MSC-derived Extracellular Vesicles Research Articles

An Update on Emerging Regenerative Medicine Applications: The Use of Extracellular Vesicles and Exosomes for the Management of Chronic Pain.

Chronic pain affects nearly two billion people worldwide, surpassing heart disease, diabetes, and cancer in terms of economic costs. Lower back pain alone is the leading cause of years lived with disability worldwide. Despite limited treatment options, regenerative medicine, particularly extracellular vesicles (EVs) and exosomes, holds early promise for patients who have exhausted other treatment options. EVs, including exosomes, are nano-sized structures released by cells, facilitating cellular communication through bioactive molecule transfer, and offering potential regenerative properties to damaged tissues. Here, we review the potential of EVs and exosomes for the management of chronic pain. In osteoarthritis, various exosomes, such as those derived from synovial mesenchymal stem cells, human placental cells, dental pulp stem cells, and bone marrow-derived mesenchymal stem cells (MSCs), demonstrate the ability to reduce inflammation, promote tissue repair, and alleviate pain in animal models. In intervertebral disc disease, Wharton's jelly MSC-derived EVs enhance cell viability and reduce inflammation. In addition, various forms of exosomes have been shown to reduce signs of inflammation in neurons and alleviate pain in neuropathic conditions in animal models. Although clinical applications of EVs and exosomes are still in the early clinical stages, they offer immense potential in the future management of chronic pain conditions. Clinical trials are ongoing to explore their therapeutic potential further, and with more research the potential applicability of EVs and exosomes will be fully understood.

Roles of extracellular vesicles from mesenchymal stem cells in regeneration.

Mesenchymal stem cells (MSCs) are highly valued in regenerative medicine due to their ability to self-renew and differentiate into various cell types. Their therapeutic benefits are primarily due to their paracrine effects, in particular through extracellular vesicles (EVs), which are related to intercellular communication. Recent advances in EV production and extraction technologies highlight the potential of MSC-derived EVs (MSC-EVs) in tissue engineering and regenerative medicine. MSC-EVs offer several advantages over traditional cell therapies, including reduced toxicity and immunogenicity compared to whole MSCs. EVs carrying functional molecules such as growth factors, cytokines and miRNAs play beneficial roles in tissue repair, fibrosis treatment and scar prevention by promoting angiogenesis, skin cell migration, proliferation, extracellular matrix remodeling and reducing inflammation. Despite the potential of MSC-EVs, there are several limitations to their use, including variability in quality, the need for standardized methods, low yield, and concerns about the composition of EVs and the potential risks. Overall, MSC-EVs are a promising alternative to cell-based therapies, and ongoing studies aim to understand their actions and optimize their use for better clinical outcomes in wound healing and skin regeneration.

Therapeutic potential of mesenchymal stem cell-derived extracellular vesicles: A focus on inflammatory bowel disease.

Mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) have emerged as key regulators of intercellular communication, orchestrating essential biological processes by delivering bioactive cargoes to target cells. Available evidence suggests that MSC-EVs can mimic the functions of their parental cells, exhibiting immunomodulatory, pro-regenerative, anti-apoptotic, and antifibrotic properties. Consequently, MSC-EVs represent a cell-free therapeutic option for patients with inflammatory bowel disease (IBD), overcoming the limitations associated with cell replacement therapy, including their non-immunogenic nature, lower risk of tumourigenicity, cargo specificity and ease of manipulation and storage. This review aims to provide a comprehensive examination of the therapeutic efficacy of MSC-EVs in IBD, with a focus on their mechanisms of action and potential impact on treatment outcomes. We examine the advantages of MSC-EVs over traditional therapies, discuss methods for their isolation and characterisation, and present mechanistic insights into their therapeutic effects through transcriptomic, proteomic and lipidomic analyses of MSC-EV cargoes. We also discuss available preclinical studies demonstrating that MSC-EVs reduce inflammation, promote tissue repair and restore intestinal homeostasis in IBD models, and compare these findings with those of clinical trials. Finally, we highlight the potential of MSC-EVs as a novel therapy for IBD and identify challenges and opportunities associated with their translation into clinical practice. The source of mesenchymal stem cells (MSCs) strongly influences the composition and function of MSC-derived extracellular vesicles (EVs), affecting their therapeutic potential. Adipose-derived MSC-EVs, known for their immunoregulatory properties and ease of isolation, show promise as a treatment for inflammatory bowel disease (IBD). MicroRNAs are consistently present in MSC-EVs across cell types and are involved in pathways that are dysregulated in IBD, making them potential therapeutic agents. For example, miR-let-7a is associated with inhibition of apoptosis, miR-100 supports cell survival, miR-125b helps suppress pro-inflammatory cytokines and miR-20 promotes anti-inflammatory M2 macrophage polarisation. Preclinical studies in IBD models have shown that MSC-EVs reduce intestinal inflammation by suppressing pro-inflammatory mediators (e.g., TNF-α, IL-1β, IL-6) and increasing anti-inflammatory factors (e.g., IL-4, IL-10). They also promote mucosal healing and strengthen the integrity of the gut barrier, suggesting their potential to address IBD pathology.

