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J. Yang, H. Klassen, M. Pries, W. Wang, M. H. Nissen; Vitreous Humor Enhances the Proliferation of Retinal Precursor Cells. Invest. Ophthalmol. Vis. Sci. 2007;48(13):4075.
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Intravitreal injection is an important delivery route for studies involving the transplantation of various types of precursor cells to the retina, however, the effect on these cells of exposure to the vitreous microenvironment has not been specifically investigated. Here vitreous humor was evaluated for the potential to stimulate the proliferation of rat retinal precursor cells (RPCs) in vitro.
Rat RPCs were isolated at embryonic day 19 and cultured in DMEM:F12 medium containing N2 supplement and 20ng/ml EGF in the presence or absence of fluid expressed from porcine vitreous humor. Cellular proliferation at different concentrations of vitreous fluid supplementation was quantified from day 0 to day 4 using a 3H-thymidine incorporation assay. Active components of vitreous fluid were partially characterized by gel filtration chromatography (GFC) and UV spectral analysis. The effect of each vitreous fraction on RPC proliferation was then determined as well.
Addition of 20% vitreous fluid to primary rat RPC cultures increased 3H-thymidine incorporation by as much as 481%, as compared to growth medium without vitreous supplementation. One of the vitreous fractions showing growth promoting activity was localized to the low molecular weight range < 1000 Da, consistent with ascorbic acid. The presence of ascorbic acid in the vitreous fluid was confirmed by UV spectral analysis.
These results show that vitreous fluid enhances the proliferation of rat RPCs in vitro. Ascorbic acid likely contributes to this effect in that we have previously shown this molecule to be pro-proliferative for this cell type. Because vitreous-associated enhancement of RPC proliferation remains a growth-factor-dependent phenomenon, the proliferative status of transplanted cells in the vitreous cavity is likely determined by a combination of these and other factors.
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