MSC-derived Extracellular Vesicles Research Articles

Mesenchymal-Stem-Cell-Derived Extracellular Vesicles Mitigate Trained Immunity in the Brain.

Trained immunity was recently discovered in innate immune cells and shown to facilitate the clearance of pathogens at the time of occurrence of the second insult. However, it exacerbates several aspects of neuropathologies, and proper therapy is needed to rectify this abnormal immune reaction. Mesenchymal-stem cells (MSCs) exhibit a distinct capability for brain repair but are associated with safety concerns. Extracellular vesicles derived from MSCs are a promising alternative therapy. In this study, we used lipopolysaccharides to activate trained immunity in the brain and examined the therapeutic potential of MSC-derived extracellular vesicles in mitigating the trained-immunity-induced exacerbated neuropathology. We found that MSC-derived extracellular vesicles showed comparable effects to those of MSCs in the mitigation of trained immunity in the brain. Moreover, the administration of MCS-derived extracellular vesicles mitigated the aggregated inflammatory responses in the acute stage of stroke and alleviated the trained-immunity-induced increased load of amyloid-β in APP/PS1 mice. We further investigated the molecular machinery of MSC-derived extracellular vesicles and found that IL-10 is important for the mediation of the therapeutic potential of MSC-derived extracellular vesicles toward the alleviation of trained immunity. Our study indicates that extracellular-vesicle-based regenerative strategies might be useful to mitigate trained immunity in the brain.

Metabolic Syndrome Alters the Cargo of Mitochondria-Related microRNAs in Swine Mesenchymal Stem Cell-Derived Extracellular Vesicles, Impairing Their Capacity to Repair the Stenotic Kidney

Background Coexisting metabolic syndrome (MetS) and renal artery stenosis (RAS) are linked to poor renal outcomes. Mesenchymal stem/stromal cell- (MSC-) derived extracellular vesicles (EVs) from lean animals show superior ability to repair the experimental MetS+RAS kidney compared to EVs from MetS pig MSCs. We hypothesized that MetS leads to selective packaging in porcine EVs of microRNAs capable of targeting mitochondrial genes, interfering with their capacity to repair the MetS+RAS kidney. Methods Five groups of pigs (n = 7 each) were studied after 16 weeks of diet-induced MetS and RAS (MetS+RAS) and MetS+RAS 4 weeks after a single intrarenal delivery of EVs harvested from allogeneic adipose tissue-derived MSCs isolated from Lean or MetS pigs, and Lean or MetS sham controls. Single-kidney blood flow (RBF) and glomerular filtration rate (GFR) were assessed in vivo with multidetector CT, whereas EV microRNA cargo, renal tubular mitochondrial structure and bioenergetics, and renal injury pathways were assessed ex vivo. Results microRNA sequencing revealed 19 dysregulated microRNAs capable of targeting several mitochondrial genes in MetS-EVs versus Lean-EVs. Lean- and MetS-EVs were detected in the stenotic kidney 4 weeks after administration. However, only MetS-EVs failed to improve renal mitochondrial density, structure, and function or attenuate oxidative stress, tubular injury, and fibrosis. Furthermore, Lean-EVs but not MetS-EVs restored RBF and GFR in MetS+RAS. Conclusion MetS alters the cargo of mitochondria-related microRNAs in swine MSC-derived EVs, which might impair their capacity to repair the poststenotic kidney in MetS+RAS. These observations may contribute to develop approaches to improve the efficacy of MSC-EVs for patients with MetS.

Preclinical Studies of MSC-Derived Extracellular Vesicles to Treat or Prevent Graft Versus Host Disease: a Systematic Review of the Literature.

