Abstract：
OBJECTIVE: Non-tuberculous mycobacterial pulmonary disease (NTM-PD) is caused by an imbalance between pathogens and impaired host immune responses. Mycobacterium avium complex (MAC) and Mycobacterium abscessus (MAB) are the two major pathogens that cause NTM-PD. In this study, we sought to dissect the transcriptomes of peripheral blood immune cells at the single-cell resolution in NTM-PD patients and explore potential clinical markers for NTM-PD diagnosis and treatment.
METHODS: Peripheral blood samples were collected from six NTM-PD patients, including three MAB-PD patients, three MAC-PD patients, and two healthy controls. We employed single-cell RNA sequencing (scRNA-seq) to define the transcriptomic landscape at a single-cell resolution. A comprehensive scRNA-seq analysis was performed, and flow cytometry was conducted to validate the results of scRNA-seq.
RESULTS: A total of 27,898 cells were analyzed. Nine T-cells, six mononuclear phagocytes (MPs), and four neutrophil subclusters were defined. During NTM infection, naïve T-cells were reduced, and effector T-cells increased. High cytotoxic activities were shown in T-cells of NTM-PD patients. The proportion of inflammatory and activated MPs subclusters was enriched in NTM-PD patients. Among neutrophil subclusters, an IFIT1+ neutrophil subcluster was expanded in NTM-PD compared to healthy controls. This suggests that IFIT1+ neutrophil subcluster might play an important role in host defense against NTM. Functional enrichment analysis of this subcluster suggested that it is related to interferon response. Cell-cell interaction analysis revealed enhanced CXCL8-CXCR1/2 interactions between the IFIT1+ neutrophil subcluster and NK cells, NKT cells, classical mononuclear phagocytes subcluster 1 (classical Mo1), classical mononuclear phagocytes subcluster 2 (classical Mo2) in NTM-PD patients compared to healthy controls.
CONCLUSIONS: Our data revealed disease-specific immune cell subclusters and provided potential new targets of NTM-PD. Specific expansion of IFIT1+ neutrophil subclusters and the CXCL8-CXCR1/2 axis may be involved in the pathogenesis of NTM-PD. These insights may have implications for the diagnosis and treatment of NTM-PD.
METHODS: Peripheral blood samples were collected from six NTM-PD patients, including three MAB-PD patients, three MAC-PD patients, and two healthy controls. We employed single-cell RNA sequencing (scRNA-seq) to define the transcriptomic landscape at a single-cell resolution. A comprehensive scRNA-seq analysis was performed, and flow cytometry was conducted to validate the results of scRNA-seq.
RESULTS: A total of 27,898 cells were analyzed. Nine T-cells, six mononuclear phagocytes (MPs), and four neutrophil subclusters were defined. During NTM infection, naïve T-cells were reduced, and effector T-cells increased. High cytotoxic activities were shown in T-cells of NTM-PD patients. The proportion of inflammatory and activated MPs subclusters was enriched in NTM-PD patients. Among neutrophil subclusters, an IFIT1+ neutrophil subcluster was expanded in NTM-PD compared to healthy controls. This suggests that IFIT1+ neutrophil subcluster might play an important role in host defense against NTM. Functional enrichment analysis of this subcluster suggested that it is related to interferon response. Cell-cell interaction analysis revealed enhanced CXCL8-CXCR1/2 interactions between the IFIT1+ neutrophil subcluster and NK cells, NKT cells, classical mononuclear phagocytes subcluster 1 (classical Mo1), classical mononuclear phagocytes subcluster 2 (classical Mo2) in NTM-PD patients compared to healthy controls.
CONCLUSIONS: Our data revealed disease-specific immune cell subclusters and provided potential new targets of NTM-PD. Specific expansion of IFIT1+ neutrophil subclusters and the CXCL8-CXCR1/2 axis may be involved in the pathogenesis of NTM-PD. These insights may have implications for the diagnosis and treatment of NTM-PD.
摘要：
目的：非结核分枝杆菌肺病（NTM-PD）是由病原体之间的失衡和宿主免疫反应受损引起的。鸟分枝杆菌复合体(MAC)和脓肿分枝杆菌(MAB)是引起NTM-PD的两种主要病原体。在这项研究中,我们试图在NTM-PD患者中以单细胞分辨率解剖外周血免疫细胞的转录组,并探索NTM-PD诊断和治疗的潜在临床标志物.
方法：收集6名NTM-PD患者的外周血样本，包括三名MAB-PD患者,三名MAC-PD患者,和两个健康的对照。我们采用单细胞RNA测序（scRNA-seq）以单细胞分辨率定义转录组景观。进行了全面的scRNA-seq分析，和流式细胞术验证scRNA-seq的结果。
结果:总共分析了27,898个细胞。九个T细胞,六个单核吞噬细胞（MPs），并定义了四个中性粒细胞亚簇。在NTM感染期间，原始T细胞减少，效应T细胞增加。在NTM-PD患者的T细胞中显示出高细胞毒性活性。NTM-PD患者中炎症和活化的MPs亚簇的比例富集。在中性粒细胞亚簇中，与健康对照组相比,NTM-PD中的IFIT1+中性粒细胞亚簇扩大.这表明IFIT1中性粒细胞亚簇可能在宿主对NTM的防御中起重要作用。该亚簇的功能富集分析表明它与干扰素反应有关。细胞相互作用分析显示，IFIT1+中性粒细胞亚簇和NK细胞之间的CXCL8-CXCR1/2相互作用增强，NKT细胞,经典单核吞噬细胞亚簇1（经典Mo1），与健康对照相比，NTM-PD患者的经典单核吞噬细胞亚簇2（经典Mo2）。
结论：我们的数据揭示了疾病特异性免疫细胞亚簇，并提供了NTM-PD的潜在新靶标。IFIT1+中性粒细胞亚簇和CXCL8-CXCR1/2轴的特异性扩增可能与NTM-PD的发病机制有关。这些见解可能对NTM-PD的诊断和治疗有影响。
方法：收集6名NTM-PD患者的外周血样本，包括三名MAB-PD患者,三名MAC-PD患者,和两个健康的对照。我们采用单细胞RNA测序（scRNA-seq）以单细胞分辨率定义转录组景观。进行了全面的scRNA-seq分析，和流式细胞术验证scRNA-seq的结果。
结果:总共分析了27,898个细胞。九个T细胞,六个单核吞噬细胞（MPs），并定义了四个中性粒细胞亚簇。在NTM感染期间，原始T细胞减少，效应T细胞增加。在NTM-PD患者的T细胞中显示出高细胞毒性活性。NTM-PD患者中炎症和活化的MPs亚簇的比例富集。在中性粒细胞亚簇中，与健康对照组相比,NTM-PD中的IFIT1+中性粒细胞亚簇扩大.这表明IFIT1中性粒细胞亚簇可能在宿主对NTM的防御中起重要作用。该亚簇的功能富集分析表明它与干扰素反应有关。细胞相互作用分析显示，IFIT1+中性粒细胞亚簇和NK细胞之间的CXCL8-CXCR1/2相互作用增强，NKT细胞,经典单核吞噬细胞亚簇1（经典Mo1），与健康对照相比，NTM-PD患者的经典单核吞噬细胞亚簇2（经典Mo2）。
结论：我们的数据揭示了疾病特异性免疫细胞亚簇，并提供了NTM-PD的潜在新靶标。IFIT1+中性粒细胞亚簇和CXCL8-CXCR1/2轴的特异性扩增可能与NTM-PD的发病机制有关。这些见解可能对NTM-PD的诊断和治疗有影响。
