title: Analysis of insulin-producing cells during in vitro differentiation from feeder-free embryonic stem cells.
abstract: Embryonic stem (ES) cells can differentiate into many cell types and are expected to be useful for tissue engineering. Recent reports have shown that ES cells can differentiate into insulin-producing cells in response to the transient expression of the pdx-1 gene, after the removal of feeder cells. To investigate the lineage of insulin-producing cells and their in vitro differentiation, we introduced the betageo gene, encoding a beta-galactosidase-neomycin phosphotransferase fusion protein under the control of the mouse insulin 2 promoter, into ES cells that had been adapted to feeder-free culture, and analyzed insulin gene expression during their in vitro differentiation. We also examined the expression of transcription factors that are related to the differentiation of the pancreas. X-gal staining analysis revealed beta-galactosidase-positive cells on the surface and in the center of the embryoid body that proliferated during differentiation. Glucose-responsive insulin-producing cells, derived from our feeder-free ES cells, expressed insulin 2, pdx-1, Pax4, and Isl1 and also the glucagon, somatostatin, and PP genes. Moreover, the genes encoding p48, amylase, and carboxypeptidase A were also expressed. These results suggest that ES cells can differentiate not only into endocrine cells but also into exocrine cells of the pancreas, without the initiation of pdx-1 expression.
authors.affiliation: Division of Stem Cell Regulation Research, Osaka University Graduate School of Medicine, 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan.
authors.forename: Yusuke
authors.initials: Y
authors.lastname: Moritoh
authors.forename: Eiji
authors.initials: E
authors.lastname: Yamato
authors.forename: Yumiko
authors.initials: Y
authors.lastname: Yasui
authors.forename: Satsuki
authors.initials: S
authors.lastname: Miyazaki
authors.forename: Jun-ichi
authors.initials: J
authors.lastname: Miyazaki
mesh_terms: D000818:Animals; D001483:Base Sequence; D002454:Cell Differentiation / Q000502:physiology*; D017931:DNA Primers; D018507:Gene Expression Regulation, Developmental; D005947:Glucose / Q000494:pharmacology; D018398:Homeodomain Proteins; D007328:Insulin / Q000096:biosynthesis / Q000235:genetics*; D019868:Kanamycin Kinase / Q000235:genetics; D051379:Mice; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D015534:Trans-Activators / Q000235:genetics; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D001617:beta-Glucosidase / Q000235:genetics
chemical_list: D017931:DNA Primers; D018398:Homeodomain Proteins; D007328:Insulin; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; D014157:Transcription Factors; C084010:pancreatic and duodenal homeobox 1 protein; D019868:Kanamycin Kinase; D001617:beta-Glucosidase; D005947:Glucose