Extracellular vesicles (EVs) are promising biomarkers for neurodegeneration. Alu elements are retrotransposons increasingly expressed with age and may be involved in aging-related diseases.

To determine the potential of Alu RNA in plasma-derived EVs as a biomarker for brain aging and neuronal injury.

EVs were isolated from plasma samples across different age groups. EV Alu RNA levels were measured and their associations with biomarkers of brain aging, including plasma neurofilament light chain (NfL), plasma amyloid-beta (Aβ42 and Aβ40), and plasma phosphorylated tau (p-Tau181), were analyzed.

EV Alu RNA levels were increased significantly with age and were strongly correlated with plasma NfL, suggesting a strong association between EV Alu RNA and neuronal injury. Significant correlations were also found between EV Alu RNA and plasma amyloid-beta levels, while no significant association was observed with tau pathology.

EV Alu RNA levels are elevated with age and associated with neuronal injury, highlighting their potential as a novel, non-invasive biomarker for brain aging and neurodegeneration.

Extracellular vesicles (EVs) are nanoscale membrane vesicles actively released by cells into a variety of biofluids. EVs carry myriad molecular cargoes; these include classical genetic biomarkers inherited from the parent cells as well as EV modifications by other entities (e.g., small molecule drugs). Aided by these diverse cargoes, EVs enable long-distance intercellular communication and have been directly implicated in various disease pathologies. As such, EVs are being increasingly recognized as a source of valuable biomarkers for minimally-invasive disease diagnostics and prognostics. Despite the clinical potential, EV molecular profiling remains challenging, especially in clinical settings. Due to the nanoscale dimension of EVs as well as the abundance of contaminants in biofluids, conventional EV detection methods have limited resolution, require extensive sample processing and can lose rare biomarkers. To address these challenges, new micro- and nanotechnologies have been developed to discover EV biomarkers and empower clinical applications. In this review, we introduce EV biogenesis for different cargo incorporation, and discuss the use of various EV biomarkers for clinical applications. We also assess different chip-based integrated technologies developed to measure genetic and modified biomarkers in EVs. Finally, we highlight future opportunities in technology development to facilitate the clinical translation of various EV biomarkers.

Messenger RNA (mRNA) has emerged as a highly promising therapeutic approach with successful applications across various diseases. Developing safe and efficient carriers to preserve mRNA integrity during in vivo delivery is critical for ensuring the therapeutic efficacy of mRNA-based treatments. Exosomes, natural nanovesicles secreted by cells, possess several favorable properties, including excellent biocompatibility, low immunogenicity, the unique ability to traverse physiological barriers and to target specific cell types, which make them attractive candidates for mRNA delivery. Moreover, exosomes can be engineered to increase their yield, enhance targeting specificity, improve cellular uptake efficiency, and extend their circulation time in the bloodstream. However, several challenges must be addressed urgently, including exosome heterogeneity, scalable production methods, and the efficient encapsulation of mRNA cargo. This review highlights the advantages and limitations of exosome-based mRNA delivery systems, discusses current strategies for engineering exosomes to improve mRNA delivery, and describes their applications in disease treatment-emphasizing their potential to advance precision medicine and targeted therapies.

Chemotherapy resistance represents a formidable obstacle in oncological therapeutics, substantially compromising the efficacy of adjuvant chemotherapy regimens and contributing to unfavorable clinical prognoses. Emerging evidence has elucidated the pivotal involvement of exosomes in the dissemination of chemoresistance phenotypes among tumor cells and within the tumor microenvironment. This review delineates two distinct intra-tumoral resistance mechanisms orchestrated by exosomes: (1) the exosome-mediated sequestration of chemotherapeutic agents coupled with enhanced drug efflux in neoplastic cells, and (2) the horizontal transfer of chemoresistance to drug-sensitive cells through the delivery of bioactive molecular cargo, thereby facilitating the propagation of resistance phenotypes across the tumor population. Furthermore, the review covers current in vivo experimental data focusing on targeted interventions against specific genetic elements and exosomal secretion pathways, demonstrating their potential in mitigating chemotherapy resistance. Additionally, the therapeutic potential of inhibiting exosome-mediated transporter transfer strategy is particularly examined as a promising strategy to overcome tumor resistance mechanisms.

Extracellular vesicles (EVs) are membrane vesicles released or secreted from almost all cell types. EVs are derived from multivesicular bodies or from the plasma membrane and contain a subset of proteins, lipids, and nucleic acids (e.g., DNA, RNA, and microRNA [miRNA]) derived from the parent cell. EVs play important roles in intercellular communication by efficiently transferring the content between cells both locally and systemically. Given their natural ability to transfer cargo to cells, sometimes in a targeted manner, and their apparent lack of immunogenicity, EVs are being engineered for delivery of therapeutic RNAs, DNAs, miRNAs, viral particles, drugs, and even proteins. In addition, many of the therapeutic effects of stem cell treatments are mediated by stem cell-derived EVs, which are safer and potentially more effective than the parental stem cells. Here we provide an overview of the use of EVs for delivery of different therapeutic nucleic acids, viruses, and drugs, as well as the use of therapeutic stem cell-derived EVs.

