title: Heparanase and vascular endothelial growth factor expression is increased in hypoxia-induced retinal neovascularization.
abstract: OBJECTIVE
  Heparanase and VEGF are related closely to angiogenesis in cancer. The purpose of our study was to evaluate the expression and correlation of heparanase and VEGF in hypoxia-induced retinal neovascularization.
  
  
  METHODS
  C57BL/6 oxygen-induced retinopathy (OIR) mice and human retinal microvascular endothelial cells (HRECs) were treated with the hypoxia mimetic agent cobalt chloride (CoCl₂), and in the presence of the heparanase inhibitor phosphomannopentaose sulfate (Muparfostat, PI-88). Heparanase activity was assayed in HRECs, and the expression of heparanase, VEGF protein and mRNA were evaluated by immunofluorescence, ELISA, Western blot, and real-time PCR while retinal flat mounts were used to evaluate the area of neovascularization of mice retina.
  
  
  RESULTS
  HREC heparanase activity was increased by treatment with CoCl₂, but was decreased by PI-88. Immunofluorescence showed that heparanase and VEGF staining was intense in hypoxia-treated HRECs and OIR mice retina, while VEGF staining was faint in the normoxia and PI-88-treated ones. Western blot and real-time PCR results indicated that the expression of heparanase and VEGF was increased under hypoxic conditions, and the increase of VEGF was inhibited by PI-88. Retinal flat mounts showed that the area of new vessels in retina of OIR mice was increased compared to the normoxic mice, and this effect was inhibited by PI-88.
  
  
  CONCLUSIONS
  Heparanase is upregulated and associated with the VEGF expression in hypoxia-induced retinal diseases. Heparanase is involved in hypoxia-induced neovascularization through promoting VEGF expression and may be a new therapeutic target for hypoxia-induced neovascularization retinal diseases.
authors.affiliation: State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, China. hehujie@126.com
authors.forename: Jie
authors.initials: J
authors.lastname: Hu
authors.forename: Xin
authors.initials: X
authors.lastname: Song
authors.forename: Yi Qing
authors.initials: YQ
authors.lastname: He
authors.forename: Craig
authors.initials: C
authors.lastname: Freeman
authors.forename: Christopher R
authors.initials: CR
authors.lastname: Parish
authors.forename: Ling
authors.initials: L
authors.lastname: Yuan
authors.forename: Honghua
authors.initials: H
authors.lastname: Yu
authors.forename: Shibo
authors.initials: S
authors.lastname: Tang
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D015153:Blotting, Western; D004195:Disease Models, Animal; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005786:Gene Expression Regulation; D005966:Glucuronidase / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D000860:Hypoxia / Q000150:complications; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D012313:RNA / Q000096:biosynthesis / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D012160:Retina / Q000378:metabolism / Q000473:pathology*; D015861:Retinal Neovascularization / Q000209:etiology / Q000235:genetics / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A / Q000096:biosynthesis / Q000235:genetics*; D000099175:Heparanase
chemical_list: D005966:Glucuronidase; D012313:RNA; D042461:Vascular Endothelial Growth Factor A; D000099175:Heparanase