title: TNF-α Activates High-Mobility Group Box 1 - Toll-Like Receptor 4 Signaling Pathway in Human Aortic Endothelial Cells.
abstract: OBJECTIVE
  Toll-like receptor 4 (TLR4) interacts with endogenous substances as well as lipopolysaccharide. We explored whether TLR4 is implicated in tumor necrosis factor-α (TNF-α) signal transduction in human aortic endothelial cells.
  
  
  METHODS
  The pathway was evaluated by transfection of siRNAs, immunoprecipitation and Western blot analysis.
  
  
  RESULTS
  TNF-α activated spleen tyrosine kinase (Syk) within 10 min, which led to endothelin-1 (ET-1) production. TLR4 was also rapidly activated by TNF-α stimulation, as shown by recruitment of interleukin-1 receptor-associated kinase 1 to TLR4 and its adaptor molecule, myeloid differentiation factor 88 (MyD88). siRNA depletion of TLR4 markedly attenuated TNF-α-induced Syk activation and ET-1 production. TLR4 inhibitor (CLI-095), TLR4-neutralizing antibody and siRNA depletion of MyD88 also attenuated TNF-α-induced Syk activation. Syk was co-immunoprecipitated with TLR4, and TNF-α activated Syk bound to TLR4. High-mobility group box 1 (HMGB1) was rapidly released and associated with TLR4 after TNF-α stimulation with a peak at 5 min, which was prevented by N-acetylcysteine, an antioxidant. Glycyrrhizin (HMGB1 inhibitor), HMGB1-neutralizing antibody and siRNA depletion of HMGB1 all suppressed TNF-α-induced Syk activation and ET-1 production.
  
  
  CONCLUSIONS
  Upon TNF-α stimulation, TLR4 is activated by HMGB1 that is immediately released after the generation of reactive oxygen species, and plays a crucial role in the signal transduction.
authors.forename: Won Seok
authors.initials: WS
authors.lastname: Yang
authors.forename: Nam Jeong
authors.initials: NJ
authors.lastname: Han
authors.forename: Jin Ju
authors.initials: JJ
authors.lastname: Kim
authors.forename: Mee Jeong
authors.initials: MJ
authors.lastname: Lee
authors.forename: Su-Kil
authors.initials: SK
authors.lastname: Park
mesh_terms: D001011:Aorta / Q000166:cytology* / Q000378:metabolism; D002460:Cell Line; D042783:Endothelial Cells / Q000166:cytology* / Q000378:metabolism; D019332:Endothelin-1 / Q000378:metabolism; D024243:HMGB1 Protein / Q000378:metabolism*; D006801:Humans; D053594:Myeloid Differentiation Factor 88 / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction; D000072377:Syk Kinase / Q000378:metabolism; D051197:Toll-Like Receptor 4 / Q000378:metabolism*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism*
chemical_list: D019332:Endothelin-1; D024243:HMGB1 Protein; C585491:HMGB1 protein, human; C507779:MYD88 protein, human; D053594:Myeloid Differentiation Factor 88; D017382:Reactive Oxygen Species; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha; D000072377:Syk Kinase