低频脉冲磁场对大鼠骨髓源内皮祖细胞增殖和体外血管化的影响

Effects of low frequency pulse magnetic fields on proliferation and tube formation of rat bone marrow-derived endothelial progenitor cells

目的观察不同强度低频脉冲磁场对体外培养大鼠骨髓源内皮祖细胞增殖、细胞周期、迁移和成血管能力的影响。方法密度梯度离心法获得大鼠骨髓源内皮祖细胞,随机分为4组:对照组,1.0mT组,1.4mT组和1.8mT组。除对照组外,其余各组用频率为15Hz不同强度的方波脉冲磁场刺激大鼠骨髓源内皮祖细胞,2h/d,持续刺激5d。曝磁5d后,四甲基偶氮唑盐微量酶反应比色(MTT)法检测细胞增殖情况,流式细胞仪检测细胞周期,划痕试验检测细胞迁移能力,管状结构形成试验和3维培养检测细胞成血管能力。结果1.0mT组和1.4mT组磁场促进内皮祖细胞增殖,使其细胞周期分布发生改变,DNA合成期(S期)和合成后期(G2期)细胞比例增加,1.8mT组磁场也能促进内皮祖细胞增殖,但对细胞周期分布影响不明显。体外培养内皮祖细胞迁移能力差,各强度磁场对其迁移能力也无明显影响。不同强度磁场均能够提高内皮祖细胞成血管能力,与对照组相比差异均有统计学意义[(14.0±1.6)比(11.0±1.6);(19.2±1.9)比(11.0±1.6);(15.4±1.1)比(11.0±1.6),P均<0.05]。1.0mT和1.4mT磁场作用强于1.8mT磁场。结论磁场的生物学作用和磁场强度相关,1.0mT和1.4mT低频脉冲磁场促进大鼠骨髓源内皮祖细胞增殖和DNA合成并提高其成血管能力。

Objectives To investigate the effects of low frequency pulse magnetic fields （LF-PMF） on proliferation, cell cycle, migration and angiogenic potential of rat bone marrow-derived endothelial progenitor cells （EPCs） in vitro. Methods Density gradient centrffugation was used to isolate EPCs from rat bone marrow. Cells were randomly divided into 4 groups（control group, 1.0 mT group, 1.4 mT group and 1.8 mT group）. EPCs, except for the control group, were exposed to low frequency square pulse magnetic fields （ 15 Hz） for 2 hours every day and cuhured for 5 days. After 5 days, we determined the effects of LF-PMF on EPCs＇ proliferation ability by MTT assay, cell cycle by flow cytometry, migration ability by cell scratch assav and angiogenie potential by tube fornmtion assay and 3D- culture assay. Results After 5 days of intervention, 1.0 mT and 1.4 mT magnetic fields accelerated EPCs＇ proliferation. The pereentage of cells at the （ S＋ G2） phase was increased significantly. 1.8 mT magnetic field also accelerated EPCs＇ proliferation, however, it didn＇t change the distribution of cell cycle significantly. EPCs had poor migratory ability in vitro both in control group and in groups exposed to magnetic field. LF-PMF of 1.0 mT, 1.4 mT and 1.8 mT elevated angiogenie potential of EPCs and increased the numbers of tube-like structures compared with control group [（14.0±1.6） vs. （11.0±1.6）, P〈0.05; （19.2±1.9） vs （11.0±1.6）, P〈0.05; （15.4±1.1） vs （11.0±1.6）, P 〈 0.05], while LF-PMF of 1.0 mT and 1.4 mT had more significant effects than magnetic fields of 1.8 mT. Conclusions LF-PMF of 1.0 mT and 1.4 mT accelerate proliferation and tube formation of EPCs.