"Peripheral nerve injury" refers to damage or trauma affecting nerves outside the brain and spinal cord. Peripheral nerve injury results in movements or sensation impairments, and represents a serious public health problem. Although severed peripheral nerves have been effectively joined and various therapies have been offered, recovery of sensory or motor functions remains limited, and efficacious therapies for complete repair of a nerve injury remain elusive. The emerging field of mesenchymal stem cells and their exosome-based therapies hold promise for enhancing nerve regeneration and function. Mesenchymal stem cells, as large living cells responsive to the environment, secrete various factors and exosomes. The latter are nano-sized extracellular vesicles containing bioactive molecules such as proteins, microRNA, and messenger RNA derived from parent mesenchymal stem cells. Exosomes have pivotal roles in cell-to-cell communication and nervous tissue function, offering solutions to changes associated with cell-based therapies. Despite ongoing investigations, mesenchymal stem cells and mesenchymal stem cell-derived exosome-based therapies are in the exploratory stage. A comprehensive review of the latest preclinical experiments and clinical trials is essential for deep understanding of therapeutic strategies and for facilitating clinical translation. This review initially explores current investigations of mesenchymal stem cells and mesenchymal stem cell-derived exosomes in peripheral nerve injury, exploring the underlying mechanisms. Subsequently, it provides an overview of the current status of mesenchymal stem cell and exosome-based therapies in clinical trials, followed by a comparative analysis of therapies utilizing mesenchymal stem cells and exosomes. Finally, the review addresses the limitations and challenges associated with use of mesenchymal stem cell-derived exosomes, offering potential solutions and guiding future directions.

Ischemic stroke is a secondary cause of mortality worldwide, imposing considerable medical and economic burdens on society. Extracellular vesicles, serving as natural nano-carriers for drug delivery, exhibit excellent biocompatibility in vivo and have significant advantages in the management of ischemic stroke. However, the uncertain distribution and rapid clearance of extracellular vesicles impede their delivery efficiency. By utilizing membrane decoration or by encapsulating therapeutic cargo within extracellular vesicles, their delivery efficacy may be greatly improved. Furthermore, previous studies have indicated that microvesicles, a subset of large-sized extracellular vesicles, can transport mitochondria to neighboring cells, thereby aiding in the restoration of mitochondrial function post-ischemic stroke. Small extracellular vesicles have also demonstrated the capability to transfer mitochondrial components, such as proteins or deoxyribonucleic acid, or their sub-components, for extracellular vesicle-based ischemic stroke therapy. In this review, we undertake a comparative analysis of the isolation techniques employed for extracellular vesicles and present an overview of the current dominant extracellular vesicle modification methodologies. Given the complex facets of treating ischemic stroke, we also delineate various extracellular vesicle modification approaches which are suited to different facets of the treatment process. Moreover, given the burgeoning interest in mitochondrial delivery, we delved into the feasibility and existing research findings on the transportation of mitochondrial fractions or intact mitochondria through small extracellular vesicles and microvesicles to offer a fresh perspective on ischemic stroke therapy.

Epidemiological studies on melanoma have revealed significant gender disparities, with the incidence and mortality rates being higher in males than in females. Recent studies indicate that androgen contributing to T cell exhaustion and promoting cancer cell proliferation. While clinical androgen deprivation therapies (ADT),particularly the use of androgen receptor (AR) antagonists to block AR signaling, has been employed in clinical settings to reduce androgen levels, antiandrogen drugs often encounter challenges such as poor targeting and selectivity, increased toxicity, low stability, short half-life and the emergence of drug resistance. Here, we establish a nanoantagonists for efficient AR signaling blockade by arming antigen-activated dendritic cells (DCs) nanovesicles with AR antibodies (aAR-NVOVA). This innovative approach demonstrates dual therapeutic efficacy: aAR-NVOVA effectively disrupts androgen-AR interactions in both melanoma cells and T cells, simultaneously inhibiting tumor proliferation and reversing T cell exhaustion. Furthermore, aAR-NVOVA retains the inherent immunostimulatory properties of DCs, facilitating T cell activation and enhancing cytotoxic T lymphocyte infiltration within tumor tissues. As a result, a synergistic effect has been observed in boosting T cell-based immunotherapy by simultaneously enhancing T cell activity and reducing its exhaustion. Our study using aAR-NVOVA to antagonize androgen effects offers a promising new strategy for enhancing melanoma immunotherapy.

The diagnosis of malignant and benign pulmonary nodules has always been a prominent research topic. Accurately distinguishing between these types of lesions, particularly small or ground glass nodules, is crucial for the early detection and proactive treatment of lung cancer, ultimately leading to improved patient survival. Although various imaging methods and tissue biopsies have advanced the diagnostic efficacy of pulmonary nodules, each approach possesses inherent limitations. In recent years, there has been a growing interest in liquid biopsy as a non‑invasive and easily obtainable alternative. Furthermore, in‑depth investigations into the mechanisms underlying tumor initiation and progression have contributed to the development of circulating biomarkers for monitoring treatment response and efficacy in lung cancer. This review provides a comprehensive overview of the current landscape of pulmonary nodule diagnosis while highlighting the latest advancements in liquid biopsy techniques, such as extracellular vesicles, tumor‑educated platelets, non‑coding RNA, circulating tumor DNA and circulating antibodies.

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.

Cancer affects millions of people, and early detection and efficient treatment are two strong levers to hurdle this disease. Recent studies on exosomes, a subset of extracellular vesicles, have deliberately shown the potential to function as a biomarker or treatment tool, thereby attracting the attention of researchers who work on developing biosensors. Due to the ability of computational methods to predict of the behavior of biomolecules, the combination of experimental and computational methods would enhance the analytical performance of the biosensor, including sensitivity, accuracy, and specificity, even detecting such vesicles from bodily fluids. In this regard, the role of computational methods such as molecular docking, molecular dynamics simulation, and density functional theory is overviewed in the development of biosensors. This review highlights the investigations and studies that have been reported using these methods to design exosome-based biosensors. This review concludes with the role of the quantum mechanics/molecular mechanics method in the investigation of chemical processes of biomolecular systems and the deficiencies in using this approach to develop exosome-based biosensors. In addition, the artificial intelligence theory is explained briefly to show its importance in the study of these biosensors.

