Characterization of NADPH oxidase 5 expression in human tumors and tumor cell lines with a novel mouse monoclonal antibody.

Abstract

Reactive oxygen species generated by NADPH oxidase 5 (Nox5) have been implicated in physiological and pathophysiological signaling pathways, including cancer development and progression. However, because immunological tools are lacking, knowledge of the role of Nox5 in tumor biology has been limited; the expression of Nox5 protein across tumors and normal tissues is essentially unknown. Here, we report the characterization and use of a mouse monoclonal antibody against a recombinant Nox5 protein (bp 600-746) for expression profiling of Nox5 in human tumors by tissue microarray analysis. Using our novel antibody, we also report the detection of endogenous Nox5 protein in human UACC-257 melanoma cells. Immunofluorescence, confocal microscopy, and immunohistochemical techniques were employed to demonstrate Nox5 localization throughout UACC-257 cells, with perinuclear enhancement. Tissue microarray analysis revealed, for the first time, substantial Nox5 overexpression in several human cancers, including those of prostate, breast, colon, lung, brain, and ovary, as well as in malignant melanoma and non-Hodgkin lymphoma; expression in most nonmalignant tissues was negative to weak. This validated mouse monoclonal antibody will promote further exploration of the functional significance of Nox5 in human pathophysiology, including tumor cell growth and proliferation.

• Authors: Smitha Antony, Yongzhong Wu, Stephen M Hewitt, Miriam R Anver, Donna Butcher, Guojian Jiang, Jennifer L Meitzler, Han Liu, Agnes Juhasz, Jiamo Lu, Krishnendu K Roy, James H Doroshow
• Journal: Free radical biology & medicine (Free Radic Biol Med) Vol. 65 Pg. 497-508 (Dec 2013) ISSN: 1873-4596 [Electronic] United States
• PMID: 23851018 (Publication Type: Journal Article, Research Support, N.I.H., Extramural)
• Copyright: Copyright © 2013. Published by Elsevier Inc.

Chemical References

• Antibodies, Monoclonal
• Biomarkers, Tumor
• Membrane Proteins
• NADPH Oxidase 5
• NADPH Oxidases
• NOX5 protein, human

Topics

• Animals
• Antibodies, Monoclonal
• Biomarkers, Tumor (analysis)
• Blotting, Western
• Cell Line, Tumor
• Female
• Fluorescent Antibody Technique
• Humans
• Immunohistochemistry
• Male
• Membrane Proteins (analysis, biosynthesis)
• Mice
• Microscopy, Confocal
• NADPH Oxidase 5
• NADPH Oxidases (analysis, biosynthesis)
• Neoplasms (enzymology)
• Real-Time Polymerase Chain Reaction
• Tissue Array Analysis