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This Article Accepted manuscript (PDF) Supplementary data All Versions of this Article: JOE-08-0482v1 JOE-08-0482v2 201/1/37    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Nagaya, M. Articles by Weir, G. Search for Related Content PubMed PubMed Citation Articles by Nagaya, M. Articles by Weir, G.

M Nagaya, Islet Transplantation and Cell Biology , Joslin Diabetes Center , Boston, 02215 , United States
H Katsuta, Islet Transplantation and Cell Biology, Joslin Diabetes Center, Boston, United States
H Kaneto, Department of Internal Medicine and Therapeutics, Osaka University Graduate School of Medicine, Osaka, Japan
S Bonner-Weir, Islet Transplantation and Cell Biology, Joslin Diabetes Center, Boston, United States
G Weir, Boston, United States

Correspondence: Masaki Nagaya, Email: Masaki.Nagaya{at}joslin.harvard.edu

Abstract

Transdifferentiation of cells from a patient's own liver into pancreatic β-cells could be useful for β-cell replacement. We hypothesized that intrahepatic biliary epithelial cells (IHBECs) could become a new source of insulin-producing cells.

IHBECs isolated from adult mice were expanded using our novel culture method, termed collagen-embedded floating culture method (CEFCM). With CEFCM, IHBECs formed three-dimensional ductal cysts and rapidly expanded their number about 15-fold within 2 wk. Over 90% of cells were positive for cytokeratin 7 and 19. At day14, IHBECs were transfected with adenoviral (Ad)- pancreas duodenum homeobox 1 (Pdx-1), NeuroD, or Pdx-1/VP16. After 7 additional days in serum- and insulin-free differentiation medium (DM), cell phenotypes were determined by RT-PCR, immunostaining and ELISA for insulin.

In DM Control IHBECs started to express some endocrine progenitor genes (Neurogenin3, NeuroD, Nkx6.1 and Pdx-1) but lacked Insulin mRNA. Transduced expression of Pdx-1, NeuroD or Pdx-1/VP16 led to expression of not only Insulin but also GLUT2 and Prohormone convertase 1 and 2. About 3% of 4000 cells counted in Pdx-1/VP16 transduced cultures stained strongly for C-peptide suggesting that a subpopulation may have the capacity for differentiation. Transduced cells released insulin (Ad-Pdx-1 0.08±0.05, Ad-NeuroD 0.33±0.09, Ad-Pdx-1/VP16 0.37±0.14 ng/1X105 cells after 48h in culture).

IHBECs can be markedly expanded, and then with molecular manipulation a subpopulation of these cells can differentiate towards a β-cell phenotype. This approach may lead to a new source of β-cells that can be used for transplantation in diabetes.

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