| Budget Amount *help |
¥3,300,000 (Direct Cost: ¥3,300,000)
Fiscal Year 2001: ¥600,000 (Direct Cost: ¥600,000)
Fiscal Year 2000: ¥1,200,000 (Direct Cost: ¥1,200,000)
Fiscal Year 1999: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Research Abstract |
The heterocyclic amine, 3-ambo-l,4-dimethyl-5 H-pyrido[4,3-,b]indole (Trp-P-l) is derived from tryptophan in cooked food. This compound reveals carcinogenicity in the liver after administration of it to rodents. In this research project, we investigated the mechanism of apoptosis induced by Trp-P-1 in primary cultured rat hepatocytes and found that Trp-P-1 induced caspase-dependent apoptosis. In the case of parenchymal hepatocytes, Trp-P-1 induced a mitochondria-dependent apoptotic pathway in which caspase-9 acted as an apical enzyme, resulting in the activation of Nuc-18-like endonuclease to form DNA ladder structure. When nonparenchymal liver cells were cocultured with parenchymal cells, apoptosis was occurred rapidly. Trp-P-1 mainly drove the caspase-8-mediated pathway that involved in Bid accompanied by a delay in the p53/NF-κB-mediated side pathway that involved Bax, Bcl-2, and caspase-9. Trp-P-1 also induced apoptosis in immunocytes such as thymocytes, splenocytes, and monocytes.
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In both thymocytes and splenocytes, Trp-P-1 caused the activation of caspase-3-like proteases, but it did not cause DNA fragmentation in thymocytes although Trp-P-1 activated 32-34 kDa nucleases. These results indicated that Trp-P-1 induced apoptosis in both splenocytes and thymocytes by different mechanisms. In mononuclear cells, Trp-P-1 induced caspase-8-dependent apoptosis. It was noteworthy that Trp-P-1 at 1 nM was sufficient to cause DNA fragmentation after 24 h in culture, We, next, investigated uptake of Trp-P-1 into the target cells and found that this compound was incorporated into rat splenocytes, thymocytes, and hepatocytes through monoamine transporters arid induced apoptosis. Regarding with the relationship between apoptosis and carcinogenicity, Trp-P-1 did not require the metabolic activation to induce apoptosis, although the metabolic activation is necessary for carcinogenicity. We also found an increase in DNA binding activity of transcription factors concerning both with apoptosis and carcinogenicity. However, it needs further studies on the involvement of reactive oxygen species and of DNA damage in apoptosis. Less
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