Extracellular zinc and ATP restore chloride secretion across cystic fibrosis airway epithelia by triggering calcium entry.
Cystic fibrosis (CF) is caused by defective cyclic AMP-dependent cystic fibrosis transmembrane conductance regulator Cl(-) channels. Thus, CF epithelia fail to transport Cl(-) and water. A postulated therapeutic avenue in CF is activation of alternative Ca(2+)-dependent Cl(-) channels. We hypothesized that stimulation of Ca(2+) entry from the extracellular space could trigger a sustained Ca(2+) signal to activate Ca(2+)-dependent Cl(-) channels. Cytosolic [Ca(2+)](i) was measured in non-polarized human CF (IB3-1) and non-CF (16HBE14o(-)) airway epithelial cells. Primary human CF and non-CF airway epithelial monolayers as well as Calu-3 monolayers were used to assess anion secretion. In vivo nasal potential difference measurements were performed in non-CF and two different CF mouse (DeltaF508 homozygous and bitransgenic gut-corrected but lung-null) models. Zinc and ATP induced a sustained, reversible, and reproducible increase in cytosolic Ca(2+) in CF and non-CF cells with chemistry and pharmacology most consistent with activation of P2X purinergic receptor channels. P2X purinergic receptor channel-mediated Ca(2+) entry stimulated sustained Cl(-) and HCO(3)(-) secretion in CF and non-CF epithelial monolayers. In non-CF mice, zinc and ATP induced a significant Cl(-) secretory response similar to the effects of agonists that increase intracellular cAMP levels. More importantly, in both CF mouse models, Cl(-) permeability of nasal epithelia was restored in a sustained manner by zinc and ATP. These effects were reversible and reacquirable upon removal and readdition of agonists. Our data suggest that activation of P2X calcium entry channels may have profound therapeutic benefit for CF that is independent of cystic fibrosis transmembrane conductance regulator genotype.
Zsembery
Akos
A
Department of Physiology and Biophysics, Gregory Fleming James Cystic Fibrosis Research Center, University of Alabama, Birmingham 35294-0005, USA. zsembery@physiology.uab.edu
Fortenberry
James A
JA
Liang
Lihua
L
Bebok
Zsuzsa
Z
Tucker
Torry A
TA
Boyce
Amanda T
AT
Braunstein
Gavin M
GM
Welty
Elisabeth
E
Bell
P Darwin
PD
Sorscher
Eric J
EJ
Clancy
J P
JP
Schwiebert
Erik M
EM
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D000838:Anions; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D002712:Chlorides / Q000378:metabolism*; D002713:Chlorine / Q000378:metabolism; D004396:Coloring Agents / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D003550:Cystic Fibrosis / Q000188:drug therapy / Q000378:metabolism*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics; D003600:Cytosol / Q000378:metabolism; D004847:Epithelial Cells / Q000378:metabolism*; D016257:Fura-2 / Q000494:pharmacology; D005838:Genotype; D006720:Homozygote; D006801:Humans; D006863:Hydrogen-Ion Concentration; D051379:Mice; D009154:Mutation; D018048:Receptors, Purinergic P2 / Q000378:metabolism; D013997:Time Factors; D014132:Trachea / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry / Q000494:pharmacology*
D000838:Anions; C505032:CFTR protein, human; D002712:Chlorides; D004396:Coloring Agents; D018048:Receptors, Purinergic P2; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D002713:Chlorine; D000255:Adenosine Triphosphate; D000242:Cyclic AMP; D015032:Zinc; D002118:Calcium; D016257:Fura-2