Proliferation, migration, and ERK activation in human retinal endothelial cells through A2B adenosine receptor stimulation

M. B. Grant, M. I. Davis, S. Caballero, I. Feoktistov, I. Biaggioni, L. Belardinelli

Research output: Contribution to journalArticle

102 Citations (Scopus)

Abstract

PURPOSE. The nucleoside adenosine has been implicated in angiogenesis. A previous study demonstrated that activation of the A2B adenosine receptor (AdoR) increases cAMP accumulation, cell proliferation, and VEGF expression in human retinal endothelial cells (HRECs). In the present study, the role of this receptor was further characterized by examination of the effects of the selective A2B AdoR antagonists 3-N-propylxanthine (enprofylline) and 3-isobutyl-8-pyrrolidinoxanthine (IPDX) on AdoR-mediated HREC proliferation, capillary tube formation, and signal-transduction pathways. METHODS. HRECs were exposed to the adenosine analogue 5′-N-ethylcarboxamido-adenosine (NECA) in the absence or presence of AdoR antagonists. Migration was measured using Boyden chambers. Proliferation was assessed by counting cells. Western analysis was used to assess extracellular signal-related kinase (ERK) and cAMP response element-binding protein (CREB) in cell lysates. The effect of AdoR activation on tube formation was studied using cells grown on a synthetic basement membrane matrix. RESULTS. NECA induced proliferation in a concentration-dependent manner that was inhibited by enprofylline and IPDX. NECA stimulated chemotaxis in a concentration-dependent manner that was also blocked by both A2B AdoR antagonists. NECA activated ERK and CREB in HRECs. Both A2B AdoR antagonists diminished activation of ERK by NECA exposure. ERK activation was also blocked by the ERK-mitogen-activated protein kinase (MAPK) inhibitor PD98059, but not by the protein kinase A (PKA) inhibitor H-89. CREB activation was blocked by H-89, but not by PD98059, suggesting that ERK activation is independent of PKA. NECA enhanced tube formation on the matrix, whereas both A2B AdoR antagonists attenuated this effect. CONCLUSIONS. The selective A2B AdoR antagonists, enprofylline and IPDX, inhibited NECA-stimulated proliferation, ERK activation, cell migration, and capillary tube formation. A2B AdoR inhibition may offer a way to inhibit retinal angiogenesis and provide a novel therapeutic approach to treatment of diseases associated with aberrant neovascularization, such as diabetic retinopathy and retinopathy of prematurity.

Original languageEnglish (US)
Pages (from-to)2068-2073
Number of pages6
JournalInvestigative Ophthalmology and Visual Science
Volume42
Issue number9
StatePublished - 2001
Externally publishedYes

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Adenosine A2B Receptors
Adenosine
Adenosine A2 Receptor Antagonists
Phosphotransferases
Endothelial Cells
Cyclic AMP Response Element-Binding Protein
Purinergic P1 Receptors
Protein Kinase Inhibitors
Cyclic AMP-Dependent Protein Kinases
Cell Proliferation
Purinergic P1 Receptor Antagonists
Retinopathy of Prematurity
Diabetic Retinopathy
Chemotaxis
Mitogen-Activated Protein Kinases
Nucleosides
Basement Membrane
Vascular Endothelial Growth Factor A
Cell Movement
Signal Transduction

ASJC Scopus subject areas

  • Ophthalmology

Cite this

Grant, M. B., Davis, M. I., Caballero, S., Feoktistov, I., Biaggioni, I., & Belardinelli, L. (2001). Proliferation, migration, and ERK activation in human retinal endothelial cells through A2B adenosine receptor stimulation. Investigative Ophthalmology and Visual Science, 42(9), 2068-2073.

Proliferation, migration, and ERK activation in human retinal endothelial cells through A2B adenosine receptor stimulation. / Grant, M. B.; Davis, M. I.; Caballero, S.; Feoktistov, I.; Biaggioni, I.; Belardinelli, L.

