title: The negative cell cycle regulators, p27(Kip1), p18(Ink4c), and GSK-3, play critical role in maintaining quiescence of adult human pancreatic β-cells and restrict their ability to proliferate.
abstract: Adult human pancreatic β-cells are primarily quiescent (G0) yet the mechanisms controlling their quiescence are poorly understood. Here, we demonstrate, by immunofluorescence and confocal microscopy, abundant levels of the critical negative cell cycle regulators, p27(Kip1) and p18(Ink4c), 2 key members of cyclin-dependent kinase (CDK) inhibitor family, and glycogen synthase kinase-3 (GSK-3), a serine-threonine protein kinase, in islet β-cells of adult human pancreatic tissue. Our data show that p27(Kip1) localizes primarily in β-cell nuclei, whereas, p18(Ink4c) is mostly present in β-cell cytosol. Additionally, p-p27(S10), a phosphorylated form of p27(Kip1), which was shown to interact with and to sequester cyclinD-CDK4/6 in the cytoplasm, is present in substantial amounts in β-cell cytosol. Our immunofluorescence analysis displays similar distribution pattern of p27(Kip1), p-p27(S10), p18(Ink4c) and GSK-3 in islet β-cells of adult mouse pancreatic tissue. We demonstrate marked interaction of p27(Kip1) with cyclin D3, an abundant D-type cyclin in adult human islets, and vice versa as well as with its cognate kinase partners, CDK4 and CDK6. Likewise, we show marked interaction of p18(Ink4c) with CDK4. The data collectively suggest that inhibition of CDK function by p27(Kip1) and p18(Ink4c) contributes to human β-cell quiescence. Consistent with this, we have found by BrdU incorporation assay that combined treatments of small molecule GSK-3 inhibitor and mitogen/s lead to elevated proliferation of human β-cells, which is caused partly due to p27(Kip1) downregulation. The results altogether suggest that ex vivo expansion of human β-cells is achievable via increased proliferation for β-cell replacement therapy in diabetes.
authors.affiliation: Section of Endocrinology; Diabetes and Metabolism; Department of Medicine; University of Chicago; Chicago, IL USA.
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Stein
authors.forename: Wieslawa M
authors.initials: WM
authors.lastname: Milewski
authors.forename: Arunangsu
authors.initials: A
authors.lastname: Dey
mesh_terms: D000328:Adult; D000368:Aged; D000818:Animals; D002452:Cell Count; D002453:Cell Cycle / Q000235:genetics*; D049109:Cell Proliferation; D002478:Cells, Cultured; D050763:Cyclin-Dependent Kinase Inhibitor p18 / Q000502:physiology*; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000502:physiology*; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D050416:Glucagon-Secreting Cells / Q000166:cytology / Q000378:metabolism; D038362:Glycogen Synthase Kinase 3 / Q000502:physiology*; D000071679:Glycogen Synthase Kinase 3 beta; D006801:Humans; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000502:physiology*; D051379:Mice; D008875:Middle Aged
chemical_list: D050763:Cyclin-Dependent Kinase Inhibitor p18; D050760:Cyclin-Dependent Kinase Inhibitor p27; D000071679:Glycogen Synthase Kinase 3 beta; D018844:Cyclin-Dependent Kinases; D038362:Glycogen Synthase Kinase 3; C448021:glycogen synthase kinase 3 alpha
keywords: CDK inhibitors; GSK-3; adult human islets; adult pancreatic ?-cell; p18(Ink4c); p27(Kip1); proliferation; quiescence
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