Primitive neural stem cells and method for differentiation of stem cells to neural cells
First Claim
1. A method for differentiating one or more pluripotent mammalian embryonic stem (ES) cells comprising:
- a. culturing the mammalian ES cells at low cell density in a serum-free and feeder-layer free media comprising leukemia inhibitory factor; and
b. allowing said mammalian ES cells to differentiate to primitive neural stem cells.
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Abstract
Described are a novel cell type in the neural lineage, and method of producing the same based on the degree of neural commitment and growth factor responsiveness in vitro and the potential to give rise to neural and non-neural progeny in vivo. The novel vell type of neural lineage and cells derived therefrom have a number of applications including applications regarding tissue engineering, transplantation and gene therapy and drug discovery. Also described are suggested uses of the method and cell type including isolating genes that positively and negatively regulate the transition from an ES cell to a neural cell and generally for studying ES cell models of mammalian neural development.
18 Citations
29 Claims
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1. A method for differentiating one or more pluripotent mammalian embryonic stem (ES) cells comprising:
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a. culturing the mammalian ES cells at low cell density in a serum-free and feeder-layer free media comprising leukemia inhibitory factor; and b. allowing said mammalian ES cells to differentiate to primitive neural stem cells. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 16, 24, 25, 26, 27, 29)
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- 13. The method of wherein the serum free media further comprises a growth factor.
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17. A method for producing secondary mammalian primitive neural stem cell colonies comprising:
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a. culturing mammalian ES cells in low cell density serum-free and feeder-layer free media comprising leukemia inhibitory factor for a time and under conditions sufficient to differentiate the said mammalian ES cells to primary primitive neural stem cell colonies; b. dissociating and subcloning the primary primitive neural stem cell colonies generated from the said mammalian ES cells; and c. administering a growth factor or survival factor to the dissociated neural cells to produce secondary primitive neural stem cell colonies. - View Dependent Claims (18, 19)
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20. A method for screening for modulators of mammalian primitive neural stem cell differentiation comprising:
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a. culturing mammalian primitive neural stem cells in serum-free and feeder-layer free media comprising leukemia inhibitory factor under low cell density conditions in the potential modulator under conditions that produce differentiation in the absence of the potential modulator; b. detecting any differentiation of the cells and cell types generated, if any, in the presence of the modulator compared to differentiation and cell types generated in the absence of the potential modulator; c. determining whether the potential modulator affects the differentiation of the cells. - View Dependent Claims (21)
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22. A method for screening for differentiation factors of cellular development comprising:
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a. culturing mammalian pluripotent embryonic stem (ES) cells in serum free media comprising leukemia inhibitory factor at low cell density in the presence of the differentiation factor; b. allowing the cells to differentiate; c. detecting differentiation of the cells, if any. - View Dependent Claims (23)
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28. A method for producing secondary mammalian primitive neural stem cell colonies comprising:
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a. culturing mammalian ES cells in low cell density serum-free and feeder-layer free media comprising leukemia inhibitory factor for a time and under conditions sufficient to differentiate the said mammalian ES cells to primary primitive neural stem cell colonies; b. dissociating and subcloning the primary primitive neural stem cell colonies generated from the said ES cells; and c. administering LW or B27 to the dissociated primary primitive neural stem cells to produce secondary primitive neural stem cell colonies.
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Specification