Method for Producing Culture Containing Megakaryocytes, and Method for Producing Platelets Using Same

Abstract

The present invention provides a method for producing megakaryocytes that have an increased capacity to produce platelets, the method comprising a step of culturing, under conditions that cause the death of cells that do not express a gene that is specifically expressed by megakaryocytes, cells that have the capacity to differentiate into megakaryocytes.

1 . A method for producing a culture containing megakaryocytes or megakaryocyte progenitor cells, the method comprising:
 a step of culturing, under conditions that cause the death of cells that do not express a gene that is specifically expressed by megakaryocytes, a cell group that contains megakaryocytes or cells having the capacity to differentiate into megakaryocytes.   
     
     
         2 . The method according to  claim 1 , wherein the cell group has been transformed by a gene for resistance to a drug that exhibits cytotoxicity, the drug resistance gene being positioned under the control of a promoter of the gene that is specifically expressed by megakaryocytes, and the drug being added in the culture step. 
     
     
         3 . The method according to  claim 1 , wherein the cell group has been transformed by a gene that causes the death of the cells that do not express the gene that is specifically expressed by megakaryocytes, and this gene is positioned under the control of a promoter of a gene that is specifically expressed by the cells that do not express the gene that is specifically expressed by megakaryocytes. 
     
     
         4 . The method according to  claim 1 , wherein the gene that is specifically expressed by megakaryocytes is a gene that codes for a cell surface marker that is specifically expressed by megakaryocytes. 
     
     
         5 . The method according to  claim 4 , wherein the cell surface marker that is specifically expressed by megakaryocytes is CD41a, CD42a, and/or CD42b. 
     
     
         6 . The method according to  claim 1 , wherein the cells having the capacity to differentiate into megakaryocytes are at least one type selected from the group consisting of hematopoietic stem cells, hematopoietic progenitor cells, CD34-positive cells, and megakaryocyte progenitor cells. 
     
     
         7 . The method according to  claim 1 , wherein the culture step is carried out in the absence of serum and/or feeder cells. 
     
     
         8 . The method according to  claim 1 , wherein the megakaryocytes in the culture to be produced retain the differentiation phenotype thereof, and have an increased platelet production capacity. 
     
     
         9 . The method according to  claim 2 , wherein the cell group is transformed by a vector in which the promoter and the drug resistance gene or a lethal gene are operably linked. 
     
     
         10 . The method according to  claim 2 , wherein the drug resistance gene is at least one type selected from the group consisting of a puromycin resistance gene, a neomycin resistance gene, a kanamycin resistance gene, a chloramphenicol resistance gene, an erythromycin resistance gene, a tetracycline resistance gene, a hygromycin resistance gene, an ampicillin resistance gene, a zeocin resistance gene, a blasticidin S resistance gene, and a histidinol resistance gene. 
     
     
         11 . A method for producing platelets, the method using megakaryocytes produced by the method described in  claim 1 . 
     
     
         12 . A culture expansion method for megakaryocytes or megakaryocyte progenitor cells, the method comprising:
 a step of culturing, under conditions that cause the death of cells that do not express a gene that is specifically expressed by megakaryocytes, a cell group that contains megakaryocytes or cells having the capacity to differentiate into megakaryocytes.   
     
     
         13 . The method according to  claim 12 , wherein the cell group has been transformed by a gene for resistance to a drug that exhibits cytotoxicity, the drug resistance gene being positioned under the control of a promoter of the gene that is specifically expressed by megakaryocytes, and the drug being added in the culture step. 
     
     
         14 . A culture obtained by the production method described in  claim 1 , and comprising megakaryocyte or megakaryocyte progenitor cells.