Intrapulmonary itaconate as a host-protective metabolite during ER Stress and Klebsiella pneumoniae infection

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PROJECT ABSTRACT Gram negative bacteria are the most common pathogens implicated in nosocomial pneumonia in critically ill patients. Klebsiella pneumoniae (KP) in particular has grown in prominence worldwide with increasing prevalence of antibiotic resistance, hypervirulent strains, and invasive clinical syndromes. Mechanisms of host defense responsible for effective clearance of KP infection from the lung in susceptible hosts are still unclear. Cis-aconitate decarboxylase 1 (Acod1) is a mitochondrial enzyme robustly induced in murine and human alveolar macrophages that catalyzes the production of itaconate. Itaconate suppresses inflammatory responses through a proposed Nrf2-dependent mechanism. The goal of this proposal is to define the role of myeloid-derived itaconate in pulmonary host defense during KP infection and to determine how it confers protection from local tissue injury. In a murine model of pneumonia using KP as the pathogen, we have shown that Acod1 deficiency is associated with decreased survival and increased bacterial dissemination. We have also shown that itaconate depletion reduces expression of key proteins of the cellular integrated stress response (ISR) pathway during ER stress, suggesting that it may regulate this pathway. Our proposal will address the following specific aims: 1) To determine if myeloid cell-derived itaconate limits local KP infection and protects against extrapulmonary sepsis; and 2) To determine whether itaconate is host-protective through the ISR and decreases cellular injury during KP infection. We hypothesize that Acod1, and thus itaconate production, is protective against sepsis and associated tissue injury in this model and increases host resilience. We also anticipate that macrophage-derived itaconate increases protection from ER stress both in macrophages and lung epithelial cells in vitro and in vivo. These studies will utilize in vitro and translational tools including an in vivo intrapulmonary KP infection model using wild-type and Acod1-/- mice, bone marrow chimeras, RAW 264.7 macrophage cell lines (wild-type and Acod1-/-), human and mouse precision-cut lung tissue slices, and human alveolar macrophages to address our research questions. In addition, we will perform transcriptomic and metabolomic profiling across affected tissues during disseminated KP infection, immunophenotype myeloid subsets, and execute complementary cell biology studies to examine key targets of the integrated stress response pathway in the presence or absence of itaconate in myeloid and epithelial cells. This work will elucidate novel immune mechanisms that may be exploited to reduce mortality associated with this pathogen. The proposal incorporates a strong mentorship team led by Dr. Janet Lee as Director of the Acute Lung Injury Center of Excellence and a robust career development plan. Support of this proposal with a K08 Career Development Award will establish a foundation for success and independence for the candidate as a physician-scientist in pulmonary and critical care medicine.