Polycystic kidney disease is characterised by massive enlargement of fluid-filled epithelial cysts that involves cAMP-dependent cell proliferation and fluid secretion driven by the cystic fibrosis transmembrane conductance regulator (CFTR) Cl^– channel (Sullivan et al. 1998). Using MDCK cysts, we previously demonstrated that CFTR blockers inhibit cyst growth (Findlay et al. 2001). In this study, we investigated the effects of CFTR blockers on cell proliferation and cAMP-stimulated Cl^– currents to understand how drugs inhibit the growth of MDCK cysts.

We grew cysts as described previously (Findlay et al. 2001) with the exception that the effects of drugs were tested between days 6 and 12. For cell proliferation assays, MDCK cells were grown in media containing 0.01 % FBS and cAMP agonists in the absence and presence of drugs over a 6 day period and cell numbers determined using a haemocytometer. Using MDCK epithelia and the Ussing chamber technique, we quantified the effects of drugs on cAMP-stimulated Cl^– currents. We tested the effects of four drugs that inhibit CFTR including the open-channel blockers glibenclamide (100 mM) and NPPB (50 mM) and the allosteric blocker genistein (100 mM). As controls, we used first, tamoxifen (10 mM), calix[4]arene (10 mM) and DIDS (200 mM), which inhibit other Cl^– channels but not CFTR, and second, TEA (10 mM), bumetanide (100 mM) and ouabain (1 mM), which block basolateral membrane channels and transporters in MDCK cells.

There were three sets of results in this study. First, CFTR inhibitors, bumetanide and ouabain reduced dramatically the volume of cysts (n = 20-36; P < 0.05; one-way ANOVA), whereas TEA, calix[4]arene and DIDS were without effect on cyst growth (n = 31-39; P > 0.05). Second, with the exception of bumetanide, all the drugs tested inhibited potently cell proliferation (n = 4-10; P < 0.0.1). Third, CFTR inhibitors, TEA, bumetanide and ouabain all inhibited cAMP-stimulated Cl^– currents (n = 5-7; P < 0.01; Student’s paired t test), whereas calix[4]arene, DIDS and tamoxifen were without effect. To analyse further these data, we compared the magnitude of inhibition of cyst growth, cell proliferation and cAMP-stimulated Cl^– currents by different drugs. Inhibition of cyst growth and cAMP-stimulated Cl^– currents were well correlated (correlation coefficient = 0.71; P < 0.01; Pearson’s correlation test), whereas inhibition of cyst growth and cell proliferation was not well correlated (correlation coefficient = 0.40; P > 0.05; Pearson’s correlation test). We interpret these data to suggest that CFTR blockers probably inhibit cyst growth by preventing fluid accumulation within the cyst lumen.

This work was supported by the NKRF and Royal Society.