title: Betulinic acid regulates generation of neuroinflammatory mediators responsible for tissue destruction in multiple sclerosis in vitro.
abstract: OBJECTIVE
  To investigate the influences of betulinic acid (BA), a triterpenoid isolated from birch bark, on neuroinflammatory mediators involved in the pathogenesis of multiple sclerosis and experimental autoimmune encephalomyelitis in vitro.
  
  
  METHODS
  Encephalitogenic T cells were prepared from draining lymph nodes and spinal cords of Dark Agouti rats 8 to 10 d after immunization with myelin basic protein (MBP) and complete Freund's adjuvant. Macrophages were isolated from the peritoneal cavity of adult untreated rats. Astrocytes were isolated from neonatal rat brains. The cells were cultured and then treated with different agents. IFN-γ, IL-17, iNOS and CXCL12 mRNA levels in the cells were analyzed with RT-PCR. iNOS and CXCL12 protein levels were detected using immunoblot. NO and ROS generation was measured using Griess reaction and flow cytometry, respectively.
  
  
  RESULTS
  In encephalitogenic T cells stimulated with MBP (10 μg/mL), addition of BA inhibited IL-17 and IFN-γ production in a dose-dependent manner. The estimated IC(50) values for IL-17 and IFN γ were 11.2 and 63.8 μmol/L, respectively. When the macrophages were stimulated with LPS (10 ng/mL), addition of BA (50 μmol/L) significantly increased ROS generation, and suppressed NO generation. The astrocytes were stimulated with ConASn containing numerous inflammatory mediators, which mimicked the inflammatory milieu within CNS; addition of BA (50 μmol/L) significantly increased ROS generation, and blocked ConASn-induced increases in iNOS and CXCL12 mRNA levels, but did not affect iNOS and CXCL12 protein levels. Importantly, in both the macrophages and astrocytes, addition of BA (50 μmol/L) inhibited lipid peroxidation.
  
  
  CONCLUSIONS
  Besides inhibiting encephalitogenic T cell cytokines and reducing NO generation, BA induces tissue-damaging ROS generation within CNS.
authors: Department of Immunology, Institute for Biological Research Siniša Stanković, University of Belgrade, Serbia.
authors: Jana
authors: J
authors: Blaževski
authors: Filip
authors: F
authors: Petković
authors: Miljana
authors: M
authors: Momčilović
authors: Reinhard
authors: R
authors: Paschke
authors: Goran N
authors: GN
authors: Kaluđerović
authors: Marija
authors: M
authors: Mostarica Stojković
authors: Djordje
authors: D
authors: Miljković
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000008:administration & dosage / Q000302:isolation & purification / Q000494:pharmacology*; D001253:Astrocytes / Q000187:drug effects / Q000276:immunology; D018929:Cell Culture Techniques; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004681:Encephalomyelitis, Autoimmune, Experimental / Q000276:immunology* / Q000473:pathology; D004797:Enzyme-Linked Immunosorbent Assay; D015151:Immunoblotting; D018836:Inflammation Mediators / Q000276:immunology / Q000378:metabolism*; D008264:Macrophages / Q000187:drug effects / Q000276:immunology; D009103:Multiple Sclerosis / Q000276:immunology* / Q000473:pathology; D009569:Nitric Oxide / Q000378:metabolism; D053978:Pentacyclic Triterpenes; D051381:Rats; D011919:Rats, Inbred Strains; D017382:Reactive Oxygen Species / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D014315:Triterpenes / Q000008:administration & dosage / Q000302:isolation & purification / Q000494:pharmacology*; D000094062:Betulinic Acid
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D018836:Inflammation Mediators; D053978:Pentacyclic Triterpenes; D017382:Reactive Oxygen Species; D014315:Triterpenes; D009569:Nitric Oxide; D000094062:Betulinic Acid
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