Wound healing presents a significant challenge in healthcare, imposing substantial physiological and economic burdens. While traditional treatments and stem cell therapies have shown benefits, milk-derived exosomes (MDEs) offer distinct advantages as a cell-free therapeutic approach. MDEs, isolated from mammalian milk, are characterized by their biocompatibility, ease of acquisition, and high yield, making them a promising tool for enhancing wound repair. This review provides a comprehensive analysis of the composition, sources, and extraction methods of MDEs, with a focus on their therapeutic role in both acute and diabetic chronic wounds. MDEs facilitate wound healing through the delivery of bioactive molecules, modulating key processes such as inflammation, angiogenesis, and collagen synthesis. Their ability to regulate complex wound-healing pathways underscores their potential for widespread clinical application. This review highlights the importance of MDEs in advancing wound management and proposes strategies to optimize their use in regenerative medicine.

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.

Cardiovascular disease (CVD) remains one of the leading causes of high mortality and morbidity worldwide, posing a substantial threat to global health. Mesenchymal stem cell (MSC) therapy has emerged as a promising treatment approach, primarily through the secretion of various bioactive factors. Exosomes (Exos), in particular, stand out as the most effective components, as their noncoding RNA and proteins play a crucial role in promoting the repair of cardiac function, positioning them a promising cell-free therapy for CVD. However, challenges such as poor stability, low delivery efficiency, weak targeting, and rapid immune-mediated clearance hinder the broader application of Exos, presenting significant obstacles for further clinical translation. Recent advancements in biomaterials, particularly hydrogels, offer new avenues for Exos-based CVD therapies. Hydrogels, with their ability to improve stability, release control, and targeting, have gained considerable attention in the biomedical field. This review explores the latest research developments regarding the treatment of CVD using Exos, and highlights their synergistic application with hydrogels, which provide valuable insights for advancing Exos-based therapies and developing novel therapeutic strategies for CVD.

Cardiovascular diseases (CVDs) remain the leading cause of death worldwide, both in developed and developing countries. Despite the implementation of various measures in clinical practice that have shown certain curative effects, poor prognosis and irreversible pathological cardiac remodeling continue to limit the therapeutic effect of CVDs. There are still many new mechanisms worth exploring for the regulation of CVDs. Previous studies have highlighted the potential applicability of exosomes in CVDs, and significant research has been conducted in this area. In this review, we summarize the physiological mechanisms of exosomes and the basic research achievements in regulating CVDs via exosomal non-coding RNAs. We also discuss the limitations and prospects of exosome application in CVD treatment.

Infectious diseases continue to present significant global public health challenges, with pathogens such as bacteria and viruses posing substantial threats to human health. Conventional therapeutic approaches face several limitations, including the rising prevalence of drug resistance, suboptimal targeting, and adverse side effects, which collectively complicate clinical management. Cell membrane vesicles (MVs), characterized by their natural biocompatibility and outstanding drug delivery capabilities, have emerged as a promising platform for addressing these challenges in the treatment of infectious diseases. To further augment the therapeutic potential of MVs, engineering modifications have been extensively employed to enhance their functionality and efficacy. This review provides a comprehensive overview of the production and modification techniques associated with MVs, emphasizing recent advancements in the development of engineered membrane vesicles (EMVs) as versatile nanoplatforms for combating infectious diseases. Additionally, the clinical prospects and existing challenges of EMVs are critically analyzed, and recommendations are proposed to guide future research and facilitate their clinical translation into practical applications in combating infectious disease.

Adipose stem cells (ADSC) have demonstrated therapeutic potential in ameliorating obesity and metabolic disorders, with their exosomes showing comparable therapeutic effects. However, the underlying molecular mechanism remains incompletely understood. Furthermore, the limited availability and inherent heterogeneity of primary ADSC present substantial challenges for consistent therapeutic outcomes.

This study aimed to investigate the molecular mechanism underlying the unique biological effects mediated by microRNAs (miRNAs) in exosomes derived from immortalized adipose stem cells (iADSC).

We first established stable iADSC and isolated their exosomes (iADSC-EXO), which exhibited both high yield and stability. In high fat diet (HFD)-fed obese mice, iADSC-EXO administration significantly attenuated obesity, reduced blood glucose and lipid levels, and alleviated hepatic steatosis. miRNA chips analysis revealed that miR-1246 is highly enriched in exosomes derived from iADSC and ADSC. Administration of miR-1246 to HFD-fed obese mice produced beneficial effects on obesity and metabolic disorders comparable to those with iADSC-EXO. Mechanistic studies revealed that miR-1246 exerts its beneficial effects through multiple pathways: First, reducing fat mass by downregulating of the expression of fat mass and obesity-associate protein (FTO) and subsequent inhibition of adipogenesis and lipogenesis. Second, enhancing white adipose tissue (WAT) beiging by suppressing Runt-related transcription factor 1 (RUNX1T1) and FTO expression. Third, promoting M2 macrophage polarization and suppressing WAT inflammation via inhibition of TNF receptor-associated factor 6 (TRAF6)-mediated inflammatory pathway.

Our study elucidates a novel molecular mechanism through which ADSC regulates adipose tissue homeostasis and metabolism and presents a potential therapeutic approach for treating obesity and related metabolic disorders via iADSC-EXO or miR-1246-based interventions.

Recombinant proteins, cell, and gene therapies are collectively defined as biological drugs or biologics. These therapies have transformed the lives of millions of patients over the past decades, with the number of FDA-approved biologics increasing exponentially in recent years. However, out of approximately 700 biological therapies approved by the FDA in the last 20 years, less than 1% are indicated for cardiac pathologies. The application of biologics in cardiovascular disease has faced significant challenges, including short plasma half-life, the multifactorial complexity of cardiac disease, and the lack of efficient, non-invasive, and patient-friendly drug-delivery routes. This translational gap is particularly pressing given the immense socioeconomic burden of cardiovascular disease, which remains the leading cause of death globally and accounts for billions in annual healthcare costs and lost productivity. Inhalation-based drug delivery has recently emerged as a promising strategy for treating cardiovascular disease, with several proof-of-concept studies demonstrating its potential in heart failure, the most prevalent cardiac condition. This narrative review summarizes the latest experimental evidence in the novel field of Cardiovascular Inhalation, i.e., the lung-to-heart route for biologics. We discuss translational challenges, preclinical evidence, and future perspectives for bringing this innovative approach to clinical practice.

