Extracellular vesicle最新文献

{"title":"An extracellular vesicle based hypothesis for the genesis of the polycystic kidney diseases","authors":"Marie C. Hogan , Christopher J. Ward","doi":"10.1016/j.vesic.2024.100048","DOIUrl":"10.1016/j.vesic.2024.100048","url":null,"abstract":"<div><p>Autosomal dominant polycystic kidney (ADPKD) disease is the commonest genetic cause of kidney failure (affecting 1:800 individuals) and is due to heterozygous germline mutations in either of two genes, <em>PKD1</em> and <em>PKD2</em>. Homozygous germline mutations in <em>PKHD1</em> are responsible for autosomal recessive polycystic kidney (ARPKD) disease a rare (1:20,000) but severe neonatal disease. The products of these three genes, <em>PKD1</em> (polycystin-1 (PC1 4302(3)aa)), <em>PKD2</em> (polycystin-2 (PC2 968aa)) and <em>PKHD1</em> (fibrocystin (4074aa)) are all present on extracellular vesicles (EVs) termed, PKD-exosome-like vesicles (PKD-ELVs). PKD-ELVs are defined as 100<!--> <!-->nm PC1/PC2/CD133 and fibrocystin positive EVs which are shed into the urine from the apical plasma membrane of proximal tubule (PT) cells. PKD-ELVs are therefore ectosomes and are distinct from classical exosomes from the multivesicular body. PC1, PC2, fibrocystin and exosomal polycystin-1 interacting protein (EPIC, CU062) form a higher order ion channel complex termed the polycystin complex (PCC) on the surface of the PKD-ELV. We hypothesize that the PCC is involved in the generation of the PKD-ELV and is a structural component thereof. The PCC has also been detected on the primary cilium, a hair like 9+0 microtubule based structure present on all cells except hepatocytes. In kidney epithelial cells, the primary cilium protrudes into the lumen of the tubule where it regulates planar cell polarity (PCP) and tubule lumen diameter. Here we present a theory that explains the presence of the PCC on PKD-ELVs and primary cilia as well as other cryptic aspects of ADPKD and ARPKD. We suggest that the fundamental role of the PCC is to assemble PKD-ELVs on the plasma membrane and then shed them into the extracellular space or the lumen of the tubule. The resultant PKD-ELVs can have multiple functions in different biological contexts. One of the roles of the resultant PKD-ELVs is to generate a planar cell polarity (PCP) signaling gradient along kidney tubules in developing or regenerating kidney. This is mediated via an adhesion event between the PKD-ELV and primary cilium. Defects in the primary cilium or PKD-ELV assembly lead to cystogenesis, the major feature of ADPKD. The other important role for the PCC dependent PKD-ELV is the detection, packaging and extrusion of defective mitochondria. The PKD-ELV is also critical in the transfer of mRNA and miRNAs between cells and as a vector for extracellular proteinases and hyaluronidases involved in tissue remodeling. A PKD-ELV centric view of polycystic disease (EV theory) can explain the requirement for primary cilium function in ADPKD (where the primary cilium is the PKD-ELV receptor), the observation of defective mitochondria in the disease, the abnormalities detected in the extracellular matrix (ECM) as well as the resistance to carcinoma noted in ADPKD patients and individuals carrying <em>PKHD1</em> mutations,","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100048"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2773041724000155/pdfft?md5=48b923544849d6c3c1821ca5848edc6c&pid=1-s2.0-S2773041724000155-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141954453","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Stem cell-derived exosomes prevent the development of thoracic aortic aneurysm/dissection by inhibiting AIM2 inflammasome and pyroptosis","authors":"Lin Lu ,&nbsp;Feng Liu ,&nbsp;Weiliang Wu ,&nbsp;Yu Zhang ,&nbsp;Bin Liu ,&nbsp;Qingfang Han ,&nbsp;Tonggan Lu ,&nbsp;Huiling Zhang ,&nbsp;Xi-yong Yu ,&nbsp;Yangxin Li","doi":"10.1016/j.vesic.2024.100046","DOIUrl":"https://doi.org/10.1016/j.vesic.2024.100046","url":null,"abstract":"<div><p>Thoracic aortic aneurysm/dissection (TAAD) is a severe vascular condition associated with life-threatening complications, and its underlying molecular mechanisms remain largely unexplored. Previous research indicates that the aberrant activation of cytosolic DNA and its receptors plays a crucial role in vascular inflammation and dysfunction. Specifically, Absent in Melanoma 2 (AIM2), an intracellular DNA receptor, can trigger the inflammasome pathway, leading