Dataset Information

Effective endothelial cell and human pluripotent stem cell interactions generate functional insulin-producing beta cells.

ABSTRACT:

Aims/hypothesis

Endothelial cells (ECs) play an essential role in pancreatic organogenesis. We hypothesise that effective in vitro interactions between human microvascular endothelial cells (HMECs) and human pluripotent stem cells (hPSCs) results in the generation of functional pancreatic beta cells.

Methods

Embryoid bodies (EBs) derived from hPSCs were cultured alone (controls) or with ECs in collagen gels. Subsequently, cells were analysed for pancreatic beta cell markers, and then isolated and expanded. Insulin secretion in response to glucose was evaluated in vitro by static and dynamic (perifusion) assays, and in vivo by EB transplantation into immunodeficient mice.

Results

Co-cultured EBs had a higher expression of mature beta cells markers and enhanced insulin secretion in vitro, compared with controls. In mice, transplanted EBs had higher levels of human C-peptide secretion with a significant reduction in hyperglycaemia after the selective destruction of native pancreatic beta cells. In addition, there was significant in vitro upregulation of bone morphogenetic proteins 2 and 4 (BMP-2, 4) in co-cultured cells, compared with controls.

Conclusions/interpretation

ECs provide essential signalling in vitro, such as activation of the BMP pathway, for derivation of functional insulin-producing beta cells from hPSCs.

SUBMITTER: Talavera-Adame D

PROVIDER: S-EPMC5506104 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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Publications

Effective endothelial cell and human pluripotent stem cell interactions generate functional insulin-producing beta cells.

Talavera-Adame Dodanim D Woolcott Orison O OO Ignatius-Irudayam Joseph J Arumugaswami Vaithilingaraja V Geller David H DH Dafoe Donald C DC

Diabetologia 20160827 11

<h4>Aims/hypothesis</h4>Endothelial cells (ECs) play an essential role in pancreatic organogenesis. We hypothesise that effective in vitro interactions between human microvascular endothelial cells (HMECs) and human pluripotent stem cells (hPSCs) results in the generation of functional pancreatic beta cells.<h4>Methods</h4>Embryoid bodies (EBs) derived from hPSCs were cultured alone (controls) or with ECs in collagen gels. Subsequently, cells were analysed for pancreatic beta cell markers, and t ...[more]

PMID: 27567623

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Project description:BackgroundEndothelial cells generated from induced pluripotent stem cells (hiPSC-ECs) show the majority of endothelial cell characteristics and markers, such as cobblestone morphology and the expression of VEGF and VE-cadherin. However, these cells are failing to show a mature endothelial cell phenotype, which is represented by the low expression and production of von Willebrand Factor (VWF) leading to the round morphology of the Weibel Palade Bodies (WPBs). The aim of this study was to improve the maturation process of hiPSC-ECs and to increase the levels of VWF.MethodshiPSC-ECs were differentiated by a standard differentiation protocol from hiPSCs generated from healthy control donors. To induce maturation, the main focus was to increase the expression and/or production of VWF by the adjustment of potential parameters influencing differentiation and maturation. We also compared alternative differentiation protocols. Cells were analyzed for the expression of endothelial cell markers, WPB structure, and the production and secretion of VWF by flow cytometry, confocal microscopy and ELISA.ResultsThe generated hiPSC-ECs have typical endothelial cell surface expression profiles, with low expression levels of non-endothelial markers as expected. Co-culture with pericytes, varying concentrations and timing of differentiation factors, applying some level of flow, and the addition of HDAC inhibitors did not substantially improve maturation of hiPSC-ECs. Transfection with the transcription factor ETV2 to induce a faster hiPSC-EC differentiation process resulted in a limited increase in VWF production, secretion, and elongation of WPB structure. Alternative differentiation protocols had limited effect.ConclusionhiPSCs-ECs have the potential to show a more mature endothelial phenotype with elongated WPBs after >30 days in culture. However, this comes with limitations as there are very few cells detected, and cells are deteriorating after being in culture for extended periods of time.

Dataset Information

Effective endothelial cell and human pluripotent stem cell interactions generate functional insulin-producing beta cells.

Aims/hypothesis

Methods

Results

Conclusions/interpretation

Publications

Effective endothelial cell and human pluripotent stem cell interactions generate functional insulin-producing beta cells.

Similar Datasets

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