Abstract
Purpose :
Nrf2 has been shown to play a protective role in regulating oxidative stress and inflammation. Our group previously found that global Nrf2 deficiency, as well as Nrf2 deficiency in neuroretina suppresses retinal revascularization and promotes pathological neovascularization in the mouse model of oxygen-induced retinopathy (OIR). Müller cells are a major glial cell type known to be involved in regulating pathological neovascularization and retinal dysfunction in ischemia-induced retinopathy. The purpose of this study was to explore whether Müller glia-derived Nrf2 has any potential role on retinal angiogenesis in the mouse OIR model.
Methods :
Müller specific Nrf2 knockout (KO) mice were generated by crossing GlastCreER transgenic mice with Nrf2lox/lox mice. Müller-specific Nrf2 KO and littermate control mice were subjected to the mouse OIR model, and retinal flat mounts with lectin staining were prepared. The areas of avascular retina and neovascular tufts were measured. Real-time PCR was used to examine expression of multiple inflammatory cytokines and angiogenesis-related factors. Primary Müller cells from Nrf2 KO and wild-type (WT) control retinas were isolated using collagenase digestion and cultured in vitro. Tube formation assay and spheroid sprouting assay were used as in vitro complementary approaches to investigate retinal vascularization.
Results :
Müller-specific Nrf2 KO mice exhibited enhanced vaso-obliteration and retinal neovascularization at P17 compared to littermate control mice, as well as increased Müller gliosis. The mRNA levels of GFAP and TNF-alpha were increased in the knockout mice. Primary Müller cells from Nrf2 KO mice exhibited a slower proliferation rate compared to WT Müller cells. Retinal endothelial cells co-cultured with Nrf2 KO Müller cells formed fewer tubes in collagen gel than those co-cultured with WT Müller cells. HREC sprouting from the spheroid mixed with WT Müller cells migrated a longer distance than those sprouting from spheroids mixed with Nrf2 KO Müller cells. We found elevated VEGF expression, and decreased IGF1 and IL-6 expression in Nrf2 KO Müller cells compared to WT Müller cells.
Conclusions :
These results indicate that Nrf2 in Müller glia could modulate retinal revascularization and pathological neovascularization in mouse OIR model, through the regulation of angiogenesis- and inflammation-related gene expression.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.