Proteomic analysis of osteoarthritic chondrocyte reveals the hyaluronic acid-regulated proteins involved in chondroprotective effect under oxidative stress.
Osteoarthritis (OA), the most common type of arthritis, is a degenerative joint disease. Oxidative stress is well known to play important roles in cartilage degradation and pathogenesis of OA. The intra-articular injection of hyaluronic acid (IAHA) is accepted as an effective clinical therapy for OA, but we do not yet fully understand the mechanisms underlying the effects of HA on OA chondrocytes under oxidative stress. Here, we show for the first time that IAHA significantly reduces the synovial fluid levels of hydrogen peroxide (H2O2) and superoxide (O2(-)) in patients with knee OA. We also demonstrate that HA suppresses H2O2-induced cell death in human OA chondrocytes. Proteomic approaches (2-DE combined with mass spectrometry) allowed us to identify 13 protein spots corresponding to 12 non-redundant proteins as HA-regulated proteins in OA chondrocytes under oxidative stress. The expression levels of three putative HA-regulated proteins (TALDO, ANXA1 and EF2) in control, H2O2-, HA- and HA/H2O2-treated OA chondrocytes were verified by Western blotting and the results indeed support the notion that HA acts in anti-oxidation, anti-apoptosis, and the promotion of cell survival. Our results collectively demonstrate the utility of proteomic approaches and provide new insights into the chondroprotective effects of HA on OA.


CONCLUSIONS
In the present study, we show for the first time that IAHA reduces the levels of H2O2 and O2(-) in synovial fluids from OA patients. We used primary cultured human OA chondrocytes as a model, treated cells with H2O2 to partly mimic their physiological conditions under oxidative stress, and examined the protection effects of HA. The proteomic approach allowed us to identify candidate proteins regulated by H2O2 and/or HA in OA chondrocytes. We found that proteins functioning in stress responses, apoptosis and protein synthesis were consistently regulated by HA in chondrocytes under oxidative stress. These novel results contribute to our understanding of the molecular mechanisms underlying HA-mediated chondroprotection.
Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan; Department of Cell and Molecular Biology, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan; Molecular Medicine Research Center, Chang Gung University, Tao-Yuan, Taiwan. Electronic address: yucj1124@mail.cgu.edu.tw.
Chia-Jung
CJ
Yu
Graduate Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Taipei, Taiwan.
Chun-Jung
CJ
Ko
Department of Orthopaedic Surgery, National Taiwan University Hospital, National Taiwan University, Taipei, Taiwan.
Chang-Hsun
CH
Hsieh
Department of Life Science, National Taiwan Normal University, Taipei, Taiwan.
Chiang-Ting
CT
Chien
Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan.
Lien-Hung
LH
Huang
Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Tao-Yuan, Taiwan.
Chien-Wei
CW
Lee
Department of Orthopaedic Surgery, National Taiwan University Hospital, National Taiwan University, Taipei, Taiwan.
Ching-Chuan
CC
Jiang
D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D017305:Annexin A1 / Q000378:metabolism; D000975:Antioxidants / Q000008:administration & dosage*; D016923:Cell Death; D019902:Chondrocytes / Q000378:metabolism* / Q000473:pathology; D005260:Female; D006801:Humans; D006820:Hyaluronic Acid / Q000008:administration & dosage*; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008297:Male; D008875:Middle Aged; D010003:Osteoarthritis / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects*; D020652:Peptide Elongation Factor 2 / Q000378:metabolism; D040901:Proteomics; D055675:Viscosupplements / Q000008:administration & dosage*
D017305:Annexin A1; D000975:Antioxidants; D016877:Oxidants; D020652:Peptide Elongation Factor 2; D055675:Viscosupplements; D006820:Hyaluronic Acid; D006861:Hydrogen Peroxide
Chondrocyte; Hyaluronic acid; Osteoarthritis; Oxidative stress; Proteomic