title: Tissue kallikrein attenuates choroidal neovascularization via cleavage of vascular endothelial growth factor.
abstract: OBJECTIVE
  To investigate the antiangiogenic properties of tissue kallikrein in a murine model of laser-induced choroidal neovascularization (CNV).
  
  
  METHODS
  CNV was induced in male C57BL/6J mice by laser photocoagulation. The animals received daily subcutaneous injections of tissue kallikrein (50 μg/kg) or vehicle control for 2 days before the laser photocoagulation, and this treatment continued until sample collection. Seven days after laser injury, the CNV size was quantified. The levels of monocyte chemoattractant protein (MCP)-1, intercellular adhesion molecule (ICAM)-1, and interleukin (IL)-6 were assessed by enzyme-linked immunosorbent assay 3 days after laser injury. Cleavage of mouse VEGF with tissue kallikrein was assessed in vivo and in vitro. The protein levels of bradykinin were assessed in the RPE-choroid complexes and hearts.
  
  
  RESULTS
  A significant decrease in CNV size was observed in animals treated with tissue kallikrein (27,168.3 ± 2432.2 μm(2)) compared with vehicle-treated controls (36,374.6 ± 3204.1 μm(2), P < 0.05). Tissue kallikrein treatment significantly reduced MCP-1, ICAM-1, and IL-6 levels in RPE-choroid complexes. Furthermore, immunoblotting showed the bands, presumably corresponding to the fragmented VEGF(164) protein, in the samples of both mouse VEGF preincubated with tissue kallikrein and RPE-choroid complexes obtained from animals treated with tissue kallikrein. In addition, bradykinin was unchanged in the RPE-choroid complexes of animals treated with tissue kallikrein, whereas the level of bradykinin was increased in the heart obtained from these experimental animals.
  
  
  CONCLUSIONS
  The current data indicate that kallikrein exhibits antiangiogenic properties by cleaving VEGF(164) in a laser-induced CNV model.
authors.affiliation: Laboratory of Ocular Cell Biology & Visual Science, Department of Ophthalmology, Hokkaido University Graduate School of Medicine, Sapporo, Japan.
authors.forename: Junichi
authors.initials: J
authors.lastname: Fukuhara
authors.forename: Kousuke
authors.initials: K
authors.lastname: Noda
authors.forename: Miyuki
authors.initials: M
authors.lastname: Murata
authors.forename: Shiho
authors.initials: S
authors.lastname: Namba
authors.forename: Satoshi
authors.initials: S
authors.lastname: Kinoshita
authors.forename: Zhenyu
authors.initials: Z
authors.lastname: Dong
authors.forename: Ryo
authors.initials: R
authors.lastname: Ando
authors.forename: Anton
authors.initials: A
authors.lastname: Lennikov
authors.forename: Atsuhiro
authors.initials: A
authors.lastname: Kanda
authors.forename: Susumu
authors.initials: S
authors.lastname: Ishida
mesh_terms: D020533:Angiogenesis Inhibitors / Q000008:administration & dosage*; D000818:Animals; D001920:Bradykinin / Q000378:metabolism; D018932:Chemokine CCL2 / Q000378:metabolism; D002829:Choroid / Q000378:metabolism; D020256:Choroidal Neovascularization / Q000473:pathology / Q000517:prevention & control*; D004195:Disease Models, Animal; D004797:Enzyme-Linked Immunosorbent Assay; D018799:Intercellular Adhesion Molecule-1 / Q000378:metabolism; D015850:Interleukin-6 / Q000378:metabolism; D017075:Laser Coagulation / Q000009:adverse effects; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D009206:Myocardium / Q000378:metabolism; D020840:Tissue Kallikreins / Q000008:administration & dosage*; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism*
chemical_list: D020533:Angiogenesis Inhibitors; C491768:Ccl2 protein, mouse; D018932:Chemokine CCL2; D015850:Interleukin-6; D042461:Vascular Endothelial Growth Factor A; D018799:Intercellular Adhesion Molecule-1; D020840:Tissue Kallikreins; D001920:Bradykinin