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X. Zhang; Characterization of Retinal Stem Cells and Their Math5-Induced Differentiation by Lentivirus-Mediated Gene Transfer. Invest. Ophthalmol. Vis. Sci. 2010;51(13):729.
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RSCs were isolated from neural retina of postnatal day 10 mice and grown as neurospheres in a defined serum-free culture medium. math5 coding sequence was subcloned into feline immunodeficiency virus (FIV) vector with N-terminal c-myc tag. RSCs were infected with FIV expressing c-myc-tagged math5 or control c-myc FIV. The expression of math5, undifferentiated progenitor-specific marker Nestin and neuron-specific DNA-binding protein NeuN (Neuronal Nuclei) were characterized in RSCs by immunocytochemistry using marker-specific antibodies.
c-myc expression was detected by immunocytochemistry in FIV-infected RSCs, suggesting that RSCs were successfully infected by FIV with the expression of c-myc-math5. Immunocytochemical analysis revealed that both FIV-math5-infected and mock infected RSCs expressed stem cell marker Nestin. However, NeuN, a neuron-specific marker, was expressed only in the RSCs infected with math5, but not in mock infected RSCs.
These results demonstrate that expression of math5 induces NeuN expression in RSCs, suggesting that math5 promotes RSC differentiation toward RGCs by upregulating neuron-specific gene expression. This study also reveals that virus-mediated gene transfer can facilitate the investigation of gene-specific regulation of RSC differentiation.
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