Medium composition comprising neuropeptide Y for the generation, maintenance, prolonged undifferentiated growth of pluripotent stem cells and method of culturing pluripotent stem cell using the same

• US 8,628,963 B2
• Filed: 06/16/2010
• Issued: 01/14/2014
• Est. Priority Date: 06/16/2009
• Status: Active Grant

• Current Assignee: Korea Research Institute of Bioscience & Biotechnology
• Original Assignee: Korea Research Institute of Bioscience & Biotechnology
• Inventors: Cho, Yee Sook, Son, Mi-Young
• Primary Examiner(s): BERTOGLIO, VALARIE E
• Application Number:
  US13/378,912
• Publication Number:
  US 20120171766A1
• Time in Patent Office: 1,308 Days
• Field of Search: None
• US Class Current: 435/384
• CPC Class Codes: C12N 2501/15 Transforming growth factor beta (TGF-β)
• CPC Class Codes: C12N 2501/835 Neuropeptide Y [NPY]; Peptide YY [PYY]
• CPC Class Codes: C12N 5/0606 Pluripotent embryonic cells, e.g. embryonic stem cells [ES] embryonic germ cells C12N5/0611, induced pluripotent stem cells C12N5/0696
• CPC Class Codes: C12N 5/0696 Artificially induced pluripotent stem cells, e.g. iPS

Abstract

The present invention relates to a medium composition comprising neuropeptide Y, effective for proliferation and maintenance of undifferentiated pluripotent stem cells, and a method for culturing undifferentiated pluripotent stem cells using the same. The present invention improves the culture conditions for undifferentiated pluripotent stem cells, and ultimately, the present invention can be effectively used for the development of large-scale culture systems, thereby acquiring clinically applicable pluripotent stem cells. Further, the present invention relates to a dedifferentiation medium composition comprising neuropeptide Y (NPY), and a method for inducing dedifferentiation (or reprogramming) using the same. The present invention improves the culture conditions for dedifferentiation and contributes to develop technology of producing clinically applicable induced pluripotent stem cells, thereby being used for the development of stem cell therapy.

Claims

1. A method for maintaining undifferentiated human pluripotent stem cells, comprising culturing human pluripotent stem cells in a culture medium comprising neuropeptide Y whereby the undifferentiated pluripotent stem cells are maintained.

2. The method according to claim 1, wherein the pluripotent stem cells are embryonic stem cells or induced pluripotent stem cells.

3. The method according to claim 1, wherein the maintenance of the undifferentiated pluripotent stem cells is confirmed by the increased expression of one or more genes selected from the group consisting of ALP (alkaline phosphatase), OCT4, SOX2, hTERT (human telomerase reverse transcriptase), TDGF (teratocarcinoma-derived growth factor) and SSEA-4 as compared to the level of expression of the one or more genes in pluripotent cells cultured in the absence of neuropeptide Y.

4. The method according to claim 1, wherein the concentration of the neuropeptide Y is in the range of 0.01 to 100 μ

M.

5. The method according to claim 1, wherein the culture medium further comprises one or more factors selected from the group consisting of N2 supplement, B27 supplement, bFGF and TGFβ

without animal serum and feeder cell-derived factors.

6. The method according to claim 5, wherein the N2 and B27 supplements are provided in a ratio of 1:

1, and bFGF and TGFβ

are provided at a concentration of 4-100 ng/ml and 1-10 ng/ml, respectively.

7. An in vitro cell culture comprising undifferentiated human pluripotent stem cells cultured according to the method of claim 1.

8. The in vitro cell culture according to claim 7, wherein the pluripotent stem cells are embryonic stem cells or induced pluripotent stem cells.