6 research outputs found
Lubiprostone Stimulates Duodenal Bicarbonate Secretion in Rats
Lubiprostone, a bicyclic fatty acid, is used for the treatment of chronic constipation. No published study has addressed the effect of lubiprostone on intestinal ion secretion in vivo.
The aim of this study was to test the hypothesis that lubiprostone augments duodenal HCO3
â secretion (DBS).
Rat proximal duodenal loops were perfused with pH 7.0 Krebs, control vehicle (medium-chain triglycerides), or lubiprostone (0.1â10Â ÎŒM). We measured DBS with flow-through pH and CO2 electrodes, perfusate [Clâ] with a Clâ electrode, and water flux using a non-absorbable ferrocyanide marker. Some rats were pretreated with a potent, selective CFTR antagonist, CFTRinh-172 (1Â mg/kg, ip), 1Â h before experiments.
Perfusion of lubiprostone concentration dependently increased DBS, whereas net Clâ output and net water output were only increased at 0.1Â ÎŒM, compared with vehicle. CFTRinh-172 reduced lubiprostone (10Â ÎŒM)-induced DBS increase, whereas net Clâ output was also unchanged. Nevertheless, CFTRinh-172 reduced basal net water output, which was reversed by lubiprostone. Furthermore, lubiprostone-induced DBS was inhibited by EP4 receptor antagonist, not by an EP1/2 receptor antagonist or by indomethacin pretreatment.
In this first study of the effect of lubiprostone on intestinal ion secretion in vivo, lubiprostone stimulated CFTR-dependent DBS without changing net Clâ secretion. This effect supports the hypothesis that Clâ secreted by CFTR is recycled across the apical membrane by anion exchangers. Recovery of water output during CFTR inhibition suggests that lubiprostone may improve the intestinal phenotype in CF patients. Furthermore, increased DBS suggests that lubiprostone may protect the duodenum from acid-induced injury via EP4 receptor activation
Expression of the chloride channel CLC-K in human airway epithelial cells
Airway submucosal gland function is severely disrupted in cystic fibrosis (CF), as a result of genetic mutation
of the cystic fibrosis transmembrane conductance regulator (CFTR), an apical membrane Clâ channel. To identify other
Clâ channel types that could potentially substitute for lost CFTR function in these cells, we investigated the functional and
molecular expression of Clâ channels in Calu-3 cells, a human cell line model of the submucosal gland serous cell. Whole
cell patch clamp recording from these cells identified outwardly rectified, pH- and calcium-sensitive Clâ currents that resemble
those previously ascribed to ClC-K type chloride channels. Using reverse transcription â polymerase chain reaction,
we identified expression of mRNA for ClC-2, ClC-3, ClC-4, ClC-5, ClC-6, ClC-7, ClC-Ka, and ClC-Kb, as well as the
common ClC-K channel b subunit barttin. Western blotting confirmed that Calu-3 cells express both ClC-K and barttin
protein. Thus, Calu-3 cells express multiple members of the ClC family of Clâ channels that, if also expressed in native
submucosal gland serous cells within the CF lung, could perhaps act to partially substitute lost CFTR function. Furthermore,
this work represents the first evidence for functional ClC-K chloride channel expression within the lung
Incomplete rescue of cystic fibrosis transmembrane conductance regulator deficient mice by the human CFTR cDNA
We have used a mouse model to study the ability of human CFTR to correct the defect in mice deficient of the endogenous protein. In this model, expression of the endogenous Cftr gene was disrupted and replaced with a human CFTR cDNA by a gene targeted 'knock-in' event. Animals homozygous for the gene replacement failed to show neither improved intestinal pathology nor survival when compared to mice completely lacking CFTR. RNA analyses showed that the human CFTR sequence was transcribed from the targeted allele in the respiratory and intestinal epithelial cells. Furthermore, in vivo potential difference measurements showed that basal CFTR chloride channel activity was present in the apical membranes of both nasal and rectal epithelial cells in all homozygous knock-in animals examined. Ussing chamber studies showed, however, that the cAMP-mediated chloride channel function was impaired in the intestinal tract among the majority of homozygous knock-in animals. Hence, failure to correct the intestinal pathology associated with loss of endogenous CFTR was related to inefficient functional expression of the human protein in mice. These results emphasize the need to understand the tissue-specific expression and regulation of CFTR function when animal models are used in gene therapy studies.link_to_OA_fulltex