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Jehwi Jeon, Sang-hoon Kim, Soojin Kim, Eunji Kong, Jee Myung Yang, Joo Yong Lee, Junyeop Lee, You-me Kim, Pilhan Kim; Natural intravital branch retinal artery occlusion model for investigating retinal microglia in ischemic reperfusion injury. Invest. Ophthalmol. Vis. Sci. 2022;63(7):4101 – F0065.
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© ARVO (1962-2015); The Authors (2016-present)
To establish a natural and reproducible branch retinal artery occlusion (BRAO) mouse model by using intravital confocal retinal microscopy and to analyze pathophysiologic changes of the retinal microglia in Ischemic-Reperfusion (IR) injury after BRAO modeling.
Rose bengal (75mg/kg) was injected to 7 weeks-old transgenic mice (CX3CR1-GFP, for microglial visualization). 561nm laser was projected to a specified single vessel to induce photo-thrombosis for 27 seconds by custom-built video-rate confocal microscopy. Thrombosis was recorded in real time and intravital retinal images were longitudinally acquired for 7 days. Immunohistochemistry, fluorescence associated cell sorting (FACS) analysis and RT-qPCR were performed to observe IR injury related reactions of retinal microglia.
Establishment of reproducible BRAO modeling was archived. Blocked perfusion in the targeted artery by embolism occlusion and the formation of ischemic area were reliably observed in all BRAO induced model. Dynamic alteration in the molecular profiles of immune cell infiltrated to the ischemic lesion were prominently activated at 3 days after BRAO modeling. Especially, Nox2 RNA expression, CD86 expression and microglial morphology changes were peaked at that time. The time point is co-related with the reperfusion with thrombus self-resolution. At day 7 with restored reperfusion in large vessel, CX3CR1 signal was slightly decreased but the number of CX3CR1+ microglia were significantly increased around ischemic area. These microglia were focally recruited from optic disc through nerve fibers and large veins. And most of the CX3CR1-GFP (+) cells had no CCR2 expression.
Our study successfully demonstrated a reproducible BRAO modeling with technical advantages of precise control of the time points and selection of a specific single target vessel. The role of retinal microglia in IR injury after BRAO could be analyzed. It will be a useful experimental tool to investigate the pathophysiology of BRAO.
This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.
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