Single-cell RNA sequencing of peripheral blood reveals that monocytes with high cathepsin S expression aggravate cerebral ischemia–reperfusion injury
Single-cell RNA sequencing of peripheral blood reveals that monocytes with high cathepsin S expression aggravate cerebral ischemia–reperfusion injury
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DOI：
10.1016/j.bbi.2022.11.001
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发表时间：
2022-11
期刊：
影响因子：
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通讯作者：
Lexing Xie;Shuang Zhang;Li Huang;Z. Peng;Hui Lu;Qian He;Ru Chen;Linlin Hu;Bingqiao Wang;Baoliang Sun;Qin Yang;Q. Xie
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文献类型：
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作者：
Lexing Xie;Shuang Zhang;Li Huang;Z. Peng;Hui Lu;Qian He;Ru Chen;Linlin Hu;Bingqiao Wang;Baoliang Sun;Qin Yang;Q. Xie
背景：中风是世界范围内发病率和死亡率的主要原因。脑缺血后，外周免疫细胞浸润大脑，引发炎症反应。然而，目前尚不清楚这些外周免疫细胞何时以及如何影响中枢炎症反应，以及针对这些过程的干预是否可以减轻缺血-再灌注（I/R）损伤。方法对I/R后不同时间小鼠外周血进行单细胞转录组测序和生物信息学分析，分析细胞亚群的关键分子。然后通过定量RT-PCR、western blot、ELISA、原位杂交等多种生物学实验，确定该分子的表达模式。接下来，使用敲除小鼠和相应的抑制剂来评估该分子的功能。结果单细胞转录组测序显示外周单核细胞亚群在I/R后显著增加。通过伪时间轨迹分析和基因功能分析，发现组织蛋白酶S （Ctss）是调控单核细胞活化的关键分子。接下来，证实Cathepsin S在单核细胞中表达，I/R后3天表达量最高。ctss敲除后，梗死面积（p < 0.05）、神经功能评分（p < 0.05）、细胞凋亡和血管渗漏率均显著降低。此外，CTSS通过与连接粘附分子（JAM）家族蛋白结合导致其降解，从而破坏血脑屏障（BBB）。结论肝组织蛋白酶S抑制可减轻脑I/R损伤；因此，组织蛋白酶S可作为脑卒中后药物干预的新靶点。
BackgroundStroke is a major cause of morbidity and mortality worldwide. After cerebral ischemia, peripheral immune cells infiltrate the brain and elicit an inflammatory response. However, it is not clear when and how these peripheral immune cells affect the central inflammatory response, and whether interventions that target these processes can alleviate ischemia–reperfusion (I/R) injury.MethodsSingle-cell transcriptomic sequencing and bioinformatics analysis were performed on peripheral blood of mice at different times after I/R to analyze the key molecule of cell subsets. Then, the expression pattern of this molecule was determined through various biological experiments, including quantitative RT-PCR, western blot, ELISA, and in situ hybridization. Next, the function of this molecule was assessed using knockout mice and the corresponding inhibitor.ResultsSingle-cell transcriptomic sequencing revealed that peripheral monocyte subpopulations increased significantly after I/R. Cathepsin S (Ctss)was identified as a key molecule regulating monocyte activation by pseudotime trajectory analysis and gene function analysis. Next, Cathepsin S was confirmed to be expressed in monocytes with the highest expression level 3 days after I/R. Infarct size (p < 0.05), neurological function scores (p < 0.05), and apoptosis and vascular leakage rates were significantly reduced afterCtssknockout. In addition, CTSS destroyed the blood–brain barrier (BBB) by binding to junctional adhesion molecule (JAM) family proteins to cause their degradation.ConclusionsCathepsin S inhibition attenuated cerebral I/R injury; therefore, cathepsin S can be used as a novel target for drug intervention after stroke.