RNA-seq transcriptional analysis of mouse lung tissue with and without Coccidioides immitis lung infection. RNA-seq transcriptional analysis of mouse lung tissue with and without Coccidioides immitis lung infection

Our previous data suggested that CXC ELR+ chemokines might be involved in neutrophil recruitment to the lungs during Coccidioides infection. To test that hypothesis, we infected IL-8R2 (Cxcr2) KO mice on a BALB/c background. IL-8R2 KO mice had fewer neutrophils in infected lungs than BALB/c controls, but unexpectedly the IL-8R2 KO mice also had 10-fold fewer organisms in their lungs than did control mice. To better understand the differences in IL-8R2 KO mouse lungs during Coccidioides infection we performed RNA-seq on lung tissue from uninfected and Coccidioides immitis infected mouse lungs in control and IL-8R2 (Cxcr2) KO mice. IL-8R2 KO mouse lungs had higher expression of genes associated with lymphocyte activation, including Th1 and Th17-related genes Ifnγ and Il17a and transcription factors Stat1 and Rorc. Bronchial alveolar lavage (BAL) fluid from infected IL-8R2 KO mice similarly contained more IL-17A and IFNγ. Overall design: RNA-seq of lung tissue from BALB/c DBA/2 Nramp1 congenic (Potter M et al. (1983) PMID: 6343242) (wild-type or WT) and Balb/c CXCR2tm (IL-8R2) knockout (IL-8R2 KO) mice with or without Coccidioides immitis infection on day 14 after infection.

本研究团队前期实验数据表明，在球孢子菌（Coccidioides）感染过程中，CXC ELR+趋化因子（CXC ELR+ chemokines）可能参与了中性粒细胞向肺部的招募过程。为验证该假说，我们对BALB/c背景下的IL-8R2（Cxcr2）基因敲除（KO）小鼠实施了粗球孢子菌（Coccidioides immitis）感染。与BALB/c野生型对照组小鼠相比，IL-8R2敲除小鼠的感染肺部中中性粒细胞数量更少，但令人意外的是，此类敲除小鼠肺部的病原体载量仅为对照组的1/10。为深入解析球孢子菌感染过程中IL-8R2敲除小鼠肺部的差异变化，我们对对照组及IL-8R2（Cxcr2）敲除小鼠的未感染与感染粗球孢子菌的肺部组织开展了RNA测序（RNA-seq）。结果显示，IL-8R2敲除小鼠肺部中与淋巴细胞活化相关的基因表达水平显著升高，其中包括与Th1、Th17亚群相关的Ifnγ、Il17a基因，以及转录因子Stat1与Rorc。感染小鼠的支气管肺泡灌洗液（BAL）检测结果同样证实，IL-8R2敲除小鼠的灌洗液中IL-17A与IFNγ含量更高。整体实验设计：采集BALB/c背景的DBA/2 Nramp1同源导入近交系（Potter M等，1983，PMID: 6343242）野生型（WT）小鼠，以及Balb/c背景的CXCR2tm（IL-8R2）基因敲除（IL-8R2 KO）小鼠的肺部组织，两组小鼠均设置未感染与感染粗球孢子菌组别，于感染后第14天取样进行RNA-seq测序。