IntroductionTreating and preventing graft-versus-host disease (GVHD) after allogeneic hematopoietic cell transplant (HCT) remains a significant challenge. The use of mesenchymal stromal cell-derived extracellular vesicles (MSC-EVs) appears promising and a systematic review of preclinical studies is needed to accelerate the design of translational studies.MethodsWe identified 4 eligible studies from a systematic review performed on December 1, 2018. In brief, eligible studies included the treatment or prevention of GVHD in animal models and the use of MSC-EVs. Study design and outcome data were extracted and reporting was evaluated using the SYRCLE tool to identify potential bias.ResultsTwo studies assessed the efficacy of MSC-EVs in treatment of GVHD and 2 studies address prevention. Mice treated with MSC-EVs showed improved median survival, GVHD clinical scores and histology scores as compared to untreated mice with GVHD. Prophylactic treatment with MSC-EVs attenuated GVHD severity and improved median survival as compared to no treatment or saline.ConclusionOur systematic review provides important insight regarding the potential of MSC-EVs to treat or prevent GVHD. Although few studies were identified, improved survival and attenuated histologic findings of GVHD were observed in mice after MSC-EV administration for the treatment and prevention of GVHD. Dosing of EVs and route of administration remain inconsistent, however, and scalability of EV isolation for clinical studies remains a challenge. Standardized outcome reporting is needed to pool results for metanalysis. Graphical abstract

Extracellular Vesicle-Dependent Communication Between Mesenchymal Stromal Cells and Immune Effector Cells.

Mesenchymal stem/stromal cells (MSCs) are multipotent cells residing in the stromal tissues of the body and capable of promoting tissue repair and attenuating inflammatory processes through their immunomodulatory properties. Preclinical and clinical observations revealed that not only direct intercellular communication mediates MSC properties; in fact, a pivotal role is also played by the release of soluble and bioactive factors, such as cytokines, growth factor and extracellular vesicles (EVs). EVs are membrane-coated vesicles containing a large variety of bioactive molecules, including lipids, proteins, and nucleic acids, such as RNA. EVs release their contents into target cells, thus influencing cell fate through the control of intracellular processes. In addition, MSC-derived EVs can mediate modulatory effects toward different effector cells belonging to both innate and adaptive immunity. In this review, we will discuss the literature data concerning MSC-derived EVs, including the current standardized methods for their isolation and characterization, the mechanisms supporting their immunoregulatory properties, and their potential clinical application as alternative to MSC-based therapy for inflammatory reactions, such as graft-versus-host disease (GvHD).

Human Adipose Tissue-Derived Stromal Cells Suppress Human, but Not Murine Lymphocyte Proliferation, via Indoleamine 2,3-Dioxygenase Activity.

Over recent years, mesenchymal stromal cells (MSC) have gained immense attraction in immunotherapy, regenerative medicine and tissue engineering. MSC microenvironment modulation occurs through synergy of direct cell–cell contact, and secreted soluble factors and extracellular vesicles (EV). MSC-derived EV have been suggested as cell-free immunomodulatory alternative to MSC; however, previous findings have challenged this. Furthermore, recent data suggest that evaluating the mechanism of action of human MSC (hMSC) in animal models might promote adverse immune reactions or lack of functionality due to xeno-incompatibilities. In this study, we first assessed the immunomodulatory strength of different human MSC sources on in vitro stimulated T cells and compared this to interferon-gamma (IFNγ) primed MSC conditioned medium (CM) and EV. Second, we addressed the main molecular mechanisms, and third, we assessed the MSC in vitro immunosuppressive effect across interspecies barriers. We identified human adipose tissue-derived stromal cells (ASC) with strongest immunomodulatory strength, followed by bone marrow (BM) and cord blood-derived MSC (CB). Whilst CM from primed ASC managed to exert analogous effects as their cellular counterpart, EV derived thereof did not, reproducing previous findings. IFNγ-induced indoleamine 2,3-dioxygenase (IDO) activity was identified as key mechanism to suppress human lymphocyte proliferation, as in the presence of the IDO inhibitor epacadostat (Epac) a stimulation of proliferation was seen. In addition, we revealed MSC immunosuppressive effects to be species-specific, because human cells failed to suppress murine lymphocyte proliferation. In summary, ASC were the strongest immunomodulators with the IDO-kynurenine pathway being key within the human system. Importantly, the in vitro lack of interspecies immunomodulatory strength suggests that preclinical data need to be carefully interpreted especially when considering a possible translation to clinical field.

