Inhibition of NADPH Oxidases Prevents Chronic Ethanol-Induced Bone Loss in Female Rats

Previous in vitro data suggest that ethanol (EtOH) activates NADPH oxidase (Nox) in osteoblasts leading to accumulation of reactive oxygen species (ROS). This might be a mechanism underlying inhibition of bone formation and increased bone resorption observed in vivo after EtOH exposure. In a rat model in which cycling females were infused intragastrically with EtOH-containing liquid diets, EtOH significantly decreased bone formation and stimulated osteoblast-dependent osteoclast differentiation. These effects were reversed by exogenous 17-β-estradiol coadministration. Moreover, coadministration of N-acetyl cysteine (NAC), an antioxidant, or diphenylene iodonium (DPI), a specific Nox inhibitor, also abolished chronic EtOH-associated bone loss. EtOH treatment up-regulated mRNA levels of Nox1, 2, 4, and the receptor activator of nuclear factor-κB ligand (RANKL), an essential factor for differentiation of osteoclasts in bone. Protein levels of Nox4, a major Nox isoform expressed in nonphagocytic cells, was also up-regulated by EtOH in bone. 17-β-Estradiol, NAC, and DPI were able to normalize EtOH-induced up-regulation of Nox and RANKL. In vitro experiments demonstrated that EtOH directly up-regulated Nox expression in osteoblasts. Pretreatment of osteoblasts with DPI eliminated EtOH-induced RANKL promoter activity. Furthermore, EtOH induced RANKL gene expression, and RANKL promoter activation in osteoblasts was ROS-dependent. These data suggest that inhibition of Nox expression and activity may be critical for prevention of chronic EtOH-induced osteoblast-dependent bone loss.

[1]  Phillip H. Roper,et al.  Long-term modulations in the vertebral transcriptome of adolescent-stage rats exposed to binge alcohol. , 2010, Alcohol and alcoholism.

[2]  S. Manolagas.  From estrogen-centric to aging and oxidative stress: a revised perspective of the pathogenesis of osteoporosis. , 2010, Endocrine reviews.

[3]  Hongli,et al.  Upregulation of Nox4 by Hypertrophic Stimuli Promotes Apoptosis and Mitochondrial Dysfunction in Cardiac Myocytes , 2010 .

[4]  N. Fukagawa,et al.  Age-related changes in redox signaling and VSMC function. , 2010, Antioxidants & redox signaling.

[5]  C. Rosen,et al.  Effects of alcohol on skeletal response to growth hormone in hypophysectomized rats. , 2010, Bone.

[6]  Michael L. Blackburn,et al.  A Role for Ethanol-Induced Oxidative Stress in Controlling Lineage Commitment of Mesenchymal Stromal Cells Through Inhibition of Wnt/β-Catenin Signaling , 2009, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[7]  T. Kawai,et al.  Interleukin 10 inhibits interferon γ- and tumor necrosis factor α-stimulated activation of NADPH oxidase 1 in human colonic epithelial cells and the mouse colon , 2009, Journal of Gastroenterology.

[8]  F. Martinez,et al.  NADPH Oxidase-4 Mediates Myofibroblast Activation and Fibrogenic Responses to Lung Injury , 2009, Nature Medicine.

[9]  J. Wadstein,et al.  Serum ethanol, hepatic enzymes and length of debauch in chronic alcoholics. , 2009, Acta medica Scandinavica.

[10]  R. Brandes,et al.  Nox4 Acts as a Switch Between Differentiation and Proliferation in Preadipocytes , 2009, Arteriosclerosis, thrombosis, and vascular biology.

[11]  Michael L. Blackburn,et al.  Ethanol Impairs Estrogen Receptor Signaling Resulting in Accelerated Activation of Senescence Pathways, Whereas Estradiol Attenuates the Effects of Ethanol in Osteoblasts , 2008, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[12]  F. Wezeman,et al.  Identification of novel bone-specific molecular targets of binge alcohol and ibandronate by transcriptome analysis. , 2008, Alcoholism, clinical and experimental research.

[13]  K. Shankar,et al.  Chronic ethanol consumption leads to disruption of vitamin D3 homeostasis associated with induction of renal 1,25 dihydroxyvitamin D3-24-hydroxylase (CYP24A1). , 2008, Endocrinology.

[14]  Yan-Hsiung Wang,et al.  Ethanol May Suppress Wnt/β-catenin Signaling on Human Bone Marrow Stroma Cells , 2008, Clinical orthopaedics and related research.

[15]  K. Shankar,et al.  Protective Effects of Estradiol on Ethanol-Induced Bone Loss Involve Inhibition of Reactive Oxygen Species Generation in Osteoblasts and Downstream Activation of the Extracellular Signal-Regulated Kinase/Signal Transducer and Activator of Transcription 3/Receptor Activator of Nuclear Factor-κB Ligan , 2008, Journal of Pharmacology and Experimental Therapeutics.

