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J Cell Physiol. 2018 Feb;233(2):1558-1573. doi: 10.1002/jcp.26058. Epub 2017 Jul 11.

Osteogenic commitment and differentiation of human mesenchymal stem cells by low-intensity pulsed ultrasound stimulation.

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Rizzoli Orthopedic Institute, Innovative Technological Platforms for Tissue Engineering, Theranostic and Oncology, Palermo, Italy.
Biology and Genetics Unit, Department of Biopathology and Medical Biotechnologies, University of Palermo, Italy.
Rizzoli Orthopedic Institute, Laboratory of Preclinical and Surgical Studies, Bologna, Italy.
Rizzoli Orthopedic Institute, Laboratory BITTA, Bologna, Italy.
IGEA SpA, Carpi (Modena), Italy.
Rizzoli Orthopedic Institute, 2nd Orthopaedic and Traumatologic Clinic, Bologna, Italy.


Low-intensity pulsed ultrasound (LIPUS) as an adjuvant therapy in in vitro and in vivo bone engineering has proven to be extremely useful. The present study aimed at investigating the effect of 30 mW/cm2 LIPUS stimulation on commercially available human mesenchymal stem cells (hMSCs) cultured in basal or osteogenic medium at different experimental time points (7, 14, 21 days). The hypothesis was that LIPUS would improve the osteogenic differentiation of hMSC and guarantying the maintenance of osteogenic committed fraction, as demonstrated by cell vitality and proteomic analysis. LIPUS stimulation (a) regulated the balance between osteoblast commitment and differentiation by specific networks (activations of RhoA/ROCK signaling and upregulation of Ribosome constituent/Protein metabolic process, Glycolysis/Gluconeogenesis, RNA metabolic process/Splicing and Tubulins); (b) allowed the maintenance of a few percentage of osteoblast precursors (21 days CD73+/CD90+: 6%; OCT-3/4+/NANOG+/SOX2+: 10%); (c) induced the activation of osteogenic specific pathways shown by gene expression (early: ALPL, COL1A1, late: RUNX2, BGLAP, MAPK1/6) and related protein release (COL1a1, OPN, OC), in particular in the presence of osteogenic soluble factors able to mimic bone microenvironment. To summarize, LIPUS might be able to improve the osteogenic commitment of hMSCs in vitro, and, at the same time, enhance their osteogenic differentiation.


low-intensity pulsed ultrasound; mesenchymal stem cells; osteoblast differentiation; osteogenic commitment; proteomic analysis

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