Cisplatin (DDP) resistance is a key factor hindering esophageal cancer (ESCA) treatment. Exosomes have been reported to confer resistance to DDP in various tumor cells. However, the effects of ESCA cell-derived exosomes and exosomal human antigen R (HuR) on DDP resistance in cancer cells have not been elucidated. In this study, isolated exosomes were identified by transmission electron microscopy, nanoparticle tracking analysis, and western blotting. CCK-8 and flow cytometry were employed to assess the functional role of exosomes in ESCA DDP-resistant cells and their parental cells. Bioinformatics analysis was performed to identify molecules that were positively associated with HuR and validated using dual-luciferase reporter analysis and RNA immunoprecipitation assays. We found that exosomes from ESCA cells enhance the resistance of drug-resistant cells to DDP. Importantly, HuR protein, but not mRNA, was directly transferred into DDP-resistant cells via exosomes, thereby increasing the level of HuR protein. Mechanistically, HuR positively correlated with Lamin B2 (LMNB2) in ESCA cells, and ESCA DDP-resistant cells transfected with siRNA targeting LMNB2 exhibited reduced cell viability and elevated apoptosis rates. Moreover, the role of ESCA cell-derived exosomes in the transmission of DDP resistance in vivo was validated using a nude mouse model. Collectively, our results revealed that exosomes exposed to ESCA cells induced greater drug resistance in DDP-resistant ESCA cells via HuR delivery. Targeting HuR or its positively related target LMNB2 may present new therapeutic opportunities for treating patients with DDP-resistant ESCA.

Exosomes are nanoscale extracellular vesicles (EVs) secreted by most eukaryotic cells and can be found in nearly all human body fluids. Increasing evidence has revealed their pivotal roles in intercellular communication, and their active participation in myriad physiological and pathological activities. Exosomes' functions rely on their contents that are closely correlated with the biological characteristics of parental cells, which may provide a rich resource of molecular information for accurate and detailed diagnosis of a diverse array of diseases, such as differential diagnosis of Alzheimer's disease, early detection and subtyping of various tumors. As a category of sensitive detection devices, biosensors can fully reveal the molecular information and convert them into actionable clinical information. In this review, recent advances in biosensor-based methods for the detection of exosomes are summarized. We have described the fabrication of various biosensors based on the analysis of exosomal proteins, RNAs or glycans for accurate diagnosis, with respect to their elaborate recognition designs, signal amplification strategies, sensing properties, as well as their application potential. The challenges along with corresponding technologies in the future development and clinical translation of these biosensors are also discussed.

Exosomes are small extracellular vesicles produced by most cell types and contain proteins, lipids, and nucleic acids (non-coding RNAs, mRNA, and DNA) that can be released by donor cells to influence the function of recipient cells. Skin photoaging is the premature aging of skin structures caused by prolonged exposure to ultraviolet (UV), as demonstrated by depigmentation, roughness, rhytides, elastosis, and precancerous alterations. Exosomes are associated with aging processes such as oxidative damage, inflammation, and senescence. Exosomes' anti-aging properties have been linked to various in vitro and preclinical investigations. There are still several unanswered questions about the use of MSC exosomes for skin rejuvenation, despite encouraging results. Uncertainty surrounds the precise processes by which exosomes stimulate the creation of collagen, skin tissue via a variety of mechanisms, including reduced matrix metalloproteinase (MMP) expression, increased collagen and elastin production, and modulation of intracellular signaling pathways and intercellular communication. These findings suggest the therapeutic potential of exosomes in skin aging. This review provides information on the molecular mechanisms and consequences of exosome anti-aging.

Oral submucosal fibrosis is a highly malignant oral condition that necessitates the use of sophisticated therapeutic procedures. OSF is a multifactorial precancerous condition induced by areca nut chewing, deficiencies in vitamins and trace minerals, immunological aspects, and hereditary factors. Adipose tissue-derived mesenchymal stem cells possess the capability for multidirectional activation and are extensively distributed throughout the body. They have minimal immunogenicity and are extensively utilized in cancer treatment. Exosomes are extracellular vesicles produced by the intracellular route. They are biological carriers comprising microRNA, messenger RNA, lipids and proteins crucial for intercellular communication. ADSC exosomes, serving as a vehicle for miRNA, possess accessibility and little immunogenicity. They can significantly contribute to adipose tissue regrowth, angiogenesis, immunological modulation, and tissue repair. ADSC-Exo exhibits antifibrotic properties and may serve as a potential treatment for OSF. This review presents a novel therapeutic approach and clarifies the precise mechanisms involved in the clinical management of OSF using ADSC-Exo.

Most eukaryotic cells maintain a large disparity in lipid compositions between the cytosolic and external leaflets of the plasma membrane (PM) bilayer. This lipid asymmetry is maintained by energy-consuming flippase enzymes that selectively drive phospholipids into the cytosolic leaflet, often against large concentration gradients. Scramblases, activated by intracellular Ca2+ or apoptotic signaling, shuttle phospholipids down their concentration gradient to release lipid asymmetry. Such scrambling is typically evidenced by exposure of phosphatidylserine (PS) to the external leaflet and is associated with many physiological processes, most notably blood clotting and cell death, but also activation of immune cells. Here, we show that both PS and phosphatidylethanolamine (PE) appear on the PM external leaflet following immune receptor-mediated activation of mast cells. We also observe similar effects in T cells. Importantly, in contrast to wholesale release of PM asymmetry induced by calcium ionophores or apoptosis, we show that scrambling in activated immune cells is focal, with small, stable regions of surface exposed PS. These scrambled foci are calcium dependent, have lower lipid packing than their surrounding outer leaflet, and are reversible. These observations of local, transient scrambling during physiological activation of healthy immune cells suggest important roles for the lateral and transbilayer organization of membrane lipids.

