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Malia M. Edwards, D S. McLeod, Imran A. Bhutto, Carol Merges, Takayuki Baba, Vikash Juriasinghani, Gerard A. Lutty; Angioblasts Differentiate Into Endothelial Cells Or Pericytes In Vitro But Favor A Pericyte Position When Injected Intravitreally. Invest. Ophthalmol. Vis. Sci. 2012;53(14):2992.
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Cultured neonatal canine retinal angioblasts (NCRA) differentiate into endothelial cells (EC) when given basic fibroblast growth factor (bFGF) or pericytes in the presence of platelet derived growth factor BB (PDGF-BB) . The present study investigated tube formation by these differentiated cells using 3-D collagen gels. The influence of different growth factors on tube formation was investigated as was the position of cells when co-cultured. The incorporation of NCRA into developing vessels and neovascular tufts was also investigated using a model for oxygen induced retinopathy.
NCRA were cultured and trained from neonatal canine retinas as previously described  and labeled with PKH dyes (26 or 67) according to the manufacturer’s instructions. For the collagen tube assay, labeled cells were mixed with collagen solution and plated into a 96 well plate . After 1 hr, media containing different growth factors was added and cells were imaged every 24 hrs. The incorporation of differentiated angioblasts into developing retinal vessels was tested by injecting labeled cells intravitreally into post-natal day (P) 2 dogs. Animals were sacrificed at P5, retinas dissected and stained with anti-von Willebrands Factor (vWF). The localization of cells in relation to blood vessels was observed. For the oxygen-induced retinopathy studies, pups were exposed to 100% oxygen for 5 days prior to the injection of trained angioblasts at P8. At P21, pups were sacrificed and the vitreous and retinas labeled with anti-vWF.
NCRA trained with bFGF but not PDGF-BB formed tubes. Tube formation by bFGF-trained cells was enhanced by VEGF and bFGF. When the two cell types were cultured together, the bFGF-trained cells formed tubes and the PDGF-BB-trained cells were observed almost exclusively on the outside walls of these tubes.Both types of NCRA injected into the developing canine vitreous were found in a pericyte position at P5. Similarly, trained cells injected following exposure to oxygen took a pericyte position on intravitreal neovascularization and retinal blood vessels.
The present study demonstrates that cultured canine angioblasts trained to be either pericytes or endothelial cells function as these cell types during in vitro tube formation but favored a pericytic position in vivo.1. Lutty, G. et al., Exp Eye Res, 2006. 83: 183.2. Liu H. et al Angiogenesis, 2008. 11:223.
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