The amiloride-inhibitable epithelial Na⁺ channel (ENaC) is a critical component of the fluid reabsorption pathway in the lung and so contributes to the natural removal of lumenal fluid during birth and the maintenance of fluid homeostasis in the healthy postnatal lung (Barker and Olver. 2002). However, work by others has shown that this delicate balance is disrupted during lung inflammation through the nitric oxide (NO)-dependent inhibition of amiloride-sensitive Na⁺ transport (Ding et al 1999), an effect that is absent in mice lacking the gene for inducible nitric oxide synthase (iNOS) (Hardiman et al 2001). We have previously shown that exposure of H441 lung epithelial cells to moderate hypoxia (PO₂ 23 mmHg) significantly evokes the activation of the inducible isoform of nitric oxide synthase (iNOS) and release of NO (Laird et al 2002). Importantly, this effect was potentiated during hypoxia by the pro-inflammatory bacterial endotoxin, lipopolysaccharide (LPS). The aim of these studies was to determine if there is a link between hypoxia/LPS-evoked NO synthesis and the transcriptional regulation of αENaC in H441 bronchial epithelial cells.

Competitive reverse transcriptase-polymerase chain reaction (comp RT-PCR) found hypoxia (PO₂ 23 mmHg) activated αENaC gene expression 2.06e+07 ± 2.06+06 transcript copies (mean ± S.E.M.; n = 6) though remained unchanged from basal (PO₂ 142 mmHg) expression (1.42e+07 ± 2.26e+06; n = 6). Potent NOS activator, E. coli lipopolysaccharide (LPS 50 µg ml^-1) was associated with an inducible NO synthase (iNOS)-dependent increase in the lung production of NO but also resulted in unaltered αENaC gene expression at both 142mmHg and 23mmHg from control levels (1.63+07 ± 4.01+06; n = 5), (1.46+07 ± 3.23+06; n = 5). Inhibition of iNOS by the specific iNOS inhibitor, L-N⁶-(1-Iminoethyl)lysine hydrochloride (L-Nil), maintained normal αENaC gene expression at atmospheric and fetal PO₂ (1.25+07 ± 5.26+06; n = 5), (1.71+07 ± 5.06+06; n = 5). No significant differences were observed between groups.These data suggest that the inhibition of αENaC by hypoxia/LPS evoked NO production is not explained by a parallel change in gene expression.

This work was supported by the Wellcome Trust.