Limits...
Requirement for endogenous heat shock factor 1 in inducible nitric oxide synthase induction in murine microglia.

Huang C, Lu X, Tong L, Wang J, Zhang W, Jiang B, Yang R - J Neuroinflammation (2015)

Bottom Line: These inhibitory effects of HSF1 inhibition on iNOS expression were confirmed in brain tissues from endotoxemic mice.In addition, HSF1 inhibition reduced NF-κB and STAT1 bindings to iNOS promoter inside the LPS/IFN-γ-stimulated cells.This preventing effect of HSF1 inhibition on iNOS mRNA transcription presents the necessary role of HSF1 in iNOS induction.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 5887218@qq.com.

ABSTRACT

Background: Inducible 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.

Methods: Western 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.

Results: HSF1 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.

Conclusions: This preventing effect of HSF1 inhibition on iNOS mRNA transcription presents the necessary role of HSF1 in iNOS induction.

No MeSH data available.


Related in: MedlinePlus

HSF1 inhibition reduces iNOS expression in brains from endotoxemic mice. Mice were divided into sham (saline + DMSO), endotoxemia (LPS 25 mg/kg + DMSO), KRIBB11 alone treatment (5 mg/kg), and LPS + KRIBB11 groups. Mice were sacrificed 20 h after injections and iNOS protein and mRNA levels were measured in brain tissues (a–c). As shown, KRIBB11 treatment significantly suppressed both iNOS protein (n = 3, *P < 0.05 vs. LPS alone-treated group; **P < 0.01 vs. control or KRIBB11 alone-treated group) and mRNA (n = 3, *P < 0.05 vs. control or LPS alone-treated group) expressions in brain tissues from endotoxemic mice. All data were shown as mean ± S.E. NS, no significance
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4607096&req=5

Fig4: HSF1 inhibition reduces iNOS expression in brains from endotoxemic mice. Mice were divided into sham (saline + DMSO), endotoxemia (LPS 25 mg/kg + DMSO), KRIBB11 alone treatment (5 mg/kg), and LPS + KRIBB11 groups. Mice were sacrificed 20 h after injections and iNOS protein and mRNA levels were measured in brain tissues (a–c). As shown, KRIBB11 treatment significantly suppressed both iNOS protein (n = 3, *P < 0.05 vs. LPS alone-treated group; **P < 0.01 vs. control or KRIBB11 alone-treated group) and mRNA (n = 3, *P < 0.05 vs. control or LPS alone-treated group) expressions in brain tissues from endotoxemic mice. All data were shown as mean ± S.E. NS, no significance

Mentions: The above studies revealed the inhibitory role of HSF1 inhibition in iNOS induction in cultured microglia under pro-inflammatory stimuli. To establish the relevance of these biological findings in vivo, the effect of HSF1 inhibition on iNOS expression in brains was investigated using endotoxemic mice. No iNOS was detectable in the brain tissue in the sham group. Injection of LPS into mice induced a marked increase in iNOS protein and mRNA expression in brains (Fig. 4a–c). Consistent with the results observed in cultured microglia, pre-treating mice with KRIBB11 (5 mg/kg, 1 h) markedly prevented the iNOS protein and mRNA expression in brain tissues from endotoxemic mice (Fig. 4a–c).Fig. 4


Requirement for endogenous heat shock factor 1 in inducible nitric oxide synthase induction in murine microglia.

Huang C, Lu X, Tong L, Wang J, Zhang W, Jiang B, Yang R - J Neuroinflammation (2015)

HSF1 inhibition reduces iNOS expression in brains from endotoxemic mice. Mice were divided into sham (saline + DMSO), endotoxemia (LPS 25 mg/kg + DMSO), KRIBB11 alone treatment (5 mg/kg), and LPS + KRIBB11 groups. Mice were sacrificed 20 h after injections and iNOS protein and mRNA levels were measured in brain tissues (a–c). As shown, KRIBB11 treatment significantly suppressed both iNOS protein (n = 3, *P < 0.05 vs. LPS alone-treated group; **P < 0.01 vs. control or KRIBB11 alone-treated group) and mRNA (n = 3, *P < 0.05 vs. control or LPS alone-treated group) expressions in brain tissues from endotoxemic mice. All data were shown as mean ± S.E. NS, no significance
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4607096&req=5

Fig4: HSF1 inhibition reduces iNOS expression in brains from endotoxemic mice. Mice were divided into sham (saline + DMSO), endotoxemia (LPS 25 mg/kg + DMSO), KRIBB11 alone treatment (5 mg/kg), and LPS + KRIBB11 groups. Mice were sacrificed 20 h after injections and iNOS protein and mRNA levels were measured in brain tissues (a–c). As shown, KRIBB11 treatment significantly suppressed both iNOS protein (n = 3, *P < 0.05 vs. LPS alone-treated group; **P < 0.01 vs. control or KRIBB11 alone-treated group) and mRNA (n = 3, *P < 0.05 vs. control or LPS alone-treated group) expressions in brain tissues from endotoxemic mice. All data were shown as mean ± S.E. NS, no significance
Mentions: The above studies revealed the inhibitory role of HSF1 inhibition in iNOS induction in cultured microglia under pro-inflammatory stimuli. To establish the relevance of these biological findings in vivo, the effect of HSF1 inhibition on iNOS expression in brains was investigated using endotoxemic mice. No iNOS was detectable in the brain tissue in the sham group. Injection of LPS into mice induced a marked increase in iNOS protein and mRNA expression in brains (Fig. 4a–c). Consistent with the results observed in cultured microglia, pre-treating mice with KRIBB11 (5 mg/kg, 1 h) markedly prevented the iNOS protein and mRNA expression in brain tissues from endotoxemic mice (Fig. 4a–c).Fig. 4

Bottom Line: These inhibitory effects of HSF1 inhibition on iNOS expression were confirmed in brain tissues from endotoxemic mice.In addition, HSF1 inhibition reduced NF-κB and STAT1 bindings to iNOS promoter inside the LPS/IFN-γ-stimulated cells.This preventing effect of HSF1 inhibition on iNOS mRNA transcription presents the necessary role of HSF1 in iNOS induction.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 5887218@qq.com.

ABSTRACT

Background: Inducible 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.

Methods: Western 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.

Results: HSF1 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.

Conclusions: This preventing effect of HSF1 inhibition on iNOS mRNA transcription presents the necessary role of HSF1 in iNOS induction.

No MeSH data available.


Related in: MedlinePlus