Spatial and temporal analysis of retinal microglia in mouse oxygen-induced retinopathy model

Abstract

PURPOSE: The association between retinal microglia and retinopathy of prematurity has been noted and yet unclear. We performed a more detailed quantitative temporal and spatial analysis of retinal microglia in a mouse oxygen-induced retinopathy (OIR) model. METHODS: Postnatal day (P) 7 C57BL/6J mouse pups with their nursing mother were exposed to 75% oxygen for 5 days to induce OIR. Room air (RA)-treated pups served as controls. On P12, P17, P21, P25, and P30, retinal microglia and vessels were visualized by immunostaining under a confocal microscope. Activated (amoeboid) and resting (ramified) microglia in different retinal areas (superficial central/mid-peripheral, deep central/mid-peripheral) were identified, categorized and counted. RESULTS: On P12, OIR retina showed more amoeboid microglia in superficial central (RA: 16.9±7.0 vs. OIR: 44.8±10.4 /mm2) and mid-peripheral (RA: 18.0±2.7 vs. 64.2±16.4 /mm2) areas. On P17, OIR retina showed increased amoeboid (RA: 12.8±4.6 vs. OIR: 148.3±32.8 /mm2) and total (RA: 155.6±11.4 vs. OIR: 213.3±24.9 / mm2), but reduced ramified (RA: 142.8±13.0 vs. OIR: 52.8±15.6 / mm2) microglia in superficial central areas; in superficial midperipheral retina, OIR led to an increase in amoeboid (RA: 14.6±2.4 vs. OIR: 189.5±11.1 /mm2) and total (RA: 161.6±16.1 vs. OIR: 276.8±36.0 /mm2) microglia, but a decrease in ramified (RA: 147.0±17.4 vs. OIR: 79.7±31.8 /mm2) microglia in tufts areas; in non-tufts areas, OIR led to increased amoeboid (40.1±22.3 /mm2), ramified (209.0±48.0 /mm2) and total (258.9±38.5 /mm2) microglia; more ramified microglia in deep retina were observed. On P21, more microglia were observed in central (RA: 141.9±8.1 vs. OIR: 340.4±78.5 /mm2) and mid-peripheral (RA: 140.6±18.7 vs. OIR 255.3±91.3 /mm2) superficial retina with more amoeboid form (central: RA: 9.5±5.3 vs. OIR: 263.2±71.4 /mm2; mid-peripheral: RA: 9.1±3.6 vs. OIR: 151.6±79.3 /mm2) in OIR retina. On P25, OIR led to increased microglia in superficial central (RA: 109.1±9.3 vs. OIR: 260.2±20.6 /mm2) and mid-peripheral (RA: 115.3±7.5 vs. OIR: 241.9±20.7 /mm2) retina, as well as in deep retina. Similar patterns were observed on P30. CONCLUSIONS: Retinal microglial activation continues throughout OIR. Activated microglia are associated with vascular abnormalities. The activation of microglia upon OIR challenge last until after the recovery of retinal vessels.