Exosomes, a subtype of extracellular vesicles, are increasingly recognized as promising biomarkers for human cancers. Rapid detection and classification of esophageal cancer-associated exosomes could significantly improve non-invasive screening for potential patients. This study aims to establish a label-free, direct surface-enhanced Raman scattering (SERS) method to capture characteristic molecular information from both esophageal cancer cells and their corresponding exosomes using confocal Raman microscopy. The results revealed distinct Raman spectra for esophageal cancer cells and their exosomes within the range of 500-1600 cm-1, with notable signal similarities observed at 506-622, 778-832, 1079-1098, and 1572-1630 cm-1. In contrast, significant differences were identified in Raman peaks related to nucleic acids (723, 654, 1354 cm-1) and proteins (998, 1028, 1354, 1560 cm-1). An orthogonal partial least squares discriminant analysis (OPLS-DA) model was utilized to discern subtle variations among these highly similar samples, achieving an accuracy rate of 100%. By comparing the spectral correlations between esophageal cancer cells and their exosomes, this study provides valuable insights into the molecular composition and cellular origins of exosomes. The findings demonstrate the potential of integrating SERS with OPLS-DA for the precise and rapid detection and monitoring of esophageal cancer through exosomal biomarkers, offering a powerful tool for diagnostic applications.

Plant-derived extracellular vesicles (PDEVs) have emerged as innovative nanocarriers for drug delivery, offering advantages such as biocompatibility, stability, and cost-effectiveness. This study explores light-mediated strategies to optimize cargo encapsulation into PDEVs while preserving structural integrity. Leveraging the intrinsic photosensitizing properties of PDEVs, light irradiation (LED) triggered reactive oxygen species (ROS) generation, including superoxide anions and singlet oxygen, which transiently enhanced membrane permeability for controlled drug loading. Using FITC-dextran (70 kDa) as a model cargo, we optimized light-induced loading efficiency, achieving a peak (~ 80%) at 10 min of irradiation. Prolonged exposure (15 min) reduced efficiency (~ 50%), likely due to excessive ROS-induced membrane destabilization. The optimal PDEVs-to-cargo ratio (1:30) ensured maximal loading while maintaining stability over 30 days. Lipid peroxidation analysis further confirmed ROS-induced membrane modifications through malondialdehyde (MDA) accumulation. These findings demonstrate that PLDENs (Pueraria lobate-derived exosomes-like nanovesicles) function as light-responsive nanocarriers, balancing ROS-mediated permeability enhancement with structural integrity. This light-triggered strategy balances permeability modulation and structural integrity, advancing PDEVs as scalable, non-invasive platforms for precision drug delivery and photodynamic applications.

Despite decades of research, effective disease-modifying treatments for Amyotrophic Lateral Sclerosis (ALS) remain scarce. The emergence of regenerative medicine presents a new frontier for ALS treatment.

This review is based on a comprehensive literature search using PubMed, Scopus and clinical trials databases on the recent therapeutic advancements in ALS, giving focus to regenerative medicine. The article includes coverage of stem cell-based therapies, including mesenchymal, neural and induced pluripotent stem cells; all of which may offer potential neuroprotective and immunomodulatory effects. Gene therapy, particularly antisense oligonucleotides targeting ALS-related mutations, has gained traction, with tofersen becoming the first FDA-approved genetic therapy for ALS. The article also covers emerging approaches such as extracellular vesicles, immune-modulating therapies, and bioengineering techniques, including CRISPR-based gene editing and cellular reprogramming, that hold promise for altering disease progression.

While regenerative medicine provides hope for ALS patients, significant challenges remain. Biomarkers will play a crucial role in guiding personalized treatment strategies, ensuring targeted interventions. Future research should prioritize optimizing combinatory approaches, integrating different therapy strategies to maximize patient outcomes. Although regenerative medicine is still in its early clinical stages, its integration into ALS treatment paradigms could redefine disease management and alter its natural course.

Extracellular vesicles (EVs) are heterogeneous nanoscale membrane vesicles released by almost all cell types into the circulation. Depending on their biogenesis and cells of origin, EVs show considerable heterogeneity in their biophysical and biomolecular composition and can serve as reflective and dynamic blood biomarkers for personalized medicine. Conventional analytical technologies, however, often lack the compatibility to reveal nanoscale EV features and resolve vesicle heterogeneity. The past decade has since witnessed the development of various nanoplasmonic technologies to empower EV analysis, through bulk and single-vesicle characterization, at an unprecedented scale and resolution. These platforms achieve versatile measurements that are not only size-matched to EV dimensions but can also probe multiplexed biomolecular contents, thereby providing new insights into EV heterogeneity and enabling transformative clinical opportunities. In this review, key characteristics of EVs and their remarkable heterogeneity are introduced. The sensing principles of plasmonic platforms are also discussed, with recent technology developments highlighted to resolve EV heterogeneity, through bulk analyses of EV subpopulations as well as high-resolution single-EV measurements. An outlook is further provided on emerging opportunities, at the interface of biomarker discovery and technology innovation, to develop empowering nanoplasmonic EV platforms for personalized medicine. biosensing; bulk analysis; extracellular vesicles; nanoplasmonics; single-vesicle analysis.

Exosomes, nanosized extracellular vesicles carrying proteins, lipids, and nucleic acids, hold great potential in therapeutic applications. Cryopreservation, a widely used method for their preservation and transport, often causes irreversible damage. Understanding the molecular mechanisms underlying biomembrane resistance to cryodamage is crucial for advancing cryopreservation techniques. In this study, we find that tetraspanin 4 (TSPAN4) and other tetraspanin family proteins play an essential role in protecting exosomes from cryodamage, likely due to their role in cholesterol binding and membrane microdomain formation. Furthermore, we engineered TSPAN4-loaded exosomes, which demonstrated enhanced cryoprotection while maintaining a similar protein composition and uptake efficiency compared to wild-type exosomes. Our novel cryopreservation strategy, which does not rely on external agents, offers a promising approach for advancing the clinical translation of exosomes as therapeutic agents.

