title: Triptolide exhibits anti-inflammatory, anti-catabolic as well as anabolic effects and suppresses TLR expression and MAPK activity in IL-1β treated human intervertebral disc cells.
abstract: BACKGROUND
  Increased levels of proinflammatory cytokines seem to play a pivotal role in the development of back pain in a subpopulation of patients with degenerative intervertebral disc (IVD) disease. As current treatment options are mostly limited to surgical interventions or conservative treatment, anti-inflammatory substances might offer a novel, more target-orientated therapeutic approach. Triptolide (TPL), a natural substance found in the Chinese medicinal herb Tripterygium wilfordii Hook, has been demonstrated to possess anti-inflammatory effects in various cells, but no studies exist so far for the IVD. Therefore, the aim of this study was to determine the effects of TPL on human IVD cells by analyzing changes in gene expression and underlying molecular mechanisms.
  
  
  METHODS
  In order to investigate the anti-inflammatory, anabolic and anti-catabolic effect of TPL, dose-dependency experiments (n = 5) and time course experiments (n = 5) were performed on IL-1β prestimulated human IVD cells and changes in gene expression of IL-6/-8, TNF-α, PGE2S, MMP1/2/3/13, aggrecan and collagen-I/-II were analyzed by real-time RT-PCR. The molecular mechanisms underlying the effects observed upon TPL treatment were investigated by analyzing involvement of Toll-like receptors TLR2/4 (real-time RT-PCR, n = 5), NF-κB, MAP kinases p38, ERK and JNK (immunoblotting and immunocytochemistry, n = 4) as well as RNA polymerase II (immunoblotting, n = 3).
  
  
  RESULTS
  Results showed that 50 nM TPL exhibited an anti-inflammatory, anti-catabolic and anabolic effect on the mRNA level for IL-6/-8, PGE2S, MMP1/2/3/13, aggrecan, collagen-II and TLR2/4, with most pronounced changes after 18 h for proinflammatory cytokines and MMPs or 30 h for TLRs and matrix proteins. However, we also observed an up-regulation of TNF-α at higher concentrations. The effects of TPL did not seem to be mediated via an inhibition of NF-κB or a decrease of RNA polymerase II levels, but TPL influenced activity of MAP kinases p38 and ERK (but not JNK) and expression of TLR2/4.
  
  
  CONCLUSIONS
  In conclusion, TPL may possess promising potential for the treatment of inflammation-related discogenic back pain in vitro, but its analgetic effect will need to be confirmed in an appropriate in vivo animal model.
authors.affiliation: Spine Research Group, Competence Center for Applied Biotechnology and Molecular Medicine, University of Zurich, Winterthurerstrasse 190 (17L28), 8057, Zurich, Switzerland.
authors.forename: M
authors.initials: M
authors.lastname: Klawitter
authors.forename: L
authors.initials: L
authors.lastname: Quero
authors.forename: J
authors.initials: J
authors.lastname: Klasen
authors.forename: T
authors.initials: T
authors.lastname: Liebscher
authors.forename: A
authors.initials: A
authors.lastname: Nerlich
authors.forename: N
authors.initials: N
authors.lastname: Boos
authors.forename: K
authors.initials: K
authors.lastname: Wuertz
mesh_terms: D000328:Adult; D053674:Aggrecans / Q000378:metabolism; D045930:Anabolic Agents / Q000494:pharmacology*; D000893:Anti-Inflammatory Agents / Q000494:pharmacology*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D016207:Cytokines / Q000378:metabolism; D004224:Diterpenes / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004852:Epoxy Compounds / Q000494:pharmacology; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D066298:In Vitro Techniques; D053583:Interleukin-1beta / Q000494:pharmacology*; D007403:Intervertebral Disc / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D008297:Male; D008660:Metabolism / Q000187:drug effects*; D008875:Middle Aged; D020929:Mitogen-Activated Protein Kinase Kinases / Q000235:genetics / Q000378:metabolism*; D010616:Phenanthrenes / Q000494:pharmacology*; D013997:Time Factors; D051193:Toll-Like Receptors / Q000235:genetics / Q000378:metabolism*
chemical_list: D053674:Aggrecans; D045930:Anabolic Agents; D000893:Anti-Inflammatory Agents; D016207:Cytokines; D004224:Diterpenes; D004852:Epoxy Compounds; D053583:Interleukin-1beta; D010616:Phenanthrenes; D051193:Toll-Like Receptors; C001899:triptolide; D003094:Collagen; D020929:Mitogen-Activated Protein Kinase Kinases