusion (AO: 0ml/min) as well as upregulation of MKP-1. Activation of p38MAPK (fold increase vs untreated:0.44±0.08) remained reduced, while Akt activation was significantly lower in presence of insulin and L-NAME (fold increase:insulin:1.52±0; insulin+L-NAME: 0.81±0.11; p<0.05).
Conclusion: Insulin-induced NOS activation associated with upregulation of the phosphatase MKP-1, contributes significantly to its cardioprotective effects. The reduced activation of the proapoptotic p38MAPK after 30 min reperfusion may be due to upregulation of MKP-1 by insulin. Inhibition of NOS activation abolished the effects of insulin on functional recovery and MKP-1 upregulation. The data obtained with L-NAME suggest that upregulation of Akt by insulin is NO dependent.