Rotational co-culture of clonal beta-cells with endothelial cells: effect of PPAR-gamma agonism in vitro on insulin and VEGF secretion

M. B. Paget, H. E. Murray, C. J. Bailey, Peter Flatt, R. Downing

Research output: Contribution to journalArticle

11 Citations (Scopus)

Abstract

Aim: Delayed graft revascularization impedes the success of human islet transplantation. This study utilized rotational co-culture of insulin secreting beta-cells with human umbilical vein endothelial cells (HUVECs) and a peroxisome proliferator-activated receptor gamma (PPAR-gamma) agonist to promote insulin and vascular endothelial growth factor (VEGF) secretory function. Methods: Clonal BRIN-BD11 (D11) cells were maintained in static culture (SC) and rotational culture (RC) +/- HUVEC and +/- the TZD (thiazolidinedione) rosiglitazone (10 mmol/l) as a specific PPAR-gamma agonist. HUVECs were cultured in SC and RC +/- D11 and +/- TZD. D11 insulin secretion was induced by static incubation with low glucose (1.67 mmol/l), high glucose (16.7 mmol/l) and high glucose with 10 mmol/l theophylline (G+T) and assessed by enzyme-linked immunosorbent assay (ELISA). HUVEC proliferation was determined by ATP luminescence, whereas VEGF secretion was quantified by ELISA. Co-cultured cells were characterized by immunostaining for insulin and CD31. Results: D11 SC and RC showed enhanced insulin secretion in response to 16.7 mmol/l and G+T (p < 0.01); without significant alteration by the TZD. Co-culture with HUVEC in SC and RC also increased D11 insulin secretion when challenged with 16.7 mmol/l and G+T (p < 0.01), and this was slightly enhanced by the TZD. The presence of HUVEC increased D11 SC and RC insulin secretion in response to high glucose and G+T, respectively (p < 0.01). Addition of the TZD increased SC and RC HUVEC ATP content (p < 0.01) and VEGF production (p < 0.01) in the presence and absence of D11 cells. Conclusions: Rotational co-culture of insulin secreting cells with endothelial cells, and exposure to a PPAR gamma agonist may improve the prospects for graft revascularization and function after implantation.
LanguageEnglish
Pages662-668
JournalDIABETES OBESITY &amp; METABOLISM
Volume13
Issue number7
DOIs
Publication statusPublished - Jul 2011

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PPAR gamma
Human Umbilical Vein Endothelial Cells
Coculture Techniques
Vascular Endothelial Growth Factor A
Endothelial Cells
Insulin
Glucose
rosiglitazone
Insulin-Secreting Cells
Adenosine Triphosphate
Enzyme-Linked Immunosorbent Assay
Transplants
Islets of Langerhans Transplantation
Theophylline
Luminescence
In Vitro Techniques
Cultured Cells
Cell Proliferation
2,4-thiazolidinedione

Cite this

@article{3da4dfd2c8cb45f9b99ae19e9e1d884c,
title = "Rotational co-culture of clonal beta-cells with endothelial cells: effect of PPAR-gamma agonism in vitro on insulin and VEGF secretion",
abstract = "Aim: Delayed graft revascularization impedes the success of human islet transplantation. This study utilized rotational co-culture of insulin secreting beta-cells with human umbilical vein endothelial cells (HUVECs) and a peroxisome proliferator-activated receptor gamma (PPAR-gamma) agonist to promote insulin and vascular endothelial growth factor (VEGF) secretory function. Methods: Clonal BRIN-BD11 (D11) cells were maintained in static culture (SC) and rotational culture (RC) +/- HUVEC and +/- the TZD (thiazolidinedione) rosiglitazone (10 mmol/l) as a specific PPAR-gamma agonist. HUVECs were cultured in SC and RC +/- D11 and +/- TZD. D11 insulin secretion was induced by static incubation with low glucose (1.67 mmol/l), high glucose (16.7 mmol/l) and high glucose with 10 mmol/l theophylline (G+T) and assessed by enzyme-linked immunosorbent assay (ELISA). HUVEC proliferation was determined by ATP luminescence, whereas VEGF secretion was quantified by ELISA. Co-cultured cells were characterized by immunostaining for insulin and CD31. Results: D11 SC and RC showed enhanced insulin secretion in response to 16.7 mmol/l and G+T (p < 0.01); without significant alteration by the TZD. Co-culture with HUVEC in SC and RC also increased D11 insulin secretion when challenged with 16.7 mmol/l and G+T (p < 0.01), and this was slightly enhanced by the TZD. The presence of HUVEC increased D11 SC and RC insulin secretion in response to high glucose and G+T, respectively (p < 0.01). Addition of the TZD increased SC and RC HUVEC ATP content (p < 0.01) and VEGF production (p < 0.01) in the presence and absence of D11 cells. Conclusions: Rotational co-culture of insulin secreting cells with endothelial cells, and exposure to a PPAR gamma agonist may improve the prospects for graft revascularization and function after implantation.",
author = "Paget, {M. B.} and Murray, {H. E.} and Bailey, {C. J.} and Peter Flatt and R. Downing",
year = "2011",
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Rotational co-culture of clonal beta-cells with endothelial cells: effect of PPAR-gamma agonism in vitro on insulin and VEGF secretion. / Paget, M. B.; Murray, H. E.; Bailey, C. J.; Flatt, Peter; Downing, R.

