The effects of hypercapnia on CFTR-dependent HCOb3p-s secretion in human airway epithelia

Abstract

Hypercapnia is clinically defined as an arterial blood partial pressure of CO2 of above 40mm Hg and is a symptom of chronic lung disease. In renal epithelia, hypercapnia can reduce agonist-stimulated cAMP levels and impair regulation of cAMP-dependent ion transporters. In the airways, elevations in intracellular cAMP in serous cells of the submucosal glands, activates CFTR-mediated HCO3- and fluid secretion, which contributes to airway surface liquid homeostasis. The aim of the current work was to investigate the effects of both acute and chronic hypercapnia on cAMP-regulated ion and fluid transport in Calu-3 cells, a model of human serous cells. Acute hypercapnia significantly reduced both forskolin-stimulated elevations in intracellular cAMP and forskolin-stimulated increase in short-circuit current, suggesting CO2 reduced cAMP-regulated, CFTR-dependent anion secretion. Stimulation of Calu-3 cells with cAMP agonists induced a reversible intracellular acidification that was a result of CFTR-dependent HCO3- secretion. In acute hypercapnia, this intracellular acidification was significantly augmented yet neither CFTR-dependent HCO3- efflux, nor NBC-dependent HCO3- influx, were found to be CO2-sensitive. However, ouabain blocked the augmentation induced by hypercapnia implicating the Na+/K+-ATPase in mediating the effects of raised CO2 on cAMP-mediated cytosolic acidification. In addition, both BAPTA-AM and the phospholipase C inhibitor, U77312, also blocked the effect of hypercapnia, implying a role for PLC-regulated Ca2+ mobilization in the pH response to hypercapnia. Although addition of exogenous ATP in normocapnia mimicked the effect of hypercapnia, there was little evidence that CO2-induced ATP release from Calu-3 cells, which therefore suggests the effect of hypercapnia is not due to ATP/Ca2+ signalling via purinergic receptors. Exposure of Calu-3 cells to 24 hours hypercapnia caused a significant reduction in the volume of fluid secreted but did not affect the HCO3- or mucus content of this secreted fluid. These data suggest that transporters involved in regulating the volume of secreted fluid have differential CO2 sensitivity compared to transporters involved in regulating its composition. These findings reveal that both acute and chronic hypercapnia affected ion and fluid transport in human airway epithelial cells, and suggests hypercapnia could have pronounced effects on the properties of the airway surface liquid which would be predicted to have important consequences for the innate defence mechanisms of the lungs.