Dissecting how rapid microbial pathogen containment occurs in vivo

Abstract Inflammatory Ly6C+ monocytes are a subset of highly plastic myeloid cells that are rapidly mobilized during multiple types of microbial pathogen infections (bacteria, virus, fungi, parasites) and differentiate into robust microbicidal effector cells that can secrete TNF and produce nitric oxide and reactive oxygen species directly contributing to pathogen killing and elimination. We and others have also reported that Ly6C+ monocytes accumulate and form clusters at foci of microbial pathogen infections. In recent work using mice infected with the intracellular bacterium Listeria monocytogenes and intravital multi-photon imaging, we also revealed that Ly6C+ monocytes arrest at infection foci, exhibit a highly activated phenotype and deliver local effector functions. While it is well established that these cells egress from the bone marrow to the blood via CCR2 in response to the rapid release of CCL2 that occurs upon microbial product sensing, how they reach infected tissues, home to foci of infections and form stable clusters is still largely unknown. Many chemotactic and adhesion mechanisms have been ruled out to regulate these processes, suggesting either other mechanisms or redundancy. Here we formally test the hypothesis that Ly6C+ monocytes need to home to foci of infection and form stable clusters to deliver local and powerful effector functions to help clear pathogens. This exploratory proposal will define the chemotactic cues and receptors involved, the initiating and orchestrating target cells that orchestrate Ly6C+ monocytes homing to infection foci and the molecules involved in monocyte arrest and formation of stable clusters. We will use combinations of genetic gain and loss of function experiments in vivo, bacterial, parasitic and viral infection models, cutting-edge intravital microscopy imaging and in vitro functional chemotactic assays.