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松茸多糖对 1-甲基-4-苯基-吡啶离子诱导 PC12 细胞损伤的影响
吕海燕 沈西雅 赵富生 朱梅*
Effects of tricholoma matsutake polysaccharides on 1-methy-4-pehnyl-pyridine ion-induced PC12 cell damage
LÜ Hai-yan SHEN Xi-ya ZHAO Fu-sheng ZHU Mei*
目的 探讨松茸多糖（TMP ）对 1-甲基-4-苯基-吡啶离子（MPP+）诱导 PC12 细胞损伤的保护作用机制。 方法 选用 MPP+ 体外构建PC12 细胞帕金森病（PD ）模型，将 PC12 细胞随机分为5组：对照组（Ctrl）、模型组(MDL)、TMP高浓度（250 μg/L）组(TMP-H)、TMP中浓度（50 μg/L）组(TMP-M)和TMP低浓度（10 μg/L）组(TMP-L)，每组 3 个复孔。TMP处理 PC12 细胞24 h 后，250 μmol/L MPP+ 诱导 PC12 细胞 24 h制备 PD 细胞模型。用CCK-8法检测各组 PC12 细胞的细胞存活率，乳酸脱氢酶(LDH)法检测各组 PC12 细胞的 LDH 释放量，荧光显微镜摄片检测线粒体活性，Image J 1.53 r软件计算线粒体网络分支平均长度及网络分支数，JC-1 法检测线粒体膜电位，Western blotting检测蛋白 Bcl-2/Bax 比值。 结果 与Ctrl组相比，MDL组 PC12 细胞存活率下降，LDH 释放水平升高，线粒体膜电位（ΔΨm ）及 Bcl-2/Bax 比值下降(均P＜0.05)；加入不同浓度的TMP后，与MDL组相比，TMP-H组和TMP-M组细胞存活率均上升，LDH释放水平降低，ΔΨm及Bcl-2/Bax 比值增高(P＜0.05)，线粒体活性增加，线粒体网络分支平均长度及网络分支数量增多，拮抗膜电位降低(P＜0.05)，其中TMP-H组在拮抗膜电位的降低作用中更为明显。 结论 TMP对 MPP+ 所致的 PC12 细胞损伤具有保护作用，可能是通过提高细胞存活率，降低 LDH 释放量，拮抗线粒体膜电位的降低，修复线粒体网络形态，从而减少细胞凋亡而实现的。
Objective To investigate the protective mechanism of tricholoma matsutake polysaccharides（TMP）against 1-methy-4-pehnyl-pyridine ion (MPP+)-induced PC12 cell damage. Methods An Parkinson’s disease (PD) PC12 cell model in vitro was constructed using MPP+, and PC12 cell were randomly divided into 5 groups: control group（Ctrl）, model group(MDL), TMP high concentration (250 μg/L) group (TMP-H), TMP medium concentration (50 μg/L) group (TMP-M), and TMP low concentration (10 μg/L)group (TMP-L), 3 complex wells in each group. Matsutake polysaccharide treated PC12 cells for 24 hours, 250 μmol/L MPP+ induced PC12 cells were prepared for 24 hours to prepare PD cell models. The cell viability of each group of PC12 cell was detected by CCK-8 method, the lactate dehydrogenase (LDH) release of each group of PC12 cell was detected by LDH method, mitochondrial activity was detected by fluorescence microscopy, the average length of mitochondrial network branches and the number of network branches were calculated by Image J 1.53 r software, the mitochondrial membrane potential was detected by JC-1 method, and the protein Bcl-2/Bax ratio was detected by Western blotting method. Results Compared with the Ctrl group, the survival rate of PC12 cell in the MDL group decreased, the LDH release level increased, and the mitochondrial membrane potential (MMP, ΔΨm) and Bcl-2/Bax ratio decreased (all P<0.05). Compared with the MDL group, the cell survival rate of matsutake polysaccharide in the TMP-H group and the TMP-M group increased, the LDH release level decreased, the ΔΨm and Bcl-2/Bax ratio increased (P<0.05), mitochondrial activity increased, the average length of mitochondrial network branches and the number of network branches increased, and the antagonistic membrane potential decreased (P<0.05), among which the matsutake polysaccharide group was more obvious in the reduction of antagonistic membrane potential. Conclusion TMP have a protective effect against MPP+-induced PC12 cell damage, possibly by improving cell viability, reducing LDH release, antagonizing the reduction of mitochondrial membrane potential, and repairing mitochondrial network morphology, thereby reducing apoptosis.
松茸多糖 / 线粒体膜电位 / 帕金森病 / PC12细胞 / 免疫印迹法
Matsutake polysaccharide / Mitochondrial membrane potential / Parkinson’s disease / PC12 cell / Western blotting
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