Composition comprising cell and biocompatible polymer

Abstract

It is an object of the present invention to provide a cell-containing composition capable of suppressing the outflow of the cells after transplantation and improving the survival rate of the cells. The present invention provides a composition which comprises any of bone marrow stromal cell-derived neural precursor cells, bone marrow stromal cell-derived Schwann cells, or bone marrow stromal cell-derived skeletal muscle cells; and a biocompatible polymer.

1 . A composition which comprises: (A) any cells of the following (a), (b) and (c), and (B) a biocompatible polymer:
 (a) bone marrow stromal cell-derived neural precursor cells which are obtained by (1) introducing a nucleic acid comprising Notch sequences, wherein said Notch sequences consist of sequences encoding a Notch intracellular domain, and (2) culturing said bone marrow stromal cells such that said bone marrow stromal cells differentiate into neural precursor cells, wherein the resultant differentiated cells are offspring of bone marrow stromal cells into which said nucleic acid has been introduced;   (b) bone marrow stromal cell-derived Schwann cells which are obtained by (1) collecting bone marrow stromal cells from bone marrow, and culturing said cells in a standard essential culture medium supplemented with a serum; (2) adding a reducing agent to said culture medium, and further culturing said cells; (3) adding retinoic acid to said culture medium, and further culturing said cells; and (4) adding forskolin, and/or a differentiation, survival and growth stimulating factor which acts on nerves and glial cells to said culture medium, and further culturing said cells to obtain said bone marrow stromal cell-derived Schwann cells; and   (c) bone marrow stromal cell-derived skeletal muscle cells which are obtained by (1) adding (i) a cyclic AMP (cAMP) increasing agent or a cAMP analogue, and (ii) a cell differentiation stimulating factor comprising bFGF, PDGF-AA, and neuregulin to a culture of bone marrow stromal cells wherein said bone marrow stromal cells are not treated with a demethylating agent, and culturing the cells; (2) introducing a Notch gene into the cells obtained in (1), and culturing the cells to obtain a culture of myoblasts, provided that said culture does not contain a co-culture of the cells introduced with the gene and non-introduced cells; and (3) adding a Notch ligand to the culture of the myoblasts obtained in (2), and culturing the cells such that skeletal muscle cells are induced.   
     
     
         2 . The composition according to  claim 1 , wherein the biocompatible polymer is a gelatin. 
     
     
         3 . The composition according to  claim 1 , wherein the biocompatible polymer is a recombinant gelatin having an amino acid sequence derived from a partial amino acid sequence of collagen. 
     
     
         4 . The composition according to  claim 3 , wherein the recombinant gelatin has a repetition of a sequence characteristic for collagen represented by Gly-X-Y wherein each of X and Y independently represents any given amino acid, wherein a plurality of Gly-X-Y may be identical to or different from one another, and the recombinant gelatin has a molecular weight of 2 KDa to 100 KDa. 
     
     
         5 . The composition according to  claim 3 , wherein the recombinant gelatin has a repetition of a sequence characteristic for collagen represented by Gly-X-Y wherein each of X and Y independently represents any given amino acid, wherein a plurality of Gly-X-Y may be identical to or different from one another, and the recombinant gelatin has a molecular weight of 10 KDa to 90 KDa. 
     
     
         6 . The composition according to  claim 3 , wherein the recombinant gelatin has a repetition of a sequence characteristic for collagen represented by Gly-X-Y wherein each of X and Y independently represents any given amino acid, wherein a plurality of Gly-X-Y may be identical to or different from one another, and the recombinant gelatin comprises two or more sequences of cell adhesion signals in a single molecule. 
     
     
         7 . The composition according to  claim 6 , wherein the cell adhesion signal has an amino acid sequence represented by Arg-Gly-Asp. 
     
     
         8 . The composition according to  claim 3 , wherein the amino acid sequence of the recombinant gelatin does not comprise serine and threonine. 
     
     
         9 . The composition according to  claim 3 , wherein the amino acid sequence of the recombinant gelatin does not comprise serine, threonine, asparagine, tyrosine, and cysteine. 
     
