PROJECT SUMMARY/ABSTRACT This proposal describes a five-year career development plan designed to provide Scott Denstaedt, MD with training in new techniques through mentored research and didactics to facilitate a career as an independent physician-scientist studying mechanisms of long-term pulmonary complications in patients surviving sepsis. Research plan: Despite modern supportive care, recovery from sepsis is associated with increased risk for hospital readmission and death. Nearly one in ten patients with sepsis will be re-hospitalized within 90-days with pulmonary conditions including pneumonia, aspiration pneumonitis, and/or respiratory failure. We do not understand the pathophysiological mechanisms underlying these complications and have no targeted therapies to prevent them. Dr. Denstaedt has developed a mouse model in which survivors of sepsis have enhanced (primed) immune responses to lung injury. Specifically, twenty-one days after sepsis there is enhanced lung permeability and neutrophil recruitment following intranasal lipopolysaccharide. Dr. Denstaedt’s prior work demonstrated that chronic brain dysfunction after sepsis is associated with innate immune priming and persistent elevations of the danger signal protein S100A8/A9. The central hypothesis of this proposal is that persistent S100A8/A9 signaling through TLR4 primes inflammatory monocytes predisposing to enhanced lung injury. Aim 1 investigates how changes in inflammatory monocytes persisting in the lung after sepsis predispose to enhanced alveolar damage after sterile or infectious lung injury. Immunostaining and fluorescent microscopy with stereology, gene expression, and epigenetics assays will be performed to evaluate pathways of inflammation and cell death. A causal link between primed monocytes and enhanced lung injury will be examined in vivo by monocyte depletion and in vitro by monocyte/epithelial cell co-culture. In Aim 2, S100A8/A9 inhibition and cell-specific TLR4 deletion will determine the effect of S100A8/A9-TLR4 signaling on enhanced lung injury and priming after sepsis. Inflammatory phenotypes predisposing to injury will be assessed by microscopy, gene expression, and epigenetics assays. The role of plasma S100A8/A9 from post- sepsis patients in priming human monocytes will be examined by gene expression and epigenetics assays. Applicant: Scott Denstaedt, MD has completed training in Pulmonary and Critical Care Medicine. He has acquired extensive research experience in the last four years studying chronic brain dysfunction in a model of long-term sepsis survival. The proposed training will provide new knowledge in lung injury and epigenetics to examine the underlying mechanisms of post-sepsis pulmonary complications. Technical training includes advanced microscopy and stereology, primary cell co-culture, state-of-the-art epigenetics assays, cell-specific inducible knockout mouse modeling, and in vitro human translational studies. The training plan, mentorship ...