Leishmaniasis continues to be a critical international health issue due to the scarcity of efficient treatment and the development of drug tolerance. New developments in the research of extracellular vesicles (EVs), especially exosomes, have revealed novel disease management approaches. Exosomes are small vesicles that transport lipids, nucleic acids, and proteins in cell signalling. Its biogenesis depends on several cellular processes, and their functions in immune response, encompassing innate and adaptive immunity, underline their function in the pathogen-host interface. Exosomes play a significant role in the pathogenesis of some parasitic infections, especially Leishmaniasis, by helping parasites escape host immunity and promote disease progression. This article explains that in the framework of parasitic diseases, exosomes can act as master regulators that define the pathogenesis of the disease, as illustrated by the engagement of exosomes in the Leishmaniasis parasite and immune escape processes. Based on many published articles on Leishmaniasis, this review aims to summarize the biogenesis of exosomes, the properties of the cargo in exosomes, and the modulation of immune responses. We delve deeper into the prospect of using exosomes for the therapy of Leishmaniasis based on the possibility of using these extracellular vesicles for drug delivery and as diagnostic and prognostic biomarkers. Lastly, we focus on the recent research perspectives and future developments, underlining the necessity to continue the investigation of exosome-mediated approaches in Leishmaniasis treatment. Thus, this review intends to draw attention to exosomes as a bright new perspective in the battle against this disabling affliction.

Extracellular vesicles (EVs), isolated through techniques such as liquid biopsy, have emerged as crucial biomarkers in various diseases, including cancer. EVs were dismissed initially as cellular debris, EVs are now recognized for their role in intercellular communication, carrying proteins, RNAs, and other molecules between cells. Their stability in biofluids and ability to mirror their parent cells' molecular composition make them attractive candidates for non-invasive diagnostics. EVs, including microvesicles and exosomes, contribute to immune modulation and cancer progression, presenting both therapeutic challenges and opportunities. However, despite advances in analytical techniques like high-resolution microscopy and nanoparticle tracking analysis (NTA), standardization in EV isolation and characterization remains a hurdle. Cutting-edge technologies, such as atomic force microscopy and Raman tweezers microspectroscopy, have enhanced our understanding of single EVs, yet issues like low throughput and high technical complexity limit their widespread application. Other technologies like transmission electron microscopy, cryogenic transmission electron microscopy, super-resolution microscopy, direct stochastic optical reconstruction microscopy, single-molecule localization microscopy, tunable resistive pulse sensing, single-particle interferometric reflectance imaging sensor, flow cytometry, droplet digital analysis, total internal reflection fluorescence also contribute to EV analysis. Future research must focus on improving detection methods, developing novel analytical platforms, and integrating artificial intelligence to enhance the specificity of EV characterization. The future of EV research holds promise for breakthroughs in precision medicine, with a collaborative effort needed to translate these advancements into clinical practice.

Extracellular vesicles (EVs) are released by all kind of cells into the extracellular space, where they shuttle parental cell-derived molecular cargoes (DNA, RNA, proteins) to both adjacent and distant cells, influencing the physiology of target cells. Their specific cargo content and abundance in liquid biopsies make them excellent candidates for biomarker studies. Indeed, EVs isolated from various body fluids, including blood, pleural fluid, urine, cerebrospinal fluid, saliva, milk, ascites, and tears, have been recognized for their potential as biomarkers in diagnosis, monitoring treatment, and predicting outcomes for various diseases. Increasing studies suggest that tears have great promise as a noninvasive liquid biopsy source for EVs. Our aim here is to provide a comprehensive review of the exploration of tears as a noninvasive reservoir of EVs and their contents, evaluating their accessibility and potential utility as a liquid biopsy method. Additionally, the potential of tear EVs in various cancers, including ocular cancer, is discussed. Finally, the advantages and challenges of employing tear-based liquid biopsy for EVs for the disease's biomarker studies are evaluated.

Pancreatic cancer has a high mortality rate, and both the incidence and mortality are continuing to increase in many countries globally. The poor prognosis of pancreatic cancer is in part due to the challenges in early diagnosis. Improving early-stage pancreatic cancer diagnosis would improve survival outcomes. Aptamer-based biosensors provide an alternative technological approach for the analysis of serum biomarkers with several potential advantages. This review summarizes the major pancreatic cancer serum biomarkers, as well as discusses recent progress in biomarker exploration and aptasensor development. Here, we review both established and novel serum biomarkers identified recently, emphasizing their potential for early-stage pancreatic cancer diagnosis. We also propose strategies for further expanding multiplex biomarker panels beyond the established CA19-9 biomarker to enhance diagnostic performance. We discuss technological advancements in aptamer-based sensors for pancreatic cancer-related biomarkers over the last decade. Optical and electrochemical sensors are highlighted as two primary modalities in aptasensor design, each offering unique advantages. Finally, we propose steps towards clinical application using aptamer-based sensors with multiplexed biomarker detection for improved pancreatic cancer diagnostics.

Background: Liquid biopsy, analyzing the expression and variation of circulating tumor cell (CTC) or circulating tumor nucleic acid genes in peripheral blood, can obtain the information of tumorigenesis, metastasis and treatment, which has a potentially broader and complementary applications in the precision cancer medicine. Small vesicles, like exosomes which belongs to extracellular vesicles (EVs), which released by living cells into the vicinal surrounding biofluids, especially cells from carcinoma. Tumor-derived materials were contained by exosomes, totally including proteins, nucleic acid genes, and lipid, etcetera for instance metabolites. Besides, molecules which were synchronously belongings and carried on exosomal surface can tolerable provides crucial clues regarding their origin. Thus, classified the types of vesicles and enrich features from tissue-specific sources become presumably achieved. Therefore, exosomes and other EVs have emerged as a liquid biopsy platform for early screening and diagnosis of cancers, which constantaneously carry-over multitudinous surface molecules also throw clues regarding their origin. It is feasible to sort vesicles types, intake with approach channel to enrich the characteristics from tissue-specific origin. Based on the property that exosomes are remarkably stable in body fluids, which can amenity gathered in plasma or urine, it can be used for meticulous clinical evaluation particularly in the early stages of carcinoma. Therefore, exosomes have aroused immense interest in the study of biomarkers. Conclusions and Perspectives: This review aims to summarize the potential exosomal biomarkers especially exosomal miRNAs for exosome-based liquid biopsy in early screening and diagnostic of cancers, including lung cancer, breast cancer (BC), kidney cancer, prostate cancer (PCa), colorectal cancer (CRC), and summarizes the traditional and novel technologies for isolating exosomes and for detecting exosomal-based biomarkers, including exosomal proteins and exosomal nucleic acids. Finally, the limitations and prospects of exosome-based liquid biopsy in the early screening and diagnosis of cancers were discussed briefly.

