Establishment of inducible cAMP early repressor transgenic fibroblasts in a porcine model of human type 1 diabetes mellitus

초록

Diabetes mellitus is a metabolic disease caused by impaired insulin secretion from the pancreatic beta cells and increased insulin resistance in peripheral tissues. Recently, the overexpression of inducible cyclic AMP (cAMP) early repressor (ICER) I gamma in rodent pancreatic beta cells was found to induce insulin deficiency and glucagon overproduction similar to that found in human diabetes mellitus. ICER I gamma with only a DNA binding domain interrupts the transcriptional regulation of the cAMP responsive element-binding protein (CREB) target genes. Based on this information, we hypothesized that the overexpression of ICER I gamma, the most powerful competitor to CREB, could be useful for generating a pig model of diabetes. First, we evaluated the promoter activities of the human insulin gene for the beta cell-specific overexpression of ICER I gamma in the pig pancreas. The maximum promoter activity region [-1,431 nucleotides (nt) to +1 nt, +1 = the transcriptional start site] of the insulin gene presented an activity level 3-fold higher than a promoterless construct. Second, ICER I gamma overexpression controlled by this promoter region significantly blocked the glucose-mediated insulin transcription, such as that regulated by the viral promoter in the pancreatic beta cell line, MIN6. This suggests that the human insulin promoter may facilitate the overexpression of ICER I gamma in porcine pancreatic beta cells. In addition, the overexpression of ICER I gamma in porcine beta cells may induce human-like type 1 diabetes mellitus in pigs. In the present study, we generated transgenic fibroblasts containing ICER I gamma cDNA controlled by the human insulin promoter, as well as two screening markers, the green fluorescence protein and the neomycin resistance gene. These fibroblasts may provide a source for somatic cell nuclear transfer to generate a pig model that mimics human diabetes mellitus.

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