The utilization of biological therapeutic agents in the treatment of tendon injuries represents a promising avenue, with particular attention drawn to adipose mesenchymal stem-cell-derived exosomes (ADSCs-exos) owing to their pivotal role in regenerative medicine. Identifying a therapeutic strategy to prolong exosome retention at the injury site for effective tendon repair remains challenging. In this study, we explored the potential of ADSC-exosomes in vitro, demonstrating their ability to promote the behavior of tendon stem/progenitor cells (TSPCs). Additionally, we designed a fibroin (SF) sponge as a biodegradable platform for enzyme-responsive exosome delivery. Subsequently, we used biodegradable SF sponges to deliver ADSC exosomes into the patellar tendon defect in rats. The results showed that, in vivo, exosomes were gradually released from the SF sponges, remained in the defect area for an extended period, and exerted functional benefits locally. These findings were supported by the upregulation of tendon-associated protein expression and improved mechanical properties observed in the in vivo specimens. In summary, we substantiated the advantageous role of ADSCs-exos in facilitating tendon regeneration. Moreover, the utilization of a SF-exos sponge delivery system emerged as an efficacious local treatment strategy for exosome delivery. These findings hold promise for the future application of exosomes in innovative therapies tailored to address tendon injuries.

Osteoarthritis (OA) is a chronic degenerative joint disease characterized by low-grade inflammation, cartilage breakdown, and persistent pain. Despite its prevalence, current therapeutic strategies primarily focus on symptom management rather than modifying disease progression. Monoclonal antibodies and cytokine inhibitors targeting inflammatory pathways, including TNF-α and IL-1, have shown promise but remain limited by inconsistent efficacy and safety concerns. Long-acting biologic therapies-ranging from extended-release formulations, such as monoclonal antibodies and cytokine inhibitors, to gene therapy approaches-have emerged as promising strategies to enhance treatment durability and improve patient outcomes. Extracellular vesicles (EVs) have gained particular attention as a novel delivery platform due to their inherent stability, biocompatibility, and ability to transport therapeutic cargo, including biologics and immunomodulatory agents, directly to joint tissues. This review explores the evolving role of EVs in OA treatment, highlighting their ability to extend drug half-life, improve targeting, and modulate inflammatory responses. Additionally, strategies for EV engineering, including endogenous and exogenous cargo loading, genetic modifications, and biomaterial-based delivery systems, are discussed.

Immune regulation is recognized as a cornerstone therapeutic strategy for the treatment of various autoimmune diseases. These disorders, driven by dysregulated immune responses, contribute significantly to morbidity and mortality. Although conventional immunosuppressive therapies provide symptomatic relief, their prolonged use is often associated with severe adverse effects, underscoring the need for safer and more effective treatment approaches. Extracellular vesicles (EVs), derived from immunoregulatory cells such as regulatory T cells, dendritic cells, mesenchymal stem cells, and neutrophils, have emerged as promising candidates for targeted immunomodulation. These nanoscale vesicles inherit the immunosuppressive properties of their parental cells, thereby facilitating immune homeostasis while mitigating the risks associated with other cell-based therapies. This review provides a comprehensive overview of recent advances in the application of immunoregulatory cell-derived EVs for autoimmune disease treatment, with a particular focus on their mechanisms of action within the immune microenvironment. Finally, we discuss the challenges and potential future directions in the development of EV-based therapies for autoimmune diseases.

Surface plasmons, a unique optical phenomenon arising at the interface between metals and dielectrics, have garnered significant interest across fields such as biochemistry, materials science, energy, optics, and nanotechnology. Recently, plasmonics is evolving from a focus on "classical plasmonics," which emphasizes fundamental effects and applications, to "integrative plasmonics," which explores the integration of plasmonics with multidisciplinary technologies. This review explores this evolution, summarizing key developments in this technological shift and offering a timely discussion on the fusion mechanisms, strategies, and applications. First, we examine the integration mechanisms of plasmons within the realm of optics, detailing how fundamental plasmonic effects give rise to optical multi-effects, such as plasmon-phonon coupling, nonlinear optical effects, electromagnetically induced transparency, chirality, nanocavity resonance, and waveguides. Next, we highlight strategies for integrating plasmons with technologies beyond optics, analyzing the processes and benefits of combining plasmonics with acoustics, electronics, and thermonics, including comprehensive plasmonic-electric-acousto-thermal integration. We then review cutting-edge applications in biochemistry (molecular diagnostics), energy (harvesting and catalysis), and informatics (photonic integrated circuits). These applications involve surface-enhanced Raman scattering (SERS), surface-enhanced infrared absorption (SEIRA), surface-enhanced fluorescence (SEF), chirality, nanotweezers, photoacoustic imaging, perovskite solar cells, photocatalysis, photothermal therapy, and triboelectric nanogenerators (TENGs). Finally, we conclude with a forward-looking perspective on the challenges and future of integrative plasmonics, considering advances in mechanisms (quantum effects, spintronics, and topology), materials (Dirac semimetals and hydrogels), technologies (machine learning, edge computing, in-sensor computing, and neuroengineering), and emerging applications (5G, 6G, virtual reality, and point-of-care testing).

The increasing number of new cancer cases and cancer-related deaths worldwide highlights the urgent need to develop novel anti-tumor treatment methods to alleviate the current challenging situation. Nearly all organisms are capable of secreting extracellular vesicles (EVs), and these nano-scale EVs carrying biological molecules play an important role in intercellular communication, further affecting various physiological and pathological processes. Notably, EVs from different sources have differences in their characteristics and functions. Consequently, diverse EVs have been utilized as drug or vaccine delivery carriers for improving anti-tumor treatment due to their good safety, ease of modification and unique properties, and achieved satisfactory results. Meanwhile, the clinical trials of EV-based platform for tumor therapy are also continuously being conducted. Therefore, in this review, we summarize the recent research progress of EV-based tumor treatment methods, including the introduction of main sources and unique functions of EVs, the application of EVs in tumor treatment as well as their prospects and challenges. Additionally, considering the unique advantages of artificial EVs over natural EVs, we also highlighted their characteristics and applications in tumor treatments. We believe that this review will help researchers develop novel EV-based anti-tumor platforms through a bottom-up design and accelerate the development in this field.