Cancer remains one of the leading causes of death worldwide. Its complex pathogenesis and metastasis pose significant challenges for early diagnosis, underscoring the urgent need for innovative and non-invasive tumor screening methods. Exosomes, small extracellular vesicles that reflect the physiological and pathological states of their parent cells, are uniquely suited for cancer liquid biopsy due to their molecular cargo, including RNA, DNA, and proteins. However, traditional methods for exosome isolation and detection are often limited by inadequate sensitivity, specificity, and efficiency. Nanopore technology, characterized by high sensitivity and single-molecule resolution, offers powerful tools for exosome analysis. This review highlights its diverse applications in tumor screening, such as magnetic nanopores for high-throughput sorting, electrochemical sensing for real-time detection, nanomaterial-based assemblies for efficient capture, and plasmon resonance for ultrasensitive analysis. These advancements have enabled precise exosome detection and demonstrated promising potential in the early diagnosis of breast, pancreatic, and prostate cancers, while also supporting personalized treatment strategies. Additionally, this review summarizes commercialized products for exosome-based cancer diagnostics and examines the technical and translational challenges in clinical applications. Finally, it discusses the future prospects of nanopore technology in advancing liquid biopsy toward clinical implementation. The continued progress of nanopore technology not only accelerates exosome-based precision medicine but also represents a significant step forward in next-generation liquid biopsy and tumor screening.

Plants release extracellularly lipid bilayer-enclosed vesicles of nanometric size that can be retrieved in their fluids. Plant-derived extracellular vesicles (PDEVs) have mostly been involved in modulating host-pathogen interaction, making them a tool for cross-kingdom communication with a key role in plant immunity. In addition, PDEVs have demonstrated promising therapeutic features, not only in terms of intrinsic nutraceutical properties but also of active molecules' delivery. Transgenic plants have been developed for a variety of purposes, i.e., to improve their functional properties like crops, but also to produce therapeutic molecules. However, it is unclear whether transgenes can end up in PDEVs, thus making them a vehicle for their cross-kingdom diffusion into the environment.

Here, we investigated the association of transgenic DNA and RNA with PDEVs secreted by tobacco (Nicotiana tabacum) engineered to express the neomycine phosphotransferase II (Npt-II) gene. PDEVs were isolated from leaf apoplastic fluid by ultracentrifugation and characterized for their morphology and size. The association of DNA and RNA was assessed by qRT-PCR and their immunomodulatory properties by assaying PDEVs-induced IL1β and IL10 on THP1 monocytes.

Npt-II RNA, but not DNA, could be amplified from PDEVs, whereas no differences were observed between wt and transgenic tobacco PDEVs in terms of immunomodulatory properties.

Although a different behaviour by other types of RNAs or DNAs could still be possible, our findings indicate that in this model, PDEVs are not associated with transgenic DNA, but they can protect RNA, including transgenic RNA, from degradation, contributing to their cross-kingdom spreading.

The ability of small nucleic acids to modulate gene expression via a range of processes has been widely explored. Compared with conventional treatments, small nucleic acid therapeutics have the potential to achieve long-lasting or even curative effects via gene editing. As a result of recent technological advances, efficient small nucleic acid delivery for therapeutic and biomedical applications has been achieved, accelerating their clinical translation. Here, we review the increasing number of small nucleic acid therapeutic classes and the most common chemical modifications and delivery platforms. We also discuss the key advances in the design, development and therapeutic application of each delivery platform. Furthermore, this review presents comprehensive profiles of currently approved small nucleic acid drugs, including 11 antisense oligonucleotides (ASOs), 2 aptamers and 6 siRNA drugs, summarizing their modifications, disease-specific mechanisms of action and delivery strategies. Other candidates whose clinical trial status has been recorded and updated are also discussed. We also consider strategic issues such as important safety considerations, novel vectors and hurdles for translating academic breakthroughs to the clinic. Small nucleic acid therapeutics have produced favorable results in clinical trials and have the potential to address previously "undruggable" targets, suggesting that they could be useful for guiding the development of additional clinical candidates.

Small-interfering RNA (siRNA) therapy holds significant potential for treating cardiac injury; however, its clinical application is constrained by poor blood stability and insufficient cellular uptake. Extracellular vesicles (EVs) have emerged as an effective delivery system for siRNA in vivo; but their lack of specific cell or tissue-targeting ability remains a major challenge. Thus, we aimed to develop an EV-based delivery system capable of targeted delivery of therapeutic siRNA to injured cardiac tissue for cardiac repair.

To identify fibroblast activation protein (FAP) as a potential target for delivery to injured cardiac tissue, we analyzed cardiac tissues from patients with heart failure and angiotensin II (Ang II)-treated mice. Injured cardiac tissue-targeting EVs were developed by embedding a cholesterol-conjugated FAP aptamer, which specifically targets FAP, onto human serum-derived EVs (hEV).

