Norrin treatment improves ganglion cell survival in an oxygen-induced retinopathy model of retinal ischemia

Abstract

Treatment of a mouse model of oxygen-induced retinopathy (OIR) with recombinant human Norrin (Norrie Disease Protein, gene: NDP) accelerates regrowth of the microvasculature into central ischemic regions of the neural retina, which are generated after treatment with 75% oxygen. While this reduces the average duration and severity of ischemia overall, we do not know if this accelerated recovery of the microvasculature results in any significant survival of retinal ganglion cells (RGCs). The purpose of this study was to investigate ganglion cell survival with and without the intravitreal injection of Norrin in the murine model of oxygen induced retinopathy (OIR), using two strains of mice: C57BL/6J and Thy1-YFP mice. Intravitreal injections of Norrin or vehicle were done after five days of exposure to 75% oxygen from ages P7 to P12. The C57BL/J mice were followed by Spectral-Domain Optical Coherence Tomography (SD-OCT), and the average nerve fiber layer (NFL) and inner-plexiform layer (IPL) thicknesses were measured at twenty-four locations per retina at P42. Additionally, some C57BL/J retinas were flat mounted and immunostained for the RGC marker, Brn3a, to compare the population density of surviving retinal ganglion cells. Using homozygous Thy1-YFP mice, single intrinsically fluorescent RGCs were imaged in live animals with a Micron-III imaging system at ages P21, 28 and P42. The relative percentage of YFP-fluorescent RGCs with dendritic arbors were compared. At age P42, the NFL was thicker in Norrin-injected OIR eyes, 14.4 μm, compared to Vehicle-injected OIR eyes, 13.3 μm (p = 0.01). In the superior retina, the average thickness of the IPL was greater in Norrin-injected OIR eyes, 37.7 μm, compared to Vehicle-injected OIR eyes, 34.6 μm (p = 0.04). Retinas from Norrin injected OIR mice had significantly more surviving RGCs (p = 0.03) than vehicle-injected mice. Based upon NFL thickness and counts of RGCs, we conclude that Norrin treatment, early in the ischemic phase, increased the relative population density of surviving RGCs in the central retinas of OIR mice.

Keywords: Ganglion cell; Neovascular; Nerve fiber layer; Norrin; Optical Coherence Tomography; Oxygen induced retinopathy; Retinal ischemia.