This study aimed to evaluate the potential of plasma-derived extracellular vesicles (EVs) as drug delivery carriers by employing two drug-loading techniques: coincubation and freeze-thaw cycles.

EVs isolated via the polyethylene glycol (PEG) precipitation method were characterized via nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM). The size of the particles was 200.1 ± 66.6 nm. The isolated vesicles were loaded with 1000 µg/ml snake venom L amino acid oxidase (SVLAAO) via the coincubation method and subjected to freeze-thaw cycles to prepare a novel formulation. The encapsulation efficiency (EE) of the loaded EVs was analysed at 30 and 60 min, and in vitro drug release profiles were evaluated for both methods and kinetic model for the same was determined.

The coincubation method achieved an EE of 58.08 ± 0.060% after 60 min, which was greater than that of the freeze-thaw method (55.80 ± 0.060%). Drug release studies demonstrated that 93% of the drug was released in 8.5 h by the coincubation method, whereas the freeze-thaw method resulted in faster release (99% in 6.5 h) due to membrane disruption. The best fit value (R2) was highest for zero order kinetics model.

In conclusion, the coincubation method preserves EV membrane integrity, enabling sustained drug release, making it a promising strategy for targeted drug delivery applications. This study highlights plasma-derived EVs as innovative carriers for therapeutic delivery.

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.

Metabolic-dysfunction-associated steatohepatitis (MASH) remains a major health challenge. Herein, we identify sphingomyelin phosphodiesterase 3 (SMPD3) as a key driver of hepatic ceramide accumulation through increasing sphingomyelin hydrolysis at the cell membrane. Hepatocyte-specific Smpd3 gene disruption or pharmacological inhibition of SMPD3 alleviates MASH, whereas reintroducing SMPD3 reverses the resolution of MASH. Although healthy livers express low-level SMPD3, lipotoxicity-induced DNA damage suppresses sirtuin 1 (SIRT1), triggering an upregulation of SMPD3 during MASH. This disrupts membrane sphingomyelin-ceramide balance and promotes disease progression by enhancing caveolae-dependent lipid uptake and extracellular vesicle secretion from steatotic hepatocytes to exacerbate inflammation and fibrosis. Consequently, SMPD3 acts as a central hub integrating key MASH hallmarks. Notably, we discovered a bifunctional agent that simultaneously activates SIRT1 and inhibits SMPD3, which shows significant therapeutic potential in MASH treatment. These findings suggest that inhibition of hepatic SMPD3 restores membrane sphingolipid metabolism and holds great promise for developing novel MASH therapies.

Blood transfusion is an irreplaceable clinical treatment. Blood components are differentiated and stored according to specific guidelines. Storage temperatures and times vary depending on the blood component, but they all release extracellular vesicles (EVs) during storage. Although blood transfusions can be life-saving, they can also cause many adverse transfusion reactions, among which the effects of EVs are of increasing interest to researchers. EVs are submicron particles that vary in size, composition, and surface biomarkers, are encapsulated by a lipid bilayer, and are not capable of self-replication. EVs released by blood cells are important contributors to pathophysiologic states through proinflammatory, coagulant, and immunosuppressive effects, which in turn promote or inhibit the associated disease phenotype. Therefore, this review explores the potential mechanisms of hematopoietic-derived EVs in transfusion-associated adverse reactions and discusses the potential of the latest proteomics tools to be applied to the analysis of EVs in the field of transfusion medicine with a view to reducing the risk of blood transfusion.

Exosome acts as an outstanding biomarker for ongoing studies, diagnosis and prognosis of multiples diseases. Therefore, the call for economically and efficiently isolating a large number of exosomes is an active area of investigation. However, to date, the challenges including complex isolated procedure, uneconomical equipment, low protein content and distinct loss in the particle number of exosomes etc. still encounter in exosome isolation. Polyethylene glycol (PEG)-induced exosome isolation increasingly attracts wide attention of scientists. PEG precipitation reveals higher performance in the yield of exosomes among multiple common isolation techniques. PEG-based precipitation is a temporarily low-purity, but inexpensive, time-save, labor-less, convenient and high-yield technique to gain exosomes with high biological activities. Hence, the PEG-based exosome isolation approach wins the endorsement of experimental workers. Herein, we summary the existing knowledge on procedures of PEG-based exosomes separation from different biospecimens, the binding process of PEG to exosomes, some notices, demerits, merits of PEG-based exosome isolation, and at last the advantages by combining PEG-precipitation to other techniques for exosome isolation, with a view to eliciting profound insights for investigators who recruit PEG for exosome separation, and advancing references for the standardization of PEG-based exosome isolation in future.

Extracellular vesicles (EVs) are small particles released by various cells, including cancer cells. They play a significant role in the development of different cancers, including brain metastasis. These EVs transport biomolecular materials such as RNA, DNA, and proteins from tumour cells to other cells, facilitating the spread of primary tumours to the brain tissue. EVs interact with the endothelial cells of the blood-brain barrier (BBB), compromising its integrity and allowing metastatic cells to pass through easily. Additionally, EVs interact with various cells in the brain's microenvironment, creating a conducive environment for incoming metastatic cells. They also influence the immune system within this premetastatic environment, promoting the growth of metastatic cells. This review paper focuses on the research regarding the role of EVs in the development of brain metastasis, including their impact on disrupting the BBB, preparing the premetastatic environment, and modulating the immune system. Furthermore, the paper discusses the potential of EVs as diagnostic and prognostic biomarkers for brain metastasis.

The 26S proteasome holoenzyme comprises 20S catalytic and 19S regulatory complexes. Accumulating evidence suggests that the majority of proteasomes in the extracellular space exist as free 20S proteasomes; however, their origin and pathophysiological function remain to be determined. Here, we report that cellular proteasomes are effectively packaged into the lumen of extracellular vesicles (EVs) and secreted in a structurally intact and enzymatically active 20S form. We further demonstrate that EV-encapsulated 20S proteasomes are delivered to recipient cells and facilitate the degradation of overexpressed tau proteins without disrupting global proteolytic pathways. These findings highlight a novel cell-to-cell communication system that transports the proteasomes to target cells for the clearance of proteotoxic substrates. Further characterisation of this homeostatic mechanism will improve our understanding of organismal stress response mechanisms and may provide a therapeutic approach to treat various proteinopathies, including Alzheimer's disease.