Our findings revealed that FAP is upregulated after cardiac injury, highlighting its potential as a target for siRNA delivery to injured cardiac tissues. We successfully developed FAP aptamer-functionalized hEV (hEV@FAP) and confirmed their typical EV characteristics, including morphology, size distribution, zeta potential, and marker protein expression. In addition, hEV@FAP demonstrated high targeting selectivity to FAP-positive regions both in vitro and in vivo. To treat cardiac injury, hEV@FAP were loaded with TGFβ1 siRNA (siTGFβ1), identified as a molecular target for cardiac repair. In Ang II-treated mice, intravenous administration of hEV@FAP-siTGFβ1 effectively reduced Ang II-induced TGFβ1 expression in cardiac tissues, attributed to the protective and targeting capabilities of hEV@FAP. Consequently, hEV@FAP-siTGFβ1 significantly improved cardiac function, reduced myocardial fibrosis, and decreased cardiomyocyte cross-sectional area (P < 0.05) without inducing systemic toxicity.

hEV@FAP represents a novel approach for targeted delivery of therapeutic siRNA to injured cardiac tissues, providing a promising nanomedicine for cardiac repair.

The inhibition of epigenetic regulators activates endogenous retrovirus (ERV) expression, which can stimulate a viral mimicry response in cancer cells. ERV elements are aberrantly expressed in hepatocellular carcinoma (HCC); however, the expression of ERVs regulated by histone modifications and their clinical significance in HCC remain unclear. Here, we identified specific human endogenous retrovirus (HERV) elements epigenetically suppressed by the histone methyltransferase SETDB1 in HCC.

The Cancer Genome Atlas (TCGA) dataset was analyzed to identify HERV elements based on SETDB1 expression levels. SETDB1 knockdown (KD) was performed in mouse and human HCC cells to investigate the resulting biological effects and changes in HERV expression, both in vitro and in vivo.

TCGA analysis revealed an inverse correlation between SETDB1 and retroelements in human HCC (R = -0.723, p = 2.297 × 10-40), identifying four specific HERV elements in SETDB1-high expressing HCC cases. Low expression of these four HERVs was associated with poor prognosis, and their combined expression provided additional prognostic insight (p <0.001). Increased expression of the four HERV elements and decreased H3K9me3 levels at these regions were detected in human HCC cells with SETDB1-KD. In murine HCC cells, Setdb1-KD impaired in vivo tumor growth with increasing CD8-positive T-cell infiltration. Moreover, the interferon α response pathway and multiple ERV elements were activated in mouse HCC cells with Setdb1-KD. The expression of interferon-stimulated genes, as indicators of a viral mimicry response, was elevated in both murine and human SETDB1-KD HCC cells.

The suppression of four novel HERV elements by SETDB1 serves as a prognostic marker in HCC. Activation of these SETDB1-regulated HERVs could represent a promising therapeutic strategy for HCC.

An inverse relationship between retroelements including human endogenous retrovirus (HERV) elements and SETDB1 expression was observed in human hepatocellular carcinoma (HCC) by The Cancer Genome Atlas data analysis. We identified four HERV elements downregulated by SETDB1-dependent H3K9me3 in HCC cells, with low expression levels of these HERV elements correlating with poor prognosis in patients with HCC. SETDB1 depletion resulted in upregulation of various interferon-stimulated genes associated with viral mimicry in HCC cells. These findings suggest that the four SETDB1-regulated HERVs could serve as prognostic markers and potential therapeutic targets for HCC.

Naturally occurring extracellular vesicles (EVs) and synthetic nanoparticles like liposomes have revolutionized precision diagnostics and medicine. EVs excel in biocompatibility and cell targeting, while liposomes offer enhanced drug loading capacity and scalability. The clinical translation of EVs is hindered by challenges including low yield and heterogeneity, whereas liposomes face rapid immune clearance and limited targeting efficiency. To bridge these gaps, biomimetic synthetic vesicles (SVs) have emerged as innovative platforms, combining the advantageous properties of EVs and liposomes. This review emphasizes critical aspects of EV biology, such as mechanisms of EV-cell interaction and source-dependent functionalities in targeting, immune modulation, and tissue regeneration, informing biomimetic SV engineering. We reviewed a broad array of biomimetic SVs, with a focus on lipid bilayered vesicles functionalized with proteins. These include cell-derived nanovesicles, protein-functionalized liposomes, and hybrid vesicles. By addressing current challenges and highlighting opportunities, this review aims to advance biomimetic SVs for transformative biomedical applications.

Extracellular vesicles (EVs), nanoscale vesicles secreted by cells, have attracted considerable attention in recent years due to their role in tumor immunomodulation. These vesicles facilitate intercellular communication by transporting proteins, nucleic acids, and other biologically active substances, and they exhibit a dual role in tumor development and immune evasion mechanisms. Specifically, EVs can assist tumor cells in evading immune surveillance and attack by impairing immune cell function or modulating immunosuppressive pathways, thereby promoting tumor progression and metastasis. Conversely, they can also transport and release immunomodulatory factors that stimulate the activation and regulation of the immune system, enhancing the body's capacity to combat malignant diseases. This dual functionality of EVs presents promising avenues and targets for tumor immunotherapy. By examining the biological characteristics of EVs and their influence on tumor immunity, novel therapeutic strategies can be developed to improve the efficacy and relevance of cancer treatment. This review delineates the complex role of EVs in tumor immunomodulation and explores their potential implications for cancer therapeutic approaches, aiming to establish a theoretical foundation and provide practical insights for the advancement of future EVs-based cancer immunotherapy strategies.

