The role of α-, β-, and γ-ENaC subunits in distal lung epithelial fluid absorption induced by pulmonary edema fluid

Abstract

[537][538][539][540][541][542][543][544][545][546][547][548] 2002]. We now show that EF increases fluid absorption across monolayers of rat FDLE in a dose-dependent manner. To study the role of subunits of the epithelial Na ϩ channel (ENaC) in the phenomena, we cultured explants from the distal lungs of 16-day gestational age wild-type (WT) or ␣-, ␤-, or ␥-ENaC knockout or heterozygote (HT) mice. WT explants cultured in media continuously expanded over time as a result of net fluid secretion. In contrast, when explants were exposed to EF for 24 h, net fluid absorption occurred. EF-exposed explants had normal histology, but marked changes were seen after Triton X-100 or staurosporine exposure. Transmission electron microscopy showed EF promoted lamellar body formation and abundant surfactant in the explants' lumens. EF-induced changes in explant size were similar in ␣-ENaC knockout, WT, and HT littermate fetal lung explants (P Ͼ 0.05). In contrast, EF's effect was attenuated in ␤-and ␥-ENaC knockouts (P Ͻ 0.05) vs. WT and HT littermate fetal lung explants. EF exposure slightly decreased or had no effect on mRNA levels for ␣-ENaC in various mouse genotypes but decreased expression of ␤-and ␥-ENaC subunit mRNAs (P Ͻ 0.01) across all genotype groups. We conclude that ␤-and ␥-, but not ␣-, ENaC subunits are essential for EF to exert its maximal effect on net fluid absorption by distal lung epithelia. epithelium; Na ϩ transport; lung development; amiloride-sensitive epithelial Na ϩ channel PULMONARY EDEMA can mainly occur as a result of one or both of the following mechanisms: increased transvascular pressure in the pulmonary microvasculature, as occurs in congestive heart failure, or increased permeability of the alveolar-capillary membrane to solutes, as occurs in adult respiratory distress syndrome. The ability of the lungs to clear this air space fluid has important clinical implications. For example, others have demonstrated that the active absorption of air space fluid in adults with pulmonary edema correlates with improved survival (20). Air space fluid clearance is actively driven by epithelial Na ϩ transport, which crosses the apical membrane and is then extruded across the basolateral membrane by Na ATPase (18, 19). The rate-limiting step in active Na ϩ transport by epithelia is the number and activity of the apical membrane Na ϩ channels. One of these Na ϩ channels is the amiloridesensitive epithelial Na ϩ channel (ENaC) that is composed of ␣-, ␤-, and ␥-subunits (6). However, it has been shown that normal mammalian lung epithelia have both an amiloridesensitive and an amiloride-insensitive component to their active Na ϩ transport both in vivo (24) and in vitro (8, 25). Presently, the molecular basis of this amiloride-insensitive Na ϩ transport is poorly understood. Regardless, if one could determine mechanisms underlying the regulation of this amiloride-sensitive and -insensitive Na ϩ transport, one could in the future identify novel approaches for the therapy of patients with pulmonary edema.It has been...

Deletion of the ubiquitin ligase Nedd4L in lung epithelia causes cystic fibrosis-like disease

“…Pups were sacrificed by cervical dislocation, lungs inflated, fixed, sectioned (5 μm), and processed for H&E, PAS, or collagen (Trichrome) staining, as detailed in SI Text. EM analysis of trachea and some bronchi were processed as described (29).…”

“…Wet∕Dry weight ratio of lungs from 4-h-old newborns was carried out as described (29). Lung explant growth assay was performed on explants from E16 embryos in the presence/absence of amiloride (10 μM), as described (29) and detailed in SI Text.…”

“…4A, lungs of newborn Nedd4L f ∕f ;SPCrtTA;tet o -Cre mice were drier than controls (Nedd4L f ∕f ;SPC-rtTA or Nedd4L f ∕f ;tet o -Cre), suggesting that the knockout mice are better able to absorb lung fluid at birth. Next, we tested fluid absorption in distal lung explants harvested from E16 embryos and grown in culture, as described previously (29). Fig.…”

Pulmonary Neuroendocrine Cell-Secreted Factors May Alter Fetal Lung Liquid Clearance

“…Fetal distal lung epithelium (FDLE) were isolated and grown in submersion culture on porous Transwell filters, as previously described (21,22). Cells prepared using similar methods mainly contain fetal type II epithelial cells (23); however, because precursor type II cells or airways cells may also be present, we refer to these cells as FDLE.…”

Expression and function of the epithelial sodium channel δ-subunit in human respiratory epithelial cells in vitro

“…The roles of β-and γ-ENaC subunits in the alveoli appear less critical. Mice lacking β-or γ-ENaC were able to clear their lungs at birth but died from abnormal renal electrolyte re-absorption [4,19,36].…”