Engineered EVs Mitigate Endothelial Cell Dysfunction

Background: Endothelial cell (EC) dysfunction driven by inflammation is a key pathological feature of cardiovascular diseases (CVDs). Developing anti-inflammatory strategies to improve EC function represents a critical therapeutic target. However, there are currently no EC-specific therapies for CVDs. Extracellular vesicles (EVs) are emerging as promising drug carriers, due to their ability to be engineered for targeted delivery through modification of their cargo and surface molecules. In this study, we explored the potential of engineered EVs carrying a novel therapeutic miRNA, selected from milk EV cargo with anti-inflammatory properties, to restore EC function. Methods: HEK293 cell-derived EVs were engineered by fusing a vascular cell adhesion molecule-1-targeted peptide (VTP) and GFP to the extra-exosomal N-terminus of the Lamp2b protein, producing VTP-EVs. Control EVs and VTP-EVs were isolated through centrifugation and characterized by western blotting and nanoparticle tracking analysis. The localization of VTP-EVs was examined in LPS-treated human umbilical vein endothelial cells (HUVECs) and C57BL/6 mice. To evaluate therapeutic potential, the anti-inflammatory effects of candidate miRNAs identified from milk EV miRNAs were tested. Subsequently, miR-494-3p was transfected into producer HEK293 cells, and encapsulated within VTP-EVs (VTP-EV miR-494 ). The EC targeting and therapeutic effects of VTP-EV miR-494 were assessed in LPS-treated HUVECs and high-fat diet-induced obese C57BL/6 mice. Results: VTP-EVs exhibited similar biophysical characteristics to control EVs but demonstrated significantly enhanced targeting of ECs in both LPS-treated HUVECs and LPS-administered C57BL/6 mice. VTP-EV miR-494 effectively reduced the expression of inflammatory markers in LPS-treated HUVECs and partially restored EC function in the mesenteric arteries of obese mice. Conclusion: VTP-EVs offers a promising therapeutic strategy for CVDs associated with EC dysfunction. Future studies will focus on evaluating the therapeutic efficacy of these engineered EVs in additional CVD animal models and to enhance the targeting capabilities by incorporating additional EC-specific markers. Fraternal Order of Eagles Diabetes Research Center This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.