Wednesday, May 8, 2024
5/8/2024
PROJECT SUMMARY Cells employ nanoscale mechanisms to secrete extracellular vesicles (EVs) that contain a variety of functional ligands on the lipid membrane. This mode of delivery offers an effective strategy to systemically activate membrane receptors over distance without a need for cell migration and juxtacrine interactions. However, the intrinsic properties of EVs that enable this function and their mechanisms of action remain generally unclear. To address these questions, we developed a facile strategy to functionalize EVs with defined multivalent extracellular ligands that can activate specific cell membrane receptors to induce therapeutic effects. As a proof-of-concept, we turned to the Notch pathway, since it employs a mechanistically direct, simple mechanism of activation with direct transcriptional outcomes in the development and homeostasis of most vascularized organs. Our preliminary data show that Notch ligand functionalized EVs but not synthetic liposomes with a more rigid formulation reverse edema and vascular permeability after endotoxemia-induced injury in mice. We also show that the ability of Notch ligand functionalized EVs to restore endothelial barrier function requires the activation of the canonical Notch signaling pathway in endothelial cells. We will build on these results to test the hypothesis that Notch ligand functionalized EVs therapeutically restore the endothelial barrier and tissue function. In Aim 1, we will determine the ideal characteristics that enable Notch ligand functionalized EVs to restore endothelial barrier integrity. We predict that EVs can sustain deformation during adhesion to cell membrane receptors under mechanical force due to their unique lipid compositions and water flux capability, thereby resulting in potent, yet specific activation of downstream pathways. In Aim 2, we will investigate how Notch ligand loaded EVs restore the endothelial barrier after injury, which Notch receptor is activated, and what gene targets are expressed. The project is highly multidisciplinary in that it will employ a combination of expertise in nanotechnology, chemistry, membrane biophysics, cellular and molecular biology, and in vivo approaches to address the specific aims. Success of the project will enable the generalization of EVs as nanoscale therapeutic modalities to activate specific molecular pathways via multivalent ligand presentation that lead to tissue repair and regeneration.
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