title: Association of endothelin-1 expression and cartilaginous endplate degeneration in humans.
abstract: BACKGROUND
  Inflammatory cytokines are involved in intervertebral disc (IVD) degeneration. Endothelin-1 (ET-1), a 21-amino-acid cytokine implicated with cartilage degradation, is secreted by vascular endothelial cells and also by many other cell types. The expression of ET-1 in human IVD cartilage endplate (CEP) and its role in disc degeneration have not been explored.
  
  
  RESULTS
  The expression of ET-1 in degenerated CEP was analyzed by immunohistochemical staining and Western blotting; ET-1 was demonstrated in cartilaginous endplate cells (CECs) by immunofluorescent staining. The ET-1 mRNA expression and protein production by CECs stimulated by tumor necrosis factor alpha (TNF-α), a pro-inflammatory cytokine, were determined by real-time PCR analysis and Western blotting, respectively. The matrix metalloprotease-1 (MMP-1), MMP-13 and tissue inhibitor of metalloproteases-1 (TIMP-1) levels in the supernatant of cultured CECs treated with ET-1 were determined using enzyme-linked immunosorbent assays. Nitric oxide (NO) release and nitric oxide synthase (NOS) activity were measured using a spectrophotometric assay. The apoptosis of CECs by ET-1 was measured by an Annexin V-FITC detection assay. The production of ET-1 in degenerated cartilage endplate was significantly higher than normal CEP. The results showed that ET-1 was expressed by CECs and modulated by TNF-α in a dose-dependent manner. ET-1 increased production of MMP-1 and MMP-13, decreased TIMP-1 production, and induced NO and NOS release by cultured CECs. The direct stimulation of CECs by ET-1 did not promote cell apoptosis.
  
  
  CONCLUSIONS
  The study results suggest that ET-1 played a pivotal role in human CEP degeneration, and may be a new target for development of therapies for this condition.
authors.affiliation: Department of Orthopedic Surgery, Zhongshan Hospital, Fudan University, Shanghai, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Yuan
authors.forename: Ming-Dong
authors.initials: MD
authors.lastname: Zhao
authors.forename: Feng-Lai
authors.initials: FL
authors.lastname: Yuan
authors.forename: Wu
authors.initials: W
authors.lastname: Che
authors.forename: Ping-Guo
authors.initials: PG
authors.lastname: Duan
authors.forename: Yi
authors.initials: Y
authors.lastname: Liu
authors.forename: Jian
authors.initials: J
authors.lastname: Dong
mesh_terms: D000328:Adult; D000368:Aged; D017209:Apoptosis / Q000187:drug effects; D002356:Cartilage / Q000378:metabolism* / Q000473:pathology*; D019902:Chondrocytes / Q000378:metabolism / Q000473:pathology; D019332:Endothelin-1 / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D005260:Female; D006801:Humans; D055959:Intervertebral Disc Degeneration / Q000235:genetics* / Q000378:metabolism; D008297:Male; D020781:Matrix Metalloproteinase 1 / Q000378:metabolism; D053509:Matrix Metalloproteinase 13 / Q000378:metabolism; D008875:Middle Aged; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000378:metabolism; D014176:Protein Biosynthesis / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology; D055815:Young Adult
chemical_list: D019332:Endothelin-1; D012333:RNA, Messenger; D019715:Tissue Inhibitor of Metalloproteinase-1; D014409:Tumor Necrosis Factor-alpha; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D053509:Matrix Metalloproteinase 13; D020781:Matrix Metalloproteinase 1
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