Neural regenerating cells with alterations in DNA methylation

Abstract

Disclosed herein are cells, that are descendents of marrow adherent stem cells (MASCs), capable of rescuing and/or reversing various neural disorders after transplantation into sites of central nervous system (CNS) or peripheral nervous system (PNS) injury. The cells contain alterations in the methylation state of certain genes, compared to their methylation state in MASCs. Methods of making cells capable of rescuing and/or reversing various neural disorders after transplantation into sites of CNS or PNS injury, by alteration of the methylation status of certain genes, are also provided.

What is claimed is: 
     
       1. A method for converting a marrow adherent stromal cell (MASC) to a neural regenerating cell (NRC); the method comprising:
 altering the methylation state of one or more genes in the MASC, wherein the alterations comprise:
 (a) increased methylation of the PITX2, DNMT3b, IGF2R and SDF4 genes; and 
 (b) decreased methylation of the ROPN1L and TMEM179 genes; 
 
 wherein the method does not include transfection of the MASC with a polynucleotide comprising sequences encoding a Notch intracellular domain. 
 
     
     
       2. The method of  claim 1 , wherein the methylation state of the PITX2, DNMT3b, IGF2R and SDF4 genes is increased by contacting the MASC with one or more fusion proteins comprising a methylation domain and a DNA-binding domain, or with one or more nucleic acids encoding a fusion protein comprising a methylation domain and a DNA-binding domain, wherein the DNA-binding domains are engineered to bind to one or more sequences in each of the PITX2, DNMT3b, IGF2R and SDF4 genes. 
     
     
       3. The method of  claim 1 , wherein the methylation state of the ROPN1L and TMEM179 genes is decreased by contacting the MASC with one or more fusion proteins comprising a demethylation domain and a DNA-binding domain, or with one or more nucleic acids encoding a fusion protein comprising a demethylation domain and a DNA-binding domain, wherein the DNA-binding domains are engineered to bind to one or more sequences in each of the RPON1L and TMEM179 genes. 
     
     
       4. A neural regenerating cell that is descended from a MASC in vitro, wherein:
 (a) the cell supports the growth and/or regeneration of neural tissue;   (b) the methylation state of one or more genes in the cell is altered compared to the MASC, wherein the alterations in methylation comprise:
 (i) increased methylation of the PITX2, DNMT3b, IGF2R and SDF4 genes, and 
 (ii) decreased methylation of the ROPN1L and TMEM179 genes; and 
   (c) during culture in vitro, neither the MASC nor any of its descendants were transfected with a polynucleotide comprising sequences encoding a Notch intracellular domain.   
     
     
       5. The neural regenerating cell of  claim 4  A neural regenerating cell that is descended from a MASC in vitro, wherein:
 (a) the cell supports the growth and/or regeneration of neural tissue; 
 (b) the methylation state of one or more genes in the cell is altered compared to the MASC, wherein the alterations in methylation comprise:
 (i) increased methylation of the PITX2, DNMT3b, IGF2R and SDF4 genes, and 
 (ii) decreased methylation of the ROPN1L and TMEM179 genes; and 
 
 (c) during culture in vitro, neither the MASC nor any of its descendants were transfected with a polynucleotide comprising sequences encoding a Notch intracellular domain,  
 wherein the methylation state of the PITX2, DNMT3b, IGF2R and SDF4 genes is increased by contacting the MASC with one or more fusion proteins comprising a methylation domain and a DNA-binding domain, or with one or more nucleic acids encoding a fusion protein comprising a methylation domain and a DNA-binding domain, wherein the DNA-binding domains are engineered to bind to one or more sequences in each of the PITX2, DNMT3b, IGF2R and SDF4 genes. 
 
     
     
       6. The neural regenerating cell of  claim 4  A neural regenerating cell that is descended from a MASC in vitro, wherein:
 (a) the cell supports the growth and/or regeneration of neural tissue; 
 (b) the methylation state of one or more genes in the cell is altered compared to the MASC, wherein the alterations in methylation comprise:
 (i) increased methylation of the PITX2, DNMT3b, IGFZR and SDF4 genes, and 
 (ii) decreased methylation of the ROPNIL and TMEM179 genes; and 
 
 (c) during culture in vitro, neither the MASC nor any of its descendants were transfected with a polynucleotide comprising sequences encoding a Notch intracellular domain, 
 wherein the methylation state of the ROPNIL and TMEM179 genes is decreased by contacting the MASC with one or more fusion proteins comprising a demethylation domain and a DNA-binding domain, or with one or more nucleic acids encoding a fusion protein comprising a demethylation domain and a DNA-binding domain, wherein the DNA-binding domains are engineered to bind to one or more sequences in each of the RPON1L and TMEM179 genes.