IκBα glutathionylation and reduced histone H3 phosphorylation inhibit eotaxin and RANTES

Airway smooth muscle cells (ASMCs) secrete eotaxin and RANTES (regulated on activation, normal T-cell expressed and secreted) in response to tumour necrosis factor (TNF)-alpha, which is inhibited by the nuclear factor (NF)-kappa B inhibitor dimethylfumarate (DMF). NF-kappa B/I kappa B (inhibitor of NF-kappa B) glutathionylation and changes in chromatin remodelling can inhibit NF-kappa B activity. In this study, we determined whether NF-kappa B/I kappa B glutathionylation and reduced histone H3 phosphorylation might underlie the inhibitory effect of DMF on NF-kappa B activity, and eotaxin and RANTES secretion. Primary human ASMCs were treated with DMF, diamide and/or glutathione (GSH) ethylester (OEt) prior to TNF-alpha stimulation and were subsequently analysed by ELISA, electrophoretic mobility shift assay, immunofluorescence, co-immunoprecipitation or immunoblotting. DMF reduced intracellular GSH and induced I kappa B alpha glutathionylation (I kappa B alpha-SSG), which inhibited I kappa B alpha degradation, NF-kappa B p65 nuclear entry and NF-kappa B/DNA binding. In addition, DMF inhibited the phosphorylation of histone H3, which was possibly mediated by the inhibitory effect of DMF on mitogen-and stress-activated protein kinase (MSK)-1. However, p38 mitogen-activated protein kinase (MAPK), extracellular signal-regulated kinase MAPK and MAPK phosphatase-1, upstream of MSK-1, were not inhibited by DMF. Importantly, DMF-mediated effects on NF-kappa B, histone H3, eotaxin and RANTES were reversed by addition of GSH-OEt. Our data suggest that DMF inhibits NF-kappa B-dependent eotaxin and RANTES secretion by reduction of GSH with subsequent induction of I kappa B alpha-SSG and inhibition of histone H3 phosphorylation. Our findings offer new potential drug targets to reduce airway inflammation in asthma.