Journal
GRAEFES ARCHIVE FOR CLINICAL AND EXPERIMENTAL OPHTHALMOLOGY
Volume 246, Issue 10, Pages 1413-1422Publisher
SPRINGER
DOI: 10.1007/s00417-008-0858-8
Keywords
retinal pigment epithelium; choroidal microvascular endothelial cell; choroidal neovascularization; growth factors
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Funding
- National Natural Science Foundation of China [30371516, 30672291]
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Background Retinal pigment epithelial (RPE) cells and choroidal microvascular endothelial cells (CECs) are the main cells involved in choroidal neovascularization (CNV), and hypoxia plays an important role in CNV formation via hypoxia inducible factor 1 (HIF-1). Our aim was to evaluate the role of HIF-1 in human RPE cells with regard to proliferation, migration and tube formation of CECs under hypoxia. Methods RPE cells were cultured under chemical hypoxia induced by 200 mu M CoCl(2), and RNA interference (RNAi) technique was used to knock down HIF-1 alpha gene in RPE cells. mRNA and protein expression of HIF-1 alpha and VEGF in RPE cells were investigated by real-time RT-PCR and Western blot. Three kinds of coculture models were used to observe the effects of RPE cells transfected by short hairpin RNA (shRNA)-expressing plasmid DNA (pDNA) (pshHIF-1 alpha) on the proliferation, migration and tube formation of CECs respectively. Results Transfection of shRNA-expressing pDNA targeting HIF-1 alpha to RPE cells resulted in the knock down of HIF-1 alpha gene and reduction of the corresponding mRNA and protein of HIF-1 alpha and VEGF under hypoxia. Consequently, the proliferation, migration and tube formation of CECs were significantly inhibited by the knocked-down RPE cells compared with the control in the coculture system. The proliferation rates of CECs decreased by 40.2%, 36.6% and 36.8% on days 3, 4 and 5 respectively. Migration reduced by 49.6% at 5 h, and tube formation decreased by 40.4% at 48 h. Conclusions RNAi of HIF-1 alpha in RPE cells can inhibit angiogenesis in vitro and provide a possible strategy for treatment of choroidal neovascularization diseases by targeting HIF-1 alpha.
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