Vascular endothelial growth factor up-regulates the expression of intracellular adhesion molecule-1 in retinal endothelial cells via reactive oxygen species, but not nitric oxide.
- Author:
Xiao-ling ZHANG
1
;
Liang WEN
;
Yan-jiong CHEN
;
Yi ZHU
Author Information
- Publication Type:Journal Article
- MeSH: Animals; Blotting, Western; Cattle; Cells, Cultured; Endothelial Cells; cytology; drug effects; metabolism; Gene Expression; drug effects; Intercellular Adhesion Molecule-1; metabolism; Nitric Oxide; metabolism; Reactive Oxygen Species; metabolism; Retina; cytology; Vascular Endothelial Growth Factor A; pharmacology
- From: Chinese Medical Journal 2009;122(3):338-343
- CountryChina
- Language:English
-
Abstract:
BACKGROUNDThe vascular endothelial growth factor (VEGF) is involved in the initiation of retinal vascular leakage and nonperfusion in diabetes. The intracellular adhesion molecule-1 (ICAM-1) is the key mediator of the effect of VEGFs on retinal leukostasis. Although the VEGF is expressed in an early-stage diabetic retina, whether it directly up-regulates ICAM-1 in retinal endothelial cells (ECs) is unknown. In this study, we provided a new mechanism to explain that VEGF does up-regulate the expression of ICAM-1 in retinal ECs.
METHODSBovine retinal ECs (BRECs) were isolated and cultured. Immunohistochemical staining was performed to identify BRECs. The cultured cells were divided into corresponding groups. Then, VEGF (100 ng/ml) and other inhibitors were used to treat the cells. Cell lysate and the cultured supernatant were collected, and then, the protein level of ICAM-1 and phosphorylation of the endothelial nitric oxide synthase (eNOS) were detected using Western blotting. Griess reaction was used to detect nitric oxide (NO).
RESULTSWestern blotting showed that the VEGF up-regulated the expression of ICAM-1 protein and increased phosphorylation of the eNOS in retinal ECs. Neither the block of NO nor protein kinase C (PKC) altered the expression of ICAM-1 or the phosphorylation of eNOS. The result of the Western blotting also showed that inhibition of phosphatidylinositol 3-kinase (PI3K) or reactive oxygen species (ROS) significantly reduced the expression of ICAM-1. Inhibition of PI3K also reduced phosphorylation of eNOS. Griess reaction showed that VEGF significantly increased during NO production. When eNOS was blocked by L-NAME or PI3K was blocked by LY294002, the basal level of NO production and the increment of NO caused by VEGF could be significantly decreased.
CONCLUSIONROS-NO coupling in the retinal endothelium may be a new mechanism that could help to explain why VEGF induces ICAM-1 expression and the resulting leukostasis in diabetic retinopathy.