title: Early vessel destabilization mediated by Angiopoietin-2 and subsequent vessel maturation via Angiopoietin-1 induce functional neovasculature after ischemia.
abstract: BACKGROUND
  We assessed whether Angiopoietin-2 (Ang2), a Tie2 ligand and partial antagonist of Angiopoietin-1 (Ang1), is required for early vessel destabilization during postischemic angiogenesis, when combined with vascular growth factors.
  
  
  METHODS
  In vitro, matrigel co-cultures assessed endothelial-cell tube formation and pericyte recruitment after stimulation of VEGF-A, Apelin (APLN), Ang1 with or without Ang2. In a murine hindlimb ischemia model, adeno-associated virus (rAAV, 3×10(12) virusparticles) transduction of VEGF-A, APLN and Ang1 with or without Ang2 (continuous or early expression d0-3) was performed intramuscularly (d-14). Femoral artery ligation was performed at d0, followed by laser doppler perfusion meassurements (LDI) 7 and 14. At d7 (early timepoint) and d14 (late timepoint), histological analysis of capillary/muscle fiber ratio (CMF-R, PECAM-1) and pericyte/capillary ratio (PC-R, NG2) was performed.
  
  
  RESULTS
  In vitro, VEGF-A, APLN and Ang1 induced ring formation, but only APLN and Ang1 recruited pericytes. Ang2 did not affect tube formation by APLN, but reduced pericyte recruitment after APLN or Ang1 overexpression. In vivo, rAAV.VEGF-A did not alter LDI-perfusion at d14, consistent with an impaired PC-R despite a rise in CMF-R. rAAV.APLN improved perfusion at d14, with or without continuous Ang2, increasing CMF-R and PC-R. rAAV.Ang1 improved perfusion at d14, when combined with rAAV.Ang2 (d0-3), accompanied by an increased CMF-R and PC-R.
  
  
  CONCLUSIONS
  The combination of early vessel destabilization (Ang2 d0-3) and continuous Ang1 overexpression improves hindlimb perfusion, pointing to the importance of early vessel destabilization and subsequent vessel maturation for enhanced therapeutic neovascularization.
authors.affiliation: Medizinische Klinik und Poliklinik I, Klinikum Großhadern, Munich, Germany.
authors.forename: Di
authors.initials: D
authors.lastname: Qin
authors.forename: Teresa
authors.initials: T
authors.lastname: Trenkwalder
authors.forename: Seungmin
authors.initials: S
authors.lastname: Lee
authors.forename: Omary
authors.initials: O
authors.lastname: Chillo
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Deindl
authors.forename: Christian
authors.initials: C
authors.lastname: Kupatt
authors.forename: Rabea
authors.initials: R
authors.lastname: Hinkel
mesh_terms: D042683:Angiopoietin-1 / Q000494:pharmacology*; D042702:Angiopoietin-2 / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D018920:Coculture Techniques; D006614:Hindlimb / Q000473:pathology; D006801:Humans; D007511:Ischemia / Q000378:metabolism* / Q000473:pathology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018919:Neovascularization, Physiologic / Q000187:drug effects*; D020286:Pericytes / Q000166:cytology / Q000187:drug effects
chemical_list: D042683:Angiopoietin-1; D042702:Angiopoietin-2