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Sahar Balagholi, Sedigheh Amini Kafiabadi, Zahra-Soheila Soheili, Mozhgan Rezaeikanavi, Shahram Samiei; Investigating the Effect of PAX6 Gene Expression on Differentiation of CD133+Human Cord Blood Stem Cells. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3581. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
To assess the influence of PAX6 gene expression on differentiation of CD133+ human cord blood stem cells.
Human cord blood mononuclear cells were isolated by ficoll. CD133+ cells were obtained using the CD133MicroBead Kit in combination with the autoMACS Separator and then cultured in the Stem Span media. HEK293T packaging cells were co-transfected with PLEX-MCS, PsPAX2, and pMD2G via calcium phosphate method. The resultant lentiviral vectors were collected and concentrated by PEG 6000. The appropriate amount of viruses was used for infecting CD133+ cells. The successful transduced cells were selected by puromycin resistance. After two weeks, the expression of rhodopsin, CHX10, Thy1, nestin, and PAX6 proteins were assayed by immunocytochemistry.
Two weeks after transfection, CD133+ cells expressed rhodopsin, CHX10, Thy1, nestin, and PAX6 proteins, as compared to the HEK293T cells that expressed rhodopsin and nestin.
Transfected CD133+ human cord blood stem cells with PAX6 gene are able to differentiate into progenitor retinal neural- and ganglion-like cells.
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