Adult human bone marrow-derived mesenchymal progenitor cells are capable of adhesion-independent survival and expansion.
BACKGROUND
We show the existence of adult human mesenchymal progenitor cells (hMPCs) that can proliferate, in a cytokine-dependent manner, as individual cells in stirred suspension cultures (SSC) while maintaining their ability to form functional differentiated mesenchymal cell types.


METHODS
Ficolled human bone marrow (BM)-derived cells were grown in SSC (and adherent controls) in the presence and absence of exogenously added cytokines. Phenotypic, gene expression, and functional assays for hematopoietic and nonhematopoietic cell populations were used to kinetically track cell production. Limiting-dilution analysis was used to relate culture-produced cells to input cell populations.


RESULTS
Cytokine cocktail influenced total and progenitor cell expansion, as well as the types of cells generated upon plating. Flow cytometric analysis of CD117, CD123, and CD45 expression showed that cytokine supplementation influenced SSC output. The concomitant growth of CD45(+) and CD45(-) cells in the cultures that exhibited the greatest hMPC expansions suggests that the growth of these cells may benefit from interactions with hematopoietic cells. Functional assays demonstrated that the SSC-derived cells (input CFU-O number: 1990+/-377) grown in the presence of SCF+IL-3 resulted, after 21 days, in the generation of a significantly greater number (p<0.05) of bone progenitors (33,700+/-8763 CFU-O) than similarly initiated adherent cultures (214+/-75 CFU-O). RT-PCR analysis confirmed that the SSC-derived cells grown in osteogenic conditions express bone-specific genes (Cbfa1/Runx2, bone sialoprotein, and osteocalcin).


CONCLUSIONS
Our approach not only provides an alternative strategy to expand adult BM-derived nonhematopoietic progenitor cell numbers in a scalable and controllable bioprocess, but also questions established biological paradigms concerning the properties of connective-tissue stem and progenitor cells.
Baksh
Dolores
D
Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, ON, Canada
Davies
John E
JE
Zandstra
Peter W
PW
D000328:Adult; D001854:Bone Marrow Cells / Q000166:cytology* / Q000187:drug effects; D002448:Cell Adhesion; D018929:Cell Culture Techniques / Q000379:methods; D002455:Cell Division / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured / Q000166:cytology / Q000187:drug effects; D016207:Cytokines / Q000494:pharmacology; D006410:Hematopoiesis; D006801:Humans; D008648:Mesoderm / Q000166:cytology*; D008875:Middle Aged; D039902:Multipotent Stem Cells / Q000166:cytology* / Q000187:drug effects; D013535:Suspensions
D016207:Cytokines; D013535:Suspensions