The Potential of Mesenchymal Stromal Cell as Therapy in Neonatal Diseases.

Mesenchymal stromal cells (MSCs) can be derived from various tissue sources, such as the bone marrow (BMSCs), adipose tissue (ADSCs), umbilical cord (UC-MSCs) and umbilical cord blood (UCB-MSCs). Clinical trials have been conducted to investigate the potential of MSCs in ameliorating neonatal diseases, including bronchopulmonary dysplasia (BPD), intraventricular hemorrhage (IVH) and necrotizing enterocolitis (NEC). In preclinical studies, MSC therapy has been tested for the treatment of various neonatal diseases affecting the heart, eye, gut, and brain as well as sepsis. Up to date, the number of clinical trials using MSCs to treat neonatal diseases is still limited. The data reported thus far positioned MSC therapy as safe with positive outcomes. However, most of these trials are still preliminary and generally smaller in scale. Larger trials with more appropriate controls and a longer follow-up period need to be conducted to prove the safety and efficacy of the therapy more conclusively. This review discusses the current application of MSCs in treating neonatal diseases, its mechanism of action and future direction of this novel therapy, including the potential of using MSC-derived extracellular vesicles instead of the cells to treat various clinical conditions in the newborn.

Scalable Production of Human Mesenchymal Stromal Cell-Derived Extracellular Vesicles Under Serum-/Xeno-Free Conditions in a Microcarrier-Based Bioreactor Culture System.

Mesenchymal stromal cells (MSC) hold great promise for tissue engineering and cell-based therapies due to their multilineage differentiation potential and intrinsic immunomodulatory and trophic activities. Over the past years, increasing evidence has proposed extracellular vesicles (EVs) as mediators of many of the MSC-associated therapeutic features. EVs have emerged as mediators of intercellular communication, being associated with multiple physiological processes, but also in the pathogenesis of several diseases. EVs are derived from cell membranes, allowing high biocompatibility to target cells, while their small size makes them ideal candidates to cross biological barriers. Despite the promising potential of EVs for therapeutic applications, robust manufacturing processes that would increase the consistency and scalability of EV production are still lacking. In this work, EVs were produced by MSC isolated from different human tissue sources [bone marrow (BM), adipose tissue (AT), and umbilical cord matrix (UCM)]. A serum-/xeno-free microcarrier-based culture system was implemented in a Vertical-WheelTM bioreactor (VWBR), employing a human platelet lysate culture supplement (UltraGROTM-PURE), toward the scalable production of MSC-derived EVs (MSC-EVs). The morphology and structure of the manufactured EVs were assessed by atomic force microscopy, while EV protein markers were successfully identified in EVs by Western blot, and EV surface charge was maintained relatively constant (between −15.5 ± 1.6 mV and −19.4 ± 1.4 mV), as determined by zeta potential measurements. When compared to traditional culture systems under static conditions (T-flasks), the VWBR system allowed the production of EVs at higher concentration (i.e., EV concentration in the conditioned medium) (5.7-fold increase overall) and productivity (i.e., amount of EVs generated per cell) (3-fold increase overall). BM, AT and UCM MSC cultured in the VWBR system yielded an average of 2.8 ± 0.1 × 1011, 3.1 ± 1.3 × 1011, and 4.1 ± 1.7 × 1011 EV particles (n = 3), respectively, in a 60 mL final volume. This bioreactor system also allowed to obtain a more robust MSC-EV production, regarding their purity, compared to static culture. Overall, we demonstrate that this scalable culture system can robustly manufacture EVs from MSC derived from different tissue sources, toward the development of novel therapeutic products.

A narrative review of research advances in mesenchymal stem cell therapy for asthma.

