Changes in the expression of transcription factors in pancreatic AR42J cells during differentiation into insulin-producing cells.

Abstract

Pancreatic AR42J cells possess both exocrine and neuroendocrine properties and convert to insulin-producing cells upon treatment with activin A and hepatocyte growth factor (HGF). We studied changes in the mRNA expression of various transcription factors during the course of differentiation. Among the transcription factors studied, expression levels of Pax4 and neurogenin3 changed significantly. These two factors were not detected in naive cells, whereas their mRNA levels were markedly increased after treatment with activin A and HGF. Thus, these two factors were induced by activin A. P ancreatic endocrine and exocrine cells are thought to arise from common precursor cells located in the epithelium of pancreatic anlage during development (1-3). Recent genetic studies indicate that pancreatic development depends on an integrated network of distinct transcription factors operating at various levels. Several homeodomain and basic helix-loop-helix transcription factors have been postulated to play important roles in regulating differentiation of pancreatic endocrine cells. A mouse homeobox protein, insulin promoter factor-1 (IPF-1/PDX-1), is required for development of the murine pancreas (4,5). Islet-1 (Isl-1), a LIM homeodomain-containing protein, is necessary for the development of the dorsal pancreas and is required for generation of islet cells (6). In addition, inactivation of Beta-2/ NeuroD causes a striking reduction in the number of insulinproducing cells, and mice lacking the functional Pax4 gene lack differentiated ␤-and ␦-cells and fail to develop mature islets (7,8). In contrast, the number of all differentiated endocrine cell types, especially the number of ␣-cells, is markedly reduced in mice lacking the Pax6 gene (9). These transcription factors are thought to be involved in a complex regulatory network and cascade to exert their functions (10,11). However, because differentiation of pancreatic ␤-cells involves many steps, it is not clear how these steps are regulated by transcription factors.Pancreatic AR42J cells are derived from a chemically induced pancreatic tumor and express both exocrine and neuroendocrine properties (12). Upon treatment with activin A, AR42J cells stop growing, and their morphology changes significantly by extending neurites (13). In addition, activin-treated cells express mRNA for GLUT2, ATP-sensitive potassium channel, and pancreatic polypeptide (PP). Thus, activin A converts AR42J cells into endocrine cells (13,14). Furthermore, in the presence of betacellulin (14) or hepatocyte growth factor (HGF) (15), activin-treated AR42J cells further convert to insulin-producing cells. Thus, AR42J cells provide a model system to study the molecular mechanism involved in ␤-cell differentiation. The present study was conducted to investigate how the expression of islet-associated transcription factors is changed during the differentiation of AR42J cells. We also assessed the role of such transcription factors in differentiation. RESEARCH DESIGN AND METHODS Materials.Recombinant human a...

The transcription factor PAX4 acts as a survival gene in INS-1E insulinoma cells

“…Indeed, Pax4 mRNA is detected in the mouse pancreatic bud at E9.5, becoming maximal at E13.5-15.5 during the period of massive b-cell expansion and thereafter declining. In mature islets, low levels of Pax4 expression are detected in human, mouse and rat b-cells (Zhang et al, 2001;Heremans et al, 2002;Kojima et al, 2003;Zalzman et al, 2003;Brun et al, 2004Brun et al, , 2005Theis et al, 2004;Gunton et al, 2005). Consistent with its tissue-specific expression pattern, targeted disruption of the pax4 gene in mice results in the absence of mature pancreatic b-and d-cells with a commensurate increase in the glucagon-producing a-cells (Sosa-Pineda et al, 1997;Wang et al, 2004).…”

Regulation of Pax4 Paired Homeodomain Gene by Neuron-restrictive Silencer Factor

“…In the presence of hepatocyte growth factor, activin A treatment causes AR42J cells to differentiate into insulin-producing cells (18,19). Under these conditions, pax4 gene expression is substantially enhanced (18). Moreover, activin A has been shown to increase pax4 gene expression in pancreatic ␤-cell lines, independent of changes in the expression of other transcription factors (20).…”

“…Thus, up-regulation of synaptophysin gene expression in the activin A-treated AR42J cells is likely to result from decreased NRSF activity, an event that is known to occur during neuronal differentiation (17). In the presence of hepatocyte growth factor, activin A treatment causes AR42J cells to differentiate into insulin-producing cells (18,19). Under these conditions, pax4 gene expression is substantially enhanced (18).…”

p38 MAPK Is Involved in Activin A- and Hepatocyte Growth Factor-mediated Expression of Pro-endocrine Gene Neurogenin 3 in AR42J-B13 Cells

“…It has been shown already that activin A plus HGF can induce the expression of ngn3 in this cell line (12). Although HGF alone, but not activin A, induced a modest increase of NEUROG3 promoter activity, the addition of activin A plus HGF induces a marked increase of its activity (Fig.…”

“…Thus, AR42J cells provide a model system for studying the molecular mechanisms involved in ␤ cell differentiation. Probably reflecting the physiological differentiation process, activin A and HGF can also provoke the expression of ngn3 (12).…”