褪黑素通过抑制NLRP3减轻MPP^(+)诱导的HT22细胞焦亡/炎性损害  

Melatonin alleviates MPP^(+)-induced pyroptosis/inflammatory damage of HT22 cells by inhibiting NOD-like receptor protein 3

作　　者：姬丽娅 柴佳林 饶维 王彦刚 程俊凯 王凯 王利 JI Li-Ya;CHAI Jia-Lin;RAO Wei;WANG Yang-Gang;CHENG Jun-Kai;WANG Kai;WANG Li(Neurological Department,Xi’an Third Hospital.Xi’an,Shaanxi 710032,China;Shaanxi University of Chinese Medicine,Xianyang,Shaanxi 712046,China;Senior Department of Neurosurgery,The First Medical Center,General Hospital of PLA,Beijing 100048,China;Department of Neurosurgery,Xijing Hospital,Air Force Medical University of PLA,Xi’an,Shaanxi 710032,China)

机构地区：[1]陕西省西安市第三医院,陕西西安710032 [2]陕西中医药大学,陕西咸阳712046 [3]中国人民解放军总医院第一医学中心神经外科医学部,北京100048 [4]中国人民解放军空军军医大学第一附属医院神经外科,陕西西安710032

出　　处：《国际神经病学神经外科学杂志》2022年第2期1-5,共5页Journal of International Neurology and Neurosurgery

基　　金：国家自然科学基金青年项目资助(81801165);陕西省自然科学基础研究计划项目(2017JM8073,2020JM-338);西安市卫健委青年培育科研项目(2022qn02);空军军医大学第一附属医院学科助推计划(XJZT19Z23)。

摘　　要：目的 研究褪黑素(MT)预处理抑制NOD样受体热蛋白结构域相关蛋白3(NLRP3)水平,减轻神经细胞焦亡,改善帕金森病症状的作用及分子机制。方法 正常培养的HT22细胞记为空白对照组(Sham组);细胞进行10 mmol/L 1-甲基-4-苯基-吡啶离子(MPP^(+))处理24 h,制成帕金森病细胞模型后,记为MPP^(+)组;先用5μmmol/L MT预处理6 h后,再造模后,记为MPP^(+)+MT组;各组细胞处理后经培养24 h后取材。采用CCK8检测细胞死亡变化;采用荧光免疫组化法和Western-blotting方法检测NLRP3分布和表达变化;使用标准ELISA试剂盒检测IL-1β和IL-18表达变化。结果 与Sham组相比,MPP^(+)组神经细胞大量死亡,NLRP3在存活细胞的胞膜和胞浆中高表达,相应的炎症因子IL-1β和IL-18也大量释放,两组间差异有统计学意义(P<0.05)。相较于MPP^(+)组,MPP^(+)+MT组细胞存活率升高,细胞中NLRP3染色阳性率降低,表达量也降低,相应的IL-1β和IL-18表达量也降低,两组间差异有统计学意义(P<0.05)。结论 MT可有效抑制NLRP3表达和炎症因子的释放,减轻神经细胞焦亡,从而减轻MPP^(+)造成的HT22细胞损伤。Objective To investigate the role and molecular mechanism of melatonin(MT) pretreatment in inhibiting the level of NOD-like receptor protein 3(NLRP3),reducing neuronal pyroptosis,and improving Parkinson’s disease.Methods Normally cultured HT22 cells were established as blank control group(sham group);the cells were treated with 10 mmol/L MPP^(+)for 24 hours to establish a cell model of Parkinson’s disease(MPP^(+)group);the cells pretreated with 5μmmol/L MT for 6 hours before modeling were established as MPP^(+)+MT group;samples were collected after the cells were cultured for 24 hours after treatment.CCK8 assay was used to measure the change in cell death;fluorescence immunohistochemistry and Western blotting were used to measure the changes in the distribution and expression of NLRP3;a standard ELISA kit was used to measure the changes in the expression of interleukin-1β(IL-1β) and interleukin-18(IL-18).Results Compared with the sham group,the MPP^(+)group had the death of a large number of nerve cells,a high expression level of NLRP3 in the membrane and cytoplasm of the surviving cells,and the secretion of large amounts of the corresponding inflammatory factors IL-1 β and IL-18(P<0.05).Compared with the MPP^(+)group,the MPP^(+)+MT group had a significant increase in cell viability and significant reductions in the positive rate of NLRP3 staining and the expression levels of NLRP3,IL-1β,and IL-18(P<0.05).Conclusions MT can effectively inhibit the expression of NLRP3 and the release of inflammatory factors,reduce nerve cell pyroptosis,and thus improve the damage of HT22 cells caused by MPP^(+).

关 键 词：帕金森病 褪黑素 NOD样受体热蛋白结构域相关蛋白3 白介素-1Β 白介素-18 

分 类 号：R742.5[医药卫生—神经病学与精神病学]