In: Investigative Ophthalmology and Visual Science, Vol. 42, No. 9, 2001, p. 2068-2073.

Research output: Contribution to journalArticle

Grant, MB, Davis, MI, Caballero, S, Feoktistov, I, Biaggioni, I & Belardinelli, L 2001, 'Proliferation, migration, and ERK activation in human retinal endothelial cells through A2B adenosine receptor stimulation', Investigative Ophthalmology and Visual Science, vol. 42, no. 9, pp. 2068-2073.
Grant, M. B. ; Davis, M. I. ; Caballero, S. ; Feoktistov, I. ; Biaggioni, I. ; Belardinelli, L. / Proliferation, migration, and ERK activation in human retinal endothelial cells through A2B adenosine receptor stimulation. In: Investigative Ophthalmology and Visual Science. 2001 ; Vol. 42, No. 9. pp. 2068-2073.
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abstract = "PURPOSE. The nucleoside adenosine has been implicated in angiogenesis. A previous study demonstrated that activation of the A2B adenosine receptor (AdoR) increases cAMP accumulation, cell proliferation, and VEGF expression in human retinal endothelial cells (HRECs). In the present study, the role of this receptor was further characterized by examination of the effects of the selective A2B AdoR antagonists 3-N-propylxanthine (enprofylline) and 3-isobutyl-8-pyrrolidinoxanthine (IPDX) on AdoR-mediated HREC proliferation, capillary tube formation, and signal-transduction pathways. METHODS. HRECs were exposed to the adenosine analogue 5′-N-ethylcarboxamido-adenosine (NECA) in the absence or presence of AdoR antagonists. Migration was measured using Boyden chambers. Proliferation was assessed by counting cells. Western analysis was used to assess extracellular signal-related kinase (ERK) and cAMP response element-binding protein (CREB) in cell lysates. The effect of AdoR activation on tube formation was studied using cells grown on a synthetic basement membrane matrix. RESULTS. NECA induced proliferation in a concentration-dependent manner that was inhibited by enprofylline and IPDX. NECA stimulated chemotaxis in a concentration-dependent manner that was also blocked by both A2B AdoR antagonists. NECA activated ERK and CREB in HRECs. Both A2B AdoR antagonists diminished activation of ERK by NECA exposure. ERK activation was also blocked by the ERK-mitogen-activated protein kinase (MAPK) inhibitor PD98059, but not by the protein kinase A (PKA) inhibitor H-89. CREB activation was blocked by H-89, but not by PD98059, suggesting that ERK activation is independent of PKA. NECA enhanced tube formation on the matrix, whereas both A2B AdoR antagonists attenuated this effect. CONCLUSIONS. The selective A2B AdoR antagonists, enprofylline and IPDX, inhibited NECA-stimulated proliferation, ERK activation, cell migration, and capillary tube formation. A2B AdoR inhibition may offer a way to inhibit retinal angiogenesis and provide a novel therapeutic approach to treatment of diseases associated with aberrant neovascularization, such as diabetic retinopathy and retinopathy of prematurity.",
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T1 - Proliferation, migration, and ERK activation in human retinal endothelial cells through A2B adenosine receptor stimulation

AU - Grant, M. B.

AU - Davis, M. I.

AU - Caballero, S.

AU - Feoktistov, I.

AU - Biaggioni, I.

AU - Belardinelli, L.

