Cathepsin B contributes to Na⁺ hyperabsorption in cystic fibrosis airway epithelial cultures

Abstract

Key pointsr The epithelial Na + channel (ENaC) is hyperactive in cystic fibrosis (CF) airway epithelia, and contributes to excessive Na + absorption and dehydration of the airway surface liquid (ASL) that lines the lungs -a major cause of CF lung disease.r The CF gene product (CFTR) is a cAMP-regulated anion channel and how CFTR dysfunction contributes to ENaC hyperactivity is poorly understood. Of note, ENaC must be proteolytically cleaved to be active.r CF ASL is moderately acidic, due to the absence of HCO 3 − secretion through CFTR. The acid protease cathepsin B is present in the apical membrane of airway epithelia and is secreted into the ASL, suggesting that this protease may be active in the acidic CF environment.r We have found that cathepsin B induces activation of ENaC in both Xenopus laevis oocytes and in airway epithelia. Cathepsin B is also highly expressed in airway epithelia derived from patients with CF.r Inhibition of cathepsin B prevents Na + /ASL hyperabsorption in CF airway cultures, suggesting a hitherto unrecognized role for this protease in CF pathogenesis. Treatments directed at normalizing CF ASL pH or inhibiting cathepsin B may be useful in the treatment of CF lung disease.Abstract In cystic fibrosis (CF) lung disease, the absence of functional CF transmembrane conductance regulator results in Cl − /HCO 3 − hyposecretion and triggers Na + hyperabsorption through the epithelial Na + channel (ENaC), which contribute to reduced airway surface liquid (ASL) pH and volume. Prostasin, a membrane-anchored serine protease with trypsin-like substrate specificity has previously been shown to activate ENaC in CF airways. However, prostasin is typically inactive below pH 7.0, suggesting that it may be less relevant in acidic CF airways. Cathepsin B (CTSB) is present in both normal and CF epithelia and is secreted into ASL, but little is known about its function in the airways. We hypothesized that the acidic ASL seen in CF airways may stimulate CTSB to activate ENaC, contributing to Na + hyperabsorption and depletion of CF ASL volume. In Xenopus laevis oocytes, CTSB triggered α-and γENaC cleavage and induced an increase in ENaC activity. In bronchial epithelia from both normal and CF donor lungs, CTSB localized to the apical membrane. In normal and CF human bronchial epithelial cultures, CTSB was detected at the apical plasma membrane and in the ASL. CTSB activity was significantly elevated in acidic ASL, which correlated with increased abundance of ENaC in the plasma membrane and a reduction in ASL volume. This acid/CTSB-dependent activation of ENaC was ameliorated with the cell impermeable, CTSB-selective inhibitor CA074, suggesting that CTSB inhibition may have therapeutic relevance. Taken together, our data suggest that CTSB is a pathophysiologically relevant protease that activates ENaC in CF airways.

Cathepsin B increases ENaC activity leading to hypertension early in nephrotic syndrome

“…Using trypsin as positive control, we observed a very mild processing by cathepsin D and legumain by in vitro protease assays. Although Tan et al reported that cathepsin B cleaves αENaC and γENaC, we did not observe γENaC processing which may be due to differences in techniques and models used. The very faint cleavage of γENaC by cathepsin D at 70 kDa and legumain at 72 kDa corresponds to the increase in relative currents in mpkCCD cl14 cells at 2 hours.…”

Inhibition of Protease–Epithelial Sodium Channel Signaling Improves Mucociliary Function in Cystic Fibrosis Airways

“…3A). Although not all cleavage products are functionally relevant our observation is consistent with another recent study, also conducted using normal and CF primary human bronchial epithelial cells, wherein processed ENaCγ is detected at the same molecular weight, and moreover shown to be modulated by cathepsin B activity [47]. As such additional studies are warranted to fully understand the regulation and functional role of this pool of ENaCγ in CF airways.…”

Sodium Retention and Volume Expansion in Nephrotic Syndrome: Implications for Hypertension

“…This group of channel activating proteases includes prostasin 21,24,47 , transmembrane protease serine 4 (TMPRSS4) 24,48,49 , matriptase 24,50 , cathepsin B 28,51 , elastase 25,26,52 , kallikrein 27,53 and plasmin 31,32 .…”