The overall goal of this proposal is to define mechanisms governing pancreatic ?-cell proliferation. Variations in insulin demand resulting from physiological and pathological states require adaptive functional changes in pancreatic ?-cells. These adaptive changes include increased insulin synthesis and secretion as well as hyperplasia and possibly formation of new ?-cells from progenitor cells. Of these mechanisms, ?-cell proliferation and regulation of cell cycle appear predominant in establishing and determining ?-cell mass. Inability of adequate ?-cell response to metabolic challenges results in hyperglycemia and frank diabetes mellitus. The incretin hormone glucagon-like peptide-1 (GLP-1) stimulates murine pancreatic ?-cell proliferation. Upon binding to its receptor on ?-cells, GLP-1 stimulates both cyclic AMP (cAMP) and phosphoinositol 3 kinase (PI3K) activation. Both cAMP and PI3K signaling pathways are implicated in ?-cell proliferation. The cAMP signal activates protein kinase A (PKA), which phosphorylates the nuclear cAMP response element binding protein (CREB). Phosphorylated CREB recruits the nuclear co-activators CREB binding protein (CBP), and the related protein p300, which stimulate gene transcription through histone deacetylase activity and stimulating RNA polymerase. On the other hand, PI3 kinase activates protein kinase B (PKB = Akt). PKB phosphorylates transcription factor FoxO1, which leads to its nuclear exclusion, thereby altering gene expression including derepressing transcription of pancreas duodenum homeobox-1 (PDX-1). PDX-1 is required for GLP-1 effects on ?-cell proliferation and for ?-cell mass adaptation to insulin resistance. Our preliminary results indicate that GLP-1 stimulates, via the cAMP-PKA-CREB pathway, transcription of the cell cycle positive regulator cyclin A2. Cyclin A2 upregulation is sufficient to augment mouse ?-cell proliferation in vitro. Furthermore, in ?-cells, PDX-1 is required for GLP-1 stimulated cAMP production, subsequent PKA activation and cyclin A2 expression. Thus, our findings suggest an interdependence of cAMP and PI3K mediated GLP-1 receptor signaling and a mechanistic link between PDX-1 and GLP-1 induced ?-cell proliferation.
The specific aims of this proposal are to further elucidate the role of GLP-1 induced cAMP/PKA signaling and of cyclin A2 in regulating ?-cell proliferation.
Aim 1 or our proposal is to examine in vivo cAMP/PKA-signaling effects in ?-cell proliferation and cycle regulatory proteins by ?- cell-specific ablation of the PKA regulatory subunit.
Aim 2 is to examine the in vivo effects of ?-cell specific cyclin A2 upregulation on islet mass and ?-cell function.
Aim 3 is to assess the consequences of ?-cell specific cyclin A2 ablation on ?-cell proliferation during development and in adult animals in response to GLP-1 and as adaptation to high-fat diet-induced insulin resistance. These studies of in vivo mechanisms of ?-cell proliferation may provide therapeutic approaches for diabetes mellitus in humans.

Public Health Relevance

Diabetes mellitus results from failure of insulin-producing ?-cells to meet metabolic demands, and adaptation to the demands includes ?-cell proliferation (i.e., duplication). In this proposal we aim to understand mechanism(s) governing pancreatic ?-cell proliferation, in particular those mediated by the incretin hormone glucagon-like peptide-1. The studies proposed herein are significant because they may lead to therapeutic approaches for treating diabetes mellitus in humans.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK081472-04
Application #
8272634
Study Section
Cellular Aspects of Diabetes and Obesity Study Section (CADO)
Program Officer
Appel, Michael C
Project Start
2009-06-10
Project End
2014-04-30
Budget Start
2012-05-01
Budget End
2013-04-30
Support Year
4
Fiscal Year
2012
Total Cost
$390,151
Indirect Cost
$152,254
Name
Johns Hopkins University
Department
Pediatrics
Type
Schools of Medicine
DUNS #
001910777
City
Baltimore
State
MD
Country
United States
Zip Code
21218
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