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Your search keyword '"Sheppard, David"' showing total 26 results
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26 results on '"Sheppard, David"'

2. Experimental pharmacology in precision medicine

Author

Urbaniak, Alicja, primary, Thummel, Kenneth E., additional, Alade, Ayoade N., additional, Rettie, Allan E., additional, Prasad, Bhagwat, additional, De Nicolò, Amedeo, additional, Martin, Jennifer H., additional, Sheppard, David N., additional, and Jarvis, Michael F., additional

Published
2023
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4. Correlating genotype with phenotype using CFTR‐mediated whole‐cell Cl − currents in human nasal epithelial cells

Author

Noel, Sabrina, primary, Servel, Nathalie, additional, Hatton, Aurélie, additional, Golec, Anita, additional, Rodrat, Mayuree, additional, Ng, Demi R. S., additional, Li, Hongyu, additional, Pranke, Iwona, additional, Hinzpeter, Alexandre, additional, Edelman, Aleksander, additional, Sheppard, David N., additional, and Sermet‐Gaudelus, Isabelle, additional

Published
2021
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6. Partial rescue of F508del-cystic fibrosis transmembrane conductance regulator channel gating with modest improvement of protein processing, but not stability, by a dual-acting small molecule

Author

Liu, Jia, primary, Bihler, Hermann, additional, Farinha, Carlos M, additional, Awatade, Nikhil T, additional, Romão, Ana M, additional, Mercadante, Dayna, additional, Cheng, Yi, additional, Musisi, Isaac, additional, Jantarajit, Walailak, additional, Wang, Yiting, additional, Cai, Zhiwei, additional, Amaral, Margarida D, additional, Mense, Martin, additional, and Sheppard, David N, additional

Published
2018
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7. Impact of the F508del mutation on ovine CFTR, a Cl- channel with enhanced conductance and ATP-dependent gating

Author

Cai, Zhiwei, Palmai-Pallag, Timea, Khuituan, Pissared, Mutolo, Michael J, Boinot, Clément, Liu, Beihui, Scott-Ward, Toby S, Callebaut, Isabelle, Harris, Ann, and Sheppard, David N

Subjects
congenital, hereditary, and neonatal diseases and abnormalities, C700 Molecular Biology, Biophysics and Biochemistry, C130 Cell Biology, respiratory system, C440 Molecular Genetics, digestive system diseases, respiratory tract diseases
Abstract

Cross-species comparative studies are a powerful approach to understand the epithelial Cl- channel cystic fibrosis transmembrane conductance regulator (CFTR), which is defective in the genetic disease cystic fibrosis (CF). Here, we investigate the single-channel behaviour of ovine CFTR and the impact of the most common CF mutation, F508del-CFTR, using excised inside-out membrane patches from transiently transfected CHO cells. Like human CFTR, ovine CFTR formed a weakly inwardly rectifying Cl- channel regulated by PKA-dependent phosphorylation, inhibited by the open-channel blocker glibenclamide. However, for three reasons, ovine CFTR was noticeably more active than human CFTR. First, single-channel conductance was increased. Second, open probability was augmented because the frequency and duration of channel openings were increased. Third, with enhanced affinity and efficacy, ATP more strongly stimulated ovine CFTR channel gating. Consistent with these data, the CFTR modulator phloxine B failed to potentiate ovine CFTR Cl- currents. Like its impact on human CFTR, the F508del mutation caused a temperature-sensitive folding defect, which disrupted ovine CFTR protein processing and reduced membrane stability. However, the F508del mutation had reduced impact on ovine CFTR channel gating in contrast to its marked effects on human CFTR. We conclude that ovine CFTR forms a regulated Cl- channel with enhanced conductance and ATP-dependent channel gating. This phylogenetic analysis of CFTR structure and function demonstrates that subtle changes in structure have pronounced effects on channel function and the consequences of the CF mutation F508del. This article is protected by copyright. All rights reserved.

