Title : Utilizing plant derived extracellular vesicles for drug delivery and therapeutic development
Abstract:
Introduction
Plant-Derived Extracellular Vesicles (PDEVs), nanosized lipid bilayer-enclosed vesicles, have recently emerged as an exciting and versatile platform with significant potential for treating various gastrointestinal disorders and supporting overall human health. Despite their promise, the therapeutic potential and characteristic properties of PDEVs remain underexplored, largely due to challenges in establishing efficient, reproducible purification protocols and the relatively limited scope of current research.
Methods
Using citrullus lanatus (watermelon) as an initial model, EVs were isolated using differential ultracentrifugation. Comprehensive structural analysis was performed using transmission electron microscopy (TEM), while nanoparticle tracking analysis (NTA) provided quantitative size distributions and concentrations. This isolation method was applied to various plant sources (citrus sinensis, brassica oleracea, solanum lycopersicum, vaccinium corymbosum, and capsicum annuum) to further demonstrate its reproducibility. The functional uptake of these WMEVs was demonstrated in vitro using triple-negative breast cancer cells. WMEVs stained with BioTracker 555 orange dye and incubated with MDA-MB-231 cells were fixed at various time points (1 hour, 2 hours, 4 hours, and 6 hours) and imaged using light microscopy. To assess the biodistribution of PDEVs, 100 μL of WMEVs stained with DiR were injected into BALB/c mice and tracked at various time points. To evaluate their potential as drug delivery vehicles, a microfluidic-based loading strategy was used to encapsulate doxorubicin (DOX) within Nicotiana tabacum protoplast-derived extracellular vesicles.
Results
A robust and reproducible methodology for isolating PDEVs was developed, enabling the recovery of high-purity PDEVs suitable for both characterization and functional assays, ensuring consistency and scalability for future studies. These PDEVs were found to be within compatible ranges of mammary-derived EV morphology. WMEVs have demonstrated successful cellular uptake in MDA-MB-231 cells between 2 and 4 hours and localization within the cells at 6 hours. DiR-stained WMEVs injected into BALB/c mice were localized within the liver and the spleen.
Summary/Conclusion:
These findings not only validate our optimized isolation protocol but also provide a proof-of-concept for using PDEVs as a safe, biocompatible, and plant-based drug-delivery platform. By advancing both the isolation and application of PDEVs, this work lays an important foundation for developing novel therapeutic strategies targeting cancer, gastrointestinal diseases, and potentially other systemic disorders.
