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NADPH oxidase isoform selective regulation of endothelial cell proliferation and survival

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Abstract

Proliferation and apoptosis of endothelial cells are crucial angiogenic processes that contribute to carcinogenesis and tumor progression. Emerging evidence implicates the regulation of proliferation and apoptosis by reactive oxygen species (ROS) such as superoxide and hydrogen peroxide (H2O2). In the present study, we investigated the roles of the ROS-generating Nox4- and Nox2-containing reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidases in proliferation of human endothelial cells by examining the impact of these enzyme systems on (1) specific proliferative and tumorigenic kinases, extracellular regulated kinase1/2 (ERK1/2) and Akt, (2) cytoskeletal organization, and (3) the mechanisms that influence cellular apoptosis. ROS production and the expression of NADPH oxidase subunit Nox4, but not Nox2, were markedly higher in proliferating than in quiescent endothelial cells. Addition of the H2O2 scavenger catalase or downregulation of Nox4 protein with specific siRNA reduced ROS levels, cell proliferation, and ERK1/2 phosphorylation but had no effect on either cell morphology or caspase 3/7 activity. Although downregulation of Nox2 protein with siRNA also reduced ROS production and cell proliferation, it caused an increase in caspase 3/7 activity, reduced Akt phosphorylation, and caused cytoskeletal disorganization. Therefore, in endothelial cells, Nox4-derived H2O2 activates ERK1/2 to promote proliferation, whereas Nox2-containing NADPH oxidase maintains the cytoskeleton and prevents apoptosis to support cell survival. Our study provides a new understanding of the molecular mechanisms that underpin endothelial cell survival and a rationale for the combined suppression of Nox4- and Nox2-containing NADPH oxidases for unwanted angiogenesis in cancer.

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Abbreviations

DPI:

Diphenylene iodonium

FAD:

Flavin adenine dinucleotide

FBS:

Fetal bovine serum

H2-DCFDA:

Dichlorodihydrofluorescein-diacetate

HMECs:

Human microvascular endothelial cells

HUVEC:

Human umbilical vein endothelial cells

H2O2 :

Hydrogen peroxide

ERK1/2:

Extracellular regulated kinase1/2

l-NAME:

NG-Nitro-l-arginine methyl ester

170-ODYA:

17-Octadecynoic acid

ROS:

Reactive oxygen species

PI-3 K:

Phosphatidyl inositol 3-kinase

TNF-α:

Tumor necrosis factor-alpha

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Acknowledgments

Supported by the National Health and Medical Research Council (NHMRC) of Australia. Fellowships from the NHMRC supported HP (327401), GJD (400303), GRD (465109), CGS (350327) and SS (232324).

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There are no competing financial interests for all authors involved.

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Authors and Affiliations

  1. Bernard O’Brien Institute of Microsurgery, The University of Melbourne, Fitzroy, Victoria, 3065, Australia

    Hitesh Peshavariya, Gregory J. Dusting & Fan Jiang

  2. Department of Pharmacology, Monash University, Clayton, Victoria, 3800, Australia

    Lesley R. Halmos, Christopher G. Sobey, Grant R. Drummond & Stavros Selemidis

Authors
  1. Hitesh Peshavariya
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  2. Gregory J. Dusting
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  3. Fan Jiang
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  4. Lesley R. Halmos
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  5. Christopher G. Sobey
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  6. Grant R. Drummond
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  7. Stavros Selemidis
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Corresponding author

Correspondence to Grant R. Drummond.

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Peshavariya, H., Dusting, G.J., Jiang, F. et al. NADPH oxidase isoform selective regulation of endothelial cell proliferation and survival. Naunyn-Schmied Arch Pharmacol 380, 193–204 (2009). https://doi.org/10.1007/s00210-009-0413-0

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  • DOI: https://doi.org/10.1007/s00210-009-0413-0

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