From idiopathic pulmonary hemosiderosis to ENaC channelopathy: a novel SCNN1G mutation

A 24-year-old woman presented with a 7-year history of recurrent hemoptysis, showed no response to immunosuppressants, and was initially misdiagnosed with idiopathic pulmonary hemosiderosis (IPH) based on hemosiderin-laden macrophages (Figure 1A, B). High-resolution computed tomography revealed acute alveolar hemorrhage without bronchiectasis (Figure 1C). Secondary causes, including autoimmune and cardiovascular etiologies, were excluded (Table S1).

Figure 1.

Cytological and radiological findings of the patient. (A) Low-magnification (100×) hematoxylin and eosin staining of the patient’s bronchoalveolar lavage fluid demonstrating numerous hemosiderin-laden macrophages scattered throughout the field. Scale bar = 100 μm. (B) High-magnification (400×) image showing characteristic intracytoplasmic golden-brown hemosiderin granules (arrows). Scale bar = 25 μm. (C) High-resolution computed tomography (HRCT) at presentation, demonstrating diffuse bilateral ground-glass opacities and focal consolidation in the right lower lobe, indicating acute alveolar hemorrhage. (D) Follow-up HRCT after airway-directed therapy, showing significant resolution of the pulmonary lesions and infiltrates.

Whole-exome sequencing identified a novel heterozygous variant (c.1126G > A; p.Gly376Arg) in SCNN1G, which encodes the γ subunit of the epithelial sodium channel (ENaC) (Figure 2A), and this was confirmed by Sanger sequencing (Figure 2B). Following confirmation of mutant γ-ENaC expression (Figure 2C) and complex assembly via α/β-subunit co-transfection (Figure 2D), patch-clamp electrophysiology revealed a gain-of-function phenotype with amplified amiloride-sensitive sodium currents (Figure 2E).

Figure 2.

Genetic and functional validation of the SCNN1G c.1126G > A (p.Gly376Arg) variant. (A) Integrative Genomics Viewer snapshot of whole-exome sequencing data, identifying a heterozygous missense mutation (c.1126G > A, p.Gly376Arg) in the SCNN1G gene (NM_001039.4). (B) Sanger sequencing chromatogram confirming the heterozygous c.1126G > A variant (shaded gray box). (C). Western blot analysis of HEK293T cell lines (293T/NC, 293T/SCNN1G wild-type [wt], and 293T/SCNN1G mutant [mut]). Both wild-type and mutant γ-epithelial sodium channel (ENaC) proteins showed robust and comparable expression at approximately 75 kDa, with GAPDH as the loading control. (D) Immunofluorescence microscopy confirming the co-expression and proper membrane localization of α-ENaC (green) and β-ENaC (red) subunits in the γ-ENaC established cell lines. (E) Representative whole-cell patch-clamp electrophysiology traces. In cells co-expressing α/β subunits with mutant γ-ENaC (G376R), the amiloride-sensitive sodium current (activated by chymotrypsin and inhibited by amiloride) was significantly increased compared to the wild-type group, confirming a gain-of-function effect.

ENaC regulates airway surface liquid volume and mucociliary clearance.¹ We hypothesize a novel channelopathy wherein hyperactive ENaC might drive fluid absorption and mucociliary impairment.^2,3 Though requiring further validation, the ensuing chronic inflammation could injure the alveolar-capillary membrane, leading to recurrent alveolar hemorrhage.

Following the initiation of an exploratory regimen aimed at this mechanism—inhaled hypertonic saline (3%) targeting airway hydration, alongside oral eucalyptol and azithromycin intended to inhibit ENaC⁴—we observed a reduction in hemoptysis frequency from 41% to 17.2% over 6 months, coinciding with radiographic resolution (Figure 1D).

This case proposes a potential link between ENaC gain-of-function and alveolar hemorrhage, suggesting genetic evaluation for refractory IPH. While a temporal clinical improvement was noted, in vivo models remain essential to confirm if ENaC hyperactivity directly mediates capillary injury. Furthermore, ENaC’s role in alveolar type 2 cell lineage⁵ highlights its underexplored function in distal alveolar homeostasis.

Author contributions

Q.P. and X.L. were involved in study conception and genetic analysis. Z.J., F.Z., and X.Z. performed the experimental studies. Y.C. and Y.L. contributed to clinical data acquisition and patient management. L.Y. and X.H. performed pathological examination and interpretation. W.L. provided and interpreted radiological images. L.W. assisted in electrophysiological experiments and data analysis. B.Z. and X.L. supervised the project and critically revised the manuscript. All authors reviewed and approved the final version. B.Z. and X.L. are the guarantors.

Supplementary material

Supplementary material is available at American Journal of Respiratory and Critical Care Medicine online.

Conflicts of interest

Please see the ICMJE disclosure forms, which have been provided as supplementary material. None declared.

Funding

This work was supported by the Guangdong Basic and Applied Basic Research Fund (Guangdong-Dongguan Joint Fund) (2024A1515140175), Dongguan Eighth People’s Hospital doctoral research start-up fund project (DBBS2023003).