Researcher(s)
- Paige Turner, Applied Molecular Biology & Biotechnology, University of Delaware
Faculty Mentor(s)
- Mona Batish, Medical & Molecular Sciences, University of Delaware
Abstract
Extracellular vesicles (EVs) are small, membrane-enclosed particles naturally released by cells into the extracellular space. Extracellular vesicles carry cargo such as proteins, drugs, and Ribose Nucleic Acid (RNA). Once released, these particles can be absorbed and their cargo utilized by adjacent cells. Exporting RNA molecules from cells via EVs could improve intercellular communication. This research project aims to investigate the uptake of extracellular vesicle cargo to gain a deeper understanding of their role in mediating intercellular communication. The study examined whether extracellular vesicles carrying envelope proteins increase their uptake by target cells.
Cultured cells were transfected with specific plasmids to naturally produce EVs under desired conditions. The produced extracellular vesicles contain mCherry RNA (a fluorescent reporter of gene expression) and proteins to aid in their uptake by the receptor cells. The extracellular vesicles were released by the donor HEK-293T cells into media from which EVs were isolated and added to fresh media to create a mix. This mix was then added to a 12-well plate containing recipient HEK-293T cells on coverslips. The technique Single-molecule Fluorescence in situ Hybridization (smFISH) was utilized to image mCheery RNA and to determine if the cells took the EVs.
Preliminary results showed that EVs with upregulated envelope protein show an increase in the uptake of EVs cargo as seen by the mCherry signal via smFISH. Further work needs to be performed to determine the efficient transfer and uptake of cargo via envelope protein overexpression. EV-based cargo delivery systems can be explored for various applications such as gene therapy where different regulatory molecules can be packed as cargo into EVs.