In: DIABETES OBESITY &amp; METABOLISM, Vol. 13, No. 7, 07.2011, p. 662-668.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Rotational co-culture of clonal beta-cells with endothelial cells: effect of PPAR-gamma agonism in vitro on insulin and VEGF secretion

AU - Paget, M. B.

AU - Murray, H. E.

AU - Bailey, C. J.

AU - Flatt, Peter

AU - Downing, R.

PY - 2011/7

Y1 - 2011/7

N2 - Aim: Delayed graft revascularization impedes the success of human islet transplantation. This study utilized rotational co-culture of insulin secreting beta-cells with human umbilical vein endothelial cells (HUVECs) and a peroxisome proliferator-activated receptor gamma (PPAR-gamma) agonist to promote insulin and vascular endothelial growth factor (VEGF) secretory function. Methods: Clonal BRIN-BD11 (D11) cells were maintained in static culture (SC) and rotational culture (RC) +/- HUVEC and +/- the TZD (thiazolidinedione) rosiglitazone (10 mmol/l) as a specific PPAR-gamma agonist. HUVECs were cultured in SC and RC +/- D11 and +/- TZD. D11 insulin secretion was induced by static incubation with low glucose (1.67 mmol/l), high glucose (16.7 mmol/l) and high glucose with 10 mmol/l theophylline (G+T) and assessed by enzyme-linked immunosorbent assay (ELISA). HUVEC proliferation was determined by ATP luminescence, whereas VEGF secretion was quantified by ELISA. Co-cultured cells were characterized by immunostaining for insulin and CD31. Results: D11 SC and RC showed enhanced insulin secretion in response to 16.7 mmol/l and G+T (p < 0.01); without significant alteration by the TZD. Co-culture with HUVEC in SC and RC also increased D11 insulin secretion when challenged with 16.7 mmol/l and G+T (p < 0.01), and this was slightly enhanced by the TZD. The presence of HUVEC increased D11 SC and RC insulin secretion in response to high glucose and G+T, respectively (p < 0.01). Addition of the TZD increased SC and RC HUVEC ATP content (p < 0.01) and VEGF production (p < 0.01) in the presence and absence of D11 cells. Conclusions: Rotational co-culture of insulin secreting cells with endothelial cells, and exposure to a PPAR gamma agonist may improve the prospects for graft revascularization and function after implantation.

AB - Aim: Delayed graft revascularization impedes the success of human islet transplantation. This study utilized rotational co-culture of insulin secreting beta-cells with human umbilical vein endothelial cells (HUVECs) and a peroxisome proliferator-activated receptor gamma (PPAR-gamma) agonist to promote insulin and vascular endothelial growth factor (VEGF) secretory function. Methods: Clonal BRIN-BD11 (D11) cells were maintained in static culture (SC) and rotational culture (RC) +/- HUVEC and +/- the TZD (thiazolidinedione) rosiglitazone (10 mmol/l) as a specific PPAR-gamma agonist. HUVECs were cultured in SC and RC +/- D11 and +/- TZD. D11 insulin secretion was induced by static incubation with low glucose (1.67 mmol/l), high glucose (16.7 mmol/l) and high glucose with 10 mmol/l theophylline (G+T) and assessed by enzyme-linked immunosorbent assay (ELISA). HUVEC proliferation was determined by ATP luminescence, whereas VEGF secretion was quantified by ELISA. Co-cultured cells were characterized by immunostaining for insulin and CD31. Results: D11 SC and RC showed enhanced insulin secretion in response to 16.7 mmol/l and G+T (p < 0.01); without significant alteration by the TZD. Co-culture with HUVEC in SC and RC also increased D11 insulin secretion when challenged with 16.7 mmol/l and G+T (p < 0.01), and this was slightly enhanced by the TZD. The presence of HUVEC increased D11 SC and RC insulin secretion in response to high glucose and G+T, respectively (p < 0.01). Addition of the TZD increased SC and RC HUVEC ATP content (p < 0.01) and VEGF production (p < 0.01) in the presence and absence of D11 cells. Conclusions: Rotational co-culture of insulin secreting cells with endothelial cells, and exposure to a PPAR gamma agonist may improve the prospects for graft revascularization and function after implantation.

U2 - 10.1111/j.1463-1326.2011.01392.x

DO - 10.1111/j.1463-1326.2011.01392.x

M3 - Article

VL - 13

SP - 662

EP - 668

JO - Diabetes, Obesity and Metabolism

T2 - Diabetes, Obesity and Metabolism

JF - Diabetes, Obesity and Metabolism

SN - 1463-1326

IS - 7

ER -