Culture method of embryoid bodies and/or neural stem cells derived from human differentiated cell-derived pluripotent stem cells

Patent metadata

Field	Value
Publication number	US-10041040-B2
Application number	US-201013147558-A
Country	US
Kind code	B2
Filing date	Feb 3, 2010
Priority date	Feb 3, 2009
Publication date	Aug 7, 2018
Grant date	Aug 7, 2018

Abstract

A method for differentiating a human differentiated cell-derived pluripotent stem cell into a neural stem cell is provided, which includes the steps of: making an embryoid body from the human differentiated cell-derived pluripotent stem cell; and culturing the embryoid body in a medium containing LIF to differentiate into a neural stem cell, so that, when the neural stem cell is allowed to differentiate in vitro after multiple subculturing of the neural stem cell, it differentiate mainly into neurons but substantially not into glial cells.

First claim

The invention claimed is:

1. A method for culturing a human neural stem cell, comprising the step of providing a human neural stem cell that has been determined to express CD56 and to not express TRA-1-60 and TRA-1-81, and culturing the human stem cell in a medium containing LIF (Leukemia Inhibitory Factor).

2. The method of claim 1 , wherein the LIF concentration is from 10 to 100 ng/ml.

3. The method of claim 1 , wherein the human neural stem cell is obtained by differentiation of a human differentiated cell-derived pluripotent stem cell or a human embryonic stem cell.

4. The method of claim 1 , further comprising differentiating the cultured neural stem cell into a neuronal cell.

5. A method for obtaining a secondary or higher-order neurosphere comprising a human neural stem cell, comprising
culturing an embryoid body derived from a human pluripotent stem cell in a medium containing LIF to obtain a primary neurosphere comprising a human neural stem cell and determining that the human stem cell expresses CD56 and does not express TRA-1-60 and TRA-1-81;
culturing the primary neurosphere according to claim 1 ;
subculturing the primary neurosphere in a medium containing LIF at least one time; and
obtaining a secondary or higher-order neurosphere comprising a human neural stem cell.

6. The method of claim 5 , wherein the LIF concentration is from 10 to 100 ng/ml.

7. The method of claim 5 , wherein the human neural stem cell is obtained by differentiation of a human differentiated cell-derived pluripotent stem cell or a human embryonic stem cell.

8. The method of claim 5 , further comprising differentiating the cultured neural stem cell into a neuronal cell.

Assignees

• Okano Hideyuki
• Okada Yohei
• Nakamura Masaya
• Univ Keio

Inventors

• Okano Hideyuki
• Okada Yohei
• Nakamura Masaya

Classifications

• A61P25/28

  for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia · CPC title

• A61P25/16

  Anti-Parkinson drugs · CPC title

• A61P25/14

  for treating abnormal movements, e.g. chorea, dyskinesia · CPC title

• A61P25/00

  Drugs for disorders of the nervous system · CPC title

• A61K35/12

  Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells (vaccines or medicinal preparations containing antigens or antibodies A61K39/00) · CPC title