Santiago Juliet V, Natu Aditya, Ramelow Christina C, Rayaprolu Sruti, Xiao Hailian, Kumar Vishnu, Seyfried Nicholas T, Rangaraju Srikant
Department of Neurology, Emory University, 201 Dowman Drive Atlanta, Georgia, 30322, United States of America.
Center for Neurodegenerative Diseases, Emory University, Atlanta, GA 30322, USA.
bioRxiv. 2023 Jul 29:2023.07.28.551012. doi: 10.1101/2023.07.28.551012.
Microglia are resident immune cells of the brain that play important roles in mediating inflammatory responses in several neurological diseases via direct and indirect mechanisms. One indirect mechanism may involve extracellular vesicle (EV) release, so that the molecular cargo transported by microglia-derived EVs can have functional effects by facilitating intercellular communication. The molecular composition of microglia-derived EVs, and how microglial activation states impacts EV composition and EV-mediated effects in neuroinflammation, remain poorly understood. We hypothesize that microglia-derived EVs have unique molecular profiles that are determined by microglial activation state. Using size-exclusion chromatography to purify EVs from BV2 microglia, combined with proteomic (label-free quantitative mass spectrometry or LFQ-MS) and transcriptomic (mRNA and non-coding RNA seq) methods, we obtained comprehensive molecular profiles of microglia-derived EVs. LFQ-MS identified several classic EV proteins (tetraspanins, ESCRT machinery, and heat shock proteins), in addition to over 200 proteins not previously reported in the literature. Unique mRNA and microRNA signatures of microglia-derived EVs were also identified. After treating BV2 microglia with lipopolysaccharide (LPS), interleukin-10, or transforming growth factor beta, to mimic pro-inflammatory, anti-inflammatory, or homeostatic states, respectively, LFQ-MS and RNA seq revealed novel state-specific proteomic and transcriptomic signatures of microglia-derived EVs. Particularly, LPS treatment had the most profound impact on proteomic and transcriptomic compositions of microglia-derived EVs. Furthermore, we found that EVs derived from LPS-activated microglia were able to induce pro-inflammatory transcriptomic changes in resting responder microglia, confirming the ability of microglia-derived EVs to relay functionally-relevant inflammatory signals. These comprehensive microglia-EV molecular datasets represent important resources for the neuroscience and glial communities, and provide novel insights into the role of microglia-derived EVs in neuroinflammation.
小胶质细胞是大脑中的常驻免疫细胞，通过直接和间接机制在多种神经疾病中介导炎症反应中发挥重要作用。一种间接机制可能涉及细胞外囊泡（EV）的释放，因此小胶质细胞衍生的EV所运输的分子货物可通过促进细胞间通讯而产生功能效应。小胶质细胞衍生的EV的分子组成，以及小胶质细胞激活状态如何影响神经炎症中EV的组成和EV介导的效应，仍知之甚少。我们假设小胶质细胞衍生的EV具有由小胶质细胞激活状态决定的独特分子谱。利用尺寸排阻色谱法从BV2小胶质细胞中纯化EV，并结合蛋白质组学（无标记定量质谱或LFQ-MS）和转录组学（mRNA和非编码RNA测序）方法，我们获得了小胶质细胞衍生的EV的全面分子谱。LFQ-MS除了鉴定出文献中先前未报道的200多种蛋白质外，还鉴定出几种经典的EV蛋白（四跨膜蛋白、ESCRT机制和热休克蛋白）。还鉴定了小胶质细胞衍生的EV独特的mRNA和微小RNA特征。在用脂多糖（LPS）、白细胞介素-10或转化生长因子β分别处理BV2小胶质细胞以模拟促炎、抗炎或稳态状态后，LFQ-MS和RNA测序揭示了小胶质细胞衍生的EV新的状态特异性蛋白质组学和转录组学特征。特别是，LPS处理对小胶质细胞衍生的EV的蛋白质组学和转录组学组成影响最为深远。此外，我们发现源自LPS激活的小胶质细胞的EV能够在静息的反应性小胶质细胞中诱导促炎转录组学变化，证实了小胶质细胞衍生的EV传递功能相关炎症信号的能力。这些全面的小胶质细胞-EV分子数据集是神经科学和神经胶质学界的重要资源，并为小胶质细胞衍生的EV在神经炎症中的作用提供了新的见解。