Cell communication is crucial for coordinating physiological functions in multicellular organisms, with exosomes playing a significant role. Exosomes mediate intercellular communication by transporting proteins, lipids, and nucleic acids between cells. These small, membrane-bound vesicles, derived from the endosomal pathway, are integral to various biological processes, including signal transmission and cellular behavior modulation. Recent advances highlight the potential of exosomes, especially dendritic cell-derived exosomes (DEXs), for diagnostic and therapeutic applications, particularly in cancer immunotherapy. DEXs are distinguished by their ability to present antigens and stimulate immune responses more effectively than exosomes from other cell types. They carry a cargo rich in immunostimulatory molecules and MHC-peptide complexes, which facilitate robust T-cell activation and enhance tumor-specific immune responses. The unique properties of DEXs, such as their ability to cross biological barriers and resist tumor-induced immunosuppression, position them as promising candidates for therapeutic applications. Here, I review the reports on the bidirectional interaction between dendritic cells and T cells through exosomes and their role in medicine.

Glioblastoma is the most common and aggressive brain tumor with a low survival rate. Due to its heterogeneous composition, high invasiveness, and frequent recurrence after surgery, treatment success has been limited. In addition, due to the brain's unique immune status and the suppressor tumor microenvironment (TME), glioblastoma treatment has faced more challenges. Exosomes play a critical role in cancer metastasis by regulating cell-cell interactions that promote tumor growth, angiogenesis, metastasis, treatment resistance, and immunological regulation in the tumor microenvironment. This review explores the pivotal role of exosomes in the development of glioblastoma, with a focus on their potential as non-invasive biomarkers for prognosis, early detection and real-time monitoring of disease progression. Notably, exosome-based drug delivery methods hold promise for overcoming the blood-brain barrier (BBB) and developing targeted therapies for glioblastoma. Despite challenges in clinical translation, the potential for personalized exosome = -054321`therapies and the capacity to enhance therapeutic responses in glioblastoma, present intriguing opportunities for improving patient outcomes. It seems that getting a good and current grasp of the role of exosomes in the fight against glioblastoma would properly serve the scientific community to further their understanding of the related potentials of these biological moieties.

Glioblastoma is a brain cancer with a poor prognosis. Failure of classical chemotherapy and surgical treatments indicates that new therapeutic approaches are needed. Among cell-free options, exosomes are versatile extracellular vesicles (EVs) that carry important cargo across barriers such as the blood-brain barrier (BBB) to their target cells. This makes exosomes an interesting option for the treatment of glioblastoma. Moreover, exosomes can comprise many therapeutic cargos, including lipids, proteins, and nucleic acids, sampled from special intercellular compartments of their origin cell. Cells exposed to various immunomodulatory stimuli can generate exosomes enriched in specific therapeutic molecules. Notably, the secretion of exosomes could modify the immune response in innate and adaptive immune systems. For instance, glioblastoma-associated exosomes (GBex) uptake by macrophages could influence macrophage dynamics (e.g., shifting CD markers expression). Expression of critical immunoregulatory proteins such as cytotoxic T-lymphocyte antigen-1 (CTLA1) and programmed death-1 (PD-1) on GBex indicates the direct crosstalk of these nano-size vesicles with the immune system. The present study reviews the role of exosomes in immune system cells, including B cells, T cells, natural killer (NK) cells, and dendritic cells (DCs), as well as novel technologies in the field.

Human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-ex) have emerged as a promising alternative to whole-cell therapies due to their minimal immunogenicity and tumorigenicity. Pentraxin 3 (PTX3) acts as an inflammatory marker and pattern recognition receptor, playing a critical role in promoting tumor progression and inflammatory diseases. Fibrotic scars resulting from cerebral hemorrhage can impair motor and sensory functions, leading to poor prognosis. This study aimed to investigate whether hUCMSC-ex regulate matrix metalloproteinase-3 (MMP3) expression via the PTX3/Toll-like receptor 4 (TLR4)/nuclear factor κB (NF-κB) pathway, thereby inhibiting inflammation and fibrotic scar formation following intracerebral hemorrhage and ultimately promoting the recovery of nerve function. A stereotactic technique was used to inject type IV collagenase (1 μL) into the striatum of rats, establishing an animal model of hemorrhagic stroke. Concurrently, hUCMSC-ex were administered via the tail vein at a dosage of 200 μg. In vitro, primary astrocytes were treated with hUCMSC-ex and subsequently stimulated with Hemin (20 μmol/mL) to create a cellular model of cerebral hemorrhage. The expression levels of PTX3, TLR4/NF-κB/MMP3 pathway proteins, and inflammatory factors, including tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-10 (IL-10), were assessed both in vivo and in vitro to investigate the effects of hUCMSC-ex on the inflammatory response and fibroblast migration. Neurological function in rats with cerebral hemorrhage was evaluated on days 1, 3, and 5 using the corner turn test, forelimb placement test, Longa score, and Bederson score. Additionally, real-time PCR was utilized to measure PTX3 mRNA expression following treatment with hUCMSC-ex. hUCMSC-ex inhibited MMP3 expression by downregulating the protein levels of PTX3, TLR4, NF-κB/P65, and p-P65. This action resulted in a reduction of pro-inflammatory cytokines TNF-α and IL-1β while simultaneously increasing the expression of the anti-inflammatory cytokine IL-10. Furthermore, hUCMSC-ex suppressed the inflammatory response, prevented fibroblast migration, and decreased MMP3 expression in the conditioned medium derived from primary astrocytes. Importantly, hUCMSC-ex improved behavioral performance in rats with intracerebral hemorrhage (ICH). hUCMSC-ex modulated the expression of MMP3 through the downregulation of PTX3, TLR4, NF-κB/P65, and p-P65. This regulatory mechanism contributed to a decrease in pro-inflammatory cytokines TNF-α and IL-1β, while concurrently enhancing the expression of the anti-inflammatory cytokine IL-10. Additionally, hUCMSC-ex effectively suppressed the inflammatory response, inhibited fibroblast migration, and reduced MMP3 expression in primary astrocyte-conditioned medium. Overall, hUCMSC-ex significantly improved behavioral performance in rats with ICH.

