BMP-2 (bone morphogenetic protein-2) promotes differentiation of osteoblast precursor cells to mature osteoblasts that form healthy bone. In the present study, we demonstrate a novel mechanism of BMP-2-induced osteoblast differentiation. The antioxidant NAC (N-acetyl-L-cysteine) and the flavoprotein enzyme NAD(P)H oxidase inhibitor DPI (diphenyleneiodonium) prevented BMP-2-stimulated alkaline phosphatase expression and mineralized bone nodule formation in mouse 2T3 pre-osteoblasts. BMP-2 elicited a rapid generation of ROS (reactive oxygen species) concomitant with increased activation of NAD(P)H oxidase. NAC and DPI inhibited BMP-2-induced ROS production and NAD(P)H oxidase activity respectively. NAD(P)H oxidases display structurally similar catalytic subunits (Nox1–5) with differential expression in various cells. We demonstrate that 2T3 pre-osteoblasts predominantly express the Nox4 isotype of NAD(P)H oxidase. To extend this finding, we tested the functional effects of Nox4. Adenovirus-mediated expression of dominant-negative Nox4 inhibited BMP-2-induced alkaline phosphatase expression. BMP-2 promotes expression of BMP-2 for maintenance of the osteoblast phenotype. NAC and DPI significantly blocked BMP-2-stimulated expression of BMP2 mRNA and protein due to a decrease in BMP2 gene transcription. Dominant-negative Nox4 also mimicked this effect of NAC and DPI. Our results provide the first evidence for a new signalling pathway linking BMP-2-stimulated Nox4-derived physiological ROS to BMP-2 expression and osteoblast differentiation.
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Research Article|
December 22 2010
Reactive oxygen species derived from Nox4 mediate BMP2 gene transcription and osteoblast differentiation
Chandi C. Mandal;
Chandi C. Mandal
*Department of Pathology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, U.S.A.
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Suthakar Ganapathy;
Suthakar Ganapathy
*Department of Pathology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, U.S.A.
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Yves Gorin;
Yves Gorin
†Department of Medicine, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, U.S.A.
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Kalyankar Mahadev;
Kalyankar Mahadev
‡Department of Medicine, Institute for Diabetes, Obesity, and Metabolism, University of Pennsylvania, Philadelphia, PA 191904, U.S.A.
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Karen Block;
Karen Block
†Department of Medicine, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, U.S.A.
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Hanna E. Abboud;
Hanna E. Abboud
†Department of Medicine, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, U.S.A.
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Stephen E. Harris;
Stephen E. Harris
§VA Research, South Texas Veterans Health Care System, San Antonio, TX 78229, U.S.A.
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Goutam Ghosh-Choudhury;
Goutam Ghosh-Choudhury
†Department of Medicine, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, U.S.A.
∥Geriatric Research, Education and Clinical Center, South Texas Veterans Health Care System, San Antonio, TX 78229, U.S.A.
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Nandini Ghosh-Choudhury
Nandini Ghosh-Choudhury
1
*Department of Pathology, University of Texas Health Science Center at San Antonio, San Antonio, TX 78229, U.S.A.
∥Geriatric Research, Education and Clinical Center, South Texas Veterans Health Care System, San Antonio, TX 78229, U.S.A.
1To whom correspondence should be addressed (email choudhury@uthscsa.edu).
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Publisher: Portland Press Ltd
Received:
March 08 2010
Revision Received:
September 09 2010
Accepted:
October 29 2010
Accepted Manuscript online:
October 29 2010
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2011 Biochemical Society
2011
Biochem J (2011) 433 (2): 393–402.
Article history
Received:
March 08 2010
Revision Received:
September 09 2010
Accepted:
October 29 2010
Accepted Manuscript online:
October 29 2010
Citation
Chandi C. Mandal, Suthakar Ganapathy, Yves Gorin, Kalyankar Mahadev, Karen Block, Hanna E. Abboud, Stephen E. Harris, Goutam Ghosh-Choudhury, Nandini Ghosh-Choudhury; Reactive oxygen species derived from Nox4 mediate BMP2 gene transcription and osteoblast differentiation. Biochem J 15 January 2011; 433 (2): 393–402. doi: https://doi.org/10.1042/BJ20100357
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