Extracellular vesicles (EVs) are nanosized lipid bilayer vesicles that are released by almost all cell types. They range in diameter from 30 nm to several micrometres and have the ability to carry ...biologically active molecules such as proteins, lipids, RNA, and DNA. EVs are natural vectors and play an important role in many physiological and pathological processes. The amount and composition of EVs in human biological fluids serve as biomarkers and are used for diagnosing diseases and monitoring the effectiveness of treatment. EVs are promising for use as therapeutic agents and as natural vectors for drug delivery. However, the successful use of EVs in clinical practice requires an understanding of their biodistribution in an organism. Numerous studies conducted so far on the biodistribution of EVs show that, after intravenous administration, EVs are mostly localized in organs rich in blood vessels and organs associated with the reticuloendothelial system, such as the liver, lungs, spleen, and kidneys. In order to improve resolution, new dyes and labels are being developed and detection methods are being optimized. In this work, we review all available modern methods and approaches used to assess the biodistribution of EVs, as well as discuss their advantages and limitations.
The immunosuppressive potential of mesenchymal stem cells has been extensively investigated in many studies
and
. In recent years, a variety preclinical and clinical studies have demonstrated that ...mesenchymal stem cells ameliorate immune-mediated disorders, including autoimmune diseases. However, to date mesenchymal stem cells have not become a widely used therapeutic agent due to safety challenges, high cost and difficulties in providing long term production. A key mechanism underpinning the immunomodulatory effect of MSCs is the production of paracrine factors including growth factors, cytokines, chemokines, and extracellular vesicles (EVs). MSCs derived EVs have become an attractive therapeutic agent for immunomodulation and treatment of immune-mediated disorders. In addition to many preclinical studies of MSCs derived EVs, their beneficial effects have been observed in patients with both acute graft-vs.-host disease and chronic kidney disease. In this review, we discuss the current findings in the field of MSCs derived EVs-based therapies in immune-mediated disorders and approaches to scale EV production for clinical use.
Ultrahigh-throughput screening (uHTS) techniques can identify unique functionality from millions of variants. To mimic the natural selection mechanisms that occur by compartmentalization in vivo, we ...developed a technique based on single-cell encapsulation in droplets of a monodisperse microfluidic double water-in-oil-in-water emulsion (MDE). Biocompatible MDE enables in-droplet cultivation of different living species. The combination of droplet-generating machinery with FACS followed by next-generation sequencing and liquid chromatography-mass spectrometry analysis of the secretomes of encapsulated organisms yielded detailed genotype/phenotype descriptions. This platform was probed with uHTS for biocatalysts anchored to yeast with enrichment close to the theoretically calculated limit and cell-to-cell interactions. MDE–FACS allowed the identification of human butyrylcholinesterase mutants that undergo self-reactivation after inhibition by the organophosphorus agent paraoxon. The versatility of the platform allowed the identification of bacteria, including slow-growing oral microbiota species that suppress the growth of a common pathogen, Staphylococcus aureus, and predicted which genera were associated with inhibitory activity.
Strategies for Engineering of Extracellular Vesicles Danilushkina, Anna A; Emene, Charles C; Barlev, Nicolai A ...
International journal of molecular sciences,
09/2023, Letnik:
24, Številka:
17
Journal Article
Recenzirano
Odprti dostop
Extracellular vesicles (EVs) are membrane vesicles released by cells into the extracellular space. EVs mediate cell-to-cell communication through local and systemic transportation of biomolecules ...such as DNA, RNA, transcription factors, cytokines, chemokines, enzymes, lipids, and organelles within the human body. EVs gained a particular interest from cancer biology scientists because of their role in the modulation of the tumor microenvironment through delivering bioactive molecules. In this respect, EVs represent an attractive therapeutic target and a means for drug delivery. The advantages of EVs include their biocompatibility, small size, and low immunogenicity. However, there are several limitations that restrict the widespread use of EVs in therapy, namely, their low specificity and payload capacity. Thus, in order to enhance the therapeutic efficacy and delivery specificity, the surface and composition of extracellular vesicles should be modified accordingly. In this review, we describe various approaches to engineering EVs, and further discuss their advantages and disadvantages to promote the application of EVs in clinical practice.