Multiplex analysis of cancer cells treated with induced mesenchymal stem cell membrane vesicles

BACKGROUND: Extracellular vesicles (EVs) are membrane-derived vesicles released by cells into the extracellular space, playing a crucial role in intercellular communication and regulating a range of biological processes. These vesicles are found in tumor tissue, where they serve as mediators in signal transduction between tumor cells and the cells of the microenvironment. Similar to their parent mesenchymal stem cells (MSCs), extracellular vesicles demonstrate contradictory effects on tumor development. Studies have shown that MSC-derived EVs promote tumor growth; however, some research has also demonstrated their inhibitory role. AIM: The aim is to assess the effect of mesenchymal stem cell-derived membrane vesicles on the molecular composition of cancer cells. METHODS: Induced membrane vesicles were obtained from mesenchymal stem cells through treatment with cytochalasin B. Mesenchymal stem cells were initially sourced from adipose tissue. To simulate intercellular communication between tumor cells and MSCs, induced membrane vesicles were applied at varying protein concentrations to recipient cells (SH-SY5Y, PC3, MCF7). The bicinchoninic acid method was used to measure the total protein concentration isolated from human cells/induced membrane vesicles. Subsequently, the molecular composition of the recipient cells after the application of induced membrane vesicles was analyzed using multiplex assays. RESULTS: We determined that after the application of MSC-derived membrane vesicles to cancer cells, significant alterations occur in the expression of numerous biologically active molecules, including cytokines, chemokines, and growth factors. Specifically, increased concentrations of growth factor FGF-2, cytokines G-CSF, Fractalkine, IL-12p40, IL-9, IL-4, IL-6, IL-8, and chemokines IP-10, MCP-1, among others, were observed. The analysis also revealed that most of these molecules are associated with cell proliferation, migration, and immune response. CONCLUSION: Mesenchymal stem cell-derived membrane vesicles are capable of altering the molecular profile of cancer cells, increasing the concentration of molecules linked to cell survival and migration.

Human adipose and umbilical cord mesenchymal stem cell-derived extracellular vesicles mitigate photoaging via TIMP1/Notch1

UVB radiation induces oxidative stress, DNA damage, and inflammation, leading to skin wrinkling, compromised barrier function, and an increased risk of carcinogenesis. Addressing or preventing photoaging may offer a promising therapeutic avenue for these conditions. Recent research indicated that mesenchymal stem cells (MSCs) exhibit significant therapeutic potential for various skin diseases. Given that extracellular vesicles (EV) can deliver diverse cargo to recipient cells and elicit similar therapeutic effects, we investigated the roles and underlying mechanisms of both adipose-derived MSC-derived EV (AMSC-EV) and umbilical cord-derived MSC-derived EV (HUMSC-EV) in photoaging. Our findings indicated that in vivo, treatment with AMSC-EV and HUMSC-EV resulted in improvements in wrinkles and skin hydration while also mitigating skin inflammation and thickness alterations in both the epidermis and dermis. Additionally, in vitro studies using human keratinocytes (HaCaTs), human dermal fibroblast cells (HDFs), and T-Skin models revealed that AMSC-EV and HUMSC-EV attenuated senescence, reduced levels of reactive oxygen species (ROS) and DNA damage, and alleviated inflammation induced by UVB. Furthermore, EV treatment enhanced cell viability and migration capacity in the epidermis and promoted extracellular matrix (ECM) remodeling in the dermis in photoaged cell models. Mechanistically, proteomics results showed that TIMP1 was highly expressed in both AMSC-EV and HUMSC-EV and could exert similar effects as MSC-EV. In addition, we found that EV and TIMP1 could inhibit Notch1 and downstream targets Hes1, P16, P21, and P53. Collectively, our data suggests that both AMSC-EV and HUMSC-EV attenuate skin photoaging through TIMP1/Notch1.

