Astrocytes respond to injury and disease through a process known as reactive astrogliosis, of which inflammatory signaling is one subset. This inflammatory response is heterogeneous with respect to the inductive stimuli and the afflicted central nervous system region. This is of plausible importance in e.g. traumatic axonal injury (TAI), where lesions in the brainstem carries a particularly poor prognosis. In fact, astrogliotic forebrain astrocytes were recently suggested to cause neuronal death following axotomy. We therefore sought to assess if ventral brainstem- or rostroventral spinal astrocytes exert similar effects on motor neurons in vitro. We derived brainstem/rostroventral spinal astrocyte-like cells (ES-astrocytes) and motor neurons using directed differentiation of mouse embryonic stem cells (ES). We activated the ES-astrocytes using the neurotoxicity-eliciting cytokines interleukin- (IL-) 1α and tumor necrosis factor-(TNF-)α and clinically relevant inflammatory mediators. In co-cultures with reactive ES-astrocytes and motor neurons, we assessed neurotoxic ES-astrocyte activity, similarly to what has previously been shown for other central nervous system (CNS) regions. We confirmed the brainstem/rostroventral ES-astrocyte identity using RNA-sequencing, immunocytochemistry, and by comparison with primary subventricular zone-astrocytes. Following cytokine stimulation, the c-Jun N-terminal kinase pathway down-stream product phosphorylated c-Jun was increased, thus demonstrating ES-astrocyte reactivity. These reactive ES-astrocytes conferred a contact-dependent neurotoxic effect upon co-culture with motor neurons. When exposed to IL-1β and IL-6, two neuroinflammatory cytokines found in the cerebrospinal fluid and serum proteome following human severe traumatic brain injury (TBI), ES-astrocytes exerted similar effects on motor neurons. Activation of ES-astrocytes by these cytokines was associated with pathways relating to endoplasmic reticulum stress and altered regulation of MYC. Ventral brainstem and rostroventral spinal cord astrocytes differentiated from mouse ES can exert neurotoxic effects in vitro. This highlights how neuroinflammation following CNS lesions can exert region- and cell-specific effects. Our in vitro model system, which uniquely portrays astrocytes and neurons from one niche, allows for a detailed and translationally relevant model system for future studies on how to improve neuronal survival in particularly vulnerable CNS regions following e.g. TAI.

中文翻译：

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干细胞来源的脑干小鼠星形胶质细胞在神经炎症时在体外获得神经毒性表型

星形胶质细胞通过一种称为反应性星形胶质细胞增生的过程对损伤和疾病做出反应，炎症信号传导是其中的一个子集。这种炎症反应对于诱导刺激和受影响的中枢神经系统区域是异质的。这对于创伤性轴突损伤（TAI）等脑干损伤的预后特别差来说似乎很重要。事实上，最近有人认为，星形胶质细胞前脑星形胶质细胞在轴索切除术后会导致神经元死亡。因此，我们试图评估腹侧脑干或口腹脊髓星形胶质细胞是否对体外运动神经元发挥类似的作用。我们利用小鼠胚胎干细胞（ES）的定向分化衍生出脑干/前腹脊髓星形胶质细胞（ES-星形胶质细胞）和运动神经元。我们使用引起神经毒性的细胞因子白细胞介素-(IL-)1α和肿瘤坏死因子-(TNF-)α以及临床相关的炎症介质激活ES-星形胶质细胞。在与反应性 ES 星形胶质细胞和运动神经元的共培养中，我们评估了神经毒性 ES 星形胶质细胞的活性，类似于之前对其他中枢神经系统 (CNS) 区域的研究。我们通过 RNA 测序、免疫细胞化学并与原发性室下区星形胶质细胞进行比较，确认了脑干/前腹侧 ES 星形胶质细胞的身份。细胞因子刺激后，c-Jun N 端激酶途径下游产物磷酸化的 c-Jun 增加，从而证明 ES 星形胶质细胞的反应性。这些反应性 ES 星形胶质细胞在与运动神经元共培养时产生接触依赖性神经毒性作用。当暴露于人类严重创伤性脑损伤 (TBI) 后脑脊液和血清蛋白质组中发现的两种神经炎症细胞因子 IL-1β 和 IL-6 时，ES 星形胶质细胞对运动神经元产生类似的影响。这些细胞因子对 ES 星形胶质细胞的激活与内质网应激和 MYC 调节改变相关的途径有关。小鼠ES分化的腹侧脑干和前腹脊髓星形胶质细胞在体外可发挥神经毒性作用。这凸显了中枢神经系统损伤后的神经炎症如何发挥区域和细胞特异性作用。我们的体外模型系统独特地描绘了来自一个利基的星形胶质细胞和神经元，