title: Requirement for endogenous heat shock factor 1 in inducible nitric oxide synthase induction in murine microglia.
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.
authors.affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 5887218@qq.com.|Key Laboratory of Inflammation and Molecular Drug Targets of Jiangsu Province, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 5887218@qq.com.
authors.forename: Chao
authors.initials: C
authors.lastname: Huang
authors.affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 418796584@qq.com.|Key Laboratory of Inflammation and Molecular Drug Targets of Jiangsu Province, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 418796584@qq.com.
authors.forename: Xu
authors.initials: X
authors.lastname: Lu
authors.affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 815964117@qq.com.|Key Laboratory of Inflammation and Molecular Drug Targets of Jiangsu Province, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 815964117@qq.com.
authors.forename: Lijuan
authors.initials: L
authors.lastname: Tong
authors.affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 372899487@qq.com.|Key Laboratory of Inflammation and Molecular Drug Targets of Jiangsu Province, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 372899487@qq.com.
authors.forename: Jili
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. asicampa@aliyun.com.|Key Laboratory of Inflammation and Molecular Drug Targets of Jiangsu Province, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. asicampa@aliyun.com.
authors.forename: Wei
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Department of Pharmacology, School of Pharmacy, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 532059984@qq.com.|Key Laboratory of Inflammation and Molecular Drug Targets of Jiangsu Province, Nantong University, #19 Qixiu Road, Nantong, Jiangsu Province, 226001, China. 532059984@qq.com.
authors.forename: Bo
authors.initials: B
authors.lastname: Jiang
authors.affiliation: Department of Anesthesiology, Affiliated Hospital of Nantong University, Jiangsu Province, #20Xisi Road, Nantong, Jiangsu Province, 226001, China. 172916178@qq.com.
authors.forename: Rongrong
authors.initials: R
authors.lastname: Yang
mesh_terms: D000631:Aminopyridines / Q000494:pharmacology; D000818:Animals; D000831:Animals, Newborn; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D000076249:Heat Shock Transcription Factors; D007191:Indazoles / Q000494:pharmacology; D007371:Interferon-gamma / Q000494:pharmacology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D017628:Microglia / Q000187:drug effects / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D050794:STAT1 Transcription Factor / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D014157:Transcription Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*
chemical_list: D000631:Aminopyridines; D004268:DNA-Binding Proteins; D000076249:Heat Shock Transcription Factors; C490017:Hsf1 protein, mouse; D007191:Indazoles; D008070:Lipopolysaccharides; C556094:N2-(1H-indazole-5-yl)-N6-methyl-3-nitropyridine-2,6-diamine; D016328:NF-kappa B; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D050794:STAT1 Transcription Factor; C494080:Stat1 protein, mouse; D014157:Transcription Factors; D007371:Interferon-gamma; D052247:Nitric Oxide Synthase Type II