title: Secretome analysis of chondroitin sulfate-treated chondrocytes reveals anti-angiogenic, anti-inflammatory and anti-catabolic properties.
abstract: BACKGROUND
  Chondroitin sulfate (CS) is a symptomatic slow-acting drug for osteoarthritis (OA) widely used in the clinic. The aim of this work is to find proteins whose secretion from cartilage cells under proinflammatory stimuli (IL-1β) is regulated by CS, employing a novel quantitative proteomic approach.
  
  
  METHODS
  Human articular chondrocytes released from three normal cartilages were grown in SILAC medium. When complete incorporation of the heavy isotope was achieved, chondrocytes were stimulated with IL-1β 5 ng/ml with or without CS pretreatment (200 µg/ml). Forty-eight hours later, chondrocyte secretomes were analyzed by nano-scale liquid chromatography-mass spectrometry. Real-time PCR, western blot and immunohistochemistry analyses were employed to confirm some of the results.
  
  
  RESULTS
  We could identify 75 different proteins in the secretome of human articular chondrocytes. Eighteen of these were modulated by CS with statistical significance (six increased and 12 decreased). In normal chondrocytes stimulated with IL-1β, CS reduces inflammation directly by decreasing the presence of several complement components (CFAB, C1S, CO3, and C1R) and also indirectly by increasing proteins such as TNFα-induced protein (TSG6). TSG6 overexpression correlates with a decrease in pro-matrix metalloproteinase activation (observed in MMP1 and MMP3 levels). Finally, we observed a strong CS-dependent increase of an angiogenesis inhibitor, thrombospondin-1.
  
  
  CONCLUSIONS
  We have generated a quantitative profile of chondrocyte extracellular protein changes driven by CS in the presence of IL-1β. We have also provided novel evidences of its anti-angiogenic, anti-inflammatory, and anti-catabolic properties. Demonstration of the anti-angiogenic action of CS might provide a novel therapeutic approach for OA targeting.
authors.forename: Valentina
authors.initials: V
authors.lastname: Calamia
authors.forename: Lucía
authors.initials: L
authors.lastname: Lourido
authors.forename: Patricia
authors.initials: P
authors.lastname: Fernández-Puente
authors.forename: Jesús
authors.initials: J
authors.lastname: Mateos
authors.forename: Beatriz
authors.initials: B
authors.lastname: Rocha
authors.forename: Eulalia
authors.initials: E
authors.lastname: Montell
authors.forename: Josep
authors.initials: J
authors.lastname: Vergés
authors.forename: Cristina
authors.initials: C
authors.lastname: Ruiz-Romero
authors.forename: Francisco J
authors.initials: FJ
authors.lastname: Blanco
mesh_terms: D045930:Anabolic Agents / Q000378:metabolism*; D020533:Angiogenesis Inhibitors / Q000378:metabolism*; D000893:Anti-Inflammatory Agents / Q000378:metabolism*; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002478:Cells, Cultured; D019902:Chondrocytes / Q000187:drug effects* / Q000378:metabolism*; D002809:Chondroitin Sulfates / Q000494:pharmacology*; D015923:Complement C1r / Q000378:metabolism; D003173:Complement C1s / Q000378:metabolism; D006801:Humans; D053583:Interleukin-1beta / Q000494:pharmacology; D020781:Matrix Metalloproteinase 1 / Q000378:metabolism; D019278:Matrix Metalloproteinase 3 / Q000378:metabolism; D019700:Thrombospondin 1 / Q000378:metabolism
chemical_list: D045930:Anabolic Agents; D020533:Angiogenesis Inhibitors; D000893:Anti-Inflammatory Agents; D015815:Cell Adhesion Molecules; D053583:Interleukin-1beta; C072273:TNFAIP6 protein, human; D019700:Thrombospondin 1; D002809:Chondroitin Sulfates; D015923:Complement C1r; D003173:Complement C1s; D019278:Matrix Metalloproteinase 3; D020781:Matrix Metalloproteinase 1