Asthma is a chronic inflammatory disease of the airways that involves multiple cells, including inflammatory cells, structural cells, and cellular components. Glucocorticoids and beta-receptor agonists are still the first choices for asthma treatment. However, the asthma symptoms may still be poorly controlled in some patients after an optimal treatment. Mesenchymal stem cells (MSCs) are characterized by the potential for multi-directional differentiation and can exert immunomodulatory and anti-inflammatory effects. Its role in treating asthma has increasingly been recognized in recent years. In this review article, we sought to summarize the recent advances in the therapeutic effects of MSCs on several types of asthma and explain the relevant mechanisms. Articles on asthma treatment with MSCs as of January 2020 were searched in PubMed, Google Scholar, and Web of Science databases. It was found that MSCs have therapeutic effects on allergic asthma, non-allergic asthma and occupational asthma; gene-modified or pretreated MSCs improves the therapeutic effects of MSCs in asthma; MSC-derived conditioned medium or extracellular vesicles possess the considerable curative effect as MSC on asthma; and MSCs exert their therapeutic effects on asthma by restoring Th1/Th2 balance, reversing Th17/Tregs imbalance, inhibiting DC maturation, and promoting the switch of M1 to M2 and repairing epithelial injury. Thus, MSCs may be a promising treatment for asthma.

Immunomodulation of MSCs and MSC-Derived Extracellular Vesicles in Osteoarthritis.

Osteoarthritis (OA) has become recognized as a low-grade inflammatory state. Inflammatory infiltration of the synovium by macrophages, T cells, B cells, and other immune cells is often observed in OA patients and plays a key role in the pathogenesis of OA. Hence, orchestrating the local inflammatory microenvironment and tissue regeneration microenvironment is important for the treatment of OA. Mesenchymal stem cells (MSCs) offer the potential for cartilage regeneration owing to their effective immunomodulatory properties and anti-inflammatory abilities. The paracrine effect, mediated by MSC-derived extracellular vehicles (EVs), has recently been suggested as a mechanism for their therapeutic properties. In this review, we summarize the interactions between MSCs or MSC-derived EVs and OA-related immune cells and discuss their therapeutic effects in OA. Additionally, we discuss the potential of MSC-derived EVs as a novel cell-free therapy approach for the clinical treatment of OA.

Rejuvenation of Senescent Endothelial Progenitor Cells by Extracellular Vesicles Derived From Mesenchymal Stromal Cells

Mesenchymal stromal cell (MSC) transplantation is a form of the stem-cell therapy that has shown beneficial effects for many diseases. The use of stem-cell therapy, including MSC transplantation, however, has limitations such as the tumorigenic potential of stem cells and the lack of efficacy of aged autologous cells. An ideal therapeutic approach would keep the beneficial effects of MSC transplantation while circumventing the limitations associated with the use of intact stem cells. This study provides proof-of-concept evidence that MSC-derived extracellular vesicles represent a promising platform to develop an acellular therapeutic approach that would just do that. Extracellular vesicles are membranous vesicles secreted by MSCs and contain bioactive molecules to mediate communication between different cells. Extracellular vesicles can be taken up by recipient cells, and once inside the recipient cells, the bioactive molecules are released to exert the beneficial effects on the recipient cells. This study, for the first time to our knowledge, shows that extracellular vesicles secreted by MSCs recapitulate the beneficial effects of MSCs on vascular repair and promote blood vessel regeneration after ischemic events. Furthermore, MSCs from aged donors can be engineered to produce extracellular vesicles with improved regenerative potential, comparable to MSCs from young donors, thus eliminating the need for allogenic young donors for elderly patients.

Rejuvenation of mesenchymal stem cells by extracellular vesicles inhibits the elevation of reactive oxygen species

