Control of mitochondrial function in neurons by microRNAs during inflammation-induced neurodegeneration

多发性硬化症(MS)是最常见的中枢神经系统炎症性疾病。炎症性损伤会导致轴突和神经元的进行性退化，而轴突和神经元是发展成永久性神经功能障碍的关键。虽然很明显，炎症有助于神经元应激反应，导致线粒体功能障碍和能量耗竭，但哪些因素稳定这些反应网络仍未确定。MicroRNAs(MiRNAs)调控是调节应激反应中mRNAs翻译活性和稳定性的重要机制。因此，我们的目的是了解炎症过程中神经元基因表达的变化是如何由miRNAs调节的，以及这种调节是否影响对线粒体功能至关重要的蛋白质表达，从而加强或改善神经退行性变。因此，确定miRNAs在炎症诱导的神经元表达网络调控中的作用的第一个必要步骤是通过排除其他类型的细胞来全面表征匹配的mRNA和miRNA表达谱。因此，我们获得了健康小鼠不同神经元水平的mRNA和miRNA图谱，并将它们与实验性自身免疫性脑脊髓炎(EAE)小鼠进行比较，利用新的遗传方法建立的MS动物模型允许我们直接从炎症和健康的中枢神经系统匀浆中分离神经元mRNAs和miRNAs。对这些神经元特异的mRNA和miRNA分离株进行下一代测序，给了我们一个前所未有的量化中枢神经系统炎症期间神经元转录成分的方法。值得注意的是，在EAE模型期间，mRNA图谱显示神经元中线粒体转录本的表达明显不足。与此同时，这些转录本被预测为新发现的炎症诱导的神经元miRNAs的靶标。因此，我们现在建议进一步缩小我们的结果，通过紫外线交联和免疫沉淀的mRNA靶标及其来自神经元的miRNAs，以揭示直接相互作用的位置。为了从功能上表征miRNAs组成改变的影响，我们将使用体外和体内对调控最显著的miRNAs的研究，破译它们对线粒体代谢功能和神经元保护或退化的贡献。总之，这将提供一种新的方法来理解miRNA对控制线粒体功能的mRNA网络的控制，并有助于在中枢神经系统炎症过程中促进神经退化或保护。了解miRNAs在病理生理机制中的作用可能为疾病治疗提供一种新的治疗模式。

Multiple sclerosis (MS) is the most frequent inflammatory disease of the central nervous system (CNS). Inflammatory insults lead to progressive degeneration of axons and neurons that are key for the development of permanent neurological disability. While it is clear that inflammation contributes to neuronal stress responses resulting in mitochondrial dysfunction and energy depletion, it remains ill defined which factors stabilise these response networks. Regulation by microRNAs (miRNAs) is an important mechanism of regulating translational activity and stability of mRNAs during stress responses. Therefore, we aim at understanding how the changes of neuronal gene expression during inflammation are regulated by miRNAs and whether this regulation impacts on protein expression that are critical for mitochondrial function thereby reinforcing or ameliorating neurodegeneration. Accordingly, the first necessary step in the characterisation of the role of miRNAs in inflammation-induced regulation of expression networks in neurons is a comprehensive characterisation of matched mRNA and miRNA expression profiles by excluding other cell types. Hence, we acquired mRNA and miRNA profiles at the level of distinct neurons in healthy mice and compared them to mice with experimental autoimmune encephalomyelitis (EAE), the animal model of MS. By taking advantage of novel genetic methods allowed us to directly isolate neuronal mRNAs and miRNAs from inflamed and healthy CNS homogenates. Using next generation sequencing on these neuron-specific mRNA and miRNA isolates gave us a never before acquired quantification of transcript compositions in neurons during CNS inflammation. Notably, the mRNA profiles showed a significant underrepresentation of mitochondrial transcripts in neurons during the EAE model. At the same time these transcripts where predicted to be targets of newly identified inflammation-induced neuronal miRNAs. Therefore, we now propose to further narrow our results by UV crosslinking and immunoprecipitation of mRNA targets together with their miRNAs specifically from neurons to reveal direct interaction sites. In order to functionally characterise the impact of an altered composition of miRNAs, we will use in vitro and in vivo studies of the most significantly regulated miRNAs, deciphering their contribution to mitochondrial metabolic function and neuronal preservation or degeneration. Together, this will offer a new approach of understanding miRNA control of mRNA networks that govern mitochondrial function and contribute to neurodegeneration or -protection during CNS inflammation. Understanding the role of miRNAs in the pathophysiologic mechanisms may offer a new therapeutic modality for disease modification.