Signals Through Tumor Necrosis Factor Alpha Receptor p75 Exacerbate CNV Formation in Mice

Abstract

Purpose: : Tumor necrosis factor alpha (TNF-a), a proinflammatory cytokine is highly expressed in fibrovascular membranes isolated from AMD patients and is known to modulate choroidal neovascularization (CNV). Little is known about the role of TNF-a receptors (TNFRp55 and TNFRp75) in angiogenic signaling in vivo and their role in the development of CNV. We investigated the role of both receptors in the development of neovascularization in a mouse model of laser-photocoagulation-induced CNV.

Methods: : Five separate laser burns were applied to induce ruptures of Bruch's membrane and subsequent choroidal neovascularization in TNFRp55^-/- (KO) and TNFRp75^-/- (KO) as well as the respective wild type (WT) mice. CNV formation was compared with respect to fluorescein angiographic leakage and also by histological appearance. CNV lesions were assessed on choroidal flatmounts after labeling of the blood vessels with isolectin IB4 or CD31 antibody. The pathological infiltration with inflammatory cells was investigated by immunocytochemistry with F4/80 and CD11b. Further, we investigated whether signals through TNFRp55 can inhibit choroidal neovascularization via endothelial cell apoptosis. TUNEL assays on paraffin sections and Western blots for active caspase-3 and caspase-8 were performed in TNFR KOs and WT mice.

Results: : Pathological vascular leakage investigated by fluorescein angiography was decreased by almost 50% in TNFRp75^-/- KO compared to WT mice, but no difference between TNFRp55^-/- KO and WT mice was observed. Furthermore, on choroidal flat-preparations and histological sections the number of the laser scars as well as their size and fluorescence intensity was reduced in TNFRp75^-/- KO but not in TNFRp55^-/- KO mice in comparison to WT animals. High infiltration with F4/80-positive macrophages was observed within and near to the laser burn in TNFRp55^-/- mice. Increased apoptotic activity (active caspase-3 and -8) was determined in choroid from TNFRp55 KO mice.

Conclusions: : These results demonstrate that TNFRp55 and TNFRp75 play probably differential roles in CNV formation after laser photocoagulation. This might be due to their opposite effects on different inflammatory signals and on the vascular growth.

Keywords: age-related macular degeneration • inflammation • neovascularization