Bladder resident macrophage subsets differently shape the response to urinary tract infection

Resident macrophages are highly abundant in the bladder, playing key roles in directing immunity to uropathogens. Yet, whether they are heterogeneous, where they come from, and precisely how they respond to infection remain largely unknown. We identified two macrophage subsets in mouse bladders with distinct localization, protein expression, and transcriptomes. Using a model of urinary tract infection, we validated our transcriptomics analyses finding that one macrophage subset phagocytosed more bacteria and polarized to a more anti-inflammatory profile, whereas the other subset died rapidly after infection. After resolution of infection, tissue-resident macrophage subsets were partially replaced by monocyte-derived cells with distinct transcriptional profiles. Elimination of these macrophages led to a type 1 biased immune response to challenge infection. Our study brings considerably more knowledge about the biology of bladder resident macrophages and their response to primary and recurrent infection, which may have broader implications for macrophage subsets in other mucosal tissues. Overall design: Samples were obtained from the whole bladders of naïve and 6 week post-infected (uropathogenic E coli) female C57BL/6J mice. Four separate fluorescence-activated cell sorts (FACS) were performed to generate biological replicates, each sort was a pool of 10 mouse bladders. Total RNA was extracted using the RNAeasy micro kit following the manufacturer's instructions. Directional libraries were prepared using the Smarter Stranded Total RNA-Seq kit Pico Input Mammalian following manufacturer's instructions. The quality of libraries was assessed with the DNA-1000 kit on a 2100 Bioanalyzer and quantification was performed with Quant-It assays on a Qubit 3.0 fluorometer. Clusters were generated for the resulting libraries with Illumina HiSeq SR Cluster Kit v4 reagents. Sequencing was performed with the Illumina HiSeq 2500 system and HiSeq SBS kit v4 reagents.