Cervical cancer (CC) remains a critical public health issue, highlighting the importance of early detection. However, current methods such as cytological and HPV testing face challenges of invasiveness and low patient compliance. Exosomes, emerging as crucial in cancer diagnosis, offer promise due to their noninvasive, highly specificity, and abundant biomarkers. However, isolating exosomes efficiently remains challenging. In this study, we designed and synthesized a bifunctional affinity nanomaterial Fe3O4 @CD63-CLIKKPF, based on the synergistic interaction between its modified aptamer CD63 and peptide CLIKKPF, and CD63 protein and PS of exosomes which can achieve high specificity and high yield separation of urinary exosomes. Notably, the co-modified aptamer CD63 and peptide CLIKKPF not only enable efficient exosome isolation by leveraging dual-affinity mechanisms through a synergistic "AND" logic analysis, but also could be achieved on the Fe3O4 in one-step reaction at room temperature via Fe-S bonding. Combined with LC-MS/MS, we conducted exosome metabolomics analysis in healthy individuals and CC patients across various stages, and machine learning models demonstrated accurate classification (accuracy >0.822) and prediction capabilities for CC. Furthermore, six key metabolites indicative of CC progression were identified and validated in additional patient samples, highlighting their potential as biomarkers. Overall, this study establishes a novel method for exosome metabolomics in CC, offering insights for non-invasive early diagnosis and progression prediction on a large scale.

In the recent decade, extracellular vesicles (EVs) have been released from nearly all the kingdoms, modulating intercellular communication and maintaining the human body's homeostasis by regulating different cellular processes. Among EVs, exosomes are the emerging field in biopharmaceuticals. They have lipid bilayer ranging from 30 to 150 nm in size and encompass DNA, RNA, protein lipids, etc. Their sources are widespread, easy to acquire, and cost-effective in manufacturing. This review focuses on the detailed classification of exosomes existing in nature, knowledge and application of omics, therapeutic, diagnostic and theranostic application of exosomes. It covers diseases such as cancer, infectious diseases (viral, bacterial, fungal infections), neurodegenerative diseases, metabolic diseases, lifestyle diseases (diabetes, cardiovascular, gastric disorder (IBD)), autoimmune disorders and their biodistribution. This article unfolds the recent progress in the exosomes arena and covers all the regulatory considerations (FDA, EMA, and other nations) involved with it. Moreover, a detailed discussion about clinical trials and its manifestation with exosomes and challenges associated with their isolation procedures, reproducibility, and safety concerns.

Hypoxic-ischemic (HI) brain damage poses a high risk of death or lifelong disability, yet effective treatments remain elusive. Here, we demonstrated that miR-100-5p levels in the lesioned cortex increased after HI insult in neonatal mice. Knockdown of miR-100-5p expression in the brain attenuated brain injury and promoted functional recovery, through inhibiting the cleaved-caspase-3 level, microglia activation, and the release of proinflammation cytokines following HI injury. Engineered extracellular vesicles (EVs) containing neuron-targeting rabies virus glycoprotein (RVG) and miR-100-5p antagonists (RVG-EVs-Antagomir) selectively targeted brain lesions and reduced miR-100-5p levels after intranasal delivery. Both pre- and post-HI administration showed therapeutic benefits. Mechanistically, we identified protein phosphatase 3 catalytic subunit alpha (Ppp3ca) as a novel candidate target gene of miR-100-5p, inhibiting c-Fos expression and neuronal apoptosis following HI insult. In conclusion, our non-invasive method using engineered EVs to deliver miR-100-5p antagomirs to the brain significantly improves functional recovery after HI injury by targeting Ppp3ca to suppress neuronal apoptosis.

Reproductive disorders, including preeclampsia (PE), endometriosis, premature ovarian failure (POF), and polycystic ovary syndrome (PCOS), present substantial challenges to women's reproductive health. Exosomes (EXOs) are cell-derived vesicles containing molecules that influence target cells' gene expression and cellular behavior. Among their cargo, microRNAs (miRNAs)-short, non-coding RNAs typically 19-25 nucleotides in length-play a crucial role in post-transcriptional gene regulation and have been extensively studied for their therapeutic potential. miRNAs are considered therapeutic targets because they regulate key cellular pathways such as proliferation, apoptosis, angiogenesis, and tissue repair. This review examines the role of exosomal miRNAs from sources such as mesenchymal stem cells (MSCs), plasma, and amniotic fluid in female reproductive disorders, including PE, POF, PCOS, and endometriosis. We discuss their biological origins, mechanisms of miRNA sorting and packaging, and their therapeutic applications in modulating disease progression. By categorizing miRNAs according to their beneficial or detrimental effects in specific conditions, we aim to simplify the understanding of their roles in female infertility.

Extracellular vesicles (EVs) are emerging as critical mediators of intercellular communication in the tumor microenvironment (TME), profoundly influencing cancer progression. These nano-sized vesicles, released by both tumor and stromal cells, carry a diverse cargo of proteins, nucleic acids, and lipids, reflecting the dynamic cellular landscape and mediating intricate interactions between cells. This review provides a comprehensive overview of the biogenesis, composition, and functional roles of EVs in cancer, highlighting their significance in both basic research and clinical applications. We discuss how cancer cells manipulate EV biogenesis pathways to produce vesicles enriched with pro-tumorigenic molecules, explore the specific contributions of EVs to key hallmarks of cancer, such as angiogenesis, metastasis, and immune evasion, emphasizing their role in shaping TME and driving therapeutic resistance. Concurrently, we submit recent knowledge on how the cargo of EVs can serve as a valuable source of biomarkers for minimally invasive liquid biopsies, and its therapeutic potential, particularly as targeted drug delivery vehicles and immunomodulatory agents, showcasing their promise for enhancing the efficacy and safety of cancer treatments. By deciphering the intricate messages carried by EVs, we can gain a deeper understanding of cancer biology and develop more effective strategies for early detection, targeted therapy, and immunotherapy, paving the way for a new era of personalized and precise cancer medicine with the potential to significantly improve patient outcomes.

