2022 Annual Meeting and Alumni Reunion
Category : Basic and Translational Research Candidate : Zhengping Hu Poster #: B10
Endomucin Deletion Delays Retinal Vascular Development and Inhibits Neovascularization in Oxygen-Induced Retinopathy
Zhengping Hu , Issahy Cano , Magali Saint-Geniez , Yin Shan Eric Ng , Patricia A . D ’ Amore
Purpose : Endomucin ( EMCN ) is a type I integral membrane glycoprotein selectively expressed by venous and capillary endothelium . We have previously shown that EMCN knockdown in vitro significantly inhibits VEGF165-induced VEGFR2 internalization and downstream activities including proliferation , migration and tube formation . The goal of this study is to characterize the role of EMCN in normal retinal vascular development and in pathologic neovascularization using the EMCN knock-out mice .
Methods : Homozygous EMCN knock-out ( EMCN- / - ) mice were obtained by crossing EMCN-floxed mice with the ROSA26-Cre strain . Eyes from adult ( 8-16 weeks ) EMCN- / - mice and EMCN +/+ control littermates were collected and retinas and RPE / choroids complex were dissected for RNA extraction , and qPCR to determine gene expression . Eyes from P5 mice and adult mice ( 12-16 weeks ) were collected for retinal flat mounts ; retinal vasculature was stained with Isolectin-B4 ( IB4 ). For the oxygen-induced retinopathy ( OIR ) model , P7 mice were housed in 75 % oxygen for five consecutive days and returned to room air at P12 , eyes at P12 and P17 were collected and the retinal vasculature stained with IB4 and the avascular and neovascular areas were quantified using photoshop .
Results : EMCN mRNA in both retinas and RPE / choroids from the EMCN- / - mice was undetectable ( n > 4 , p < 0.0001 ), compared to EMCN +/+ mice by qPCR . The area of retinal vascularization at P5 was significantly lower in the EMCN- / - pups compared to EMCN +/+ controls ( 0.14 ± 0.01 vs 0.2 ± 0.013 , p < 0.0001 , n > 10 for both groups ). Adult retinal vascular density remained lower in the EMCN- / - mice compared to EMCN +/+ mice ( 0.135 0.015 ± vs 0.154 ± 0.017 , p < 0.05 , n = 13 ). In the OIR model , the avascular area generated by high oxygen exposure at P12 was similar in the EMCN- / - and EMCN +/+ mice ( 24.57 ± 1.4 % vs 23.18 ± 1.0 %, p = 0.9 , n > 6 ). However , pathological neovascularization at P17 was significantly reduced in the EMCN- / - mice compared to EMCN +/+ ( 8.98 ± 2.9 % vs 11.98 ± 1.4 %, p < 0.05 , n > 10 ) while the avascular area at P17 was comparable between the EMCN- / - and controls ( 9.9 ± 1.5 % vs 8.85 ± 1.0 %, p > 0.5 , n > 10 ).
Conclusions : Genetic ablation of the EMCN gene reduces retinal vascularization under normal and pathological conditions . As a critical regulator of retinal angiogenesis , EMCN represents a novel therapeutic target for ocular diseases characterized by pathological blood vessel growth .