Rheumatoid arthritis (RA) is a systemic autoimmune disorder characterized by chronic inflammation and progressive damage to connective tissue. It is driven by dysregulated cellular homeostasis, often leading to autoimmune destruction and permanent disability in severe cases. Over the past decade, various drug delivery systems have been developed to enable targeted therapies for disease prevention, reduction, or suppression. As an emerging therapeutic platform, extracellular vesicles (EVs) offer several advantages over conventional drug delivery systems, including biocompatibility and low immunogenicity. Consequently, an increasing number of studies have explored EV-based delivery systems in the treatment of RA, leveraging their natural ability to evade phagocytosis, prolong in vivo half-life, and minimize the immunogenicity of therapeutic agents. In this review, we first provide an in-depth overview of the pathogenesis of RA and the current treatment landscape. We then discuss the classification and biological properties of EVs, their potential therapeutic mechanisms, and the latest advancements in EVs as drug delivery platforms for RA therapy. We emphasize the significance of EVs as carriers in RA treatment and their potential to revolutionize therapeutic strategies. Furthermore, we examine key technological innovations and the future trajectory of EV research, focusing on the challenges and opportunities in translating these platforms into clinical practice. Our discussion aims to offer a comprehensive understanding of the current state and future prospects of EV-based therapeutics in RA.

Extracellular vesicles (EVs) have emerged as promising biomarkers and therapeutic agents, yet their quantification remains technically challenging due to the limitations of conventional methods. Here, a low-cost, fluorescence-based, paper-strip immunoassay is presented for rapid and semi-quantitative estimation of EV concentration, inspired by pH strips. The assay utilizes nitrocellulose membranes functionalized with capture antibodies (anti-CD63, CD9, CD81) and fluorescent dye (ExoBrite™) for EV detection. Systematic optimization of assay parameters-including dye application sequence, incubation time, antibody configuration, and dye concentration-revealed that labeling EVs with dye and incubating on the nitrocellulose paper strips for 20 min yielded the strongest and most reproducible signal. A 200× dilution of ExoBrite™ dye was determined to provide the best balance between sensitivity and specificity. A standard curve generated through twofold serial dilution of EVs from ovarian cancer cell culture medium confirmed a positive, concentration-dependent fluorescence response, establishing a usable dynamic range. Compared to existing technologies, this platform enables fast, simple-to-implement EV quantification using minimal sample volume and equipment. The simplicity and scalability of the method offer strong potential for use in clinical diagnostics and EV research applications.

RNA-mediated toxicity, which can be controlled by alteration of gene expression, is considered a key event in Amyotrophic Lateral Sclerosis (ALS). Transcriptomic deregulation of miRNAs expression can spread via "horizontal" RNA transfer through extracellular vesicles (EVs) to act in conjunction with proteins, leading to changes in mRNA, which can provide early signals to indicate forthcoming neuropathological changes in the brain. The aim of this work is to compare expression profiles (obtained by miRNA-seq) from different tissues to highlight commonly expressed and tissue-specific miRNAs. miRNA species from plasma EVs were correlated with miRNA profiles obtained from peripheral blood mononuclear cells (PBMCs). Each tissue from ALS patients was compared to controls, revealing 159 deregulated (DE) miRNAs in Exosomes (EXOs), 247 DE miRNAs in PBMCs and 162 DE miRNAs in Microvesicles (MVs). Next, data were filtered to include only miRNAs expressed in disease samples (not in healthy subjects), to reduce the number of tissue- and ALS- specific miRNAs (EXO n = 22, MV = 11, PBMCs n = 8). We identified specific miRNAs and pathways related to each tissue. Interestingly, in PBMCs we found mainly neuro-linked pathways, such as neurotransmitters, brain and neuron development, while in EXOs, we found miRNAs implicated in MAPK and ERB signaling. In contrast, the altered pathways in MVs were not specific. This study shows that the composition of small RNA differs significantly between blood cells and its respective EVs fraction. Differentially expressed miRNAs can target definite transcripts in different cellular and molecular fractions. It is evident that, in terms of miRNAs cargo, MVs are not specific to ALS. Therefore, future studies will focus on the interaction between cells and EXOs.

For solid tumors, hypoxia is associated with disease aggressiveness and poor outcomes. In addition to undergoing broad intracellular molecular and metabolic adaptations, hypoxic tumor cells extensively communicate with their microenvironments to facilitate conditions favorable for their survival, growth, and metastasis. This communication is mediated by diverse secretory factors, including exosomes (extracellular vesicles of endosomal origin). Exosomal cargo is altered considerably by hypoxia, with significant impacts on tumor-cell communication with both local and distant microenvironments. Exosomes released by cancer cells influence the tumor environment to accelerate metastasis. While tumor-derived exosomes have been identified as a major driver of premetastatic niche formation at distant sites, this mechanism in lung adenocarcinoma (LUAD) remains unclear. We found that miR-671-3p in exosomes derived from H1975 under hypoxic conditions target Krüppel-like factor 2 (KLF2) to regulate VEGFR2 expression in endothelial cells to promote angiogenesis. In addition, miR-671-3p is expressed at high levels in circulating exosomes isolated from patients with LUAD. Our study suggests that exosome miR-671-3p is involved in the formation of premetastatic niche and may serve as a blood-based biomarker for LUAD metastasis.

