title: Endoplasmic reticulum stress induces the expression of COX-2 through activation of eIF2α, p38-MAPK and NF-κB in advanced glycation end products stimulated human chondrocytes.
abstract: BACKGROUND
  During aging, advanced glycation end products (AGEs) accumulate in articular cartilage. In this study we determined whether AGEs induce endoplasmic reticulum (ER) stress and studied the ER stress-activated pathways that stimulate cyclooxygenase-2 (COX-2) expression in human chondrocytes.
  
  
  METHODS
  Chondrocytes were stimulated with AGE-BSA. Gene expression was determined by quantitative PCR and protein expression was studied by immunoblotting. Studies to elucidate involved pathways were executed using siRNAs and specific inhibitors of eukaryotic initiation factor-2α (eIF2α), MAPKs and NF-κB.
  
  
  RESULTS
  AGE-BSA induced expression of GRP78 with concomitant increase in COX-2 expression was observed in human chondrocytes. In addition, expression of Bag-1, an ER stress marker was also increased by AGE-BSA. RAGE knockdown inhibited AGE-BSA-induced expression of GRP78 and COX-2. Treatment with eIF2α inhibitor or eIF2α knockdown inhibited AGE-BSA-induced expression of GRP78 and COX-2 with decreased PGE(2) production. Treatment with SB202190 inhibited AGE-BSA-induced expression of GRP78 and COX-2, while treatment with PD98051 inhibited AGE-BSA-induced GRP78 protein expression but had no effect on COX-2 protein expression. SP600125 had no effect on either GRP78 or COX-2 protein expression. Bay 11-7082 suppressed AGE-BSA-induced GRP78 and COX-2 expression. AGE-BSA-induced activation of NF-κB was inhibited by treatment with SB202190 and by eIF2α knockdown, but was not inhibited when chondrocytes were treated with SP600125 or PD98059.
  
  
  CONCLUSIONS
  This study demonstrates that AGEs induce ER stress and stimulate the expression of COX-2 through eIF2α, p38-MAPK and NF-κB pathways in human chondrocytes. Our results provide important insights into cartilage degradation in osteoarthritis associated with latent ER stress.
authors.affiliation: Department of Medicine, Division of Rheumatology, MetroHealth Medical Center, Cleveland, OH 44109, United States. zafarrasheed@rocketmail.com
authors.forename: Zafar
authors.initials: Z
authors.lastname: Rasheed
authors.forename: Tariq M
authors.initials: TM
authors.lastname: Haqqi
mesh_terms: D000328:Adult; D000368:Aged; D015153:Blotting, Western; D002358:Cartilage, Articular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D049109:Cell Proliferation; D019902:Chondrocytes / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism*; D000091342:Endoplasmic Reticulum Chaperone BiP; D059865:Endoplasmic Reticulum Stress / Q000502:physiology*; D004797:Enzyme-Linked Immunosorbent Assay; D015852:Eukaryotic Initiation Factor-2 / Q000235:genetics / Q000378:metabolism*; D005260:Female; D017127:Glycation End Products, Advanced / Q000494:pharmacology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism*; D010003:Osteoarthritis / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012710:Serum Albumin, Bovine / Q000494:pharmacology*; D015398:Signal Transduction / Q000187:drug effects; D048051:p38 Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism*
chemical_list: D000091342:Endoplasmic Reticulum Chaperone BiP; D015852:Eukaryotic Initiation Factor-2; D017127:Glycation End Products, Advanced; C000717247:HSPA5 protein, human; D016328:NF-kappa B; D012333:RNA, Messenger; C081628:advanced glycation end products-bovine serum albumin; D012710:Serum Albumin, Bovine; D051546:Cyclooxygenase 2; C496540:PTGS2 protein, human; D048051:p38 Mitogen-Activated Protein Kinases