Sema3E-PlexinD1 signaling selectively suppresses disoriented angiogenesis in ischemic retinopathy in mice
Yoko Fukushima, Mitsuhiro Okada, Hiroshi Kataoka, Masanori Hirashima, Yutaka Yoshida, Fanny Mann, Fumi Gomi, Kohji Nishida, Shin-Ichi Nishikawa, Akiyoshi Uemura
Yoko Fukushima, Mitsuhiro Okada, Hiroshi Kataoka, Masanori Hirashima, Yutaka Yoshida, Fanny Mann, Fumi Gomi, Kohji Nishida, Shin-Ichi Nishikawa, Akiyoshi Uemura
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Research Article Ophthalmology

Sema3E-PlexinD1 signaling selectively suppresses disoriented angiogenesis in ischemic retinopathy in mice

Abstract

During development, the retinal vasculature grows toward hypoxic areas in an organized fashion. By contrast, in ischemic retinopathies, new blood vessels grow out of the retinal surfaces without ameliorating retinal hypoxia. Restoration of proper angiogenic directionality would be of great benefit to reoxygenize the ischemic retina and resolve disease pathogenesis. Here, we show that binding of the semaphorin 3E (Sema3E) ligand to the transmembrane PlexinD1 receptor initiates a signaling pathway that normalizes angiogenic directionality in both developing retinas and ischemic retinopathy. In developing mouse retinas, inhibition of VEGF signaling resulted in downregulation of endothelial PlexinD1 expression, suggesting that astrocyte-derived VEGF normally promotes PlexinD1 expression in growing blood vessels. Neuron-derived Sema3E signaled to PlexinD1 and activated the small GTPase RhoJ in ECs, thereby counteracting VEGF-induced filopodia projections and defining the retinal vascular pathfinding. In a mouse model of ischemic retinopathy, enhanced expression of PlexinD1 and RhoJ in extraretinal vessels prevented VEGF-induced disoriented projections of the endothelial filopodia. Remarkably, intravitreal administration of Sema3E protein selectively suppressed extraretinal vascular outgrowth without affecting the desired regeneration of the retinal vasculature. Our study suggests a new paradigm for vascular regeneration therapy that guides angiogenesis precisely toward the ischemic retina.

Authors

Yoko Fukushima, Mitsuhiro Okada, Hiroshi Kataoka, Masanori Hirashima, Yutaka Yoshida, Fanny Mann, Fumi Gomi, Kohji Nishida, Shin-Ichi Nishikawa, Akiyoshi Uemura

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Figure 3

Neuron-derived Sema3E restricts vascular pathfinding in developing retinas.

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Neuron-derived Sema3E restricts vascular pathfinding in developing retin...
(A) ISH in P4 retinal sections. Plxnd1 is expressed in vascular ECs and in 7.4% of the neuronal populations in the GCL. Sema3e is expressed in 37% of neurons in the GCL. INL, inner nuclear layer. (B) High-magnification image of the advancing vascular front in AP-stained retina after intravitreal injection of Sema3E-AP fusion protein. (C) Whole-mount PECAM-1 IHC in P4 retinas. In the Sema3e–/– retina, the vascular network is disorganized in the upper-right quadrant. Note the attachment of hyaloid vessels to the retinal surface (arrowhead). (D) Quantification of filopodium length (n = 18 per group) and filopodium number/100 μm vessel length (n = 7 per group) at the sprouting vascular fronts, and vascular density (n = 4 per group) in P4 Sema3e-deficient retinas. Error bars represent SEM; ***P < 0.001, *P < 0.05. (E) Triple labeling for neurofilaments (green), PECAM-1 (red), and nuclei (blue) in retinal sections of P4 Sema3e-deficient mice. The superficial vessels (arrowheads) over the neuronal axons ectopically invade the GCL of Sema3e–/– mice (arrow). (F) A schematic diagram representing VEGF-expressing astrocyte (AC, red), PlexinD1-expressing EC (green), and Sema3E-expressing neuron (blue) in the superficial area of the developing mouse retina. Scale bars: 20 μm (A, B, and E); 100 μm (C).