Extracellular vesicles, which are predominantly classified into ectosomes and exosomes, are released by all cells under both physiological conditions and in response to acquired pathological states. Exosomes demonstrate multifaceted functions: they regulate cellular homeostasis through the elimination of redundant or detrimental intracellular components, function as mediators in intercellular signaling pathways, and serve as potential vectors for both diagnostic and therapeutic applications. Intestinal ischemia-reperfusion injury (IRI), a prevalent form of tissue and organ injury in surgical settings, has been extensively investigated. Emerging evidence indicates a crucial relationship between exosomes and intestinal IRI, specifically regarding how exosomes derived from either intestinal tissue or distant organs can modulate the pathophysiological progression of intestinal IRI. This review systematically evaluates the mechanistic roles of exosomes in intestinal IRI and their involvement in post-intestinal IRI multiple organ dysfunction, aiming to establish a theoretical foundation for therapeutic interventions and future research directions.

Breast cancer is one of the most prevalent malignant tumors among women and ranks as the second leading cause of cancer-related deaths in females, primarily due to delays in diagnosis and shortcomings in treatment strategies. Consequently, there is a pressing need to identify reliable therapeutic targets and strategies. In recent years, the identification of effective biomarkers-particularly novel molecular therapeutic targets-has become a focal point in breast cancer research, aimed at predicting disease aggressiveness and monitoring treatment responses. Simultaneously, advancements in understanding the molecular mechanisms underlying cellular programmed death have opened new avenues for targeting kinase-regulated programmed cell death as a viable therapeutic strategy. This review summarizes the latest research progress regarding kinase-regulated programmed death (including apoptosis, pyroptosis, autophagy, necroptosis, and ferroptosis) in breast cancer treatment. It covers the key kinases involved in this mechanism, their roles in the onset and progression of breast cancer, and strategies for modulating these kinases through pharmacological interventions.

Extracellular vesicles (EVs) are lipid-bound particles released by tumor cells and widely explored in cancer development, progression, and treatment response, being considered as valuable components to be explored as biomarkers or cellular targets to modulate the effect of therapies. The mechanisms underlying the production and profile of EVs during radiotherapy (RT) require addressing radiobiological aspects to determine cellular responses to specific radiation doses and fractionation. In this review, we explore the role of EVs in the 7 Rs of radiobiology, known as the molecular basis of a biological tissue response to radiation, supporting EVs as a shared player in all the seven processes. We also highlight the relevance of EVs in the context of liquid biopsy and resistance to immunotherapy, aiming to establish the connection and utility of EVs as tools in contemporary and precision radiotherapy.

Extracellular vesicles (EVs) originating from testicular somatic cells act as pivotal intermediaries in cell signaling crosstalk between spermatogenic cells and the testicular microenvironment. The intricate balance between palmitoylation and depalmitoylation governs the positioning of protein cargos on the membrane, thereby influencing cellular activities by concentrating these proteins in EVs for delivery to recipient cells. Here, we reveal that GNA13 undergoes specific S-palmitoylation at Cys14 and Cys18 residues in Sertoli cells (SCs), a modification essential for its localization to the plasma membrane. We identify DHHC13, a member of the zinc finger DHHC-type palmitoyltransferase family that catalyzes protein S-palmitoylation, as the enzyme responsible for this critical post-translational modification. Additionally, GNA13 palmitoylation is indispensable for its selective enrichment in EVs emanating from SCs. Intriguingly, we discovered the presence of palmitoylated GNA13 in SC-derived EVs significantly downregulates autophagy levels in spermatogonial stem cells (SSCs), and the inhibition of GNA13 palmitoylation attenuates its interaction with ARHGEF12 which leads to diminished RhoA activity and consequent elevation of autophagy in SSCs. Our results illuminate the crucial role of DHHC13-mediated GNA13 S-palmitoylation in modulating autophagy levels in SSCs through SCs-derived EVs, suggesting that PM-GNA13-EV may serve as a potential candidate for further exploration in addressing fertility-related challenges during spermatogenesis.

Extracellular vesicles (EVs) are nanoscale luminal vesicles, which play an important role in intercellular communication through surface signaling and molecular cargo delivery (proteins, lipids, nucleic acids, etc.). Recently, plant-derived extracellular vesicles (PDVs) containing multiple biological activities have received increasing attention due to their better biocompatibility and lower cytotoxicity in healthy tissues. In the biomedical field, PDVs are employed as cargo delivery vehicles, enabling diverse functionalities through engineering modification techniques. Nonetheless, there are certain issues with the study of PDVs, such as the lack of standardization in the identification and isolation criteria. This review provides a quick overview of the biogenesis, physicochemical properties, isolation techniques, and biomedical applications of PDVs in current studies, while critically analyzing the current challenges and opportunities. This paper is expected to provide some theoretical guidance for the development of PDVs and further biomedical applications.

Yersinia pestis is the etiologic agent of the plague. A hallmark of plague is subversion of the host immune response by disrupting host signalling pathways required for inflammation. This non-inflammatory environment permits bacterial colonization and has been shown to be essential for disease manifestation. Previous work has shown that Y. pestis inhibits phagocytosis and degranulation by neutrophils. Manipulation of these key vesicular trafficking pathways suggests that Y. pestis influences extracellular vesicle (EV) secretion, cargo selection, trafficking and/or maturation. Our goals were to define the EV population produced by neutrophils in response to Y. pestis and determine how these vesicles might influence inflammation. Towards these goals, EVs were isolated from human neutrophils infected with Y. pestis or a mutant lacking bacterial effector proteins known to manipulate host cell signalling. Mass spectrometry data revealed that cargoes packaged in EVs isolated from mutant infected cells were enriched with antimicrobial and cytotoxic proteins, contents which differed from uninfected and Y. pestis infected cells. Further, EVs produced in response to Y. pestis lacked inflammatory properties observed in those isolated from neutrophils responding to the mutant. Together, these data demonstrate that Y. pestis actively inhibits the production of antimicrobial EVs produced by neutrophils, likely contributing to immune evasion.

