Characterization of endocrine progenitor cells and critical factors for their differentiation in human adult pancreatic cell culture.

We have reproduced a previously described method for the in vitro generation of endocrine cells in adult human pancreatic tissue culture. The aim of this study was to characterize the nature of pancreatic progenitor cells and to identify the factors necessary for their differentiation in this model. During monolayer expansion, two types of cells proliferated sequentially; first cytokeratin 19 (CK19)-positive ductal epithelial cells and then nestin-positive fibroblastoid cells. After the bromodeoxyuridine-labeled cells were traced in differentiated islet buds, some of the proliferating ductal cells had differentiated into endocrine cells, whereas nestin-positive cells could not give rise to endocrine tissue. Serum-free culture was found to be an absolute requirement for the endocrine differentiation to occur. Also, overlay of the cells with Matrigel was essential, whereas nicotinamide had a potentiating effect. The in vitro-generated islet buds released insulin in response to glucose nearly as efficiently as native islets. When transplanted under the kidney capsule of nude mice, only one of five grafts demonstrated further growth with foci of both endocrine and exocrine differentiation. Our results support the previous notion that pancreatic progenitor cells represent a subpopulation of ductal epithelial cells. No evidence was found for the development of endocrine cells from nestin-positive stem cells.

[1]  B. Spencer‐Dene,et al.  The IIIb isoform of fibroblast growth factor receptor 2 is required for proper growth and branching of pancreatic ductal epithelium but not for differentiation of exocrine or endocrine cells , 2003, Mechanisms of Development.

[2]  Jayaraj Rajagopal,et al.  Insulin Staining of ES Cell Progeny from Insulin Uptake , 2003, Science.

[3]  A. Shapiro,et al.  Successful islet transplantation: continued insulin reserve provides long-term glycemic control. , 2002, Diabetes.

[4]  K. Ekdahl,et al.  Inhibition of thrombin abrogates the instant blood-mediated inflammatory reaction triggered by isolated human islets: possible application of the thrombin inhibitor melagatran in clinical islet transplantation. , 2002, Diabetes.

[5]  L. Turnpenny,et al.  Beta-cell differentiation during human development does not rely on nestin-positive precursors: implications for stem cell-derived replacement therapy , 2002, Diabetologia.

[6]  Lars Selander,et al.  Nestin is expressed in mesenchymal and not epithelial cells of the developing mouse pancreas , 2002, Mechanisms of Development.

[7]  Shulamit Levenberg,et al.  Endothelial cells derived from human embryonic stem cells , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[8]  R. Scharfmann,et al.  Role for FGFR2IIIb-mediated signals in controlling pancreatic endocrine progenitor cell proliferation , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[9]  E. Montanya,et al.  Stem cells and diabetes. , 2001, Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie.

[10]  R. McKay,et al.  Differentiation of Embryonic Stem Cells to Insulin-Secreting Structures Similar to Pancreatic Islets , 2001, Science.

[11]  J. Habener,et al.  Multipotential nestin-positive stem cells isolated from adult pancreatic islets differentiate ex vivo into pancreatic endocrine, exocrine, and hepatic phenotypes. , 2001, Diabetes.

[12]  B. Tyrberg,et al.  Stimulated endocrine cell proliferation and differentiation in transplanted human pancreatic islets: effects of the ob gene and compensatory growth of the implantation organ. , 2001, Diabetes.

[13]  O. Madsen,et al.  Islet and stem cell transplantation for treating diabetes , 2001, BMJ : British Medical Journal.

[14]  F. Pattou,et al.  Adult human cytokeratin 19-positive cells reexpress insulin promoter factor 1 in vitro: further evidence for pluripotent pancreatic stem cells in humans. , 2000, Diabetes.

[15]  S. Bonner-Weir,et al.  In vitro cultivation of human islets from expanded ductal tissue. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[16]  I. Rooman,et al.  Modulation of rat pancreatic acinoductal transdifferentiation and expression of PDX-1 in vitro , 2000, Diabetologia.

[17]  M. Longaker,et al.  Expression and role of laminin-1 in mouse pancreatic organogenesis. , 2000, Diabetes.

[18]  E. Hunziker,et al.  Nestin-expressing cells in the pancreatic islets of Langerhans. , 2000, Biochemical and biophysical research communications.

[19]  L. Bouwens,et al.  Adult human pancreatic duct and islet cells exhibit similarities in expression and differences in phosphorylation and complex formation of the homeodomain protein Ipf-1. , 2000, Diabetes.

[20]  D. Schatz,et al.  Reversal of insulin-dependent diabetes using islets generated in vitro from pancreatic stem cells , 2000, Nature Medicine.

