Endogenous hydrogen sulfide sulfhydrates IKKβ at cysteine 179 to control pulmonary artery endothelial cell inflammation.

Background: Pulmonary artery endothelial cell (PAEC) inflammation is a critical event in the development of pulmonary arterial hypertension (PAH). However, the pathogenesis of PAEC inflammation remains unclear. Methods: Purified recombinant human inhibitor of kappa B kinase subunit beta (IKK beta) protein, human PAECs and monocrotaline-induced pulmonary hypertensive rats were employed in the study. Site-directed mutagenesis, gene knockdown or overexpression were conducted to manipulate the expression or activity of a target protein. Results: We showed that hydrogen sulfide (H2S) inhibited IKk beta activation in the cell model of human PAEC inflammation induced by monocrotaline pyrrole-stimulation or knockdown of cystathionine gamma-lyase (CSE), an H2S generating enzyme. Mechanistically, H2S was proved to inhibit IKk beta activity directly via sulfhydrating IWO at cysteinyl residue 179 (C179) in purified recombinant IKk beta protein in vitro, whereas thiol reductant dithiothreitol (DTT) reversed H2S-induced MO inactivation. Furthermore, to demonstrate the significance of li to serine (C179S) in iwp. In purified IKk beta protein, C179S mutation of IKk beta abolished H2S-induced IKk beta sulfhydration and the subsequent imp inactivation. In human PAECs, C179S mutation of IKk beta blocked H2S-inhibited IKk beta activation and PAEC inflammatory response. In pulmonary hypertensive rats, C1795 mutation of IKk beta abolished the inhibitory effect of H2S on IKk beta activation and pulmonary vascular inflammation and remodeling. Conclusion: Collectively, our in vivo and in vitro findings demonstrated, for the first time, that endogenous H2S directly inactivated IKK beta via sulfhydrating IKK beta at Cys(179) to inhibit nuclear factor-kappa B (NF-kappa B) pathway activation and thereby control PAEC inflammation in PAH.