During sexual reproduction, the freshwater ciliate Tetrahymena thermophila sheds membrane-bound vesicles into the extracellular environment (cEMVs: ciliary extracellular micro-vesicles). We provide evidence that 100 nm vesicles shed from the cilia of starved cells promote mating between cells of complementary mating types. A proteomic analysis revealed that these EMVs are decorated with mating-type proteins expressed from the MAT locus, proteins that define a cell's sex (one of seven). Once the mating junction is established between cells, smaller 60 nm vesicles (junction vesicles) appear within the extracellular gap that separates mating partners. Junction vesicles (jEMVs) may play a role in remodeling the mating junction through which gametic pronuclei are exchanged. Evidence is presented demonstrating that cells must be able to internalize extracellular signals via some form of endocytosis in order to trigger conjugation. Finally, an evolutionarily conserved fusogen (Hap2) implicated in pore formation also appears necessary for jEMV processing. This system offers an excellent opportunity for studies on ectosome shedding, intercellular signaling and shed vesicle uptake by macro-pinocytosis, as they relate to sexual reproduction in the ciliate Tetrahymena thermophila.

Extracellular particles (EPs), including extracellular vesicles (EVs) and non-vesicular extracellular particles (NVEPs), are multimolecular biomaterials released by cells that play a crucial role in intercellular communication. Recently, new subtypes of EPs associated with central nervous system (CNS), such as exophers and supermeres have been identified. These EPs provide new perspectives for understanding the pathological progression of CNS disorders and confer potential diagnostic value for liquid biopsies in neurodegenerative diseases (NDs). Moreover, EPs have emerged as promising drug delivery vehicles and targeted platforms for CNS-specific therapies. In this review, we delineate the landscape of EP subtypes and their roles in the pathophysiology of CNS diseases. We also review the recent advances of EP-based diagnosis in NDs and highlight the importance of analytical platforms with single-particle resolution in the exploitation of potential biomarkers. Furthermore, we summarize the application of engineered EVs in the treatment of CNS diseases and outline the underexplored potential of NVEPs as novel therapeutic agents.

Extracellular vesicles (EVs) are small particles with phospholipid bilayers that carry a diverse range of cargoes including nucleic acids, proteins and metabolites. EVs have important roles in various cellular processes and are increasingly recognized for their ubiquitous role in cell-cell communications and potential applications in therapeutics and diagnostics. Although many methods have been developed for the characterization and measurement of EVs, analyzing them from biofluids remains a challenge with regard to throughput and sensitivity. Recently, we introduced an approach to facilitate high-throughput analysis of EVs from trace amounts of sample. In this method, an amphiphile-dendrimer supramolecular probe (ADSP) is coated onto a nitrocellulose membrane for array-based capture and to enable an in situ immunoblotting assay. Here, we describe the protocol for our array-based method of EV profiling. We describe an enhanced version of the method that incorporates an automated printing workstation, ensuring high throughput and reproducibility. We further demonstrate the use of our array to profile specific glycosylations on the EV surface using click chemistry of an azide group introduced by metabolic labeling. In this protocol, the synthesis of ADSP and the fabrication of ADSP nitrocellulose membrane array can be completed on the same day. EVs are efficiently captured from biological or clinical samples through a 30-min incubation, followed by an immunoblotting assay within a 3-h window, thus providing a high-throughput platform for EV isolation and in situ targeted analysis of EV proteins and their modifications.

Exosomes are actively produced extracellular vesicles, released from different cell types, that exert important regulatory roles in vital cellular functions. Tumor-derived exosomes (TDEs) have received increasing attention because they enable intercellular communication between the neoplastic and non-neoplastic cells present in the microenvironment of tumors, affecting important functions of different types of mesenchymal stem cells (MSCs) with the ability to self-renew and differentiate. MSC-derived exosomes (MSC-exos) carry a variety of bioactive molecules that can interact with specific cellular targets and signaling pathways, influencing critical processes in tumor biology, and exhibiting properties that either promote or inhibit tumor progression. They can regulate the tumor microenvironment by modulating immune responses, enhancing or suppressing angiogenesis, and facilitating tumor cells' communication with distant sites, thus altering the behavior of non-cancerous cells present in the microenvironment. Herein, we explore the main functions of TDEs and their intricate interactions with MSC-exos, in terms of enhancing cancer progression, as well as their promising clinical applications as tumor microenvironment modulators.

The therapeutic potential of extracellular vesicles (EVs) in reducing oral inflammation is thoroughly examined in this review, with an emphasis on gingivitis, periodontitis, and oral mucositis. It explains the complex relationship between microbial dysbiosis and host immune responses in the aetiology of oral inflammation. Pathophysiological mechanisms of periodontitis are examined, emphasising the roles played by periodontal pathogens and inflammatory mediators in the disease's chronic course and systemic effects. Preclinical research is providing new evidence that EVs originating from various cellular sources control immune cell dynamics towards a pro-healing phenotype, promote tissue regeneration, and have immunomodulatory qualities. EV-based therapies appear to be a promising new therapeutic technique with potential benefits over traditional methods for the treatment of oral inflammatory illnesses by specifically altering inflammatory signalling pathways. This review highlights the potential of EVs to improve patient outcomes in oral health and emphasises the need for additional clinical research to clarify the therapeutic efficacy and underlying mechanisms of EVs in periodontal therapy.