to extracellular matrix destruction. In this investigation, we delved into the mechanism underlying AIM2 activation in TAAD development and explored the potential of exosomes to impede TAAD progression by suppressing AIM2 expression. Our findings revealed that heightened AIM2 expression and activation contribute to TAAD development by fostering vascular inflammation and disrupting vascular homeostasis. Activated AIM2 induces pyroptosis through the recruitment of the deubiquitination enzyme USP21, which stabilizes AIM2 by reducing its ubiquitination and degradation. Moreover, we demonstrated that exosome-derived miR-485-5p exerts an anti-inflammatory and protective effect on the thoracic aorta by inhibiting AIM2 activation. This study introduces novel perspectives for the treatment of TAAD.</p></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100046"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2773041724000131/pdfft?md5=8a900f4bbb54766ace6f15d1f225c232&pid=1-s2.0-S2773041724000131-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141484849","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Ultrasonication outperforms electroporation for extracellular vesicle cargo depletion","authors":"Yundi Chen ,&nbsp;Le Wang ,&nbsp;Xu Yu ,&nbsp;Wenjun Mao ,&nbsp;Yuan Wan","doi":"10.1016/j.vesic.2024.100052","DOIUrl":"10.1016/j.vesic.2024.100052","url":null,"abstract":"<div><div>Extracellular vesicles (EVs), submicron-sized membranous structures released by cells, serve as vehicles of tissue-specific proteins and nucleic acids, facilitating intercellular communication and playing roles in pathophysiological processes. Leveraging their unique characteristics, EVs have emerged as promising drug delivery nanocarriers. Electroporation (EP) and ultrasonication (US) are among the prevalent techniques used for loading exogenous drugs into EVs owing to their simplicity and efficiency. However, the effectiveness of the two methods in depleting initial EV cargo has been overlooked. But this information is indispensable, as the bioactive residuals of EVs, notably derived from tumor or stem donor cells, may impact downstream therapeutic effects. Bridging this knowledge gap, therefore, can guide the selection of optimal drugs and loading methods tailored to therapeutic objectives. Here, we used high-throughput sequencing to investigate the protein and small RNA cargo of EVs treated with EP and US, respectively. We found that US exhibits higher efficacy in depleting EV cargo compared to EP, while US may also deplete essential endogenous molecules for combination therapy. Neither method demonstrated significant selectivity in cargo depletion, but they might preferentially retain few specific molecules. Additionally, membrane proteins are more prone to loss during US and EP treatments than cytoplasmic proteins.</div></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100052"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142424307","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"From isolation to detection, advancing insights into endothelial matrix-bound vesicles","authors":"Sahimy Ayus-Martinez ,&nbsp;William Meza-Morales ,&nbsp;Jesus Jimenez-Osorio ,&nbsp;Maria Buendia-Otero ,&nbsp;Luis López ,&nbsp;Lisandro Cunci ,&nbsp;Donald O. Freytes ,&nbsp;Camilo Mora","doi":"10.1016/j.vesic.2024.100060","DOIUrl":"10.1016/j.vesic.2024.100060","url":null,"abstract":"<div><div>Matrix-bound vesicles (MBVs), an integral part of the extracellular matrix (ECM), are emerging as pivotal factors in ECM-driven molecular signaling. This study is the first to report the isolation of MBVs from porcine arterial endothelial cell basement membranes (A-MBVs) and thyroid cartilage (C-MBVs), the latter serving as a negative control due to its minimal vascular characteristics. Using Transmission Electron Microscopy (TEM), Nano-Tracking Analysis (NTA), Electrochemical Impedance Spectroscopy (EIS), and Atomic Force Microscopy (AFM), we orthogonally characterized the isolated MBVs. We detected the presence and preservation of vascular endothelial cadherin (CD144) in A-MBVs, its low to non-detetcted in C-MBVs, in which SOX9, a chondrocyte marker, was detected. Moreover, we developed a prototype of an immuno-functionalized screen-printed electrode designed for the immunoadsorption of CD144+ MBVs. This device facilitated the electrochemical detection of the targeted vesicles and allowed for their subsequent topological characterization using AFM, which verified the integrity and morphology of CD144+ MBVs post-immunoadsorption. These advancements enhance our comprehension of MBVs as conveyors of tissue-specific signals and pioneer new avenues for harnessing their cargo in biomedical applications. This research sets a significant precedent for future studies on the application of MBVs in regenerative medicine and ECM signaling.