| Summary: | The implementation of infection models that approximate human disease is essential to understand infections and for testing new therapies before they enter into clinical stages. Rodents are used in most preclinical studies, although the differences between mice and humans have fueled the conclusion that murine studies are unreliable predictors of human outcomes. In this study, we have developed a whole-lung porcine model of infection using the ex vivo lung perfusion (EVLP) system established to recondition human lungs for transplant. As a proof of principle, we provide evidence demonstrating that infection of the porcine EVLP with the human pathogen Klebsiella pneumoniae recapitulates the known features of Klebsiella-triggered pneumonia. Moreover, our data revealed that the porcine EVLP model is useful to reveal features of the virulence of K. pneumoniae, including the manipulation of immune cells. Together, the findings of this study support the utility of the EVLP model using pig lungs as a surrogate host for assessing respiratory infections.The use of animal infection models is essential to understand microbial pathogenesis and to develop and test treatments. Insects and two-dimensional (2D) and 3D tissue models are increasingly being used as surrogates for mammalian models. However, there are concerns about whether these models recapitulate the complexity of host-pathogen interactions. In this study, we developed the ex vivo lung perfusion (EVLP) model of infection using porcine lungs to investigate Klebsiella pneumoniae-triggered pneumonia as a model of respiratory infections. The porcine EVLP model recapitulates features of K. pneumoniae-induced pneumonia lung injury. This model is also useful to assess the pathogenic potential of K. pneumoniae, as we observed that the attenuated Klebsiella capsule mutant strain caused less pathological tissue damage with a concomitant decrease in the bacterial burden compared to that in lungs infected with the wild type. The porcine EVLP model allows assessment of inflammatory responses following infection; similar to the case with the mouse pneumonia model, we observed an increase of il-10 in the lungs infected with the wild type and an increase of ifn-γ in lungs infected with the capsule mutant. This model also allows monitoring of phenotypes at the single-cell level. Wild-type K. pneumoniae skews macrophages toward an M2-like state. In vitro experiments probing pig bone marrow-derived macrophages uncovered the role for the M2 transcriptional factor STAT6 and that Klebsiella-induced il-10 expression is controlled by p38 and extracellular signal-regulated kinase (ERK). Klebsiella-induced macrophage polarization is dependent on the capsule. Together, the findings of this study support the utility of the EVLP model using pig lungs as a platform to investigate the infection biology of respiratory pathogens.
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