Department of Rehabilitation Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China; School of rehabilitation medicine, southern medical university, Guangzhou, China.
Department of Rehabilitation, Zhengzhou Central Hospital Affiliated to Zhengzhou University, Zhengzhou, China.
Exp Neurol. 2021 Jul;341:113700. doi: 10.1016/j.expneurol.2021.113700. Epub 2021 Mar 17.
BACKGROUND: Pyroptosis mediated by NLRP3 inflammasome plays a critical role in the pathogenesis of cerebral ischemia-reperfusion (I/R) injury. Mounting evidences have verified the efficacy of exosomes by relieving the inflammatory response during cerebral I/R injury, but the specific mechanism has not been well elucidated. This study aimed to clarify whether the neuroprotective effects of bone marrow mesenchymal stem cell-derived exosomes (BMSC-Exos) are associated with the attenuation of NLPR3-mediated neuron pyroptosis by modulating microglial polarization. METHODS: Rats were initially subjected to middle cerebral artery occlusion (MCAO) followed by reperfusion. Then, BMSC-Exos were administered intravenously 2 h after MCAO. The neuroprotective effects were measured using a modified neurological severity score(mNSS), triphenyltetrazolium chloride (TTC) staining, brain water content, Morris water maze,and CatWalk system. Western blotting and immunofluorescence staining were applied to detect NLRP3 inflammasome and pyroptosis. Microglial polarization was determined by real-time polymerase chain reaction (RT-PCR) and immunofluorescence staining. To mimic cerebral I/R injury in vitro, BV2 and PC12 cells were exposed to oxygen-glucose deprivation/reoxygenation. After treatment with PBS, BMSC-Exos, IL-4, or LPS, BV2 cells were co-cultured with PC12 cells in a Transwell system. RESULTS: BMSC-Exos reduced the brain infarct area and brain water content at 24 h dose dependently and improved the neurological function up to 5 weeks after stroke. In vivo, NLRP3 inflammasome- and pyroptosis-related proteins were mainly expressed on neurons and downregulated by BMSC-Exos. Furthermore, cerebral I/R injury-induced M1-polarized microglia could be shifted toward M2 phenotype by BMSC-Exos. In vitro, BMSC-Exos alleviated the neuron pyroptosis partially by modulating microglial polarization. CONCLUSION: BMSC-Exos could ameliorate cerebral I/R injury via suppression of NLRP3 inflammasome-mediated inflammation and pyroptosis by modulating microglial polarization.
背景：NLRP3 炎性体介导热缺血再灌注（I/R）损伤中的细胞焦亡发挥着关键作用。越来越多的证据证实了外泌体通过减轻脑 I/R 损伤中的炎症反应的功效，但具体机制尚不清楚。本研究旨在阐明骨髓间充质干细胞源性外泌体（BMSC-Exos）的神经保护作用是否与通过调节小胶质细胞极化来减轻 NLPR3 介导的神经元细胞焦亡有关。
方法：大鼠首先进行大脑中动脉闭塞（MCAO），然后再进行再灌注。MCAO 后 2 小时静脉给予 BMSC-Exos。通过改良神经功能缺损评分（mNSS）、氯化三苯基四氮唑（TTC）染色、脑水含量、Morris 水迷宫和 CatWalk 系统来测量神经保护作用。Western blot 和免疫荧光染色用于检测 NLRP3 炎性体和细胞焦亡。通过实时聚合酶链反应（RT-PCR）和免疫荧光染色来确定小胶质细胞极化。体外模拟脑 I/R 损伤，将 BV2 和 PC12 细胞进行氧葡萄糖剥夺/复氧。用 PBS、BMSC-Exos、IL-4 或 LPS 处理后，将 BV2 细胞在 Transwell 系统中与 PC12 细胞共培养。
结果：BMSC-Exos 呈剂量依赖性地减少脑梗死面积和脑水含量，并且在卒中后 5 周内改善神经功能。在体内，NLRP3 炎性体和细胞焦亡相关蛋白主要在神经元上表达，并被 BMSC-Exos 下调。此外，BMSC-Exos 可以将脑 I/R 损伤诱导的 M1 极化小胶质细胞向 M2 表型转化。体外，BMSC-Exos 通过调节小胶质细胞极化部分缓解神经元细胞焦亡。
结论：BMSC-Exos 通过调节小胶质细胞极化来抑制 NLRP3 炎性体介导的炎症和细胞焦亡，从而改善脑 I/R 损伤。
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