Journal of extracellular biology最新文献

{"title":"Bone marrow-fibroblast progenitor cell-derived small extracellular vesicles promote cardiac fibrosis via miR-21-5p and integrin subunit αV signalling","authors":"Prabhat Ranjan,&nbsp;Roshan Kumar Dutta,&nbsp;Karen Colin,&nbsp;Jing Li,&nbsp;Qinkun Zhang,&nbsp;Hind Lal,&nbsp;Gangjian Qin,&nbsp;Suresh Kumar Verma","doi":"10.1002/jex2.152","DOIUrl":"https://doi.org/10.1002/jex2.152","url":null,"abstract":"<p>Cardiac fibrosis is the hallmark of cardiovascular disease (CVD), which is leading cause of death worldwide. Previously, we have shown that interleukin-10 (IL10) reduces pressure overload (PO)-induced cardiac fibrosis by inhibiting the recruitment of bone marrow fibroblast progenitor cells (FPCs) to the heart. However, the precise mechanism of FPC involvement in cardiac fibrosis remains unclear. Recently, exosomes and small extracellular vesicles (sEVs) have been linked to CVD progression. Thus, we hypothesized that pro-fibrotic miRNAs enriched in sEV-derived from IL10 KO FPCs promote cardiac fibrosis in pressure-overloaded myocardium. Small EVs were isolated from FPCs cultured media and characterized as per MISEV-2018 guidelines. Small EV's miRNA profiling was performed using Qiagen fibrosis-associated miRNA profiler kit. For functional analysis, sEVs were injected in the heart following TAC surgery. Interestingly, TGFβ-treated IL10-KO-FPCs sEV increased profibrotic genes expression in cardiac fibroblasts. The exosomal miRNA profiling identified miR-21a-5p as the key player, and its inhibition with antagomir prevented profibrotic signalling and fibrosis. At mechanistic level, miR-21a-5p binds and stabilizes <i>ITGAV (</i>integrin av) mRNA. Finally, miR-21a-5p-silenced in sEV reduced PO-induced cardiac fibrosis and improved cardiac function. Our study elucidates the mechanism by which inflammatory FPC-derived sEV exacerbate cardiac fibrosis through the miR-21a-5p/ITGAV/Col1α signalling pathway, suggesting miR-21a-5p as a potential therapeutic target for treating hypertrophic cardiac remodelling and heart failure.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 6","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.152","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141441300","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":"Amniotic fluid-derived small extracellular vesicles for predicting postnatal severe outcome of congenital diaphragmatic hernia","authors":"Seiko Matsuo,&nbsp;Akira Yokoi,&nbsp;Kosuke Yoshida,&nbsp;Masami Kitagawa,&nbsp;Eri Asano-Inami,&nbsp;Mayo Miura,&nbsp;Takao Yasui,&nbsp;Sho Tano,&nbsp;Takafumi Ushida,&nbsp;Kenji Imai,&nbsp;Hiroaki Kajiyama,&nbsp;Tomomi Kotani","doi":"10.1002/jex2.160","DOIUrl":"https://doi.org/10.1002/jex2.160","url":null,"abstract":"<p>Congenital diaphragmatic hernia (CDH) is a life-threatening condition with high morbidity and mortality rates. The survival rate of neonates with severe CDH is reportedly only 10%–15%. However, prenatal prediction of severe cases is difficult, and the discovery of new predictive markers is an urgent issue. In this study, we focused on microRNAs (miRNAs) in amniotic fluid-derived small EVs (AF-sEVs). We identified four miRNAs (hsa-miR-127-3p, hsa-miR-363-3p, hsa-miR-493-5p, and hsa-miR-615-3p) with AUC &gt; 0.8 to classify good prognosis group and poor prognosis group in human study. The AUC for hsa-miR-127-3p and hsa-miR-615-3p, for predicting the poor prognosis, were 0.93 and 0.91, respectively. In addition, in the in vivo study, the miRNA profiles of the lung tissues of CDH rats were different from those of control rats. Additionally, two elevated miRNAs (rno-miR-215-5p and rno-miR-148a-3p) in the lung tissues of CDH rats were increased in the AF-sEVs of CDH rats. Our results suggest that severe CDH neonates can be predicted prenatally with high accuracy using miRNAs contained in AF-sEVs. Furthermore, miRNA profile changes in AF-sEVs reflected the lung status in CDH. Our findings may contribute to the development of advanced perinatal care for patients with CDH.