Published
2015

22. Impact of the F508del mutation on ovine CFTR, a Cl- channel with enhanced conductance and ATP-dependent gating

Author

Cai, Zhiwei, Palmai-Pallag, Timea, Khuituan, Pissared, Mutolo, Michael J, Boinot, Clément, Liu, Beihui, Scott-Ward, Toby S, Callebaut, Isabelle, Harris, Ann, Sheppard, David N, Cai, Zhiwei, Palmai-Pallag, Timea, Khuituan, Pissared, Mutolo, Michael J, Boinot, Clément, Liu, Beihui, Scott-Ward, Toby S, Callebaut, Isabelle, Harris, Ann, and Sheppard, David N

Abstract

Cross-species comparative studies are a powerful approach to understand the epithelial Cl- channel cystic fibrosis transmembrane conductance regulator (CFTR), which is defective in the genetic disease cystic fibrosis (CF). Here, we investigate the single-channel behaviour of ovine CFTR and the impact of the most common CF mutation, F508del-CFTR, using excised inside-out membrane patches from transiently transfected CHO cells. Like human CFTR, ovine CFTR formed a weakly inwardly rectifying Cl- channel regulated by PKA-dependent phosphorylation, inhibited by the open-channel blocker glibenclamide. However, for three reasons, ovine CFTR was noticeably more active than human CFTR. First, single-channel conductance was increased. Second, open probability was augmented because the frequency and duration of channel openings were increased. Third, with enhanced affinity and efficacy, ATP more strongly stimulated ovine CFTR channel gating. Consistent with these data, the CFTR modulator phloxine B failed to potentiate ovine CFTR Cl- currents. Like its impact on human CFTR, the F508del mutation caused a temperature-sensitive folding defect, which disrupted ovine CFTR protein processing and reduced membrane stability. However, the F508del mutation had reduced impact on ovine CFTR channel gating in contrast to its marked effects on human CFTR. We conclude that ovine CFTR forms a regulated Cl- channel with enhanced conductance and ATP-dependent channel gating. This phylogenetic analysis of CFTR structure and function demonstrates that subtle changes in structure have pronounced effects on channel function and the consequences of the CF mutation F508del. This article is protected by copyright. All rights reserved.

26. Loop diuretics are open-channel blockers of the cystic fibrosis transmembrane conductance regulator with distinct kinetics.

Author

Ju M, Scott-Ward TS, Liu J, Khuituan P, Li H, Cai Z, Husbands SM, and Sheppard DN

Subjects
Animals, Bumetanide pharmacology, Cricetinae, Cystic Fibrosis Transmembrane Conductance Regulator genetics, Cystic Fibrosis Transmembrane Conductance Regulator metabolism, Dose-Response Relationship, Drug, Furosemide pharmacology, Humans, Kinetics, Membrane Potentials, Mice, Molecular Structure, Rats, Sodium Potassium Chloride Symporter Inhibitors chemistry, Structure-Activity Relationship, Sulfonamides pharmacology, Xipamide pharmacology, Chlorides metabolism, Cystic Fibrosis Transmembrane Conductance Regulator antagonists & inhibitors, Sodium Potassium Chloride Symporter Inhibitors pharmacology
Abstract

Background and Purpose: Loop diuretics are widely used to inhibit the Na(+), K(+), 2Cl(-) co-transporter, but they also inhibit the cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel. Here, we investigated the mechanism of CFTR inhibition by loop diuretics and explored the effects of chemical structure on channel blockade., Experimental Approach: Using the patch-clamp technique, we tested the effects of bumetanide, furosemide, piretanide and xipamide on recombinant wild-type human CFTR., Key Results: When added to the intracellular solution, loop diuretics inhibited CFTR Cl(-) currents with potency approaching that of glibenclamide, a widely used CFTR blocker with some structural similarity to loop diuretics. To begin to study the kinetics of channel blockade, we examined the time dependence of macroscopic current inhibition following a hyperpolarizing voltage step. Like glibenclamide, piretanide blockade of CFTR was time and voltage dependent. By contrast, furosemide blockade was voltage dependent, but time independent. Consistent with these data, furosemide blocked individual CFTR Cl(-) channels with 'very fast' speed and drug-induced blocking events overlapped brief channel closures, whereas piretanide inhibited individual channels with 'intermediate' speed and drug-induced blocking events were distinct from channel closures., Conclusions and Implications: Structure-activity analysis of the loop diuretics suggests that the phenoxy group present in bumetanide and piretanide, but absent in furosemide and xipamide, might account for the different kinetics of channel block by locking loop diuretics within the intracellular vestibule of the CFTR pore. We conclude that loop diuretics are open-channel blockers of CFTR with distinct kinetics, affected by molecular dimensions and lipophilicity., (© 2013 The British Pharmacological Society.)

Published
2014
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