title: PDX1- and NGN3-mediated in vitro reprogramming of human bone marrow-derived mesenchymal stromal cells into pancreatic endocrine lineages.
abstract: OBJECTIVE
  Reprogramming of multipotent adult bone marrow (BM)-derived mesenchymal stromal/stem cells (MSC) (BM-MSC) represents one of several strategies for cell-based therapy of diabetes. However, reprogramming primary BM-MSC into pancreatic endocrine lineages has not yet been consistently demonstrated.
  
  
  METHODS
  To unravel the role and interaction of key factors governing this process, we used well-characterized telomerase-immortalized human MSC (hMSC-TERT). Pancreatic endocrine differentiation in hMSC-TERT was induced by two major in vitro strategies: (i) endocrine-promoting culture conditions and (ii) ectopic expression of two master regulatory genes of the endocrine lineage, human neurogenin 3 (NGN3) and human pancreatic duodenal homeobox 1 (PDX1).
  
  
  RESULTS
  Both approaches triggered pancreatic endocrine gene expression, notably insulin, glucose-transporter 2 and somatostatin. Transgenic overexpression of NGN3 and/or PDX1 proteins not only induced direct target genes, such as NEUROD1 and insulin, and but also triggered parts of the gene expression cascade that is involved in pancreatic endocrine differentiation. Notably, ectopic NGN3 alone was sufficient to initiate the expression of specific beta-cell lineage-associated genes, most importantly PDX1 and insulin. This was demonstrated both transcriptionally by mRNA expression and reporter gene analyzes and at a protein level by Western blotting. Such reprogramming of hMSC-TERT cells induced glucose-insensitive insulin biosynthesis and secretion.
  
  
  CONCLUSIONS
  Our results indicate that establishment of glucose-dependent insulin secretion in partially reprogrammed human MSC may depend on additional maturation factors. Moreover, hMSC-TERT provides a suitable cell model for investigating further the molecular mechanisms of reprogramming and maturation of adult MSC towards pancreatic endocrine lineages.
authors.affiliation: Division of Endocrinology and Diabetology, Department of Internal Medicine II, University Hospital of Freiburg , Freiburg, Germany.
authors.forename: Catarina
authors.initials: C
authors.lastname: Limbert
authors.forename: Günter
authors.initials: G
authors.lastname: Päth
authors.forename: Regina
authors.initials: R
authors.lastname: Ebert
authors.forename: Veit
authors.initials: V
authors.lastname: Rothhammer
authors.forename: Moustapha
authors.initials: M
authors.lastname: Kassem
authors.forename: Franz
authors.initials: F
authors.lastname: Jakob
authors.forename: Jochen
authors.initials: J
authors.lastname: Seufert
mesh_terms: D000328:Adult; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics* / Q000378:metabolism; D015415:Biomarkers; D001854:Bone Marrow Cells / Q000166:cytology*; D002096:C-Peptide / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D002461:Cell Line, Transformed; D002478:Cells, Cultured; D003470:Culture Media; D005786:Gene Expression Regulation; D051273:Glucose Transporter Type 2 / Q000235:genetics; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000235:genetics / Q000378:metabolism; D000078790:Insulin Secretion; D007515:Islets of Langerhans / Q000166:cytology*; D008297:Male; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000276:immunology / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D013004:Somatostatin / Q000235:genetics; D019098:Telomerase / Q000235:genetics / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics* / Q000378:metabolism
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D015415:Biomarkers; D002096:C-Peptide; D003470:Culture Media; D051273:Glucose Transporter Type 2; D018398:Homeodomain Proteins; D007328:Insulin; D009419:Nerve Tissue Proteins; D013004:Somatostatin; D019098:Telomerase; D015534:Trans-Activators; C093970:NEUROD1 protein, human; C498695:NEUROG3 protein, human; C495680:SLC2A2 protein, human; C084010:pancreatic and duodenal homeobox 1 protein; C509186:TERT protein, human