Aging induces numerous cellular disorders, such as the elevation of reactive oxygen species (ROS), in a number type of cells, including mesenchymal stem cells (MSCs). However, the correlation of ROS and impaired healing abilities as well as whether or not the inhibition of elevating ROS results in the rejuvenation of elderly MSCs is unclear. The rejuvenation of aged MSCs has thus recently received attention in the field of regenerative medicine. Specifically, extracellular vesicles (EVs) act as a novel tool for stem cell rejuvenation due to their gene transfer ability with systemic effects and safety. In the present study, we examined the roles of aging-associated ROS in the function and rejuvenation of elderly MSCs by infant EVs. The data clearly showed that elderly MSCs exhibited the downregulation of superoxide dismutase (SOD)1 and SOD3, which resulted in the elevation of ROS and downregulation of the MEK/ERK pathways, which are involved in the impairment of the MSCs’ ability to decrease necrotic area in the skin flap model. Furthermore, treatment with the antioxidant Edaravone or co-overexpression of SOD1 and SOD3 rescued elderly MSCs from the elevation of ROS and cellular senescence, thereby improving their functions. Of note, infant MSC-derived EVs rejuvenated elderly MSCs by inhibiting ROS production and the acceleration of cellular senescence and promoting the proliferation and in vivo functions in both type 1 and type 2 diabetic mice.

Mesenchymal Stromal Cells' Therapy for Polyglutamine Disorders: Where Do We Stand and Where Should We Go?

Polyglutamine (polyQ) diseases are a group of inherited neurodegenerative disorders caused by the expansion of the cytosine-adenine-guanine (CAG) repeat. This mutation encodes extended glutamine (Q) tract in the disease protein, resulting in the alteration of its conformation/physiological role and in the formation of toxic fragments/aggregates of the protein. This group of heterogeneous disorders shares common molecular mechanisms, which opens the possibility to develop a pan therapeutic approach. Vast efforts have been made to develop strategies to alleviate disease symptoms. Nonetheless, there is still no therapy that can cure or effectively delay disease progression of any of these disorders. Mesenchymal stromal cells (MSC) are promising tools for the treatment of polyQ disorders, promoting protection, tissue regeneration, and/or modulation of the immune system in animal models. Accordingly, data collected from clinical trials have so far demonstrated that transplantation of MSC is safe and delays the progression of some polyQ disorders for some time. However, to achieve sustained phenotypic amelioration in clinics, several treatments may be necessary. Therefore, efforts to develop new strategies to improve MSC’s therapeutic outcomes have been emerging. In this review article, we discuss the current treatments and strategies used to reduce polyQ symptoms and major pre-clinical and clinical achievements obtained with MSC transplantation as well as remaining flaws that need to be overcome. The requirement to cross the blood-brain-barrier (BBB), together with a short rate of cell engraftment in the lesioned area and low survival of MSC in a pathophysiological context upon transplantation may contribute to the transient therapeutic effects. We also review methods like pre-conditioning or genetic engineering of MSC that can be used to increase MSC survival in vivo, cellular-free approaches—i.e., MSC-conditioned medium (CM) or MSC-derived extracellular vesicles (EVs) as a way of possibly replacing the use of MSC and methods required to standardize the potential of MSC/MSC-derived products. These are fundamental questions that need to be addressed to obtain maximum MSC performance in polyQ diseases and therefore increase clinical benefits.

Mesenchymal Stromal Cell-Derived Extracellular Vesicles: Their Features and Impact on Fibrosis and Myogenesis in Vitro

Extracellular vesicles (EVs) and, particularly, exosomes are becoming a promising material for “cell-free therapy” of many pathologic conditions. In the recent years therapeutic effects of mesenchymal stromal cells (MSCs) have been attributed to their secretory factors, including EVs. In this study we aim to investigate how EVs produced by MSCs of different tissue origin can influence myogenesis and fibrosis–the main processes that accompany skeletal muscle regeneration. Bone marrow, adipose tissue, intact muscle, and injured muscle-derived rat MSCs were obtained and cultured according to standard protocols. EVs were obtained by ultracentrifugation from the MSC-conditioned medium of cell passages 2 and 3. The effects of EVs were estimated on the in vitro models of myogenesis and fibrosis. Samples of isolated EVs contained nano-sized vesicles that carried some exosomal markers. Promyogenic microRNA were found in EVs from bone marrow and muscle MSCs. We found that MSC-derived EVs from all sources significantly increased the number of newly formed myotubes in myoblast culture in vitro and also reduced the number and size of fibrotic nodules in muscle fibroblast culture in vitro. Our results suggest that MSC-derived EVs indeed possess antifibrotic and promyogenic potentials. However, the role of microRNA in these processes is yet to be determined, and the effect of EVs on skeletal muscle regeneration is yet to be tested in vivo.

