Neutrophils promote experimental choroidal neovascularization

Mol Vis. 2005 Jun 16;11:414-24.

Abstract

Purpose: To investigate the role of neutrophils in the development of laser induced experimental choroidal neovascularization (CNV).

Methods: CNV was induced by laser photocoagulation in adult male C57BL/6J mice. Neutrophil infiltration was evaluated by histology and confocal immunohistology. The expression of neutrophil chemotactic chemokines in the regions of laser injury was determined by quantitative real-time PCR. Animals were treated with NIMP-R14, an anti-murine neutrophil monoclonal antibody (mAb), intraperitoneally to deplete neutrophils. The specific neutrophil depletion was confirmed by flow cytometry. The CNV responses were compared between neutropenic and untreated control mice on the basis of fluorescein angiography (FA), CNV lesion volume and lesion histology, and vascular endothelial growth factor (VEGF) expression by ELISA. Expression of VEGF and Angiopoietin-1 and Angiopoietin-2 protein by murine neutrophils was evaluated by confocal immunohistochemistry.

Results: Neutrophils infiltrated the sites of laser injury as early as day 1 after laser treatment and peaked at day 3. The neutrophil infiltration correlated with enhanced mRNA expression of neutrophil chemotactic chemokines MIP-2 and KC in the lesions. Administration of NIMP-R14 mAb specifically depleted neutrophils. Analysis of FA, CNV volume, and lesion histology, all demonstrated a moderate decrease in the CNV response in neutropenic mice compared to control mice (p<0.01). The reduction in the CNV response in neutropenic mice was associated with decreased VEGF protein levels in the ocular posterior segment. Murine neutrophils contained VEGF and Angiopoietin-1 and Angiopoietin-2 proteins.

Conclusions: Neutrophil invasion was part of early inflammatory responses during laser induced CNV. Neutrophil depletion correlated with reduced CNV responses and decreased VEGF protein expression. These data suggest that neutrophils promoted the early development of CNV possibly via secretion of angiogenic growth factors.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Angiopoietin-1 / metabolism
  • Angiopoietin-2 / metabolism
  • Animals
  • Chemokine CXCL1
  • Chemokine CXCL2
  • Chemokines / genetics
  • Chemokines, CXC
  • Choroidal Neovascularization / metabolism
  • Choroidal Neovascularization / physiopathology*
  • Cytokines / genetics
  • Disease Models, Animal
  • Enzyme-Linked Immunosorbent Assay
  • Flow Cytometry
  • Fluorescein Angiography
  • Immunoenzyme Techniques
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Neutrophil Infiltration / physiology*
  • Neutrophils / physiology*
  • RNA, Messenger / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Vascular Endothelial Growth Factor A / metabolism

Substances

  • Angiopoietin-1
  • Angiopoietin-2
  • Chemokine CXCL1
  • Chemokine CXCL2
  • Chemokines
  • Chemokines, CXC
  • Cxcl1 protein, mouse
  • Cxcl2 protein, mouse
  • Cytokines
  • RNA, Messenger
  • Vascular Endothelial Growth Factor A
  • vascular endothelial growth factor A, mouse
  • keratinocyte-derived chemokines