Hyperoxia causes reduced density of retinal astrocytes in the central avascular zone in the mouse model of oxygen-induced retinopathy.

The mouse model of oxygen-induced retinopathy (OIR) is commonly used to investigate various aspects of the pathogenesis of the retinopathy of prematurity (ROP) as well as angiogenesis in general. Retinal astrocytes were suggested to be involved in retinal angiogenesis. This study aimed to describe their localization and cell density during the course of physiological vascularization and pathological revascularization. Mice expressing H2B-GFP (green fluorescent protein fused to histone 2B) from the endogenous Pdgfra promoter were kept in 75% oxygen from P7 (post natal day 7) to P12 (mouse model of OIR). Retinal flatmounts or cryosections were immunostained for glial fibrillary acidic protein (Gfap), glutamine synthetase (Glul), collagen IV (Col IV), desmin (Des), caspase 3 (Casp3), paired box 2 (Pax2), or Ki67. Astrocytic nuclei were counted with the ImageJ macro AuTOCellQuant. The hypoxic state of the retina was investigated by Hypoxyprobe. The GFP signal of the Pdgfra reporter mice co-localized with Pax2, a nuclear marker for retinal astrocytes. This bright label was much easier to quantify than Gfap or Pax2 staining. Quantification of the cell density of astrocytes during physiological development specified the spreading of astrocytes in a concentrical wave from the optic nerve head towards the periphery. Astrocyte density was reduced during the remodelling of the primary vascular plexus into a hierarchical vascular tree (maximal astrocyte density at P1: 2800 astrocytes/mm2, final astrocyte density: 800 astrocytes/mm2). In the OIR model, cell density of astrocytes was elevated in the peripheral vascularized zone. In contrast, astrocyte density dropped to a half (400 astrocytes/mm2) of the normal value in the central avascular zone during the hyperoxic phase between P8 and P10 by apoptosis and rose only after P17 as the retinal network normalized. An additional drop of astrocyte density was observed within the angles between the large vessels of the central avascular zone during hypoxia between P12 and P17. Astrocyte density was not altered at vascular tufts. The hyperoxia effect on astrocytes including the reduced astrocyte density is not the reason for vascular tuft formation. Hypoxia-affected astrocytes in combination with a reduced astrocytic network in the central avascular zone during the hypoxic phase are important determinants in the formation of pathological features during retinal revascularization.

Bucher F ，Stahl A ，Agostini HT ，Martin G ... -  《-》

Effects of sustained hyperoxia on revascularization in experimental retinopathy of prematurity.

Gu X ，Samuel S ，El-Shabrawey M ，Caldwell RB ，Bartoli M ，Marcus DM ，Brooks SE ... -  《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》

Localization of adenosine A2a receptor in retinal development and oxygen-induced retinopathy.

Taomoto M ，McLeod DS ，Merges C ，Lutty GA ... -  《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》

Adenosine A1 Receptors Selectively Modulate Oxygen-Induced Retinopathy at the Hyperoxic and Hypoxic Phases by Distinct Cellular Mechanisms.

We critically evaluated the role of the adenosine A1 receptor (A1R) in normal development of retinal vasculature and pathogenesis of retinopathy of prematurity (ROP) by using the A1R knockout (KO) mice and oxygen-induced retinopathy (OIR) model. Mice deficient in A1Rs and their wild-type (WT) littermates were examined during normal postnatal development or after being subjected to 75% oxygen from postnatal day (P) 7 to P12 and to room air from P12 to P17 (OIR model of ROP). Retinal vascularization was examined by whole-mount fluorescence and cross-sectional hematoxylin-eosin staining. Cellular proliferation, astrocyte and microglial activation, and tip cell function were determined by isolectin staining and immunohistochemistry. Apoptosis was determined by TUNEL assay. Genetic deletion of the A1R did not affect normal retinal vascularization during postnatal development with indistinguishable three-layer vascularization patterns in retina between WT and A1R KO mice. In the OIR model, genetic deletion of the A1R resulted in stage-specific effects: reduced hyperoxia-induced retinal vaso-obliteration at P12, but reduced avascular area and attenuated hypoxia-induced intraretinal revascularization without affecting intravitreal neovascularization at P17 and reduced avascular areas in retina at P21. These distinct effects of A1Rs on OIR were associated with A1R control of apoptosis mainly in inner and outer nuclear layers at the vaso-obliterative phase (P12) and the growth of endothelium tip cells at the vasoproliferative phase (P17), without modification of cellular proliferation, astrocytic activation, and tissue inflammation. Adenosine A1 receptor activity is not required for normal postnatal development of retinal vasculature but selectively controls hyperoxia-induced vaso-obliteration and hypoxia-driven revascularization by distinct cellular mechanisms.

Zhang S ，Li H ，Li B ，Zhong D ，Gu X ，Tang L ，Wang Y ，Wang C ，Zhou R ，Li Y ，He Y ，Chen M ，Huo Y ，Liu XL ，Chen JF ... -  《-》

T2-TrpRS inhibits preretinal neovascularization and enhances physiological vascular regrowth in OIR as assessed by a new method of quantification.

Banin E ，Dorrell MI ，Aguilar E ，Ritter MR ，Aderman CM ，Smith AC ，Friedlander J ，Friedlander M ... -  《INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE》