TY - JOUR
T1 - Insulin-Producing cells generated from dedifferentiated human pancreatic beta cells expanded in vitro
AU - Russ, Holger A.
AU - Sintov, Elad
AU - Anker-Kitai, Leeat
AU - Friedman, Orr
AU - Lenz, Ayelet
AU - Toren, Ginat
AU - Farhy, Chen
AU - Pasmanik-Chor, Metsada
AU - Oron-Karni, Varda
AU - Ravassard, Philippe
AU - Efrat, Shimon
N1 - Funding Information:
Human islets were provided through the JDRF award 31-2008-413 (ECIT Islets for Basic Research Program) and the Integrated Islet Distribution Program. We thank Yael Bar, Tel Aviv University, for helpful discussions, Sarah Knoller, Tel Aviv University, for technical assistance, Palle Serup, Hagedorn Research Institute, for NKX6.1 antibody, Julie Kerr-Conte, University Lille-Nord de France, for human pancreas sections, Alberto Hayek, UC San Diego, and John Hutton, University of Colorado Health Sciences Center, for fetal pancreas sections, Ran Elkon, Tel Aviv University, for assistance with statistical analysis, and Yuval Dor, Hebrew University, for comments on the manuscript. This work was performed in partial fulfillment of the requirements for a Ph.D. degree of HAR.
PY - 2011/9/30
Y1 - 2011/9/30
N2 - Background: Expansion of beta cells from the limited number of adult human islet donors is an attractive prospect for increasing cell availability for cell therapy of diabetes. However, attempts at expanding human islet cells in tissue culture result in loss of beta-cell phenotype. Using a lineage-tracing approach we provided evidence for massive proliferation of beta-cell-derived (BCD) cells within these cultures. Expansion involves dedifferentiation resembling epithelial-mesenchymal transition (EMT). Epigenetic analyses indicate that key beta-cell genes maintain open chromatin structure in expanded BCD cells, although they are not transcribed. Here we investigated whether BCD cells can be redifferentiated into beta-like cells. Methodology/Principal Finding: Redifferentiation conditions were screened by following activation of an insulin-DsRed2 reporter gene. Redifferentiated cells were characterized for gene expression, insulin content and secretion assays, and presence of secretory vesicles by electron microscopy. BCD cells were induced to redifferentiate by a combination of soluble factors. The redifferentiated cells expressed beta-cell genes, stored insulin in typical secretory vesicles, and released it in response to glucose. The redifferentiation process involved mesenchymal-epithelial transition, as judged by changes in gene expression. Moreover, inhibition of the EMT effector SLUG (SNAI2) using shRNA resulted in stimulation of redifferentiation. Lineage-traced cells also gave rise at a low rate to cells expressing other islet hormones, suggesting transition of BCD cells through an islet progenitor-like stage during redifferentiation. Conclusions/Significance: These findings demonstrate for the first time that expanded dedifferentiated beta cells can be induced to redifferentiate in culture. The findings suggest that ex-vivo expansion of adult human islet cells is a promising approach for generation of insulin-producing cells for transplantation, as well as basic research, toxicology studies, and drug screening.
AB - Background: Expansion of beta cells from the limited number of adult human islet donors is an attractive prospect for increasing cell availability for cell therapy of diabetes. However, attempts at expanding human islet cells in tissue culture result in loss of beta-cell phenotype. Using a lineage-tracing approach we provided evidence for massive proliferation of beta-cell-derived (BCD) cells within these cultures. Expansion involves dedifferentiation resembling epithelial-mesenchymal transition (EMT). Epigenetic analyses indicate that key beta-cell genes maintain open chromatin structure in expanded BCD cells, although they are not transcribed. Here we investigated whether BCD cells can be redifferentiated into beta-like cells. Methodology/Principal Finding: Redifferentiation conditions were screened by following activation of an insulin-DsRed2 reporter gene. Redifferentiated cells were characterized for gene expression, insulin content and secretion assays, and presence of secretory vesicles by electron microscopy. BCD cells were induced to redifferentiate by a combination of soluble factors. The redifferentiated cells expressed beta-cell genes, stored insulin in typical secretory vesicles, and released it in response to glucose. The redifferentiation process involved mesenchymal-epithelial transition, as judged by changes in gene expression. Moreover, inhibition of the EMT effector SLUG (SNAI2) using shRNA resulted in stimulation of redifferentiation. Lineage-traced cells also gave rise at a low rate to cells expressing other islet hormones, suggesting transition of BCD cells through an islet progenitor-like stage during redifferentiation. Conclusions/Significance: These findings demonstrate for the first time that expanded dedifferentiated beta cells can be induced to redifferentiate in culture. The findings suggest that ex-vivo expansion of adult human islet cells is a promising approach for generation of insulin-producing cells for transplantation, as well as basic research, toxicology studies, and drug screening.
UR - http://www.scopus.com/inward/record.url?scp=80053608936&partnerID=8YFLogxK
U2 - 10.1371/journal.pone.0025566
DO - 10.1371/journal.pone.0025566
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AN - SCOPUS:80053608936
SN - 1932-6203
VL - 6
JO - PLoS ONE
JF - PLoS ONE
IS - 9
M1 - e25566
ER -
