Isolation of mesenchymal stem cells
View Patent ↗Methods and compositions are provided for the identification and isolation of mammalian mesenchymal stem cells. The methods of the invention provide a means to obtain substantially homogeneous MSC populations. In some embodiments, the homogeneous MSC composition is stable in non-differentiating culture conditions, where the proportion of cells in the composition that have an MSC phenotype are maintained over multiple passages.
1. A method for the isolation of mammalian mesenchymal stem cells (MSC) from bone marrow, the method comprising:
introducing into an in vitro population of mammalian cells isolated from bone marrow a nucleic acid construct comprising sequences encoding a detectable marker, which marker is operably linked to a transcriptional response element regulated by β-catenin, wherein said transcriptional response element is a LEF-1/TCF binding sequence;
detecting the presence of expression of said detectable marker;
selecting for cells expressing said detectable marker; and
wherein expression of said marker is indicative that a cell is an MSC.
2. The method according to claim 1 , wherein said marker is a fluorescence producing protein.
3. The method according to claim 1 , further comprising the step of culturing cells expressing said detectable marker in vitro.
4. The method according to claim 3 ,, wherein said in vitro culture provides non-differentiating conditions.
5. The method according to claim 3 , wherein said in vitro culture provides differentiating conditions.
6. The method according to claim 5 , wherein said culture conditions induce differentiation of at least one of chondrocytes, adipocytes, osteocytes and myocytes.
7. The method according to claim 4 , further comprising the steps of:
culturing said cells for at least one passage; and
selecting for cells expressing said detectable marker.
8. The method according to claim 7 , wherein the selected cells remain non-differentiated for at least five passages in culture.