Inflammation is a ubiquitous component of lung disease involving accumulation of leukocytes in the airspaces. In order for inflammation to resolve, dead and dying leukocytes must be removed and production of inflammatory cytokines must be turned off. Macrophages (M?) are key orchestrators of these processes, however the triggers that reprogram inflammatory M? to perform these roles remain incompletely understood. Clearance of dead cells has been shown to provide an important reprogramming signal, but we have limited knowledge of the precise mechanisms by which dead cell ingestion facilitates redirection of M? function. Recent work has demonstrated a key role for cellular metabolism in determining M? function. Ingested target cells provide a clear source of varied macromolecules that must be digested by M?. However, little research has been done to assess intracellular metabolites produced by this process or to consider the subsequent immune consequence. We propose the first comprehensive study of M? metabolism following target cell ingestion and degradation, and hypothesize that changes in the levels of intracellular metabolites control ingestion-driven M? reprogramming. Our preliminary studies have identified a promising molecular target, polyamines, which are dramatically increased in M? following the ingestion of apoptotic target cells. Polyamines have a known anti- inflammatory function including suppression of numerous pro-inflammatory cytokines. We propose to test the hypothesis that upregulation of polyamine synthesis by M? following target cell ingestion is critical to suppress a key M? cytokine IL-1?, and important for the resolution of lung inflammation in vivo. During the K99 mentored phase, I will build upon my experience studying M? biology and target cell clearance by learning techniques related to cellular metabolism including unbiased metabolomics and the use of isotope metabolites to measure metabolic flux. During this phase, I will: 1) complete a comprehensive, integrated assessment of the metabolites produced following ingestion of target cells, focusing on polyamines, and assess whether these metabolites derive from digested target cell material, and 2) specifically assess the role of arginase-1 in regulating polyamine synthesis by inflammatory M?. Simultaneously, I will enrich my professional development by participating in journal clubs, seminars, national conferences, coursework, and having semi-annual evaluations by a trainee advisory committee. The R00 independent phase will allow me to establish my laboratory as I continue investigation into: 3) how M? polyamines affect cytokine production after target cell ingestion, and 4) the effects of M? polyamines on resolution of inflammation and lung repair. Collectively, this proposal will enhance my current expertise, address a critical unknown in the field of target cell clearance, and provide the necessary foundation to establish myself as an independent research scientist.
Macrophages are critical orchestrators of inflammation, however the mechanisms that reprogram macrophages to shut off production of inflammatory cytokines remain unclear. This proposal tests the hypothesis that ingestion of apoptotic cells and other target cells reprograms macrophages by altering macrophage metabolism and inducing the synthesis of anti-inflammatory polyamines that mediate suppression of pro-inflammatory cytokines, particularly IL-1?. Achieving the aims of this proposal will enhance our understanding of apoptotic cell clearance and macrophage reprogramming, and pave the way for new strategies to hasten the resolution of lung inflammation and restoration of lung function.
