title: Three Arachidonoylamide Derivatives Inhibit Pro-Inflammatory Genes Expression by Modulating NF-kB and AP1 Activities.
abstract: BACKGROUND
  Since the anti-inflammatory activity of arachidonic acid derivatives was previously reported, we synthesized three new amide derivatives of arachidonic acid (AA-Ds) and tested their anti-inflammatory effects on an in vitro skin inflammation model. Aim of our study was to find derivatives of natural compounds able to down regulate inflammatory signal transduction pathway.
  
  
  METHODS
  Human keratinocytes cell line (HaCaT) was cultured and induced by cytokines in the presence of AA-Ds. Cytokines administration elicited an inflammatory response mediated by NF-kB and STAT-1 activation that induced proinflammatory genes expression.
  
  
  RESULTS
  By real time PCR we found that 24 hours after induction all AA-Ds significantly inhibit inducible Nitric Oxide Synthase (iNOS), TNFα, Inhibitor α of NF-kB, chemokine (C-X-C motif) ligand 9 and 10 genes expression. We analyzed their molecular effects in particular on the iNOS gene expression. Since iNOS transcript half-life did not change with AA-Ds treatment, we excluded a prominent role of post-transcriptional regulation for this gene and focused our attention on its transcriptional regulation. Starting three-five hours after cytokines induction, HaCaT cells, pretreated with each compound, showed inhibition of both NF-kB DNA-binding and NF-kB p65-Ser536 phosphorylation. STAT1 activation was inhibited only by AA-D4 derivative. To explain why the inhibition of iNOS expression began late after induction we analyzed activities of others key transcription factors. AA-Ds treatment elicited early increases of AP1 DNA binding as well as c-Jun, c-Fos and Fra-1 mRNA levels. Our data agree with the repressing effects of AP1 on human iNOS promoter previously described in others cell systems (Kleinert et al.).
  
  
  CONCLUSIONS
  AA-Ds shown to be good candidates as inhibitors of several pro-inflammatory genes induction and our study provides indications for their possible use as new antiinflammatory drugs.
authors.forename: Alex
authors.initials: A
authors.lastname: Gregorelli
authors.forename: Anna
authors.initials: A
authors.lastname: Sgarbossa
authors.forename: Shahbaz
authors.initials: S
authors.lastname: Khan
authors.forename: Annunziata
authors.initials: A
authors.lastname: Soriente
authors.forename: Margherita
authors.initials: M
authors.lastname: De Rosa
authors.affiliation: Department of Pharmacy, University of Salerno, 84084 Fisciano (SA), Italy. saturnino@unisa.it.
authors.forename: Carmela
authors.initials: C
authors.lastname: Saturnino
authors.affiliation: Department of Neurosciences, Biomedicine and Movement Sciences, Biochemistry Section, University of Verona, Strada Le Grazie, 8, 37134 Verona, Italy. marta.menegazzi@univr.it.
authors.forename: Marta
authors.initials: M
authors.lastname: Menegazzi
mesh_terms: D000577:Amides / Q000494:pharmacology; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D001095:Arachidonic Acids / Q000235:genetics / Q000494:pharmacology*; D002460:Cell Line; D054357:Chemokine CXCL10 / Q000235:genetics; D054370:Chemokine CXCL9 / Q000235:genetics; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000235:genetics / Q000378:metabolism; D000072000:NF-KappaB Inhibitor alpha / Q000235:genetics; D052247:Nitric Oxide Synthase Type II / Q000235:genetics; D010766:Phosphorylation; D016760:Proto-Oncogene Proteins c-fos / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D050794:STAT1 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction; D018808:Transcription Factor AP-1 / Q000378:metabolism*; D051996:Transcription Factor RelA / Q000378:metabolism*; D014158:Transcription, Genetic; D015533:Transcriptional Activation; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics; D000099169:Fos-Related Antigen 1
chemical_list: D000577:Amides; D000894:Anti-Inflammatory Agents, Non-Steroidal; D001095:Arachidonic Acids; D054357:Chemokine CXCL10; D054370:Chemokine CXCL9; D048031:JNK Mitogen-Activated Protein Kinases; D000072000:NF-KappaB Inhibitor alpha; D052247:Nitric Oxide Synthase Type II; D016760:Proto-Oncogene Proteins c-fos; D012333:RNA, Messenger; D050794:STAT1 Transcription Factor; D018808:Transcription Factor AP-1; D051996:Transcription Factor RelA; D014409:Tumor Necrosis Factor-alpha; D000099169:Fos-Related Antigen 1; C114753:CXCL10 protein, human; C080810:CXCL9 protein, human; C494079:STAT1 protein, human; C496319:NOS2 protein, human