RESUMEN
Objective To study in vitro the effects of low-intensity pulsed ultrasound (LIPUS) on the proliferation and differentiation of cultured myoblasts,and to explore the cellular and molecular mechanisms behind any therapeutic effect of LIPUS.Methods Myoblasts were isolated from the skeletal muscles of mice and cultured in vitro.Treatment and control groups of proliferating and differentiating myoblasts were defined.The treatment groups were exposed to LIPUS at 1.5 MHz and a spatial and temporal average intensity of 30 mW/cm2,for 20 min daily,the proliferation group for 6 consecutive days and the differentiation group for 4 consecutive days.The cell proliferation kinetics of the proliferation group were analyzed using flow cytometry.The expression of myogenic regulation factor MyoD and heme oxygenase-1 (HO-1) in the proliferation group,and of myosin heavy chain (MHC) in the differentiation group were examined by immunofluorescent staining.Myoblast fusion indexes were analyzed.Results In the LIPUS treatment groups the proliferating myoblasts had a higher ratio of active cells in the G2 and S phases (19.30％ ±5.14％,37.00％ ±8.72％),compared with the controls (10.33％ ± 1.53％,25.00％ ±4.36％),and the proliferation index increased significantly.The expression of HO-1 was up-regulated,while MyoD staining was unchanged.During the induction of differentiation,the myoblasts of the treatment group fused into smaller myotubes and the myoblast fusion index (18.73％ ± 6.81％) was significantly lower than that of the control group (37.52％ ± 11.23％),while MHC expression did not change markedly.Conclusion LIPUS can promote myoblast proliferation while inhibiting their differentiation,but it does not affect the cells' myogenic properties.HO-1 may be involved in the regulation process.