| 主权项 |
1. A method for producing a stable cell line of human neural precursor cells in vitro, comprising the steps of:
a) preparing a culture of neural precursor cells in a serum-free medium; b) culturing the neural precursor cells in the presence of a first mitogen, wherein said first mitogen is selected from the group consisting of aFGF, bFGF, EGF, TGFα and combinations thereof; c) introducing into the neural precursor cells a recombinant DNA construct comprising a receptor ligand-regulated c-myc gene, wherein the c-myc gene is fused with DNA encoding a ligand-binding domain of a nuclear receptor; d) contacting the neural precursor cells with an agent that binds the ligand-binding domain of the nuclear receptor, wherein the agent comprises a myc-activating chemical selected from the group consisting of β-estradiol, RU38486, dexamethasone, thyroid hormones, retinoids, and ecdysone; e) further culturing the neural precursor cells in a medium containing the first mitogen and a second mitogen to produce a stable cell line of neural precursor cells, wherein said second mitogen is selected from the group consisting of aFGF, bFGF, EGF, TGFα, serum and combinations thereof, with the proviso that the second mitogen is other than the first mitogen, and f) expanding the neural precursor cells of the stable cell line beyond thirty cell doublings in the medium containing the first mitogen and the second mitogen, wherein the expanded neural precursor cells of the stable cell line of neural precursor cells are capable of differentiating into neurons, astrocytes, and oligodendrocytes after removal of the first and second mitogen from the medium. |