Myocardial infarction (MI) with resulting congestive heart failure is one of the leading causes of death worldwide. Current therapies for treating MI, such as devices, traditional medicine, and surgeries, come with many limitations as patients in their final stages of heart failure have little chances of experiencing any reversible changes. In recent decades, Mesenchymal stem cell (MSC) based therapy has become one of the most popular and rapidly developing fields in treating MI. Their supremacy for clinical applications is partially due to their unique properties and encouraging pre-clinical outcomes in various animal disease models. However, the majority of clinical trials registered for MSC therapy for diverse human diseases, including MI, have fallen short of expectations. This review intends to discuss the recent advances in the clinical application of using MSCs for cardiac repair and discuss challenges facing the clinical translation of MSCs for cardiac regeneration such as restoration of endothelial-cardiomyocyte crosstalk, immunomodulation and immune rejection, poor homing and migration, as well as low retention and survival. Furthermore, we will discuss recent strategies being investigated to help overcome some of these challenges.

Neurodegenerative diseases (NDs) like Alzheimer's, Parkinson's, and ALS rank among the most challenging global health issues, marked by substantial obstacles in early diagnosis and effective treatment. Current diagnostic techniques frequently demonstrate inadequate sensitivity and specificity, whilst conventional treatment strategies encounter challenges related to restricted bioavailability and insufficient blood-brain barrier (BBB) permeability. Recently, exosomes-nanoscale vesicles packed with proteins, RNAs, and lipids-have emerged as promising agents with the potential to reshape diagnostic and therapeutic approaches to these diseases. Unlike conventional drug carriers, they naturally traverse the BBB and can deliver bioactive molecules to affected neural cells. Their molecular cargo can influence cell signaling, reduce neuroinflammation, and potentially slow neurodegenerative progression. Moreover, exosomes serve as non-invasive biomarkers, enabling early and precise diagnosis while allowing real-time disease monitoring. Additionally, engineered exosomes, loaded with therapeutic molecules, enhance this capability by targeting diseased neurons and overcoming conventional treatment barriers. By offering enhanced specificity, reduced immunogenicity, and an ability to bypass physiological limitations, exosome-based strategies present a transformative advantage over existing diagnostic and therapeutic approaches. This review examines the multifaceted role of exosomes in NDDs, emphasizing their diagnostic capabilities, intrinsic therapeutic functions, and transformative potential as advanced treatment vehicles.

Exercise as a non-pharmacological intervention can exert beneficial effects directly through exosomes crossing the blood-brain barrier and reduce apoptosis after cerebral ischaemia/reperfusion injury (CI/RI). miRNA-124 (miR-124) is present in exosomes and plays an important role in regulating cerebral neurological activity; however, the mechanism of the relationship between exercise and the activity of exosomes and apoptosis after CI/RI remains unclear. Therefore, the present study investigated the effects of exercise preconditioning on CI/RI from the perspective of exosomal miR-124 and apoptosis.

The middle cerebral artery occlusion/reperfusion (MCAO/R) model was established by blocking the middle cerebral artery, and a motorized running wheel was chosen as the method of exercise preconditioning for rats, the morphology, particle concentration and particle size distribution of the exosome samples were identified at the 6 h, 12 h, and 24 h time points. RT-PCR, western blotting, immunohistochemistry, TUNEL staining, TTC staining and mNSS scores were used to investigate the effects of exercise preconditioning on apoptosis in MCAO/R rats.

The results showed exercise reduced neurological dysfunction and infarct size, increased the content of plasma exocrine miR-124 at 24 h, which inhibited the expression of STAT3, increased the expression of the anti-apoptotic BCL-2, and decreased the expression of the pro-apoptotic BAX, thereby reducing apoptosis.

Our findings indicated that exercise preconditioning can enhance the anti-apoptotic capacity of tissues in the rat ischemic penumbra and reduce apoptosis after CI/RI via the exosomal miR-124, STAT3, BCL-2/BAX pathway.

Small extracellular vesicles (sEVs), including exosomes as a subtype, with a diameter typically less than 200 nm and originating from the endosomal system, are capable of transporting a diverse array of bioactive molecules, including proteins, nucleic acids, and lipids, thereby facilitating intercellular communication and modulating cellular functions. Vascular dementia (VaD) represents a form of cognitive impairment attributed to cerebrovascular disease, characterized by a complex and multifaceted pathophysiological mechanism. Currently, the therapeutic approach to VaD predominantly emphasizes symptom management, as no specific pharmacological treatment exists to cure the condition. Recent investigations have illuminated the significant role of sEVs in the pathogenesis of vascular dementia. This review seeks to provide a comprehensive analysis of the characteristics and functions of sEVs, with a particular focus on their involvement in vascular dementia and its underlying mechanisms. The objective is to advance the understanding of the interplays between sEVs and vascular dementia, thereby offering novel insights for future research and therapeutic strategies.

Extracellular vesicles (EVs) are membrane vesicles secreted by all types of cells, including bacteria, animals, and plants. These vesicles contain proteins, nucleic acids, and lipids from their parent cells and can transfer these components between cells. EVs have attracted attention for their potential use in diagnosis and therapy due to their natural properties, such as low immunogenicity, high biocompatibility, and ability to cross the blood-brain barrier. They can also be engineered to carry therapeutic molecules. EVs can be delivered via various routes. The intranasal route is particularly advantageous for delivering them to the central nervous system, making it a promising approach for treating neurological disorders.

