Abstract
Purpose:
To assess the influence of PAX6 gene expression on differentiation of CD133+ human cord blood stem cells.
Methods:
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.
Results:
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.
Conclusions:
Transfected CD133+ human cord blood stem cells with PAX6 gene are able to differentiate into progenitor retinal neural- and ganglion-like cells.