Phosphorylation of GSK-3b is Required for Intralipid to Protect the Heart Against Ischemia/Reperfusion Injury

Abstract

Recently we found that administration of Intralipid (ILP) during reperfusion significantly improves post-ischemic cardiac function and reduces the myocardial infarct size by ∼70% , both in the isolated mouse heart and in-vivo rat heart. Here we investigated whether ILP-induced cardioprotection is mediated through the inhibition of GSK-3b. Wild type (WT) C57BL/6 male mice and GSK-3b Knockin (KI) were used. The isolated hearts were subjected to 20 min of global normothermic ischemia followed by reperfusion with 1% ILP (40 min for heart function and infarct size and 10 min for calcium retention capacity (CRC) experiments). The left ventricular (LV) systolic pressure, LV end-diastolic pressure (LVDP), heart rate, maximum velocity of contraction (dP/dt max) and maximum velocity of relaxation (dP/dt min) were recorded. Myocardial necrosis was assessed using TTC staining. Mitochondria were isolated to measure CRC by calculating the number of pulses required to trigger the opening of the mitochondrial transition permeability pore as a result of calcium overload. Before ischemia, the baseline RPP, LVDP, dP/dtmax and dP/dtmin in GSK-3b KI mice were similar to WT. However, the functional recovery during reperfusion was very poor in GSK-3b KI mice. At the end of 40 min of reperfusion, the RPP was 1990±499 in GSK-3b KI mice vs. 15405±1011mmHg∗beats/min in WT, the LV dP/dtmax was 239.7±17.8 in GSK-3b KI vs. 2703±145 mmHg/s in WT and the LV dP/dtmin was 219±14 in GSK-3b KI vs. 1683±66 mmHg/s in WT. The infarct size was significantly larger compared to WT (45.3±10.3 vs. 16.7±2.33% in WT, P<0.001). Postischemic administration of ILP in GSK-3b KI mice demonstrated lower CRC than WT (1.3±0.1 vs. 2.7±0.06 µM/mg-mitochondrial protein in WT). In conclusions, these data demonstrate that phosphoryltaion of GSK-3b is required for the cardioprotective action of ILP.