This review delves into the promising potential of intranasally administered EVs-based therapies for various medical conditions, with a particular focus on those affecting the brain and central nervous system. Additionally, the potential use of these therapies for pulmonary conditions, cancer, and allergies is examined, offering a hopeful outlook for the future of medical treatments.

The intranasal administration of EVs offers significant advantages over other delivery methods. By directly delivering EVs to the brain, specifically targeting areas that have been injured, this administration proves to be highly efficient and effective, providing reassurance about the progress in medical treatments. Intranasal delivery is not limited to brain-related conditions. It can also benefit other organs like the lungs and stimulate a mucosal immune response against various pathogens due to the highly vascularized nature of the nasal cavity and airways. Moreover, it has the added benefit of minimizing toxicity to non-targeted organs and allows the EVs to remain longer in the body. As a result, there is a growing emphasis on conducting clinical trials for intranasal administration of EVs, particularly in treating respiratory tract pathologies such as coronavirus disease.

Exosomes (Exos), nanosized membranous vesicles (30-160 nm), have been validated as an effective drug delivery system capable of traversing biological barriers. Mesenchymal stem cells (MSCs), due to their near-limitless self-renewal capabilities, provide a plentiful source of exosomes for clinical applications. In this study, we utilized an exosome-encapsulated rapamycin (Exo-Rapa) delivery strategy, which permits the use of smaller drug dosages to achieve effects typically seen with higher dosages, thus enhancing drug efficacy. Moreover, Exos can transport pharmaceuticals across the blood-brain barrier (BBB) to the brain, and further penetrate GL261 cells to exert their effects. Within the tumor microenvironment, Exo-Rapa is released more rapidly and efficiently at the tumor site. The acidic conditions in tumors accelerate the release of Exo-Rapa, a characteristic that may make it a promising targeted therapeutic in future cancer research. Additionally, a series of in vivo experiments have further demonstrated the permeability of Exo-Rapa across the BBB, enabling it to accumulate at tumor sites; it also ameliorates inflammatory responses in Glioblastoma multiforme (GBM) mouse models and enhances anti-tumor activity through the regulation of angiogenesis via the VEGF/VEGFRs axis. Our results indicate that MSC-derived exosomes are a potent therapeutic carrier for GBM, offering an effective strategy for enhancing drug delivery across the BBB and providing a scientific foundation for the use of exosomes in the treatment of GBM and other diseases.

This altmetric analysis aimed to identify and describe the top 100 papers on prostate cancer-associated exosomes in the Middle East and North Africa (MENA) region cited on social media.

As an applied study with an altmetric approach, this research included all Science Citation Index (SCI) Expanded indexed papers on prostate cancer-associated exosomes in the MENA region during 1970-2023. Altmetric Attention Scores (AASs) were extracted from the Altmetric Explorer, and Excel and SPSS were used for data analysis.

Twitter ranked first in mentioning 73.55 of the top 100 studied papers. The highest score of mentions on Twitter equaled 187, and that of AAS was 516, which belonged to an original research article. However, the top paper in citation counts was a guideline (AAS = 116; citation count = 5,664 =). The Journal of Urology published most papers (n = 21), with total AAS = 1,094. Most papers were international collaborations (n = 82). There was no significant relationship between the AASs of papers and those of Web of Science (WoS) citation counts (R2 = 0.1284, p-value = 0.2054).

Showing a broad perspective on the research priorities and new directions in prostate cancer-associated exosomes, this study can be a guideline for finding main papers on diagnosing, treating, and preventing prostate cancer. It helps researchers, professionals, and policymakers in developing the use of social media in disseminating related information.

By providing helpful information on prostate cancer-associated exosomes, this study can inform researchers and administrators of the state of research on the topic and consequent health promotion among the public.

A serum-free conditioning period is a crucial step during small extracellular vesicle (sEV) preparation ranging from 12 to 72h. There is a paucity of knowledge about downstream effects of serum-free conditioning on sEVs and the optimal duration of the conditioning period. The aim of this study was to investigate the influence of the serum-free conditioning period on the sEVs derived from primary adipose stromal cells (AdSCs) and their regenerative potential. Primary AdSCs were conditioned in serum-free medium for 72h. Conditioned medium was collected and refreshed every 24h obtaining three fractions, namely sEVs released after 24h (early), 24h to 48h (intermediate) and 48h to 72h (late). After sEV enrichment with ultracentrifugation, the sEV fractions were analyzed by their size, phenotypic expression, and morphology. Proliferation assays of primary Schwann cells after treatment with sEVs were performed. Particles meeting criteria to be classified as sEVs were detected in all fractions. However, sEVs differed by their size and phenotypic expression. A long conditioning period led to a heterogenous population of larger sEVs and increased protein per particle ratio. Moreover, the expression of tetraspanines was affected. Lastly, the proliferative effect of sEVs on Schwann cells decreased with increasing conditioning period. In conclusion, particles meeting the criteria of EVs are released by primary AdSCs over 72h under serum free conditioning. Nonetheless, they significantly differ in their proliferative effect on Schwann cells cultures.

Extracellular vesicles (EVs) have gained traction as potential cell-free therapeutic candidates. Development of purification methods that are scalable and robust is a major focus of EV research. Yet there is still little in the literature that evaluates purification methods against potency of the EV product. In the present study, we examined two monolith chromatography methods with a focus on assessing the ability of purified EVs to retain stimulatory effects on fibroblasts to connect scalable purification methods with product outputs.

We characterized EVs recovered from CTX0E03 (CTX) neural stem cell-conditioned medium in terms of biomarker distribution, functional capacity and purity. We evaluated the ability of EVs to promote wound closure in an in vitro scratch assay prior to and following two monolith chromatography steps (anion exchange and hydrophobic interaction) to determine whether these options may better serve EV bioprocessing.

