Small extracellular vesicles (sEVs) isolated from animal sources are among the most investigated types of cell-free therapeutic tools to cure different diseases. sEVs have been isolated from a variety of sources, ranging from prokaryotes to animals and plants. Human-derived sEVs have many uses in pre- and clinical studies in medicine and drug delivery, while plant-derived EVs, also known as plant-derived nanovesicles (PDNVs), have not been widely investigated until the second decade of the 21st century. For the past five years, there has been a rapid rise in the use of plant EVs as a therapeutic tool due to the ease of massive production with high efficacy and yield of preparation. Plant EVs contain various active biomolecules such as proteins, regulatory RNAs, and secondary metabolites and play a key role in inter-kingdom communications. Many studies have already investigated the potential application of plant EVs in preventing and treating cancer, inflammation, infectious diseases, and tissue regeneration with no sign of toxicity and are therefore considered safe. However, due to a lack of universal markers, the properties of plant EVs have not been extensively studied. Concerns regarding the safety and therapeutic function of plant EVs derived from genetically modified plants have been raised. In this paper, we review the physiological role of EVs in plants. Moreover, we focus on molecular and cellular mechanisms involved in the therapeutic effects of plant EVs on various human diseases. We also provide detailed information on the methodological aspects of plant EV isolation and analysis, which could pave the way for future clinical translation.