Proceedings of The Physiological Society
University of Oxford (2011) Proc Physiol Soc 23, C64
Membrane expression of deltaF508 mutant increases epithelial HCO3- secretion via functional Cl-/HCO3- exchanger(s)
F. Xiao1, J. Li1,2, A. Sultan1, J. Wang1, B. Riederer1, R. Engelhardt1, B. Rausch1, G. Lamprecht3, H. De Jonge4, B. Scholte5, U. Seidler1
1. Dept. of Gastoenterology, Hepatology & Endocrinology, Hannover Medical School, Germany, Hannover, Germany. 2. Dept. of Nephrology,, Tongji Hospital, Huazhong University, China, Wuhan, China. 3. 1st. Dept. of Medicine, University of T
Background: The CF-associated HCO3- secretory defect interferes with mucus clearance, nutrient absorption and fertility(1). “Corrector” drug therapy allows the most frequent mutation in CF, the delF508 mutant, to partially escape proteasom degradation and reach the apical membrane. But does this restore the HCO3- secretory defect? Aims: To find out whether, and by what molecular mechanism, delF508 membrane expression can promote epithelial HCO3- secretion in mouse intestine. Method and results: Basal and forskolin (FSK) stimulated duodenal HCO3- secretion was studied in vivo in luminally perfused (duodenum and colon) mice, which were anesthetized by spontaneous inhalation of isoflurane (a mixture of 2.2±0.2% isoflurane in ～30-40% oxygen, ～60-70% nitrogen) and were kept in physiological conditions following standard experimental procedure as previously described(2), as well as in vitro in isolated duodenal and colonic mucosa of CFTR KO(2), delF508 mutant(3), and WT littermates, congenic on FVB/N background. Western blot analysis was performed in isolated brush border membranes (BBM) and demonstrated approx 5%, and 0% glycosylated band C CFTR in the intestinal BBM of delF508, and CFTR KO, respectively, compared to WT BBM. The FSK-induced, CFTR dependent Isc response in the delF508 mutant intestinal mucosa was also approx. 5-10% of that in WT mucosa, whereas the CFTR-dependent HCO3- secretory response in delF508 was almost half of the WT response. Similar results were obtained in vivo. This markedly higher HCO3- than Isc response in the delF508 intestine was completely dependent on the presence of luminal Cl- both in the duodenal as well as colonic mucosa and can be inhibited by CFTR inhibitors Cftr(inh)-172(4) (20µM) together with GlyH101(5) (10µM). Conclusion: The data suggest that even a very low Cl- membrane conductance significantly enhances apical Cl-/HCO3- exchange. This suggests that delF508 “corrector” therapy has the potential to significantly enhance epithelial HCO3- secretion.
Where applicable, experiments conform with Society ethical requirements