title: Adult Human Biliary Tree Stem Cells Differentiate to β-Pancreatic Islet Cells by Treatment with a Recombinant Human Pdx1 Peptide.
abstract: Generation of β-pancreatic cells represents a major goal in research. The aim of this study was to explore a protein-based strategy to induce differentiation of human biliary tree stem cells (hBTSCs) towards β-pancreatic cells. A plasmid containing the sequence of the human pancreatic and duodenal homeobox 1 (PDX1) has been expressed in E. coli. Epithelial-Cell-Adhesion-Molecule positive hBTSCs or mature human hepatocyte cell line, HepG2, were grown in medium to which Pdx1 peptide was added. Differentiation toward pancreatic islet cells were evaluated by the expression of the β-cell transcription factors, Pdx1 and musculoapo-neurotic fibrosarcoma oncogene homolog A, and of the pancreatic hormones, insulin, glucagon, and somatostatin, investigated by real time polymerase chain reaction, western blot, light microscopy and immunofluorescence. C-peptide secretion in response to high glucose was also measured. Results indicated how purified Pdx1 protein corresponding to the primary structure of the human Pdx1 by mass spectroscopy was efficiently produced in bacteria, and transduced into hBTSCs. Pdx1 exposure triggered the expression of both intermediate and mature stage β-cell differentiation markers only in hBTSCs but not in HepG2 cell line. Furthermore, hBTSCs exposed to Pdx1 showed up-regulation of insulin, glucagon and somatostatin genes and formation of 3-dimensional islet-like structures intensely positive for insulin and glucagon. Finally, Pdx1-induced islet-like structures exhibited glucose-regulated C-peptide secretion. In conclusion, the human Pdx1 is highly effective in triggering hBTSC differentiation toward functional β-pancreatic cells.
authors.affiliation: Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy.
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Cardinale
authors.affiliation: Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy.
authors.forename: Rosa
authors.initials: R
authors.lastname: Puca
authors.affiliation: Department of Health Sciences, University of Rome "Foro Italico", Rome, Italy.
authors.forename: Guido
authors.initials: G
authors.lastname: Carpino
authors.affiliation: Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy.
authors.forename: Gaia
authors.initials: G
authors.lastname: Scafetta
authors.affiliation: Department of Anatomical, Histological, Forensic Medicine and Orthopedics Sciences, Sapienza University of Rome, Rome, Italy.
authors.forename: Anastasia
authors.initials: A
authors.lastname: Renzi
authors.affiliation: Departments of Science and Chemical Technologies, University of Tor Vergata, Rome, Italy.
authors.forename: Michele
authors.initials: M
authors.lastname: De Canio
authors.affiliation: Departments of Science and Chemical Technologies, University of Tor Vergata, Rome, Italy.
authors.forename: Francesca
authors.initials: F
authors.lastname: Sicilia
authors.affiliation: Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy.
authors.forename: Lorenzo
authors.initials: L
authors.lastname: Nevi
authors.affiliation: Italian Federation of Juvenile Diabetes (FDG), Rome, Italy.
authors.forename: Domenico
authors.initials: D
authors.lastname: Casa
authors.affiliation: Italian Federation of Juvenile Diabetes (FDG), Rome, Italy.
authors.forename: Rocco
authors.initials: R
authors.lastname: Panetta
authors.affiliation: Department of General Surgery and Organ Transplantation, Sapienza University of Rome, Rome, Italy.
authors.forename: Pasquale Bartolomeo
authors.initials: PB
authors.lastname: Berloco
authors.affiliation: Departments of Cell and Molecular Physiology, Program in Molecular Biology and Biotechnology, University of North Carolina School of Medicine, Chapel Hill, North Carolina, United States of America.
authors.forename: Lola M
authors.initials: LM
authors.lastname: Reid
authors.affiliation: Departments of Science and Chemical Technologies, University of Tor Vergata, Rome, Italy.
authors.forename: Giorgio
authors.initials: G
authors.lastname: Federici
authors.affiliation: Department of Anatomical, Histological, Forensic Medicine and Orthopedics Sciences, Sapienza University of Rome, Rome, Italy.
authors.forename: Eugenio
authors.initials: E
authors.lastname: Gaudio
authors.affiliation: Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy.
authors.forename: Marella
authors.initials: M
authors.lastname: Maroder
authors.affiliation: Department of Medico-Surgical Sciences and Biotechnologies, Sapienza University of Rome, Rome, Italy; Eleonora Lorillard Spencer-Cenci Foundation, Rome, Italy.
authors.forename: Domenico
authors.initials: D
authors.lastname: Alvaro
mesh_terms: D053687:Adult Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D001659:Biliary Tract / Q000166:cytology*; D002454:Cell Differentiation / Q000187:drug effects*; D048430:Cell Shape / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002845:Chromatography; D004705:Endocytosis / Q000187:drug effects; D056945:Hep G2 Cells; D018398:Homeodomain Proteins / Q000494:pharmacology*; D006801:Humans; D050417:Insulin-Secreting Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology*; D015534:Trans-Activators / Q000494:pharmacology*
chemical_list: D018398:Homeodomain Proteins; D011994:Recombinant Proteins; D015534:Trans-Activators; C084010:pancreatic and duodenal homeobox 1 protein