title: FGF7 and cell density are required for final differentiation of pancreatic amylase-positive cells from human ES cells.
abstract: The major molecular signals of pancreatic exocrine development are largely unknown. We examine the role of fibroblast growth factor 7 (FGF7) in the final induction of pancreatic amylase-containing exocrine cells from induced-pancreatic progenitor cells derived from human embryonic stem (hES) cells. Our protocol consisted in three steps: Step I, differentiation of definitive endoderm (DE) by activin A treatment of hES cell colonies; Step II, differentiation of pancreatic progenitor cells by re-plating of the cells of Step I onto 24-well plates at high density and stimulation with all-trans retinoic acid; Step III, differentiation of pancreatic exocrine cells with a combination of FGF7, glucagon-like peptide 1 and nicotinamide. The expression levels of pancreatic endodermal markers such as Foxa2, Sox17 and gut tube endoderm marker HNF1β were up-regulated in both Step I and II. Moreover, in Step III, the induced cells expressed pancreatic markers such as amylase, carboxypeptidase A and chymotrypsinogen B, which were similar to those in normal human pancreas. From day 8 in Step III, cells immunohistochemically positive for amylase and for carboxypeptidase A, a pancreatic exocrine cell product, were induced by FGF7. Pancreatic progenitor Pdx1-positive cells were localized in proximity to the amylase-positive cells. In the absence of FGF7, few amylase-positive cells were identified. Thus, our three-step culture protocol for human ES cells effectively induces the differentiation of amylase- and carboxypeptidase-A-containing pancreatic exocrine cells.
authors.affiliation: Laboratory for Advanced Health Sciences, Bourbon Institutes of Health, BOURBON Corporation, 4-2-14 Matsunami, Kashiwazaki, Niigata, 945-8611, Japan.
authors.forename: Sakiko
authors.initials: S
authors.lastname: Takizawa-Shirasawa
authors.forename: Susumu
authors.initials: S
authors.lastname: Yoshie
authors.forename: Fengming
authors.initials: F
authors.lastname: Yue
authors.forename: Akimi
authors.initials: A
authors.lastname: Mogi
authors.forename: Tadayuki
authors.initials: T
authors.lastname: Yokoyama
authors.forename: Daihachiro
authors.initials: D
authors.lastname: Tomotsune
authors.forename: Katsunori
authors.initials: K
authors.lastname: Sasaki
mesh_terms: D028341:Activins / Q000494:pharmacology; D000681:Amylases / Q000096:biosynthesis*; D002450:Cell Communication / Q000187:drug effects / Q000502:physiology; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D003584:Cytological Techniques; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D051523:Fibroblast Growth Factor 7 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D052216:Glucagon-Like Peptide 1 / Q000494:pharmacology; D006801:Humans; D009536:Niacinamide / Q000494:pharmacology; D046790:Pancreas, Exocrine / Q000166:cytology* / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D014212:Tretinoin / Q000494:pharmacology
chemical_list: C496453:FGF7 protein, human; D028341:Activins; D051523:Fibroblast Growth Factor 7; D009536:Niacinamide; D014212:Tretinoin; D052216:Glucagon-Like Peptide 1; D000681:Amylases