The purpose of this study was to evaluate the functional and biochemical effects in immature cardiac myocytes under hypoxic and hypothermic conditions. Cardiac myocytes were isolated from neonatal rat ventricles and cultured for 4 days, after which 12.5×10
5 myocytes/flask were incubated under 3% hypoxic conditions at 4°C, 10°C, 15°C, 20°C, 25°C, and 37°C for 6, 12, and 24h. After each hypoxic incubation, creatine kinase (CK) and lactate dehydrogenase (LDH) were measured in the incubation medium. The myocytes were then cultured for an additional 24h at 37°C to evaluate the recovery of the myocyte beating rate. In the 4°C and 37°C groups, the myocyte beating rate recovery markedly decreased with increasing incubation times from 78.1% and 97.2% at 6h to 0.0% and 38.4% of the control, which was the beating rate prior to hypoxic incubation, at 24h, respectively. However, in the 10°C, 15°C, and 25°C groups, this value decreased significantly only at 24h. In the 20°C group, beating rate completely recovered in 24h. A marked increase was found in the release of CK and LDH in the 4°C group from 28.5mIU/flask and 232.9mIU/flask at 6h to 83.7mIU/flask and 640.7 mIU/flask at 24h, respectively. However, in the 25°C and 37°C groups, this release was significantly increased only at 24h. In the 15°C and 20°C groups, no significant increases were observed over 24h. Below 15°C, hypothermia induced cellular damage both functionally and biochemically, and the greatest damage was observed at 4°C. Above 25°C, the damage was due to hypoxia. Thus, a temperature of 15°C to 20°C appears to be suitable for hypothermic preservation of immature myocardium.
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