Signaling pathways mediating a selective induction of nitric oxide synthase II by tumor necrosis factor alpha in nerve growth factor-responsive cells.

backgroundInflammation and oxidative stress play a critical role in neurodegeneration associated with acute and chronic insults of the nervous system. Notably, affected neurons are often responsive to and dependent on trophic factors such as nerve growth factor (NGF). We previously showed in NGF-responsive PC12 cells that tumor necrosis factor alpha (TNFalpha) and NGF synergistically induce the expression of the free-radical producing enzyme inducible nitric oxide synthase (iNOS). We proposed that NGF-responsive neurons might be selectively exposed to iNOS-mediated oxidative damage as a consequence of elevated TNFalpha levels. With the aim of identifying possible therapeutic targets, in the present study we investigated the signaling pathways involved in NGF/TNFalpha-promoted iNOS induction.

methodsWestern blotting, RT-PCR, transcription factor-specific reporter gene systems, mutant cells lacking the low affinity p75NTR NGF receptor and transfections of TNFalpha/NGF chimeric receptors were used to investigate signalling events associated with NGF/TNFalpha-promoted iNOS induction in PC12 cells.

resultsOur results show that iNOS expression resulting from NGF/TNFalpha combined treatment can be elicited in PC12 cells. Mutant PC12 cells lacking p75NTR did not respond, suggesting that p75NTR is required to mediate iNOS expression. Furthermore, cells transfected with chimeric TNFalpha/NGF receptors demonstrated that the simultaneous presence of both p75NTR and TrkA signaling is necessary to synergize with TNFalpha to mediate iNOS expression. Lastly, our data show that NGF/TNFalpha-promoted iNOS induction requires activation of the transcription factor nuclear factor kappa B (NF-kappaB).

conclusionCollectively, our in vitro model suggests that cells bearing both the high and low affinity NGF receptors may display increased sensitivity to TNFalpha in terms of iNOS expression and therefore be selectively at risk during acute (e.g. neurotrauma) or chronic (e.g. neurodegenerative diseases) conditions where high levels of pro-inflammatory cytokines in the nervous system occur pathologically. Our results also suggest that modulation of NFkappaB-promoted transcription of selective genes could serve as a potential therapeutic target to prevent neuroinflammation-induced neuronal damage.