Natural killer (NK) cells are widely involved in the field of tumor immunotherapy due to their unique killing ability. However, the durability and efficacy of NK-cell monotherapy are facing great challenges owing to the limitation of immunosuppressive tumor microenvironment (TME). NK cell-derived exosomes (Neo) not only play an innate immunomodulatory role similar to NK cells but also emerge as promising antitumor nanocarriers. In this study, an engineered Neo (R@Neo-MN) was designed that encapsulates the multifunctional antitumor drug (Raddeanin a, RA) and modified with maleimide (Mal, M) and mannose (Man, N). The obtained R@Neo-MN could not only exert NK cell-like antitumor function but also induce the immunogenic cell death of tumors to release tumor-associated antigens (TAAs). Furthermore, R@Neo-MN activated the cyclic guanosine monophosphate-adenosine monophosphate synthase/interferon gene stimulator (cGAS/STING) to release type I interferons (IFN). Then, R@Neo-MN could capture TAAs through Mal and specifically target dendritic cells (DCs) through Man, thereby promoting the maturation of DCs and enhancing tumor-specific cytotoxic T-cell (CTL)-mediated adaptive immunity. The released IFN further promoted the infiltration and activition of NK cells and CTLs at the tumor site. Our study suggested a novel strategy that harnesses both innate and adaptive immunity for enhanced tumor immunotherapy.

Cancer vaccines hold promise for tumor immunotherapy, with their success hinging on effective systems to boost anti-tumor immunity. Biological membranes are not only a delivery vehicle but also a source of antigens and adjuvants, garnering growing interest in vaccine research. This review starts with an introduction to the composition and mechanisms of cancer vaccines and describes the sources, advantages/disadvantages, engineering strategies, and applications of these membrane-based platforms for cancer vaccine development. This review also offers a critical analysis and discusses the further direction of the vaccine platform in view of clinical translation for tumor immunotherapy.

Exosome-based therapeutics have garnered significant attention for intracerebral hemorrhage (ICH) treatment due to their capacity to regulate metabolic dysregulation, restore cellular homeostasis, and modulate the injury microenvironment via bioactive cargoes such as microRNAs and proteins. However, rapid systemic clearance and enzymatic degradation critically limit their therapeutic efficacy. To address this challenge, we engineered a three-dimensional (3D) bioprinted scaffold capable of encapsulating and sustaining the release of human umbilical cord mesenchymal stem cell-derived exosomes (hUCMSC-exos).

Based on previous research [1-3], the scaffold was composed of a decellularized brain matrix (dECM), gelatin-methacryloyl (GelMA), and silk fibroin (SF) crosslinked with a photoinitiator. hUCMSC-exos were precisely incorporated via extrusion-based 3D bioprinting. Release kinetics were assessed via in vitro elution and in vivo imaging. An ICH rat model received stereotaxic implantation of the exosome-laden scaffold (dECM@exo). Neuroinflammatory markers (IL-6, TNF-α, IL-10) and apoptotic activity (JC-1, Annexin V/PI, TUNEL) were quantified. Neurological outcomes were longitudinally evaluated using the modified Longa scale, Bederson scoring, and sensorimotor tests (rotarod, forelimb placement) at 1, 4, 7 and 14 days post-ICH.

dECM@exo demonstrated sustained exosome release over 14 days, significantly promoting neural tissue regeneration while attenuating perihematomal edema. Mechanistically, the scaffold modulated pathological MMP activity and inflammatory cytokine expression, thereby restoring extracellular matrix homeostasis and reducing neuronal apoptosis.

The findings demonstrate that the 3D biological scaffold dECM@exo effectively maintains microenvironmental homeostasis in the early stages of ICH and improves outcomes associated with the condition. dECM@exo is poised to serve as a robust platform for drug delivery and biotherapy in ICH treatment, offering a viable alternative for managing this condition.

Migrasomes are newly identified organelles that were first discovered in 2015. Since then, their biological structure, formation process, and physiological functions have been gradually elucidated. Research in recent years has expanded our understanding of these aspects, highlighting their significance in various physiological and pathological processes. Migrasomes have been found to play crucial roles in normal physiological functions, including embryonic development, vascular homeostasis, material transport, and mitochondrial quality control. Additionally, emerging evidence suggests their involvement in various diseases; however, clinical research on their roles remains limited. Current studies indicate that migrasomes may contribute to disease pathogenesis and hold potential for diagnostic and therapeutic applications. This review consolidates existing clinical research on migrasomes, focusing on their role in disease mechanisms and their use in medical applications. By examining their biological structure and function, this review aims to generate insights that encourage further research, ultimately contributing to advancements in disease prevention and treatment.

Extracellular vesicles can play an important role in the processes occurring after stem cell transplantation, preventing cell apoptosis, stimulating immunological processes, and promoting the synthesis of extracellular matrix. Human follicular fluid (FF) can be a source of a subpopulation of cells with mesenchymal stem cells (MSCs) properties. Moreover these subpopulations of FF cells can differentiate into osteoblasts. In presented studies flow cytometry of ovarian FF cells confirmed positive expression of MSCs markers such as: CD44, CD90, CD105, CD73 and negative expression of a hematopoietic marker: CD45. The CD90+, CD105+, CD45- cell subpopulation has been obtained during magnetic separation using appropriate antibodies conjugated with microbeads. The extracellular vesicles (EVs) secreted by the cells during osteodifferentiation process differed from those secreted by cells culture in the basal medium. Based on the previous and current electron microscopy research, changes in size, number, and shape would support the notion that released EVs could be crucial to the ovarian FF cell subpopulation differentiation process. Osteogenic differentiation has been confirmed via Alizarin red staining. Therefore, follicular fluid (FF) can be a new source of a cell subpopulation with MSC properties, with the cells capable of differentiating into the osteogenic lineage. EVs could play a key role as mediators in tissue regeneration, especially bone tissue regeneration.

