Excessive reactive oxygen species are therapeutic targets for intervertebral disc degeneration

Satoshi Suzuki, Nobuyuki Fujita, Naobumi Hosogane, Koota Watanabe, Ken Ishii, Yoshiaki Toyama, Keiyo Takubo, Keisuke Horiuchi, Takeshi Miyamoto, Masaya Nakamura, Morio Matsumoto

Research output: Contribution to journalArticle

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Abstract

Introduction: Oxidative stress has been reported to be involved in numerous human diseases, including musculoskeletal disorders such as osteoarthritis. However, the interaction between intervertebral disc (IVD) degeneration and oxidative stress is not well understood. The purpose of the present study was to elucidate the contribution of oxidative stress to IVD degeneration and the efficacy of antioxidant treatment for degenerative discs. Methods: The expression level of an oxidative stress marker, nitrotyrosine, was assessed by immunohistochemistry and Western blotting. For evaluating intracellular reactive oxygen species (ROS) levels and oxidative stress in rat annulus fibrosus (AF) cells, flow cytometry and luciferase assay with an OKD48 construct were performed. The grade of IVD degeneration was assessed by magnetic resonance imaging and histological analysis. Results: A high frequency of nitrotyrosine-positive cells was observed in rat and human degenerative discs. mRNA expression of catabolic factors such as tumor necrosis factor-alpha (TNF-alpha), matrix metalloprotease-3 (MMP-3), and cyclooxygenase-2 (COX-2) was significantly induced by treatment with H2O2 or buthionine sulfoximine, whereas that of aggrecan, an important chondrogenic proteoglycan, was reduced in a dose-dependent manner. Treatment with mitogen-activated protein kinase (MAPK) inhibitors blocked the inductive effect of excessive ROS on COX-2 mRNA expression. Western blotting confirmed the phosphorylation of MAPKs in H2O2 and BSO-treated AF cells. Conversely, we showed that TNF-aα induced oxidative stress with increased intracellular ROS levels in AF cells. Treatment with the antioxidant N-acetyl cysteine (NAC) abrogated the catabolic effect of excessive ROS and TNF-alpha in vitro. Finally, we showed that oral administration of NAC prevented IVD degeneration in rat degenerative model. Conclusions: A positive feedback loop was formed between excessive ROS and TNF-alpha in AF cells. Thus, oxidative stress contributes to the progression of IVD degeneration and NAC can be a therapeutic option for IVD degeneration.

Original languageEnglish
Article number316
JournalArthritis Research and Therapy
Volume17
Issue number1
DOIs
Publication statusPublished - 2015 Nov 5

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Intervertebral Disc Degeneration
Reactive Oxygen Species
Oxidative Stress
Cysteine
Tumor Necrosis Factor-alpha
Cyclooxygenase 2
Therapeutics
Antioxidants
Western Blotting
Musculoskeletal Diseases
Buthionine Sulfoximine
Aggrecans
Messenger RNA
Metalloproteases
Proteoglycans
Protein Kinase Inhibitors
Mitogen-Activated Protein Kinases
Luciferases
Osteoarthritis
Oral Administration

Keywords

  • Antioxidant
  • Intervertebral disc degeneration
  • Oxidative stress
  • Reactive oxygen species

ASJC Scopus subject areas

  • Rheumatology
  • Immunology
  • Immunology and Allergy

Cite this

Excessive reactive oxygen species are therapeutic targets for intervertebral disc degeneration. / Suzuki, Satoshi; Fujita, Nobuyuki; Hosogane, Naobumi; Watanabe, Koota; Ishii, Ken; Toyama, Yoshiaki; Takubo, Keiyo; Horiuchi, Keisuke; Miyamoto, Takeshi; Nakamura, Masaya; Matsumoto, Morio.

In: Arthritis Research and Therapy, Vol. 17, No. 1, 316, 05.11.2015.

