Gene expression analysis of mouse alveolar macrophages infected with Mycobacterium tuberculosis isolated from BCG-vaccinated mice [balRNAseq]. Gene expression analysis of mouse alveolar macrophages infected with Mycobacterium tuberculosis isolated from BCG-vaccinated mice [balRNAseq]

The aim of this study was to measure the immune response of alveolar macrophages (AMs) from BCG-vaccinated mice to intracellular Mtb infection in vivo. We characterized the transcriptional profile of murine Mtb-infected AMs after aerosol infection by sorting cells from bronchoalveolar lavage fluid and performing low-input RNA-sequencing. Overall design: Mice were vaccinated sub-cutaneously with BCG-Pasteur and allowed to rest for 4 weeks. To characterize the early macrophage transcriptional response to Mtb infection, we infected mice with 2000-4000 CFU of mEmerald-H37Rv, and isolated AMs (CD45+, Zombie Violet-, CD3-/CD19-, Siglec-F+, CD11bmid, and CD64+) from BAL by fluorescence activated cell sorting at 24 hours following infection. Three populations were sorted for analysis by RNA-seq: Mtb-infected AMs (mEmerald+), bystander AMs (mEmerald-), and AMs from naïve mice.

本研究旨在测定卡介苗（Bacillus Calmette-Guérin, BCG）免疫小鼠肺泡巨噬细胞（alveolar macrophages, AMs）在体内感染胞内结核分枝杆菌（Mycobacterium tuberculosis, Mtb）后的免疫应答。我们通过分选支气管肺泡灌洗液（bronchoalveolar lavage fluid, BALF）中的细胞并进行低起始量RNA测序（low-input RNA-sequencing），表征了气溶胶感染后结核分枝杆菌感染小鼠肺泡巨噬细胞的转录组特征。总体实验设计：小鼠经皮下接种卡介苗巴斯德株（BCG-Pasteur），并静置饲养4周。为表征巨噬细胞对结核分枝杆菌感染的早期转录应答，我们以2000~4000菌落形成单位（colony-forming unit, CFU）的mEmerald标记H37Rv毒株（mEmerald-H37Rv）感染小鼠，并于感染后24小时通过荧光激活细胞分选（fluorescence activated cell sorting, FACS）从支气管肺泡灌洗液中分选肺泡巨噬细胞（表型为CD45+、Zombie Violet-、CD3-/CD19-、Siglec-F+、CD11bmid及CD64+）。我们共分选三类群体用于RNA测序分析：结核分枝杆菌感染的肺泡巨噬细胞（mEmerald+）、旁观者肺泡巨噬细胞（mEmerald-），以及未感染小鼠的肺泡巨噬细胞。