Mesenchymal stem cell-derived small extracellular vesicles and bone regeneration.

Mesenchymal stem cells (MSCs) and MSC‐derived small extracellular vesicles (sEVs) are promising candidates for cell‐based and cell‐free regenerative medicine, respectively. By virtue of their multiple lineage differentiation capacity, MSCs have been implicated as an ideal tool for bone and cartilage regeneration. However, later observations attributed such regenerative effects to MSC‐secreted paracrine factors. Exosomes, endosomal originated sEVs carrying lipid, protein and nucleic acid cargoes, were identified as components of the MSC secretome and propagated the key regenerative and immunoregulatory characteristics of parental MSCs. Here, exosome biogenesis, the molecular composition of exosomes, sEV‐cell interactions and the effects on key bone homeostasis cells are reviewed. MSC‐derived sEVs show to promote neovascularization and bone and cartilage regeneration in preclinical disease models. The mechanisms include the transfer of molecules, including microRNAs, mRNAs and proteins, to other key cells. MSC‐derived sEVs are interesting candidates as biopharmaceuticals for drug delivery and for the engineering of biologically functionalized materials. Although major exploratory efforts have been made for therapeutic development, the secretion, distribution and biological effects of MSC‐derived sEVs in bone and cartilage regeneration are not fully understood. Moreover, techniques for high‐yield production, purity and storage need to be optimized before effective and safe MSC‐derived sEVs therapies are realized.

MicroRNA-130b-3p contained in MSC-derived EVs promotes lung cancer progression by regulating the FOXO3/NFE2L2/TXNRD1 axis

This study aimed to explore the molecular mechanism by which mesenchymal stem cells (MSCs) mediate lung cancer progression. Extracellular vesicles (EVs) were isolated from transfected or untransfected MSCs, and were co-cultured with lung cancer cells with/without microRNA-130b-3p (miR-130b-3p) inhibitor, mimic, overexpression plasmids of FOXO3/NFE2L2, or shRNAs. CCK-8 assay, colony formation, transwell assay, and flow cytometry were carried out to determine the biological functioning of lung cancer cells. Furthermore, FOXO3, Keap1, NFE2L2, and TXNRD1 expression was determined by qRT-PCR and western blot analysis. A tumor xenograft mouse model was used to determine role of EVs-miR-130b-3p and its target FOXO3 in lung cancer progression in vivo. miR-130b-3p was highly expressed in lung cancer tissues and MSC-derived EVs. Moreover, the MSC-derived EVs transferred miR-130b-3p to lung cancer cells to promote cell proliferation, migration, and invasion while repress cell apoptosis. miR-130b-3p directly targeted FOXO3, and FOXO3 elevated Keap1 expression to downregulate NFE2L2, thus inhibiting TXNRD1. FOXO3 overexpression or silencing of NFE2L2 or TXNRD1 diminished lung cancer cell proliferation, invasion, and migration but enhanced apoptosis. EV-delivered miR-130b-3p or FOXO3 silencing promoted lung cancer progression in vivo. In summary, MSC-derived EVs with upregulated miR-130b-3p suppressed FOXO3 to block the NFE2L2/TXNRD1 pathway, thus playing an oncogenic role in lung cancer progression.

