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Joshua M. Barnett, Ashwath Jayagopal, Gary W. McCollum, John S. Penn; Tube Formation and Proliferative Capacities of Endothelial Progenitor Cell Subpopulations: a Systematic Analysis. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3185.
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Bone marrow-derived endothelial progenitor cells (EPCs) play a significant role in the neovascular component of retinal diseases such as diabetic retinopathy and age-related macular degeneration. A variety of cell surface antigens have been shown to modulate this neovascular capacity, but little is known about the potential synergistic or competitive roles of these subpopulations in ocular pathology. This study sought to evaluate and compare different EPC subpopulations in tube formation and proliferation assays, under conditions relevant to ocular disease.
Bone marrow was isolated from 4-6 week-old Brown Norway rats. EPC subpopulations were identified by fluorescence-activated cell sorting, using antibodies directed against CD34, CD133, Tie2 and CXCR4. Distinct EPC subpopulations were labeled with quantum dot-conjugated acetylated LDL, and mature endothelial cells (ECs) were labeled with Qtracker®. Tube formation and proliferation assays were used to compare ECs and EPC subpopulations under conditions of VEGF or serum induction.
CD34+/CD133+ cells demonstrated increased tube formation relative to ECs in serum-free, VEGF and 10% serum-stimulated conditions; tube formation was increased 612% (p<0.001), 78% (p<0.01) and 67% (p<0.01), respectively. Tie2+/CD133+ cells also demonstrated increased tube formation in VEGF and serum-stimulated conditions relative to ECs; tube formation increased 43% (p<0.05) and 22% (p<0.05), respectively. The proliferative capacity of CD34+/CD133+ cells was greatest, with a 70% (p<0.01) increase in VEGF-induced proliferation and a 44% increase in serum-induced proliferation, relative to ECs. Compared to EC controls, Tie2+/CD133+ cells demonstrated a 38% reduction (p<0.05) in VEGF-induced proliferation, while the Tie2-/CD133+ population demonstrated a 17% reduction in serum-induced proliferation.
We have shown various EPC populations to exhibit different neovascular responses using two in vitro models of retinal angiogenic cell behavior. In both assays, the CD34+/CD133+ cells demonstrated the greatest neovascular capacity, suggesting that this EPC population may provide the greatest contribution to retinal disease. These results emphasize the importance of characterizing the angiogenic capacity of diverse EPC subpopulations in retinal vascular disease, as the knowledge may lead to better-targeted therapies for ocular diseases.
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