Stimulation of Human and Rat Islet β-Cell Proliferation with Retention of Function by the Homeodomain Transcription Factor Nkx6.1

Type 1 diabetes results from autoimmune destruction of insulin-producing β cells in the islets of Langerhans, whereas type 2 diabetes involves loss of glucose-stimulated insulin secretion (GSIS) and a gradual diminution of β-cell mass (45). Insulin injection therapy has been the standard treatment for type 1 diabetes since the discovery of the hormone more than 80 years ago. Islet transplantation has been investigated as an alternative to insulin injection, but a major obstacle to broad application of this approach has been an inadequate supply of human islets (21). Pharmacotherapy of type 2 diabetes includes administration of agents that enhance insulin secretion, but these drugs often lose efficacy over time and cause complications such as hypoglycemia (30). Moreover, no controlled strategy for restoration of β-cell mass has been identified for the type 2 disease. Thus, a more complete understanding of the mechanisms that control islet β-cell growth and function is required in order to develop more effective therapies for both major forms of diabetes. Several members of the homeodomain family of transcription factors, including Pdx1, Hb9/Hlxb9, Nkx2.2, Nkx6.1, Isl-1, Pax6, and Pax4, are involved in the development of the various pancreatic islet cell types (α, β, and δ) and maintenance of their differentiated functions (9, 19). Nkx6.1 was initially isolated in a screen for homeodomain-containing transcripts in a hamster insulinoma cell cDNA library (40). Subsequently, functional roles for Nkx6.1 in development of the nervous system (38) and islet β cells (41) were described. Disruption of Nkx6.1 in transgenic mice resulted in a >90% decrease in functional β-cell mass, with no apparent defect in the development of other islet cell types (41). The dearth of β cells in this model was attributed to the absence of the secondary transition, in which insulin-positive cells normally undergo rapid proliferation at approximately embryonic day 15 in the developing mouse embryo. The paucity of β cells in Nkx6.1-knockout mice limits further phenotypic characterization, leaving potential roles of the transcription factor in maintenance of mature β-cell function and growth unexplored. To address this issue, we have recently begun to manipulate Nkx6.1 expression in mature β cells, resulting in the demonstration of new roles in suppression of glucagon expression and control of GSIS (42). In the present study, we showed that Nkx6.1 overexpression strongly stimulates indices of islet cell replication, such as [methyl-3H]thymidine and 5-bromo-2′-deoxyuridine (BrdU) incorporation, as well as increasing cell numbers in both rat and human islets, whereas small interfering RNA (siRNA)-mediated suppression of Nkx6.1 has the opposite effect. We also demonstrate that the growth-promoting effect of Nkx6.1 is mediated by upregulation of a broad array of cell cycle control genes, some of which are shown by chromatin immunoprecipitation (ChIP) assays to be direct targets for Nkx6.1 binding. Finally, we show that the β-cell-mitogenic effects of Nkx6.1 are accompanied by retention or enhancement of GSIS in human and rat islets, respectively.