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Despite the approval of several artificial nanotherapeutics for the treatment of triple-negative breast cancer (TNBC), significant challenges, including unsatisfactory therapeutic outcomes, severe side effects, and the high cost of large-scale production, still restrict their long-term application. In contrast, plant-derived extracellular vesicles (PEVs) exhibit promising potential in cancer therapy due to their negligible systemic toxicity, high bioavailability and cost- effectiveness. In this study, we developed an alternative strategy to inhibit TNBC via Platycodon grandiflorum (PG)-derived extracellular vesicles (PGEVs). The PGEVs were isolated by ultracentrifugation and sucrose gradient centrifugation method and contained adequate functional components such as proteins, lipids, RNAs and active molecules. PGEVs exhibited remarkable stability, tolerating acidic digestion and undergoing minimal changes in simulated gastrointestinal fluid. They were efficiently taken up by tumor cells and induced increased production of reactive oxygen species (ROS), leading to tumor cell proliferation inhibition and apoptosis, particularly in the TNBC cell line 4T1. Additionally, PGEVs facilitated the polarization of tumor-associated macrophages (TAMs) toward M1 phenotype and increased the secretion of pro-inflammatory cytokines. Further in vivo investigations revealed that PGEVs efficiently accumulated in 4T1 tumors and exerted significant therapeutic effects through boosting systemic anti-tumor immune responses and modulating the gut microbiota whether administered orally or intravenously (i.v.). In conclusion, these findings highlight PGEVs as a promising natural, biocompatible and efficient nanotherapeutic candidate for treating TNBC. [ABSTRACT FROM AUTHOR]

Background/Objectives: Stress protein HSP70 administered exogenously has demonstrated high potential as an efficient adjuvant in antitumor immune response. To enhance the antigen-presenting activity, bioavailability, and stability of exogenous recombinant human HSP70, we propose incorporating it into plant extracellular vesicles. Earlier, we found that grapefruit-derived extracellular vesicles (GEV) were able to store the protein with no loss of its major function, chaperone activity. Methods: In this study, we tested whether HSP70 loaded into GEV (GEV-HSP70) could elicit an antitumor immune response in cellular and animal models of colorectal cancer. Results: To test the hypothesis in vitro, human and mouse colorectal cancer cell lines were used. We have shown that the addition of HSP70, either in free form or as part of GEVs, increases the sensitivity of human (HCT-116, DLD1) or mouse (CT-26) colon cancer cells to mouse cytotoxic lymphocytes and human NK-92 cells. Moreover, the amount of protein in the form of GEV-HSP70 required to cause the same activation of antitumor immunity was 20 times less than when HSP70 was added in free form. In a colon carcinoma model in vivo, GEV-HSP70 were inoculated subcutaneously into BALB/c mice together with CT-26 cells to form a tumor node. As compared with the control groups, we observed an increase in the lifespan of animals and a decrease in the tumor size, as well as a decrease in the level of TGFB1 IL-10 factors in the blood plasma. In vitro analysis of the immunomodulatory activity of GEV-HSP70 showed that antitumor response in GEV-HSP70-treated mice was associated with the accumulation of CD8+ cells. Conclusions: These results demonstrate the high feasibility and efficacy of the new technique based on HSP70 encapsulated in plant vesicles in activation of the specific response to colon tumors. [ABSTRACT FROM AUTHOR]

Abstract Rheumatoid arthritis (RA), a form of autoimmune inflammation, is marked by enduring synovial inflammation and the subsequent impairment of joint function. Despite the availability of conventional treatments, they are often marred by significant side effects and the associated high costs. Plant-derived extracellular vesicles (PEVs) offer a compelling alternative, owing to their abundant availability, affordability, low immunogenicity, high biocompatibility, and feasibility for large-scale production. These vesicles enhance intercellular communication by transferring intrinsic bioactive molecules. In our research, we delve into the capacity of PEVs to treat RA, highlighting the role of ginger-derived extracellular vesicles (GDEVs). By conjugating GDEVs with folic acid (FA), we have developed FA-GDEVs that maintain their inherent immunomodulatory properties. FA-GDEVs are designed to selectively target M1 macrophages in inflamed joints via the folate receptors (FRs). Our in vitro findings indicate that FA-GDEVs promote the polarization towards a reparative M2 macrophage phenotype by modulating the PI3K-AKT pathway. Further corroboration comes from in vivo studies, which demonstrate that FA-GDEVs not only concentrate efficiently in the affected joints but also markedly reduce the manifestations of RA. Synthesizing these findings, it is evident that FA-GDEVs emerge as a hopeful candidate for RA treatment, offering benefits such as safety, affordability, and therapeutic efficacy. Graphical abstract

Exosomes are extracellular nanovesicles secreted by all cell types and have been studied to understand and treat many human diseases. Exosomes are involved in numerous physiological and pathological processes, intercellular communication, and the transfer of substances. Over the years, several studies have explored mammalian-derived exosomes for therapeutic and diagnostic uses. Only recently have plant-derived extracellular vesicles (EVs) attracted attention for their ability to overcome many defects associated with using mammalian-derived extracellular vesicles, such as safety and scale-up issues. The ease of large-scale production, low toxicity, low immunogenicity, efficient cellular uptake, high biocompatibility, and high stability of these nanovesicles make them attractive for drug delivery systems. In addition, their native contents of proteins, miRNAs and secondary metabolites could be exploited for pharmaceutical applications in combination with other drugs. The present review intends to provide adequate tools for studying and developing drug delivery systems based on plant-derived EVs. Therefore, indications concerning extraction methods, characterisation, and drug loading will be offered. Their biological composition and content will also be reported. Finally, the current applications of these systems as nanocarriers for pharmacologically active substances will be shown. [ABSTRACT FROM AUTHOR]

