Distinct signaling pathways for induction of type II NOS by IFNγ and LPS in BV-2 microglial cells

Nitric oxide (NO) release upon microglial cell activation has been implicated in the tissue injury and cell death in many neurodegenerative diseases. Recent studies have indicated the ability of interferon-γ (IFNγ) and lipopolysaccharides (LPS) to independently induce type II nitric oxide synthase (iNOS) expression and NO production in BV-2 microglial cells. However, a detailed comparison between the signaling pathways activating iNOS by these two agents has not been accomplished. Analysis of PKC isoforms revealed mainly the presence of PKCδ, ι and λ in BV-2 cells. Although both IFNγ and LPS could specifically enhance the tyrosine phosphorylation of PKCδ, treatment with IFNγ induced a steady increase of phospho-PKCδ for up to 1 h, whereas treatment with LPS elevated phospho-PKCδ levels only transiently, with peak activity at 5 min. Rottlerin, a specific inhibitor for PKCδ, dose-dependently inhibited IFNγ- and LPS-induced NO production. Despite the common involvement of PKCδ, IFNγ- but not LPS-induced NO production involved extracellular signal-regulated kinases (ERK1/2) cascade and IFNγ-induced phosphorylation of ERK1/2 was mediated through PKC. On the other hand, LPS- but not IFNγ-induced NO production was through stimulation of NF-κB activation and nuclear translocation to interact with DNA. These results demonstrated distinct signaling pathways for induction of iNOS by IFNγ and LPS in BV-2 microglial cells.