Long-term correction of diabetes in mice by in vivo reprogramming of pancreatic ducts
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE109113 | GEO | 2018/05/01
REPOSITORIES: GEO
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Project description:The adult pancreas is capable of limited regeneration after injury, but has no defined stem cell population. The cell types and molecular signals that govern the production of new pancreatic tissue are not well understood. Here we show that inactivation of the SCF-type E3 ubiquitin ligase substrate recognition component Fbw7 induces pancreatic ductal cells to reprogram into β-cells. The induced β-cells resemble islet β-cells in morphology and histology, express genes essential for β-cell function, and release insulin upon glucose challenge. Thus, loss of Fbw7 appears to reawaken an endocrine developmental differentiation program in adult pancreatic ductal cells. Our study highlights the plasticity of seemingly differentiated adult cells, identifies Fbw7 as a master regulator of cell fate decisions in the pancreas, and reveals adult pancreatic duct cells as a latent multipotent cell type. We used microarray to compare adult mouse fbw7 knock out ductal cells with bonafide beta cells In order to isolate fbw7 ko, fbw7 wt ductal cells and beta cells we used adult mice with the following genotypes: MIP-GFP (beta cells are constitutively labelled with GFP), Ck19-CreERT; R26-LSL-YFP; Fbw7 +/+ (ductal cells are labelled with YFP upon tamoxifen injection) and Ck19-CreERT; R26-LSL-YFP; Fbw7 F/F mice (ductal cells are labelled with YFP upon tamoxifen injection). 30000 GFP+ cells were isolated by FACS-sorting. RNA was extracted and amplified using NuGEN kit.
2014-08-07 | E-GEOD-58969 | biostudies-arrayexpress
Project description:This experiment used RNA-Seq technology to explore gene expression in mouse FACS-purified pancreatic P60 Acinar cells, P60 Beta-cells, 1 day 3TF (MafA/Neurog3/Pdx1)-induced Acinar cells and 7 days 3TF-induced Acinar cells.ďż˝
2016-07-06 | E-MTAB-3921 | biostudies-arrayexpress
Project description:The adult pancreas is capable of limited regeneration after injury, but has no defined stem cell population. The cell types and molecular signals that govern the production of new pancreatic tissue are not well understood. Here we show that inactivation of the SCF-type E3 ubiquitin ligase substrate recognition component Fbw7 induces pancreatic ductal cells to reprogram into β-cells. The induced β-cells resemble islet β-cells in morphology and histology, express genes essential for β-cell function, and release insulin upon glucose challenge. Thus, loss of Fbw7 appears to reawaken an endocrine developmental differentiation program in adult pancreatic ductal cells. Our study highlights the plasticity of seemingly differentiated adult cells, identifies Fbw7 as a master regulator of cell fate decisions in the pancreas, and reveals adult pancreatic duct cells as a latent multipotent cell type. We used microarray to compare adult mouse fbw7 knock out ductal cells with bonafide beta cells
2014-08-07 | GSE58969 | GEO
Project description:Direct lineage conversion of adult cells is a promising approach for regenerative medicine. A major challenge of lineage conversion is to generate specific subtypes of cells, closely related cells with distinct properties. The pancreatic islets contain three major hormone-secreting endocrine subtypes: insulin+ β-cells, glucagon+ α-cells, and somatostatin+ δ-cells. We previously reported that a combination of three transcription factors, Ngn3, Mafa, and Pdx1, directly reprogram pancreatic acinar cells to β-cells. We now show that acinar cells can be converted to δ-like and α-like cells by Ngn3 and Ngn3+Mafa respectively. Thus, three major islet endocrine subtypes can be derived by acinar reprogramming. Ngn3 promotes establishment of a generic endocrine state in acinar cells at the onset of reprogramming in addition to promoting δ-specification. Mafa and Pdx1 suppress δ-specification in α- and β-cell formation. These studies identify a set of defined factors whose combinatorial actions reprogram acinar cells to distinct islet endocrine subtypes in vivo. induced beta cells samples at day 10 collected for the microarray
2013-11-20 | E-GEOD-52522 | biostudies-arrayexpress
Project description:During pancreatic development, Neurogenin3 (Ngn3) promotes the differentiation of endocrine cells, including insulin-producing β-cells. Our previous work has shown that Ngn3 phosphorylation on multiple serine-proline (SP) sites increases protein stability and endocrine differentiation (Azzarelli et al., DevCell 2017). Here, we aim to exploit our recent data to investigate whether manipulation of Ngn3 phosphostatus might improve in vitro generation of β-cells for replacement therapies in diabetes. A potential source of β-cells comes from fate conversion of exocrine pancreatic cells into the endocrine lineage, by overexpression of 3 transcription factors, Ngn3, Pdx1 and MafA. Using this approach we show that pancreatic ductal cells grown in vitro as 3D organoids can be reprogrammed to express insulin. The efficiency can be strongly enhanced by substituting wild type Ngn3 with a the un(der)phosphorylated and more active Ngn3 form (6S-A Ngn3). Here we perform transcriptomic analysis of a low number of pancreatic organoid cells expressing wild type or phosphomutant Ngn3, alone or in various combination Pdx1 and/or MafA. The analysis revealed endocrine differentiation and in particular β-like cell gene expression in the conditions where Ngn3 was in combination with both Pdx1 and MafA.
