Compositions and methods for propagation of neural progenitor cells
First Claim
Patent Images
1. A cell culture comprising:
- (a) a culture medium, wherein the calcium concentration of the medium is about 0.03 mM to 0.15 mM;
(b) about 20-100 ng/ml epidermal growth factor (EGF);
(c) about 10-50 ng/ml basic fibroblast growth factor (bFGF);
(d) about 1-150 ng/ml transforming growth factor-alpha (TGFα
);
(e) mammalian neural progenitor cells (NPC) immunopositive for Oct4 and nestin, and wherein the cells continue to proliferate for at least 1 year in vitro.
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Abstract
Compositions and methods for the culturing, propagation, cryopreservation and manipulation of neural progenitor cells (NPC) and pluripotent stem cells (PSC) are provided. The cells exhibit rapid doubling times and can be maintained in vitro for extended periods. Also provided is a method of propagating neural progenitor cells, and a method of transplanting human NPC and/or PSC to a host. The cells can be genetically modified to express a therapeutic agent prior to the transplanting.
41 Citations
29 Claims
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1. A cell culture comprising:
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(a) a culture medium, wherein the calcium concentration of the medium is about 0.03 mM to 0.15 mM; (b) about 20-100 ng/ml epidermal growth factor (EGF); (c) about 10-50 ng/ml basic fibroblast growth factor (bFGF); (d) about 1-150 ng/ml transforming growth factor-alpha (TGFα
);(e) mammalian neural progenitor cells (NPC) immunopositive for Oct4 and nestin, and wherein the cells continue to proliferate for at least 1 year in vitro. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 19, 20, 21, 22, 23, 24, 25, 26)
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17. A method of propagating NPC that are immunopositive for nestin and Oct4, comprising culturing primary human fetal brain tissue in a culture medium, wherein the culture medium comprises:
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(a) 0.03 to 0.09 mM calcium; (b) about 20-100 ng/ml epidermal growth factor (EGF); (c) about 10-50 ng/ml fibroblast growth factor basic (bFGF); and (d) about 1-150 ng/ml transforming growth factor-alpha (TGFα
),wherein the cells continue to proliferate for at least 1 year in vitro. - View Dependent Claims (18)
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27. A cell culture comprising human neural progenitor cells (NPC) that are immunopositive for nestin and Oct4 suspended in a medium, the medium consisting of:
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(a) a culture medium, wherein the calcium concentration of the medium is about 0.03 mM to about 0.09 mM; (b) about 20-100 ng/ml epidermal growth factor (EGF); (c) about 10-50 ng/ml basic fibroblast growth factor (bFGF); (d) about 1-150 ng/ml transforming growth factor-alpha (TGFα
);(e) about 7-30 ng/ml leukemia inhibiting factor (LIF); and
, optionally;(f) 2% B27 supplement; (g) about 0.11 mg/ml;
sodium pyruvate.
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28. A method of transplanting human NPC to a mammalian host, comprising:
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(a) obtaining a cell culture of undifferentiated NPC that are immunopositive for nestin and Oct4 suspended in a culture medium comprising; (i) about 20-100 ng/ml EGF; (ii) about 10-50 ng/ml bFGF; (iii) about 1-150 ng/ml TGFα
;(iv) about 0.03 mM to 0.15 mM calcium; and (b) transplanting the undifferentiated cell culture to the central nervous system (CNS) of the host, wherein the host has Parkinson'"'"'s disease or epilepsy.
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29. A method of transplanting human NPC to a mammalian host, comprising:
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(a) obtaining a cell culture of NPC that are immunopositive for nestin and Oct4 suspended in a culture medium comprising; (i) about 20-100 ng/ml EGF; (ii) about 10-50 ng/ml bFGF; (iii) about 1-150 ng/ml TGFα
; and(iv) about 0.03 mM to 0.15 mM calcium; and (b) transplanting the cell culture to the central nervous system (CNS) of the host, wherein the host has Parkinson'"'"'s disease or epilepsy; wherein the NPC are not genetically modified.
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Specification