Method for producing myocardial sheet from embryonic stem cell

Patent metadata

Field	Value
Publication number	US-9623052-B2
Application number	US-201214009018-A
Country	US
Kind code	B2
Filing date	Mar 30, 2012
Priority date	Mar 30, 2011
Publication date	Apr 18, 2017
Grant date	Apr 18, 2017

Abstract

The present invention provides a method for producing a myocardial sheet using a group of cells derived from embryonic stem cells. This method is characterized by mixing Flk/KDR positive cells, cardiomyocytes, endothelial cells, and mural cells, all derived from embryonic stem cells, and culturing the mixed cells. Furthermore, the myocardial sheet can be used as a therapeutic agent for heart diseases since VEGF is released from the sheet.

First claim

The invention claimed is:

1. A method for producing a myocardial sheet, comprising the following steps (a) to (c):
(a) producing Flk/DR positive cells, cardiomyocytes, endothelial cells, and mural cells from embryonic stem cells; and
(b) mixing the Flk/KDR positive cells with the cardiomyocytes, endothelial cells, and mural cells to obtain a cell mixture of FLk/KDR positive cells, cardiomyocytes, endothelial cells, and mural cells; and
(c) forming a sheet by culturing the mixed cells obtained in step (b) in a culture medium comprising VEGF in a culture vessel coated with a temperature-responsive polymer.

2. The method according to claim 1 , wherein the Flk/KDR positive cells are induced by culturing embryonic stem cells on a gelatin-coated culture vessel.

3. The method according to claim 1 , wherein in the step (a), the cardiomyocytes are produced by culturing the Flk/KDR positive cells in a state in which the Flk/KDR positive cells are contacted with cyclosporin A.

4. The method according to claim 1 , wherein in the step (a), the endothelial cells and the mural cells are produced by culturing the Flk/KDR positive cells in a state in which the Flk/KDR positive cells are contacted with VEGF and cAMP.

5. The method according to claim 1 , wherein in the step (b), the Flk/KDR positive cells are cultured for 1 to 7 days and then mixed with cardiomyocytes, endothelial cells, and mural cells.

6. The method according to claim 1 , further comprising a step of laminating the sheet to at least one additional myocardial sheet produced according to the same method to form a laminated sheet.

7. The method according to claim 6 , wherein the laminated sheet consists of three layers of myocardial sheets.

Assignees

• Okano Teruo
• Shimizu Tatsuya
• Yamashita Jun
• Masumoto Hidetoshi
• Tokyo Women'S Medical Univ
• Univ Kyoto

Inventors

• Okano Teruo
• Shimizu Tatsuya
• Yamashita Jun
• Masumoto Hidetoshi

Classifications

• A61P9/00

  Drugs for disorders of the cardiovascular system · CPC title

• A61P43/00

  Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00 · CPC title

• A61K9/70

  Web, sheet or filament bases {; Films; Fibres of the matrix type containing drug (hollow drug-filled fibres A61K9/0092)} · CPC title

• A61L27/54

  Biologically active materials, e.g. therapeutic substances {(A61L27/227 takes precedence)} · CPC title

• C12N2502/1329

  Cardiomyocytes · CPC title