title: Cysteine-Mediated Redox Regulation of Cell Signaling in Chondrocytes Stimulated With Fibronectin Fragments.
abstract: OBJECTIVE
  Oxidative posttranslational modifications of intracellular proteins can potentially regulate signaling pathways relevant to cartilage destruction in arthritis. In this study, oxidation of cysteine residues to form sulfenic acid (S-sulfenylation) was examined in osteoarthritic (OA) chondrocytes and investigated in normal chondrocytes as a mechanism by which fragments of fibronectin (FN-f) stimulate chondrocyte catabolic signaling.
  
  
  METHODS
  Chondrocytes isolated from OA and normal human articular cartilage were analyzed using analogs of dimedone that specifically and irreversibly react with protein S-sulfenylated cysteines. Global S-sulfenylation was measured in cell lysates with and without FN-f stimulation by immunoblotting and in fixed cells by confocal microscopy. S-sulfenylation in specific proteins was identified by mass spectroscopy and confirmed by immunoblotting. Src activity was measured in live cells using a fluorescence resonance energy transfer biosensor.
  
  
  RESULTS
  Proteins in chondrocytes isolated from OA cartilage were found to have elevated basal levels of S-sulfenylation relative to those of chondrocytes from normal cartilage. Treatment of normal chondrocytes with FN-f induced increased levels of S-sulfenylation in multiple proteins, including the tyrosine kinase Src. FN-f treatment also increased the levels of Src activity. Pretreatment with dimedone to alter S-sulfenylation function or with Src kinase inhibitors inhibited FN-f-induced production of matrix metalloproteinase 13.
  
  
  CONCLUSIONS
  These results demonstrate for the first time the presence of oxidative posttranslational modification of proteins in human articular chondrocytes by S-sulfenylation. Due to the ability to regulate the activity of a number of cell signaling pathways, including catabolic mediators induced by fibronectin fragments, S-sulfenylation may contribute to cartilage destruction in OA and warrants further investigation.
authors: University of North Carolina School of Medicine, Chapel Hill.
authors: Scott T
authors: ST
authors: Wood
authors: Wake Forest University School of Medicine, Winston-Salem, North Carolina.
authors: David L
authors: DL
authors: Long
authors: Wake Forest University School of Medicine, Winston-Salem, North Carolina.
authors: Julie A
authors: JA
authors: Reisz
authors: Wake Forest University School of Medicine, Winston-Salem, North Carolina.
authors: Raghunatha R
authors: RR
authors: Yammani
authors: Wake Forest University School of Medicine, Winston-Salem, North Carolina.
authors: Elizabeth A
authors: EA
authors: Burke
authors: Wake Forest University School of Medicine, Winston-Salem, North Carolina.
authors: Chananat
authors: C
authors: Klomsiri
authors: Wake Forest University School of Medicine, Winston-Salem, North Carolina.
authors: Leslie B
authors: LB
authors: Poole
authors: Wake Forest University School of Medicine, Winston-Salem, North Carolina.
authors: Cristina M
authors: CM
authors: Furdui
authors: University of North Carolina School of Medicine, Chapel Hill.
authors: Richard F
authors: RF
authors: Loeser
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D002358:Cartilage, Articular / Q000166:cytology*; D016022:Case-Control Studies; D019902:Chondrocytes / Q000187:drug effects / Q000378:metabolism*; D003512:Cyclohexanones / Q000494:pharmacology; D003545:Cysteine / Q000378:metabolism*; D005260:Female; D005353:Fibronectins / Q000494:pharmacology; D049690:History, Ancient; D006801:Humans; D015151:Immunoblotting; D013058:Mass Spectrometry; D053509:Matrix Metalloproteinase 13 / Q000187:drug effects / Q000378:metabolism; D018613:Microscopy, Confocal; D008875:Middle Aged; D010003:Osteoarthritis / Q000378:metabolism*; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000494:pharmacology; D011499:Protein Processing, Post-Translational; D015398:Signal Transduction; D013434:Sulfenic Acids / Q000378:metabolism*; D019061:src-Family Kinases / Q000187:drug effects / Q000378:metabolism*
chemical_list: D003512:Cyclohexanones; D005353:Fibronectins; D010446:Peptide Fragments; D013434:Sulfenic Acids; C004774:dimedone; D019061:src-Family Kinases; C507079:MMP13 protein, human; D053509:Matrix Metalloproteinase 13; D003545:Cysteine