Effects of polyamines on apoptosis induced by simulated ischemia/reperfusion injury in cultured neonatal rat cardiomyocytes

We incubated neonatal Sprague—Dawley rat cardiomyocytes in primary culture in a medium simulating ischemia (consisting of hypoxia plus serum deprivation) for 2 h, then re‐incubated them for 24 h in normal culture medium to establish a model of simulated ischemia/reperfusion (I/R) injury. Apoptotic cell death was measured by MTT assay, TUNEL staining and flow cytometry. Morphological alterations were assessed by transmission electron microscopy, the expression of caspases‐3 and −9 and Bcl‐2 and the release of cytochrome c by Western blotting, and the intracellular free‐calcium concentration ([Ca 2+ ] i ) by laser confocal scanning microscopy. The results showed that pretreatment with 10 μmol/l spermine or spermidine significantly inhibited apoptosis in the I/R cells, suppressed the expression of caspases‐3 and −9 and cytochrome c release, up‐regulated Bcl‐2 expression and decreased [Ca 2+ ] i . However, pretreatment with 10 μmol/l putrescine had the opposite effects. Evidence for this “double‐edged” effect of polyamines on apoptosis in I/R‐injured cardiomyocytes is presented for the first time. It may suggest a novel pharmacological target for preventing and treating cardiac ischemia/reperfusion injury.