TLR2/NF⁃κB通路介导MPP+诱导的RSC96细胞凋亡与自噬  

TLR2/NF⁃κB pathway mediates MPP+⁃induced apoptosis and autophagy in RSC96 cells

作　　者：李杨夏 佟晴[1] 程越 耿铫 王田田 张克忠[1] LI Yangxia;TONG Qing;CHENG Yue;GENG Yao;WANG Tiantian;ZHANG Kezhong(Department of Neurology,the First Affiliated Hospital of Nanjing Medical University,Nanjing 210029;Department of Pharmacology,School of Basic Medical Sciences,Nanjing Medical University,Nanjing 211166,China)

机构地区：[1]南京医科大学第一附属医院神经内科,江苏南京210029 [2]南京医科大学基础医学院药理学系,江苏南京211166

出　　处：《南京医科大学学报（自然科学版）》2023年第12期1623-1629,共7页Journal of Nanjing Medical University(Natural Sciences)

基　　金：国家自然科学基金(8207051941)。

摘　　要：目的:探讨1⁃甲基⁃4⁃苯基⁃吡啶离子(1⁃methyl⁃4⁃phenyl⁃pyridinium,MPP+)诱导的RSC96细胞凋亡与自噬功能障碍与TLR2/NF⁃κB信号通路的关系。方法:将RSC96细胞分为PBS组、MPP+组和MPP++CU⁃CPT22组;CCK⁃8检测不同MPP+浓度(0.1、0.3、0.5、0.7、0.9 mmol/L)处理后细胞存活率;TUNEL检测细胞凋亡水平;RT⁃qPCR检测Toll样受体2(TLR2)mRNA转录水平;Western blot检测凋亡相关指标Bcl⁃2/Bax、cleaved caspase⁃3/caspase⁃3,自噬相关指标LC3Ⅱ/LC3Ⅰ和P62,以及TLR2、p⁃NF⁃κB/NF⁃κB蛋白表达水平。结果:与PBS组相比,MPP+组细胞存活率下降且呈浓度依赖性,TUNEL染色阳性细胞数增多,Bcl⁃2/Bax蛋白比值水平下降,cleaved caspase⁃3/caspase⁃3比值增高。同时LC3Ⅱ/LC3Ⅰ比值下降,P62表达增加,p⁃NF⁃κB/NF⁃κB比值表达增加。RT⁃qPCR和Western blot结果显示,MPP+上调TLR2的表达。此外,与MPP+组相比,MPP++CU⁃CPT22组TUNEL阳性细胞数目减少,Bcl⁃2/Bax比值升高,cleaved caspase⁃3/caspase⁃3比值降低;同时,LC3Ⅱ/LC3Ⅰ比值上升,P62水平下降。结论:MPP+刺激诱导RSC96细胞凋亡和自噬水平失衡,发生机制可能与TLR2/NF⁃κB通路的激活有关。Objective:To investigate the relationship between TLR2/NF⁃κB signaling pathway and the apoptosis and autophagy dysfunction of RSC96 cells induced by 1⁃methyl⁃4⁃phenyl⁃pyridinium(MPP+).Methods:RSC96 cells were divided into the PBS group,MPP+group,and MPP++CU⁃CPT22 group.Cell survival rate was detected using CCK⁃8 after treatment with different MPP+concentrations(0.1,0.3,0.5,0.7,0.9 mmol/L).Cell apoptosis was detected by TUNEL staining.RT⁃qPCR was performed to detect the TLR2 mRNA level.Western blot was performed to detect the expression levels of apoptosis related indicators Bcl⁃2/Bax,cleaved caspase⁃3/caspase⁃3,autophagy⁃related indicators LC3II/LC3I and P62,as well as TLR2 and p⁃NF⁃κB/NF⁃κB.Results:Compared with the PBS group,the cell viability of the MPP+group decreased in a concentration⁃dependent manner,the number of TUNEL⁃staining positive cells increased,the ratio of Bcl⁃2/Bax decreased while cleaved caspase⁃3/caspase⁃3 ratio increased,as well as had a decrease in the ratio of LC3Ⅱ/LC3Ⅰand an increase in P62 and p⁃NF⁃κB/NF⁃κB ratio elvel.RT⁃qPCR and Western blot results showed that MPP+upregulated the expression of TLR2.In addition,compared with the MPP+group,the MPP++CU⁃CPT22 group showed a decrease in the number of TUNEL⁃staining positive cells,an increase in Bcl⁃2/Bax level,and a decrease in cleaved caspase⁃3/caspase⁃3 ratio.Meanwhile,LC3Ⅱ/LC3Ⅰratio was increased,and the P62 expression level was decreased.Conclusion:MPP+stimulation induced apoptosis and the imbalance of autophagy in RSC96 cells,and the mechanism may be related to the activation of the TLR2/NF⁃κB signaling pathway.

关 键 词：帕金森病 雪旺细胞 TOLL样受体2 自噬 凋亡 

分 类 号：R285.5[医药卫生—中药学]