Low-flow ischemia and hypoxia stimulate apoptosis in perfused hearts independently of reperfusion.

Post-ischemic reperfusion leads to apoptosis-linked loss of myocytes in cultured cells and in vivo. We tested the hypothesis that apoptosis develops without reperfusion in Langendorff-perfused hearts exposed to either low-flow ischemia (LFI) or hypoxia (H). Rat hearts were perfused with amino-acid-enriched Krebs-Henseleit buffer and exposed for 6 h to LFI (flow=2 ml/min, PO(2)=500+/-50mmHg, mean+/-SD), H (10ml/min, 120+/-15mmHg), or control conditions (C, 10ml/min, 500+/-50mmHg). At selected times, DNA-fragmentation was measured by agarose-gel electrophoresis and in situ TUNEL assay. After 6 h, the ratio (TUNEL-positive)/(total nuclei) was 0.620+/-0.027, 0.615+/-0.005, 0.404+/-0.021 in LFI, H and C, respectively. The ratio was 0.813+/-0.021 in hearts exposed to 90 min global no-flow ischemia and reperfused (5 h). To assess the role of membrane-diffusible factors, separate experiments were performed recirculating the medium and exposing hearts to LFI or H as above. The degree of apoptosis was the same in both the recirculating and non-recirculating modes. Thus, apoptosis develops by similar extents and in a time-dependent fashion in crystalloid-perfused rat hearts during LFI or H at the same oxygen shortage (flow.PO(2)), even without the reperfusion.