A Systematic Review of Mesenchymal Stem Cell-Derived Extracellular Vesicles: A Potential Treatment for Glioblastoma

Background: Glioblastoma (GBM) is an extremely aggressive brain tumor that has few available treatment options and a dismal prognosis. Recent research has highlighted the potential of extracellular vesicles (MSC-EVs) produced from mesenchymal stem cells as a potential treatment approach for GBM. MSC-EVs, including exosomes, microvesicles, and apoptotic bodies, perform a significant function in cellular communication and have shown promise in mediating anti-tumor effects. Purpose: This systematic literature review aims to consolidate current findings on the therapeutic potential of MSC-EVs in GBM treatment. Methods: A systematic search was conducted across major medical databases (PubMed, Web of Science, and Scopus) up to September 2024 to identify studies investigating the use of MSC-derived EVs in GBM therapy. Keywords included “extracellular vesicles”, “mesenchymal stem cells”, “targeted therapies”, “outcomes”, “adverse events”, “glioblastoma”, and “exosomes”. Inclusion criteria were studies published in English involving GBM models both in vivo and in vitro and those reporting on therapeutic outcomes of MSC-EVs. Data were extracted and analyzed based on EV characteristics, mechanisms of action, and therapeutic efficacy. Results: The review identified several key studies demonstrating the anti-tumor effects of MSC-EVs in GBM models. A total of three studies were included, focusing on studies conducted between 2021 and 2023. The review included three studies that collectively enrolled a total of 18 patients. These studies were distributed across two years, with two trials published in 2023 (66.7%) and one in 2021 (33.3%). The mean age of the participants ranged from 37 to 57 years. In terms of gender distribution, males were the predominant group in all studies. Prior to receiving MSC-EV therapy, all patients had undergone standard treatments for GBM, including surgery, chemotherapy (CT), and, in some cases, radiation therapy (RT). In all three studies, the targeted treatment involved the administration of herpes simplex virus thymidine kinase (HSVtk) gene therapy delivered to the tumor site, then 14 days of ganciclovir treatment. Outcomes across the studies indicated varying levels of efficacy for the MSC-EV-based therapy. The larger 2023 study reported fewer encouraging outcomes, with a median PFS of 11.0 months (95% CI: 8.3–13.7) and a median OS of 16.0 months (95% CI: 14.3–17.7). Adverse effects were reported in only one of the studies, the 2021 trial, where patients experienced mild-to-moderate side effects, including fever, headache, and cerebrospinal fluid leukocytosis. A total of 11 studies on preclinical trials, using in vitro and in vivo models, were included, covering publications from 2010 to 2024. The studies utilized MSCs as delivery systems for various therapeutic agents (interleukin 12, interleukin 7, doxorubicin, paclitaxel), reflecting the versatility of these cells in targeted cancer therapies. Conclusions: MSC-derived EVs represent a promising therapeutic approach for GBM, offering multiple mechanisms to inhibit tumor growth and enhance treatment efficacy. Their ability to deliver bioactive molecules and modulate the tumor microenvironment underscores their potential as a novel, cell-free therapeutic strategy. Future studies should optimize EV production and delivery methods and fully understand their long-term effects in clinical settings to harness their therapeutic potential in GBM treatment.

Molecular and Cellular Mechanisms of the Therapeutic Effect of Mesenchymal Stem Cells and Extracellular Vesicles in Corneal Regeneration

The cornea is a vital component of the visual system, and its integrity is crucial for optimal vision. Damage to the cornea resulting from trauma, infection, or disease can lead to blindness. Corneal regeneration using mesenchymal stem cells (MSCs) and MSC-derived extracellular vesicles (MSC-EVs) offers a promising alternative to corneal transplantation. MSCs are multipotent stromal cells that can differentiate into various cell types, including corneal cells. They can also secrete a variety of anti-inflammatory cytokines and several growth factors, promoting wound healing and tissue reconstruction. This review summarizes the current understanding of the molecular and cellular mechanisms by which MSCs and MSC-EVs contribute to corneal regeneration. It discusses the potential of MSCs and MSC-EV for treating various corneal diseases, including corneal epithelial defects, dry eye disease, and keratoconus. The review also highlights finalized human clinical trials investigating the safety and efficacy of MSC-based therapy in corneal regeneration. The therapeutic potential of MSCs and MSC-EVs for corneal regeneration is promising; however, further research is needed to optimize their clinical application.

Efficient Treatment of Psoriasis Using Conditioned Media from Mesenchymal Stem Cell Spheroids Cultured to Produce Transforming Growth Factor-β1-Enriched Small-Sized Extracellular Vesicles.

Psoriasis is a common chronic inflammatory disease in which keratinocytes proliferate abnormally due to excessive immune action. Psoriasis can be associated with various comorbidities and has a significant impact on health-related quality of life. Although many systemic treatments, including biologic agents, have been developed, topical treatment remains the main option for psoriasis management. Consequently, there is an urgent need to develop topical treatments with minimal side effects and high efficacy. Mesenchymal stem cells (MSCs) exhibit excellent immune regulation, anti-inflammatory activities, and therapeutic effects, and MSC-derived extracellular vesicles (EVs) can serve as crucial mediators of functional transfer from MSCs. Therefore, this study aimed to develop a safe and easy-to-use emulsion cream for treating psoriasis using MSC conditioned media (CM) containing EVs. We developed an enhanced Wharton's jelly MSC (WJ-MSC) culture method through a three-dimensional (3D) culture containing exogenous transforming growth factor-β3. Using the 3D culture system, we obtained CM from WJ-MSCs, which yielded a higher EV production compared to that of conventional WJ-MSC culture methods, and investigated the effect of EV-enriched 3D-WJ-MSC-CM cream on psoriasis-related inflammation. Administration of the EV-enriched 3D-WJ-MSC-CM cream significantly reduced erythema, thickness, and scaling of skin lesions, alleviated imiquimod-induced psoriasiform lesions in mice, and ameliorated histopathological changes in mouse skin. The upregulated mRNA expression of inflammatory cytokines, including IL-17a, IL-22, IL-23, and IL-36, decreased in the lesions. In conclusion, we present here a new topical treatment for psoriasis using an MSC EV-enriched cream.