Migrasomes are a newly discovered type of extracellular vesicle (EV) formed during cell migration, playing a pivotal role in intercellular communication. These vesicles are generated by retracting fibers of migrating cells and encapsulate various molecules, such as proteins, lipids, and RNA, allowing the transfer of biochemical signals to neighboring cells. Current evidence suggests that migrasomes are involved in a wide range of physiological processes such as embryogenesis, angiogenesis, immune modulation, and mitochondrial quality control. Moreover, migrasomes are implicated in pathological conditions, including cancer metastasis, cardiovascular diseases, and viral infections. To fully understand their significance, it is critical to first explore the molecular mechanisms underlying their formation and function. Recent studies have shed light on the biogenesis, release, and biological properties of migrasomes, all of which are key to understanding their role in cell-to-cell communication. In this review, we provide an up-to-date summary of migrasome biogenesis, release, characterization, and their biological activities in intercellular communication, while also proposing potential new functions for these vesicles.

Cancer is the second leading cause of death in children, and solid tumors make up 30% of childhood cancers. Molecular profiling of pediatric solid tumors allows a personalized approach to therapy, but this approach mostly relies on surgical biopsy, which is invasive and carries the risk of complications. Liquid biopsy serves as a reliable alternative and a minimally invasive tool for diagnosing, prognosticating, and residual disease monitoring in childhood cancers. This review outlines the potential of exosomes as informative liquid biopsies in pediatric solid tumors. Studies highlighting the potential applications and clinical utility of exosomes and their molecular constituents as prognosticators and therapies in common childhood solid tumors, including neuroblastoma, medulloblastoma, sarcoma, and hepatoblastoma, have been overviewed. We also discuss the limitations and technical challenges of utilizing exosomes for pediatric solid tumors.

The EZH2 gene encodes an enzyme that is part of the epigenetic factor Polycomb Repressive Complex 2 (PRC2). In order to control gene expression, PRC2 mainly modifies chromatin structure. In this complex process, EZH2 methylates histone proteins, which in turn suppresses further RNA transcriptions. As a result, EZH2 dysregulations can occasionally induce abnormal gene expression patterns, which can aid in the development and progression of cancer. Non-coding RNAs significantly impact the expression of EZH2 through epigenetic mechanisms. Meanwhile, normal and cancerous cells frequently release vesicles into the extracellular matrix, also known as exosomes, that occasionally carry RNA molecules from their origin cells, including messenger RNAs, microRNAs, and other non-coding RNAs. Thus exosomes are granted the ability to regulate numerous physiological functions and act as crucial messengers between cells by influencing gene expression in the recipient cell. We conducted this review to focus on EZH2's substantial biological role and the mechanisms that regulate it, driven by the desire to understand the possible impact of exosomal RNAs on EZH2 expression.

Extracellular vesicles (EVs) are nanovesicles that facilitate intercellular communication by carrying essential biomolecules under physiological and pathological conditions including microRNAs (miRNAs). They are found in various body fluids, such as blood, urine, and saliva, and their levels fluctuate with disease progression, making them valuable diagnostic tools. However, isolating EVs is challenging due to their small size and biological complexity. Here, we summarize the principles behind the most common EV isolation methods including ultracentrifugation, precipitation, immunoaffinity, sorting, ultrafiltration, size exclusion chromatography, and microfluidics while highlighting protocol strengths and weaknesses. We also review the main strategies to identify and quantify circulating miRNAs with a particular focus on EV-encapsulated miRNAs. Since these miRNAs hold special clinical interest derived from their superior stability and therapeutic potential, the information provided here should provide valuable guidance for future research initiatives in the promising field of disease diagnostic and treatment based on EV-encapsulated miRNAs.

Living organisms must adapt and communicate effectively in their environment to survive. Cells communicate through various mechanisms, including releasing growth factors, chemokines, small bioactive molecules, and cell-cell contact. In recent years, a new and sophisticated cell communication mechanism based on extracellular vesicles (EVs) has been described in all three domains of life: archaea, bacteria, and eukaryotes. EVs are small, bilayer proteolipid vesicles released by cells into the extracellular space. This review aims to analyze and compare the current literature on bacterial, archaeal, and eukaryotic EVs and their possible clinical applications. This framework will address three key points: (a) The role of EVs in bacteria, eukaryotes, and archaea. (b) What is the impact of EVs in archaea on disease?

Extracellular vesicles (EVs) are secreted by most cell types and play a central role in cell-cell communication. These naturally occurring nanoparticles have been particularly implicated in cancer, but EV heterogeneity and lengthy isolation methods with low yield make them difficult to study. To circumvent the challenges in EV research, we aimed to develop a unique synthetic model by engineering bioinspired liposomes to study EV properties and their impact on cellular uptake. We produced EV-like liposomes mimicking the physicochemical properties as cancer EVs. First, using a panel of cancer and non-cancer cell lines, small EVs were isolated by ultracentrifugation and characterized by dynamic light scattering (DLS) and nanoparticle tracking analysis (NTA). Cancer EVs ranged in mean size from 107.9 to 161 nm by NTA, hydrodynamic diameter from 152 to 355 nm by DLS, with a zeta potential ranging from - 25 to -6 mV. EV markers TSG101 and CD81 were positive on all EVs. Using a microfluidics bottom-up approach, liposomes were produced using the nanoprecipitation method adapted to micromixers developed by our group. A library of liposome formulations was created that mimicked the ranges of size (90-222 nm) and zeta potential (anionic [-47 mV] to neutral [-1 mV]) at a production throughput of up to 41 mL/h and yielding a concentration of 1 × 1012 particles per mL. EV size and zeta potential were reproduced by controlling the flow conditions and lipid composition set by a statistical model based on the response surface methodology. The model was fairly accurate with an R-squared > 70% for both parameters between the targeted EV and the obtained liposomes. Finally, the internalization of fluorescently labeled EV-like liposomes was assessed by confocal microscopy and flow cytometry, and correlated with decreasing liposome size and less negative zeta potential, providing insights into the effects of key EV physicochemical properties. Our data demonstrated that liposomes can be used as a powerful synthetic model of EVs. By mimicking cancer cell-derived EV properties, the effects on cellular internalization can be assessed individually and in combination. Taken together, we present a novel system that can accelerate research on the effects of EVs in cancer models.

