Susumu Ishida

VEGF164-mediated Inflammation Is Required for Pathological, but Not Physiological, Ischemia-induced Retinal Neovascularization

Hypoxia-induced VEGF governs both physiological retinal vascular development and pathological retinal neovascularization. In the current paper, the mechanisms of physiological and pathological neovascularization are compared and contrasted. During pathological neovascularization, both the absolute and relative expression levels for VEGF164 increased to a greater degree than during physiological neovascularization. Furthermore, extensive leukocyte adhesion was observed at the leading edge of pathological, but not physiological, neovascularization. When a VEGF164-specific neutralizing aptamer was administered, it potently suppressed the leukocyte adhesion and pathological neovascularization, whereas it had little or no effect on physiological neovascularization. In parallel experiments, genetically altered VEGF164-deficient (VEGF120/188) mice exhibited no difference in physiological neovascularization when compared with wild-type (VEGF+/+) controls. In contrast, administration of a VEGFR-1/Fc fusion protein, which blocks all VEGF isoforms, led to significant suppression of both pathological and physiological neovascularization. In addition, the targeted inactivation of monocyte lineage cells with clodronate-liposomes led to the suppression of pathological neovascularization. Conversely, the blockade of T lymphocyte–mediated immune responses with an anti-CD2 antibody exacerbated pathological neovascularization. These data highlight important molecular and cellular differences between physiological and pathological retinal neovascularization. During pathological neovascularization, VEGF164 selectively induces inflammation and cellular immunity. These processes provide positive and negative angiogenic regulation, respectively. Together, new therapeutic approaches for selectively targeting pathological, but not physiological, retinal neovascularization are outlined.

Leukocytes mediate retinal vascular remodeling during development and vaso-obliteration in disease

Retinal ischemia can cause vision-threatening pathological neovascularization. The mechanisms of retinal ischemia are not fully understood, however. Here we have shown that leukocytes prune the retinal vasculature during normal development and obliterate it in disease. Beginning at postnatal day 5 (P5) in the normal rat, vascular pruning began centrally and extended peripherally, leaving behind a less dense, smaller-caliber vasculature. The pruning was correlated with retinal vascular expression of intercellular adhesion molecule-1 (ICAM-1) and coincided with an outward-moving wave of adherent leukocytes composed in part of cytotoxic T lymphocytes. The leukocytes adhered to the vasculature through CD18 and remodeled it through Fas ligand (FasL)-mediated endothelial cell apoptosis. In a model of oxygen-induced ischemic retinopathy, this process was exaggerated. Leukocytes used CD18 and FasL to obliterate the retinal vasculature, leaving behind large areas of ischemic retina. In vitro, T lymphocytes isolated from oxygen-exposed neonates induced a FasL-mediated apoptosis of hyperoxygenated endothelial cells. Targeting these pathways may prove useful in the treatment of retinal ischemia, a leading cause of vision loss and blindness.

Reply to “FasL, leukocytes and vascular modeling”

NATURE MEDICINE VOLUME 10 | NUMBER 1 | JANUARY 2004 13 cal, as vascular modeling occurs quickly. When placed in a hyperoxic environment for 72 hours, the CD18-deficient mice showed vaso-obliteration similar to that seen in wildtype mice (S.I., K.Y., T.U. and A.P.A., unpublished data), but significantly less vaso-obliteration was seen in CD18-deficient mice at 48 hours1. Both vascular pruning and vaso-obliteration may represent defense mechanisms aimed at protecting the sensory retina from oxidative stress. As noted in our paper, compensatory systems may be used in vivo to preserve this important defense mechanism. We hypothesize that the gld mice data2,3 reflect this biological redundancy. FasL blockade did not suppress leukocyte adhesion in our experiments1. Under prolonged oxidative stress, we speculate that known leukocyte products, such as perforin and TRAIL, have a similar role in endothelial cell apoptosis in gld mice. We also noted1 that FasL-bearing blood-borne cells of nonleukocytic origin may have a role in pruning and vaso-obliteration4,5. It is therefore possible that leukocyte cytotoxicity does not depend on a single molecular pathway, and that FasL expression in the retina is not from a single cell type. A temporal examination of the role of blood-borne cells in pruning and vasoobliteration is required in the genetically altered mice that Ferguson and Stuart describe. The fact that gld mice show increased pathological neovascularization after hyperoxia-induced retinal ischemia2,3 is also consistent with our published data6. In our model of oxygen-induced retinopathy, T lymphocytes served as negative regulators of pathological neovascularization.