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Introduction: Patients who survive early stage sepsis may progress to late stage sepsis which is characterised by immunosuppression believed to be due, in part, to immune cell paralysis. We have previously shown that Mesenchymal Stem Cells (MSCs) are effective in preclinical models of early stage sepsis (Devaney et al., 2015) and to restore macrophage function in early clinical sepsis. We sought to develop in vivo and in vitro late stage sepsis models to assess if MSCs could restore immune cell homeostasis in this setting. Methods: We established an in vivo model of pulmonary sepsis by intratracheal administration of K.pneumoniae to Sprague-Dawley rats. MSCs were administered at day two and immune cell and cytokine secretion profiles in the Bronchoalveolar Lavage (BAL) and blood were analysed at day five. An in vitro endotoxin tolerance model was then established using immune cell lines and primary macrophages from blood and BAL exposed to repeated high levels of endotoxin. Results: MSCs improve paralysed immune cell responsiveness to an endotoxin stimulus ex vivo as demonstrated by a restoration of cytokine secretion in cells isolated from treated animal models (Figure 1). Conclusions: MSCs have the potential to reverse late sepsis induced immune cell dysfunction caused by prolonged exposure to bacterial endotoxin ex vivo.

Acute respiratory distress syndrome (ARDS) continues to have a high mortality with no effective specific therapy. Anti-inflammatory and anti-oxidant therapeutic proteins have previously shown promise in in vivo ARDS models. Similarly, targeting cell permeability (‘leakiness’) associated with ARDS has shown potential. A synthetic mRNA vector for rapid production of protein, coupled with direct delivery to the injured lung via nebulisation, could overcome these limitations. mRNA complexed with a novel transfection reagent (Factor Bioscience Ltd.) encoding green fluorescent protein (GFP), anti-inflammatory and anti-oxidant therapeutic proteins was administered to aseptic or septic rat lungs through a vibrating mesh nebuliser (Aerogen), via the flexiVent (SCIREQ). Animals were harvested and physiological and histological parameters were assessed. An in vitro model of cell permeability employing Calu-3 epithelial cells was also examined using TIE2 and ANGPT1 mRNA targets. A system incorporating Ussing chambers facilitating the measurement of specific ionic transport mechanisms, and by extension fluid dynamics across cell monolayers, was enrolled. GFP mRNA expressed rapidly in lung epithelial cells (