Atopic dermatitis (AD) is a global concern marked by inflammation, skin barrier dysfunction, and immune dysregulation. Current treatments primarily address symptoms without offering a cure, underscoring the need for innovative therapeutic approaches. Mesenchymal stromal cell-derived extracellular vesicles (MSC-EVs) have attracted attention for their potential in immunomodulation and tissue repair, similar to their parent cells. This review provides a comprehensive analysis of the current landscape of MSC-EV research for AD management. We identified 12 studies that met our predefined inclusion criteria. We thoroughly reviewed both human and animal studies, analyzing aspects such as the source, isolation, and characterization of MSC-EVs, as well as the animal and disease models, dosage strategies, efficacy, mechanisms, and adverse effects. While this review highlights the promising potential of MSC-EV therapy for AD, it also emphasizes significant challenges, including heterogeneity and insufficient reporting. Given that this research area is still in its early stages, addressing these uncertainties will require collaborative efforts among researchers, regulatory bodies, and international societies to advance the field and improve patient outcomes.

Exosomes have emerged as promising tools for targeted drug delivery in biomedical applications and medicine. This review delves into the scientific advancements, challenges, and future prospects specifically associated with these technologies. In this work, we trace the research milestones that led to the discovery and characterization of exosomes and extracellular vesicles, and discuss strategies for optimizing the synthetic yield and the loading of these particles with various therapeutics. In addition, we report the current major issues affecting the field and hampering the clinical translation of these technologies. Highlighting the pivotal role of imaging techniques, we explore how they drive exosome therapy and development by offering insights into biodistribution and cellular trafficking dynamics. Methodologies for vesicle isolation, characterization, loading, and delivery mechanisms are thoroughly examined, alongside strategies aimed at enhancing their therapeutic efficacy. Special emphasis was dedicated to their therapeutic properties, particularly to their ability to deliver biologics into the cytoplasm. Furthermore, we delve into the intricate balance between surface modifications and targeting properties including also transgenic methods aimed at their functionalization and visualization within biological systems. This review underscores the transformative potential of these carriers in targeted drug delivery and identifies crucial areas for further research and clinical translation.

Enzyme-powered micro/nanomotors (MNMs) (EMNMs) use natural enzymes to facilitate the decomposition of fuels, including hydrogen peroxide (H2O2), glucose, triglycerides, and urea to provide power. EMNMs can achieve self-propulsion through the in situ utilization of biofuels without additional fuels, exhibiting excellent biocompatibility and significant potential for application in the biomedical field. Compared with H2O2, which may cause oxidative damage to the body, urea exhibits superior biosafety characteristics. Presently, urease-powered MNMs (UMNMs) have made notable progress in their applications in the biomedical field and have garnered considerable attention from researchers. In this review, we present the latest advancements in the biomedical field of UMNMs, primarily focusing on: 1) diverse materials used for constructing the fundamental framework of motors; 2) control of motor movement through the regulation of enzymatic reaction rates; and 3) research directions for the clinical application of motors, including in vivo imaging, biomarker detection, cancer treatment, optical therapy, overcoming biological barriers, antibacterial interventions, antithrombotic strategies, and gastric disease management. Despite showing immense potential in biomedical applications, there are still several challenges impeding its practical implementation, such as maintaining activity in the in vivo environment while accurately targeting specific sites to achieve the desired clinical therapeutic effects.

Non-healing wounds pose significant challenges and require effective therapeutic interventions. Extracellular vesicles (EVs) have emerged as promising cell-free therapeutic agents in tissue regeneration. However, the functional differences between different subpopulations of EVs in wound healing remain understudied. This study aimed to evaluate the effects of two distinct subpopulations of clonal mesenchymal stromal cells (cMSC)-derived EVs (cMSC-EVs), namely 20 K and 110K-cMSC-EVs, primarily on in vitro wound healing process, providing fast and cost-effective alternatives to animal models.

In vitro assays were conducted to compare the effects of 20 K and 110K-cMSC-EVs, isolated through high-speed centrifugation and differential ultracentrifugation, respectively. For evaluation the main mechanisms of wound healing, including cell proliferation, cell migration, angiogenesis, and contraction. Human dermal fibroblasts (HDF) were considered as the main cells for analysis of these procedures. Moreover, gene expression analysis was performed to assess the impact of these EV subpopulations on the related process of wound healing on HDF.

The results demonstrated that both 20 K and 110K-cMSC-EVs exhibited beneficial effects on cell proliferation, cell migration, angiogenesis, and gel contraction. RT-qPCR revealed that both EV types downregulated interleukin 6 (IL6), induced proliferation by upregulating proliferating cell nuclear antigen (PCNA), and regulated remodeling by upregulating matrix metallopeptidase 1 (MMP1) and downregulating collagen type 1 (COL1).

This study highlights the effects of both 20 K and 110K-cMSC-EVs on the potency of HDFs in wound healing-related process. As the notable finding, 20K-cMSC-EVs offer a more feasible and cost-effective subpopulation for isolation and follow the GMP standard, recommended to utilize this fraction for therapeutic application.