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 6","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.160","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141439550","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":"Characterisation of sRNAs enriched in outer membrane vesicles of pathogenic Flavobacterium psychrophilum causing Bacterial Cold Water Disease in rainbow trout","authors":"Pratima Chapagain,&nbsp;Ali Ali,&nbsp;Destaalem T. Kidane,&nbsp;Mary Farone,&nbsp;Mohamed Salem","doi":"10.1002/jex2.161","DOIUrl":"https://doi.org/10.1002/jex2.161","url":null,"abstract":"<p><i>Flavobacterium psychrophilum</i> (<i>Fp</i>) causes Bacterial Cold Water Disease in salmonids. During host-pathogen interactions, gram-negative bacteria, such as <i>Fp</i>, release external membrane vesicles (OMVs) harbouring cargos, such as DNA, RNA and virulence factors. This study aimed to characterise the potential role of the OMVs’ small RNAs (sRNAs) in the <i>Fp</i>-rainbow trout host-pathogen interactions. sRNAs carried within OMVs were isolated from <i>Fp</i>. RNA-Seq datasets from whole-cell <i>Fp</i> and their isolated OMVs indicated substantial enrichment of specific sRNAs in the OMVs compared to the parent cell. Many of the OMV-packaged sRNAs were located in the pathogenicity islands of <i>Fp</i>. Conservation of sRNAs in 65 strains with variable degrees of virulence was reported. Dual RNA-Seq of host and pathogen transcriptomes on day 5 post-infection of <i>Fp</i> -resistant and -susceptible rainbow trout genetic lines revealed correlated expression of OMV-packaged sRNAs and their predicted host's immune gene targets. In vitro, treatment of the rainbow trout epithelial cell line RTgill-W1 with OMVs showed signs of cytotoxicity accompanied by dynamic changes in the expression of host genes when profiled 24 h following treatment. The OMV-treated cells, similar to the <i>Fp</i> -resistant fish, showed downregulated expression of the suppressor of cytokine signalling 1 (SOCS1) gene, suggesting induction of phagosomal maturation. Other signs of modulating the host gene expression following OMV-treatment include favouring elements from the phagocytic, endocytic and antigen presentation pathways in addition to HSP70, HSP90 and cochaperone proteins, which provide evidence for a potential role of OMVs in boosting the host immune response. In conclusion, the study identified novel microbial targets and inherent characteristics of OMVs that could open up new avenues of treatment and prevention of fish infections.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 6","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.161","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141435646","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":"Proteomic profiling of mesenchymal stem cell-derived extracellular vesicles: Impact of isolation methods on protein cargo","authors":"Morteza Abyadeh,&nbsp;Shahab Mirshahvaladi,&nbsp;Sara Assar Kashani,&nbsp;Joao A. Paulo,&nbsp;Ardeshir Amirkhani,&nbsp;Fatemeh Mehryab,&nbsp;Homeyra Seydi,&nbsp;Niloufar Moradpour,&nbsp;Sheyda Jodeiryjabarzade,&nbsp;Mehdi Mirzaei,&nbsp;Vivek Gupta,&nbsp;Faezeh Shekari,&nbsp;Ghasem Hosseini Salekdeh","doi":"10.1002/jex2.159","DOIUrl":"https://doi.org/10.1002/jex2.159","url":null,"abstract":"<p>Extracellular vesicles (EVs) are nanosized vesicles with a lipid bilayer that are secreted by cells and play a critical role in cell-to-cell communication. Despite the promising reports regarding their diagnostic and therapeutic potential, the utilization of EVs in the clinical setting is limited due to insufficient information about their cargo and a lack of standardization in isolation and analysis methods. Considering protein cargos