Sentinel Functions of Tissue Resident Macrophages During Early Fungal Infections

Abstract

Invasive fungal infections result in high morbidity and mortality. Host organs targeted by fungal pathogens vary depending on the route of infection and fungal species encountered. Across different tissues in hosts, tissue-resident macrophages (TRMs) are one of the most abundant cells of the innate immune system and are in the front-line sensing invading fungi and elicit anti-fungal response. Neutrophil recruitment to the site of infection is one of the hallmarks of immune defense. In this thesis I found that TRMs are potent producers of neutrophil chemokine CXCL2. I used CXCL2 reporter animal model and uncovered two layers of TRMs heterogeneity: differential CXCL2 expression among TRMs of various tissues and transcriptional heterogeneity within alveolar macrophages (AMs). Different fungal infection models were utilized to examine TRMs in tissues with specific fungal tropism. Peritoneum macrophages, kupffer cells, kidney macrophages and alveolar macrophages were all capable of producing CXCL2 at the early hours post fungal infection. On the contrary, spleen macrophages and microglia were unable to produce CXCL2. Chromatin status of TRMs from naïve mice revealed that CXCL2-non-expressing TRMs also had less accessible Cxcl2 promoter than CXCL2-expressing TRMs, suggesting the regulation of Cxcl2 in TRMs at the chromatin level.Among the CXCL2-expressing TRMs, heterogeneous transcriptional states were identified within alveolar macrophages (AMs) populations. Although Cxcl2 promoter in AMs are constantly accessible, some AMs could not produce CXCL2 protein regardless the fungal inocula, spatial distribution or ontogenic origins. In addition to be a chemokine, CXCL2 was also a biological marker that differentiated AM cell states based on unbiased single cell RNA-seq. CXCL2+ and CXCL2- AMs expressed pro- and anti-inflammatory molecules, respectively, as well as distinct metabolic profiles and phagocytic capacity. CXCL2-divided AMs cell states also have distinct chromatin status enriched with differential transcription factor motifs. Long-term study of AM populations by ex vivo or in vivo transfer revealed CXCL2+ pro-inflammatory states were plastic and could be reset in AMs. However, one exception was the anti-inflammatory C1q molecule, expressed only by the pre-CXCL2- AMs. Promoters of C1q genes become accessible during early fungal infection only in CXCL2- AMs leading to their long-term gene expression.Fungal infections revealed quite heterogeneous CXCL2 responses both among TRMs and within AM populations. I used biologic reporter system and multiple system biology approaches to expand our knowledge of TRMs diversity and AM cell states. Particularly, my thesis demonstrated the co-existence of heterogenous AMs states with pro- and anti-inflammatory profiles, distinguished by CXCL2 during pulmonary Cryptococcus infection in vivo. The simultaneous appearance of functionally opposite AM states in the lung implies the balance of inflammation and tolerance even within a highly defined population of AMs.