[21]  O. Korsgren,et al.  Differentiation and maturation of porcine fetal islet cells in vitro and after transplantation. , 1999, Transplantation.

[22]  Ana D. Lopez,et al.  Sustained proliferation of PDX-1+ cells derived from human islets. , 1999, Diabetes.

[23]  D. Parry,et al.  A High Molecular Weight Intermediate Filament-associated Protein in BHK-21 Cells Is Nestin, a Type VI Intermediate Filament Protein , 1999, The Journal of Biological Chemistry.

[24]  L. Harrison,et al.  Laminin-1 promotes differentiation of fetal mouse pancreatic beta-cells. , 1999, Diabetes.

[25]  S. Bonner-Weir,et al.  The homeodomain protein IDX-1 increases after an early burst of proliferation during pancreatic regeneration. , 1999, Diabetes.

[26]  K. Docherty,et al.  Glucose Stimulates Translocation of the Homeodomain Transcription Factor PDX1 from the Cytoplasm to the Nucleus in Pancreatic β-Cells* , 1999, The Journal of Biological Chemistry.

[27]  D. Parry,et al.  A high molecular weight intermediate filament-associated protein in BHK-21 cells is nestin, a type VI intermediate filament protein. Limited co-assembly in vitro to form heteropolymers with type III vimentin and type IV alpha-internexin. , 1999, The Journal of biological chemistry.

[28]  L. Bouwens,et al.  Extra-insular beta cells associated with ductules are frequent in adult human pancreas , 1998, Diabetologia.

[29]  L. Bouwens.  Cytokeratins and cell differentiation in the pancreas , 1998, The Journal of pathology.

[30]  L. Bouwens,et al.  Culture of Adult Human Islet Preparations With Hepatocyte Growth Factor and 804G Matrix Is Mitogenic for Duct Cells but Not for β-Cells , 1998, Diabetes.

[31]  V. Tchernev,et al.  In Vitro-Generation of Islets in Long-Term Cultures of Pluripotent Stem Cells From Adult Mouse Pancreas , 1997, Hormone and metabolic research = Hormon- und Stoffwechselforschung = Hormones et metabolisme.

[32]  L. Bouwens,et al.  Proliferation and differentiation in the human fetal endocrine pancreas , 1997, Diabetologia.

[33]  Ana D. Lopez,et al.  Functional β-Cell Mass After Transplantation of Human Fetal Pancreatic Cells: Differentiation or Proliferation? , 1997, Diabetes.

[34]  J. Rubin,et al.  Regulation of Proliferation and Differentiation of Human Fetal Pancreatic Islet Cells by Extracellular Matrix, Hepatocyte Growth Factor, and Cell-Cell Contact , 1996, Diabetes.

[35]  L. Baxter,et al.  A Second Pathway for Regeneration of Adult Exocrine and Endocrine Pancreas: A Possible Recapitulation of Embryonic Development , 1993, Diabetes.

[36]  C. Ricordi,et al.  Nicotinamide is a potent inducer of endocrine differentiation in cultured human fetal pancreatic cells. , 1993, The Journal of clinical investigation.

[37]  J. C. Jones,et al.  The matrix secreted by 804G cells contains laminin-related components that participate in hemidesmosome assembly in vitro. , 1993, Journal of cell science.

[38]  N. Sarvetnick,et al.  Epithelial cell proliferation and islet neogenesis in IFN-g transgenic mice. , 1993, Development.

[39]  J. Rubin,et al.  Characterization of the receptor for keratinocyte growth factor. Evidence for multiple fibroblast growth factor receptors. , 1990, The Journal of biological chemistry.

[40]  Camillo Ricordi,et al.  Automated Method for Isolation of Human Pancreatic Islets , 1988, Diabetes.

[41]  R. Alejandro,et al.  A simple method of staining fresh and cultured islets. , 1988, Transplantation.

[42]  J. Koudstaal,et al.  Nesidioblastosis and endocrine hyperplasia of the pancreas: a secondary phenomenon. , 1986, Human pathology.

[43]  H. Okamoto,et al.  Amelioration of Diabetes Mellitus in Partially Depancreatized Rats by Poly(ADP-ribose) Synthetase Inhibitors: Evidence of Islet B-Cell Regeneration , 1984, Diabetes.

[44]  W. Rutter,et al.  5-bromodeoxyuridine may alter the differentiative program of the embryonic pancreas , 1976, The Journal of cell biology.

[45]  R. Hinegardner,et al.  An improved fluorometric assay for DNA. , 1971, Analytical biochemistry.