The growing interest in plant-origin active molecules with medicinal properties has led to a revaluation of plants in the pharmaceutical field. Plant-derived extracellular vesicles (PDEVs) have emerged as promising candidates for next-generation drug delivery systems due to their ability to concentrate and deliver a plethora of bioactive molecules. These bilayer membranous vesicles, whose diameter ranges from 30 to 1000 nm, are released by different cell types and play a crucial role in cross-kingdom communication between plants and humans. Notably, PDEVs have demonstrated efficacy in treating various diseases, including cancer, alcoholic liver disease, and inflammatory bowel disease. However, further research on plant vesicles is necessary to fully understand their traits and purposes. This study investigates the phototoxic effects of extracellular vesicles (EVs) from Aloe arborescens, Aloe barbadensis, and Aloe chinensis on the human melanoma cell line SK-MEL-5, focusing on their anthraquinone content, recognized as natural photosensitizers. The phototoxic impact of Aloe EVs is associated with ROS production, leading to significant oxidative stress in melanoma cells, as validated by a metabolome analysis. These findings suggest that EVs from Aloe arborescens, Aloe barbadensis, and Aloe chinensis hold promise as potential photosensitizers, thus highlighting their potential for future application in photodynamic cancer therapy and providing valuable insights into the possible utilization of PDEVs for therapeutic purposes. [ABSTRACT FROM AUTHOR]

Plant-Derived Extracellular Vesicles extracellular vesicles (PDEVs) from organic agriculture (without the use of pesticides and microbicides) contain high levels of antioxidants. Organic PDEVs have shown an increased antioxidant power compared to PDEVs from single plants, suggesting a synergistic effect of the bioactives constitutively expressed in the PDEVs from single fruits. With this study, we wanted to investigate the beneficial effects of a mix of PDEVs on human skin cells. We found detectable levels of citric acid, ascorbic acid, glutathione, catalase, and SOD in a mix of PDEVs deriving from five different fruits (grape, red orange, papaya, pomegranate, and tangerine). We then treated H2O2-conditioned fibroblasts with the mix of PDEVs. The results showed that the PDEVs' mixture reverted the H2O2-induced redox imbalance, restoring mitochondrial homeostasis, with a strong reduction of mitochondrial anion superoxide and an increase in sirtuin levels. The antioxidant action was consistent with wound repair on a lesion produced in a fibroblast's monolayer. This result was consistent with an increased level of vimentin and matrix metalloproteinase-9, whose expression is directly related to the efficiency of the reparative processes. These data support a beneficial role of PDEVs in both preventing and treating skin injuries through their potent antioxidant and reparative activities. [ABSTRACT FROM AUTHOR]

This systematic review offers a comprehensive analysis of plant-derived extracellular vesicles (PDEVs) as emerging drug delivery systems, focusing on original research articles published between 2016 and 2024 that exclusively examine the use of PDEVs for drug delivery. After a rigorous search across multiple databases, 20 relevant studies out of 805 initial results were selected for analysis. This review systematically summarizes the critical data on PDEV components, isolation methods, and drug-loading techniques. It highlights the potential of PDEVs to significantly enhance drug safety and efficacy, reduce dosage and toxicity, and align drug development with sustainable and environmentally friendly biotechnological processes. This review also emphasizes the advantages of PDEVs over mammalian-derived vesicles, such as cost-effectiveness, higher yield, and reduced immunogenicity. Additionally, it explores the synergistic potential between encapsulated drugs and bioactive compounds naturally present in PDEVs. This study acknowledges the challenges in standardizing isolation and formulation methods for clinical use. Overall, this review provides valuable insights into the current state and future directions of PDEV-based drug delivery systems, highlighting their promising role in advancing pharmaceutical research and development. [ABSTRACT FROM AUTHOR]

Food‐derived extracellular vesicles (FEVs) are nanoscale membrane vesicles obtained from dietary materials such as breast milk, plants and probiotics. Distinct from other EVs, FEVs can survive the harsh degrading conditions in the gastrointestinal tract and reach the intestines. This unique feature allows FEVs to be promising prebiotics in health and oral nanomedicine for gut disorders, such as inflammatory bowel disease. Interestingly, therapeutic effects of FEVs have recently also been observed in non‐gastrointestinal diseases. However, the mechanisms remain unclear or even mysterious. It is speculated that orally administered FEVs could enter the bloodstream, reach remote organs, and thus exert therapeutic effects therein. However, emerging evidence suggests that the amount of FEVs reaching organs beyond the gastrointestinal tract is marginal and may be insufficient to account for the significant therapeutic effects achieved regarding diseases involving remote organs such as the liver. Thus, we herein propose that FEVs primarily act locally in the intestine by modulating intestinal microenvironments such as barrier integrity and microbiota, thereby eliciting therapeutic impact remotely on the liver in non‐gastrointestinal diseases via the gut‐liver axis. Likewise, drugs delivered to the gastrointestinal system through FEVs may act via the gut‐liver axis. As the liver is the main metabolic hub, the intestinal microenvironment may be implicated in other metabolic diseases. In fact, many patients with non‐alcoholic fatty liver disease, obesity, diabetes and cardiovascular disease suffer from a leaky gut and dysbiosis. In this review, we provide an overview of the recent progress in FEVs and discuss their biomedical applications as therapeutic agents and drug delivery systems, highlighting the pivotal role of the gut‐liver axis in the mechanisms of action of FEVs for the treatment of gut disorders and metabolic diseases. [ABSTRACT FROM AUTHOR]

Current mRNA vaccines are mainly administered via intramuscular injection, which induces good systemic immunity but limited mucosal immunity. Achieving mucosal immunity through mRNA vaccination could diminish pathogen replication at the entry site and reduce interhuman transmission. However, delivering mRNA vaccines to mucosae faces challenges like mRNA degradation, poor entry into cells, and reactogenicity. Encapsulating mRNA in extracellular vesicles may protect the mRNA and reduce reactogenicity, making mucosal mRNA vaccines possible. Plant-derived extracellular vesicles from edible fruits have been investigated as mRNA carriers. Studies in animals show that mRNA vehiculated in orange-derived extracellular vesicles can elicit both systemic and mucosal immune responses when administered by the oral, nasal, or intramuscular routes. Once lyophilized, these products show remarkable stability. The optimization of mRNA to improve translation efficiency, immunogenicity, reactogenicity, and stability can be obtained through adjustments of the 5′cap region, poly-A tail, codons selection, and the use of nucleoside analogues. Recent studies have also proposed self-amplifying RNA vaccines containing an RNA polymerase as well as circular mRNA constructs. Data from parenterally primed animals demonstrate the efficacy of nasal immunization with non-adjuvanted protein, and studies in humans indicate that the combination of a parenteral vaccine with the natural exposure of mucosae to the same antigen provides protection and reduces transmission. Hence, mucosal mRNA vaccination would be beneficial at least in organisms pre-treated with parenteral vaccines. This practice could have wide applications for the treatment of infectious diseases. [ABSTRACT FROM AUTHOR]