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Project description:Direct lineage conversion of adult cells is a promising approach for regenerative medicine. A major challenge of lineage conversion is to generate specific subtypes of cells, closely related cells with distinct properties. The pancreatic islets contain three major hormone-secreting endocrine subtypes: insulin+ β-cells, glucagon+ α-cells, and somatostatin+ δ-cells. We previously reported that a combination of three transcription factors, Ngn3, Mafa, and Pdx1, directly reprogram pancreatic acinar cells to β-cells. We now show that acinar cells can be converted to δ-like and α-like cells by Ngn3 and Ngn3+Mafa respectively. Thus, three major islet endocrine subtypes can be derived by acinar reprogramming. Ngn3 promotes establishment of a generic endocrine state in acinar cells at the onset of reprogramming in addition to promoting δ-specification. Mafa and Pdx1 suppress δ-specification in α- and β-cell formation. These studies identify a set of defined factors whose combinatorial actions reprogram acinar cells to distinct islet endocrine subtypes in vivo.
2013-11-20 | GSE52522 | GEO
Project description:Bromodomain and Extra-terminal (BET) proteins are epigenetic readers that interact with acetylated lysines of histone tails. Recent studies have demonstrated their role in cancer progression, as they recruit key components of the transcriptional machinery to modulate gene expression. However, their role during embryonic development of the pancreas has never been studied. Using mouse embryonic pancreatic explants and human IPSC, we show that inhibition of BET proteins with either I-BET151 or JQ1 dramatically enhances the number of neurogenin3 (NEUROG3) endocrine progenitors, leading to an increased number of endocrine cells in the pancreatic explants. Despite inducing several β cell markers, BETi also strongly down-regulated Ins1 expression in developed explants and adult β cells. However, removal of BETi from explants prior to β cell development, as NEUROG3 expression peaks, further led to enhanced β cell development with higher expression of insulin and maturation markers UCN3 and MAFA. Altogether, these results show that BET proteins play a major role in the pancreas endocrine differentiation. Furthermore, they highlight the potential use of BETi to improve β cell replacement therapies.
2020-01-01 | GSE109858 | GEO
Project description:MafA and MafB transcription factors have been shown to be key regulators of insulin and glucagon transcription. MafB is essential for alpha and beta cell differentiation, as MafB deficient mice produced fewer insulin+ and glucagon+ cells during development, with MafA expressed in remaining insulin+ cells. In contrast, beta cell development was reported to be normal in a total MafA knock out, although the animals developed beta cell dysfunction and diabetes as adults. However, we have found that MafB expression is elevated during development and retained in adult insulin+ cells after conditional removal of MafA in the pancreas. These studies will evaluate the broader significance of these insulin and glucagon regulators in alpha and beta cell development and function. Our efforts will focus on determining if the concerted actions of MafA and MafB factors are significant to beta cell formation, and we specifically plan to: Determine how alpha and beta cell differentiation is affected in MafA/MafB compound mutant mice during pancreas development. cDNA microarray studies (pancchip 6.0) with wild type, MafAKO, MafB-/-, and MafAKOMafB-/- mutant E18.5 pancreata will be performed to comprehensively identify genes controlled by MafA and MafB in developing alpha and beta cells.
2014-11-04 | E-MTAB-3070 | biostudies-arrayexpress
Project description:Three master regulatory transcription factors Pdx1, MafA and Ngn3 have the ability to transdifferentiate pancreatic acinar cells to insulin-producing beta cells in mice. BRD7552 was identified as a small-molecule inducer that can upregulate the expression of Pdx1 in PANC-1 cells by high-throughput qPCR screening. We used microarrays to illustrate the mechnism of BRD7552 for PDX1 and insulin induction.
2013-03-10 | GSE44946 | GEO
Project description:Neurog3 and NeuroD1 share the ability to induce endocrine differentiation when expressed ectopically in vivo (Apelqvist et al., 1999; Schwitzgebel et al., 2000; Grapin-Botton et al., 2001) and in vitro (Heremans et al., 2002; Gasa et al., 2004), suggesting that they target a common set of genes. To test this hypothesis, we directly compared in the same cells the effects of Neurog3 and NeuroD1 on global gene expression patterns. Normalized M and A values [limmaGUI output] for the neurogenin3 (Ngn3) and neuroD1 (Nd1) experiments (individual or combined) have been linked below as Supplementary files. Note that M=log2(test/ref), eg, log2(Nd1/Bgal); A=[log2(test*ref)]/2, eg, [log2(Nd1*Bgal)]/2. Additional measures of reliability/probability are also included within the supplementary files. Mouse pancreatic ductal cells (mPAC cells) were infected with equal titers of recombinant adenoviruses encoding human Neurogenin3 (AdCMV-NEUROG3) or mouse NeuroD1 (AdCMV-NeuroD1), and 48 hrs later their transcriptomes were compared with that of control cells infected with an adenovirus encoding betagalactosidase (beta-gal; AdCMV-Bgal) using cDNA microarrays. Differences in the activation patterns of the two transcription factors were then determined by comparing the fold-change (FC) values of the pair-wise hybridizations Neurog3/Bgal and NeuroD1/beta-gal.
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