Extracellular vesicles (EVs) are heterogeneous vesicles released by donor cells that can be taken up by recipient cells, thus inducing cellular phenotype changes. Since their discovery decades ago, roles of EVs in modulating initiation, growth, survival and metastasis of cancer have been revealed. Recent studies from multifaceted perspectives have further detailed the contribution of EVs to cancer drug resistance; however, the role of EV uptake in conferring drug resistance seems to be overlooked. In this comprehensive review, we update the EV subtypes and approaches for determining EV uptake. The biological basis of EV uptake is systematically summarized. Moreover, we focus on the diverse uptake mechanisms by which EVs carry out the intracellular delivery of functional molecules and drug resistance signaling. Furthermore, we highlight how EV uptake confers drug resistance and identify potential strategies for targeting EV uptake to overcome drug resistance. Finally, we discuss the research gap on the role of EV uptake in promoting drug resistance. This updated knowledge provides a new avenue to overcome cancer drug resistance by targeting EV uptake.

Pregnancy involves extracellular vesicles (EVs) through mechanisms that are poorly understood to date. Furthermore, it is not surprising that EVs may also be involved in the pathophysiology of pre-eclampsia (PE) and gestational hypertension, two clinical conditions with high morbidity and mortality, given their capacity to mediate intracellular communications and regulate inflammation and angiogenesis. We searched major online scientific search engines (PubMed, Google Scholar, Scopus, WES, Embase, etc.) using the terms "Preeclampsia", "Pregnancy", "Hypertension", "Pregnancy-related hypertension", "Extracellular vesicles", "Biomarkers", "Gestation" AND "Obstetrics". Finding potential early biomarkers of risk or illness progression would be essential for the optimum care of expectant mothers with the aforementioned conditions. Nevertheless, none of the various screening assays that have been discovered recently have shown high predictive values. The analysis of EVs in the peripheral blood starting from the first trimester of pregnancy may hold great promise for the possible correlation with gestational hypertension problems and represent a marker of the early stages of the disease. EVs use may be a novel therapeutic approach for the management of various illnesses, as well. In order to define EVs' function in the physiopathology of pregnancy-associated hypertension and PE, as well as their potential as early biomarkers and therapeutic tools, we have compiled the most recent data in this review.

The activation of astrocytes in the injured lesion induces the progression of spinal cord injury (SCI). However, adverse side-effects during systemic administration have limited applications. Exosomes (Exos) are an emerging clinical treatment method that exerts anti-inflammatory effects by reducing pro-inflammatory factors and promoting functional recovery. Exosomes exhibit great potential as carriers of traditional Chinese medicine, attributed to their high delivery efficiency to internalized and targeted accumulation in inflammatory tissues. Herein, We synthesized resveratrol loaded microglia-derived exosomes (R-MDEs) for highly efficient accumulation and infiltration in the injured spinal cord. In vitro and in vivo experiments suggested that R-MDEs effectively accumulated in A1 astrocytes, inhibited reactive oxygen species (ROS) and glial scar formation by reprogramming the metabolic astrocytes. R-MDEs achieve a synergistic therapeutic effect of immunomodulation and neuroprotection, thereby shedding new light on the application of Exos and provides great potential for SCI.

Major depressive disorder (MDD) is considered a psychiatric disorder and have a relationship with stressful events. Although the common therapeutic approaches against MDD are diverse, a large number of patients do not present an adequate response to antidepressant treatments. On the other hand, effective non-pharmacological treatments for MDD and their tolerability are addressed. Several affective treatments for MDD are used but non-pharmacological strategies for decreasing the common depression-related drugs side effects have been focused recently. However, the potential of extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs), microRNAs (miRNAs) as cell-based therapeutic paradigms, besides other non-pharmacological strategies including mitochondrial transfer, plasma, transcranial direct current stimulation (tDCS), transcranial magnetic stimulation (TMS), and exercise therapy needs to further study. This review explores the therapeutic potential of cell-based therapeutic non-pharmacological paradigms for MDD treatment. In addition, plasma therapy, mitotherapy, and exercise therapy in several in vitro and in vivo conditions in experimental disease models along with tDCS and TMS will be discussed as novel non-pharmacological promising therapeutic approaches.

Exosomes are nanoscale vesicles secreted by cells that play vital regulatory roles in intercellular communication and immune responses. Listeria monocytogenes (L. Monocytogenes, LM) is a notable Gram-positive intracellular parasitic bacterium that infects humans and diverse animal species. However, the specific biological function of exosomes secreted by macrophages during L. Monocytogenes infection (hereafter EXO-LM) remains elusive. Here, we discovered that EXO-LM stimulated the secretion of inflammation-associated cytokines by macrophages, facilitating the intracellular survival of L. monocytogenes within macrophages. Transcriptomic analysis shows that EXO-LM significantly upregulates immune recognition and inflammation-related signaling pathways in macrophages. Furthermore, a ceRNA regulatory network comprising exosomal ncRNAs and macrophage RNAs was constructed through EXO-LM transcriptome sequencing. Utilizing bioinformatics and dual-luciferase reporter assays, we identified two potential binding sites between lncRNA Rpl13a-213 and miR-132-3p. Cell transfection experiments demonstrated that Rpl13a-213 overexpression augmented pro-inflammatory cytokine expression in macrophages, in contrast to the suppression by miR-132-3p overexpression. The decrease in Rpl13a-213 upon EXO-LM stimulation enhances miR-132-3p expression, dampening the inflammatory response in macrophages and aiding L. monocytogenes intracellular survival. This study unveils the immunomodulatory function of exosomal ncRNAs originating from macrophages, which provides fresh perspectives into the mechanisms underlying macrophage inflammatory response regulation by L. monocytogenes-infected cell-derived exosomes.

