Dependence of regulatory volume decrease on transient receptor potential vanilloid 4 (TRPV4) expression in human corneal epithelial cells.
TRPV4 is a non-selective cation channel with moderate calcium permeability, which is activated by exposure to hypotonicity. Such a stress induces regulatory volume decrease (RVD) behavior in human corneal epithelial cells (HCEC). We hypothesize that TRPV4 channel mediates RVD in HCEC. Immunohistochemistry revealed centrally and superficially concentrated TRPV4 localization in the corneal tissue. Immunocytochemical and fluorescence activated cell sorter (FACS) analyses identified TRPV4 membrane surface and cytosolic expression. RT-PCR and Western blot analyses identified TRPV4 gene and protein expression in HCEC, respectively. In addition, 4alpha-PDD or a 50% hypotonic medium induced up to threefold transient intracellular Ca2+ ([Ca2+]i) increases. Following TRPV4 siRNA HCEC transfection, its protein expression level declined by 64%, which abrogated these [Ca2+]i transients. Similarly, exposure to either ruthenium red or Ca(2+)-free Ringer's solution also eliminated this response. In these transfected cells, RVD declined by 51% whereas in the non-transfected counterpart, ruthenium red and Ca(2+)-free solution inhibited RVD by 54 and 64%, respectively. In contrast, capsazepine, a TRPV1 antagonist, failed to suppress [Ca2+]i transients and RVD. TRPV4 activation contributes to RVD since declines in TRPV4 expression and activity are associated with suppression of this response. In conclusion, there is TRPV4 functional expression in HCEC.
Pan
Zan
Z
Department of Biological Sciences, State University of New York, College of Optometry, 33 West 42nd Street, New York, NY 10036, USA.
Yang
Hua
H
Mergler
Stefan
S
Liu
Hongshan
H
Tachado
Souvenir D
SD
Zhang
Fan
F
Kao
Winston W Y
WW
Koziel
Henry
H
Pleyer
Uwe
U
Reinach
Peter S
PS
D002118:Calcium / Q000378:metabolism*; D002211:Capsaicin / Q000031:analogs & derivatives / Q000494:pharmacology; D002460:Cell Line; D004396:Coloring Agents / Q000494:pharmacology; D019573:Epithelium, Corneal / Q000187:drug effects / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006801:Humans; D007038:Hypotonic Solutions; D007552:Isotonic Solutions / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000378:metabolism; D000077331:Ringer's Solution; D012430:Ruthenium Red / Q000494:pharmacology; D050916:TRPV Cation Channels / Q000037:antagonists & inhibitors / Q000187:drug effects / Q000378:metabolism*
D004396:Coloring Agents; D007038:Hypotonic Solutions; D007552:Isotonic Solutions; D034741:RNA, Small Interfering; D050916:TRPV Cation Channels; C416986:TRPV4 protein, human; D012430:Ruthenium Red; D000077331:Ringer's Solution; C071423:capsazepine; D002211:Capsaicin; D002118:Calcium