LV infarct size and AAR were determined as previously described (34). The ligature around the LAD was retied through the previous ligation, and 0.2 ml of 2% Evans blue dye was injected retrogradely through the aorta after 24 hours of reperfusion. The dye distributed in the heart to areas perfused by the nonligated coronary arteries. The heart was then excised and sliced into five 1.2-mm-thick sections in the short-axis orientation. The sections were then stained with 2% TTC (Sigma-Aldrich) in phosphate-buffered saline (PBS), followed by PBS-only incubation, and digitally photographed. Infarct area was the TTC-negative staining region. The AAR consisted of the Evans blue–negative region (TTC-positive and TTC-negative regions). The ANAR consisted of Evans blue–positive regions. These regions were quantified by SigmaScan Pro (SPSS Science). Myocardial percentage of AAR was computed as the infarct area/AAR, and the AAR was calculated as a percentage total LV [AAR/(AAR + ANAR)].

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