Abstract
Extracellular vesicles (EVs) are naturally generated nanovesicles which potentially mediate the intercellular communication and interorgan crosstalk. EVs have recently gained significant interest as a promising material for delivery of therapeutics. Small RNAs, including small interfering RNA (siRNA) and microRNA (miRNA), provide a great therapeutic strategy for treating human diseases. However, it remains a challenge to deliver unconjugated small RNAs to the target tissue or cells. The delivery of small RNAs in an EV-encapsulating manner has a number of advantages, such as enhancing the concentration of small RNAs, improving the uptake of small RNAs by the recipient cells, and potentially achieving a cell-specific delivery. In this chapter, a protocol is provided for EV preparation and loading with small RNAs. Additionally, a detailed experimental protocol for tracking and validating small RNA delivery into the lungs is described. Overall, the described protocols are valuable for delivering functional small RNAs both in vitro and in vivo.