Gene expression analysis of mouse alveolar macrophages infected with Mycobacterium tuberculosis

The aim of this study was to measure the immune response of alveolar macrophages (AMs) 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. To determine the effect of the transcription factor Nrf2 on this response we also measured the transcriptional profiles of AMs from mice lacking Nrf2. Overall design: 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 various time points over the first 10 days following infection. Three populations were sorted for analysis by RNA-seq: Mtb-infected AMs (mEmerald+), bystander AMs (mEmerald-), and AMs from naïve mice. As a comparison, we repeated the high-dose infections using an mEmerald-expressing strain of Mtb lacking the RD1 virulence locus (deltaRD1-H37Rv) and using fluorescent 1uM carboxylate-coated latex beads.