Xiaoliang Liu,1,* Kecheng Lou,2,* Yunmeng Zhang,3 Chuanxiao Li,1 Shenghong Wei,1 Shangzhi Feng1 1Department of Urology, Jiujiang University Clinic College/Hospital, Jiujiang, Jiangxi, People’s Republic of China; 2Department of Urology, Lanxi People’s Hospital, Jinhua, Zhejiang, People’s Republic of China; 3Department of Anesthesiology, Jiujiang College Hospital, Jiujiang, Jiangxi, People’s Republic of China*These authors contributed equally to this workCorrespondence: Shangzhi Feng, Email 1131718266@qq.comAbstract: Botanical preparations for herbal medicine have received more and more attention from drug researchers, and the extraction of active ingredients and their successful clinical application have become an important direction of drug research in major pharmaceutical companies, but the complexity of extracts, multiple side effects, and significant individual differences have brought many difficulties to the clinical application of herbal preparations. It is noteworthy that extracellular vesicles as active biomolecules extracted from medicinal plants are believed to be useful for the treatment of a variety of diseases, including cancer, inflammation, regenerative-restorative and degenerative diseases, which may provide a new direction for the clinical utilization of herbal preparations. In this review, we sort out recent advances in medicinal plant extracellular vesicles and discuss their potential as disease therapeutics. Finally, future challenges and research directions for the clinical translation of medicinal plant extracellular vesicles are also discussed, and we expect that continued development based on medicinal plant extracellular vesicles will facilitate the clinical application of herbal preparations.Keywords: plant-derived extracellular vesicles, Chinese medicine preparation, nanoparticle, drug delivery nanoplatforms, nanotherapeutics

Background/Objectives: Stress protein HSP70 administered exogenously has demonstrated high potential as an efficient adjuvant in antitumor immune response. To enhance the antigen-presenting activity, bioavailability, and stability of exogenous recombinant human HSP70, we propose incorporating it into plant extracellular vesicles. Earlier, we found that grapefruit-derived extracellular vesicles (GEV) were able to store the protein with no loss of its major function, chaperone activity. Methods: In this study, we tested whether HSP70 loaded into GEV (GEV-HSP70) could elicit an antitumor immune response in cellular and animal models of colorectal cancer. Results: To test the hypothesis in vitro, human and mouse colorectal cancer cell lines were used. We have shown that the addition of HSP70, either in free form or as part of GEVs, increases the sensitivity of human (HCT-116, DLD1) or mouse (CT-26) colon cancer cells to mouse cytotoxic lymphocytes and human NK-92 cells. Moreover, the amount of protein in the form of GEV-HSP70 required to cause the same activation of antitumor immunity was 20 times less than when HSP70 was added in free form. In a colon carcinoma model in vivo, GEV-HSP70 were inoculated subcutaneously into BALB/c mice together with CT-26 cells to form a tumor node. As compared with the control groups, we observed an increase in the lifespan of animals and a decrease in the tumor size, as well as a decrease in the level of TGFB1 IL-10 factors in the blood plasma. In vitro analysis of the immunomodulatory activity of GEV-HSP70 showed that antitumor response in GEV-HSP70-treated mice was associated with the accumulation of CD8+ cells. Conclusions: These results demonstrate the high feasibility and efficacy of the new technique based on HSP70 encapsulated in plant vesicles in activation of the specific response to colon tumors.

Background/aim: Small extracellular vesicles (SEVs) are known to have an impact on the physiological conditions of target cells, are a critical component of cell-to-cell communication, and have been implicated in a variety of diseases. Although it has been proposed that edible plant-derived nanoparticles have an effect on communication with mammalian cells, the influence of these nanoparticles on cancer cell development has yet to be explored. Materials and methods: In order to characterize small extracellular vesicles obtained from garlic, specific SEV surface markers, antibodies, and size detections were identified using scanning electron microscopy and nanoparticle tracking analysis. Human hepatoma (Hep3B), human neuroblastoma (SH-SY5Y), human pancreatic adenocarcinoma (Panc-1a), human glioblastoma (U87), prostate cancer (PC-3), and human umbilical vein endothelial (HUVEC) cell lines were treated with garlic SEVs to examine their anticancer properties. Results: Annexin V FITC/PI staining for apoptosis, mRNA, and protein expression levels via RT-PCR and ELISA indicated that garlic SEVs triggered apoptosis by activating the intrinsic pathway. Our findings support the idea that SEVs produced from garlic may trigger apoptotic cell death in cancer cells while having no effect on healthy cells. Conclusion: It was discovered that plant SEVs had anti-cancer effects by activating caspase-mediated apoptosis. [ABSTRACT FROM AUTHOR]

Plant-derived extracellular vesicles (PDEV) constitute nanoscale entities comprising lipids, proteins, nucleic acids and various components enveloped by the lipid bilayers of plant cells. These vesicles play a crucial role in facilitating substance and information transfer not only between plant cells but also across different species. Owing to its safety, stability, and the abundance of raw materials, this substance has found extensive utilization in recent years within research endeavors aimed at treating various diseases. This article provides an overview of the pathways and biological characteristics of PDEV, along with the prevalent methods employed for its isolation, purification, and storage. Furthermore, we comprehensively outline the therapeutic implications of diverse sources of PDEV in musculoskeletal system disorders. Additionally, we explore the utilization of PDEV as platforms for engineering drug carriers, aiming to delve deeper into the significance and potential contributions of PDEV in the realm of the musculoskeletal system.