EVs from CTX cells were successful in initiating wound repair in a fibroblast scratch assay over 72 h with a single 20-μg dose. EV preparations presented the markers CD9, CD81 and CD63 but also contained culture albumin and DNA as process impurities. EVs recovered by tangential flow filtration could be successfully purified further by both monolith chromatography steps. Post-monolith EV stimulation was conserved.

The results indicate that monolith chromatography is a viable purification method for EVs derived from cell culture that does not detract from the product's ability to stimulate fibroblasts, suggesting that product functionality is conserved. Further work is needed in developing suitable downstream processes and analytics to achieve clinically relevant purities for injectable biologics.

Deer antler blastema progenitor cells (ABPCs) are promising for regenerative medicine due to their role in annual antler regeneration, the only case of complete organ regeneration in mammals. ABPC-derived signals show great potential for promoting regeneration in tissues with limited natural regenerative ability. Our findings demonstrate the capability of extracellular vesicles from ABPCs (EVsABPC) to repair spinal cord injury (SCI), a condition with low regenerative capacity. EVsABPC significantly enhanced the proliferation of neural stem cells (NSCs) and activated neuronal regenerative potential, resulting in a 5.2-fold increase in axonal length. Additionally, EVsABPC exhibited immunomodulatory effects, shifting macrophages from M1 to M2. Engineered with activated cell-penetrating peptides (ACPPs), EVsABPC significantly outperformed EVs from rat bone marrow stem cells (EVsBMSC) and neural stem cells (EVsNSC), promoting a 1.3-fold increase in axonal growth, a 30.6% reduction in neuronal apoptosis, and a 2.6-fold improvement in motor function recovery. These findings support ABPC-derived EVs as a promising therapeutic candidate for SCI repair.

Osteoarthritis (OA) presents a significant therapeutic challenge, with few options for preserving joint cartilage and repairing associated tissue damage. Inflammation is a pivotal factor in OA-induced cartilage deterioration and synovial inflammation. Recently, exosomes derived from human umbilical cord mesenchymal stem cells (HucMSCs) have gained recognition as a promising noncellular therapeutic modality, but their use is hindered by the challenge of harvesting a sufficient number of exosomes with effective therapeutic efficacy. Given that HucMSCs are highly sensitive to microenvironmental signals, we hypothesized that priming HucMSCs within a proinflammatory environment would increase the number of exosomes secreted with enhanced anti-inflammatory properties. Subsequent miRNA profiling and pathway analysis confirmed that interleukin-1 beta (IL-1β)-induced exosomes (C-Exos) exert positive effects through miRNA regulation and signaling pathway modulation. In vitro experiments revealed that C-Exos enhance chondrocyte functionality and cartilage matrix production, as well as macrophage polarization, thereby enhancing cartilage repair. C-Exos were encapsulated in hyaluronic acid hydrogel microspheres (HMs) to ensure sustained release, leading to substantial improvements in the inflammatory microenvironment and cartilage regeneration in a rat OA model. This study outlines a strategy to tailor exosome cargo for anti-inflammatory and cartilage regenerative purposes, with the functionalized HMs demonstrating potential for OA treatment.

Recently, the various regulative functions of long non-coding RNAs (LncRNAs) have been well determined. Recently, the vital role of LncRNAs as gene regulators has been identified in the immune system, especially in the inflammatory response. All cells of the immune system are governed by a complex and ever-changing gene expression program that is regulated through both transcriptional and post-transcriptional processes. LncRNAs regulate gene expression within the cell nucleus by influencing transcription or through post-transcriptional processes that affect the splicing, stability, or translation of messenger RNAs (mRNAs). Recent studies in immunology have revealed substantial alterations in the expression of lncRNAs during the activation of the innate immune system as well as the development, differentiation, and activation of T cells. These lncRNAs regulate key aspects of immune function, including the manufacturing of inflammatory molecules, cellular distinction, and cell movement. They do this by modulating protein-protein interactions or through base pairing with RNA and DNA. Here we review the current understanding of the mechanism of action of lncRNAs as novel immune-related regulators and their impact on physiological and pathological processes related to the immune system, including autoimmune diseases. We also highlight the emerging pattern of gene expression control in important research areas at the intersection between immunology and lncRNA biology.

Regenerative medicine endeavors to restore damaged tissues and organs utilizing biological approaches. Utilizing biomaterials to target and regulate the pathophysiological processes of injured tissues stands as a crucial method in propelling this field forward. The Extracellular Vesicles-in-Hydrogel (EViH) system amalgamates the advantages of extracellular vesicles (EVs) and hydrogels, rendering it a prominent biomaterial in regenerative medicine with substantial potential for clinical translation. This review elucidates the development and benefits of the EViH system in tissue regeneration, emphasizing the interaction and impact of EVs and hydrogels. Furthermore, it succinctly outlines the pathophysiological characteristics of various types of tissue injuries such as wounds, bone and cartilage injuries, cardiovascular diseases, nerve injuries, as well as liver and kidney injuries, underscoring how EViH systems target these processes to address related tissue damage. Lastly, it explores the challenges and prospects in further advancing EViH-based tissue regeneration, aiming to impart a comprehensive understanding of EViH. The objective is to furnish a thorough overview of EViH in enhancing regenerative medicine applications and to inspire researchers to devise innovative tissue engineering materials for regenerative medicine.