     
         10 . The composition according to  claim 3 , wherein the amino acid sequence of the recombinant gelatin does not comprise an amino acid sequence represented by Asp-Arg-Gly-Asp. 
     
     
         11 . The composition according to  claim 3 , wherein the recombinant gelatin is represented by the following formula:
   A-[(Gly-X-Y) n ] m -B   wherein A represents any given amino acid or amino acid sequence; B represents any given amino acid or amino acid sequence; each of an n number of X independently represents any given amino acid; each of an n number of Y independently represents any given amino acid; n represents an integer of 3 to 100; m represents an integer of 2 to 10; and further, an n number of Gly-X-Y may be identical to or different from one another.   
     
     
         12 . The composition according to  claim 3 , wherein the recombinant gelatin is represented by the following formula:
   Gly-Ala-Pro-[(Gly-X-Y) 63 ] 3 -Gly   wherein each of 63 X units independently represents any given amino acid; each of 63 Y units independently represents any given amino acid; and 63 Gly-X-Y units may be identical to or different from one another.   
     
     
         13 . The composition according to  claim 3 , wherein the recombinant gelatin has: (1) the amino acid sequence shown in SEQ ID NO: 1, or (2) an amino acid sequence showing a homology of 80% or more with the amino acid sequence shown in SEQ ID NO: 1 and having the property of adhering to a nerve cell or a cell capable of differentiating a nerve cell. 
     
     
         14 . The composition according to  claim 3 , wherein the recombinant gelatin is crosslinked. 
     
     
         15 . The composition according to  claim 3 , wherein the crosslinking is carried out with an aldehyde, a condensing agent, or an enzyme. 
     
     
         16 . A method for treating nervous disease, which comprises administering to a patient suffering from nervous disease a therapeutically effective amount of a composition comprising (A) any cells of the following (a) and (b), and (B) a biocompatible polymer.
 (a) bone marrow stromal cell-derived neural precursor cells which are obtained by (1) introducing a nucleic acid comprising Notch sequences, wherein said Notch sequences consist of sequences encoding a Notch intracellular domain, and (2) culturing said bone marrow stromal cells such that said bone marrow stromal cells differentiate into neural precursor cells, wherein the resultant differentiated cells are offspring of bone marrow stromal cells into which said nucleic acid has been introduced; and   (b) bone marrow stromal cell-derived Schwann cells which are obtained by (1) collecting bone marrow stromal cells from bone marrow, and culturing said cells in a standard essential culture medium supplemented with a serum; (2) adding a reducing agent to said culture medium, and further culturing said cells; (3) adding retinoic acid to said culture medium, and further culturing said cells; and (4) adding forskolin, and/or a differentiation, survival and growth stimulating factor which acts on nerves and glial cells to said culture medium, and further culturing said cells to obtain said bone marrow stromal cell-derived Schwann cells; or   
     
     
         17 . The method according to  claim 16 , wherein the composition is administered locally. 
     
     
         18 . The method according to  claim 16 , wherein the composition is administered to the central nervous system of the patient. 
     
     
         19 . A method for treating muscular disease, which comprises administering to a patient suffering from muscular disease a therapeutically effective amount of a composition comprising (A) any cells of the following (c), and (B) a biocompatible polymer.
 (c) bone marrow stromal cell-derived skeletal muscle cells which are obtained by (1) adding (i) a cyclic AMP (cAMP) increasing agent or a cAMP analogue, and (ii) a cell differentiation stimulating factor comprising bFGF, PDGF-AA, and neuregulin to a culture of bone marrow stromal cells wherein said bone marrow stromal cells are not treated with a demethylating agent, and culturing the cells; (2) introducing a Notch gene into the cells obtained in (1), and culturing the cells to obtain a culture of myoblasts, provided that said culture does not contain a co-culture of the cells introduced with the gene and non-introduced cells; and (3) adding a Notch ligand to the culture of the myoblasts obtained in (2), and culturing the cells such that skeletal muscle cells are induced.   
     
     
         20 . The method according to  claim 19 , wherein the composition is administered locally.