Large and small extracellular vesicles from Wharton’s jelly MSCs: Biophysics, function, and strategies to improve immunomodulation

Extracellular vesicles (EVs) have emerged as mediators of immunosuppression and pro-regenerative processes, particularly through mesenchymal stromal cells (MSCs) across various disease models. Despite significant progress, there is still a need for a deeper understanding of EV content and functionality to fully harness their biomedical potential. Moreover, strategies to enhance EV production for clinical scalability are still under development. This study aimed to characterize two distinct types of EV-large EV (lgEV) and small EV (smEV)-secreted by Wharton's jelly MSCs (WJ-MSCs). Strategies were explored to augment both EV production and their immunoregulatory effects. Both lgEV and smEV displayed typical EV markers and demonstrated inhibition of human lymphocyte proliferation. Furthermore, analysis of IsomiR content revealed a pronounced immunomodulating signature within MSC-derived EVs, validated by a dual-fluorescence reporter system. MSC primed with pro-inflammatory cytokines yielded increased production of lgEV and smEV, enhancing their immunomodulatory potency. Finally, genetically engineering WJ-MSC to express CD9 resulted in lgEV and smEV with heightened efficacy in suppressing lymphocyte proliferation. This study successfully isolated, characterized, and demonstrated the potent immunosuppressive effect of WJ-MSC-derived lgEV and smEV. We propose cytokine preconditioning and genetic manipulation as viable strategies to enhance the therapeutic potential of WJ-MSC-derived EV in inflammatory conditions.

Therapeutic Potential of MSC-Derived Extracellular Vesicles Carrying miRNAs for Modulating Autophagy and Cellular Degeneration in Intervertebral Disc Degeneration.

This research utilized bioinformatics and in vitro modeling to assess the effects of mesenchymal stem cell-derived extracellular vesicles (MSC-EVs) containing specific microRNAs (miRNAs) on the autophagy and degeneration of nucleus pulposus (NP) cells in intervertebral disc degeneration (IDD). To determine the therapeutic potential of MSC-EVs loaded with miRNAs in modulating pathological changes in IDD. IDD is characterized by changes in gene expression that contribute to cell degeneration and reduced disc integrity. MSC-EVs are known for their role in cellular communication and potentially reversing these degenerative processes. Key differentially expressed miRNAs and mRNAs in MSC-EVs were identified using bioinformatics analysis. Three miRNAs (miR-486-5p, miR-3648, miR-1827) typically down-regulated in IDD were selected for further study. The bone marrow-derived MSC-EVs used in this study were cultured in a 2D environment. These MSC-EVs were isolated with the purpose of delivering the selected miRNAs to IDD NP cells in vitro, targeting specifically the upregulated genes (SMAD2, ESR1, MAVS, MMP14) associated with autophagy and degeneration. MSC-EV treatment led to significant downregulation of target genes, enhanced cellular proliferation, and decreased apoptosis and autophagy. Overexpression of these target genes produced the opposite effects, confirming the miRNAs' regulatory roles. MSC-EVs carrying specific miRNAs can effectively modulate gene expression, reduce degenerative processes, and promote cellular proliferation in IDD, indicating a promising therapeutic strategy for treating IDD and potentially other degenerative diseases. Further investigations are warranted to explore MSC-EV applications in regenerative medicine.

Effect of Mesenchymal Stem Cell–Derived Extracellular Vesicles Induced by Advanced Glycation End Products on Energy Metabolism in Vascular Endothelial Cells