</div></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100060"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11759483/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143048614","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Therapeutic potential of human stem cell-derived extracellular vesicles in idiopathic pulmonary fibrosis","authors":"Kyoung Soo Lee ,&nbsp;Seung Ho Yeom ,&nbsp;Min Kang Kim ,&nbsp;Chang Hee Woo ,&nbsp;Young Chan Choi ,&nbsp;Ji Suk Choi ,&nbsp;Yong Woo Cho","doi":"10.1016/j.vesic.2024.100045","DOIUrl":"https://doi.org/10.1016/j.vesic.2024.100045","url":null,"abstract":"<div><p>Idiopathic pulmonary fibrosis (IPF) is a lethal and chronic lung disease that occurs due to persistent epithelial cell injury and abnormal extracellular matrix (ECM) deposition. Extracellular vesicles (EVs) have been proposed as a potential therapeutic option of IPF because of their functions, such as anti-inflammation, anti-fibrosis, microenvironment regulation, and tissue repair. In this study, we investigated the therapeutic potential of human adipose-derived mesenchymal stem cell (AD-MSC) EVs in IPF model. AD-MSC EVs were isolated by a multi-filtration system based on the tangential flow filtration (TFF) and characterized using transmission electron microscopy (TEM), dynamic light scattering (DLS), flow cytometry analysis, zeta potential, and small RNA sequencing. <em>In vitro</em> analysis reveals that AD-MSC EVs treatments inhibited migration of pulmonary fibroblasts and myofibroblast differentiation through down regulation of fibrosis-associated TGF-β and WNT signaling. In addition, inhalation treatment of AD-MSC EVs significantly alleviated bleomycin (BLM)-induced pulmonary fibrosis. These results reveal that AD-MSC EVs are highly promising as a treatment option for pulmonary fibrosis.</p></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100045"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S277304172400012X/pdfft?md5=d3ec1dc4b1a879c67ab2d843a901c474&pid=1-s2.0-S277304172400012X-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141484868","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Application of miR-29a-Exosome and multifunctional scaffold for full-thickness cartilage defects","authors":"Fan Yang ,&nbsp;Zewen Wang ,&nbsp;Mingjian Wu,&nbsp;Jingyi Xu,&nbsp;Junlei Li,&nbsp;Jiahe Liu,&nbsp;Ting He,&nbsp;Tao Zhang,&nbsp;Baoyi Liu","doi":"10.1016/j.vesic.2024.100055","DOIUrl":"10.1016/j.vesic.2024.100055","url":null,"abstract":"<div><h3>Background</h3><div>Full-thickness cartilage defect is a common refractory disease in orthopedics. In this study, we designed a novel composite scaffold composed of silk fibroin-chitosan for the cartilage layer and porous tantalum for the subchondral bone layer, loaded with engineered bone mesenchymal stem cell exosomes, to evaluate its efficacy in repairing full-thickness cartilage defect.</div></div><div><h3>Methods</h3><div>Porous tantalum was 3D printed and combined with silk fibroin-chitosan to form a composite scaffold. Chondrocytes were cultured on the scaffold, and their growth was assessed using the CCK-8 method. Toluidine blue staining confirmed cell morphology, while immunofluorescence revealed collagen type Ⅱ expression. Engineered exosomes loaded with miR-29a were created and characterized using various techniques. Co-culturing with chondrocytes demonstrated their proliferation over 10 days. Immunofluorescence revealed staining for the nucleus, collagen type II, and Aggrecan. In vivo experiments were performed on rats to assess cartilage defect repair, utilizing histological staining and micro-CT scanning at 4 and 8 weeks post-operation.</div></div><div><h3>Results</h3><div>The silk fibroin-chitosan scaffold demonstrated good biocompatibility, supporting chondrocyte adhesion, growth, and cartilage tissue formation. Engineered exosomes exhibited promising biological activity, conducive to bone and cartilage regeneration. The implantation of the silk fibroin-chitosan/porous tantalum composite scaffold loaded with engineered exosomes promoted integration with the surrounding bone and cartilage tissues, facilitating repair and regeneration.