Department of Neurology, Beijing Chao-yang Hospital, Capital Medical University, No. 8, Gongti South Road, Beijing, 100020, China.
J Mol Neurosci. 2020 Dec;70(12):1977-1986. doi: 10.1007/s12031-020-01599-4. Epub 2020 Jul 2.
The objective of this study was to understand if RNF13 can affect Parkinson's disease (PD) model mice by modulating the endoplasmic reticulum stress (ERS)-mediated IRE1α-TRAF2-ASK1-JNK pathway. C57BL/6 mice injected with MPTP to establish PD mice models were divided into Control, MPTP, MPTP + sh-RNF13, and MPTP + sh-NC groups. Rotarod, balance beam, and open-field tests were used to assess the behavioral changes of experimental mice. Immunofluorescence assay was used to determine TH-positive expression in substantia nigra, TUNEL staining to detect apoptosis, and Western blotting to measure the expression of IRE1α-TRAF2-ASK1-JNK pathway. Besides, SH-SY5Y cells treated with MPP were assigned into Control, MPP, MPP + sh-RNF13, and MPP + sh-NC groups in vitro to detect cell viability, apoptosis and Ca level. When compared with those Control mice, MPTP mice showed decreased retention time spent on rotarod performance and prolonged time on balance beam test, as well as evident reductions in floor plane (FP) movements, moving time, moving distance, and mean velocity in open-field test, which had an obvious increase of TUNEL-positive cells, significant decrease of TH-positive cells, and remarkable up-regulations of RNF13, p-IRE1α/IRE1α, TRAF2, ASK1, and p-JNK/JNK. Meanwhile, MPTP mice treated with sh-RNF13 were improved in all above indexes. In vitro, MPP treated SH-SY5Y cells had decreased cell viability and increased cell apoptosis, as well as the upregulated IRE1α-TRAF2-ASK1-JNK pathway proteins and Ca level. RNF13 knockdown improved all above indexes in SH-SY5Y cells treated with MPP. Silencing RNF13 can alleviate motor dysfunction and dopamine neuronal damage in PD mice by inhibiting ERS-mediated IRE1α-TRAF2-ASK1-JNK pathway.
本研究旨在探讨 RNF13 是否可以通过调节内质网应激（ERS）介导的 IRE1α-TRAF2-ASK1-JNK 通路来影响帕金森病（PD）模型小鼠。将 C57BL/6 小鼠注射 MPTP 建立 PD 小鼠模型，分为对照组、MPTP 组、MPTP+sh-RNF13 组和 MPTP+sh-NC 组。采用转棒、平衡木和旷场实验评估实验小鼠的行为变化。免疫荧光法检测黑质中 TH 阳性表达，TUNEL 染色检测细胞凋亡，Western blot 检测 IRE1α-TRAF2-ASK1-JNK 通路的表达。此外，体外将 MPP 处理的 SH-SY5Y 细胞分为对照组、MPP 组、MPP+sh-RNF13 组和 MPP+sh-NC 组，检测细胞活力、细胞凋亡和 Ca 水平。与对照组相比，MPTP 组小鼠在转棒实验中的保留时间减少，平衡木测试时间延长，旷场实验中的 FP 运动减少，运动时间、运动距离和平均速度明显降低，TUNEL 阳性细胞增多，TH 阳性细胞减少，RNF13、p-IRE1α/IRE1α、TRAF2、ASK1 和 p-JNK/JNK 表达明显上调。同时，MPTP 组经 sh-RNF13 处理后，上述指标均有所改善。体外，MPP 处理的 SH-SY5Y 细胞活力降低，细胞凋亡增加，IRE1α-TRAF2-ASK1-JNK 通路蛋白和 Ca 水平上调。MPP 处理的 SH-SY5Y 细胞中沉默 RNF13 可改善上述所有指标。沉默 RNF13 可通过抑制 ERS 介导的 IRE1α-TRAF2-ASK1-JNK 通路，减轻 PD 小鼠的运动功能障碍和多巴胺神经元损伤。