[16]  P. Roberson,et al.  Skeletal Involution by Age-associated Oxidative Stress and Its Acceleration by Loss of Sex Steroids* , 2007, Journal of Biological Chemistry.

[17]  F. Muller,et al.  Trends in oxidative aging theories. , 2007, Free radical biology & medicine.

[18]  Kutty Selva Nandakumar,et al.  Ethanol prevents development of destructive arthritis , 2007, Proceedings of the National Academy of Sciences.

[19]  K. Shankar,et al.  Estradiol Protects against Ethanol-Induced Bone Loss by Inhibiting Up-Regulation of Receptor Activator of Nuclear Factor-κB Ligand in Osteoblasts , 2006, Journal of Pharmacology and Experimental Therapeutics.

[20]  C. O’Brien,et al.  Parathyroid Hormone Controls Receptor Activator of NF-κB Ligand Gene Expression via a Distant Transcriptional Enhancer , 2006, Molecular and Cellular Biology.

[21]  V. Thakur,et al.  Chronic ethanol feeding increases activation of NADPH oxidase by lipopolysaccharide in rat Kupffer cells: role of increased reactive oxygen in LPS‐stimulated ERK1/2 activation and TNF‐α production , 2006, Journal of leukocyte biology.

[22]  J. Gimble,et al.  Playing with bone and fat , 2006, Journal of cellular biochemistry.

[23]  A. Patwardhan,et al.  Binge alcohol treatment increases vertebral bone loss following ovariectomy: compensation by intermittent parathyroid hormone. , 2006, Alcoholism, clinical and experimental research.

[24]  R. Brandes,et al.  Regulation of NAD(P)H Oxidase by Associated Protein Disulfide Isomerase in Vascular Smooth Muscle Cells* , 2005, Journal of Biological Chemistry.

[25]  D. Chakkalakal.  Alcohol-induced bone loss and deficient bone repair. , 2005, Alcoholism, clinical and experimental research.

[26]  Soo Young Lee,et al.  A crucial role for reactive oxygen species in RANKL-induced osteoclast differentiation. , 2005, Blood.

[27]  Z. Urry,et al.  A crucial role for thiol antioxidants in estrogen-deficiency bone loss. , 2003, The Journal of clinical investigation.

[28]  Jian Zhang,et al.  Osteoprotegerin Abrogates Chronic Alcohol Ingestion‐Induced Bone Loss in Mice , 2002, Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research.

[29]  I. Rusyn,et al.  Diphenyleneiodonium sulfate, an NADPH oxidase inhibitor, prevents early alcohol-induced liver injury in the rat. , 2001, American journal of physiology. Gastrointestinal and liver physiology.

[30]  MasayoshiHashimoto,et al.  Estrogen Prevents Oxidative Stress–Induced Endothelial Cell Apoptosis in Rats , 2001 .

[31]  R. Turner.  Skeletal response to alcohol. , 2000, Alcoholism, clinical and experimental research.

[32]  J. Dai,et al.  Chronic alcohol ingestion induces osteoclastogenesis and bone loss through IL-6 in mice. , 2000, The Journal of clinical investigation.

[33]  H. W. Sampson,et al.  Alcohol consumption inhibits osteoblastic cell proliferation and activity in vivo. , 1998, Alcohol.

[34]  G Shimamoto,et al.  Osteoprotegerin: A Novel Secreted Protein Involved in the Regulation of Bone Density , 1997, Cell.

[35]  A. Nègre-Salvayre,et al.  Ethinylestradiol does not enhance the expression of nitric oxide synthase in bovine endothelial cells but increases the release of bioactive nitric oxide by inhibiting superoxide anion production. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[36]  D. Rader,et al.  Oestrogen and inhibition of oxidation of low-density lipoproteins in postmenopausal women , 1994, The Lancet.

[37]  A. Wakeling,et al.  A potent specific pure antiestrogen with clinical potential. , 1991, Cancer research.

[38]  R. Turner,et al.  Chronic alcohol treatment results in disturbed vitamin D metabolism and skeletal abnormalities in rats. , 1988, Alcoholism, clinical and experimental research.

[39]  秀行 佐々木.  NADPH oxidase-derived reactive oxygen species are essential for differentiation of a mouse macrophage cell line (RAW264.7) into osteoclasts , 2009 .

[40]  J. Aronson,et al.  Different molecular mechanisms underlie ethanol-induced bone loss in cycling and pregnant rats. , 2006, Endocrinology.

[41]  H. W. Sampson,et al.  Alcohol and Other Factors Affecting Osteoporosis Risk in Women , 2002, Alcohol research & health : the journal of the National Institute on Alcohol Abuse and Alcoholism.

[42]  M. Ingelman-Sundberg,et al.  Effects of chronic ethanol on growth hormone secretion and hepatic cytochrome P450 isozymes of the rat. , 1993, The Journal of pharmacology and experimental therapeutics.