Histones are conserved nuclear proteins that function as part of the nucleosome in the regulation of chromatin structure and gene expression. Interestingly, extracellular histones populate biofluids from healthy individuals, and when elevated, may contribute to various acute and chronic diseases. It is generally assumed that most extracellular histones exist as nucleosomes, as components of extracellular chromatin. We analysed cell culture models under normal and stressed conditions to identify pathways of histone secretion. We report that core and linker histones localize to extracellular vesicles (EVs) and are secreted via the multivesicular body/exosome pathway. Upregulation of EV histone secretion occurs in response to cellular stress, with enhanced vesicle secretion and a shift towards a population of smaller EVs. Most histones were membrane associated with the outer surface of EVs. Degradation of EV-DNA did not impact significantly on EV-histone association. Individual histones  and histone octamers bound strongly to liposomes and EVs, but nucleosomes did not, showing histones do not require DNA for EV binding. Histones colocalized to tetraspanin positive EVs but using genetic or pharmacological intervention, we found that all known pathways of exosome biogenesis acted positively on histone secretion. Inhibition of autophagy and lysosomal degradation had a strong positive effect on EV histone release. Unexpectedly, EV-associated histones lacked the extensive post-translational modification of their nuclear counterparts, suggesting loss of PTMs may be involved in their trafficking or secretion. Our data does not support a significant role for EV-histones existing as nucleosomes. We show for the first time that histones are secreted from cells as membrane proteins via EVs/exosomes. This fundamental discovery provides support for further investigation of the biological activity of exosome associated histones and their role in disease.

Extracellular vesicles (EVs), particularly exosomes (EXOs) of various skeletal and stem cells, were shown to delay osteoarthritis (OA) progression, and apoptotic bodies (ABs), another EV subtype, of osteoclasts showed osteoanabolic actions and were involved in the osteoclastic-regulation of local bone formation. Moreover, this study demonstrates that microvesicles (MVs) released by osteoclasts displayed potent pro-chondrogenic, pro-osteogenic, and anti-inflammatory activities. These activities were unique to osteoclastic MVs and were not shared by osteoclastic ABs and EXOs or MVs of other cell types. Because chronic synovial inflammation, progressive articular cartilage erosion, abnormal subchondral bone remodeling, and inability to regenerate articular cartilage are key etiologies of OA, we postulate that the foregoing activities of osteoclastic MVs could simultaneously target multiple etiologies of OA and could thereby be an effective therapy for OA. Accordingly, this study sought to assess the feasibility of an osteoclastic MV-based strategy for OA with a mouse tibial plateau injury model of OA. Briefly, tibial plateau injuries were created on the right knees of adult C57BL/6J mice, MVs were intraarticularly injected into the injured joints biweekly, and the OA progression was monitored histologically at five weeks post-injury. The MV treatment reduced the OA-induced losses of articular cartilage area and thickness, decreased irregularity in the articular cartilage surface, reduced loss of gliding/intermediate zone of articular cartilage, reduced osteophyte formation, suppressed synovial inflammation, and decreased the OARSI OA score. In summary, treatment with osteoclastic MVs delayed or reversed OA progression. Thus, this study supports the feasibility of an osteoclastic MV-based therapy for OA.

Oral contraceptives (OCs) are approved for use after onset of menarche, which is well before brain maturation is complete. OC use may induce biochemical changes in the brain, especially during the neurobiologically dynamic adolescent/young adult years. MicroRNA cargo in L1CAM-associated extracellular vesicles was measured from serum samples collected from young women using the miRCURY LNA miRNA Focus PCR Panel (Qiagen) and validated using quantitative PCR. Linear regression and F-tests were applied to identify differentially expressed microRNAs by OC use (never versus current), and PANTHER pathway analysis was conducted on the gene targets of significantly differentially expressed microRNAs. Twelve microRNAs had significant differential expression variability by OC use (Bonferroni adjusted p < 0.002). Pathway analysis revealed that the 1254 unique genes targeted by the significant microRNAs were most enriched for the Gonadotropin-releasing hormone receptor pathway (FDR q = 5 × 10-7), which is associated with the release of gonadotropins, pubertal development, and reproduction. The results are consistent with the hypothesis that microRNA cargo in circulating extracellular vesicles may reflect brain-related biological activity and that OC use may influence extracellular vesicle cargo composition. The significant difference in expression variability may have implications for designing future studies, including power calculations.

Cell-cell interactions and communication represent the fundamental cornerstone of cells' collaborative efforts in executing diverse biological processes. A profound understanding of how cells interface through various mediators is pivotal across a spectrum of biological systems. Recent strides in microfluidic technologies have significantly bolstered the precision and prowess in capturing and manipulating cells with exceptional spatial and temporal resolution. These advanced methodologies converge with multi-signal mediator detection systems, furnishing potent, high-throughput platforms for dissecting cell-cell interactions at the single-cell level. This approach empowers researchers to delve into intricate cellular dynamics with unprecedented accuracy and efficiency. Here, we present a critical evaluation of the latest advancements in microfluidics-driven techniques for detecting signal mediators involved in cell-cell interactions and communication at the single-cell level. We underscore notable biological applications that have benefited from these technologies and identify pressing challenges that must be addressed in future endeavors leveraging microfluidic tools for single-cell interaction studies.

Metastasis is the primary cause of mortality in small cell lung cancer (SCLC), with the liver being a predominant site for distal metastasis. Despite this clinical significance, mechanisms underlying the interaction between SCLC and liver microenvironment, fostering metastasis, remain unclear.

SCLC patient tissue array, bioinformatics analysis were performed to demonstrate the role of periostin (POSTN) in SCLC progression, metastasis, and prognosis. Cell migration, invasion and sphere formation assay were performed to determine the oncogenic role of POSTN. RNA sequencing analysis was utilized to identify the key signaling pathway regulated by POSTN. Immunoprecipitation, immunofluorescence and co-culture system were used to clarify the mechanism of POSTN-NOTCH1 axis in tumor cells-hepatic stellate cells (HSCs) crosstalk. Subcutaneous xenograft model and liver metastasis model were established to examine the anti-tumor growth and metastases effect of targeting POSTN-NOTCH1 signaling axis.

