Population RNA-seq of in vitro polarized CD4 T cells

To understand the molecular programs driving Th17 heterogeneity and CD4 T cell fate, we profiled in vitro polarized non-pathogenic Th17, pathogenic Th17, Th1, and Treg cells using RNA-seq. We profiled in vitro polarized and in vivo mouse primary CD4 T cells in the following independent experiments: 1) non-pathogenic and pathogenic Th17 cells harvested polarized from naïve CD4 T cells isolated from Il17-eGFP reporter mice, sorted for GFP+ and all cells (Il17eGFP); 2) non-pathogenic Th17, pathogenic Th17, Th1, and Treg cells harvested after 72h polarization from naive mouse CD4 T cells (CD4_diff); 3) non-pathogenic Th17, pathogenic Th17, and Th1 harvested at time points ranging from 0 to 48 hr after polarization from naive mouse CD4 T cells (Time_course); 4) Il17-expressing Th17 cells from the draining lymph node and central nervous system of mice at the peak of experimental autoimmune encephalomyelitis response (EAE)

为解析驱动Th17细胞异质性与CD4 T细胞命运的分子程序，我们利用RNA测序（RNA-seq）技术，对体外极化的非致病性Th17细胞、致病性Th17细胞、Th1细胞及调节性T细胞（Treg）进行了转录组分析。此外，我们还通过四项独立实验，对体外极化及体内来源的小鼠原代CD4 T细胞开展了转录组分析，具体实验如下：1. 实验1：从IL17-eGFP报告基因小鼠中分离初始CD4 T细胞，经体外极化后收集非致病性与致病性Th17细胞，并按GFP阳性细胞及总细胞进行分选（对应样本集Il17eGFP）；2. 实验2：从小鼠初始CD4 T细胞体外极化72小时后，收集非致病性Th17细胞、致病性Th17细胞、Th1细胞及调节性T细胞（Treg）（对应样本集CD4_diff）；3. 实验3：从小鼠初始CD4 T细胞体外极化后的0至48小时多个时间点取样，收集非致病性Th17细胞、致病性Th17细胞及Th1细胞（对应样本集Time_course）；4. 实验4：在实验性自身免疫性脑脊髓炎（EAE，experimental autoimmune encephalomyelitis）反应峰值期，从小鼠引流淋巴结及中枢神经系统中分离表达IL17的Th17细胞。