Acute lung injury (ALI) is an inflammatory lung disease, which manifests itself in patients as acute respiratory distress syndrome (ARDS). Previous studies have implicated alveolar-epithelial succinate in ALI protection. Therefore, we hypothesized that targeting alveolar succinate dehydrogenase SDH A would result in elevated succinate levels and concomitant lung protection. Wild-type (WT) mice or transgenic mice with targeted alveolar-epithelial Sdha or hypoxia-inducible transcription factor Hif1a deletion were exposed to ALI induced by mechanical ventilation. Succinate metabolism was assessed in alveolar-epithelial via mass spectrometry as well as redox measurements and evaluation of lung injury. In WT mice, ALI induced by mechanical ventilation decreased SDHA activity and increased succinate in alveolar-epithelial. In vitro, cell-permeable succinate decreased epithelial inflammation during stretch injury. Mice with inducible alveolar-epithelial Sdha deletion (Sdha loxp/loxp SPC-CreER mice) revealed reduced lung inflammation, improved alveolar barrier function, and attenuated histologic injury. Consistent with a functional role of succinate to stabilize HIF, Sdha loxp/loxp SPC-CreER experienced enhanced Hif1a levels during hypoxia or ALI. Conversely, Hif1a loxp/loxp SPC-CreER showed increased inflammation with ALI induced by mechanical ventilation. Finally, wild-type mice treated with intratracheal dimethlysuccinate were protected during ALI. These data suggest that targeting alveolar-epithelial SDHA dampens ALI via succinate-mediated stabilization 2 of 18 | VOHWINKEL Et aL.

Acute lung injury (ALI) is a severe form of lung inflammation causing acute respiratory distress syndrome in patients. ALI pathogenesis is closely linked to uncontrolled alveolar inflammation.We hypothesize that specific enzymes of the glycolytic pathway could function as key regulators of alveolar inflammation. Therefore, we screened isolated alveolar epithelia from mice exposed to ALI induced by injurious ventilation to assess their metabolic responses. These studies pointed us towards a selective role for isoform 3 of the 6-phosphofructo-2-kinase/fructose-2,6bisphosphatase (PFKFB3). Pharmacologic inhibition or genetic deletion of Pfkfb3 in alveolar epithelia (Pfkfb3 loxp/loxp SPC-ER-Cre+ mice) was associated with profound increases in ALI during injurious mechanical ventilation or acid installation. Studies in genetic models linked Pfkfb3 expression and function to hypoxia-inducible factor Hif1a. Intra-tracheal pyruvate instillation not only reconstituted Pfkfb3 loxp/loxp or Hif1a loxp/loxp SPC ER Cre+ mice, but pyruvate was also effective in ALI treatment of wild-type mice. Finally, proof-of-principle studies in human lung biopsies confirmed increased PFKFB3 staining in injured lungs and co-localized PFKFB3 to alveolar epithelia. These studies reveal a specific role for PFKFB3 in counterbalancing alveolar inflammation and lay the groundwork for novel metabolic therapeutic approaches during ALI.

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