Synovial Mesenchymal Stem Cell-Derived EV-Packaged miR-31 Downregulates Histone Demethylase KDM2A to Prevent Knee Osteoarthritis

Extracellular vesicles (EVs) derived from mesenchymal stem cells (MSCs) have emerged as important mediators of intercellular communication in response to cartilage damage. In this study, we sought to characterize the inhibitory role of microRNA (miR)-31 encapsulated in synovial MSC (SMSC)-derived EVs in knee osteoarthritis (OA). The expression of miR-31, lysine demethylase 2A (KDM2A), E2F transcription factor 1 (E2F1), and pituitary tumor transforming gene 1 (PTTG1) was validated in cartilage tissues of knee OA patients. Following SMSC-EV extraction and identification, chondrocytes with the miR-31 inhibitor were added with SMSC-EVs, whereupon the effects of miR-31 on proliferation and migration of chondrocytes were assessed. The interaction among miR-31, KDM2A, E2F1, and PTTG1 in chondrocyte activities was probed in vitro, along with an in vivo mouse knee OA model. We identified downregulated miR-31, E2F1, and PTTG1 and upregulated KDM2A in cartilage tissues of knee OA patients. SMSC-EV-packaged miR-31 potentiated chondrocyte proliferation and migration as well as cartilage formation by targeting KDM2A. Mechanistically, KDM2A bound to the transcription factor E2F1 and inhibited its transcriptional activity. Enrichment of E2F1 in the PTTG1 promoter region activated PTTG1 transcription, accelerating chondrocyte proliferation and migration. SMSC-EVs and EVs from miR-31-overexpressed SMSCs alleviated cartilage damage and inflammation in knee joints in vivo. SMSC-EV-encapsulated miR-31 ameliorates knee OA via the KDM2A/E2F1/PTTG1 axis.

Pulsed focused ultrasound enhances the therapeutic effect of mesenchymal stromal cell-derived extracellular vesicles in acute kidney injury

BackgroundAcute kidney injury (AKI) is characterized by rapid failure of renal function and has no curative therapies. Mesenchymal stromal cell (MSC)-derived extracellular vesicles (EVs) are known to carry therapeutic factors, which have shown promise in regenerative medicine applications, including AKI. However, there remains an unmet need to optimize their therapeutic effect. One potential avenue of optimization lies in pulsed focused ultrasound (pFUS), where tissues-of-interest are treated with sound waves. pFUS has been shown to enhance MSC therapy via increased cell homing, but its effects on cell-free EV therapy remain largely unexplored.MethodsWe combine pFUS pretreatment of the kidney with MSC-derived EV therapy in a mouse model of cisplatin-induced AKI.ResultsEVs significantly improved kidney function, reduced injury markers, mediated increased proliferation, and reduced inflammation and apoptosis. While pFUS did not enhance EV homing to the kidney, the combined treatment resulted in a superior therapeutic effect compared to either treatment alone. We identified several molecular mechanisms underlying this synergistic therapeutic effect, including upregulation of proliferative signaling (MAPK/ERK, PI3K/Akt) and regenerative pathways (eNOS, SIRT3), as well as suppression of inflammation.ConclusionTaken together, pFUS may be a strategy for enhancing the therapeutic efficacy of MSC-derived EV treatment for the treatment of AKI.

Strategies for scalable manufacturing and translation of MSC-derived extracellular vesicles.

Mesenchymal Stem/Stromal Cells (MSCs) are a well-studied cellular therapy with many clinical trials over the last few decades to treat a range of therapeutic indications. Recently, extracellular vesicles secreted by MSCs (MSC-EVs) have been shown to recapitulate many of the therapeutic effects of the MSCs themselves. While research in MSC-EVs has exploded, it is still early in their development towards a clinical therapy. One of the main challenges in cellular therapy, which will clearly also be a challenge in MSC-EV manufacturing, is developing a scalable, cGMP-compatible manufacturing paradigm. Therefore, the focus of this review is to identify some key MSC-EV manufacturing considerations such as the selection of critical raw materials, manufacturing platforms, and critical quality attribute assays. Addressing these issues early in research and development will accelerate clinical product development, clinical trials, and commercial therapies of MSC-EVs.

