Roles of the Src-Family Kinases LynA and LynB in Macrophage Inflammatory Signaling

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ABSTRACT The autoimmune disease juvenile idiopathic arthritis (JIA), which causes joint, eye, and bone damage and chronic pain, is the most common chronic childhood disease in the United States. Joint fluid from JIA patients contains large numbers of macrophage immune cells, and macrophage secretion of inflammatory factors drives the disease. Current therapies for JIA block this inflammatory signaling but do not discriminate between disease- associated inflammatory signaling and the normal immune response to infection. Therefore, suppression of an- timicrobial immunity is a severe side effect of these existing therapies. We have discovered a signaling mecha- nism through the protein LynA that is required for hypersensitive inflammatory signaling by macrophages, but is not strictly required for pathogen recognition. We have now also generated the first LynA-/- mice and propose to test whether the LynA regulatory pathway has promise as a novel therapeutic target, to specifically suppress inflammatory macrophage signaling without suppressing the immune system generally. Our long-term goal is to develop new therapeutic approaches targeted specifically to pathological macrophages. To support the diversity mission in our lab and increase the depth of our funded investigation, we aim to test the role of LynA in isolated macrophages and in additional models of mouse inflammatory arthritis. More specifically, we propose to: A) Examine the effect of LynA and LynB knockout in the K/BxN genetic model and the SKG model of murine in- flammatory arthritis: We will breed these strains with our existing knockout moels and assess disease progres- sion. We will also obtain tissues to be used in subsequent Aims. We predict that LynA-/- mice will have less severe disease than LynB-/- mice. B) Quantify autoantibody and cytokine production in the above models: We will perform additional profiling of IgG, IgM, and inflammatory cytokine levels in sera from the above mice with autoimmune disease, which will yield clues to unique pathological profiles and cell drivers of autoimmune dis- ease. Based on our recently published work, we predict that LynA-/- mice will have a more macrophage/mono- cyte-driven profile, whereas LynB-/- mice will be more dendritic-cell driven. We predict that female mice will have more autoantibody production than male mice. C) Test polarization of myeloid cells and activation of B cells taken from arthritic mice above: We hypothesize that, as in mice, inflammation in JIA-affected joints induces LynA upregulation in human macrophages. To complement our cytokine profiling, we will use flow cytometry and qRT-PCR to characterize myeloid-cell polarization and B-cell activation This will help connect cytokine profiles to cellular drivers of disease. It may also reveal the most relevant pathways driving the autoimmune pathology. This project will benefit from our expertise in macrophage signaling and close collaborations within the Center for Immunology at the University of Minnesota. These studies will establish the feasibility of targeting LynA to develop safer, targeted treatment options for JIA and other macrophage-driven inflammatory diseases.