Functional characterization of human mesenchymal stem cells that maintain osteochondral fates.
Adult stem cells are essential for tissue renewal, regeneration and repair, and their expansion in defined culture medium is on focus for regenerative medicine and genetic pathologies. The bone marrow has been shown to be very rich is pluripotent mesenchymal stem cells (MSCs) capable of forming bone, cartilage and also may give rise, to neurons and astrocytes in vivo and in vitro. MSCs can be isolated and expanded in culture, but human cells cannot be verified for a cartilage or a bone fate by transfer experiments. Accordingly, here we used different approaches to characterize hMSCs osteoblastic differentiation in vitro. hMSCs grown in culture in the presence of fetal bovine serum (FBS) expressed the bone-specific transcription factor Runx2/AML3. When cells were incubated in osteoblastic differentiation medium, cells expressed transcripts belonging to the signaling of Indian HH-PTHrP axis, GLI transcription factors, and bone target genes including osteopontin. The HH pathway proved to be functional since it induced cells to grow. Cells growing or differentiating to osteoblasts presented the Runx2/AML3 transcription factor, its partner CBFB, and Smad2/3 at the nuclei associated with the nuclear matrix. Furthermore, Runx2/AML3 was observed to co-localize with SC35 to the nuclear intermediary filaments. These data support the notion that hMSCs isolated from human bone are or become bone progenitor cells upon culture. In the absence of FBS and in the presence of insulin or prolactin, cells show cytoskeletal organization and an AP-1 transcription site activity resembling proliferative osteochondrocytes while cells in the presence of dexamethasone and added prolactin or TGF-beta resembled differentiated osteoblasts. These specific cellular conditions match those observed during endochondral bone formation.
Romero-Prado
Marina
M
Departamento de Biología Molecular, Centro de Biología Molecular, Severo Ochoa, Facultad de Ciencias, Universidad Autónoma de Madrid, Cantoblanco, 28049 Madrid, Spain.
Blázquez
Cristina
C
Rodríguez-Navas
Carmen
C
Muñoz
Jaime
J
Guerrero
Isabel
I
Delgado-Baeza
Emilio
E
García-Ruiz
Josefa P
JP
D002454:Cell Differentiation; D050659:Core Binding Factor Alpha 1 Subunit / Q000378:metabolism; D050658:Core Binding Factor beta Subunit / Q000378:metabolism; D003599:Cytoskeleton / Q000378:metabolism; D053823:Hedgehog Proteins / Q000378:metabolism*; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism* / Q000502:physiology; D015530:Nuclear Matrix / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D010006:Osteoblasts / Q000378:metabolism; D010012:Osteogenesis; D044162:Parathyroid Hormone-Related Protein / Q000378:metabolism*; D012261:Ribonucleoproteins / Q000378:metabolism; D000068103:Serine-Arginine Splicing Factors; D051899:Smad2 Protein / Q000378:metabolism; D051900:Smad3 Protein / Q000378:metabolism
C493692:CBFB protein, human; D050659:Core Binding Factor Alpha 1 Subunit; D050658:Core Binding Factor beta Subunit; D053823:Hedgehog Proteins; D009687:Nuclear Proteins; D044162:Parathyroid Hormone-Related Protein; D012261:Ribonucleoproteins; C497790:SMAD3 protein, human; D051899:Smad2 Protein; D051900:Smad3 Protein; C073288:SRSF2 protein, human; D000068103:Serine-Arginine Splicing Factors