Department of Neurosurgery (Z.C., S.S.H., T.C., A.G.F., M.Y.C., G.K.S.), Stanford University School of Medicine, CA.
Stanford Stroke Center (Z.C., S.S.H., T.C., A.G.F., M.Y.C., G.K.S.), Stanford University School of Medicine, CA.
Stroke. 2021 Jan;52(2):687-698. doi: 10.1161/STROKEAHA.120.032402. Epub 2021 Jan 8.
Stroke disrupts neuronal functions in both local and remotely connected regions, leading to network-wide deficits that can hinder recovery. The thalamus is particularly affected, with progressive development of neurodegeneration accompanied by inflammatory responses. However, the complexity of the involved inflammatory responses is poorly understood. Herein we investigated the spatiotemporal changes in the secondary degenerative thalamus after cortical stroke, using targeted transcriptome approach in conjunction with histology and flow cytometry.
Cortical ischemic stroke was generated by permanent occlusion of the left middle cerebral artery in male C57BL6J mice. Neurodegeneration, neuroinflammatory responses, and microglial activation were examined in naive and stroke mice at from poststroke days (PD) 1 to 84, in both ipsilesional somatosensory cortex and ipsilesional thalamus. NanoString neuropathology panel (780 genes) was used to examine transcriptome changes at PD7 and PD28. Fluorescence activated cell sorting was used to collect CD11c microglia from ipsilesional thalamus, and gene expressions were validated by quantitative real-time polymerase chain reaction.
Neurodegeneration in the thalamus was detected at PD7 and progressively worsened by PD28. This was accompanied by rapid microglial activation detected as early as PD1, which preceded the neurodegenerative changes. Transcriptome analysis showed higher number of differentially expressed genes in ipsilesional thalamus at PD28. Notably, neuroinflammation was the top activated pathway, and microglia was the most enriched cell type. (CD11c) was the most significantly increased gene, and its expression was highly detected in microglia. Flow-sorted CD11c microglia from degenerative thalamus indicated molecular signatures similar to neurodegenerative disease-associated microglia; these included downregulated Tmem119 and CX3CR1 and upregulated ApoE, Axl, LpL, CSF1, and Cst7.
Our findings demonstrate the dynamic changes of microglia after stroke and highlight the importance of investigating stroke network-wide deficits. Importantly, we report the existence of a unique subtype of microglia (CD11c) with neurodegenerative disease-associated microglia features in the degenerative thalamus after stroke.
中风会扰乱局部和远程连接区域的神经元功能，导致广泛的网络缺陷，从而阻碍恢复。丘脑尤其受到影响，伴随着神经退行性变的进行性发展和炎症反应。然而，涉及的炎症反应的复杂性还了解甚少。在此，我们通过靶向转录组方法结合组织学和流式细胞术，研究了皮质性中风后次级丘脑的时空变化。
通过永久性阻塞雄性 C57BL6J 小鼠左侧大脑中动脉，产生皮质性脑梗死。在中风后第 1 天至第 84 天，在同侧体感皮层和同侧丘脑，检查未受影响的和中风的小鼠中的神经退行性变、神经炎症反应和小胶质细胞激活。使用 NanoString 神经病理学面板（780 个基因）检查 PD7 和 PD28 的转录组变化。使用荧光激活细胞分选从同侧丘脑收集 CD11c 小胶质细胞，并通过定量实时聚合酶链反应验证基因表达。
在 PD7 时检测到丘脑的神经退行性变，并在 PD28 时逐渐加重。这伴随着早在 PD1 时就检测到的快速小胶质细胞激活，这先于神经退行性变。转录组分析显示，PD28 时同侧丘脑的差异表达基因数量更多。值得注意的是，神经炎症是激活的首要途径，小胶质细胞是最丰富的细胞类型。（CD11c）是增加最显著的基因，其表达在小胶质细胞中高度检测到。从变性丘脑分离的 CD11c 小胶质细胞的流式分选表明其具有与神经退行性疾病相关的小胶质细胞相似的分子特征；这些特征包括 Tmem119 和 CX3CR1 的下调和 ApoE、Axl、LpL、CSF1 和 Cst7 的上调。
我们的研究结果表明，中风后小胶质细胞的动态变化，并强调了研究中风的广泛网络缺陷的重要性。重要的是，我们报告了在中风后变性丘脑存在具有神经退行性疾病相关小胶质细胞特征的独特小胶质细胞（CD11c）亚群。
