| Budget Amount *help |
¥2,900,000 (Direct Cost: ¥2,900,000)
Fiscal Year 2000: ¥1,400,000 (Direct Cost: ¥1,400,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Research Abstract |
Although apoptosis and necrosis may appear to be distinct forms of cell death, recent studies suggest that the two may represent different outcomes of a common pathway. In ischemic myocardium, apoptosis appears early, while energy stores are presumably still available, followed only later by necrosis. The present study examined the possibility that intracellular ATP levels dictate whether hypoxic cardiac myocytes die by apoptosis or necrosis. Neonatal rat cardiac myocytes in primary culture were exposed to continuous hypoxia, during which the intracellular ATP concentration was modulated by varying the glucose content in the medium. The form of cell death was determined at the end of the hypoxic exposure. Under total glucose deprivation, ATP dropped precipitously and cell death occurred exclusively by necrosis as determined by creatine kinase release, nuclear staining with ethidium homodimer-1, and smearing on DNA agarose gets. However, with increasing glucose concentrations (10, 20, 50, 100 mg/dL) cellular ATP increased correspondingly, and apoptosis progressively replaced necrosis until it became the sole form of cell death, as determined by nuclear morphology, DNA fragmentation on agarose gets, and caspase-3 activation. The data showed a significantly positive correlation between myocyte ATP content and the percentage of apoptotic cells. Loss of mitochondrial membrane potential and cytochrome c release from the mitochondria was observed in both the apoptotic and necrotic cells. Increased lactate production and a lack of effect by the mitochondrial inhibitor oligomycin indicated that ATP was generated exclusively through glycolysis. We demonstrate that ATP, generated through glycolysis, is a critical determinant of the form of cell death in hypoxic cardiac myocytes. Our data suggest that necrosis and apoptosis represent different outcomes of the same pathway. In the absence of ATP, necrosis prevails. However, the presence of ATP promotes and favors apoptosis.
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