MKP1通过抑制JNK信号途径减轻淀粉样蛋白的神经毒性作用

MKP1 attenuates amyloid beta-induced neurotoxicity via suppressing JNK pathway

目的研究MKP1在Aβ所致MAPK激活、神经炎症和细胞凋亡中的调控作用。方法在细胞培养基中加入不同浓度的Aβ42(0μmol/L,0.1μmol/L,1μmol/L,10μmol/L和100 v),处理24 h后CCK-8检测细胞活性。向细胞培养基中加入10μmol/L的Aβ42,在不同时间点(0 h、6 h、12 h、18 h和24 h)通过qRT-PCR和Western blot检测MKP1表达。将野生型PC12细胞分为对照组(Control)和Aβ42组,敲除MKP1的细胞为MKP1 KD+Aβ42组,过表达MKP1细胞为MKP1+Aβ组,后3组加入10μmol/L Aβ42,置于培养箱中24 h,通过线粒体膜电位、DCFH-DA以及超氧化物歧化酶活性和丙二醛检测评价细胞内氧化应激水平,通过qRT-PCR检测TNF-α和IL-1β表达,Western blot检测p-JNK水平。结果发现经Aβ刺激后,PC12细胞活性受到抑制,MAPK的重要调节因子MKP1表达下调,并呈时间依赖性。过表达MKP1后,Aβ诱导的PC12细胞中p-JNK水平降低,细胞内活性氧类水平下降,TNF-α和IL-1β表达减少。而敲除MKP1可加重Aβ诱导的PC12氧化应激和炎症反应。结论 Aβ通过下调MKP1表达激活MAPK信号途径,过表达MKP1可通过抑制JNK信号途径减轻Aβ所诱导的氧化应激和神经炎症从而发挥神经保护作用。

ObjectiveTo investigate the role of MKP1in MAPK activation,neuroinflammation and apoptosis induced by Aβ.MethodsAβ42(0μmol/L0.1μmol/L,1μmol/L,10μmol/L and100μmol/L)was added to the cell culture medium.CCK8activity was measured24hours after treatment.The expression of MKP1was detected by qRT PCR and Western blot at different time points(0h,6h,12h,18h and24h)in PC12cells treated with10μmol Aβ42.The wild type PC12cells were divided into control group and Aβ42group.MKP1knockdown cells were named MKP1KD+Aβ42group,whereas MKP1overexpressed cells were named MKP1+Aβgroup.10μmol/L Aβ42cells were added to the last3groups and the cells were incubated in the incubator.The activity of cells were detected by CCK8and the levels of mitochondrial membrane potential,DCFH DA,SOD activity and malondialdehyde(MDA)were measured to evaluate the intracellular oxidative stress.ResultsAβstimulation resulted in reduced PC12cell viability and downregulation of mitogen activated protein kinase phosphatase1(MKP1).MKP1over expression inhibited Aβinduced phosphorylation of c Jun N terminal kinase(JNK),elevation of reactive oxygen species(ROS),expression of TNFαand IL1β,and apoptosis in PC12cells.In contrast,MKP1knockdown by RNA interference aggravated Aβinduced oxidative stress,inflammation and cell damage in PC12cells.ConclusionThese results showed that Aβactivated the JNK signal pathway by down regulating MKP1expression,and MKP1reactivation could alleviate Aβinduced oxidative stress and neuroinflammation by inhibiting the JNK signaling pathway to play a neuroprotective effect.