Shockwave stimulates oxygen radical-mediated osteogenesis of the mesenchymal cells from human umbilical cord blood.
Human umbilical cord blood (HUCB) mesenchymal progenitor cells expressed stro-1 or CD44 or CD29, and subsequently, differentiated toward osteogenic lineage. Physical shockwave treatment increased osteogenic activity of HUCB mesenchymal progenitor cells through superoxide-mediated TGF-beta1 induction. Transplantation of shockwave-treated HUCB mesenchymal progenitor cells enhanced healing of segmental femoral defect in severe combined immunodeficiency disease (SCID) mice.


BACKGROUND
Mesenchymal progenitor cells (MPCs) in the bone marrow are precursors to bone development. It remains uncertain whether MPCs are present in human umbilical cord blood (HUCB) and are capable of differentiating into osteogenic cell lineage. Extending from a model of shockwave (SW) promotion of bone marrow stromal cell differentiation toward osteoprogenitors in rats, we further investigated how physical SW mediated biological responses in regulating osteogenic differentiation of HUCB MPCs.


METHODS
HUCB was subjected to SW treatment at different energy flux densities and impulses. Colony-forming units-stroma (CFU-Stroma), osteogenic activities (Cbfa1/Runx2 expression, bone alkaline phosphatase activity, and bone nodule formation), and bone formation by heterologous transplantation into SCID mice were assessed.


RESULTS
Few CD34+ stem cells (1.3%) and stro-1+ cells (1.0%) were present in the freshly prepared mononuclear cells (MNCs) from HUCB. The number of stro-1+ cells, but not CD34+, increased to 72.4% in the adherent cell culture over 6 days. Stro-1+ cells co-expressed CD44 and CD29 markers and grew into CFU-Stroma that matured into bone nodules. We found that the SW treatment (0.16 mJ/mm2 energy flux density, 200 impulses) elicited superoxide production and promoted formation of CFU-Stroma, but not of hematopoietic CFU-Mix. SW also enhanced the production of transforming growth factor (TGF)-beta1, but not of interleukin (IL)-3 or granulocyte monocyte-colony stimulating factor (GM-CSF). Neutralization of TGF-beta1 significantly reduced SW-promoted CFU-Stroma formation. Superoxide scavenging by superoxide dismutase blocked SW enhancement of TGF-beta1 production and formation of CFU-Stroma. Administration of SW-treated HUCB MPCs to SCID mice with femoral segmental defects facilitated dense, bridging callus and gap closure.


CONCLUSIONS
HUCB MPCs subjected to SW treatment is a potential source for stem cells useful in the treatment of orthopedic disorders. An optimal physical SW treatment enhanced osteogenesis through superoxide-mediated TGF-beta1 production. Physical stimulation is an alternative method for extending mesenchymal stem cells of HUCB.
Wang
Feng-Sheng
FS
Department of Medical Research, Chang Gung Memorial Hospital at Kaohsiung, Chang Gung University, Niau-Sung, Kaohsiung, Taiwan.
Yang
Kuender D
KD
Wang
Ching-Jen
CJ
Huang
Hui-Cheng
HC
Chio
Chi-Chian
CC
Hsu
Te-Yao
TY
Ou
Chia-Yu
CY
D000818:Animals; D015703:Antigens, CD / Q000276:immunology; D001483:Base Sequence; D017690:Cell Transplantation; D017931:DNA Primers; D005312:Fetal Blood; D006801:Humans; D016130:Immunophenotyping; D008297:Male; D008648:Mesoderm / Q000166:cytology*; D051379:Mice; D016513:Mice, SCID; D009249:NADP / Q000378:metabolism; D010012:Osteogenesis; D010812:Physical Stimulation; D013481:Superoxides / Q000378:metabolism*; D014470:Umbilical Cord / Q000166:cytology*
D015703:Antigens, CD; D017931:DNA Primers; D013481:Superoxides; D009249:NADP