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

Effects of tricholoma matsutake polysaccharides on 1-methy-4-pehnyl-pyridine ion-induced PC12 cell damage

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(250μg/L)组(TMP-H)、TMP中浓度(50μg/L)组(TMP-M)和TMP低浓度(10μg/L)组(TMP-L),每组3个复孔。TMP处理PC12细胞24 h后,250μmol/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(250μg/L)group(TMP-H),TMP medium concentration(50μg/L)group(TMP-M),and TMP low concentration(10μg/L)group(TMP-L),3 complex wells in each group.Matsutake polysaccharide treated PC12 cells for 24 hours,250μmol/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,theΔΨm 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.