A Novel Pro-Angiogenic Function for Interferon-γ-Secreting Natural Killer Cells

PURPOSE. To explore the function of natural killer (NK) cells in inflammatory angiogenesis in mice. METHODS. To study ocular angiogenic responses we used the cornea BFGF micropellet and the laser-induced choroidal neovascularization (CNV) mouse models (C57BL/6). To deplete NK cells in these models, we injected an anti-NK1.1 antibody or an isotype antibody as a control. Corneas or choroids were immunohistochemically stained for blood vessels (CD31), macrophages (F4/80), or CNV (isolectin-IB4). Vascular endothelial growth factors (VEGF), IFN-gamma, or TNF-alpha levels were measured by real-time quantitative PCR (qPCR) or flow cytometry. A coculture assay of macrophages, NK cells, and human umbilical vein endothelial cells (HUVECs) was analyzed morphometrically to examine the ability of NK cells to induce angiogenesis in vitro. RESULTS. Our data demonstrate that in vivo depletion of NK cells leads to a significant reduction of corneal angiogenesis and CNV. Furthermore, NK cell depletion reduces macrophage infiltration into the cornea and mRNA expression levels of VEGF-A, VEGF-C, and VEGFR3 at day 7 after micropellet insertion. In the laser-induced CNV model, our data show that NK cell depletion leads to decreased areas of CNV and significantly reduced mRNA expression of VEGFs and IFN-gamma in the choroid. An in vitro coculture assay shows an IFN-gamma-dependent increase in VEGF expression levels, thereby increasing endothelial cell proliferation. CONCLUSIONS. Our findings demonstrate a novel pro-angiogenic function for NK cells, indicating that IFN-gamma-secreting NK cells can induce angiogenesis by promoting enhanced VEGF expression by macrophages.