Most bicarbonate secretion by Calu-3 cells is mediated by CFTR and independent of pendrin

Abstract

Bicarbonate plays an important role in airway host defense, however, its transport mechanisms remain uncertain. Here we examined the relative contributions of the anion channel CFTR (cystic fibrosis transmembrane conductance regulator, ABCC7) and the anion exchanger pendrin (SLC26A4) to HCO 3 − secretion by the human airway cell line Calu‐3. Pendrin and CFTR were both detected in parental Calu‐3 cells, although mRNA and protein expression appeared higher for CFTR than for pendrin. Targeting pendrin transcripts with lentiviral shRNA reduced pendrin detection by immunofluorescence staining but did not alter the rates of HCO 3 − or fluid secretion, HCO 3 − transport under pH‐stat conditions, or net HCO 3 − flux across basolaterally permeabilized monolayers. Intracellular pH varied with step changes in apical Cl− and HCO 3 − concentrations in control and pendrin knockdown Calu‐3 cells, but not in CFTR deficient cells. Exposure to the proinflammatory cytokine IL‐4, which strongly upregulates pendrin expression in airway surface epithelia, had little effect on Calu‐3 pendrin expression and did not alter fluid or HCO 3 − secretion. Similar results were obtained using air–liquid interface and submerged cultures, although CFTR and pendrin mRNA expression were both lower when cells were cultured under submerged conditions. While the conclusions cannot be extrapolated to other airway epithelia, the present results demonstrate that most HCO 3 − secretion by Calu‐3 cells is mediated by CFTR.

Inverse regulation of secretion and inflammation in human airway gland serous cells by neuropeptides upregulated in allergy and asthma

“…These data suggest that VIP receptor activation or direct cAMP elevation with forskolin can activate both Cland HCO3secretion directly through CFTR. We found no evidence for Cl -/HCO3exchanger-mediated HCO3efflux in primary serous cells (Supplemental Figure 2), suggesting CFTR is the main HCO3efflux pathway during cAMP stimulation, agreeing with recent Calu-3 studies suggesting CFTR sustains HCO3secretion (88,89) instead of the pendrin Cl -/HCO3exchanger (90,91).…”

cAMP stimulates SLC26A3 activity in human colon by a CFTR-dependent mechanism that does not require CFTR activity

“…Studies in the widely studied human airway cell line Calu-3 examined mechanisms of cAMP-stimulated HCO 3 - secretion. 36 These cells express a large amount of CFTR and much less SLC26A4 (pendrin). Forskolin-stimulated HCO 3 - secretion in Calu-3 cells was entirely CFTR-dependent and not affected by SLC26A4 knockdown, identifying an additional model of cAMP-stimulated HCO 3 - secretion in cells that contain both CFTR and members of the SLC26 family.…”

“…In HEK293/DRA cells and oocytes, there was no cAMP effect unless CFTR was also expressed, which led to modest stimulation. 9,10 In Caco-2 cells, cAMP inhibited DRA activity using 36 Cl to measure unidirectional fluxes, which was accompanied by less brush border DRA. 11 In murine duodenal brush border vesicle studies, cAMP increased Cl - /HCO 3 - exchange.…”

Extracellular pH and lung infections in cystic fibrosis

“…This secreted HCO 3 − appears to be generated by carbonic anhydrase located near the apical membrane (Shan et al, 2012). The same laboratory then reported that HCO 3 − secretion in Calu-3 cells is independent of pendrin (Huang et al, 2018), while other authors reported that CFTR acts together with pendrin channel (SLC26A4) to secrete HCO 3 − , in the same Calu-3 cells (Garnett et al, 2011). Thus, this issue is also a controversial matter.…”