| Budget Amount *help |
¥3,200,000 (Direct Cost: ¥3,200,000)
Fiscal Year 2003: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 2002: ¥1,700,000 (Direct Cost: ¥1,700,000)
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| Research Abstract |
Anatomical aspects of portal circulation are quite important for recognition of blood glucose level as well as physiological regulation of insulin secretion. In this concern, we have devised a novel strategy to control diabetes mellitus in which hepatic parenchymal cells are functionally substituted for pancreatic beta cells. We performed therapeutic experiments using the Streptozotocin-induced diabetic mouse model to transducer via intravenous injection under high pressure with insulin gene that had been modified to be cleaved by the sequence-specific proteinase, furin. The insulin gene was regulated under the ubiquitous promoter, CAG, or the rat insulin promoter (RIP). Moreover, the EBV plasmid vector was employed to achieve high level transfection and expression, and EBNA1 gene was inserted downstream of CAG or RIP. In case CAG promoter was used, a remarkable decrease in fasting blood glucose was obtained, the fasting blood glucose becoming less than 100 mg/dl while untreated contro
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l group showed 350-400 mg/dl. In contrast, fasting blood glucose was not reduced after transfection with RIP plasmid, and serum insulin level did not reach a detectable level by means of the ELISA. However, blood glucose level-dependent production of insulin was obtained after systemic administration with glucose administration. These results indicate that regulation of blood glucose can be achieved in the liver, and use of promoters may allow either high level expression to reduce fasting blood glucose, or glucose concentration-dependent regulation of insulin secretion. Moreover, by means of the luciferase assay and RT-PCR analysis, we demonstrated for the first time that RIP works in the liver in a blood glucose level-dependent fashion. This may be because the hepatic parenchymal cells and pancreatic beta cells may be derived from the same precursor cells. It was also strongly suggested that functional substitution of beta cells can be effectively achieved by regenerative gene therapy based on portal circulation. Less
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