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V. Marchetti, E. Aguilar, S. Naccache, G. Nemerow, D. Friedlander, M. Friedlander; Human Cord Blood Derived CD14+ Cells Facilitate Retinal Vascular Repair in Oxygen Induced Retinopathy. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4378.
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Human umbilical cord blood (HCB) is a rich source of stem cells that can differentiate into hematopoietic (HSC) or endothelial (EPC) cell lineages. EPCs may also be derived from the myeloid lineage of HSC. We investigated whether HCB myeloid/monocytic populations could differentiate in endothelial cells and participate in retinal vascularization.
Monocytic CD14+ cells were isolated from an HCB mononuclear cell fraction and incubated on fibronectin-coated dishes in endothelial cell medium. After 7 days the cells were analyzed by flow cytometry for the expression of myeloid and endothelial markers. The ability of the CD14+ cells to form tubes and network was tested in an in vitro matrigel assay. CD14+ cells infected with Adenovirus 5 GFP were injected on p7 in the vitreous of mice under hyperoxia conditions (75% O2) and visualized in the retina by confocal microscopy. Rescue was evaluated on p17 quantifying areas of vaso-obliteration and neovascularization (NV). Cytokine production was analyzed on p11 by analyzing total retinal lysed using an antibody-sandwich angiogenesis array (Panomics, Angiogenesis Antibody Array).
On day 0 CD14+ cells express both the myeloid progenitor surface marker, CD33, and specific endothelial antigen VEGFR-2, and form cord- and tube-like structures in vitro. The CD14+ cells form endothelial colonies between day 4-7. By day 13 these cells demonstrate characteristics of differentiated endothelial cells including receptor and enzyme expression, as well as formation of intercellular junctions. Using the mouse OIR model, we show that both freshly isolated and in vitro differentiated CD14+ can target sites of retinal NV and significantly enhance vascular repair. Intravitreal injection of these cells reduces vascular obliteration and preretinal NV by over 50% when compared to vehicle injection. Injection with CD14+ cells induces the expression of mouse angiogenic cytokines such as IL-1α, FGF-a and b, IL-6 and TGFα in the OIR retinas.
We have identified and isolated a monocytic population of cells derived from human CB that can target retina vasculature and provide trophic rescue in a model of hypoxic retinopathy. These cells may be visualized in the retina using Ad5 F16 vectors. Such visualization may permit the study and characterization of CD14+ cells function and differentiation in the retina. The use of Adenoviral vectors encoding various trophic factors could increase the trophic activity of CB derived CD14+, providing more efficient and safer methods of ocular gene delivery.
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