低强度脉冲超声保护MPP+诱导的N2a细胞损伤的实验研究

目的 观察低强度脉冲超声（LIPUS）能否减弱1-甲基-4-苯基吡啶离子（MPP⁺）诱导的N2a细胞毒性，对帕金森病（PD）细胞模型发挥保护作用。方法 以MPP⁺作用于N2a细胞24 h，建立PD细胞模型，并对N2a细胞进行超声辐照（1 MHz，50 mW/cm²，脉冲重复频率1 kHz，辐照10 min）。将细胞分为对照组、超声对照（LIPUS）组、MPP⁺组和超声处理（MPP⁺+LIPUS）组，并给予相应处理。采用CCK8法测定细胞活力，DCFH-DA染色法检测细胞内活性氧簇（ROS）水平，JC-1染色法检测线粒体膜电位变化，Annexin V-FITC/PI双染法检测细胞凋亡情况；以实时荧光定量PCR检测瞬时受体电位M7（TRPM7）的mRNA表达水平。结果 与对照组相比，MPP⁺组细胞存活率及线粒体膜电位显著降低，细胞内ROS水平和细胞凋亡明显增加（P均<0.01）；LIPUS辐照后，经MPP⁺诱导的细胞存活率和线粒体膜电位显著上升，细胞内ROS水平及细胞凋亡明显降低（P均<0.01）。MPP⁺组TRPM7的mRNA表达下降，MPP⁺+LIPUS组TRPM7表达显著升高（P均<0.01）。结论 LIPUS可增加MPP⁺诱导的N2a细胞存活率，减少线粒体膜电位损伤和ROS生成，抑制细胞凋亡，对MPP⁺的神经细胞毒性具有保护作用。

Protective effect of low-intensity pulsed ultrasound on N2a cells injury induced by MPP+

Objective To investigate whether low-intensity pulsed ultrasound (LIPUS) could attenuate the cytotoxicity of N2a induced by MPP⁺ and protect the cell model of Parkinson disease. Methods N2a cells were treated with MPP⁺ for 24 h to establish PD cell models. N2a cells were irradiated by ultrasound (1 MHz, 50 mW/cm², pulse repetition frequency 1 kHz, irradiation for 10 min). Then the cells were divided into control group, LIPUS group, MPP⁺ group and MPP⁺+LIPUS group and were processed accordingly. CCK8 assay was used to detect cell viability. The level of intracellular reactive oxygen species (ROS) was detected with DCFH-DA staining. Mitochondrial membrane potential was detected with JC-1 staining. Annexin VFITC/PI double staining method was used to observe cell apoptosis. The mRNA expression of TRPM7 was detected using real-time quantitative PCR. Results Compared with control group, cell viability and mitochondrial membrane potential of MPP⁺ group significantly decreased, the level of intracellular ROS and cell apoptosis rate significantly increased (all P<0.01). After irradiation with LIPUS, cell viability and mitochondrial membrane potential of MPP⁺-induced cells increased significantly, while ROS production and cell apoptosis rate decreased significantly (all P<0.01). The mRNA expression of TRPM7 in MPP⁺ group decreased, and the expression of TRPM7 in MPP⁺+LIPUS group increased significantly (all P<0.01). Conclusion LIPUS can increase the survival rate of N2a cells induced by MPP⁺, reduce mitochondrial membrane potential damage and ROS production, inhibit apoptosis and protect the neuronal toxicity of MPP⁺.