title: Early nuclear factor-kappaB activation and inducible nitric oxide synthase expression in injured spinal cord neurons correlating with a diffuse reduction of constitutive nitric oxide synthase activity.
abstract: OBJECTIVE
  Because of toxicity at high concentrations, nitric oxide (NO) contributes to spinal cord injury (SCI) secondary lesions. At low concentrations NO modulates nuclear factor-kappaB (NF-kappaB) activation. The authors investigated the activity of neuronal and endothelial NO synthase (nNOS and eNOS) to determine correlations with NF-kappaB activation and inducible NOS (iNOS) expression soon after SCI.
  
  
  METHODS
  In 48 adult male Wistar rats clip-based (50 g/mm2/10 seconds) SCI was induced, and spinal cords were removed at different intervals for the following evaluations: 1) assaying specific activity of nNOS and eNOS; 2) electrophoresis mobility shift assay for activated NF-kappaB; 3) Northern blotting for iNOS; 4) immunohistochemistry for iNOS and NF-kappaB; and 5) immunofluorescence for iNOS and NF-kappaB. At 15 minutes postinjury, eNOS activity decreased significantly (p < 0.001), as did nNOS activity at 1 hour compared with these levels in control animals and rats killed at 15 and 30 minutes after SCI (p < 0.001). Basal NF-kappaB levels were variable in controls and at 15 and 30 minutes after injury. One hour postinjury, NF-kappaB activation was diffuse. Inducible NOS messenger RNA localized diffusely, peaking 6 hours after injury and remaining stable until 24 hours postinjury. Immunohistochemical analysis showed diffuse iNOS and NF-kappaB staining, especially in neurons inside and around the lesion. Immunofluorescence demonstrated that injured neurons were a source of NF-kappaB and iNOS soon after injury.
  
  
  CONCLUSIONS
  Both nNOS and eNOS exhibited different regulation and roles soon after injury: nNOS correlated with NF-kappaB activation, whereas eNOS may have participated in vascular changes of the injured spinal cord. Neurons seemed to play a pivotal role in modulating and amplifying the inflammatory response in the injured spinal cord.
authors.affiliation: Dipartimento di Scienze Neurologiche, Neurochirurgia, Università di Roma La Sapienza, Italia. m.miscusi@libero.it
authors.forename: Massimo
authors.initials: M
authors.lastname: Miscusi
authors.forename: Florian
authors.initials: F
authors.lastname: Ebner
authors.forename: Sabrina
authors.initials: S
authors.lastname: Ceccariglia
authors.forename: Marta
authors.initials: M
authors.lastname: Menegazzi
authors.forename: Sofia
authors.initials: S
authors.lastname: Mariotto
authors.forename: Luigi
authors.initials: L
authors.lastname: Berra
authors.forename: Aurora
authors.initials: A
authors.lastname: Del Fa
authors.forename: Carlo
authors.initials: C
authors.lastname: Gangitano
authors.forename: Liverana
authors.initials: L
authors.lastname: Lauretti
authors.forename: Paolo
authors.initials: P
authors.lastname: Missori
authors.forename: Roberto
authors.initials: R
authors.lastname: Delfini
authors.forename: Hisanori
authors.initials: H
authors.lastname: Suzuki
mesh_terms: D000818:Animals; D004195:Disease Models, Animal; D008297:Male; D016328:NF-kappa B / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D013119:Spinal Cord Injuries / Q000378:metabolism* / Q000473:pathology*; D013904:Thoracic Vertebrae; D013997:Time Factors
chemical_list: D016328:NF-kappa B; D012333:RNA, Messenger; D019001:Nitric Oxide Synthase