Human iPSCs derived astrocytes rescue rotenone-induced mitochondrial dysfunction and dopaminergic neurodegeneration in vitro by donating functional mitochondria
Human iPSCs derived astrocytes rescue rotenone-induced mitochondrial dysfunction and dopaminergic neurodegeneration in vitro by donating functional mitochondria
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DOI：
10.1186/s40035-020-00190-6
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发表时间：
2020-04-24
影响因子：
12.6
通讯作者：
Deng, Wen-Bin
中科院分区：
文献类型：
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作者：
Cheng, Xiao-Yu;Biswas, Sangita;Deng, Wen-Bin
研究背景帕金森病（Parkinson 'sDisease，PD）是一种以脑内黑质多巴胺能（Dopaminergic，DA）神经元逐渐丧失为特征的神经退行性疾病。大量证据表明，在细胞水平上，线粒体功能障碍是导致病理特征如神经元死亡和错误折叠的α-突触核蛋白聚集体积累的关键因素。健康的纯化线粒体的自体移植已经显示出在PD的体外和体内模型中减弱表型。然而，要获得大量具有正常功能的纯化线粒体存在重大的技术困难。此外，线粒体的半衰期在几天到几周之间变化。因此，通过细胞间线粒体转移鉴定健康线粒体的连续来源是用于治疗目的的有吸引力的选择。在这项研究中，我们询问了iPSC衍生的星形胶质细胞是否可以作为供体提供功能性线粒体并在PD的体外模型中暴露于鱼藤酮后拯救受损的DA神经元。方法我们从人iPSC和hESC中产生DA神经元和星形胶质细胞。我们建立了星形胶质细胞-神经元共培养系统，研究细胞间线粒体转移，以及线粒体转移的神经保护作用。我们采用免疫细胞化学和流式细胞仪分析跟踪线粒体。结果在鱼藤酮诱导的体外PD模型中，iPSC来源的星形胶质细胞或星形胶质细胞条件培养基（ACM）可以通过线粒体转移挽救DA神经元的变性。具体来说，我们发现来自健康的iPSC衍生的星形胶质细胞自发地将功能性线粒体释放到培养基中。在ACM中检测到Mito-Tracker绿色标记的星形胶质细胞线粒体，并显示其通过磷酸化p38依赖的途径被损伤的神经元内化。转移线粒体能够显着逆转DA神经变性和轴突修剪暴露于鱼藤酮。当鱼藤酮损伤的神经元培养在ACM耗尽线粒体（超滤）的存在下，神经保护作用被取消。结论我们的研究为iPSC源性星形胶质细胞作为线粒体供体修复损伤的DA神经元并减轻病理提供了理论依据。使用iPSC衍生的星形胶质细胞作为供体可以提供一种新的策略，可以进一步开发用于PD的细胞治疗。
Background Parkinson's disease (PD) is one of the neurodegeneration diseases characterized by the gradual loss of dopaminergic (DA) neurons in the substantia nigra region of the brain. Substantial evidence indicates that at the cellular level mitochondrial dysfunction is a key factor leading to pathological features such as neuronal death and accumulation of misfolded alpha-synuclein aggregations. Autologous transplantation of healthy purified mitochondria has shown to attenuate phenotypes in vitro and in vivo models of PD. However, there are significant technical difficulties in obtaining large amounts of purified mitochondria with normal function. In addition, the half-life of mitochondria varies between days to a few weeks. Thus, identifying a continuous source of healthy mitochondria via intercellular mitochondrial transfer is an attractive option for therapeutic purposes. In this study, we asked whether iPSCs derived astrocytes can serve as a donor to provide functional mitochondria and rescue injured DA neurons after rotenone exposure in an in vitro model of PD. Methods We generated DA neurons and astrocytes from human iPSCs and hESCs. We established an astroglial-neuronal co-culture system to investigate the intercellular mitochondrial transfer, as well as the neuroprotective effect of mitochondrial transfer. We employed immunocytochemistry and FACS analysis to track mitochondria. Results We showed evidence that iPSCs-derived astrocytes or astrocytic conditioned media (ACM) can rescue DA neurons degeneration via intercellular mitochondrial transfer in a rotenone induced in vitro PD model. Specifically, we showed that iPSCs-derived astrocytes from health spontaneously release functional mitochondria into the media. Mito-Tracker Green tagged astrocytic mitochondria were detected in the ACM and were shown to be internalized by the injured neurons via a phospho-p38 depended pathway. Transferred mitochondria were able to significantly reverse DA neurodegeneration and axonal pruning following exposure to rotenone. When rotenone injured neurons were cultured in presence of ACM depleted of mitochondria (by ultrafiltration), the neuroprotective effects were abolished. Conclusions Our studies provide the proof of principle that iPSCs-derived astrocytes can act as mitochondria donor to the injured DA neurons and attenuate pathology. Using iPSCs derived astrocytes as a donor can provide a novel strategy that can be further developed for cellular therapy for PD.