Method for direct reprogramming of urine cells into neural stem cells using synthetic MRNA

A method for direct reprogramming of urine cells into neural stem cells, wherein direct reprogramming involves reprogramming of urine cells into neural stem cells without the intermediate generation of induced pluripotent stem cells (iPSCs), the method comprising:

• (a) isolating urine cells from urine and culturing the urine cells;
• (b) introducing into the cultured urine cells a composition comprising:
• (i) an Oct4 protein or a nucleic acid encoding the Oct4 protein;
• (ii) a Sox2 protein or a nucleic acid encoding the Sox2 protein;
• (iii) a Klf4 protein or a nucleic acid encoding the Klf4 protein;
• and
• (iv) a Glis1 protein or a nucleic acid encoding the Glis1 protein;
• (c) inducing direct reprogramming by culturing urine cells into which the composition has been introduced in a neural stem cell-inducing medium;
• (d) selecting a neural stem cell(s) expressing SOX1, SOX2, EN1 and PAX6 from the cultured urine cells in the neural stem cell-inducing medium, and
• wherein in step (c), the neural stem cell-inducing medium is obtained by mixing a DMEM/F12 medium and a neurobasal medium at a volume ratio of 1:1 and adding N2, B27, human LIF, a TGF-beta inhibitor, and a GSK3-beta inhibitor to the medium mixture.