IntroductionAdvanced glycation end products (AGEs) play a critical role in the development of vascular diseases in diabetes. While stem cell therapies often involve exposure to AGEs, the impact of this environment on extracellular vesicles (EVs) and endothelial cell metabolism remains unclear. MethodsHuman umbilical cord mesenchymal stem cells (MSC) were treated with either 0 ng/mL or 100 ng/mL AGEs in a serum-free medium for 48 hours, after which MSC-EVs were isolated. The EVs were characterized for morphology, particle size, and protein markers of MSC-EVs, and performed miRNA sequencing to identify differentially expressed miRNAs. MSC-EVs were co-cultured with HUVEC cells to assess effects on cell viability, metabolic activity, oxidative stress, and antioxidant capacity. Tube formation and glucose transporter protein analyses were conducted to evaluate the angiogenic ability and glucose metabolism capacity. ResultsMSC-EVs ranged from 30 to 150 nm, consistenting with exosomal properties. AGEs treatment reduced MSC viability but had minimal effect on EV morphology and protein markers. miRNAs sequencing showed downregulation of hsa-miR-223-3p, hsa-miR-126-3p_R-1, with upregulation of hsa-miR-574-5p, implicating changes in glycolytic and oxidative phosphorylation pathways. MSC-EVs treated with AGEs decreased HUVEC viability (P<0.05), pH (P<0.05), ATP metabolism (P<0.05), glucose metabolism (P<0.05), while enhancing glycolysis processes, including glycolytic activity, capacity, and reserve (P<0.05). This likely resulted from impaired mitochondrial function, including reduced ATP production, maximal respiration, basal respiration, and spare respiratory capacity (P<0.05), or increased ROS (P<0.05) and G6PD activity (P<0.05). Additionally, AGEs reduced GLUT1, GLUT3, GLUT4, and SCO2 expression (P<0.05), along with angiogenic capacity (P<0.05) in HUVEC cells. ConclusionExposure to AGEs diminishes the therapeutic potential of MSC-derived EVs by disrupting energy metabolism and promoting metabolic reprogramming in endothelial cells. These findings suggest that adjusting the dosage or frequency of MSC-EVs may enhance their efficacy for treating diabetes-related vascular conditions. Further research is warranted to evaluate AGEs' broader impact on various cell types and metabolic pathways for improved exosome-based therapies.

Immunomodulatory Effect of Adipose Stem Cell-Derived Extra-Cellular Vesicles on Cytokine Expression and Regulatory T Cells in Patients with Asthma.

Although mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) are as effective as MSCs in the suppression of allergic airway inflammation, few studies have evaluated the immunomodulatory capacity of MSC-derived EVs in patients with asthma. Thus, we assessed the effects of adipose stem cell (ASC)-derived EVs on cytokine expression and regulatory T cells (Tregs) in peripheral blood mononuclear cells (PBMCs) of asthmatic patients. PBMCs (1 × 106 cells/mL) were isolated from asthmatic patient and healthy controls and co-cultured with 1 μg/mL of ASC-derived EVs. Th (T helper) 1-, Th2-, and Treg-related cytokine expression, fluorescence-activated cell sorting analysis of CD4+CD25+FOXP3+ T cells, and co-stimulatory molecules were analyzed before and after ASC-derived EV treatment. The expression levels of IL-4 and costimulatory molecules such as CD83 and CD86 were significantly higher in PBMCs of asthmatic patients than in control PBMCs. However, ASC-derived EV treatment significantly decreased the levels of interleukin (IL)-4 and co-stimulatory molecules such as CD83 and CD86 in the phytohemagglutinin (PHA)-stimulated PBMC of asthmatic patients. Furthermore, ASC-derived EVs remarkably increased the transforming growth factor-β (TGF-β) levels and expression of Tregs in the PBMC of asthmatic patients. ASC-derived EVs induce Treg expansion and have immunomodulatory effects by downregulating IL-4 and upregulating TGF-β in PBMCs of asthmatic patients.

Urine-derived stem cells serve as a robust platform for generating native or engineered extracellular vesicles

BackgroundMesenchymal stromal cell (MSC) therapy holds great potential yet efficacy and safety concerns with cell therapy persist. The beneficial effects of MSCs are often attributed to their secretome that includes extracellular vesicles (EVs). EVs carry biologically active molecules, protected by a lipid bilayer. However, several barriers hinder large-scale MSC EV production. A serum-free culturing approach is preferred for producing clinical-grade MSC-derived EVs but this can affect both yield and purity. Consequently, new strategies have been explored, including genetically engineering MSCs to alter EV compositions to enhance potency, increase circulation time or mediate targeting. However, efficient transfection of MSCs is challenging. Typical sources of MSC include adipose tissue and bone marrow, which both require invasive extraction procedures. Here, we investigate the use of urine-derived stem cells (USCs) as a non-invasive and inexhaustible source of MSCs for EV production.MethodsWe isolated, expanded, and characterized urine-derived stem cells (USCs) harvested from eight healthy donors at three different time points during the day. We evaluated the number of clones per urination, proliferation capacity and conducted flow cytometry to establish expression of surface markers. EVs were produced in chemically defined media and characterized. PEI/DNA transfection was used to genetically engineer USCs using transposon technology.ResultsThere were no differences between time points for clone number, doubling time or viability. USCs showed immunophenotypic characteristics of MSCs, such as expression of CD73, CD90 and CD105, with no difference at the assessed time points, however, male donors had reduced CD73 + cells. Expanded USCs were incubated without growth factors or serum for 72 h without a loss in viability and EVs were isolated. USCs were transfected with high efficiency and after 10 days of selection, pure engineered cell cultures were established.ConclusionsIsolation and expansion of MSCs from urine is non-invasive, robust, and without apparent sex-related differences. The sampling time point did not affect any measured markers or USC isolation potential. USCs offer an attractive production platform for EVs, both native and engineered.