Extracellular membrane vesicles (EVs) offer promising values in various medical fields, e.g., as biomarkers in liquid biopsies or as native (or bioengineered) biological nanocarriers in tissue engineering, regenerative medicine and cancer therapy. Based on their cellular origin EVs can vary considerably in composition and diameter. Cell biological studies on mammalian prominin-1, a cholesterol-binding membrane glycoprotein, have helped to reveal new donor membranes as sources of EVs. For instance, small EVs can originate from microvilli and primary cilia, while large EVs might be produced by transient structures such as retracting cellular extremities of cancer cells during the mitotic rounding process, and the midbody at the end of cytokinesis. Here, we will highlight the various subcellular origins of prominin-1+ EVs, also called prominosomes, and the potential mechanism(s) regulating their formation. We will further discuss the molecular and cellular characteristics of prominin-1, notably those that have a direct effect on the release of prominin-1+ EVs, a process that might be directly implicated in donor cell reprogramming of stem and cancer stem cells. Prominin-1+ EVs also mediate intercellular communication during embryonic development and adult homeostasis in healthy individuals, while disseminating biological information during diseases.

Ischemic stroke and acute lung injury are prevalent life-threatening conditions marked by intricate molecular mechanisms and elevated mortality rates. Despite evident pathophysiological distinctions, a notable similarity exists in the gene responses to tissue injury observed in both pathologies. This similarity extends to both protein-encoding RNAs and non-coding RNAs. Extracellular vesicles (EVs) are nano-scale vesicles derived through cell secretion, possessing unique advantages such as high biocompatibility, low immunogenicity, intrinsic cell targeting, and facile chemical and genetic manipulation. Importantly, miRNAs, the most prevalent non-coding RNAs, are selectively concentrated within EVs. Macrophages/microglia serve as immune defense and homeostatic cells, deriving from progenitor cells in the bone marrow. They can be classified into two contrasting types: classical proinflammatory M1 phenotype or alternative anti-inflammatory M2 phenotype. However, there exists a continuum of various intermediate phenotypes between M1 and M2, and macrophages/microglia can transition from one phenotype to another. This review will investigate recent discoveries concerning the impact of EVs derived from macrophages/microglia under various states on the progression of ischemic stroke and acute lung injury. The focus will be on the involvement of miRNAs within these vesicles. The concluding remarks of this review will underscore the clinical possibilities linked to EV-miRNAs, accentuating their potential as both biomarkers and therapeutic targets.

Extracellular vesicles (EVs) are lipid bilayer particles released by virtually all cells, with prominent roles in both physiological and pathological processes. The size, number, and molecular composition of released EVs correlate to the cells of origin, modulated by the cell's environment and pathologic state. The proteins, DNA, RNA, and protein cargo carried by EVs are protected by degradation, with a prominent role in targeted intercellular signaling. These properties make EVs salient targets as both carriers of biomarkers and potential therapeutic delivery vehicles. The majority of EV research has focused on blood, urine, saliva, and cerebrospinal fluid due to easy accessibility. EVs have also been identified and studied in all ocular biofluids, including the vitreous humor, the aqueous humor, and the tear film, and the study of EVs in ocular disease is a new, promising, and underexplored direction with unique challenges and considerations. This review covers recent advances in the diagnostic and therapeutic use of ocular EVs, with a focus on human applications and key preceding in vitro and in vivo animal studies. We also discuss future directions based on the study of EVs in other organ systems and disease sates.

Osteoarthritis (OA) is a joint disease characterized by articular cartilage degradation. Persistent low-grade inflammation defines OA pathogenesis, with crucial involvement of pro-inflammatory M1-like macrophages. While mesenchymal stromal cells (MSC) and their small extracellular vesicles (sEV) hold promise for OA treatment, achieving consistent clinical-grade sEV products remains a significant challenge. This study aims to develop fully characterized, reproducible, clinical-grade batches of sEV derived from umbilical cord (UC)-MSC for the treatment of OA while assessing its efficacy and safety. Initially, a standardized, research-grade manufacturing protocol was established to ensure consistent sEV production. UC-MSC-sEV characterization under non-cGMP conditions showed consistent miRNA and protein profiles, suggesting their potential for standardized manufacturing. In vitro studies evaluated the efficacy, safety, and potency of sEV; animal studies confirmed their effectiveness and safety. In vitro, UC-MSC-sEV polarized macrophages to an anti-inflammatory M2b-like phenotype, through STAT1 modulation, indicating their potential to create an anti-inflammatory environment in the affected joints. In silico studies confirmed sEV's immunosuppressive signature through miRNA and proteome analysis. In an OA mouse model, sEV injected intra-articularly (IA) induced hyaline cartilage regeneration, validated by histological and μCT analyses. The unique detection of sEV signals within the knee joint over time highlights its safety profile by confirming the retention of sEV in the joint. The product development of UC-MSC-sEV involved refining, standardizing, and validating processes in compliance with GMP standards. The initial assessment of the safety of the clinical-grade product via IA administration in a first-in-human study showed no adverse effects after a 12 month follow-up period. These results support the progress of this sEV-based therapy in an early-phase clinical trial, the details of which are presented and discussed in this work. This study provides data on using UC-MSC-sEV as local therapy for OA, highlighting their regenerative and anti-inflammatory properties and safety in preclinical and a proof-of-principle clinical application.

We aimed to explore the therapeutic efficacy of miR-7704-modified extracellular vesicles (EVs) derived from human umbilical cord mesenchymal stem cells (HUCMSCs) for osteoarthritis (OA) treatment. In vitro experiments demonstrated the successful transfection of miR-7704 into HUCMSCs and the isolation of EVs from these cells. In vivo experiments used an OA mouse model to assess the effects of the injection of miR-7704-modified EVs intra-articularly. Walking capacity (rotarod test), cartilage morphology, histological scores, and the expression of type II collagen, aggrecan, interleukin-1 beta, and matrix metalloproteinase 13 (MMP13) in the cartilage were evaluated. The EVs were characterized to confirm their suitability for therapeutic use. IL-1beta-treated chondrocytes increased type II collagen and decreased MMP13 after treatment with miR-7704-overexpressed EVs. In vivo experiments revealed that an intra-articular injection of miR-7704-overexpressed EVs significantly improved walking capacity, preserved cartilage morphology, and resulted in higher histological scores compared to in the controls. Furthermore, the decreased expression of MMP13 in the cartilage post treatment suggests a potential mechanism for the observed therapeutic effects. Therefore, miR-7704-overexpressed EVs derived from HUCMSCs showed potential as an innovative therapeutic strategy for treating OA. Further investigations should focus on optimizing dosage, understanding mechanisms, ensuring safety and efficacy, developing advanced delivery systems, and conducting early-phase clinical trials to establish the therapeutic potential of HUCMSC-derived EVs for OA management.

