Project Summary/Abstract Sepsis is a complex, heterogeneous, global inflammatory response to an infection that precipitates critical hypotension, is beset with high mortality, and constitutes a clinical emergency. Large blood vessels (macrovessels) become atonic and fluid-permeable, while small vessels (microvascular beds) do not perfuse, causing shock and hypoxic organ damage, a condition termed vasoplegia. Dr. Wise endeavors to become an independently funded, collaborative surgeon-scientist by characterizing physiologic injury and defining cellular processes that occur during the vasoplegia precipitated by sepsis. While elimination of the infectious pathogen is most critical to survival, mitigating vasoplegic physiology prevents morbidity to vital end organs such as the lung. Examining the problem of vasoplegia from the lens of the vascular system necessarily requires career development in three key areas, identified to optimally address the research question: cell biology and multi- omics, use of intravital surveillance techniques (microscopy), and facility with large animal models to best recapitulate human physiology in sepsis. This project is well-positioned to succeed due to a complementary mentorship and advisory team led by Dr. Greg Beilman (University of Minnesota), strong institutional support, and a focused career development plan inclusive of germane coursework, didactics, seminars and meetings. Vasoplegia will be studied using a newly-piloted porcine “fecal clot” (with ischemia-reperfusion injury) model of surgically-induced sepsis. In the first Aim, fecal clot sepsis or “sham” sepsis will be induced, followed by surgical harvest of mesenteric and saphenous arteries and veins after 24 hours. Cut rings from these vessels will be suspended in a muscle bath to determine relative severity of injury of smooth muscle cells, endothelial cells and extracellular matrix. Permeability will be assessed via immunohistochemical staining for glycocalyx. Regulation of the endoplasmic reticulum stress response and P38 MAP Kinase activation by putative endogenous inhibitor Niban will be studied, to assess the role of “off-signaling” in propagating septic inflammation. In the second Aim, lung microvasculature will be imaged using intravital orthogonal polarized spectral video-microscopy, to quantify the loss of perfused vessels and blood flow induced by fecal clot sepsis. After synthesis of macrovascular and microvascular injury patterns of vasoplegia from the first two Aims, a pilot mitigation strategy will be tested in the third Aim. Intravenous “NiPp,” a rationally designed cell-permeant phospho-mimetic of Niban developed in the laboratory of Colleen Brophy (co-mentor), will be administered to septic pigs, to decrease harmful “feed-forward” activation of injury-induced pro-inflammatory pathways leading to vasoplegia. The primary outcome will be improvement in a validated clinical porcine sepsis score. All pigs will have systemic vascular tissue ext...