Brd4 Is Essential for IL-1β-Induced Inflammation in Human Airway Epithelial Cells

Abstract

BackgroundChronic inflammation and oxidative stress are key features of chronic obstructive pulmonary disease (COPD). Oxidative stress enhances COPD inflammation under the control of the pro-inflammatory redox-sensitive transcription factor nuclear factor-kappaB (NF-κB). Histone acetylation plays a critical role in chronic inflammation and bromodomain and extra terminal (BET) proteins act as “readers” of acetylated histones. Therefore, we examined the role of BET proteins in particular Brd2 and Brd4 and their inhibitors (JQ1 and PFI-1) in oxidative stress- enhanced inflammation in human bronchial epithelial cells.MethodsHuman primary epithelial (NHBE) cells and BEAS-2B cell lines were stimulated with IL-1β (inflammatory stimulus) in the presence or absence of H2O2 (oxidative stress) and the effect of pre-treatment with bromodomain inhibitors (JQ1 and PFI-1) was investigated. Pro-inflammatory mediators (CXCL8 and IL-6) were measured by ELISA and transcripts by RT-PCR. H3 and H4 acetylation and recruitment of p65 and Brd4 to the native IL-8 and IL-6 promoters was investigated using chromatin immunoprecipitation (ChIP). The impact of Brd2 and Brd4 siRNA knockdown on inflammatory mediators was also investigated.ResultH2O2 enhanced IL1β-induced IL-6 and CXCL8 expression in NHBE and BEAS-2B cells whereas H2O2 alone did not have any affect. H3 acetylation at the IL-6 and IL-8 promoters was associated with recruitment of p65 and Brd4 proteins. Although p65 acetylation was increased this was not directly targeted by Brd4. The BET inhibitors JQ1 and PFI-1 significantly reduced IL-6 and CXCL8 expression whereas no effect was seen with the inactive enantiomer JQ1(-). Brd4, but not Brd2, knockdown markedly reduced IL-6 and CXCL8 release. JQ1 also inhibited p65 and Brd4 recruitment to the IL-6 and IL-8 promoters.ConclusionOxidative stress enhanced IL1β-induced IL-6 and CXCL8 expression was significantly reduced by Brd4 inhibition. Brd4 plays an important role in the regulation of inflammatory genes and provides a potential novel anti-inflammatory target.

Functions of BET proteins in erythroid gene expression

“…However, the assignment of enhancers to target genes by genomic distance is unreliable, potentially obscuring possible relationships between enhancer BET binding and JQ1 response. 4 Significantly, the most JQ1-sensitive genes were those activated by GATA1, consistent with the notion that BETs function generally in inducible transcription, 22,63,64 and activation by GATA1 predicted JQ1 sensitivity better than BET occupancy at superenhancers.Linking ChIP-seq and transcriptome data sets requires reliable quantification of transcripts. Cell-state transitions such as maturation can dramatically alter both total RNA and mRNA content, limiting reliability of normalization with internal standards.…”

Bromodomain-Containing Protein 4

“…41 This paves the way for the use of BRD4 inhibitors to modulate the autoimmune arm of PAH pathobiology. Indeed, autoimmune diseases, such as connective tissue diseases, make a favorable seedbed for associated PAH occurrence and autoantibodies are implicated in the development of both idiopathic PAH and PAH associated with connective tissue diseases.…”

“…16,24,40 BRD2 and BRD4 are often associated to cell cycle control and inflammation in multiple inflammatory diseases and cancers, although the role of BRD3 in the cell is less understood. 16,[40][41][42][43] Further investigations are required to determine a possible role for BRD2 and BRD3 in PAH.…”

Epigenetic and current treatment approaches in chronic obstructive pulmonary disease

“…SIRT1'in fosforilasyonunu inhibe eden yeni farmakolojik tedaviler KOAH'lı hastalarda inflamasyonu iyileştirebilir ancak steroid direncini düzeltmedeki etkileri henüz bilinmemektedir (35). KOAH'ta oksidatif stresle artan ve IL-1 tarafından uyarılan IL-6, IL-8 (KC) ekspresyonu bromodomain ve ekstraterminal proteinlerce (BET) baskılanarak antiinflamatuvar tedavi sağlanabileceği söylenmiştir (36). Sonuçta KOAH'ta epigenetik mekanizmalar çevresel etkenlerle gen ifadesini değiştirmektedir.…”