DUOX1-mediated hydrogen peroxide release regulates sodium transport in H441 bronchiolar epithelial cells

Dexamethasone has been shown to induce the formation of epithelial domes by bronchiolar H441 cells. It stimulates the expression of both amiloride inhibitable epithelial sodium channels (ENaC) and dual oxidase-1 (DUOX1). We therefore ask the question whether DUOX1 expression and production of submillimolar amounts of H₂ O₂ is instrumental for the sodium channel upregulation observed in H441 cells.

In vitro cell culture, nystatin-perforated whole-cell patch-clamp technique, immunocytochemistry and RT-PCR methods have been used.

Cells forming epithelial domes induced by dexamethasone (0.1 μmol L^-1 , 24 hours) and by 5-aza-2'-deoxytidine (1 μmol L^-1 , 48 hours) expressed more DUOX1 protein compared with other cells in the monolayer. Dome formation could be inhibited by exogenous catalase in a concentration-dependent manner and by the NADPH oxidase inhibitor diphenyliodonium, which suggested the involvement of H₂ O₂ . While single application of 0.2 mmol L^-1 H₂ O₂ induced transient dome formation, lower doses were ineffective and higher doses disrupted the cell monolayer. Hydrogen peroxide (0.1 mmol L^-1 ) activated acutely amiloride-sensitive whole-cell currents from 3.91 ± 0.79 pA pF^-1 to 4.76 ± 0.98 pA pF^-1 in dome-forming cells and had no effect in cells outside of domes. ENaC but not DUOX1 transcription was potentiated by catalase in the presence of dexamethasone, which suggested negative feedback of H₂ O₂ on ENaC gene expression.

Our observations suggest that tonic production of H₂ O₂ by DUOX1 participates in maintaining the level of vectorial sodium transport by lung epithelial cells. Moreover, the system appears to be well tuned as it would allow H₂ O₂ -dependent innate immunity without inducing airway/alveolar sodium and fluid hyperabsorption.