Extracellular vesicle (EV) monitoring can complement clinical assessment of cancer response. In this study, patients with advanced non-small cell lung cancer (NSCLC) undergoing osimertinib, alectinib, pembrolizumab or platinum-based chemotherapy ± pembrolizumab were enrolled. EVs were characterized using Bradford assay to quantify the circulating cell-free EV protein content (cfEV), and dynamic light scattering to assess Rayleigh ratio excess at 90°, z-averaged hydrodynamic diameter and polydispersity index. A total of 135 plasma samples from 27 patients were collected at baseline (T0) and at the first radiological restaging (T1). A ∆cfEV < 20% was associated with improved median progression-free survival (mPFS) in responders versus non-responders. Specifically, cfEV responders on pembrolizumab had a significantly better mPFS (25.2 months) compared to those on chemotherapy plus pembrolizumab (6.1 months). EGFR-positive cfEV responders also experienced longer mPFS compared to cfEV non-responders (35.1 months, 95% CI: 14.9-35.5 vs. 20.8 months, 95% CI: 11.2-30.4). This study suggested that monitoring circulating EV could provide valuable insights into treatment efficacy in NSCLC, particularly for patients receiving pembrolizumab or osimertinib.

Colorectal cancer (CRC) continues to be a major worldwide health issue, with elevated death rates linked to late stages of the illness. Immunotherapy has made significant progress in developing effective techniques to improve the immune system's capacity to identify and eradicate cancerous cells. This study examines the most recent advancements in CAR-T cell treatment and exosome-based immunotherapy for CRC. CAR-T cell therapy, although effective in treating blood cancers, encounters obstacles when used against solid tumours such as CRC. These obstacles include the presence of an immunosuppressive tumour microenvironment and a scarcity of tumour-specific antigens. Nevertheless, novel strategies like dual-receptor CAR-T cells and combination therapy involving cytokines have demonstrated promise in surmounting these obstacles. Exosome-based immunotherapy is a promising approach for targeted delivery of therapeutic drugs to tumour cells, with high specificity and minimal off-target effects. However, there are still obstacles to overcome in the field, such as resistance to treatment, adverse effects associated with the immune system, and the necessity for more individualised methods. The current research is focused on enhancing these therapies, enhancing the results for patients, and ultimately incorporating these innovative immunotherapeutic approaches into the standard treatment protocols for CRC.

Nanotechnology-based delivery systems have brought a paradigm shift in the management of cancer. However, the main obstacles to nanocarrier-based delivery are their limited circulation duration, excessive immune clearance, inefficiency in interacting effectively in a biological context and overcoming biological barriers. This demands effective engineering of nanocarriers to achieve maximum efficacy. Nanocarriers can be maneuvered with biological components to acquire biological identity for further regulating their biodistribution and cell-to-cell cross-talk. Thus, the integration of synthetic and biological components to deliver therapeutic cargo is called a biohybrid delivery system. These delivery systems possess the advantage of synthetic nanocarriers, such as high drug loading, engineerable surface, reproducibility, adequate communication and immune evasion ability of biological constituents. The biohybrid delivery vectors offer an excellent opportunity to harness the synergistic properties of the best entities of the two worlds for improved therapeutic outputs. The major spotlights of this review are different biological components, synthetic counterparts of biohybrid nanocarriers, recent advances in hybridization techniques, and the design of biohybrid delivery systems for cancer therapy. Moreover, this review provides an overview of biohybrid systems with therapeutic and diagnostic applications. In a nutshell, this article summarizes the advantages and limitations of various biohybrid nano-platforms, their clinical potential and future directions for successful translation in cancer management.

Extracellular vesicles (EVs) are diverse nanoparticles with large heterogeneity in size and molecular composition. Although this heterogeneity provides high diagnostic value for liquid biopsy and confers many exploitable functions for therapeutic applications in cancer detection, wound healing and neurodegenerative and cardiovascular diseases, it has also impeded their clinical translation-hence heterogeneity acts as a double-edged sword. Here we review the impact of subpopulation heterogeneity on EV function and identify key cornerstones for addressing heterogeneity in the context of modern analytical platforms with single-particle resolution. We outline concrete steps towards the identification of key active biomolecules that determine EV mechanisms of action across different EV subtypes. We describe how such knowledge could accelerate EV-based therapies and engineering approaches for mimetic artificial nanovesicle formulations. This approach blunts one edge of the sword, leaving only a single razor-sharp edge on which EV heterogeneity can be exploited for therapeutic applications across many diseases.

Cell-based therapies are revolutionizing medicine by replacing or modifying dysfunctional cells with healthy cells or engineered derivatives, offering disease reversal and cure. One promising approach is using cell-derived extracellular vesicles (EVs), which offer therapeutic benefits similar to cell transplants without the biosafety risks. Although EV applications face challenges like limited production, inadequate therapeutic loading, and poor targeting efficiency, recent advances in bioengineering have enhanced their effectiveness. Herein, we summarize technological breakthroughs in EV bioengineering over the past 5 years, highlighting their improved therapeutic functionalities and potential clinical prospects. We also discuss biomanufacturing processes, regulation, and safety considerations for bioengineered EV therapies, emphasizing the significance of establishing robust frameworks to ensure translation capability, safety, and therapeutic effectiveness for successful clinical adoption.