Intratympanic injection of MSC-derived small extracellular vesicles protects spiral ganglion neurons from degeneration

Sensorineural hearing loss is one of the most prevalent sensory deficits. Spiral ganglion neurons (SGNs) exhibit very limited regeneration capacity and their degeneration leads to profound hearing loss. Mesenchymal stem cell-derived small extracellular vesicles (MSC-sEV) have been demonstrated to repair tissue damage in various degenerative diseases. However, the effects of MSC-sEV on SGN degeneration remain unclear. In this study, we investigated the efficacy of MSC-sEV for protection against ouabain-induced SGN degeneration. MSC-sEV were derived from rat bone marrow and their components related to neuron growth were determined by proteomic analysis. In primary culture SGNs, MSC-sEV significantly promoted neurite growth and growth cone development. The RNA-Seq analysis of SGNs showed that enriched pathways include neuron development and axon regeneration, consistent with proteomics. In ouabain induced SGN degeneration rat model, MSC-sEV administration via intratympanic injection significantly enhanced SGN survival and mitigated hearing loss. Furthermore, after ouabain treatment, SGNs displayed evident signs of apoptosis, including nuclei condensation and fragmentation, with numerous cells exhibiting TUNEL-positive. However, administration of MSC-sEV effectively decreased the number of TUNEL-positive cells and reduced caspase-3 activation. In conclusion, our findings demonstrate the potential of MSC-sEV in preventing SGN degeneration and promoting neural growth, suggesting intratympanic injection of MSC-sEV is a specific and efficient strategy for neural hearing loss.

Emerging Role and Mechanism of Mesenchymal Stem Cells-Derived Extracellular Vesicles in Rheumatic Disease.

Mesenchymal stem cells (MSCs) are pluripotent stem cells derived from mesoderm. Through cell-to-cell contact or paracrine effects, they carry out biological tasks like immunomodulatory, anti-inflammatory, regeneration, and repair. Extracellular vesicles (EVs) are the primary mechanism for the paracrine regulation of MSCs. They deliver proteins, nucleic acids, lipids, and other active compounds to various tissues and organs, thus facilitating intercellular communication. Rheumatic diseases may be treated using MSCs and MSC-derived EVs (MSC-EVs) due to their immunomodulatory capabilities, according to mounting data. Since MSC-EVs have low immunogenicity, high stability, and similar biological effects as to MSCs themselves, they are advantageous over cell therapy for potential therapeutic applications in rheumatoid arthritis, systemic erythematosus lupus, systemic sclerosis, Sjogren's syndrome, and other rheumatoid diseases. This review integrates recent advances in the characteristics, functions, and potential molecular mechanisms of MSC-EVs in rheumatic diseases and provides a new understanding of the pathogenesis of rheumatic diseases and MSC-EV-based treatment strategies.

Mesenchymal Stromal Cell-Derived Extracellular Vesicles for Reversing Hepatic Fibrosis in 3D Liver Spheroids.

Hepatic fibrosis, arising from prolonged liver injury, entails the activation of hepatic stellate cells (HSCs) into myofibroblast-like cells expressing alpha-smooth muscle actin (α-SMA), thereby driving extracellular matrix deposition and fibrosis progression. Strategies targeting activated HSC reversal and hepatocyte regeneration show promise for fibrosis management. Previous studies suggest that extracellular vesicles (EVs) from mesenchymal stromal cells (MSCs) can suppress HSC activation, but ensuring EV purity is essential for clinical use. This study investigated the effects of MSC-derived EVs cultured in chemically defined conditions on liver spheroids and activated HSCs. Umbilical cord- and bone marrow-derived MSCs were expanded in chemically defined media, and EVs were isolated using filtration and differential ultracentrifugation. The impact of MSC-EVs was evaluated on liver spheroids generated in Sphericalplate 5D™ and on human HSCs, both activated by transforming growth factor beta 1 (TGF-β1). MSC-EVs effectively reduced the expression of profibrotic markers in liver spheroids and activated HSCs induced by TGF-β1 stimulation. These results highlight the potential of MSC-EVs collected under chemically defined conditions to mitigate the activated phenotype of HSCs and liver spheroids, suggesting MSC-EVs as a promising treatment for hepatic fibrosis.

