Parkin通过介导PINK 1/Parkin线粒体自噬信号通路加速小鼠帕金森病发展及加剧神经炎症发生

doi:10.12122/j.issn.1673-4254.2024.12.11

南方医科大学学报 ›› 2024, Vol. 44 ›› Issue (12): 2359-2366.doi: 10.12122/j.issn.1673-4254.2024.12.11

• • 上一篇

Parkin通过介导PINK 1/Parkin线粒体自噬信号通路加速小鼠帕金森病发展及加剧神经炎症发生

姜诚诚¹(), 李洋洋¹, 段可欣¹, 詹婷婷¹, 陈子龙¹, 王永雪², 赵蕊², 马彩云², 郭俣¹^,³, 刘长青¹^,²()

^1.蚌埠医科大学，安徽省神经再生技术与医用新材料工程研究中心，安徽蚌埠 233000
^2.蚌埠医科大学，生命科学学院，安徽蚌埠 233000
^3.蚌埠医科大学，检验医学院，安徽蚌埠 233000

收稿日期:2024-06-08 出版日期:2024-12-20 发布日期:2024-12-26
通讯作者: 刘长青 E-mail:15955225712@163.com;lcq7813@bbmc.edu.cn
作者简介:姜诚诚，在读硕士研究生，E-mail: 15955225712@163.com
基金资助:
国家自然科学基金(82371382);安徽省重点研究与开发计划项目(2022e07020030);安徽省自然科学基金(2308085MH256);蚌埠医科大学重大科技项目孵育计划(2021byfy002);国家级大学生创新创业训练项目资助(202310367005)

Parkin deletion affects PINK1/Parkin-mediated mitochondrial autophagy to exacerbate neuroinflammation and accelerate progression of Parkinson's disease in mice

Chengcheng JIANG¹(), Yangyang LI¹, Kexin DUAN¹, Tingting ZHAN¹, Zilong CHEN¹, Yongxue WANG², Rui ZHAO², Caiyun MA², Yu GUO¹^,³, Changqing LIU¹^,²()

^1.Anhui Provincial Center for Neural Regeneration Technology and New Medical Materials Engineering Research, Bengbu Medical University, Bengbu 233000, China
^2.School of Life Sciences, Bengbu Medical University, Bengbu 233000, China
^3.School of Laboratory Medicine, Bengbu Medical University, Bengbu 233000, China

Received:2024-06-08 Online:2024-12-20 Published:2024-12-26
Contact: Changqing LIU E-mail:15955225712@163.com;lcq7813@bbmc.edu.cn
Supported by:
National Natural Science Foundation of China(82371382)

摘要/Abstract

摘要：

目的探讨E3 泛素连接酶Parkin缺失造成的线粒体自噬障碍在帕金森病（PD）神经炎症中的作用。方法建立MPTP-PD模型，野生型（WT）小鼠和Parkin^-/-小鼠连续腹腔注射MPTP 5 d建立PD小鼠模型，分别设置对照组：WT-PBS组、Parkin^-/--PBS组；PD建模组：WT-MPTP组、Parkin^-/--MPTP组，8只/组，对照组连续5 d注射等量的PBS。建模1周后，通过旷场实验评估WT小鼠与Parkin^-/-小鼠的运动行为；通过脑切片免疫荧光和Western blotting检测 Parkin缺失对PD发展以及神经炎症的影响；通过PINK 1/Parkin信号通路变化，探讨Parkin调控线粒体自噬对PD神经炎症发生的分子机制。结果与WT-MPTP组相比，Parkin^-/--MPTP组小鼠运动功能下降（P<0.001），脑内TH+神经元减少并且α-突触核蛋白（α-syn）积累增加；神经炎症相关GFAP和I-ba1阳性细胞数量增加（P<0.001）。Parkin缺失影响PINK1/Parkin介导的线粒体自噬，导致mtDNA增多以及炎症相关蛋白STING和NLRP3炎症小体的表达上调（P<0.01）。结论 Parkin通过调节PINK1/Parkin信号通路加速了小鼠帕金森病发展及神经炎症发生，为后期研究Parkin基因隐性遗传早发性帕金森病发病机制及其治疗奠定了实验基础。

关键词: 帕金森病, Parkin, 神经炎症, 线粒体自噬

Abstract:

Objective To investigate the role of mitochondrial autophagy disorder caused by deletion of E3 ubiquitin ligase Parkin in neuroinflammation in a mouse model of MPTP-induced Parkinson's disease (PD). Methods Wild-type (WT) male C57BL/6 mice and Parkin^-/-mice were given intraperitoneal injections with MPTP or PBS for 5 consecutive days, and the changes in motor behaviors of the mice were observed using open field test. The effects of Parkin deletion on PD development and neuroinflammation were evaluated using immunofluorescence and Western blotting. The changes of the PINK 1/Parkin signaling pathway in the midbrain substantia nigra of the mice were examined to explore the molecular mechanism of Parkin-mediated regulation of mitochondrial autophagy and its effect on neuroinflammation in PD mice. Results Compared with their WT counterparts, the Parkin^-/- mice with MPTP injections exhibited significant impairment of motor function with decreased TH⁺ neurons, increased α-synuclein (α-syn) accumulation, and increased numbers of GFAP⁺ and I-ba1⁺ cells in the midbrain substantia nigra. Parkin deletion obviously affected PINK1/Parkin-mediated mitochondrial autophagy to result in significantly increased mtDNA and upregulated expressions of STING and NLRP3 inflammatosomes in the midbrain substantia nigra of MPTP-treated transgenic mice. Conclusion Parkin deletion causes mitochondrial autophagy disorder to accelerate PD progression and exacerbates neuroinflammation in mice by affecting the PINK1/Parkin signaling pathway, suggesting the important role of Parkin in early pathogenesis of PD.

Key words: Parkinson's disease, Parkin, neuroinflammation, mitochondrial autophagy

姜诚诚, 李洋洋, 段可欣, 詹婷婷, 陈子龙, 王永雪, 赵蕊, 马彩云, 郭俣, 刘长青. Parkin通过介导PINK 1/Parkin线粒体自噬信号通路加速小鼠帕金森病发展及加剧神经炎症发生[J]. 南方医科大学学报, 2024, 44(12): 2359-2366.

Chengcheng JIANG, Yangyang LI, Kexin DUAN, Tingting ZHAN, Zilong CHEN, Yongxue WANG, Rui ZHAO, Caiyun MA, Yu GUO, Changqing LIU. Parkin deletion affects PINK1/Parkin-mediated mitochondrial autophagy to exacerbate neuroinflammation and accelerate progression of Parkinson's disease in mice[J]. Journal of Southern Medical University, 2024, 44(12): 2359-2366.

图/表 4

图1 WT及Parkin-/-小鼠注射MPTP后运动行为测试

Fig.1 Behavioral test of wild-type (WT) and Parkin-/- mice after MPTP injection. A: Flow chart of MPTP-PD modeling in the mice. B: Heat map of movement of the mice in open field tests. C: Motion trajectory graph of the mice. D, E: Statistics of total movement distance and the distance in the central area of the mice in open field test. **P<0.01, ***P<0.001.

图2 WT和Parkin-/-小鼠MPTP诱导的帕金森病变

Fig.2 MPTP-induced Parkinson's disease in WT and Parkin-/- mice. A: Immunofluorescence staining of dopaminergic neuron markers TH (red) and DAPI (blue). B: Protein expression level of TH in the midbrain substantia nigra. C: Analysis of TH protein expression. D: Immunofluorescence staining of α-syn (red) and DAPI (blue) in the mouse midbrain. **P<0.01, ***P<0.001.

图3 Parkin-/-小鼠MPTP诱导神经炎症加剧

Fig.3 Parkin deletion increased neuroinflammation induced by MPTP in mice. A: Immunofluorescence staining of astrocyte markers GFAP (red) and DAPI (blue). B: Co-staining of microglial marker I-ba1 (red) and NLRP3 (green) inflammasomes. C: Analysis of GFAP+ cell count (%). D: Analysis of I-ba1+ cell count (%).**P<0.01, ***P<0.001, ****P<0.0001.

图4 Parkin调控线粒体自噬的分子机制

Fig.4 Molecular mechanism of Parkin-mediated regulation of mitochondrial autophagy in midbrain substantia nigra. A: Immunofluorescence assay showing abnormal mitochondrial autophagy (white arrows) in Parkin-/- mice with MPTP injection. B: Western blotting for detecting protein changes in Parkin-mediated regulation of mitochondrial autophagy and neuroinflammation. C: Scanning densitometry for semi-quantitative analysis of the protein expression levels. *P<0.05, **P<0.01 vs WT-MPTP.

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Parkin通过介导PINK 1/Parkin线粒体自噬信号通路加速小鼠帕金森病发展及加剧神经炎症发生

Parkin deletion affects PINK1/Parkin-mediated mitochondrial autophagy to exacerbate neuroinflammation and accelerate progression of Parkinson's disease in mice

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