PY - 2001

Y1 - 2001

N2 - PURPOSE. The nucleoside adenosine has been implicated in angiogenesis. A previous study demonstrated that activation of the A2B adenosine receptor (AdoR) increases cAMP accumulation, cell proliferation, and VEGF expression in human retinal endothelial cells (HRECs). In the present study, the role of this receptor was further characterized by examination of the effects of the selective A2B AdoR antagonists 3-N-propylxanthine (enprofylline) and 3-isobutyl-8-pyrrolidinoxanthine (IPDX) on AdoR-mediated HREC proliferation, capillary tube formation, and signal-transduction pathways. METHODS. HRECs were exposed to the adenosine analogue 5′-N-ethylcarboxamido-adenosine (NECA) in the absence or presence of AdoR antagonists. Migration was measured using Boyden chambers. Proliferation was assessed by counting cells. Western analysis was used to assess extracellular signal-related kinase (ERK) and cAMP response element-binding protein (CREB) in cell lysates. The effect of AdoR activation on tube formation was studied using cells grown on a synthetic basement membrane matrix. RESULTS. NECA induced proliferation in a concentration-dependent manner that was inhibited by enprofylline and IPDX. NECA stimulated chemotaxis in a concentration-dependent manner that was also blocked by both A2B AdoR antagonists. NECA activated ERK and CREB in HRECs. Both A2B AdoR antagonists diminished activation of ERK by NECA exposure. ERK activation was also blocked by the ERK-mitogen-activated protein kinase (MAPK) inhibitor PD98059, but not by the protein kinase A (PKA) inhibitor H-89. CREB activation was blocked by H-89, but not by PD98059, suggesting that ERK activation is independent of PKA. NECA enhanced tube formation on the matrix, whereas both A2B AdoR antagonists attenuated this effect. CONCLUSIONS. The selective A2B AdoR antagonists, enprofylline and IPDX, inhibited NECA-stimulated proliferation, ERK activation, cell migration, and capillary tube formation. A2B AdoR inhibition may offer a way to inhibit retinal angiogenesis and provide a novel therapeutic approach to treatment of diseases associated with aberrant neovascularization, such as diabetic retinopathy and retinopathy of prematurity.

AB - PURPOSE. The nucleoside adenosine has been implicated in angiogenesis. A previous study demonstrated that activation of the A2B adenosine receptor (AdoR) increases cAMP accumulation, cell proliferation, and VEGF expression in human retinal endothelial cells (HRECs). In the present study, the role of this receptor was further characterized by examination of the effects of the selective A2B AdoR antagonists 3-N-propylxanthine (enprofylline) and 3-isobutyl-8-pyrrolidinoxanthine (IPDX) on AdoR-mediated HREC proliferation, capillary tube formation, and signal-transduction pathways. METHODS. HRECs were exposed to the adenosine analogue 5′-N-ethylcarboxamido-adenosine (NECA) in the absence or presence of AdoR antagonists. Migration was measured using Boyden chambers. Proliferation was assessed by counting cells. Western analysis was used to assess extracellular signal-related kinase (ERK) and cAMP response element-binding protein (CREB) in cell lysates. The effect of AdoR activation on tube formation was studied using cells grown on a synthetic basement membrane matrix. RESULTS. NECA induced proliferation in a concentration-dependent manner that was inhibited by enprofylline and IPDX. NECA stimulated chemotaxis in a concentration-dependent manner that was also blocked by both A2B AdoR antagonists. NECA activated ERK and CREB in HRECs. Both A2B AdoR antagonists diminished activation of ERK by NECA exposure. ERK activation was also blocked by the ERK-mitogen-activated protein kinase (MAPK) inhibitor PD98059, but not by the protein kinase A (PKA) inhibitor H-89. CREB activation was blocked by H-89, but not by PD98059, suggesting that ERK activation is independent of PKA. NECA enhanced tube formation on the matrix, whereas both A2B AdoR antagonists attenuated this effect. CONCLUSIONS. The selective A2B AdoR antagonists, enprofylline and IPDX, inhibited NECA-stimulated proliferation, ERK activation, cell migration, and capillary tube formation. A2B AdoR inhibition may offer a way to inhibit retinal angiogenesis and provide a novel therapeutic approach to treatment of diseases associated with aberrant neovascularization, such as diabetic retinopathy and retinopathy of prematurity.

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