</div></div><div><h3>Conclusions</h3><div>The silk fibroin-chitosan combined with porous tantalum scaffold carrying engineered exosomes loaded with miR-29a has good potential for full-thickness cartilage defects regeneration.</div></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100055"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142593391","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Characterization of plasma-derived small extracellular vesicle miRNA and protein cargo in hereditary angioedema","authors":"Linda Hofmann ,&nbsp;Robin Lochbaum ,&nbsp;Lutz Schütt ,&nbsp;Ralph Röth ,&nbsp;Stefanie Schmitteckert ,&nbsp;Barbara Wollenberg ,&nbsp;Thomas K. Hoffmann ,&nbsp;Cornelia Brunner ,&nbsp;Jens Greve ,&nbsp;Janina Hahn ,&nbsp;Marie-Nicole Theodoraki","doi":"10.1016/j.vesic.2024.100056","DOIUrl":"10.1016/j.vesic.2024.100056","url":null,"abstract":"<div><div>Hereditary angioedema (HAE) is an inherited disorder causing attacks of subcutaneous tissue or mucosa swelling. The disease burden and attack frequencies vary significantly among patients. This is the first pilot study investigating small extracellular vesicles (sEV) as potential disease modulators in HAE.</div><div>Plasma-derived sEVs from HAE patients and healthy donors (HD) were thoroughly characterized by Western blot, transmission electron microscopy, nanoparticle tracking and bead-based flow cytometry. The miRNA content of sEVs was examined by nCounter technology and used to predict sEV-based pathomechanisms <em>in silico</em>. All sEV readouts were analyzed regarding HAE-related changes and associations with clinical parameters and attack frequency.</div><div>Total sEV protein levels were elevated in HAE patients compared to HD. In HAE patients, lower levels of sEVs carrying CD8, CD209, CD81, CD24 and CD44 were measured. sEV miRNA profiling revealed 84 HAE-exclusive and 30 significantly HAE-upregulated candidates. Core hubs of their predicted interaction networks were AGO2, VEGF, RGS5, MTA1, IFG1 and BAX. A set of 12 and 36 sEV miRNAs were restricted to patients with absent attacks or patients with present attacks during prophylactic therapy, respectively.</div><div>sEVs, especially sEV miRNAs, could contribute to disease pathogenesis and differential attack frequencies. They emerged as disease modulators in HAE and require further study to reveal underlying mechanisms.</div></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100056"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142657127","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A bibliometric analysis of extracellular vesicles as drug delivery vehicles in disease treatment (2010–2024)","authors":"Wuli Guo ,&nbsp;Qi Shu ,&nbsp;Lina Gao,&nbsp;Na Gao,&nbsp;Zhen Wang,&nbsp;Wenjing Wei,&nbsp;Yuhan Zhang,&nbsp;Ting Huyan,&nbsp;Qi Li","doi":"10.1016/j.vesic.2024.100051","DOIUrl":"10.1016/j.vesic.2024.100051","url":null,"abstract":"<div><div>To comprehensively summarize the progress of utilizing extracellular vesicles (EVs) as drug delivery vehicles in disease treatment, bibliometric and visualization tools were employed to analyze and exhibit the key information from 6515 literatures that collected by searching “drug delivery” and “extracellular vesicles”, as well as “drug delivery” and “exosomes” as the keywords in Web of Science Core Collection. The obtained articles are from 90 countries/regions, 3343 institutions, 17,989 authors, and 591 journals. Among them, the “nanoparticles”, “cancer” and “biomarkers” are the keywords with the highest frequency. According to experimental articles, EVs are mainly used in the treatment of cancer as drug vehicles.</div></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100051"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142318709","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Role of Extracellular vesicle microRNAs and RNA binding proteins on glioblastoma dynamics and therapeutics development","authors":"Shilpi Singh ,&nbsp;Clark C. Chen ,&nbsp;Stefan Kim ,&nbsp;Amar Singh ,&nbsp;Gatikrushna Singh","doi":"10.1016/j.vesic.2024.100049","DOIUrl":"10.1016/j.vesic.2024.100049","url":null,"abstract":"<div><p>The intricate interplay between extracellular vesicles (EVs), microRNAs, and RNA binding proteins (RBPs) constitutes a crucial mechanism in glioblastoma pathogenesis. The regulated sorting of microRNAs into EVs, influenced