Research output: Contribution to journalArticle

Suzuki, Satoshi ; Fujita, Nobuyuki ; Hosogane, Naobumi ; Watanabe, Koota ; Ishii, Ken ; Toyama, Yoshiaki ; Takubo, Keiyo ; Horiuchi, Keisuke ; Miyamoto, Takeshi ; Nakamura, Masaya ; Matsumoto, Morio. / Excessive reactive oxygen species are therapeutic targets for intervertebral disc degeneration. In: Arthritis Research and Therapy. 2015 ; Vol. 17, No. 1.
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AU - Suzuki, Satoshi

AU - Fujita, Nobuyuki

AU - Hosogane, Naobumi

AU - Watanabe, Koota

AU - Ishii, Ken

AU - Toyama, Yoshiaki

AU - Takubo, Keiyo

AU - Horiuchi, Keisuke

AU - Miyamoto, Takeshi

AU - Nakamura, Masaya

AU - Matsumoto, Morio

PY - 2015/11/5

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N2 - Introduction: Oxidative stress has been reported to be involved in numerous human diseases, including musculoskeletal disorders such as osteoarthritis. However, the interaction between intervertebral disc (IVD) degeneration and oxidative stress is not well understood. The purpose of the present study was to elucidate the contribution of oxidative stress to IVD degeneration and the efficacy of antioxidant treatment for degenerative discs. Methods: The expression level of an oxidative stress marker, nitrotyrosine, was assessed by immunohistochemistry and Western blotting. For evaluating intracellular reactive oxygen species (ROS) levels and oxidative stress in rat annulus fibrosus (AF) cells, flow cytometry and luciferase assay with an OKD48 construct were performed. The grade of IVD degeneration was assessed by magnetic resonance imaging and histological analysis. Results: A high frequency of nitrotyrosine-positive cells was observed in rat and human degenerative discs. mRNA expression of catabolic factors such as tumor necrosis factor-alpha (TNF-alpha), matrix metalloprotease-3 (MMP-3), and cyclooxygenase-2 (COX-2) was significantly induced by treatment with H2O2 or buthionine sulfoximine, whereas that of aggrecan, an important chondrogenic proteoglycan, was reduced in a dose-dependent manner. Treatment with mitogen-activated protein kinase (MAPK) inhibitors blocked the inductive effect of excessive ROS on COX-2 mRNA expression. Western blotting confirmed the phosphorylation of MAPKs in H2O2 and BSO-treated AF cells. Conversely, we showed that TNF-aα induced oxidative stress with increased intracellular ROS levels in AF cells. Treatment with the antioxidant N-acetyl cysteine (NAC) abrogated the catabolic effect of excessive ROS and TNF-alpha in vitro. Finally, we showed that oral administration of NAC prevented IVD degeneration in rat degenerative model. Conclusions: A positive feedback loop was formed between excessive ROS and TNF-alpha in AF cells. Thus, oxidative stress contributes to the progression of IVD degeneration and NAC can be a therapeutic option for IVD degeneration.

AB - Introduction: Oxidative stress has been reported to be involved in numerous human diseases, including musculoskeletal disorders such as osteoarthritis. However, the interaction between intervertebral disc (IVD) degeneration and oxidative stress is not well understood. The purpose of the present study was to elucidate the contribution of oxidative stress to IVD degeneration and the efficacy of antioxidant treatment for degenerative discs. Methods: The expression level of an oxidative stress marker, nitrotyrosine, was assessed by immunohistochemistry and Western blotting. For evaluating intracellular reactive oxygen species (ROS) levels and oxidative stress in rat annulus fibrosus (AF) cells, flow cytometry and luciferase assay with an OKD48 construct were performed. The grade of IVD degeneration was assessed by magnetic resonance imaging and histological analysis. Results: A high frequency of nitrotyrosine-positive cells was observed in rat and human degenerative discs. mRNA expression of catabolic factors such as tumor necrosis factor-alpha (TNF-alpha), matrix metalloprotease-3 (MMP-3), and cyclooxygenase-2 (COX-2) was significantly induced by treatment with H2O2 or buthionine sulfoximine, whereas that of aggrecan, an important chondrogenic proteoglycan, was reduced in a dose-dependent manner. Treatment with mitogen-activated protein kinase (MAPK) inhibitors blocked the inductive effect of excessive ROS on COX-2 mRNA expression. Western blotting confirmed the phosphorylation of MAPKs in H2O2 and BSO-treated AF cells. Conversely, we showed that TNF-aα induced oxidative stress with increased intracellular ROS levels in AF cells. Treatment with the antioxidant N-acetyl cysteine (NAC) abrogated the catabolic effect of excessive ROS and TNF-alpha in vitro. Finally, we showed that oral administration of NAC prevented IVD degeneration in rat degenerative model. Conclusions: A positive feedback loop was formed between excessive ROS and TNF-alpha in AF cells. Thus, oxidative stress contributes to the progression of IVD degeneration and NAC can be a therapeutic option for IVD degeneration.

KW - Antioxidant

KW - Intervertebral disc degeneration

KW - Oxidative stress

KW - Reactive oxygen species

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