The phenomenon of mitochondria donation is found in various tissues of humans and animals and is attracting increasing attention. To date, numerous studies have described the transfer of mitochondria ...from stem cells to injured cells, leading to increased ATP production, restoration of mitochondria function, and rescue of recipient cells from apoptosis. Mitochondria transplantation is considered as a novel therapeutic approach for the treatment of mitochondrial diseases and mitochondrial function deficiency. Mitochondrial dysfunction affects cells with high energy needs such as neural, skeletal muscle, heart, and liver cells and plays a crucial role in type 2 diabetes, as well as Parkinson's, Alzheimer's diseases, ischemia, stroke, cancer, and age-related disorders. In this review, we summarize recent findings in the field of mitochondria donation and mechanism of mitochondria transfer between cells. We review the existing clinical trials and discuss advantages and disadvantages of mitochondrial transplantation strategies based on the injection of stem cells, isolated functional mitochondria, or EVs containing mitochondria.
The immune system plays a crucial role in recognizing and eliminating altered tumor cells. However, tumors develop mechanisms to evade the body's natural immune defenses. Therefore, methods for ...specifically recognizing/targeting tumor cells, for instance, through the activation, directed polarization, and training of immune cells, have been developed based on the body's immune cells. This strategy has been termed cellular immunotherapy. One promising strategy for treating tumor diseases is NK cell-based immunotherapy. NK cells have the ability to recognize and destroy transformed cells without prior activation as well as tumor cells with reduced MHC-I expression. A novel approach in immunotherapy is the use of extracellular vesicles (EVs) derived from NK cells. The main advantages of NK cell-derived EVs are their small size and better tissue penetration into a tumor. The aim of this review is to systematically present existing information on the mechanisms of antitumor immunity and the role of NK cells and extracellular vesicles in cancer immunotherapy. Clinical and preclinical studies utilizing NK cells and extracellular vesicles for anticancer therapy currently underway will provide valuable insights for researchers in the field of cancer.
Extracellular vesicles (EVs) are promising therapeutic instruments and vectors for therapeutics delivery. In order to increase the yield of EVs, a method of inducing EVs release using cytochalasin B ...is being actively developed. In this work, we compared the yield of naturally occurring extracellular vesicles and cytochalasin B-induced membrane vesicles (CIMVs) from mesenchymal stem cells (MSCs). In order to maintain accuracy in the comparative analysis, the same culture was used for the isolation of EVs and CIMVs: conditioned medium was used for EVs isolation and cells were harvested for CIMVs production. The pellets obtained after centrifugation 2300×
, 10,000×
and 100,000×
were analyzed using scanning electron microscopy analysis (SEM), flow cytometry, the bicinchoninic acid assay, dynamic light scattering (DLS), and nanoparticle tracking analysis (NTA). We found that the use of cytochalasin B treatment and vortexing resulted in the production of a more homogeneous population of membrane vesicles with a median diameter greater than that of EVs. We found that EVs-like particles remained in the FBS, despite overnight ultracentrifugation, which introduced a significant inaccuracy in the calculation of the EVs yield. Therefore, we cultivated cells in a serum-free medium for the subsequent isolation of EVs. We observed that the number of CIMVs significantly exceeded the number of EVs after each step of centrifugation (2300×
, 10,000×
and 100,000×
) by up to 5, 9, and 20 times, respectively.
