Generation of fibroblasts overexpressing liver-specific PEPCK in a miniature pig model of human type 2 diabetes mellitus

Molecular Medicine Reports
Yu-Kyung KimEui-Bae Jeung

Abstract

Type 2 diabetes mellitus (T2DM) is one of the most common complex metabolic disorders in humans, and is characterized by hyperglycemia and metabolic alterations. In T2DM, fasting hyperglycemia is attributed to excessive hepatic glucose production, and increased gluconeogenesis has been ascribed to increased transcriptional expression of phosphoenolpyruvate carboxykinase (PEPCK). In this study, we analyzed porcine PEPCK promoter activities to generate liver-specific expression vectors. We generated miniature pig fibroblasts overexpressing PEPCK via transgenes to provide an animal model of human T2DM. Various regions of the promoter showed high levels of activity in the presence of glucocorticoids, a PEPCK gene inducer, in liver cells compared to a positive control promoter. The selected promoter region for a liver-specific expression system was adopted based on the current targeting vector containing two selection markers, green fluorescence protein and a neomycin-resistance gene. The linearized vector was introduced into pig fibroblasts which facilitated liver-specific PEPCK overexpression and screening according to the two selection markers. In the present study, we used a liposome-mediated transfection protocol rather than a ...Continue Reading

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