Elevated expression of POSTN in SCLC is correlated with accelerated tumor progression and metastasis. Conditioned medium rich in POSTN derived from SCLC tumors demonstrates the ability to activate HSCs in the liver microenvironment. Mechanistically, POSTN emerges as a binding partner for the membrane receptor NOTCH1 and transducing the extracellular signals to intracellular fibroblasts. Furthermore, targeting the POSTN-NOTCH1 signaling axis proves effective in suppressing SCLC tumor growth and inhibiting liver metastasis. This study elucidates that the SCLC-derived secreted protein POSTN interacts with NOTCH1 on HSCs to promote the activation of HSCs, thereby providing a favorable microenvironment for liver metastasis.

These findings uncover the intricate communications between primary SCLC cells and HSCs in the tumor microenvironment mediated by the secreted protein POSTN in the context of liver metastasis. Consequently, targeting the POSTN-NOTCH1 signaling axis emerges as a promising therapeutic strategy for metastatic SCLC.

Exosomes are a member of extracellular vesicles. However, their biological characteristics differ from those of other vesicles, and recently, their powerful functions as information molecules, biomarkers, and carriers have been demonstrated. Malignancies are the leading cause of high morbidity and mortality worldwide. The cure rate of malignancies can be improved by improving early screening rates and therapy. Moreover, a close correlation between exosomes and malignancies has been observed. An in-depth study of exosomes can provide new methods for diagnosing and treating tumors. Therefore, this study aimed to review, sort, and summarize such achievements, and present ideas and opinions on the application of exosomes in tumor treatment.

Conventional radiation therapy can restore the ability of cells to undergo immunogenic cell death. Recent preclinical studies suggest that targeted radionuclide therapy, which delivers radiation to tumors at a continuous low dose rate, also stimulates the immune system and offers a promising approach for overcoming resistance to immune checkpoint inhibitors. In this context, we examined the growing body of preclinical and clinical findings showing that the immune system can be activated by the release of extracellular vesicles from irradiated cells, contributing to the antitumor immunity.

The MYC proto-oncogene encodes a basic helix-loop-helix leucine zipper (HLH-LZ) transcription factor, acting as a master regulator of genes involved in cellular proliferation, differentiation, and immune surveillance. Dysregulation of MYC is implicated in over 70% of human cancers, driving oncogenic processes through altered gene expression and disrupted cellular functions. Non-synonymous single nucleotide polymorphisms (nsSNPs) within coding regions can significantly impact protein structure and function, leading to abnormal cellular behaviours. This study employed 29 in silico tools to systematically evaluate the deleteriousness of nsSNPs within the MYC gene. These tools assessed the variants' effects on protein structure, disease association, functional domains, and post-translational modification sites. This study investigated if these variants may disrupt protein-protein interactions, critical for MYC's oncogenic roles and normal cellular functions. Our analysis identified 21 nsSNPs that were predicted to be deleterious and pathogenic. These variants correspond to residues D63H, D63Y, P74L, P75L, N375D, N375I, E378K, E378Q, E378A, E378G, E378V, R379P, R381K, R381T, R382W, L392P, R393C, R393H, R393P, L411H, and L411P. Stability assessments indicated that these variants could destabilise the MYC protein. None of the variants affected post-translational modifications. Protein-protein interaction and docking analysis revealed that variants within bHLH and LZ domains may disrupt MYC/MAX binding, potentially impacting MYC's oncogenic activity and transcriptional regulation. This computational assessment enhances our understanding of genetic variations within the MYC gene and prioritises candidate nsSNPs for experimental validation and therapeutic exploration.

Aberrant expression of repeat RNAs in pancreatic ductal adenocarcinoma (PDAC) mimics viral-like responses with implications on tumor cell state and the response of the surrounding microenvironment. To better understand the relationship of repeat RNAs in human PDAC, we performed spatial molecular imaging at single-cell resolution in 46 primary tumors, revealing correlations of high repeat RNA expression with alterations in epithelial state in PDAC cells and myofibroblast phenotype in cancer-associated fibroblasts (CAFs). This loss of cellular identity is observed with dosing of extracellular vesicles (EVs) and individual repeat RNAs of PDAC and CAF cell culture models pointing to cell-cell intercommunication of these viral-like elements. Differences in PDAC and CAF responses are driven by distinct innate immune signaling through interferon regulatory factor 3 (IRF3). The cell-context-specific viral-like responses to repeat RNAs provide a mechanism for modulation of cellular plasticity in diverse cell types in the PDAC microenvironment.

Extracellular vesicles (EVs) are cell-derived nanoparticles that have attracted significant attention in the investigation of human health and disease, including cancer biology and its clinical management. Concerning cancer, EVs have been shown to influence numerous aspects of oncogenesis, including tumor proliferation and metastasis. EVs can augment the immune system and have been implicated in virtually all aspects of innate and adaptive immunity. With immunotherapy changing the landscape of cancer treatment across multiple disease sites, it is paramount to understand their mechanisms of action and to further improve upon their efficacy. Despite a rapidly growing body of evidence supporting of the utility of EVs in cancer diagnostics and therapeutics, their application in clinical trials involving solid tumors and immunotherapy remains limited. To date, relatively few trials are known to incorporate EVs in this context, mainly employing them as biomarkers. To help address this gap, this review summarizes known applications of EVs in clinical trials and provides a brief overview of the roles that EVs play in cancer biology, immunology, and their proposed implications in immunotherapy. The impetus to leverage EVs in future clinical trials and correlative studies is crucial, as they are ideally positioned to synergize with advancements in multi-omics research to further therapeutic discovery and our understanding of cancer biology.