MicroRNA-based engineered mesenchymal stem cell extracellular vesicles to treat visual deficits after blast-induced trauma

Our previous studies have shown the benefit of intravitreal injection of a mesenchymal stem cell (MSC)- derived secretome to treat visual deficits in a mild traumatic brain injury (mTBI) mouse model. In this study, we have addressed whether MSC-derived extracellular vesicles (EV) overexpressing miR424, which particularly targets neuroinflammation, show similar benefits in the mTBI model. Adult C57BL/6 mice were subjected to a 50-psi air pulse on the left side, overlying the forebrain, resulting in mTBI. Sham-blast mice were controls. Within an hour of blast injury, 3 μl (∼7.5 × 108 particles) of miR424-EVs, native-EVs, or saline was delivered intravitreally. One month later, retinal morphology was observed through optical coherence tomography (OCT); visual function was assessed using optokinetic nystagmus (OKN) and electroretinogram (ERG), followed by immunohistological analysis. A separate study in adult mice tested the dose-response of EVs for safety. Blast injury mice with saline showed decreased visual acuity compared with the sham group (0.30 ± 0.03 vs. 0.39 ± 0.01 c/d, p < 0.02), improved with miR424-EVs (0.39 ± 0.02 c/d, p < 0.01) but not native-EVs (0.33 ± 0.04 c/d, p > 0.05). Contrast sensitivity thresholds of blast mice receiving saline increased compared with the sham group (85.3 ± 5.9 vs. 19.9 ± 4.8, %, p < 0.001), rescued by miR424-EVs (23.6 ± 7.3 %, p < 0.001) and native-EVs (45.6 ± 10.7 %, p < 0.01). Blast injury decreased “b” wave amplitude compared to sham mice (94.6 ± 24.0 vs. 279.2 ± 25.3 μV, p < 0.001), improved with miR424-EVs (173.0 ± 27.2 μV, p < 0.03) and native-EVs (230.2 ± 37.2 μV, p < 0.01) with a similar decrease in a-wave amplitude in blast mice improved with both miR424-EVs and native-EVs. Immunohistology showed increased GFAP and IBA1 in blast mice with saline compared with sham (GFAP: 11.9 ± 1.49 vs. 9.1 ± 0.8, mean intensity/100,000 μm2 area, p < 0.03; IBA1: 36.08 ± 4.3 vs. 24.0 ± 1.54, mean intensity/100,000 μm2 area, p < 0.01), with no changes with native-EVs (GFAP: 12.6 ± 0.79, p > 0.05; IBA1: 32.8 ± 2.9, p > 0.05), and miR424-EV (GFAP: 13.14 ± 0.76, p > 0.05; IBA1: 31.4 ± 2.7, p > 0.05). Both native-EVs and miR424-EVs exhibited vitreous aggregation, as evidenced by particulates in the vitreous by OCT, and increased vascular structures, as evidenced by αSMA and CD31 immunostainings. The number of capillary lumens in the ganglion cell layer increased with increased particles in the eye, with native EVs showing the worst effects. In conclusion, our study highlights the promise of EV-based therapies for treating visual dysfunction caused by mTBI, with miR424-EVs showing particularly strong neuroprotective benefits. Both miR424-EVs and native-EVs provided similar protection, but issues with EV aggregation and astrogliosis or microglial/macrophage activation at the current dosage call for improved delivery methods and dosage adjustments. Future research should investigate the mechanisms behind EVs' effects and optimize miR424 delivery strategies to enhance therapeutic outcomes and reduce complications.

Unveiling the multifaceted roles of microRNAs in extracellular vesicles derived from mesenchymal stem cells: implications in tumor progression and therapeutic interventions.

Mesenchymal stem/stromal cells (MSCs) have the capacity to migrate to tumor sites in vivo and transmit paracrine signals by secreting extracellular vesicles (EVs) to regulate tumor biological behaviors. MSC-derived EVs (MSC-EVs) have similar tumor tropism and pro- or anti-tumorigenesis as their parental cells and exhibit superior properties in drug delivery. MSC-EVs can transfer microRNAs (miRNAs) to tumor cells, thereby manipulating multiple key cancer-related pathways, and further playing a vital role in the tumor growth, metastasis, drug resistance and other aspects. In addition, tumor cells can also influence the behaviors of MSCs in the tumor microenvironment (TME), orchestrating this regulatory process via miRNAs in EVs (EV-miRNAs). Clarifying the specific mechanism by which MSC-derived EV-miRNAs regulate tumor progression, as well as investigating the roles of EV-miRNAs in the TME will contribute to their applications in tumor pharmacotherapy. This article mainly reviews the multifaceted roles and mechanism of miRNAs in MSC-EVs affecting tumor progression, the crosstalk between MSCs and tumor cells caused by EV-miRNAs in the TME. Eventually, the clinical applications of miRNAs in MSC-EVs in tumor therapeutics are illustrated.

The role of mesenchymal stem cells in cancer and prospects for their use in cancer therapeutics.