Introduction: Triple negative breast cancer (TNBC), a highly aggressive subtype accounting for 15–20% of all breast cancer cases, faces limited treatment options often accompanied by severe side effects. In recent years, natural extracellular nanovesicles derived from plants have emerged as promising candidates for cancer therapy, given their safety profile marked by non-immunogenicity and absence of inflammatory responses. Nevertheless, the potential anti-cancer effects of Citrus limonL.-derived extracellular nanovesicles (CLENs) for breast cancer treatment is still unexplored.Methods: In this study, we investigated the anti-cancer effects of CLENs on two TNBC cell lines (4T1 and HCC-1806 cells) under growth conditions in 2D and 3D culture environments. The cellular uptake efficiency of CLENs and their internalization mechanism were evaluated in both cells using confocal microscopy. Thereafter, we assessed the effect of different concentrations of CLENs on cell viability over time using a dual approach of Calcein-AM PI live-dead assay and CellTiter-Glo bioluminescence assay. We also examined the influence of CLENs on the migratory and evasion abilities of TNBC cells through wound healing and 3D Matrigel drop evasion assays. Furthermore, Western blot analysis was employed to investigate the effects of CLENs on the phosphorylation levels of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), and extracellular signal- regulated kinase (ERK) expression.Results: We found that CLENs were internalized by the cells via endocytosis, leading to decreased cell viability, in a dose- and time-dependent manner. Additionally, the migration and evasion abilities of TNBC cells were significantly inhibited under exposed to 40 and 80 μg/mL CLENs. Furthermore, down-regulated expression levels of phosphorylated phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), and extracellular signal-regulated kinase (ERK), suggesting that the inhibition of cancer cell proliferation, migration, and evasion is driven by the inhibition of the PI3K/AKT and MAPK/ERK signaling pathways.Discussion: Overall, our results demonstrate the anti-tumor efficiency of CLENs against TNBC cells, highlighting their potential as promising natural anti-cancer agents for clinical applications in cancer treatment.

Plant-Derived Extracellular Vesicles extracellular vesicles (PDEVs) from organic agriculture (without the use of pesticides and microbicides) contain high levels of antioxidants. Organic PDEVs have shown an increased antioxidant power compared to PDEVs from single plants, suggesting a synergistic effect of the bioactives constitutively expressed in the PDEVs from single fruits. With this study, we wanted to investigate the beneficial effects of a mix of PDEVs on human skin cells. We found detectable levels of citric acid, ascorbic acid, glutathione, catalase, and SOD in a mix of PDEVs deriving from five different fruits (grape, red orange, papaya, pomegranate, and tangerine). We then treated H2O2-conditioned fibroblasts with the mix of PDEVs. The results showed that the PDEVs’ mixture reverted the H2O2-induced redox imbalance, restoring mitochondrial homeostasis, with a strong reduction of mitochondrial anion superoxide and an increase in sirtuin levels. The antioxidant action was consistent with wound repair on a lesion produced in a fibroblast’s monolayer. This result was consistent with an increased level of vimentin and matrix metalloproteinase-9, whose expression is directly related to the efficiency of the reparative processes. These data support a beneficial role of PDEVs in both preventing and treating skin injuries through their potent antioxidant and reparative activities.

The scientific interest in cannabis plants' beneficial properties has recently sparked certain interest in the possible functional characterization of plant-derived extracellular vesicles (PDEVs). Establishing the most appropriate and efficient isolation procedure for PDEVs remains a challenge due to vast differences in the physio-structural characteristics of different plants within the same genera and species. In this study, we employed a crude but standard isolation procedure for the extraction of apoplastic wash fluid (AWF) which is known to contain the PDEVs. This method includes a detailed stepwise process of PDEV extraction from five (5) cultivars of cannabis plants, namely: Citrus (C), Henola (HA), Bialobrezenski (BZ), Southern-Sunset (SS), and Cat-Daddy (CAD). Approximately, 150 leaves were collected from each plant strain. In order to collect PDEV pellets, apoplastic wash fluid (AWF) was extracted from plants via negative pressure permeabilization and infiltration followed by high-speed differential ultracentrifugation. Particle tracking analysis of PDEVs revealed particle size distribution in the range of 20 to 200 nm from all plant strains, while PDEV total protein concentration from HA was higher than that of SS. Although HA-PDEVs' total protein was higher than SS-PDEVs, SS-PDEVs' RNA yield was higher than that of HA-PDEVs. Our result suggests that the cannabis plant strains contain EVs, and PDEV concentration from the cannabis plant could be age or strain dependent. Overall, the results provide a guide for the selection and optimization of PDEV isolation methods for future studies. [ABSTRACT FROM AUTHOR]

Abstract Background The past few years have witnessed a significant increase in research related to plant-derived extracellular vesicles (PDEVs) in biological and medical applications. Using biochemical technologies, multiple independent groups have demonstrated the important roles of PDEVs as potential mediators involved in cell-cell communication and the exchange of bio-information between species. Recently, several contents have been well identified in PDEVs, including nucleic acids, proteins, lipids, and other active substances. These cargoes carried by PDEVs could be transferred into recipient cells and remarkably influence their biological behaviors associated with human diseases, such as cancers and inflammatory diseases. Main body of the abstract This review summarizes the latest updates regarding PDEVs and focuses on its important role in nanomedicine applications, as well as the potential of PDEVs as drug delivery strategies to develop diagnostic and therapeutic agents for the clinical management of diseases, especially like cancers. Conclusion Considering its unique advantages, especially high stability, intrinsic bioactivity and easy absorption, further elaboration on molecular mechanisms and biological factors driving the function of PDEVs will provide new horizons for the treatment of human disease.

