Blocking endothelial apoptosis revascularizes the retina in a model of ischemic retinopathy
Zoe L. Grant, … , Robert C.A. Symons, Leigh Coultas
Zoe L. Grant, … , Robert C.A. Symons, Leigh Coultas
Published May 19, 2020
Citation Information: J Clin Invest. 2020;130(8):4235-4251. https://doi.org/10.1172/JCI127668.
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Research Article Angiogenesis

Blocking endothelial apoptosis revascularizes the retina in a model of ischemic retinopathy

Abstract

Aberrant, neovascular retinal blood vessel growth is a vision-threatening complication in ischemic retinal diseases. It is driven by retinal hypoxia frequently caused by capillary nonperfusion and endothelial cell (EC) loss. We investigated the role of EC apoptosis in this process using a mouse model of ischemic retinopathy, in which vessel closure and EC apoptosis cause capillary regression and retinal ischemia followed by neovascularization. Protecting ECs from apoptosis in this model did not prevent capillary closure or retinal ischemia. Nonetheless, it prevented the clearance of ECs from closed capillaries, delaying vessel regression and allowing ECs to persist in clusters throughout the ischemic zone. In response to hypoxia, these preserved ECs underwent a vessel sprouting response and rapidly reassembled into a functional vascular network. This alleviated retinal hypoxia, preventing subsequent pathogenic neovascularization. Vessel reassembly was not limited by VEGFA neutralization, suggesting it was not dependent on the excess VEGFA produced by the ischemic retina. Neutralization of ANG2 did not prevent vessel reassembly, but did impair subsequent angiogenic expansion of the reassembled vessels. Blockade of EC apoptosis may promote ischemic tissue revascularization by preserving ECs within ischemic tissue that retain the capacity to reassemble a functional network and rapidly restore blood supply.

Authors

Zoe L. Grant, Lachlan Whitehead, Vickie H.Y. Wong, Zheng He, Richard Y. Yan, Abigail R. Miles, Andrew V. Benest, David O. Bates, Claudia Prahst, Katie Bentley, Bang V. Bui, Robert C.A. Symons, Leigh Coultas

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Figure 1

Blocking apoptosis prevents loss of ECs from retinas exposed to high oxygen.

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Blocking apoptosis prevents loss of ECs from retinas exposed to high oxy...
(A and B) Representative images and quantification of EC apoptosis visualized by active caspase-3 staining (cyan) and PECAM1 (red) in control (n = 8) and Bak–/– BaxEC/EC (24 hours, n = 3; 48 hours, n = 5) retinas after 24 or 48 hours in high oxygen. Quantitative data from control mice exposed to high oxygen for 48 hours are not shown because there are no central retina capillaries remaining. Arrow indicates rare apoptotic EC in Bak–/– BaxEC/EC retina. Scale bars: 100 μm. Student’s 2-tailed t test. (C) PECAM1 staining of control and Bak–/– BaxEC/EC retinas after 48 hours in high oxygen. Scale bars: 500 μm. (D) Quantification of central retina vessel area in mice exposed to high oxygen for 24 hours (control, n = 4; Bak–/– BaxEC/EC, n = 5) or 48 hours (control, n = 6; Bak–/– BaxEC/EC, n = 6) compared with 8-day-old normoxic mice (control, n = 3; Bak–/– BaxEC/EC, n = 3). Multiple t tests using Holm-Šidák correction for multiple comparisons. (E) PECAM1 (cyan) and collagen IV (red) staining within the central retina of control and Bak–/– BaxEC/EC mice raised in room air (normoxia) or for 48 hours in high oxygen. Scale bars: 80 μm. (F and G) Quantification of vessel regression and network fragmentation in the central retina of Bak–/– BaxEC/EC mice exposed to high oxygen for 24 hours (n = 5) or 48 hours (n = 6) compared with 8-day-old normoxic mice (control, n = 3; Bak–/– BaxEC/EC, n = 3). Quantitative data from control mice exposed to high oxygen are not shown because there are no central retina capillaries remaining. One-way ANOVA with Tukey’s multiple-comparisons test. All data are mean ± SEM. Each circle represents 1 animal.