| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2001: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 2000: ¥2,400,000 (Direct Cost: ¥2,400,000)
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| Research Abstract |
Hypertonicity-induced cell damage and apoptosis were investigated in MDCK cells and peritoneal mesothelial cells. In MDCK cells, when the cells were exposed to 700 mOsm medium for 24h, TUNEL positive cells were demonstrated, indicating that apoptosis involved in the hypertonicity-induced cell death. Caspase-3 activity of the hypertonic cells increased about 20 fold the value of isotonic cells. Addition of ImM betaine, one of the predominant osmolytes in kidney medulla, inhibited the hypertonicity-induced apoptosis and activation of caspase-3. In peritoneal mesotelial cells, when the medium osmolality was increased to 700 mOsm, marked activation of caspase-3 and -9 but not caspase-8 were evident within 4h. Addition of zVAD-fmk, a broad-spectrum caspase inhibitor, partially protected the hypertonicity-induced cell death.
Cytochrome c release from mitochondria occurred at the early phase of hypertonicity-induced apoptosis, suggesting that mitochondrial pathway plays an important role in the hypertonicity-induced apoptosis. To assess the mechanism of cytochrome-c release, protein levels of p53, Bax, Bcl-2, and Bel-XL were investigated. When the cells were exposed to 700mOsm, p53 level decreased rapidly within Ih. Protein levels of Bax, Bcl-2, and Bel-XL did not change after the hypertonic exposure, suggesting that these proteins did not contribute to the induction of the apoptosis. We concluded that Betaine had a potent protective effect on the hypertonicity induced apoptosis in MDCK cells. The results suggested that mitochondrial pathway contributed to the hypertonicity induced apoptosis.
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