Induction of cardiac CD39 during murine myocardial ischemia in vivo. (A) Murine model of cardiac ischemia. To study in vivo expression of cardiac CD39 during cardiac ischemia, we exposed mice to in vivo coronary artery ligation (B) over indicated time periods (0-60 minutes) followed by 2 hours of reperfusion or (C) exposed mice to 45 minutes of coronary ischemia followed by indicated time periods of reperfusion. A indicates anesthesia induction; T, thoracotomy. (B) CD39 transcript from myocardial biopsies of the area at risk. C57BL/6 mice were subjected to myocardial ischemia over 0 to 60 minutes of ischemia followed by 2 hours of reperfusion, and cardiac tissue from the area at risk was excised, flash frozen, and CD39 transcript levels were determined by real-time RT-PCR. Data were calculated relative to an internal housekeeping gene (β-actin) and are expressed as fold change compared with control (0 minutes of myocardial ischemia ± SEM at each indicated time, n = 6; *P < .01). (C) C57BL/6 mice were subjected to 45 minutes of in situ ligation of the left coronary artery, and myocardial tissue from the area at risk was excised after indicated reperfusion times (0-120 minutes), lysed, proteins resolved by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE), and transferred to nitrocellulose. Membranes were probed with CD39 antibody. The same blot was probed for β-actin expression as a control for protein loading. One representative experiment of 3 is shown. (D) C57BL/6 mice were subjected to 45 minutes of myocardial ischemia, and cardiac tissue from the area at risk was harvested after 120 minutes of reperfusion, sectioned, and stained using CD39 antibody. Cardiac tissue from sham-operated mice served as control (original magnification ×400 or ×800, as indicated). To control for nonspecific staining, IgG was used at identical concentrations and staining conditions as the target primary antibodies for CD39.