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


Corresponding author at: Department of Pathophysiology, Harbin Medical University, Baojian Road, Harbin 150086, China. Tel.: +86 451 8667 4548; fax: +86 451 8750 3325. xucq45@126.com


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 ([Ca2+]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 [Ca2+]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.