Stimulating the process of angiogenesis in treating ischemia-related diseases is an urgent task for modern medicine, which can be achieved through the use of different cell types. Umbilical cord ...blood (UCB) continues to be one of the attractive cell sources for transplantation. The goal of this study was to investigate the role and therapeutic potential of gene-engineered umbilical cord blood mononuclear cells (UCB-MC) as a forward-looking strategy for the activation of angiogenesis. Adenovirus constructs Ad-VEGF, Ad-FGF2, Ad-SDF1α, and Ad-EGFP were synthesized and used for cell modification. UCB-MCs were isolated from UCB and transduced with adenoviral vectors. As part of our in vitro experiments, we evaluated the efficiency of transfection, the expression of recombinant genes, and the secretome profile. Later, we applied an in vivo Matrigel plug assay to assess engineered UCB-MC's angiogenic potential. We conclude that hUCB-MCs can be efficiently modified simultaneously with several adenoviral vectors. Modified UCB-MCs overexpress recombinant genes and proteins. Genetic modification of cells with recombinant adenoviruses does not affect the profile of secreted pro- and anti-inflammatory cytokines, chemokines, and growth factors, except for an increase in the synthesis of recombinant proteins. hUCB-MCs genetically modified with therapeutic genes induced the formation of new vessels. An increase in the expression of endothelial cells marker (CD31) was revealed, which correlated with the data of visual examination and histological analysis. The present study demonstrates that gene-engineered UCB-MC can be used to stimulate angiogenesis and possibly treat cardiovascular disease and diabetic cardiomyopathy.
One of the most promising approaches to stimulate regeneration and angiogenesis in traumatic or ischemic tissue damage is stem cell therapy. Embryonic and fetal stem cells have the greatest potential ...of differentiation into different cell types; however, at the same time, they carry the highest risk of teratoma formation. Adult stem cells have the potential risk of transformation during prolonged cultivation in vitro, or as a result of genetic changes during gene-cell therapy applications. In this regard, technologies that can reduce the potential risks of cell and gene-cell therapy are of particular interest. According to the paracrine hypothesis, the beneficial effect of stem cell therapy is due to stimulation of resident cells by cell-to-cell contacts, secretion of bioactive molecules, and release of extracellular vesicles. In this review, we discuss the development of regenerative medicine from cell to cell-free therapy based on extracellular vesicles.
High-dose recombinant interleukin 2 (IL2) therapy has been shown to be successful in renal cell carcinoma and metastatic melanoma. However, systemic administration of high doses of IL2 can be toxic, ...causing capillary leakage syndrome and stimulating pro-tumor immune response. One of the strategies to reduce the systemic toxicity of IL2 is the use of mesenchymal stem cells (MSCs) as a vehicle for the targeted delivery of IL2. Human adipose tissue-derived MSCs were transduced with lentivirus encoding IL2 (hADSCs-IL2) or blue fluorescent protein (BFP) (hADSCs-BFP). The proliferation, immunophenotype, cytokine profile and ultrastructure of hADSCs-IL2 and hADSCs-BFP were determined. The effect of hADSCs on activation of peripheral blood mononuclear cells (PBMCs) and proliferation and viability of SH-SY5Y neuroblastoma cells after co-culture with native hADSCs, hADSCs-BFP or hADSCs-IL2 on plastic and Matrigel was evaluated. Ultrastructure and cytokine production by hADSCs-IL2 showed modest changes in comparison with hADSCs and hADSCs-BFP. Conditioned medium from hADSC-IL2 affected tumor cell proliferation, increasing the proliferation of SH-SY5Y cells and also increasing the number of late-activated T-cells, natural killer (NK) cells, NKT-cells and activated T-killers. Conversely, hADSC-IL2 co-culture led to a decrease in SH-SY5Y proliferation on plastic and Matrigel. These data show that hADSCs-IL2 can reduce SH-SY5Y proliferation and activate PBMCs in vitro. However, IL2-mediated therapeutic effects of hADSCs could be offset by the increased expression of pro-oncogenes, as well as the natural ability of hADSCs to promote the progression of some tumors.
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