Prostate cancer (PCa) is one of the most commonly diagnosed malignancies and main causes of cancer-related deaths worldwide. It is characterized by high heterogeneity, ranging from slow-growing tumor to metastatic disease. Since both therapy selection and outcome strongly rely on appropriate patient stratification, it is crucial to differentiate benign from more aggressive conditions using new and improved diagnostic and prognostic biomarkers. Extracellular vesicles (EVs) are membrane-coated particles carrying a specific biological cargo composed of nucleic acids, proteins, and metabolites. Here, we provide an overview of the role of EVs in PCa, focusing on both their biological function and clinical value. Specifically, we summarize the oncogenic role of EVs in mediating the interactions with PCa microenvironment as well as the horizontal transfer of metastatic traits and drug resistance between PCa cells. Furthermore, we discuss the potential usage of EVs as innovative tools for PCa diagnosis and prognosis.

Endogenous viral elements (EVEs) are viral sequences integrated within the host genome that can influence gene regulation and tumor development. While EVEs have been implicated in cancer, their role in regulating key transcription factors in glioblastoma (GBM) remains underexplored. This study investigates the relationship between EVEs and the activation of OCT4, a critical transcription factor in GBM progression.

We utilized CancerHERVdb and HervD Atlas databases to identify potential interactions between EVEs and key genes involved in GBM. Data from 273 GBM patient samples in the TCGA database were analyzed to examine the correlation between OCT4 expression and mutations in glioma-related genes. Furthermore, glioblastoma stem cells (GSCs) were assessed for the expression levels of OCT4 and SOX2, and Pearson correlation analysis was performed.

Our analysis revealed that OCT4 is a pivotal gene activated by EVEs in GBM. OCT4 expression was significantly correlated with mutations in key glioma-associated genes. Higher OCT4 levels were associated with poorer patient prognosis, higher tumor grades, and older age. Additionally, GSCs exhibited elevated expression of both OCT4 and SOX2, with a positive correlation observed between these two genes in GBM patients.

This study highlights the potential role of EVEs in driving GBM progression through the activation of OCT4. The findings emphasize the importance of OCT4 in GBM malignancy and suggest that targeting EVE-mediated pathways may provide new therapeutic approaches for GBM treatment.

Extracellular vesicles (EVs) play an important role in cell-cell communication, carrying bioactive molecules including DNA. EV-associated DNA (EV-DNA) has created enormous interest in the field of biomarkers, particularly related to liquid biopsy. However, its analysis is challenging due to the nanoscale structure of EVs, the low abundance of EV-DNA, and surrounding biogenetic debate. Therefore, novel protocols to enhance the accurate detection of EV-DNA are essential to study its role in normal physiology and disease states. Here, we provide two protocols for confirming the presence of EV-DNA from biological samples. In the first protocol, ultrathin sectioning of EVs is combined with immunogold labeling to detect the presence of double-stranded (ds) DNA within the EV lumen using transmission electron microscopy (TEM). In the second protocol, whole-mount EV immunogold labeling allows detailed morphological analysis of EVs and their surface-associated DNA. Using TEM imaging, we have demonstrated that cancer-cell-derived individual EVs exhibit simultaneous positivity for dsDNA and the EV surface protein tetraspanin 9. We believe that this method can be used to label any proteins of interest inside as well as on the surface of EVs. This can aid in the characterization of single EVs and in the identification and verification of EV-associated biomarkers. © 2024 The Author(s). Current Protocols published by Wiley Periodicals LLC. Basic Protocol 1: EV isolation from cell-culture-conditioned medium, EV embedding, ultrathin sectioning, labeling, and imaging Basic Protocol 2: Whole-mount immunolabeling of EV-DNA.

In the last decade, it has become clear that extracellular vesicles (EVs) are a ubiquitous component of living systems. These small membrane-enclosed particles can confer diverse functions to the cells that release, capture, or coexist with them in an environment. We use examples across living systems to produce a conceptual framework that classifies three modes by which EVs exert functions: (a) EV release that serves a function for producing cells, (b) EV modification of the extracellular environment, and (c) EV interactions with, and alteration of, receiving cells. We provide an overview of the inherent properties of EVs (i.e., their nature) as well as factors in the environment and receiving cell (i.e., nurture) that determine whether transmission of EV cargo leads to functional cellular responses. This review broadens the context for ruminating on EV functions and highlights the emergent properties of EVs that define their role in biology and will shape their applications in medicine.

We present a series of articles proving the existence of a previously unknown mechanism of interaction between hematopoietic stem cells and extracellular double-stranded DNA (and, in particular, double-stranded DNA of the peripheral bloodstream), which explains the possibility of emergence and fixation of genetic information contained in double-stranded DNA of extracellular origin in hematopoietic stem cells. The concept of the possibility of stochastic or targeted changes in the genome of hematopoietic stem cells is formulated based on the discovery of new, previously unknown biological properties of poorly differentiated hematopoietic precursors. The main provisions of the concept are as follows. The hematopoietic stem cell takes up and internalizes fragments of extracellular double-stranded DNA via a natural mechanism. Specific groups of glycocalyx factors, including glycoproteins/proteoglycans, glycosylphosphatidylinositol-anchored proteins and scavenger receptors, take part in the internalization event. The binding sites for DNA fragments are heparin-binding domains and clusters of positively charged amino acid residues that are parts of protein molecules of these factors. Extracellular fragments delivered to the internal compartments of hematopoietic stem cells initiate terminal differentiation, colony formation, and proliferation of hematopoietic precursors. The molecular manifestation of these processes is the emergence and repair of pangenomic single-strand breaks. The occurrence of pangenomic single-strand breaks and restoration of genome (genomic DNA) integrity are associated with activation of a "recombinogenic situation" in the cell; during its active phase, stochastic homologous recombination or other recombination events between extracellular fragments localized in the nucleus and chromosomal DNA are possible. As a result, genetic material of initially extracellular localization either integrates into the recipient genome with the replacement of homologous chromosomal segments, or is transitively present in the nucleus and can manifest itself as a new genetic trait. It is assumed that as a result of stochastic acts of homologous exchange, chromosome loci are corrected in hematopoietic stem cells that have acquired mutations during the existence of the organism, which are the cause of clonal hematopoiesis associated with old age. In this regard, there is a fundamental possibility of changing the hematopoietic status of hematopoietic stem cells in the direction of polyclonality and the original diversity of clones. Such events can form the basis for the rejuvenation of the blood-forming cell system. The results of the laboratory's work indicate that other stem cells in the body capture extracellular DNA fragments too. This fact creates a paradigm for the overall rejuvenation of the body.

