کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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6197609 | 1261193 | 2008 | 7 صفحه PDF | دانلود رایگان |
The cystic fibrosis transmembrane conductance regulator (CFTR) is present on the apical membrane of corneal endothelial cells. Increasing intracellular [cAMP] with forskolin stimulates an NPPB and glibenclamide-inhibitable apical Clâ and HCO3â permeability [Sun, X.C., Bonanno, J.A., 2002. Expression, localization, and functional evaluation of CFTR in bovine corneal endothelial cells. Am. J. Physiol. Cell Physiol. 282, C673-C683]. To definitively determine that the increased permeability is dependent on CFTR, we used an siRNA knockdown approach. Apical Clâ and HCO3â permeability and steady-state HCO3â flux were measured in the presence or absence of forskolin using cultured bovine corneal endothelial cells that were transfected with CFTR siRNA or a scrambled sequence control. CFTR protein expression was reduced by â¼80% in CFTR siRNA treated cultures. Forskolin (10 μM) increased apical chloride permeability by 7-fold, which was reduced to control level in siRNA treated cells. CFTR siRNA treatment had no effect on baseline apical chloride permeability. Apical HCO3â permeability was increased 2-fold by 10 μM forskolin, which was reduced to control level in siRNA treated cultures. Similarly, there was no effect on baseline apical HCO3â permeability by knocking down CFTR expression. The steady-state apical-basolateral pH gradient (ÎpH) at 4 h in control cultures was increased â¼2.5-fold by forskolin. In CFTR siRNA treated cells, the baseline ÎpH was similar to control, however forskolin did not have a significant effect. We conclude that forskolin induced increases in apical HCO3â permeability in bovine corneal endothelium requires CFTR. However, CFTR does not have a major role in determining baseline apical chloride or HCO3â permeability.
Journal: Experimental Eye Research - Volume 86, Issue 4, April 2008, Pages 684-690