The application of extracellular vesicles (EVs) as therapeutics or nanocarriers in cell-free therapies necessitates meticulous evaluations of different features, including their identity, bioactivity, batch-to-batch reproducibility, and stability. Given the inherent heterogeneity in EV preparations, this assessment demands sensitive functional assays to provide key quality control metrics, complementing established methods to ensure that EV preparations meet the required functionality and quality standards. Here, we introduce the detectEV assay, an enzymatic-based approach for assessing EV luminal cargo bioactivity and membrane integrity. This method is fast, cost-effective, and quantifiable through enzymatic units. Utilizing microalgae-derived EVs, known as nanoalgosomes, as model systems, we optimised the assay parameters and validated its sensitivity and specificity in quantifying the enzymatic activity of esterases within the EV lumen while also evaluating EV membrane integrity. Compared to conventional methods that assess physicochemical features of EVs, our single-step analysis efficiently detects batch-to-batch variations by evaluating changes in luminal cargo bioactivity and integrity across various EV samples, including differences under distinct storage conditions and following diverse isolation and exogenous loading methods, all using small sample sizes. The detectEV assay's application to various human-derived EV types demonstrated its versatility and potential universality. Additionally, the assay effectively predicted EV functionality, such as the antioxidant activity of different nanoalgosome batches. Our findings underscore the detectEV assay's utility in comprehensive characterization of EV functionality and integrity, enhancing batch-to-batch reproducibility and facilitating their therapeutic applications.

Mesenchymal stem cells (MSCs) are pluripotent stem cells with self-renewal. They play a critical role in cell therapy due to their powerful immunomodulatory and regenerative effects. Recent studies suggest that one of the key therapeutic mechanisms of MSCs seems to derive from their paracrine product, called extracellular vesicles (EVs). The EVs contain much DNA, messenger RNA (mRNA), microRNA, and protein components, which can exert intracellular communication to target cells. In clinical applications, the MSC-EVs have been widely used in tissue repair and immune disorder diseases. However, there are serval issues need to be considered such as how to accomplish the large-scale production of EVs and how to verify the exact mechanism of EVs. In this review, we summarize the current progress of MSC-EVs and discuss the challenges and future of MSC-EVs.

Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related mortality globally. HCC is highly heterogenous with diverse etiologies leading to different driver mutations potentiating unique tumor immune microenvironments. Current therapeutic options, including immune checkpoint inhibitors and combinations, have achieved limited objective response rates for the majority of patients. Thus, a precision medicine approach is needed to tailor specific treatment options for molecular subsets of HCC patients. Lipid nanovesicle platforms, either liposome- (synthetic) or extracellular vesicle (natural)-derived present are improved drug delivery vehicles which may be modified to contain specific cargos for targeting specific tumor sites, with a natural affinity for liver with limited toxicity. This mini-review provides updates on the applications of novel lipid nanovesicle-based therapeutics for HCC precision medicine and the challenges associated with translating this therapeutic subclass from preclinical models to the clinic.

Colorectal cancer (CRC) ranks as the third most common cancer worldwide and remains a major cause of cancer-related deaths, necessitating the development of innovative therapeutic approaches beyond conventional treatment modalities. Conventional therapies, such as radiation, chemotherapy, and surgery, are hindered by challenges like imprecise targeting, substantial toxicity, and the development of resistance. Exosome-driven nano-immunotherapy has emerged as a groundbreaking approach that leverages the natural properties of exosomes-cell-derived vesicles known for their role in intercellular communication-to deliver therapeutic agents with high precision and specificity. This approach utilizes the natural ability of exosomes to serve as natural nanocarriers for various biomolecules, such as proteins, nucleic acids, and lipids, enabling precise drug delivery and immune modulation. Exosomes offer distinct advantages compared to traditional drug delivery systems, including their biocompatibility, capability to traverse biological barriers, and suitability for personalized medicine approaches. We evaluate the effectiveness of exosome-based therapies in comparison to traditional approaches, emphasizing their ability to achieve precise delivery, minimize systemic toxicity, and enhance treatment results. Despite their promise, several challenges remain, including the standardization of exosome isolation and production, optimization of cargo loading techniques, and ensuring safety and efficacy in clinical applications. By overcoming these obstacles and leveraging the distinctive characteristics of exosomes, exosome-driven nano-immunotherapy presents a promising avenue for more efficient therapeutic interventions.

Heart transplantation is a crucial intervention for severe heart failure, yet the challenge of organ rejection is significant. Bone marrow mesenchymal stem cells (BMSCs) and their exosomes have demonstrated potential in modulating T cells, dendtitic cells (DCs), and cytokines to achieve immunomodulatory effects. DCs, as key antigen-presenting cells, play a critical role in shaping immune responses by influencing T-cell activation and cytokine production. Through this modulation, BMSCs and their exosomes enhance graft tolerance and prolonging survival.

To explore the immunomodulatory effects of exosomes derived from BMSCs overexpressing microRNA-540-3p (miR-540-3p) on cardiac allograft tolerance, focusing on how these exosomes modulating DCs and T cells activity through the CD74/nuclear factor-kappaB (NF-κB) pathway.

Rat models were used to assess the impact of miR-540-3p-enhanced exosomes on immune tolerance in cardiac allografts. MiR-540-3p expression was manipulated in BMSCs, and derived exosomes were collected and administered to the rat models post-heart transplantation. The study monitored expression levels of major histocompatibility complex II, CD80, CD86, and CD274 in DCs, and quantified CD4+ and CD8+ T cells, T regulatory cells, and cytokine profiles.

