A homeodomain containing transcription factor PDX-1 can induce beta-cell-specific gene expressions in some non-beta-cells and may therefore be useful for future diabetes gene/cell therapy. Among the potential target organs or tissues for transcription factor-mediated induction of beta-cell-like differentiation are the intestinal epithelial cells. They have certain merits over other tissues and organs in terms of accessibility for gene delivery and of similarity in developmental background to the pancreatic primordium. In this study, we used an intestinal epithelium-derived cell line, IEC-6 cells, and investigated the possible effects of PDX-1 expression in those cells. By exogenous expression of the PDX-1 gene, IEC-6 cells started expressing multiple beta-cell-specific genes such as amylin, glucokinase, and Nkx6.1, which were not found in the original IEC-6 cells. Insulin gene expression, which was missing initially even in the PDX-1-transfected IEC-6 cells, became detectable when the cells were transplanted under the renal capsule of a rat. When the PDX-1(+) IEC-6 cells were kept in vitro, treatment with betacellulin could also confer insulin gene expression to them. Although insulin secretory granules became visible by electron microscopy, they were secreted regardless of glucose concentration. The in vivo or in vitro inductions of the insulin gene expression were not observed in the PDX-1(-) IEC-6 cells. Thus, our present observations demonstrate the potency of intestinal epithelial cells as a tool for diabetes gene/cell therapy and provide further support for the potency of PDX-1 in driving beta-cell-like differentiation in non-beta-cells.