Ovarian cancer (OC) is the third most common malignant tumor of women and is accompanied by an alteration of systemic metabolism. A liquid biopsy that captures and detects tumor-related biomarkers in body fluids has great potential for OC diagnosis. EVs, nanosized extracellular vesicles found in the blood, have been proposed as promising biomarkers for liquid biopsies. In this study we recruited 37 OC patients, 22 benign ovarian tumor (BE) patients, and 46 clinically healthy control patients (CON). Plasma EVs were purified from blood samples and sensitive thermal separation probe-based mass spectrometry analysis using a global untargeted metabolic profiling strategy was employed to characterize the metabolite fingerprints. Uniform manifold approximation and projection (UMAP) analysis demonstrated a distinct separation of EVs among the three groups. We screened for diagnostic biomarkers from plasma EV metabolites using seven machine learning algorithms, including artificial neural network (ANN), decision tree (DT), K nearest neighbor (KNN), logistics regression (LR), Naïve Bayes (NB), random forest (RF), and support vector machine (SVM). For the OC-CON comparison, the highest AUC values were found for RF (0.91), ANN (0.90) and NB (0.90), with the F1-scores of 0.88, 0.83, and 0.76 respectively. For the OC-BE comparison, SVM (0.94), RF (0.86), and KNN (0.86) gave the highest AUCs, with F1-scores of 0.80, 0.80, and 0.91 respectively. A total of 19 and 158 metabolic features exhibited significant differences (FC = 1.5, q < 0.01) in the OC vs BE and OC vs CON comparisons, respectively. Notably, the quantities of 9-octadecenamide and 1,4-methanobenzocyclodecene were significantly elevated, while maltol showed a significant reduction in the OC group compared to the BE group. When comparing the OC group to the CON group, the concentrations of 4-amino-furazan-3-carboxylic acid 2-hydroxy-4-methoxybenzaldehyde, N-phenylethyl, and 4-morpholineethanamine were significantly elevated, while the remaining metabolites, including hydrazine and pyridine sulfonamide, were reduced, in the OC group. The metabolites showing different abundancies are associated with cancer-related mutations, immune responses, and metabolic reprogramming. We demonstrate that the RF algorithm, combined with sensitive thermal separation probe-based mass spectrometry analysis of plasma EVs, can effectively identify OC patients with good accuracy. Thus, our study has shortlisted a set of potential biomarkers in plasma EVs, and the proposed approach could serve as a routine prescreening tool for ovarian cancer.

Treatment of solid tumors remains challenging and therapeutic strategies require continuous development. Tumor-infiltrating macrophages play a pivotal role in tumor dynamics. Here, we present a Macrophage-Drug Conjugate (MDC) platform technology that enables loading macrophages with ferritin-drug complexes. We first show that macrophages actively take up human heavy chain ferritin (HFt) in vitro via macrophage scavenger receptor 1 (MSR1). We further manifest that drug-loaded macrophages transfer ferritin to adjacent cancer cells through a process termed 'TRAnsfer of Iron-binding protein' (TRAIN). The TRAIN process requires direct cell-to-cell contact and an immune synapse-like structure. At last, MDCs with various anti-cancer drugs are formulated with their safety and anti-tumor efficacy validated in multiple syngeneic mice and orthotopic human tumor models via different routes of administration. Importantly, MDCs can be prepared in advance and used as thawed products, supporting their clinical applicability. This MDC approach thus represents a promising advancement in the therapeutic landscape for solid tumors.

The complex and continuously evolving features of the tumor microenvironment, varying between tumor histotypes, are characterized by the presence of host cells and tumor cells embedded in a milieu shaped by hypoxia and low pH, resulting from the frequent imbalance between vascularity and tumor cell proliferation. These microenvironmental metabolic stressors play a crucial role in remodeling host cells and tumor cells, contributing to the stimulation of cancer cell heterogeneity, clonal evolution, and multidrug resistance, ultimately leading to progression and metastasis. The extracellular vesicles (EVs), membrane-enclosed structures released into the extracellular milieu by tumor/host cells, are now recognized as critical drivers in the complex intercellular communication between tumor cells and the local cellular components in a hypoxic/acidic microenvironment. Understanding the intricate molecular mechanisms governing the interactions between tumor and host cells within a hypoxic and acidic microenvironment, triggered by the release of EVs, could pave the way for innovative strategies to disrupt the complex interplay of cancer cells with their microenvironment. This approach may contribute to the development of an efficient and safe therapeutic strategy to combat cancer progression. Therefore, we review the major findings on the release of EVs in a hypoxic/acidic tumor microenvironment to appreciate their role in tumor progression toward metastatic disease.

Extracellular vesicles (EVs) serve as critical mediators of intercellular communication, encompassing exosomes, microvesicles, and apoptotic vesicles that play significant roles in diverse physiological and pathological contexts. Numerous studies have demonstrated that EVs derived from mesenchymal stem cells (MSC-EVs) play a pivotal role in facilitating tissue and organ repair, alleviating inflammation and apoptosis, enhancing the proliferation of endogenous stem cells within tissues and organs, and modulating immune function-these functions have been extensively utilized in clinical applications. The precise classification, isolation, and identification of MSC-EVs are essential for their clinical applications. This article provides a comprehensive overview of the biological properties of EVs, emphasizing both their advantages and limitations in isolation and identification methodologies. Additionally, we summarize the protein markers associated with MSC-EVs, emphasizing their significance in the treatment of various diseases. Finally, this article addresses the current challenges and dilemmas in developing clinical applications for MSC-EVs, aiming to offer valuable insights for future research.