Abstract

BackgroundInducible nitric oxide synthase (iNOS) makes a great contribution to host defense and inflammation. In many settings, lipopolysaccharide (LPS) induces iNOS expression through activation of the inhibitor of κB-α (IκB-α)-nuclear factor-κB (NF-κB) cascade, whereas interferon-γ (IFN-γ) acts through Janus kinase (JAK)-signal transducer and activator of transcription 1 (STAT1) signals. Heat shock factor 1 (HSF1), a major regulator of heat shock protein transcription, has been shown to regulate the production of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), but it remains obscure whether and how HSF1 affects iNOS induction.MethodsWestern blot was used to measure the protein expression. The mRNA level was measured by real-time PCR. Silence of HSF1 was achieved by small interfering RNA. Nitric oxide (NO) content and NF-κB binding activity were assayed by commercial kits. Chromatin immunoprecipitation (ChIP) was used to measure the binding activity of NF-κB and STAT1 to iNOS promoters.ResultsHSF1 inhibition or knockdown prevented the LPS- and/or IFN-γ-stimulated iNOS protein expression in cultured microglia. HSF1 inhibition blocked iNOS mRNA transcription. These inhibitory effects of HSF1 inhibition on iNOS expression were confirmed in brain tissues from endotoxemic mice. Further analysis showed that HSF1 inhibition had no effect on IκB-α degradation and NF-κB or STAT1 phosphorylation in LPS/IFN-γ-stimulated cells. The nuclear transport of active NF-κB or STAT1 was also not affected by HSF1 inhibition, but HSF1 inhibition reduced the binding of NF-κB and STAT1 to their DNA elements. In addition, HSF1 inhibition reduced NF-κB and STAT1 bindings to iNOS promoter inside the LPS/IFN-γ-stimulated cells.ConclusionsThis preventing effect of HSF1 inhibition on iNOS mRNA transcription presents the necessary role of HSF1 in iNOS induction.

Highlights

  • Inducible nitric oxide synthase makes a great contribution to host defense and inflammation

  • Heat shock factor 1 (HSF1) inhibition attenuates Inducible nitric oxide synthase (iNOS) induction in LPS/IFN-γ-stimulated microglia To determine the role of HSF1 in iNOS induction, BV-2 cells were first treated with LPS and IFN-γ

  • HSF1 inhibition attenuates the DNA-binding activity of nuclear factor-κB (NF-κB) and signal transducer and activator of transcription 1 (STAT1) in LPS- or IFN-γ-stimulated BV-2 cells Since HSF1 inhibition did not appear to affect inhibitor of κB-α (IκB-α)– NF-κB and STAT1 signals, we investigated the possibility that HSF1 inhibition suppresses iNOS transcription by directly interfering with NF-κB and STAT1 binding to their DNA elements in promoters

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Summary

Introduction

Inducible nitric oxide synthase (iNOS) makes a great contribution to host defense and inflammation. Heat shock factor 1 (HSF1), a major regulator of heat shock protein transcription, has been shown to regulate the production of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), but it remains obscure whether and how HSF1 affects iNOS induction. Through production of nitric oxide (NO), it plays critical roles in a lot of pathophysiological processes [2]. INOS contributes to host defense and inflammation resolution through killing bacteria, tumor cells, and viruses [3,4,5]. Excessive iNOS induces self-damage in disorders associated with inflammation. Over-accumulated iNOS has been shown to damage mitochondrial functions and induce cellular apoptosis [6, 7]. INOS induction should be tightly controlled in order to balance the role of iNOS in host defense

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