In-vivo imaging of choroidal neovascularization using a fluorescent labeled multivalent polymer targeting L- and P-selectin

Abstract

Purpose: Selectins are expressed on the vascular endothelium and leukocytes, mediating the migration of leukocytes during inflammation. The novel multivalent synthetic dendritic polyglycerol sulfate (dPGS) inhibits selectins and suppresses inflammation. Aim of the study was to investigate fluorescently labeled dPGS as imaging probe in an animal model of laser-induced choroidal neovascularization (CNV).

Methods: For in-vivo imaging, dPGS was covalently attached to an indocyanine green type dye (6S-ICG), yielding soluble conjugates. Using confocal scanning laser ophthalmoscopy (cSLO), in-vivo reflectance and fluorescence imaging was performed in Dark Agouti rats that had undergone argon laser photocoagulation to induce CNV. Retinal uptake and fluorescence were recorded following intravenous and intravitreal injection of dPGS-6S-ICG. The distribution and accumulation of dPGS-6S-ICG were measured and frozen sections as well as flatmounts were prepared.

Results: Immediately following intravenous and intravitreal injection a strong fluorescence was visible. Twenty-four hours following injection an accumulation of dPGS-6S-ICG within the laser lesions were observed. Furthermore, multiple fluorescent spots were visible up to 56 days following intravenous injection and for up to 100 days following intravitreal injection of dPGS-6S-ICG. Over time, a continuous decrease of the fluorescence intensity was observed. Post-mortem analysis revealed a distinct accumulation to macrophages and/or microglia cells in frozen sections (after staining with CD68 and Iba1).

Conclusions: Pharmacokinetics of fluorescent dPGS can be investigated in-vivo following intravenous and intravitreal injection. The in-vivo and ex-vivo observations are in accordance with an immune mediated response following laser treatment. Fluorescent dPGS may be a potential biomarker for the in-vivo assessment of inflammation and leukocyte migration in retinal disorders.

Keywords: 453 choroid: neovascularization • 552 imaging methods (CT, FA, ICG, MRI, OCT, RTA, SLO, ultrasound) • 688 retina