Aquaporin-1 channels in human retinal pigment epithelium: role in transepithelial water movement.
OBJECTIVE
Aquaporin (AQP) is a hexahelical integral membrane protein that functions as a constitutive channel for water and regulated channel for cations in fluid transporting tissues, including many in the eye. Although AQP1 has been cloned from a cDNA library prepared from cultures of retinal pigment epithelial (RPE) cells isolated from human fetal tissue, three separate studies failed with various immunochemical techniques to detect AQP1 protein in adult human or rat RPE preparations. The purpose of this study was to examine specifically the expression and distribution of AQP1 in adult human RPE in situ by using alternative methodologies and model systems and to determine the contribution of AQP1 to water movement across cultured RPE cells isolated from human cadaveric and fetal eyes.


METHODS
AQP1 in human RPE in situ was determined after biotinylation of proteins on cell surfaces and streptavidin chromatography, followed by immunoblot analyses. AQP1 distribution in a polarized in vitro RPE model was determined with indirect immunofluorescence confocal microscopy. The role of channel-mediated transport of water across RPE cell monolayers on filters was assessed by osmotic challenge assay. Expression levels of AQP1 were controlled with an adenovirus expression system and monitored by immunoblot analyses.


RESULTS
AQP1 protein was detected in human RPE in situ and in cultures of human adult and fetal RPE cells. In functional assays, AQP1 facilitated water movement across RPE monolayers in an expression-dependent manner in two complementary model systems.


CONCLUSIONS
The expression of AQP1 by RPE in vivo probably contributes to the efficient transepithelial water transport across RPE, maintains retinal attachment, and prevents subretinal edema.
Stamer
W Daniel
WD
Department of Ophthalmology, The University of Arizona, Tucson, Arizona 85711-1824, USA. dstamer@eyes.arizona.edu
Bok
Dean
D
Hu
Jane
J
Jaffe
Glenn J
GJ
McKay
Brian S
BS
D000256:Adenoviridae / Q000235:genetics; D000293:Adolescent; D051398:Aquaporin 1; D020346:Aquaporins / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport; D001710:Biotin; D001789:Blood Group Antigens; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D005333:Fetus; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D006801:Humans; D018613:Microscopy, Confocal; D008875:Middle Aged; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D014867:Water / Q000378:metabolism*
C496020:AQP1 protein, human; D020346:Aquaporins; D001789:Blood Group Antigens; D014867:Water; D051398:Aquaporin 1; D001710:Biotin