摘要
目的研究低强度脉冲超声波（LIPUS）对培养成肌细胞增殖分化的影响，探讨其治疗效应的细胞分子机制。方法建立小鼠骨骼肌成肌细胞体外培养模型，将培养成肌细胞随机分增殖超声组、分化超声组、增殖对照组和分化对照组。增殖超声组和分化超声组均采用超声频率为1．5MHz，平均强度为30mW／cm^2的低强度脉冲超声波辐射，每次20rain，每日1次，增殖超声组连续处理6d，分化超声组连续处理4d。增殖超声组与增殖对照组采用流式细胞仪进行细胞增殖动力学检测，免疫荧光染色检测成肌细胞生长因子MyoD、血红素加氧酶-1（HO-1）的表达，并对分化超声组与分化对照组进行肌球蛋白重链（MHC）染色和成肌细胞融合指数分析。结果增殖超声组成肌细胞进入活性细胞周期G2／M与S期细胞比例和增殖指数（PI）分别为（19．30±5．14）％，（37．00±8．72）％和（47．93±0．87）％，与增殖对照组的（10．334-1．53）％，（25．00±4．36）％和（38．66±0．67）％比较，差异具有统计学意义（P〈0．05）；增殖超声组HO．1荧光染色阳性比例和平均荧光强度分别为（82．034±5．14）％和（152．02±4．76）％，与对照组的（60．01±3．22）％和（138．70±5．08）％比较，差异具有统计学意义（尸〈0．05）；分化超声组成肌细胞融合指数（18．73±6．81）％，与分化对照组的（37．52±11．23）％比较，差异有统计学意义（P〈0．05）。结论低强度脉冲超声波促进成肌细胞的增殖，抑制其分化，且不改变其肌原属性，血红素加氧酶一1参与该调控过程。
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 cytoroetry. 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 immunofluoreseent 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.
出处
《中华物理医学与康复杂志》
CAS
CSCD
北大核心
2013年第4期256-260,共5页
Chinese Journal of Physical Medicine and Rehabilitation
基金
国家自然科学基金资助项目（31171148）
重庆市科委自然科学基金资助项目（CSTC,2011BB5038）
