title: Normalization of NF-κB activity in dorsal root ganglia neurons cultured from diabetic rats reverses neuropathy-linked markers of cellular pathology.
abstract: OBJECTIVE
  Dorsal root ganglia (DRG) sensory neurons cultured from 3 to 5 month streptozotocin (STZ)-induced diabetic rats exhibit structural and biochemical changes seen in peripheral nerve fibers in vivo, including axonal swellings, oxidative damage, reduced axonal sprouting, and decreased NF-κB activity. NF-κB is a transcription factor required by DRG neurons for survival and plasticity, and regulates transcription of antioxidant proteins (e.g. MnSOD). We hypothesized that the diabetes-induced decrease in NF-κB activity in DRG contributes to pathological phenomena observed in cultured DRG neurons from diabetic rats.
  
  
  METHODS
  NF-κB localization was assessed in intact DRG and neuron cultures using immunostaining. NF-κB activity was manipulated in sensory neuron cultures derived from age-matched normal or 3-5 month STZ-diabetic rats using pharmacological means and lentiviral expression of shRNA. The impact of diabetes and altered NF-κB activity on neuronal phenotype involved analysis of neurite outgrowth, neurite morphology, oxidative stress (lipid peroxidation) and expression of MnSOD.
  
  
  RESULTS
  STZ-induced diabetes caused a significant decrease in nuclear localization of NF-κB subunits p50 and c-rel, but no change in p65 in intact DRG. Inhibition of NF-κB in normal neuron cultures significantly increased axonal swellings and oxidative stress, and reduced both neurite outgrowth and expression of MnSOD. These phenomena mimicked markers of pathology in cultured DRG neurons from diabetic rats. Enhancement of NF-κB activity in cultured diabetic DRG neurons ameliorated the sub-optimal neurite outgrowth and MnSOD levels triggered by diabetes. Exogenous insulin enhanced nuclear localization of p50 and c-rel but not p65 in diabetic neuronal cultures.
  
  
  CONCLUSIONS
  The diabetes-induced decrease of nuclear localization of NF-κB subunits p50 and c-rel in DRG contributes to development of in vitro markers of peripheral neuropathy, possibly through impaired mitochondrial ROS scavenging by deficient MnSOD.
authors.affiliation: Division of Neurodegenerative Disorders, St Boniface Hospital Research Centre, Winnipeg, MB, Canada.
authors.forename: A
authors.initials: A
authors.lastname: Saleh
authors.forename: J
authors.initials: J
authors.lastname: Schapansky
authors.forename: D R
authors.initials: DR
authors.lastname: Smith
authors.forename: N
authors.initials: N
authors.lastname: Young
authors.forename: G L
authors.initials: GL
authors.lastname: Odero
authors.forename: B
authors.initials: B
authors.lastname: Aulston
authors.forename: P
authors.initials: P
authors.lastname: Fernyhough
authors.forename: G W
authors.initials: GW
authors.lastname: Glazner
mesh_terms: D000074183:ATPases Associated with Diverse Cellular Activities; D000447:Aldehydes / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D001369:Axons / Q000187:drug effects / Q000473:pathology; D002478:Cells, Cultured; D004265:DNA Helicases / Q000378:metabolism; D003921:Diabetes Mellitus, Experimental / Q000150:complications / Q000473:pathology*; D004195:Disease Models, Animal; D019922:GAP-43 Protein / Q000378:metabolism; D005727:Ganglia, Spinal / Q000473:pathology*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006943:Hyperglycemia / Q000209:etiology; D007004:Hypoglycemic Agents / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology; D008297:Male; D016328:NF-kappa B / Q000378:metabolism* / Q000494:pharmacology; D009363:Neoplasm Proteins / Q000378:metabolism; D016501:Neurites / Q000187:drug effects / Q000473:pathology; D029543:Nucleocytoplasmic Transport Proteins / Q000378:metabolism; D018384:Oxidative Stress / Q000502:physiology; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011984:Sensory Receptor Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology*; D013482:Superoxide Dismutase / Q000378:metabolism; D013997:Time Factors; D051996:Transcription Factor RelA / Q000378:metabolism; D014162:Transfection
chemical_list: D000447:Aldehydes; D019922:GAP-43 Protein; D007004:Hypoglycemic Agents; D007328:Insulin; D016328:NF-kappa B; D009363:Neoplasm Proteins; D029543:Nucleocytoplasmic Transport Proteins; D034741:RNA, Small Interfering; D051996:Transcription Factor RelA; C063447:p65 oncofetal mRNA transport protein, rat; D049452:Green Fluorescent Proteins; D013482:Superoxide Dismutase; D000074183:ATPases Associated with Diverse Cellular Activities; D004265:DNA Helicases; C486550:Ruvbl1 protein, rat; C027576:4-hydroxy-2-nonenal