Plant-derived extracellular vesicles (EVs) isolated from seeds, leaves, and fruits have shown a significant therapeutic potential for their anticancer, anti-inflammatory, and antioxidant properties. The ability to transport bioactive molecules and the low toxicity give EVs remarkable versatility in the field of nanomedicine for the development of drug delivery systems. Moreover, the physicochemical stability in gastric and intestinal fluids makes them the ideal candidate as nutritional carriers in oral formulations. It is well known that the consumption of antioxidant molecules from dietary plant sources, such as fruits and vegetables, can prevent pathologies caused by oxidative damage, including inflammatory and cardiovascular disease, neurodegeneration, aging, and cancer. EVs present in plant juices are receiving a lot of interest concerning their biological relevance in terms of their health benefits. EVs from food might be new components participating in body homeostasis, as they are in contact with the intestinal tract. This review aims to report and discuss the main biological properties and nutraceutical use of plant-derived EVs as promising therapeutic tools, with a focus on anti-oxidant effect and as a basis in developing new food-derived technology. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs), which are natural nanocarriers characterized by a phospholipid bilayer structure, are released by living cells. They play a crucial role in the intercellular transport of proteins, nucleic acids, lipids, and metabolites, facilitating substance delivery and information exchange between cells. In light of recent numerous studies, EVs has been found to transcend their basic role as mere delivery vehicle. Instead, they demonstrate an impressive array of biological activities, displaying preventive and therapeutic potential in mitigating various pathological processes encompassing cancer, neoplastic proliferation, infectious diseases, and oxidative trauma. Particularly, EVs derived from edible plants (EPDEVs) have been emphasized for their extensive range of physiological regulatory functions in animals and humans, with the potential for targeted drug delivery through oral administration. Leveraging these advantages, EPDEVs are expected to have excellent competitiveness in clinical applications or preventive healthcare products. This review provides a brief overview of the biogenesis, structure, and composition of EPDEVs, and summarizes their biological functions and mechanisms. It also analyzes the methods for isolating and purifying plant-EVs, assessing their advantages and disadvantages; discusses the latest advancements in biomedical applications, and concludes with a prospective insight into the research and development directions of EPDEVs.

Plant cells release tiny membranous vesicles called extracellular vesicles (EVs), which are rich in lipids, proteins, nucleic acids, and pharmacologically active compounds. These plant-derived EVs (PDEVs) are safe and easily extractable and have been shown to have therapeutic effects against inflammation, cancer, bacteria, and aging. They have shown promise in preventing or treating colitis, cancer, alcoholic liver disease, and even COVID-19. PDEVs can also be used as natural carriers for small-molecule drugs and nucleic acids through various administration routes such as oral, transdermal, or injection. The unique advantages of PDEVs make them highly competitive in clinical applications and preventive healthcare products in the future. This review covers the latest methods for isolating and characterizing PDEVs, their applications in disease prevention and treatment, and their potential as a new drug carrier, with special attention to their commercial viability and toxicological profile, as the future of nanomedicine therapeutics. This review champions the formation of a new task force specializing in PDEVs to address a global need for rigor and standardization in PDEV research. [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) represent a diverse class of lipid bilayer membrane vesicles released by both animal and plant cells. These ubiquitous vesicles are involved in intercellular communication and transport of various biological cargos, including proteins, lipids, and nucleic acids. In recent years, interest in plant‐derived EVs has increased tremendously, as they serve as a scalable and sustainable alternative to EVs derived from mammalian sources. In vitro and in vivo findings have demonstrated that these plant‐derived vesicles (PDVs) possess intrinsic therapeutic activities that can potentially treat diseases and improve human health. In addition, PDVs can also act as efficient and biocompatible drug carriers. While preclinical studies have shown promising results, there are still several challenges and knowledge gaps that have to be addressed for the successful translation of PDVs into clinical applications, especially in view of the lack of standardised protocols for material handling and PDV isolation from various plant sources. This review provides the readers with a quick overview of the current understanding and research on PDVs, critically analysing the current challenges and highlighting the immense potential of PDVs as a novel class of therapeutics to treat human diseases. It is expected that this work will guide scientists to address the knowledge gaps currently associated with PDVs and promote new advances in plant‐based therapeutic solutions. [ABSTRACT FROM AUTHOR]

Introduction: The blood-brain barrier (BBB) is a highly selective structure that protects the central nervous system (CNS) while hindering the delivery of many therapeutic agents. This presents a major challenge in treating neurological disorders, such as multiple sclerosis, where effective drug delivery to the brain is crucial for improving patient outcomes. Innovative strategies are urgently needed to address this limitation., Areas Covered: This review explores the potential of extracellular vesicles (EVs) as innovative drug delivery systems capable of crossing the BBB. EVs are membrane-bound vesicles derived from cells, tissues, or plant materials, offering natural biocompatibility and therapeutic potential. Recent studies investigating the permeability of EVs and their mechanisms for crossing the BBB, such as transcytosis, are summarized. Special emphasis is placed on plant-derived EVs (PDEVs) due to their unique advantages in drug delivery. Challenges related to the large-scale production and therapeutic consistency of EVs are also discussed., Expert Opinion: EVs, particularly PDEVs, hold significant promise as scalable and noninvasive systems for CNS drug delivery. However, critical barriers such as improving standardization techniques, manufacturing processes and addressing scalability must be overcome to facilitate clinical translation. Collaborative efforts in research and innovation will be pivotal in realizing the therapeutic potential of EVs for neurological conditions.

Extracellular vesicles are functionally active, nanoscale, membrane-bound vesicles that can be secreted by all cells. They have a key role in most health and disease states and have gradually become a promising class of delivery vehicles for targeted therapies for a variety of diseases. Plant-derived extracellular vesicles have received increasing attention based on their easy availability, non-toxicity and high absorption. However, compared with mammalian extracellular vesicles, the role of these nanoparticles as nano-delivery systems in tumour therapy has been underestimated. In this paper, the application of plant-derived extracellular vesicles and their nano-derivatives as nano-delivery systems in tumour therapy is reviewed to illustrate their great application potential.

Oral administration is one of the most convenient and widely utilized methods of drug administration. However, many drugs were difficult to be administered orally due to their poor oral bioavailability. Designing a safe and effective oral drug delivery system is one of the basic strategies to overcome the poor oral bioavailability. Plant-derived extracellular vesicles (PDEVs) were found in a variety of plants and have similar physical and chemical properties to mammalian EVs. It has been proved that PDEVs can effectively encapsulate hydrophilic and hydrophobic drugs, remain stable in harsh gastrointestinal environments, and cross biological barriers to reach target tissues. Furthermore, the biological activity of PDEVs enables it to play a synergistic therapeutic role with drugs. In addition, the safety and high yield of PDEVs indicate their potential as oral drug carriers. In this review, we introduce the biogenesis, isolation, characterization and drug delivery methods of PDEVs, describe their stability, transport, delivery and therapeutic applications. Finally, the potential and challenges of PDEVs as drug carriers are discussed. [Display omitted] • Overview the properties of plant derived extracellular vesicles (PDEVs) • Outline of the recent applications of PDEVs in therapeutic • The potential of PDEVs as oral drug carriers were discussed • Analyses of the challenges of PDEVs as oral drug carriers [ABSTRACT FROM AUTHOR]

