Christman, J. W. | Search bioRxiv

bioRxiv2025

Specialized Pro-Resolving Mediator loaded Extracellular Vesicles Mitigate Pulmonary Inflammation

Specialized pro-resolving mediators (SPMs), including lipoxins derived from arachidonic acid and resolvins, protectins, and maresins derived from docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), orchestrate the active resolution of inflammation. These SPMs are biosynthesized through the coordinated interaction of various cells in a process known as transcellular biosynthesis, involving the sequential action of cyclooxygenase-2 (COX-2), 5-lipoxygenase (5-LOX), 12-lipoxygenase (12-LOX), and/or 15-lipoxygenase (15-LOX) enzymes. Additionally, Aspirin-triggered Resolvins are produced by acetylated COX-2, along with various lipoxygenases. Although SPMs regulate various cellular processes to actively resolve inflammation, their in vivo levels are typically low. To address this limitation, we engineered a multigene expression vector that co-expresses COX-2, 5-LOX, and 15-LOX, potentiating the synthesis of various SPMs. HEK293T cells transfected with this vector and cultured with fatty acid-free BSA-complexed DHA, EPA, and aspirin, successfully mimicked both transcellular and aspirin-triggered biosynthesis of Resolvins. These Resolvins are packaged into extracellular vesicles, which significantly inhibited neutrophil adhesion to endothelial cells, preserved endothelial monolayer barrier integrity, suppressed NF-{kappa}B reporter activity, and enhanced macrophage efferocytosis in vitro. Notably, post-injury administration of Resolvin-loaded EVs mitigated pulmonary inflammation in LPS-treated mice without causing systemic or pulmonary toxicity. In summary, we report a novel cell-based platform for generating Resolvin-loaded EVs that mitigate pulmonary inflammation in mouse models, underscoring their potential for treating other acute inflammatory diseases.

synthetic biology↗

bioRxiv2020

Mechanosensitive activation of mTORC1 mediates ventilator induced lung injury during the acute respiratory distress syndrome

Acute respiratory distress syndrome (ARDS) is a highly lethal condition that impairs lung function and causes respiratory failure. Mechanical ventilation maintains gas exchange in patients with ARDS, but exposes lung cells to physical forces that exacerbate lung injury. Our data demonstrate that mTOR complex 1 (mTORC1) is a mechanosensor in lung epithelial cells and that activation of this pathway during mechanical ventilation exacerbates lung injury. We found that mTORC1 is activated in lung epithelial cells following volutrauma and atelectrauma in mice and humanized in vitro models of the lung microenvironment. mTORC1 is also activated in lung tissue of mechanically ventilated patients with ARDS. Deletion of Tsc2, a negative regulator of mTORC1, in epithelial cells exacerbates physiologic lung dysfunction during mechanical ventilation. Conversely, treatment with rapamycin at the time mechanical ventilation is initiated prevents physiologic lung injury (i.e. decreased compliance) without altering lung inflammation or barrier permeability. mTORC1 inhibition mitigates physiologic lung injury by preventing surfactant dysfunction during mechanical ventilation. Our data demonstrate that in contrast to canonical mTORC1 activation under favorable growth conditions, activation of mTORC1 during mechanical ventilation exacerbates lung injury and inhibition of this pathway may be a novel therapeutic target to mitigate ventilator induced lung injury during ARDS.

molecular biology↗