Division of Neurosurgery, University of Cambridge, Cambridge, United Kingdom.
Department of Clinical Neuroscience, Karolinska Institutet, Stockholm, Sweden.
J Neurotrauma. 2020 Jan 1;37(1):93-105. doi: 10.1089/neu.2019.6480. Epub 2019 Sep 18.
Neuroinflammation has been shown to mediate the pathophysiological response following traumatic brain injury (TBI). Accumulating evidence implicates astrocytes as key immune cells within the central nervous system (CNS), displaying both pro- and anti-inflammatory properties. The aim of this study was to investigate how human astrocyte cultures respond to cytokines across a concentration range that approximates the aftermath of human TBI. To this end, enriched cultures of human induced pluripotent stem cell (iPSC)-derived astrocytes were exposed to interleukin-1β (IL-1β) (1-10,000 pg/mL), IL-4 (1-10,000 pg/mL), IL-6 (100-1,000,000 pg/mL), IL-10 (1-10,000 pg/mL) and tumor necrosis factor (TNF)-α (1-10,000 pg/mL). After 1, 24, 48 and 72 h, cultures were fixed and immunolabeled, and the secretome/supernatant was analyzed at 24, 48, and 72 h using a human cytokine/chemokine 39-plex Luminex assay. Data were compared to previous studies of neuronal cultures and clinical TBI studies. The secretome revealed concentration-, time- and/or both concentration- and time-dependent production of downstream cytokines (29, 21, and 17 cytokines, respectively, <0.05). IL-1β exposure generated the most profound downstream response (27 cytokines), IL-6 and TNF had intermediate responses (13 and 11 cytokines, respectively), whereas IL-4 and IL-10 only led to weak responses over time or in escalating concentration (8 and 8 cytokines, respectively). Notably, expression of IL-1β, IL-6, and TNF cytokine receptor mRNA was higher in astrocyte cultures than in neuronal cultures. Several secreted cytokines had temporal trajectories, which corresponded to those seen in the aftermath of human TBI. In summary, iPSC-derived astrocyte cultures exposed to cytokine concentrations reflecting those in TBI generated an increased downstream cytokine production, particularly IL-1β. Although more work is needed to better understand how different cells in the CNS respond to the neuroinflammatory milieu after TBI, our data shows that iPSC-derived astrocytes represent a tractable model to study cytokine stimulation in a cell type-specific manner.
神经炎症已被证明介导创伤性脑损伤 (TBI) 后的病理生理反应。越来越多的证据表明星形胶质细胞是中枢神经系统 (CNS) 中的关键免疫细胞，具有促炎和抗炎特性。本研究旨在研究人类星形胶质细胞培养物如何在接近人类 TBI 后果的浓度范围内对细胞因子作出反应。为此，使用白细胞介素-1β (IL-1β) (1-10,000 pg/mL)、IL-4 (1-10,000 pg/mL)、IL-6 (100-1,000,000 pg/mL)、IL-10 (1-10,000 pg/mL) 和肿瘤坏死因子 (TNF)-α (1-10,000 pg/mL) 处理富含人诱导多能干细胞 (iPSC) 衍生星形胶质细胞的培养物。在 1、24、48 和 72 小时后，将培养物固定并免疫标记，并在 24、48 和 72 小时使用人细胞因子/趋化因子 39 合成分析试剂盒分析分泌组/上清液。数据与先前的神经元培养物和临床 TBI 研究进行了比较。分泌组显示下游细胞因子的浓度、时间和/或浓度和时间依赖性产生 (分别为 29、21 和 17 种细胞因子，<0.05)。IL-1β 暴露产生最显著的下游反应 (27 种细胞因子)，IL-6 和 TNF 具有中等反应 (分别为 13 和 11 种细胞因子)，而 IL-4 和 IL-10 仅随时间或浓度增加产生微弱反应 (分别为 8 和 8 种细胞因子)。值得注意的是，星形胶质细胞培养物中 IL-1β、IL-6 和 TNF 细胞因子受体 mRNA 的表达高于神经元培养物。一些分泌细胞因子的时间轨迹与人类 TBI 后的时间轨迹相对应。总之，暴露于反映 TBI 中细胞因子浓度的 iPSC 衍生星形胶质细胞培养物产生了增加的下游细胞因子产生，特别是 IL-1β。尽管需要进一步的工作来更好地了解 TBI 后中枢神经系统中的不同细胞如何对神经炎症环境作出反应，但我们的数据表明，iPSC 衍生的星形胶质细胞代表了一种可行的模型，可以研究细胞类型特异性的细胞因子刺激。
