Notch signaling in ocular vasculature development and diseases.

Abstract

Ocular angiogenesis, characterized by the formation of new blood vessels in the avascular area in eyes, is a highly coordinated process involved in retinal vasculature formation and several ocular diseases such as age-related macular degeneration, proliferative diabetic retinopathy and retinopathy of prematurity. This process is orchestrated by complicated cellular interactions and vascular growth factors, during which endothelial cells acquire heterogeneous phenotypes and distinct cellular destinations. To date, while the vascular endothelial growth factor has been identified as the most critical angiogenic agent with a remarkable therapeutic value, the Notch signaling pathway appears to be a similarly important regulator in several angiogenic steps. Recent progress has highlighted the involvement, mechanisms and therapeutic potential of Notch signaling in retinal vasculature development and pathological angiogenesis-related eye disorders, which may cause irreversible blindness.

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Notch signaling in ocular vasculature development and diseases

Guo-Rui Dou1,2, Lin Wang2, Yu-Sheng Wang1,*, and Hua Han2,*
1Department of Ophthalmology, Xijing Hospital, and 2Department of Medical Genetics and
Developmental Biology, Fourth Military Medical University, Xi’an 710032, China

*Correspondence Authors: Yu-Sheng Wang, Department of Ophthalmology, Xijing
Hospital, Fourth Military Medical University, Xi’an 710032, China, Tel/Fax +86 29 847
753 71, E-Mail: wangys@fmmu.edu.cn. Or Hua Han, Department of Medical Genetics and
Developmental Biology, Fourth Military Medical University, Xi’an 710032, China,
Tel/Fax +86 29 847 744 87, E-Mail: huahan@fmmu.edu.cn.

Running Title: Notch signaling in ocular angiogenesis









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Molecular Medicine
www.molmed.org

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ABSTRACT
Ocular angiogenesis, characterized by the formation of new blood vessels in the avascular
area in eyes, is a highly coordinated process involved in retinal vasculature formation and
several ocular diseases such as age-related macular degeneration (AMD), proliferative
diabetic retinopathy (PDR) and retinopathy of prematurity (ROP). This process is
orchestrated by complicated cellular interactions and vascular growth factors, during which
endothelial cells acquire heterogeneous phenotypes and distinct cellular destinations. To
date, while vascular endothelial growth factor has been identified as the most critical
angiogenic agent with a remarkable therapeutic value, the Notch signaling pathway appears
to be a similarly important regulator in several angiogenic steps. Recent progress has
highlighted the involvement, mechanisms, and therapeutic potential of Notch signaling in
retinal vasculature development and pathological angiogenesis-related eye disorders, which
may cause irreversible blindness.
Key words: Notch signaling, ocular angiogenesis, vascular endothelial growth factor,
endothelial cells

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Molecular Medicine
www.molmed.org

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INTRODUCTION
Retinal vasculature and choroidal vasculature, as two major portions of the ocular
vasculature, constitute a finely ordered vascular system that guarantees blood supply to the
highly oxygen-consuming retina. The primary function of the choroidal vasculature is to
provide nutrients and oxygen to the outer retina, whereas the retinal vascular network
supplies the inner retina. The physiological functions of the retina are confined by this
normal ocular vasculature, which is established primarily by ocular angiogenesis in late
embryogenesis (man) or early postnatal days (mouse) (1). However, the anatomical and
physiological properties of posterior ocular segments make the ocular vasculature
vulnerable to exogenous and endogenous insults, leading to pathological ocular
angiogenesis and vision loss in a number of eye diseases (2), such as exudative age-related
macular degeneration (AMD) and proliferative diabetic retinopathy (PDR). Systemic
reasons or local microenvironmental factors in these diseases stimulate abnormal
neovascular growth from the existing retinal capillaries in and out of the inner surface of
the retina (retinal neovascularization, RNV) (1), and from the choroidal vessels breaking
into Bruch’s membrane beneath the retinal pigment epithelium (RPE) or neural retina
(choroidal neovascularization, CNV) (3). PDR is the leading cause of irreversible blindness
in working age adults (4), with 40,000 new patients per year in 16 million diabetic
Americans (5). AMD is considered as the major reason for blindness in elderly populations,
which affects as many as 15 million Americans with 200,000 new cases each year. In AMD
patients, although exudative AMD with the common complication of CNV constitutes only
10% of AMD, it accounts for nearly 90% of blindness in AMD patients. Retinopathy of
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prematurity (ROP), in contrast, occurs in premature infants with the disruption of retinal
vascular development due to exposure to hyperoxic circumstance, and is one of the most
common causes of childhood blindness worldwide (6). An inadequate supply of angiogenic
factors in the early stages of retinal vasculature formation in ROP results in
vaso-obliterative changes followed by compensatory new vessels growing between
vascularized and avascularized areas in the retina. These pathological vessels can induce
fibrotic scarring in the retina, vitreous, and lens.
Anti-angiogenic therapies have already been successfully used in treating these ocular
diseases (5). The currently representative therapy is the neutralization of vascular
endothelial growth factor (VEGF) activity (6). VEGF is not only required for retinal and
choroidal vessel formation during developmental stages, but also for the maintenance of
these capillaries after birth (7, 8). More importantly, distorted expression of VEGF in
pathological settings is the key stimulus to abnormal vessel growth (6). Blocking VEGF
signaling by using VEGF aptamer, neutralizing anti-VEGF antibody, or soluble VEGF
receptor (VEGFR) 1 (sVEGFR1 or VEGF trap), has been shown not only to stop the
pathological angiogenic process, but also to improve vision in a great proportion of AMD
patients during a 2-year follow-up (6). However, this field is now facing the challenge of
drug resistance and recurring cases while using these agents. Further investigation of
alternative signals regulating ocular angiogenesis might elicit potentially new therapeutic
targets. Recent intensive studies, mainly based on models of ocular angiogenesis and
gene-modified mice, have revealed the significance of Notch signaling in
angiogenesis-associated ocular diseases.
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Molecular Medicine
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CELLULAR AND MOLECULAR ASPECTS OF OCULAR ANGIOGENESIS
The retinal vasculature in mice develops as angiogenic sprouts protruding from the optic
disc immediately after birth. During the first postnatal week, sprouts from retinal vessels
constantly extend over the superficial retinal layer to form a two-dimensional vascular
network. Then the retinal vessels continue to sprout and penetrate into deeper retinal layers,
finally leading to the formation of a three-layered vascular system by the end of the third
postnatal week (9). Sharing the common steps and molecular mechanisms with
angiogenesis in the whole body, the postnatal retinal development in mice provides an ideal
platform to angiogenesis studies on imaging and intervention without the difficulties
associated with investigating embryonic development.
The primary signal initiating retinal angiogenesis is VEGF, which is secreted by retinal
astrocytes and forms a radial gradient of concentration to guide the direction of vessel
sprouts (12). In response to VEGF, quiescent endothelial cells (ECs) differentiate into
subpopulations with coordinating cellular behaviors to fulfill the angiogenic process. Tip
cells are specialized ECs located at the tips of angiogenic sprouts, and respond to the VEGF
gradients by migration to guide the sprout extending through the extracellular matrix
(ECM). Stalk cells are located behind tip cells in a sprout, and respond to VEGF by
proliferation, while being responsible for ECM production and lumen formation. A third
type of EC in angiogenesis is phalanx cells, which do not proliferate but align and form a
smooth monolayer of cells. Thus, during angiogenesis, in each new vascular sprout, a
single tip cell pioneers an angiogenic pathway by using filopodia to guide a sprouting
vessel to follow the VEGF gradient. This pioneering tip cell is highly motile but
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Molecular Medicine
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