Plant-derived extracellular vesicles (PLEVs), as a type of naturally occurring lipid bilayer membrane structure, represent an emerging delivery vehicle with immense potential due to their ability to encapsulate hydrophobic and hydrophilic compounds, shield them from external environmental stresses, control release, exhibit biocompatibility, and demonstrate biodegradability. This comprehensive review analyzes engineering preparation strategies for natural vesicles, focusing on PLEVs and their purification and surface engineering. Furthermore, it encompasses the latest advancements in utilizing PLEVs to transport active components, serving as a nanotherapeutic system. The prospects and potential development of PLEVs are also discussed. It is anticipated that this work will not only address existing knowledge gaps concerning PLEVs but also provide valuable guidance for researchers in the fields of food science and biomedical studies, stimulating novel breakthroughs in plant-based therapeutic options. • Plant-derived extracellular vesicles (PLEVs) are important for disease treatment. • Advances of PLEVs purification and surface modification engineering. • Latest progress in transporting bioactive substances as nano-therapeutic system. [ABSTRACT FROM AUTHOR]

Almond is a traditional food with established beneficial effects on health. Nothing is known about the presence of extracellular vesicles (EVs), recently isolated from other plant material, ingested with food, or as engineered bioactive nanovectors. Aiming to develop and optimize a method to isolate almonds derived nano vesicles (ADNVs), we tested different protocols on pure, blanched and roasted almonds, and investigated the resulting biophysical features. The most feasible and valid method was the sucrose-based ultracentrifugation (DGUC). Size distribution ranged on average 100–200 nm. A negative zeta-potential in the range of –27 to –21 mV has been measured. Microscopy showed a spheroid morphology and preserved structural integrity of isolated ADNVs. Pure almond ADNVs showed the greater amount of TET8; PEN1 was also found, although cooking treatments resulted in a decrease. Attention should be paid to sample managing during DGUC steps and to sample deposition and drying for microscopy procedures. Our findings contribute to enriching the hot issue on EVs research from edible sources, giving reason to their possible role in interspecies communication, and their exploitation for the delivery of bioactive compounds or therapeutics molecules. The potentiality of almonds' industrial residuals should be investigated in further research. • Almond contains extracellular vesicles following density-based ultracentrifugation. • ADNVs show spheroid morphology and zeta-potential suggests colloidal stability. • ADNVs from pure almonds express tetraspanin TET8 compared to treated ;almonds. • ADNVs are preserved after thermal treatments but putative microvesicles are limited. • Critical steps refer to the fat layers and procedures prior to microscopy. [ABSTRACT FROM AUTHOR]

Chronic inflammation is associated with the occurrence of several diseases. However, the side effects of anti‐inflammatory drugs prompt the identification of new therapeutic strategies. Plant‐derived extracellular vesicles (PDEVs) are gaining increasing interest in the scientific community for their biological properties. We isolated PDEVs from the juice of Citrus limon L. (LEVs) and characterized their flavonoid, limonoid and lipid contents through reversed‐phase high‐performance liquid chromatography coupled to electrospray ionization quadrupole time‐of‐flight mass spectrometry (RP‐HPLC–ESI‐Q‐TOF‐MS). To investigate whether LEVs have a protective role on the inflammatory process, murine and primary human macrophages were pre‐treated with LEVs for 24 h and then were stimulated with lipopolysaccharide (LPS). We found that pre‐treatment with LEVs decreased gene and protein expression of pro‐inflammatory cytokines, such as IL‐6, IL1‐β and TNF‐α, and reduced the nuclear translocation and phosphorylation of NF‐κB in LPS‐stimulated murine macrophages. The inhibition of NF‐κB activation was associated with the reduction in ERK1‐2 phosphorylation. Furthermore, the ability of LEVs to decrease pro‐inflammatory cytokines and increase anti‐inflammatory molecules was confirmed ex vivo in human primary T lymphocytes. In conclusion, we demonstrated that LEVs exert anti‐inflammatory effects both in vitro and ex vivo by inhibiting the ERK1‐2/NF‐κB signalling pathway. [ABSTRACT FROM AUTHOR]

Plant-derived extracellular vesicles (EVs) isolated from seeds, leaves, and fruits have shown a significant therapeutic potential for their anticancer, anti-inflammatory, and antioxidant properties. The ability to transport bioactive molecules and the low toxicity give EVs remarkable versatility in the field of nanomedicine for the development of drug delivery systems. Moreover, the physicochemical stability in gastric and intestinal fluids makes them the ideal candidate as nutritional carriers in oral formulations. It is well known that the consumption of antioxidant molecules from dietary plant sources, such as fruits and vegetables, can prevent pathologies caused by oxidative damage, including inflammatory and cardiovascular disease, neurodegeneration, aging, and cancer. EVs present in plant juices are receiving a lot of interest concerning their biological relevance in terms of their health benefits. EVs from food might be new components participating in body homeostasis, as they are in contact with the intestinal tract. This review aims to report and discuss the main biological properties and nutraceutical use of plant-derived EVs as promising therapeutic tools, with a focus on anti-oxidant effect and as a basis in developing new food-derived technology.

The escalating prevalence of obesity poses significant health challenges due to its direct association with various diseases. Most existing medications, such as appetite suppressants and fat absorption inhibitors, suffer from limited effectiveness and undesirable side effects. Here, inspired by the versatile metabolic effects of turmeric, we developed a naturally derived nanoformulation of "Reconstructed Turmeric-derived Nanovesicles (Rec-tNVs)" for obesity treatment. Employing quantitative nanoflow cytometry, a four-orders-of-magnitude increase in curcumin content (∼10 8 molecules per particle) was identified in individual Rec-tNVs compared to their ultracentrifugation-isolated counterparts. Rec-tNVs, featuring highly aggregated curcumin arrangements and other coencapsulated bioactive compounds, demonstrated a dose-dependent lipid-lowering effect in mature 3T3-L1 cells by promoting lipolysis, suppressing lipogenesis, inducing adipocyte browning, and triggering apoptosis after internalization via multiple pathways. In vivo experiments revealed that Rec-tNVs alleviated obesity more effectively than free curcumin and achieved weight reductions of 18.68 and 14.56% through intragastric and subcutaneous delivery, respectively, in high-fat-diet mouse models over a four-week treatment period. These effects were attributed to targeted actions on adipose tissues and systemic impacts on metabolism and gut microbiota composition. Overall, this study underscores the multifaceted antiobesity efficacy of Rec-tNVs, and offers a promising paradigm for developing plant-derived nanovesicle-based therapeutics.