Extracellular vesicles derived from mesenchymal stromal cells mitigate intestinal toxicity in a mouse model of acute radiation syndrome

BackgroundHuman exposure to high doses of radiation resulting in acute radiation syndrome and death can rapidly escalate to a mass casualty catastrophe in the event of nuclear accidents or terrorism. The primary reason is that there is presently no effective treatment option, especially for radiation-induced gastrointestinal syndrome. This syndrome results from disruption of mucosal barrier integrity leading to severe dehydration, blood loss, and sepsis. In this study, we tested whether extracellular vesicles derived from mesenchymal stromal cells (MSC) could reduce radiation-related mucosal barrier damage and reduce radiation-induced animal mortality.MethodsHuman MSC-derived extracellular vesicles were intravenously administered to NUDE mice, 3, 24, and 48 h after lethal whole-body irradiation (10 Gy). Integrity of the small intestine epithelial barrier was assessed by morphologic analysis, immunostaining for tight junction protein (claudin-3), and in vivo permeability to 4 kDa FITC-labeled dextran. Renewal of the small intestinal epithelium was determined by quantifying epithelial cell apoptosis (TUNEL staining) and proliferation (Ki67 immunostaining). Statistical analyses were performed using one-way ANOVA followed by a Tukey test. Statistical analyses of mouse survival were performed using Kaplan-Meier and Cox methods.ResultsWe demonstrated that MSC-derived extracellular vesicle treatment reduced by 85% the instantaneous mortality risk in mice subjected to 10 Gy whole-body irradiation and so increased their survival time. This effect could be attributed to the efficacy of MSC-derived extracellular vesicles in reducing mucosal barrier disruption. We showed that the MSC-derived extracellular vesicles improved the renewal of the small intestinal epithelium by stimulating proliferation and inhibiting apoptosis of the epithelial crypt cells. The MSC-derived extracellular vesicles also reduced radiation-induced mucosal permeability as evidenced by the preservation of claudin-3 immunostaining at the tight junctions of the epithelium.ConclusionsMSC-derived extracellular vesicles promote epithelial repair and regeneration and preserve structural integrity of the intestinal epithelium in mice exposed to radiation-induced gastrointestinal toxicity. Our results suggest that the administration of MSC-derived extracellular vesicles could be an effective therapy for limiting acute radiation syndrome.

Role of mesenchymal stem cell derived extracellular vesicles in autoimmunity: A systematic review.

BACKGROUNDMesenchymal stem cells (MSCs) have been reported to possess immune regulatory effects in innate and adaptive immune reactions. MSCs can mediate intercellular communications by releasing extracellular vesicles (EVs), which deliver functional molecules to targeted cells. MSC derived EVs (MSC-EVs) confer altering effects on many immune cells, including T lymphocytes, B lymphocytes, natural killer cells, dendritic cells, and macrophages. A large number of studies have suggested that MSC-EVs participate in regulating autoimmunity related diseases. This characteristic of MSC-EVs makes them be potential biomarkers for the diagnosis and treatment of autoimmunity related diseases.AIMTo verify the potential of MSC-EVs for molecular targeted therapy of autoimmunity related diseases.METHODSLiterature search was conducted in PubMed to retrieve the articles published between 2010 and 2020 in the English language. The keywords, such as “MSCs,” “EVs,” “exosome,” “autoimmunity,” “tumor immunity,” and “transplantation immunity,” and Boolean operator “AND” and “NOT” coalesced admirably to be used for searching studies on the specific molecular mechanisms of MSC-EVs in many immune cell types and many autoimmunity related diseases. Studies that did not investigate the molecular mechanisms of MSC-EVs in the occurrence and development of autoimmune diseases were excluded.RESULTSA total of 96 articles were chosen for final reference lists. After analyzing those publications, we found that it had been well documented that MSC-EVs have the ability to induce multiple immune cells, like T lymphocytes, B lymphocytes, natural killer cells, dendritic cells, and macrophages, to regulate immune responses in innate immunity and adaptive immunity. Many validated EVs-delivered molecules have been identified as key biomarkers, such as proteins, lipids, and nucleotides. Some EVs-encapsulated functional molecules can serve as promising therapeutic targets particularly for autoimmune disease.CONCLUSIONMSC-EVs play an equally important part in the differentiation, activation, and proliferation of immune cells, and they may become potential biomarkers for diagnosis and treatment of autoimmunity related diseases.