Retrotransposons (RTEs) have been postulated to reactivate with age and contribute to aging through activated innate immune response and inflammation. Here, we analyzed the relationship between RTE expression and aging using published transcriptomic and methylomic datasets of human blood. Despite no observed correlation between RTE activity and chronological age, the expression of most RTE classes and families except short interspersed nuclear elements (SINEs) correlated with biological age-associated gene signature scores. Strikingly, we found that the expression of SINEs was linked to upregulated DNA repair pathways in multiple cohorts. We also observed DNA hypomethylation with aging and the significant increase in RTE expression level in hypomethylated RTEs except for SINEs. Additionally, our single-cell transcriptomic analysis suggested a role for plasma cells in aging mediated by RTEs. Altogether, our multi-omics analysis of large human cohorts highlights the role of RTEs in biological aging and suggests possible mechanisms and cell populations for future investigations.

Cell-derived extracellular vesicles (EVs), which carry diverse biomolecules such as nucleic acids, proteins, metabolites, and lipids reflecting their cell of origin, are released under both physiological and pathological conditions. EVs have been demonstrated to mediate cell-to-cell communication and serve as biomarkers. EV-associated DNA (EV-DNA) comprises genomic and mitochondrial DNA (i.e., gDNA and mtDNA) fragments. Some studies have revealed that EV-DNA can represent the full nuclear genome and mitochondrial genome of parental cells. Furthermore, DNA fragments loaded into EVs are stable and can be transferred to recipient cells to regulate their biological functions. In this review, we summarized and discussed EV-DNA research advances with an emphasis on EV-DNA detection at the population-EV and single-EV levels, gene transfer-associated biological functions, and clinical applications as biomarkers for disease liquid biopsy. We hope that this review will provide potential directions or guidance for future EV-DNA investigations.

The clonal evolution model of cancer was developed in the 1950s-1970s and became central to cancer biology in the twenty-first century, largely through studies of cancer genetics. Although it has proven its worth, its structure has been challenged by observations of phenotypic plasticity, non-genetic forms of inheritance, non-genetic determinants of clone fitness and non-tree-like transmission of genes. There is even confusion about the definition of a clone, which we aim to resolve. The performance and value of the clonal evolution model depends on the empirical extent to which evolutionary processes are involved in cancer, and on its theoretical ability to account for those evolutionary processes. Here, we identify limits in the theoretical performance of the clonal evolution model and provide solutions to overcome those limits. Although we do not claim that clonal evolution can explain everything about cancer, we show how many of the complexities that have been identified in the dynamics of cancer can be integrated into the model to improve our current understanding of cancer.

Extracellular vesicles (EVs) have emerged as key players in intercellular communication, facilitating the transfer of crucial cargo between cells. Liquid biopsy, particularly through the isolation of EVs, has unveiled a rich source of potential biomarkers for health and disease, encompassing proteins and nucleic acids. A milestone in this exploration occurred a decade ago with the identification of extracellular vesicle-associated DNA (EV-DNA) in the bloodstream of a patient diagnosed with pancreatic cancer. Subsequent years have witnessed substantial advancements, deepening our insights into the molecular intricacies of EV-DNA emission, detection, and analysis. Understanding the complexities surrounding the release of EV-DNA and addressing the challenges inherent in EV-DNA research are pivotal steps toward enhancing liquid biopsy-based strategies. These strategies, crucial for the detection and monitoring of various pathological conditions, particularly cancer, rely on a comprehensive understanding of why and how EV-DNA is released. In our review, we aim to provide a thorough summary of a decade's worth of research on EV-DNA. We will delve into diverse mechanisms of EV-DNA emission, its potential as a biomarker, its functional capabilities, discordant findings in the field, and the hurdles hindering its clinical application. Looking ahead to the next decade, we envision that advancements in EV isolation and detection techniques, coupled with improved standardization and data sharing, will catalyze the development of novel strategies exploiting EV-DNA as both a source of biomarkers and therapeutic targets.

Extracellular vesicles (EVs) are cell-secreted biological nanoparticles that are critical mediators of intercellular communication. They contain diverse bioactive components, which are promising diagnostic biomarkers and therapeutic agents. Their nanosized membrane-bound structures and innate ability to transport functional cargo across major biological barriers make them promising candidates as drug delivery vehicles. However, the complex biology and heterogeneity of EVs pose significant challenges for their controlled and actionable applications in diagnostics and therapeutics. Recently, DNA molecules with high biocompatibility emerge as excellent functional blocks for surface engineering of EVs. The robust Watson-Crick base pairing of DNA molecules and the resulting programmable DNA nanomaterials provide the EV surface with precise structural customization and adjustable physical and chemical properties, creating unprecedented opportunities for EV biomedical applications. This review focuses on the recent advances in the utilization of programmable DNA to engineer EV surfaces. The biology, function, and biomedical applications of EVs are summarized and the state-of-the-art achievements in EV isolation, analysis, and delivery based on DNA nanomaterials are introduced. Finally, the challenges and new frontiers in EV engineering are discussed.