Although cancer chemopreventive agents have been confirmed to effectively protect high-risk populations from cancer invasion or recurrence, only over ten drugs have been approved by the U.S. Food and Drug Administration. Therefore, screening potent cancer chemopreventive agents is crucial to reduce the constantly increasing incidence and mortality rate of cancer. Considering the lengthy prevention process, an ideal chemopreventive agent should be nontoxic, inexpensive, and oral. Natural compounds have become a natural treasure reservoir for cancer chemoprevention because of their superior ease of availability, cost-effectiveness, and safety. The benefits of natural compounds as chemopreventive agents in cancer prevention have been confirmed in various studies. In light of this, the present review is intended to fully delineate the entire scope of cancer chemoprevention, and primarily focuses on various aspects of cancer chemoprevention based on natural compounds, specifically focusing on the mechanism of action of natural compounds in cancer prevention, and discussing in detail how they exert cancer prevention effects by affecting classical signaling pathways, immune checkpoints, and gut microbiome. We also introduce novel cancer chemoprevention strategies and summarize the role of natural compounds in improving chemotherapy regimens. Furthermore, we describe strategies for discovering anticancer compounds with low abundance and high activity, revealing the broad prospects of natural compounds in drug discovery for cancer chemoprevention. Moreover, we associate cancer chemoprevention with precision medicine, and discuss the challenges encountered in cancer chemoprevention. Finally, we emphasize the transformative potential of natural compounds in advancing the field of cancer chemoprevention and their ability to introduce more effective and less toxic preventive options for oncology.

Preclinical studies demonstrate that exercise reduces tumor incidence and growth. Rapid release of extracellular vesicles (EVs) during exercise suggests their potential role as mediators of exercise-induced systemic effects and physiological adaptation. This study investigated the impact of exercise-induced plasma EVs on tumor growth and immune tumor microenvironment in murine models of triple-negative breast cancer (TNBC): EO771 (a C57BL/6-derived TNBC cell line) and 4T1 (a BALB/c-derived TNBC cell line).

Size exclusion chromatography was used to isolate exercise-induced EVs from plasma of healthy female mice (BALB/c and C56BL/6, n = 30 per strain) that underwent ten 30-min moderate-intensity treadmill running sessions over 2 weeks. Nanoparticle tracking analysis, Western blot, and electron microscopy confirmed the presence of EVs in the samples. Tumor-bearing mice (n = 72 per strain) were administered with exercise-induced EVs before or/and after tumor implantation. Local and systemic immune responses were assessed using flow cytometry, enzyme-linked immunosorbent assay (ELISA), and quantitative polymerase chain reaction (qPCR).

Administration of exercise-induced EVs, particularly before tumor implantation, significantly suppressed tumor growth and reduced tumor burden in both TNBC models. In EO771, endpoint tumor volumes were 278-330 mm³ in treated groups compared to 799 mm³ in untreated (p < 0.0001), while in 4T1, treated groups showed volumes of 287-564 mm³ vs. 696 mm³ in untreated (p = 0.0002). Notable differences in tumor-infiltrating lymphoid and myeloid cell subpopulations indicated immunomodulatory effects of exercise-induced EVs, particularly in the 4T1 model, where their continuous administration significantly increased intratumoral cluster of differentiation 8 (CD8) T lymphocyte proportion (5.77% vs. 0.90% in untreated, p < 0.0001). Similarly, in the EO771 model, exercise-induced EVs administered before tumor implantation led to a marked rise in intratumoral CD8 T lymphocytes (2.24% vs. 1.08% in untreated, p = 0.0181).

Our findings indicate that exercise-induced EV treatment elicits a pro-inflammatory antitumor immune response, suggesting a shift of immunologically cold TNBC tumors towards a more inflamed phenotype associated with better outcomes. Our study supports the further investigation of EVs as modulators of antitumor immunity and their potential utility in enhancing the efficacy of immunotherapy.

The vast majority of cancer-related deaths are attributed to metastasis. The lung, being a common site for cancer metastasis, is highly prone to being a target for multiple cancer types and causes a heavy disease burden. Accumulating evidence has demonstrated that tumor metastasis necessitates continuous interactions between tumor cells and distant metastatic niches. Nevertheless, a comprehensive elucidation of the underlying mechanisms governing lung-specific metastasis still poses a formidable challenge. In this review, we depict the lung susceptibility and the molecular profiles of tumors with the potential for lung metastasis. Under the conceptual framework of "Reciprocal Tumor-Lung Metastatic Symbiosis" (RTLMS), we mechanistically delineate the bidirectional regulatory dynamics and coevolutionary adaptation between tumor cells and distal pulmonary niches during lung-specific metastasis, including the induction of pre-metastatic-niches, positive responses of the lung, tumor colonization, dormancy, and reawakening. An enhanced understanding of the latest mechanisms is essential for developing targeted strategies to counteract lung-specific metastasis.

The epidermal growth factor receptor (EGFR) signaling pathway is essential for cellular processes such as proliferation, survival, and migration. Dysregulation of EGFR signaling is frequently observed in non-small cell lung cancer (NSCLC) and is associated with poor prognosis. This study aims to isolate and characterize extracellular vesicles (EVs) released by mutant EGFR lung cancer cell line PC9 and compare them with wild-type EGFR lung cancer cell line A549, while also evaluating the effect of gefitinib treatment on EV secretion and cargo composition. The two lung cancer cell lines were cultured with 2% EV-free serum, and EVs were subsequently isolated by differential ultra centrifugation. EVs were characterized by nanoparticle tracking analysis (NTA) and transmission electron microscopy (TEM) for quantification size and shape determination. Western blot analysis confirmed the enrichment and purity of isolated EVs. Results showed that EGFR mutation significantly increased EV release and altered their size, compared to EVs released by wild-type EGFR cells. In addition to classical EV markers such as CD81, Flotillin- 1, and TSG101, Western blot analysis also detected phosphorylated EGFR (p-EGFR) selectively packaged into EVs from PC9 cells. Gefitinib treatment significantly reduced EV secretion in PC9 cells and led to a marked decrease in p-EGFR incorporation into EVs, indicating that EV biogenesis and compostion are modulated by active EGFR signaling. In conclusion, this study highlights the significant influence of EGFR activation on EV secretion and cargo composition while demonstrating that EGFR inhibition via gefitinib alters EV-mediated signaling in lung cancer cells. These findings provide insights into tumor behavior, EV-mediated oncogenic communication, and the potential use of EVs as biomarkers and therapeutic targets in NSCLC.