title: Netrin-4 Mediates Corneal Hemangiogenesis but Not Lymphangiogenesis in the Mouse-Model of Suture-Induced Neovascularization.
abstract: Netrin-4, a secreted protein, is found in the basement membrane of blood vessels and acts as a key regulator of angiogenesis. Here we investigated the role of Netrin-4 in the mouse-model of suture-induced corneal hem- and lymphangiogenesis.
  
  
  
  Corneal hem- and lymphangiogenesis were induced in Netrin-4-deficient (Ntn4-/-) and wild-type (WT) mice by placing three 11-0 nylon sutures intrastromally. Fourteen days after suturing, the vascularized area was analyzed via corneal flat mount immunohistochemistry. Messenger RNA levels for VEGF-A, VEGF-C, Lyve-1, Netrin-4, Unc5H2, "deleted in colon cancer" receptor, and Neogenin in treated and nontreated mouse corneas, cultured human corneal keratocytes (HCK) and epithelial cells (HCEC+HCET) were analyzed by quantitative PCR.
  
  
  
  In wild-type mice, Netrin-4 mRNA expression in the cornea decreased in growing corneal neovascularization after suturing. Correspondingly, Ntn4-/- mice showed an increased vascularized area compared to that in WT mice. Expression of VEGF-A mRNA was higher in Ntn4-/- versus WT mice. There was no Netrin-4 expression in lymphatic vessels and the area of lymphatic vascularization did not differ between Ntn4-/- and WT mice, nor did expression of VEGF-C and Lyve-1 mRNA. Human corneal epithelial cells showed mainly Netrin-4 mRNA expression, which increased after stimulation, while HCK demonstrated Unc5H2 mRNA expression. Expression of VEGF-A, Netrin-4, Unc5H2, and Neogenin mRNA in HCEC and HCK did not differ significantly between the serum-free condition and VEGF-A or Netrin-4 stimulation.
  
  
  
  Absence of Netrin-4 increased corneal hemangiogenesis but not lymphangiogenesis in the mouse-model of suture-induced neovascularization. Netrin-4 acted as an antiangiogenic factor in the cornea, with which the healthy cornea is enriched via its expression by corneal epithelial cells.
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Anna-Karina B
authors.initials: AB
authors.lastname: Maier
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Klein
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Norbert
authors.initials: N
authors.lastname: Kociok
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Aline I
authors.initials: AI
authors.lastname: Riechardt
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Enken
authors.initials: E
authors.lastname: Gundlach
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Nadine
authors.initials: N
authors.lastname: Reichhart
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Olaf
authors.initials: O
authors.lastname: Strauß
authors.affiliation: Department of Ophthalmology, Charité-Universitätsmedizin, Berlin, Germany.
authors.forename: Antonia M
authors.initials: AM
authors.lastname: Joussen
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D003315:Cornea / Q000098:blood supply* / Q000378:metabolism / Q000473:pathology; D016510:Corneal Neovascularization / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D004195:Disease Models, Animal; D005786:Gene Expression Regulation; D006801:Humans; D007150:Immunohistochemistry; D042583:Lymphangiogenesis / Q000235:genetics*; D042601:Lymphatic Vessels / Q000473:pathology*; D051379:Mice; D008810:Mice, Inbred C57BL; D009414:Nerve Growth Factors / Q000096:biosynthesis / Q000235:genetics*; D000075385:Netrins; D012333:RNA, Messenger / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D013537:Sutures / Q000009:adverse effects
chemical_list: D009414:Nerve Growth Factors; D000075385:Netrins; C416214:Ntn4 protein, mouse; D012333:RNA, Messenger