Abstract
Elucidating the regulation of glucose-stimulated insulin secretion (GSIS) in pancreatic islet β cells is important for understanding and treating diabetes. MIN6 cells, a transformed β-cell line derived from a mouse insulinoma, retain GSIS and are a popular in vitro model for insulin secretion. However, in long-term culture, MIN6 cells' GSIS capacity is lost. We previously isolated a subclone, MIN6 clone 4, from the parental MIN6 cells, that shows well-regulated insulin secretion in response to glucose, glybenclamide, and KCl, even after prolonged culture. To investigate the molecular mechanisms responsible for preserving GSIS in this subclone, we compared four groups of MIN6 cells: Pr-LP (parental MIN6, low passage number), Pr-HP (parental MIN6, high passage number), C4-LP (MIN6 clone 4, low passage number), and C4-HP (MIN6 clone 4, high passage number). Based on their capacity for GSIS, we designated the Pr-LP, C4-LP, and C4-HP cells as “responder cells.” In a DNA microarray analysis, we identified a group of genes with high expression in responder cells (“responder genes”), but extremely low expression in the Pr-HP cells. Another group of genes (“non-responder genes”) was expressed at high levels in the Pr-HP cells, but at extremely low levels in the responder cells. Some of the responder genes were involved in secretory machinery or glucose metabolism, including Chrebp, Scgn, and Syt7. Among the non-responder genes were Car2, Maf, and Gcg, which are not normally expressed in islet β cells. Interestingly, we found a disproportionate number of known imprinted genes among the responder genes. Our findings suggest that the global expression profiling of GSIS-competent and GSIS-incompetent MIN6 cells will help delineate the gene regulatory networks for insulin secretion.
Highlights
MIN6 cells, which were isolated from an insulinoma of a transgenic mouse expressing the SV40 T antigen in pancreatic islet b cells, retain some features of differentiated pancreatic b cells [1]
Several reports have described the isolation of glucose-responsive and -unresponsive MIN6 cell subclones, and these subclones have been used to identify genes associated with glucose-stimulated insulin secretion (GSIS) [5,6]
Insulin secretion and insulin content Insulin secretory capacity was compared by static incubation among parental MIN6 cells at 17–20 passages (Pr-LP) and 35–40 passages (Pr-HP) and MIN6 clone 4 cells at 17–20 passages (C4LP) and 40–50 passages (C4-HP) (Figure 1 and 2)
Summary
MIN6 cells, which were isolated from an insulinoma of a transgenic mouse expressing the SV40 T antigen in pancreatic islet b cells, retain some features of differentiated pancreatic b cells [1]. Several reports have described the isolation of glucose-responsive and -unresponsive MIN6 cell subclones, and these subclones have been used to identify genes associated with glucose-stimulated insulin secretion (GSIS) [5,6]. These reports did not rule out the possibility that this differential gene expression was attributable to clonal variation. Impairment of glucose and lipid oxidation was suggested to be involved in the loss of GSIS in high passage MIN6 cells [9]
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