by RBPs and specific RNA motifs, plays a pivotal role in intercellular communication and significantly impacts various biological processes associated with glioblastoma development. Glioblastoma cells exploit this mechanism to exchange genetic information, influencing tumor progression, heterogeneity, and treatment response. This review highlights the dysregulation of EV-encapsulated microRNAs as a modulator of key signaling the disease. The versatile role of EV-microRNAs extends to impacting glioma behavior, immune response, angiogenesis, and serving as valuable diagnostic, prognostic, and therapeutic targets. Furthermore, the involvement of RBPs in microRNA sorting into EVs presents therapeutic targets that bridge cellular signaling with EV cargo composition. This comprehensive understanding and the strategic approaches targeting the complex interactions within the EV-microRNA-RBP axis opens promising avenues for advancing diagnostics and developing targeted therapies against the heterogeneous nature of glioblastoma and enhance treatment efficacy. The personalized and effective treatment strategies that may emerge from this research have the potential to revolutionize the approach to combating this devastating brain cancer.</p></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"4 ","pages":"Article 100049"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2773041724000167/pdfft?md5=65f1184d771358a37b490b40b3ddee79&pid=1-s2.0-S2773041724000167-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142161956","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"A simple scalable extracellular vesicle isolation method using polyethylenimine polymers for use in cellular delivery","authors":"Marie Ange Djeungoue Petga ,&nbsp;Catherine Taylor ,&nbsp;Alexander Macpherson ,&nbsp;Surendar Reddy Dhadi ,&nbsp;Thomas Rollin ,&nbsp;Jeremy W. Roy ,&nbsp;Anirban Ghosh ,&nbsp;Stephen M. Lewis ,&nbsp;Rodney J. Ouellette","doi":"10.1016/j.vesic.2023.100033","DOIUrl":"https://doi.org/10.1016/j.vesic.2023.100033","url":null,"abstract":"<div><p>Extracellular vesicles (EVs) are gaining interest as efficient, biocompatible vehicles for cellular delivery of therapeutic cargo. Precipitation-based methods for the isolation of EVs remain popular due to ease of use and lack of requirements for specialized equipment. We describe here a novel charge-based EV isolation method that is simple, scalable, and uses inexpensive polyethylenimine (PEI) polymers. GFP-expressing EVs were isolated from the conditioned cell culture (CCM) media of HEK293-GFP cells using either branched 10 kDa PEI (B-PEI) or linear 25 kDa PEI (L-PEI). Isolated EVs were characterized by Western blotting, nanoparticle tracking analysis, transmission electron microscopy (TEM), and flow cytometry. Western blotting for common EV markers, including CD63, CD9, flotillin-1, and heat shock protein 70 were positive, while GRP94, a marker for cellular contamination, was negative. Isolated EVs had a mean diameter of 146 nm for B-PEI and 175 nm for L-PEI, while TEM revealed a spherical cup-shaped appearance typical of EVs. In addition, we determined that PEI-based EV isolation methods were scalable up to volumes of at least 50 mL. EVs isolated from CCM collected from SUM159 cells that express CD63 fused to a dual EGFP-Renilla-split tag were tested for their ability to reconstitute functional luciferase by delivering the CD63-EGFP-Renilla-split tag to SUM159 recipient cells loaded with a cytopermeable Renilla luciferase substrate. Although EVs isolated using L-PEI behaved similarly to EVs isolated using ultracentrifugation, we observed that EVs isolated using B-PEI produced a more rapid uptake and delivery of active luciferase. In this study we demonstrate that both branched and linear PEI polymers can precipitate EVs from CCM. Furthermore, once eluted from the polymers, the isolated EVs were able to deliver functional protein cargo to recipient cells. Overall, our data support PEI-based isolation of EVs as a simple, rapid method for the recovery of functional EVs.</p></div>","PeriodicalId":73007,"journal":{"name":"Extracellular vesicle","volume":"3 ","pages":"Article 100033"},"PeriodicalIF":0.0,"publicationDate":"2024-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2773041723000124/pdfft?md5=c7feb4086e774e807af0d19f7fcc7a6f&pid=1-s2.0-S2773041723000124-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139504234","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}