Dataset Information

Hypoxia induction in cultured pancreatic islets enhances endothelial cell morphology and survival while maintaining beta-cell function.

ABSTRACT:

Background

Pancreatic islets are heavily vascularized in vivo yet lose this vasculature after only a few days in culture. Determining how to maintain islet vascularity in culture could lead to better outcomes in transplanting this tissue for the treatment of type 1 diabetes as well as provide insight into the complex communication between beta-cells and endothelial cells (ECs). We previously showed that islet ECs die in part due to limited diffusion of serum albumin into the tissue. We now aim to determine the impact of hypoxia on islet vascularization.

Methods

We induced hypoxia in cultured mouse islets using the hypoxia mimetic cobalt chloride (100 ?M CoCl2). We measured the impact on islet metabolism (two-photon NAD(P)H and Rh123 imaging) and function (insulin secretion and survival). We also measured the impact on hypoxia related transcripts (HIF-1?, VEGF-A, PDK-1, LDHA, COX4) and confirmed increased VEGF-A expression and secretion. Finally, we measured the vascularization of islets in static and flowing culture using PECAM-1 immunofluorescence.

Results

CoCl2 did not induce significant changes in beta cell metabolism (NAD(P)H and Rh123), insulin secretion, and survival. Consistent with hypoxia induction, CoCl2 stimulated HIF-1?, PDK-1, and LDHA transcripts and also stimulated VEGF expression and secretion. We observed a modest switch to the less oxidative isoform of COX4 (isoform 1 to 2) and this switch was noted in the glucose-stimulated cytoplasmic NAD(P)H responses. EC morphology and survival were greater in CoCl2 treated islets compared to exogenous VEGF-A in both static (dish) and microfluidic flow culture.

Conclusions

Hypoxia induction using CoCl2 had a positive effect on islet EC morphology and survival with limited impact on beta-cell metabolism, function, and survival. The EC response appears to be due to endogenous production and secretion of angiogenic factors (e.g. VEGF-A), and mechanistically independent from survival induced by serum albumin.

SUBMITTER: Sankar KS

PROVIDER: S-EPMC6786522 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Hypoxia induction in cultured pancreatic islets enhances endothelial cell morphology and survival while maintaining beta-cell function.

Sankar Krishana S KS Altamentova Svetlana M SM Rocheleau Jonathan V JV

PloS one 20191010 10

<h4>Background</h4>Pancreatic islets are heavily vascularized in vivo yet lose this vasculature after only a few days in culture. Determining how to maintain islet vascularity in culture could lead to better outcomes in transplanting this tissue for the treatment of type 1 diabetes as well as provide insight into the complex communication between beta-cells and endothelial cells (ECs). We previously showed that islet ECs die in part due to limited diffusion of serum albumin into the tissue. We n ...[more]

PMID: 31600313

Similar Datasets

Project description:BackgroundHypoxia is associated with the development of pancreatic cancer (PC). However, genes associated with hypoxia response and their regulatory mechanism in PC cells were unclear. The current study aims to investigate the role of the hypoxia associated gene fucosyltransferase 11 (FUT11) in the progression of PC.MethodsIn the preliminary study, bioinformatics analysis predicted FUT11 as a key hypoxia associated gene in PC. The expression of FUT11 in PC was evaluated using quantitative real-time PCR (qRT-PCR), Western blot and immunohistochemistry. The effects of FUT11 on PC cells proliferation and migration under normoxia and hypoxia were evaluated using Cell Counting Kit 8, 5-ethynyl-2'-deoxyuridine (EDU) assay, colony formation assay and transwell assay. The effects of FUT11 in vivo was examined in mouse tumor models of liver metastasis and subcutaneous xenograft. Furthermore, Western blot, luciferase assay and immunoprecipitation were performed to explore the regulatory relationship among FUT11, hypoxia-inducible factor 1α (HIF1α) and pyruvate dehydrogenase kinase 1 (PDK1) in PC.ResultsFUT11 was markedly increased of PC cells with hypoxia, upregulated in the PC clinical tissues, and predicted a poor outcome of PC patients. Inhibition of FUT11 reduced PC cell growth and migratory ability of PC cells under normoxia and hypoxia conditions in vitro, and growth and tumor cell metastasis in vivo. FUT11 bound to PDK1 and regulated the expression PDK1 under normoxia and hypoxia. FUT11 interacted with PDK1 and decreased the ubiquitination of PDK1, lead to the activation of AKT/mTOR signaling pathway. FUT11 knockdown significantly increased the degradation of PDK1 under hypoxia, while treatment with MG132 can relieve the degradation of PDK1 induced by FUT11 knockdown. Overexpression of PDK1 in PC cells under hypoxia conditions reversed the suppressive impacts of FUT11 knockdown on PC cell growth and migration. In addition, HIF1α bound to the promoter of FUT11 and increased its expression, as well as co-expressed with FUT11 in PC tissues. Furthermore, overexpression of FUT11 partially rescued the suppressive effects of HIF1α knockdown on PC cell growth and migration in hypoxia condition.ConclusionOur data implicate that hypoxia-induced FUT11 contributes to proliferation and metastasis of PC by maintaining the stability of PDK1, thus mediating activation of AKT/mTOR signaling pathway, and suggest that FUT11 could be a novel and effective target for the treatment of pancreatic cancer.

Dataset Information

Hypoxia induction in cultured pancreatic islets enhances endothelial cell morphology and survival while maintaining beta-cell function.

Background

Methods

Results

Conclusions

Publications

Hypoxia induction in cultured pancreatic islets enhances endothelial cell morphology and survival while maintaining beta-cell function.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets