Development of the human retinal vasculature is poorly understood. In the past five years, we have contributed to our understanding of this process by demonstrating that the human superficial retinal vasculature forms by vasculogenesis. Vasculogenesis entails the differentiation and assembly of vascular progenitors, angioblasts, into primordial blood vessels. We have cultured human retinal angioblasts (CXCR4+, cKit+, CD39+) from avascular neonatal dog and fetal human retina. Our overall hypothesis is that cytokines cause differentiation of these retinal angioblasts into endothelial cells (EC) and pericytes and regulate their assembly via vasculogenesis to form the superficial retinal vasculature. Furthermore, we propose that Muller and/or ganglion cells (GC) provide the guidance for this assembly. One goal of these studies is to characterize human retinal angioblasts in vitro and investigate the potential of these human cells to differentiate into various cell types found in inner retina. We will also determine the stimulus for their formation into blood vessels in vitro. We will determine the effects of VEGF165 and VEGF165b on retinal angioblast differentiation and tube formation in vitro and determine why VEGF165b is associated with nucleus of progenitors while VEGF165 is cytoplasmic and diffusely in the perivascular milieu. We will determine the cell types and extracellular matrix components that potentially provide guidance for assembly of retinal angioblasts into the fetal human retinal vasculature in vivo by immunohistochemistry and in vitro using GC and Muller cells co-cultured with angioblasts in 3-D gels. Coated nanofibers will be used to examine the influence of individual matrix components and guidance molecules (neuropilin-1, astrocytes 3A) on angioblast alignment, patterning, and assembly of tubes. These studies will broaden our knowledge of human vascular development and characterize the pertinent progenitors. It appears from the literature that the mouse superficial vasculature develops by angiogenesis and guidance for vascular assembly is provided by an astrocytic template. There is no strocytes template in advance of the superficial human retinal vasculature while developing by vasculogenesis. Mice deficient in superficial retinal blood vessels, GC or Muller cells will be used to demonstrate functionally the importance of these cells to retinal vascular development with the ultimate outcome being delineation of which features in mouse retinal vascular development can actually be transcribed to human retinal vascular development. Finally, the potential for therapeutic use of angioblasts in the initial stage of OIR will also be determined using human angioblasts from different sources (including clinically relevant iPS from cord blood cells) and magnetic nanoparticles to increase their homing and engraftment to obliterated retinal vascular segments. If acellular vascular segments can be quickly repopulated and the blood vessels regenerated, then vaso- obliteration will be limited and subsequent neovascularization could be suppressed.

Public Health Relevance

This study will broaden our knowledge of human vascular development by characterizing the pertinent progenitors (angioblasts) and determining the cues needed to assemble the human superficial retinal vasculature. The potential of human angioblasts derived from several sources (including induced-pluripotent cells from cord blood) to be used for regenerative medicine (repopulating acellular vascular segments) will be determined using the mouse oxygen-induced retinopathy model for retinopathy of prematurity (ROP). If we want to effectively treat ROP, which is an oxidative insult of a developing vasculature, we must understand how the human retinal vasculature forms and investigate the potential for using autologous cells (angioblasts from the persons own cord blood) as a treatment.

Agency
National Institute of Health (NIH)
Institute
National Eye Institute (NEI)
Type
Research Project (R01)
Project #
5R01EY009357-19
Application #
8436246
Study Section
Special Emphasis Panel (ZRG1-CB-G (90))
Program Officer
Shen, Grace L
Project Start
1992-12-01
Project End
2015-03-31
Budget Start
2013-04-01
Budget End
2014-03-31
Support Year
19
Fiscal Year
2013
Total Cost
$448,844
Indirect Cost
$175,158
Name
Johns Hopkins University
Department
Ophthalmology
Type
Schools of Medicine
DUNS #
001910777
City
Baltimore
State
MD
Country
United States
Zip Code
21218
Park, Tea Soon; Bhutto, Imran; Zimmerlin, Ludovic et al. (2014) Vascular progenitors from cord blood-derived induced pluripotent stem cells possess augmented capacity for regenerating ischemic retinal vasculature. Circulation 129:359-72
Edwards, Malia M; Rodríguez, José J; Gutierrez-Lanza, Raquel et al. (2014) Retinal macroglia changes in a triple transgenic mouse model of Alzheimer's disease. Exp Eye Res 127:252-60
Rodrigues, Murilo; Xin, Xiaoban; Jee, Kathleen et al. (2013) VEGF secreted by hypoxic Muller cells induces MMP-2 expression and activity in endothelial cells to promote retinal neovascularization in proliferative diabetic retinopathy. Diabetes 62:3863-73
Edwards, Malia M; Lefebvre, Olivier (2013) Laminins and retinal vascular development. Cell Adh Migr 7:82-9
Edwards, Malia M; McLeod, D Scott; Li, Renzhong et al. (2012) The deletion of Math5 disrupts retinal blood vessel and glial development in mice. Exp Eye Res 96:147-56
McLeod, D Scott; Baba, Takayuki; Bhutto, Imran A et al. (2012) Co-expression of endothelial and neuronal nitric oxide synthases in the developing vasculatures of the human fetal eye. Graefes Arch Clin Exp Ophthalmol 250:839-48
Baba, Takayuki; McLeod, D Scott; Edwards, Malia M et al. (2012) VEGF 165 b in the developing vasculatures of the fetal human eye. Dev Dyn 241:595-607
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McLeod, D Scott; Ying, Howard S; McLeod, Colin A et al. (2011) Retinal and optic nerve head pathology in Susac's syndrome. Ophthalmology 118:548-52
Lutty, Gerard A; McLeod, D Scott; Bhutto, Imran et al. (2011) Effect of VEGF trap on normal retinal vascular development and oxygen-induced retinopathy in the dog. Invest Ophthalmol Vis Sci 52:4039-47

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