Oxidant and antioxidant modulation of chloride channels expressed in human retinal pigment epithelium.
Retinal pigment epithelium (RPE) possesses regulated chloride channels that are crucial for transepithelial fluid and ion transport. At present, little is known about the molecular nature of chloride channels in human adult RPE (haRPE) or the effects of oxidative stress on membrane conductance properties. In the present study, we assessed ClC channel and cystic fibrosis transmembrane conductance regulator (CFTR) expression and membrane chloride conductance properties in haRPE cells. ClC-5, ClC-3, ClC-2, and CFTR mRNA expression was confirmed with RT-PCR analysis, and protein expression was detected with Western blot analysis and immunofluorescence microscopy. Whole cell recordings of primary cultures of haRPE showed an outwardly rectifying chloride current that was inhibited by the oxidant H(2)O(2). The inhibitory effects of H(2)O(2) were reduced in cultured human RPE cells that were incubated with precursors of glutathione synthesis or that were stably transfected to overexpress glutathione S-transferase. These findings indicate a possible role for ClC channels in haRPE cells and suggest possible redox modulation of human RPE chloride conductances.
Weng
T X
TX
Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston 77555, USA.
Godley
B F
BF
Jin
G F
GF
Mangini
N J
NJ
Kennedy
B G
BG
Yu
A S L
AS
Wills
N K
NK
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000368:Aged; D000975:Antioxidants / Q000494:pharmacology*; D000090007:CLC-2 Chloride Channels; D018118:Chloride Channels / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D000242:Cyclic AMP / Q000378:metabolism / Q000494:pharmacology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000187:drug effects / Q000235:genetics / Q000378:metabolism; D005978:Glutathione / Q000494:pharmacology; D005982:Glutathione Transferase / Q000096:biosynthesis / Q000235:genetics; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects; D016877:Oxidants / Q000494:pharmacology*; D018408:Patch-Clamp Techniques; D010857:Pigment Epithelium of Eye / Q000166:cytology / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012604:Scorpion Venoms / Q000494:pharmacology
D000975:Antioxidants; C505032:CFTR protein, human; D000090007:CLC-2 Chloride Channels; C097503:CLC-5 chloride channel; D018118:Chloride Channels; C115376:ClC-3 channel; D016877:Oxidants; D012333:RNA, Messenger; D012604:Scorpion Venoms; C091539:Chlorotoxin; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D006861:Hydrogen Peroxide; D000242:Cyclic AMP; D005982:Glutathione Transferase; D005978:Glutathione; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid