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KMID : 0624219950030000039
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Focus on Genetic Science
1995 Volume.3 No. 0 p.39 ~ p.49
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Development of Insulin-Secreting Non-endocrine Cell
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Yoon, Do-jun
Kim, Jae-woo/Kim, Yu-kyong/Ahn, Yong-ho
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Abstract
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Treatment of insulin dependent diabetes mellitus (IDDM) requires insulin replacement to the patients. One of the best way to treat IDDM patients may be accomplished by transplanting a pancreatic tissue which functions normally. However, the adoption of this methods is limited by the availability of the human tissues as well as tissue rejection problems. Recently, the advent of genetic engineering technics enabled us to transfer foreign genes of interests into various cells and establish an “artificial β-cells” capable of secreting insulin in response to plasma glucose level.
In this study, we have designed a study to establish an “artificial β-cells” by transfecting liver/pancreatic β-cells type glucose transporter (GLUT2) cDNA and genomic DNA of insulin. Because GLUT2 molecules on the plasma membranes act as a sensor of glucose outside the cell and promote the secretion of insulin from the cells, cotransfection of GLUT2 cDNA along with insulin gene will translate the GLUT2 molecules necessary for glucose transport into the cells leading to insulin secretion. Therefore, we have subcloned GLUT2 cDNA and insulin gene into separate eukaryotic expression vectors and transfected them to Chinese hamster ovary cells. The stable cell lines harboring GLUT2 cDNA and insulin gene were selected by G418. The surviving clones were harvested and subjected to Southern blot analysis by digesting the chromosomal DNA either with BamHl for insulin gene detection or Xhol/Smal double digestion for GLUT2 gene detection. Trough this analysis, we were able to find out 3 clones positive for both GLUT2 gene and insulin gene. Of these clones, clone 5 cell secreted insulin 3 times as much as that of the control CHO cells.
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