Antiangiogenic effects and transcriptional regulation of pigment epithelium-derived factor in diabetic retinopathy.
The effects of the antiangiogenic cytokine PEDF on key steps in retinal angiogenesis, specifically endothelial cell proliferation and vascular tubule formation, and the regulation of PEDF expression in retinal capillary endothelial cells were evaluated. HUVECs were co-cultured with fibroblasts to construct a model of angiogenesis using the Angiokit assay, and image analysis software was used to measure the effects of PEDF and VEGF on vascular tubule formation. Quantitative real-time PCR analysis was used to determine the expression of PEDF in microvascular endothelial cells exposed to glucose 20 mM, insulin 100 nM and VEGF 10 ng/ml. PEDF inhibited endothelial cell proliferation and significantly decreased the number of tubules (629+93 AU vs 311+31, p=0.001), number of branching points (145+19 AU vs 46+5, p=0.03) and total tubule length (4848+748 AU vs 11,172+2353, p=0.001). In bovine retinal capillary endothelial cells (BRCECs), PEDF mRNA and protein expression was suppressed by insulin (22%) in a rapamycin-sensitive manner; wortmannin had no effect. PEDF mRNA expression was also significantly reduced in the presence of high glucose (23%) and VEGF (25%). In conclusion, PEDF inhibits key steps in the angiogenic response of BRCECs, including endothelial cell proliferation and vascular tubule formation. Gene expression of PEDF is negatively regulated by glucose, insulin (via an mTOR-dependent pathway) and VEGF.
Wang
Binhui
B
Vascular Medicine, School of Graduate-Entry Medicine and Health, University of Nottingham, Nottingham, UK.
Atherton
Philip
P
Patel
Rekha
R
Manning
Gillian
G
Donnelly
Richard
R
D000730:Androstadienes / Q000494:pharmacology; D043165:Angiostatic Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D002417:Cattle; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D003930:Diabetic Retinopathy / Q000378:metabolism*; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005136:Eye Proteins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D005786:Gene Expression Regulation / Q000502:physiology*; D005947:Glucose / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000494:pharmacology; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D009414:Nerve Growth Factors / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D012171:Retinal Vessels / Q000166:cytology; D015843:Serpins / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D020123:Sirolimus / Q000494:pharmacology; D014470:Umbilical Cord / Q000166:cytology; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology; D000077191:Wortmannin; D000099174:Pigment Epithelium-Derived Factor
D000730:Androstadienes; D043165:Angiostatic Proteins; D004791:Enzyme Inhibitors; D005136:Eye Proteins; D005947:Glucose; D007328:Insulin; D009414:Nerve Growth Factors; D015843:Serpins; D020123:Sirolimus; D042461:Vascular Endothelial Growth Factor A; D000077191:Wortmannin; D000099174:Pigment Epithelium-Derived Factor; C467484:VEGFA protein, human