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目的研究不同的电刺激信号对大鼠雪旺细胞(RSC96)增殖的影响。方法实验使用RSC96细胞系(中国科学院典型培养物保藏委员会细胞库),经过2次传代培养,以1.25×10~4/孔的密度种植于24孔细胞培养板中,次日电刺激信号分别采用不同的电压(10、100、500、1 000、5 000、10 000 m V/cm)、不同的频率(0.1、1.0、100.0、1 000.0、10 000.0、1 000 000.0 Hz)、不同的脉冲占空比(5%、10%、30%、50%、70%、90%)及不同的波形(正脉冲波、矩形波、三角波、正弦波、杂波、直流波)对细胞进行电刺激,每组设置3个平行样,细胞每天刺激30 min,持续刺激4 d。采用四甲基偶氮唑盐(MTT)法检测细胞增殖率。结果当电刺激频率为100.0 Hz、脉冲占空比为50%、波形为矩形波时,电压>500 m V/cm时对细胞的增殖具有抑制作用。当电刺激频率为100.0 Hz、脉冲占空比为50%、电压为500 m V/cm时,正脉冲波、正弦波、矩形波和三角波都能够促进细胞的增殖。当电刺激波形为矩形波、脉冲占空比为50%、电压为500 m V/cm时,频率100.0~1 000.0 Hz组细胞增殖率最高。当电刺激波形为矩形波、频率为100.0 Hz、电压为500 m V/cm时,不同的脉冲占空比对细胞增殖的影响不明显。结论不同的电刺激信号条件对细胞增殖的影响较大。电压≤500 m V/cm、有规律的脉冲波形、频率为100.0~1 000.0 Hz电刺激信号能够获得较好的RSC96增殖的效果。实验所获得的优化电刺激参数在外周神经修复中具有非常大的应用前景。
Objective To study the effects of different electrical stimulation signals on the proliferation of rat Schwann cells (RSC96). Methods The RSC96 cell line (cell culture library of Chinese Academy of Sciences) was used in this experiment. After 2 subcultures, cells were seeded in 24-well cell culture plates at a density of 1.25 × 10 4 / well. The next day’s electrical stimulation signals were respectively Different voltages (10,100,500,1,000,5,000,10,000 mV / cm), different frequencies (0.1,1.0,100.0,1 000.0,10,000.0,1,000,000.0 Hz), different pulses (5%, 10%, 30%, 50%, 70%, 90%) and different waveforms (positive, rectangular, triangular, sine, clutter and DC) Three parallel samples were set in each group. The cells were stimulated for 30 min every day for 4 days. Cell proliferation rate was measured by MTT assay. Results When the frequency of electrical stimulation was 100.0 Hz, the pulse duty cycle was 50% and the waveform was rectangular wave, the cell proliferation was inhibited when the voltage was> 500 mV / cm. When the electrical stimulation frequency is 100.0 Hz, the pulse duty cycle is 50% and the voltage is 500 mV / cm, positive pulse, sine wave, square wave and triangular wave can all promote cell proliferation. When the waveform of electrical stimulation was a square wave and the pulse duty cycle was 50% and the voltage was 500 mV / cm, the cell proliferation rate was highest in the frequency of 100.0-1000.0 Hz. When the waveform of electrical stimulation was a square wave with a frequency of 100.0 Hz and a voltage of 500 mV / cm, the effect of different pulse duty cycles on cell proliferation was insignificant. Conclusion Different electrical stimulation signal conditions have a greater impact on cell proliferation. Voltage ≤500 m V / cm, a regular pulse waveform, the frequency of 100.0 ~ 1 000.0 Hz electrical stimulation signal can get better RSC96 proliferation. Optimized electrical stimulation parameters obtained in the experiment in the peripheral nerve repair has a very large application prospects.