Exosomes from miR-540-3p-overexpressing BMSCs lead to reduced expression of immune activation markers CD74 and NF-κB p65 in DCs and T cells. Rats treated with these exosomes showed decreased inflammation and improved cardiac function, indicated by lower levels of pro-inflammatory cytokines (interleukin-1β, interferon-γ) and higher levels of anti-inflammatory cytokines (interleukin-10, transforming growth factor β1). Additionally, miR-540-3p skewed the profiles of DCs and T cells towards immune tolerance, increasing the ratio of T regulatory cells and shifting cytokine secretion to favor graft acceptance.

Exosomes derived from BMSCs overexpressing miR-540-3p significantly enhance immune tolerance and prolong cardiac allograft survival by modulating the CD74/NF-κB pathway, which regulates activities of DCs and T cells. These findings highlight a promising therapeutic strategy to improve heart transplantation outcomes and potentially reduce the need for prolonged immunosuppression.

Necrotizing enterocolitis (NEC) is a severe inflammatory and necrotizing disease of the intestine that primarily affects the neonates, particularly premature infants. It has a high incidence of approximately 8.9% in extremely preterm infants, with a mortality rate ranging from 20 to 30%. In recent years, exosomes, particularly those derived from breast milk, have emerged as potential candidates for NEC therapy. Human breast milk-derived exosomes (BME) have been shown to enhance intestinal barrier function, protect intestinal epithelial cells from oxidative stress, promote the proliferation and migration of intestinal epithelial cells, and reduce the severity of experimental NEC models. As a subset of extracellular vesicles, BME possess the membrane structure, low immunogenicity, and high permeability, making them ideal vehicles for the treatment of NEC. Additionally, exosomes derived from various sources, including stem cells, intestinal epithelial cells, plants, and bacteria, have been implicated in the development and protection of intestinal diseases. This article summarizes the mechanisms through which exosomes, particularly BME, exert their effects on NEC and discusses the feasibility and obstacles associated with this novel therapeutic strategy.

Extracellular vesicles (EVs) are nanosized particles secreted by cells that play crucial roles in intercellular communication, especially in the context of cardiovascular diseases (CVDs). These vesicles carry complex cargo, including proteins, lipids, and nucleic acids, that reflects the physiological or pathological state of their cells of origin. Multiomics analysis of cell-derived EVs has provided valuable insights into the molecular mechanisms underlying CVDs by identifying specific proteins and EV-bound targets involved in disease progression. Recent studies have demonstrated that engineered EVs, which are designed to carry specific therapeutic molecules or modified to enhance their targeting capabilities, hold promise for treating CVDs. Analysis of the EV proteome has been instrumental in identifying key proteins that can be targeted or modulated within these engineered vesicles. For example, proteins involved in inflammation, thrombosis, and cardiac remodeling have been identified as potential therapeutic targets. Furthermore, the engineering of EVs to increase their delivery to specific tissues, such as the myocardium, or to modulate their immunogenicity and therapeutic efficacy is an emerging area of research. By leveraging the insights gained from multiomics analyses, researchers are developing EV-based therapies that can selectively target pathological processes in CVDs, offering a novel and potentially more effective treatment strategy. This review integrates the core findings from EV multiomics analysis in the context of CVDs and highlights the potential of engineered EVs in therapeutic applications.

Cancer cells secrete extracellular vesicles (EV) encapsulating bioactive cargoes to facilitate inter-organ communication in vivo and are emerging as critical mediators of tumor progression and metastasis, a condition which is often accompanied by a dysregulated cholesterol metabolism. Whether EVs are involved in the control of cholesterol homeostasis during tumor metastasis is still undefined and warrant further investigation. Here, we find that breast cancer-derived exosomal miR-9-5p induces the expression of HMGCR and CH25H, two enzymes involved in cholesterol synthesis and the conversion of 25-hydroxycholesterol from cholesterol by targeting INSIG1, INSIG2 and ATF3 genes in the liver. Notably, in vivo miR-9-5p antagomir treatment and genetic CH25H ablation prevents tumor metastasis in a mouse model of breast cancer. Thus, our findings reveal the regulatory mechanism of tumor-derived miR-9-5p in liver metastasis by linking oxysterol metabolism and Kupffer cell polarization, shedding light on future applications for cancer diagnosis and treatment.

Extracellular vesicles (EVs) function as biomarkers and are crucial in cell communication and regulation, with therapeutic potential for intervertebral disc (IVD)-related low back pain (LBP). EV cargo is often affected by tissue health, which may affect the therapeutic potential. There is currently limited knowledge of how the cargo of IVD cell-derived EVs varies with tissue health and how differences in proteomic profile affect the predicted biological functions.

Our study purified EVs from human IVD cell conditioned media by size-exclusion chromatography. Nanoparticle tracking analysis was conducted to measure EV size and concentration. Transmission electron microscopy and Western blot were performed to examine EV structure and markers. Tandem mass tag-mass spectrometry was conducted to determine protein cargo.

Most EVs were exosomes and intermediate microvesicles with an increasing amount linked to disease progression. Of the proteins detected, 88.6% were shared across the non-degenerate, mildly-degenerate, and degenerate samples. GO and KEGG analyses revealed that cargo from the mildly-degenerate samples was the most distinct, with the proteins in high abundance strongly associated with extracellular matrix (ECM) organization and structure. Shared proteins, highly expressed in the non-degenerate and degenerate samples, showed strong associations with cell adhesion, ECM-receptor interaction, and vesicle-mediated transport, respectively.

Our findings indicate that EVs from IVD cells from tissue with different degrees of degeneration share a majority of the cargo proteins. However, the level of expression differs with degeneration grade. Cargo from the mildly-degenerate samples exhibits the most differences. A better understanding of changes in EV cargo in the degenerative process may provide novel information related to molecular mechanisms underlying IVD degeneration and suggest new potential treatment modalities for IVD-related LBP.