The release of extracellular vesicles (EVs) is a common language, used by living organisms from different kingdoms as a means of communication between them. Extracellular vesicles are lipoproteic particles that contain many biomolecules, such as proteins, nucleic acids, and lipids. The primary role of EVs is to convey information to the recipient cells, affecting their function. Plant-derived extracellular vesicles (PDEVs) can be isolated from several plant species, and the study of their biological properties is becoming an essential starting point to study cross-kingdom communication, especially between plants and mammalians. Furthermore, the presence of microRNAs (miRNAs) in PDEVs represents an interesting aspect for understanding how PDEVs can target the mammalian genes involved in pathological conditions such as cancer, inflammation, and oxidative stress. In particular, this review focuses on the history of PDEVs, from their discovery, to purification from various matrices, and on the functional role of PDEV-RNAs in cross-kingdom interactions. It is worth noting that miRNAs packaged in PDEVs can be key modulators of human gene expression, representing potential therapeutic agents.

Plant-derived extracellular vesicles (EVs) are capable of efficiency delivering mRNAs, miRNAs, bioactive lipids, and proteins to mammalian cells. Plant-derived EVs critically contribute to the ability of plants to defend against pathogen attacks at the plant cell surface. They also represent a novel candidate natural substance that shows potential to be developed for food, cosmetic, and pharmaceutical products. However, although plant-derived EVs are acknowledged as having potential for various industrial applications, little is known about how their stability is affected by storage conditions. In this study, we evaluated the stability of Dendropanax morbifera leaf-derived extracellular vesicles (LEVs) alone or combined with the preservatives, 1,3-butylene glycol (to yield LEVs-1,3-BG) or TMO (LEVs-TMO). We stored these formulations at −20, 4, 25, and 45 °C for up to 4 weeks, and compared the stability of fresh and stored LEVs. We also assessed the effect of freeze-thawing cycles on the quantity and morphology of the LEVs. We found that different storage temperatures and number of freeze-thawing cycles altered the stability, size distribution, protein content, surface charge, and cellular uptake of LEVs compared to those of freshly isolated LEVs. LEVs-TMO showed higher stability when stored at 4 °C, compared to LEVs and LEVs-1,3-BG. Our study provides comprehensive information on how storage conditions affect LEVs and suggests that the potential industrial applications of plant-derived EVs may be broadened by the use of preservatives.

Background: Plant-derived extracellular vesicles (PEVs) are studied as a natural carrier of functional biomolecules and as a potential system of targeted delivery of therapeutic agents. One of the urgent tasks in this direction is the selection of the carrier with optimal physicochemical parameters and morphology from a variety of plant sources. To date, vesicles from only a few plants were visualized using cryo-electron microscopy (cryo-EM). Here we investigated the morphology and physical parameters of extracellular vesicles from plant sources not previously studied utilizing this method. Methods: PEVs derived by ultracentrifugation from juice and cultural medium of 11 plants and mushrooms were studied using methods of cryo-EM. Samples were plunge frozen in liquid ethane with Vitrobot Mark IV and examined under cryogenic transmission electron microscope Titan Krios 60-300 (ThermoFisher Scientific, USA) in low dose mode using EPU software. Results: Most of the observed particles in each sample were classified as extracellular vesicles due to the presence of the lipid bilayer. Morphology and size characteristics of PEVs were determined and compared with each other. A variety of morphological configurations of PEVs were found: with single and multiple membranes, with different conformations and integrity state. Most of the isolated PEVs were single, round-shaped, and in a size range from 30 to 150 nm. Conclusion: Cryo-EM allowed us to obtain high-quality images of PEVs isolated from 11 plants and mushrooms (blueberry, chanterelle, cowberry, fly agaric, garlic, redcurrant, chlamydomonas, cucumber, shadberry, viburnum, gooseberry) which have been characterized by their size and morphology. From the data obtained, the most promising sources of vesicles were proposed. The approbation of the selected vesicles as effective delivery systems requires further research.

Plant-derived extracellular vesicles (PDEVs) have shown remarkable potential as sustainable, green, and efficient drug delivery nanocarriers. As natural nanoparticles containing lipids, protein, nucleic acids and secondary metabolites, they have received widespread attention as a replacement for mammalian exosomes in recent years. In this review, the advances in isolation, identification, composition, therapeutic effect, and clinical application prospect were comprehensively reviewed, respectively. In addition, the specific modification strategies have been listed focusing on the inherent drawbacks of the raw PDEVs like low targeting efficiency and poor homogeneity. With emphasis on their biology mechanism in terms of immune regulation, regulating oxidative stress and promoting regeneration in the anti-inflammatory field and application value demonstrated by citing some typical examples, this review about PDEVs would provide a broad and fundamental vision for the in-depth exploration and development of plant-derived extracellular vesicles in the in-vivo anti-inflammation and even other biomedical applications. [ABSTRACT FROM AUTHOR]

Plant-derived extracellular vesicles (PDEVs) have recently emerged as a promising area of research due to their potential health benefits and biomedical applications. Produced by various plant species, these EVs contain diverse bioactive molecules, including proteins, lipids, and nucleic acids. Increasing in vitro and in vivo studies have shown that PDEVs have inherent pharmacological activities that affect cellular processes, exerting anti-inflammatory, antioxidant, and anticancer activities, which can potentially contribute to disease therapy and improve human health. Additionally, PDEVs have shown potential as efficient and biocompatible drug delivery vehicles in treating various diseases. However, while PDEVs serve as a potential rising star in modern healthy diets and biomedical applications, further research is needed to address their underlying knowledge gaps, especially the lack of standardized protocols for their isolation, identification, and large-scale production. Furthermore, the safety and efficacy of PDEVs in clinical applications must be thoroughly evaluated. In this review, we concisely discuss current knowledge in the PDEV field, including their characteristics, biomedical applications, and isolation methods, to provide an overview of the current state of PDEV research. Finally, we discuss the challenges regarding the current and prospective issues for PDEVs. This review is expected to provide new insights into healthy diets and biomedical applications of vegetables and fruits, inspiring new advances in natural food-based science and technology.

