DataSheet_1_Application of spatial transcriptomics analysis using the Visium system for the mouse nasal cavity after intranasal vaccination.pdf

Intranasal vaccines that elicit mucosal immunity are deemed effective against respiratory tract infections such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but their ability to induce humoral immunity characterized by immunoglobulin A (IgA) and IgG production is low. It has been reported that vaccination with a mixture of a viscous base carboxyvinyl polymer (CVP) and viral antigens induced robust systemic and mucosal immune responses. In this study, we analyzed the behavior of immunocompetent cells in the nasal cavity over time by spatial transcriptome profiling induced immediately after antigen vaccination using CVP. We established a method for performing spatial transcriptomics using the Visium system in the mouse nasal cavity and analyzed gene expression profiles within the nasal cavity after intranasal vaccination. Glycoprotein 2 (Gp2)-, SRY-box transcription factor 8 (Sox8)-, or Spi-B transcription factor (Spib)-expressing cells were increased in the nasal passage (NP) region at 3–6 hr after SARS-CoV-2 spike protein and CVP (S-CVP) vaccination. The results suggested that microfold (M) cells are activated within a short period of time (3–6 hr). Subsequent cluster analysis of cells in the nasal cavity showed an increase in Cluster 9 at 3–6 hr after intranasal vaccination with the S-CVP. We found that Il6 in Cluster 9 had the highest log2 fold values within the NP at 3–6 hr. A search for gene expression patterns similar to that of Il6 revealed that the log2 fold values of Edn2, Ccl20, and Hk2 also increased in the nasal cavity after 3–6 hr. The results showed that the early response of immune cells occurred immediately after intranasal vaccination. In this study, we identified changes in gene expression that contribute to the activation of M cells and immunocompetent cells after intranasal vaccination of mice with antigen-CVP using a time-series analysis of spatial transcriptomics data. The results facilitated the identification of the cell types that are activated during the initial induction of nasal mucosal immunity.

可诱导黏膜免疫的鼻内疫苗被认为对重症急性呼吸综合征冠状病毒2（severe acute respiratory syndrome coronavirus 2，SARS-CoV-2）等呼吸道感染具有防控效果，但其诱导以免疫球蛋白A（immunoglobulin A，IgA）和免疫球蛋白G（immunoglobulin G，IgG）产生为特征的体液免疫的能力较弱。已有研究表明，将黏性基质羧基乙烯聚合物（carboxyvinyl polymer，CVP）与病毒抗原混合后进行免疫接种，可诱导强烈的系统性免疫与黏膜免疫应答。本研究借助空间转录组分析技术，对使用羧基乙烯聚合物进行抗原免疫接种后即刻的小鼠鼻腔内免疫活性细胞的动态变化进行了分析。我们建立了一套在小鼠鼻腔中使用Visium平台开展空间转录组学分析的方法，并对鼻内免疫接种后的鼻腔基因表达谱进行了分析。在接种SARS-CoV-2刺突蛋白联合羧基乙烯聚合物（S-CVP）后3~6小时，鼻道（nasal passage，NP）区域内表达糖蛋白2（Glycoprotein 2，Gp2）、SRY框转录因子8（SRY-box transcription factor 8，Sox8）或Spi-B转录因子（Spi-B transcription factor，Spib）的细胞数量显著增加。上述结果提示，微褶皱细胞（microfold cell，M细胞）可在接种后3~6小时的短时间内被激活。后续对鼻腔细胞的聚类分析显示，在S-CVP鼻内免疫接种后3~6小时，第9聚类群（Cluster 9）的细胞占比显著升高。我们发现，在接种后3~6小时的鼻道区域内，第9聚类群中的白细胞介素6（Il6）的log2倍变化值最高。通过搜索与Il6表达模式相似的基因，我们发现接种后3~6小时，鼻腔内皮素2（Edn2）、趋化因子配体20（Ccl20）与己糖激酶2（Hk2）的log2倍变化值同样显著升高。上述结果表明，免疫细胞的早期应答在鼻内免疫接种后即刻发生。本研究通过对空间转录组数据进行时序分析，明确了小鼠鼻内接种抗原-羧基乙烯聚合物制剂后，参与微褶皱细胞与免疫活性细胞激活的基因表达变化。本研究结果有助于识别鼻黏膜免疫初始诱导阶段被激活的细胞类群。