Pancreatic differentiation of stem cells reveals pathogenesis of a syndrome of Ketosis-Prone Diabetes

Artificially generated pancreatic β-cells from pluripotent stem cells are expected for cell replacement therapy for type 1 diabetes. Several strategies are adopted to direct pluripotent stem cells toward pancreatic differentiation. However, a standard differentiation method for clinical application has not been established. It is important to develop more effective and safer methods for generating pancreatic β-cells without toxic or mutagenic chemicals. In the present study, we screened several endogenous factors involved in organ development to identify the factor, which induced the efficiency of pancreatic differentiation and found that treatment with erythropoietin (EPO) facilitated the differentiation of mouse embryonic stem cells (ESCs) into definitive endoderm. At an early stage of differentiation, EPO treatment significantly increased Sox17 gene expression, as a marker of the definitive endoderm. Contrary to the canonical function of EPO, it did not affect the levels of phosphorylated JAK2 and STAT5, but stimulated the phosphorylation of ERK1/2 and Akt. The MEK inhibitor U0126 significantly inhibited EPO-induced Sox17 expression. The differentiation of ESCs into definitive endoderm is an important step for the differentiation into pancreatic and other endodermal lineages. This study suggests a possible role of EPO in embryonic endodermal development and a new agent for directing the differentiation into endodermal lineages like pancreatic β-cells.

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Differentiation into Endoderm Lineage: Pancreatic differentiation from Embryonic Stem Cells

International Journal of Stem Cells ◽

10.15283/ijsc.2011.4.1.35 ◽

2011 ◽

Vol 4 (1) ◽

pp. 35-42 ◽

Cited By ~ 6

Author(s):

Dong Hyeon Lee ◽

Hyung Min Chung

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Embryonic Stem ◽

Pancreatic Differentiation

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Transdifferentiation of α-1,3-galactosyltransferase knockout pig bone marrow derived mesenchymal stem cells into pancreatic β-like cells by microenvironment modulation

Asian-Australasian Journal of Animal Sciences ◽

10.5713/ajas.19.0796 ◽

2020 ◽

Vol 33 (11) ◽

pp. 1837-1847

Author(s):

Imran Ullah ◽

Ran Lee ◽

Keon Bong Oh ◽

Seongsoo Hwang ◽

Youngim Kim ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Cpg Island ◽

Morphological Changes ◽

Quantitative Polymerase Chain Reaction ◽

Differentiation Potential ◽

Pancreatic Differentiation ◽

Epigenetic Modifiers ◽

Pluripotent Marker

Objective: To evaluate the pancreatic differentiation potential of α-1,3-galactosyltransferase knockout (GalTKO) pig-derived bone marrow-derived mesenchymal stem cells (BM-MSCs) using epigenetic modifiers with different pancreatic induction media.Methods: The BM-MSCs have been differentiated into pancreatic β-like cells by inducing the overexpression of key transcription regulatory factors or by exposure to specific soluble inducers/small molecules. In this study, we evaluated the pancreatic differentiation of GalTKO pig-derived BM-MSCs using epigenetic modifiers, 5-azacytidine (5-Aza) and valproic acid (VPA), and two types of pancreatic induction media – advanced Dulbecco's modified Eagle's medium (ADMEM)-based and N2B27-based media. GalTKO BM-MSCs were treated with pancreatic induction media and the expression of pancreas-islets-specific markers was evaluated by real-time quantitative polymerase chain reaction, Western blotting, and immunofluorescence. Morphological changes and changes in the 5'-C-phosphate-G-3' (CpG) island methylation patterns were also evaluated.Results: The expression of the pluripotent marker (POU class 5 homeobox 1 [OCT4]) was upregulated upon exposure to 5-Aza and/or VPA. GalTKO BM-MSCs showed increased expression of neurogenic differentiation 1 in the ADMEM-based (5-Aza) media, while the expression of NK6 homeobox 1 was elevated in cells induced with the N2B27-based (5-Aza) media. Moreover, the morphological transition and formation of islets-like cellular clusters were also prominent in the cells induced with the N2B27-based media with 5-Aza. The higher insulin expression revealed the augmented trans-differentiation ability of GalTKO BM-MSCs into pancreatic β-like cells in the N2B27-based media than in the ADMEM-based media.Conclusion: 5-Aza treated GalTKO BM-MSCs showed an enhanced demethylation pattern in the second CpG island of the OCT4 promoter region compared to that in the GalTKO BM-MSCs. The exposure of GalTKO pig-derived BM-MSCs to the N2B27-based microenvironment can significantly enhance their trans-differentiation ability into pancreatic β-like cells.

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Dual Inhibition of BMP and WNT Signals Promotes Pancreatic Differentiation from Human Pluripotent Stem Cells

Stem Cells International ◽

10.1155/2019/5026793 ◽

2019 ◽

Vol 2019 ◽

pp. 1-15 ◽

Cited By ~ 2

Author(s):

Mengtian Tan ◽

Lai Jiang ◽

Yinglei Li ◽

Wei Jiang

Keyword(s):

Stem Cells ◽

Embryonic Stem Cells ◽

Beta Cells ◽

Pluripotent Stem Cells ◽

Embryonic Stem ◽

Signal Pathways ◽

Pancreatic Differentiation ◽

Dual Inhibition ◽

Induced Pluripotent ◽

Wnt Signal

Pathological or functional loss of pancreatic beta cells is the cause of diabetes. Understanding how signaling pathways regulate pancreatic lineage and searching for combinations of signal modulators to promote pancreatic differentiation will definitely facilitate the robust generation of functional beta cells for curing hyperglycemia. In this study, we first tested the effect of several potent BMP inhibitors on pancreatic differentiation using human embryonic stem cells. Next, we examined the endodermal lineage bias upon potent BMP inhibitor treatment and further checked the crosstalk between signal pathways governing endodermal lineage determination. Furthermore, we improved current pancreatic differentiation system based on the signaling pathway study. Finally, we used human-induced pluripotent stem cells to validate our finding. We found BMP inhibitors indeed not only blocked hepatic lineage but also impeded intestinal lineage from human definitive endoderm unexpectedly. Signaling pathway analysis indicated potent BMP inhibitor resulted in the decrease of WNT signal activity and inhibition of WNT could contribute to the improved pancreatic differentiation. Herein, we combined the dual inhibition of BMP and WNT signaling and greatly enhanced human pancreatic progenitor differentiation as well as beta cell generation from both embryonic stem cells and induced pluripotent stem cells. Conclusively, our present work identified the crosstalk between the BMP and WNT signal pathways during human endoderm patterning and pancreas specification, and provided an improved in vitro pancreatic directed differentiation protocol from human pluripotent stem cells.

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