Preconditioning of mesenchymal stem cells with low-intensity ultrasound for cartilage formation in vivo.
The purpose of this study was to evaluate the benefits of in vitro preconditioning of mesenchymal stem cells (MSCs) using low-intensity ultrasound (US) in the induction of chondrogenic differentiation of MSCs in vivo. After rabbit bone marrow-derived MSCs were seeded onto a polyglycolic acid (PGA) scaffold, the PGA-MSCs constructs were divided into 4 subgroups: untreated control, low-intensity US group, transforming growth factor-beta [TGF]-treated group and low-intensity US/TGF group. The chondrocyte-seeded PGA construct served as a positive control. For 1 week before implantation, the low-intensity US groups were subjected to ultrasound treatment for 20 min daily at an intensity of 200 mW/cm(2). The TGF groups were treated with 10 ng/mL TGF-beta1. The cells were then implanted into the nude mouse subcutaneously. Retrieved 1, 2, 4, and 6 weeks after implantation, each construct underwent gross examination, histology, biochemical assays, mechanical testing, and reverse transcriptase polymerase chain reaction (RT-PCR). Substantial size reduction and blood invasion were found much earlier in the groups that did not undergo low-intensity US than in those that did. Safranin O/Fast green staining revealed that the chondrogenic differentiation of MSCs was more widespread throughout the constructs in the low-intensity US groups. In the biochemical and mechanical analyses, the low-intensity US and low-intensity US/TGF groups were significantly better in forming hyaline cartilage-like tissue by 4 weeks than the non-low-intensity US groups. Presented by von Kossa staining, the development of osteogenic phenotypes was highly suppressed until 4 weeks in the low-intensity US groups, along with compressive strength comparable to the positive control. In the RT-PCR analysis before implantation, the messenger RNA levels of Sox-9, aggrecan, and tissue inhibitors of metalloproteinase-2 were higher in the low-intensity US groups, while those of type I and type X collagens and matrix metalloproteinase-13 were higher in the non-low-intensity US groups. Blood invasion into the constructs was also considerably hindered in the low-intensity US groups. These results strongly indicate that low-intensity US preconditioning in vitro could be an effective cue to upregulate chondrogenic differentiation of MSCs in vivo.
Cui
Ji Hao
JH
Department of Orthopaedic Surgery, Ajou University School of Medicine, Wonchon-dong, Youngtong-gu, Suwon, Gyeonggi, Korea.
Park
So Ra
SR
Park
Kwideok
K
Choi
Byung Hyune
BH
Min
Byoung-Hyun
BH
D000818:Animals; D002356:Cartilage / Q000166:cytology / Q000254:growth & development* / Q000528:radiation effects*; D002454:Cell Differentiation / Q000502:physiology / Q000528:radiation effects; D049109:Cell Proliferation / Q000528:radiation effects; D002478:Cells, Cultured; D020219:Chondrogenesis / Q000502:physiology / Q000528:radiation effects*; D005260:Female; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000502:physiology / Q000528:radiation effects*; D009924:Organ Culture Techniques / Q000379:methods; D011817:Rabbits; D011829:Radiation Dosage; D013010:Sonication; D023822:Tissue Engineering / Q000379:methods*