Vascular endothelial growth factor expression and signaling in the lens.
OBJECTIVE
Previous studies have identified sequences encoding vascular endothelial growth factor (VEGF)-A and one of the VEGF receptors (VEGFR2, Flk-1, KDR) in lens fiber cells. The current study was undertaken to determine the distribution of VEGF-A protein in the lens, whether signaling through VEGF receptors occurs in lens cells, the pattern of VEGF-A expression during lens development, and the effect of hypoxia on VEGF-A expression.


METHODS
VEGF-A and VEGFR2 were localized using immunocytochemistry. VEGF-A and VEGFR2 protein were identified and quantified by Western blot analysis. Activated (tyrosine phosphorylated) VEGFR2 was detected by immunoprecipitation with an anti-phosphotyrosine antibody followed by Western blot analysis with antibody to VEGFR2. Levels of VEGF-A mRNA were measured by quantitative PCR. Suturing the lids of adult mouse or rabbit eyes for 3 days was used to induce lens hypoxia.


RESULTS
VEGFR2 sequences were present in adult human lens epithelial cells, and VEGF-A transcripts were detected in chicken embryo, adult human, and mouse lens epithelial cells. VEGF-A protein localized to the ends of mouse embryo lens fiber cells at developmental stages when the fetal vasculature was forming. At later stages, VEGF-A was distributed uniformly throughout the cytoplasm of cortical fiber cells. VEGFR2 was present in mouse lens epithelial and fiber cells and was tyrosine phosphorylated at all stages examined. VEGF-A protein was barely detectable in lens epithelial cells during the first postnatal week, but increased as the capillaries of the anterior pupillary membrane regressed. VEGF-A levels were highest in adult lenses. Suturing the eyelid caused an increase in VEGF-A mRNA and protein in lens epithelial and fiber cells.


CONCLUSIONS
VEGF-A secreted by lens cells may stimulate the formation of the fetal vasculature, but regression of these vessels is not likely to be caused by a reduction in VEGF-A production by the lens. An active VEGF-A signaling system of unknown function appears to be active in the lens. It is likely that VEGF-A expression is regulated by tissue hypoxia at all stages of lens development.
Shui
Ying-Bo
YB
Department of Ophthalmology and Visual Sciences, Washington University, St. Louis, Missouri 63110, USA.
Wang
Xiaohui
X
Hu
Joan S
JS
Wang
Shui-Ping
SP
Garcia
Claudia M
CM
Potts
Jay D
JD
Sharma
Yogendra
Y
Beebe
David C
DC
D000818:Animals; D015153:Blotting, Western; D002642:Chick Embryo; D016228:Endothelial Growth Factors / Q000235:genetics / Q000378:metabolism*; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D000860:Hypoxia / Q000378:metabolism; D007150:Immunohistochemistry; D007908:Lens, Crystalline / Q000196:embryology / Q000254:growth & development / Q000378:metabolism*; D051379:Mice; D016133:Polymerase Chain Reaction; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D051381:Rats; D015398:Signal Transduction; D042461:Vascular Endothelial Growth Factor A; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics / Q000378:metabolism*
D016228:Endothelial Growth Factors; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; D040301:Vascular Endothelial Growth Factor Receptor-2