The Role of Nuclear Factor- B in Oxidative Stress Associated with Rabies Virus Infection (P03.263)

Abstract

Objective: To investigate the role of NF-κB in rabies virus-induced oxidative stress in neurons. Background Recent studies in an experimental mouse model of rabies have shown that there are major structural changes in the brain involving neuronal processes that are associated with severe clinical disease. Cultured adult dorsal root ganglion (DRG) neurons are a good in vitro model for studying mechanisms involved in rabies virus – induced degeneration of neurites, because, unlike other neuronal cell types, these neurons are permissive to rabies virus infection. DRG neurons infected with the CVS strain of rabies virus show axonal swellings and immunostaining for 4-hydroxy-2-nonenal (4-HNE), indicating evidence of lipid peroxidation associated with oxidative stress, and also reduced axonal growth vs. mock-infected DRG neurons. Design/Methods: The expression and localization of NF-κB were evaluated by Western immunoblotting and immunofluorescence, respectively, in mock- versus CVS-infected rat DRG neurons. The effects of ciliary neurotrophic factor (CNTF), a potent activator of NF-κB, and SN50, a peptide inhibitor of NF-κB, were also evaluated. Results: CNTF had no effect on mock-infected rat DRG neurons, and was neuroprotective for CVS-neurons in reducing the number of 4-HNE-labeled puncta. SN50 and CVS infection had an additive effect in producing axonal swellings, suggesting that NF-κB is neuroprotective. CVS infection induced the expression of protein for the p50 subunit of NF-κB. The fluorescent signal for p50 was quantitatively evaluated in the nucleus and cytoplasm of mock- and CVS-infected rat DRG neurons. At 24 hrs p.i. there was a significant increase in the nucleus:cytoplasm ratio, indicating increased transcriptional activity of NF-kB, perhaps as a response to stress, whereas at both 48 and 72 hrs p.i. there was significantly reduced nuclear localization of NF-kB. Conclusions: CVS infection may induce oxidative stress by preventing nuclear activation of NF-κB. A rabies virus protein may directly inhibit NF-kB activity. Supported by: Canadian Institutes of Health Research. Disclosure: Dr. Jackson has nothing to disclose. Dr. Kammouni has nothing to disclose. Dr. Hasan has nothing to disclose. Dr. Fernyhough has nothing to disclose.