Inflammatory bowel disease (IBD) encompasses a collection of idiopathic diseases characterized by chronic inflammation in the gastrointestinal (GI) tract. Patients diagnosed with IBD often experience necessitate long-term pharmacological interventions. Among the multitude of administration routes available for treating IBD, oral administration has gained significant popularity owing to its convenience and widespread utilization. In recent years, there has been extensive evaluation of the efficacy of orally administered herbal medicinal products and their extracts as a means of treating IBD. Consequently, substantial evidence has emerged, supporting their effectiveness in IBD treatment. This review aimed to provide a comprehensive summary of recent studies evaluating the effects of herbal medicinal products in the treatment of IBD. We delved into the regulatory role of these products in modulating immunity and maintaining the integrity of the intestinal epithelial barrier. Additionally, we examined their impact on antioxidant activity, anti-inflammatory properties, and the modulation of intestinal flora. By exploring these aspects, we aimed to emphasize the significant advantages associated with the use of oral herbal medicinal products in the treatment of IBD. Of particular note, this review introduced the concept of herbal plant-derived exosome-like nanoparticles (PDENs) as the active ingredient in herbal medicinal products for the treatment of IBD. The inclusion of PDENs offers distinct advantages, including enhanced tissue penetration and improved physical and chemical stability. These unique attributes not only demonstrate the potential of PDENs but also pave the way for the modernization of herbal medicinal products in IBD treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

Extracellular vesicles (EVs) are phospholipidic bi-layer enclosed nanoparticles secreted naturally by all cell types. They are attracting increasing attention in the fields of nanomedicine, nutraceutics and cosmetics as biocompatible carriers for drug delivery, with intrinsic properties beneficial to human health. Scientific work now focuses on developing techniques for isolating EVs that can translate into industrial-scale production and meet rigorous clinical requirements. The science of EVs is ongoing, and many pitfalls must be addressed, such as the requirement for standard, reproducible, inexpensive, and Good Manufacturing Practices (GMP) adherent EV processing techniques. Researchers are exploring the use of alternative sources to EVs derived from mammalian cultures, such as plant EVs, as well as the use of bacteria, algae and milk. Regarding the downstream processing of EVs, many alternative techniques to the ultracentrifugation (UC) protocols most commonly used in the laboratory are emerging. In the context of process scale-up, membrane-based processes for isolation and purification of EVs are the most promising, either as stand-alone processes or in combination with chromatographic techniques. This review discusses current trends on EVs source selection and EVs downstream processing techniques, with a focus on plant-derived EVs and membrane-based techniques for EVs enrichment. • The isolation of extracellular vesicles (EVs) and their downstream processing is a challenge. • Plant, bacteria and animal are emerging as alternative sources for human-derived EVs. • Membrane-based processes are the most promising for large scale production of EVs. • Tangential flow filtration is a flexible isolation method for continuous production, • Microfluidic technologies have a great potential in the field of EV isolation. [ABSTRACT FROM AUTHOR]

Plant-derived extracellular vesicles (EVs) are capable of efficiency delivering mRNAs, miRNAs, bioactive lipids, and proteins to mammalian cells. Plant-derived EVs critically contribute to the ability of plants to defend against pathogen attacks at the plant cell surface. They also represent a novel candidate natural substance that shows potential to be developed for food, cosmetic, and pharmaceutical products. However, although plant-derived EVs are acknowledged as having potential for various industrial applications, little is known about how their stability is affected by storage conditions. In this study, we evaluated the stability of Dendropanax morbifera leaf-derived extracellular vesicles (LEVs) alone or combined with the preservatives, 1,3-butylene glycol (to yield LEVs-1,3-BG) or TMO (LEVs-TMO). We stored these formulations at −20, 4, 25, and 45 °C for up to 4 weeks, and compared the stability of fresh and stored LEVs. We also assessed the effect of freeze-thawing cycles on the quantity and morphology of the LEVs. We found that different storage temperatures and number of freeze-thawing cycles altered the stability, size distribution, protein content, surface charge, and cellular uptake of LEVs compared to those of freshly isolated LEVs. LEVs-TMO showed higher stability when stored at 4 °C, compared to LEVs and LEVs-1,3-BG. Our study provides comprehensive information on how storage conditions affect LEVs and suggests that the potential industrial applications of plant-derived EVs may be broadened by the use of preservatives. [ABSTRACT FROM AUTHOR]

The scientific interest in the beneficial properties of natural substances has been recognized for decades, as well as the growing attention in extracellular vesicles (EVs) released by different organisms, in particular from animal cells. However, there is increasing interest in the isolation and biological and functional characterization of these lipoproteic structures in the plant kingdom. Similar to animal vesicles, these plant-derived extracellular vesicles (PDEVs) exhibit a complex content of small RNAs, proteins, lipids, and other metabolites. This sophisticated composition enables PDEVs to be therapeutically attractive. In this review, we report and discuss current knowledge on PDEVs in terms of isolation, characterization of their content, biological properties, and potential use as drug delivery systems. In conclusion, we outline controversial issues on which the scientific community shall focus the attention shortly. [ABSTRACT FROM AUTHOR]

The development of effective nanosystems for drug delivery represents a key challenge for the improvement of most current anticancer therapies. Recent progress in the understanding of structure and function of extracellular vesicles (EVs)—specialized membrane-bound nanocarriers for intercellular communication—suggests that they might also serve as optimal delivery systems of therapeutics. In addition to carrying proteins, lipids, DNA and different forms of RNAs, EVs can be engineered to deliver specific bioactive molecules to target cells. Exploitation of their molecular composition and physical properties, together with improvement in bio-techniques to modify their content are critical issues to target them to specific cells/tissues/organs. Here, we will discuss the current developments in the field of animal and plant-derived EVs toward their potential use for delivery of therapeutic agents in different pathological conditions, with a special focus on cancer. [ABSTRACT FROM AUTHOR]