Mesenchymal stem cells (MSCs) are recruited by malignant tumor cells to the tumor microenvironment (TME) and play a crucial role in the initiation and progression of malignant tumors. This role encompasses immune evasion, promotion of angiogenesis, stimulation of cancer cell proliferation, correlation with cancer stem cells, multilineage differentiation within the TME, and development of treatment resistance. Simultaneously, extensive research is exploring the homing effect of MSCs and MSC-derived extracellular vesicles (MSCs-EVs) in tumors, aiming to design them as carriers for antitumor substances. These substances are targeted to deliver antitumor drugs to enhance drug efficacy while reducing drug toxicity. This paper provides a review of the supportive role of MSCs in tumor progression and the associated molecular mechanisms. Additionally, we summarize the latest therapeutic strategies involving engineered MSCs and MSCs-EVs in cancer treatment, including their utilization as carriers for gene therapeutic agents, chemotherapeutics, and oncolytic viruses. We also discuss the distribution and clearance of MSCs and MSCs-EVs upon entry into the body to elucidate the potential of targeted therapies based on MSCs and MSCs-EVs in cancer treatment, along with the challenges they face.

P-793 New approaches for treating diminished ovarian reserve: MSC-derived extracellular vesicles and autologous platelet rich plasma

Abstract Study question Does intraovarian injection of MSC-Derived Extracellular Vesicles (EVs) and autologous platelet rich plasma (PRP) improves the outcome of IVF in women with Diminished Ovarian reserve? Summary answer EVs and PRP therapies increse Antral follicle count (AFC), lower FSH level and result in higher amount of mature oocytes and blastocysts What is known already Diminished ovarian reserve (DOR) and premature ovarian insufficiency (POI) are significant impediments to the success of in vitro fertilization (IVF). Current strategies for ovarian stimulation primarily affects only the growth of antral follicles. Even in scenarios where ovarian ovulatory function is compromised, a reservoir of dormant follicles persists, which could potentially be activated by stem cells. Mesenchymal stem cells (MSCs), owing to their multipotent differentiation capabilities and paracrine signaling properties, have emerged as leading candidates for innovative therapeutic approaches. Recent scientific inquiries have highlighted the paracrine impact of EVs secreted by stem cells, underscoring their prospective therapeutic relevance. Study design, size, duration This prospective interventional study was conducted at the V.I. Kulakov Scientific Research Center for Obstetrics, Gynecology, and Perinatology in Russia. It involved 60 infertile female participants aged 20 to 38 years, each with DOR and a history of at least two unsuccessful IVF cycles. These participants were enrolled in the study during their subsequent IVF cycle. Prior to inclusion, all patients provided written informed consent. Participants/materials, setting, methods Patients in group 1 (n = 30) received treatment with exosomes derived from human umbilical cord mesenchymal stem cells (HUC-MSCs-Exos) prior to initiating their in vitro fertilization (IVF) cycle. In contrast, participants in group 2 (n = 30) underwent intraovarian therapy with autologous platelet-rich plasma (PRP) in preparation for their IVF cycle. The comparative analysis encompassed antral follicle count (AFC), anti-Müllerian hormone (AMH) levels, follicle-stimulating hormone (FSH) levels, ovarian stimulation outcomes, and the primary characteristics of oogenesis and embryogenesis. Main results and the role of chance After transplantation of EVs and PRP the ovarian function-related hormone levels and the AFC returned to nearly normal parameteres.Meanwhile, after exosomal treatment the improvement in reproductive outcomes was more significant. FSH concentration was significantly different before and 30 days after the procedure in both groups. In group 1 FSH values were less than the initial data In 95% of cases, in group 2 – in 72% of cases. AMH levels did not change significantly in both groups. The AFC increased in group 1 compared to group 2. All Patients underwent an IVF antagonist cycle after 3 months from the treatment. The number of obtained oocytes in group 1 ranged from 2 to 8, MII from 1 to 6; in group 2: from 0-5, MII – 0-5. The number of blastocysts in group 1 ranged from 1 to 5, in group 2 – from 0 to 3. In group 1, pregnancy occurred in 9 patients (30%): 1 women ( 3,3%) conceived spontaneously in the cycle of procedures, 8 (26,7%) attempted IVF, in group 2 pregnancy occurred in 4 patients (13,3%): 4 attempted IVF. EVs and PRP injections improve outcomes of IVF results in women with low ovarian reserve. Limitations, reasons for caution Limitations may include the fact that the study is single-center and has a rather small amount of patients enrolled in the study. Fundamental approaches to further research in this area are also required to better understand the obtained results. Wider implications of the findings EVs and PRP therapies appear to enhance ovarian reserve metrics and the success rates of IVF programs in patients with DOR. The clinical implementation of these interventions may offer improved reproductive prospects for patients whose sole option was previously oocyte donation. Trial registration number 121040600410-7