in EVs as key contributors to their therapeutic potency, we conducted a tandem mass tag (TMT) quantitative proteomics analysis of three subpopulations of mesenchymal stem cell (MSC)-derived EVs obtained through three different isolation techniques: ultracentrifugation (UC), high-speed centrifugation (HS), and ultracentrifugation on sucrose cushion (SU). Subsequently, we checked EV marker expression, size distribution, and morphological characterization, followed by bioinformatic analysis. The bioinformatic analysis of the proteome results revealed that these subpopulations exhibit distinct molecular and functional characteristics. The choice of isolation method impacts the proteome of isolated EVs by isolating different subpopulations of EVs. Specifically, EVs isolated through the high-speed centrifugation (HS) method exhibited a higher abundance of ribosomal and mitochondrial proteins. Functional apoptosis assays comparing isolated mitochondria with EVs isolated through different methods revealed that HS-EVs, but not other EVs, induced early apoptosis in cancer cells. On the other hand, EVs isolated using the sucrose cushion (SU) and ultracentrifugation (UC) methods demonstrated a higher abundance of proteins primarily involved in the immune response, cell-cell interactions and extracellular matrix interactions. Our analyses unveil notable disparities in proteins and associated biological functions among EV subpopulations, underscoring the importance of meticulously selecting isolation methods and resultant EV subpopulations based on the intended application.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 6","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-06-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.159","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141286791","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":"Chemo-small extracellular vesicles released in cisplatin-resistance ovarian cancer cells are regulated by the lysosomal function","authors":"Cristóbal Cerda-Troncoso,&nbsp;Felipe Grünenwald,&nbsp;Eloísa Arias-Muñoz,&nbsp;Viviana A. Cavieres,&nbsp;Albano Caceres-Verschae,&nbsp;Sergio Hernández,&nbsp;Belén Gaete-Ramírez,&nbsp;Francisca Álvarez-Astudillo,&nbsp;Rodrigo A. Acuña,&nbsp;Matias Ostrowski,&nbsp;Patricia V. Burgos,&nbsp;Manuel Varas-Godoy","doi":"10.1002/jex2.157","DOIUrl":"https://doi.org/10.1002/jex2.157","url":null,"abstract":"<p>Chemoresistance is a common problem in ovarian cancer (OvCa) treatment, where resistant cells, in response to chemotherapy, secrete small extracellular vesicles (sEVs), known as chemo-sEVs, that transfer resistance to recipient cells. sEVs are formed as intraluminal vesicles (ILVs) within multivesicular endosomes (MVEs), whose trafficking is regulated by Ras-associated binding (RAB) GTPases that mediate sEVs secretion or lysosomal degradation. A decrease in lysosomal function can promote sEVs secretion, but the relationship between MVEs trafficking pathways and sEVs secretion in OvCa chemoresistance is unclear. Here, we show that A2780cis cisplatin (CCDP) resistant OvCa cells had an increased number of MVEs and ILVs structures, higher levels of Endosomal Sorting Complex Required for Transport (ESCRTs) machinery components, and RAB27A compared to A2780 CDDP-sensitive OvCa cells. CDDP promoted the secretion of chemo-sEVs in A2780cis cells, enriched in DNA damage response proteins. A2780cis cells exhibited poor lysosomal function with reduced levels of RAB7, essential in MVEs-Lysosomal trafficking. The silencing of RAB27A in A2780cis cells prevents the Chemo-EVs secretion, reduces its chemoresistance and restores lysosomal function and levels of RAB7, switching them into an A2780-like cellular phenotype. Enhancing lysosomal function with rapamycin reduced chemo-sEVs secretion. Our results suggest that adjusting the balance between secretory MVEs and lysosomal MVEs trafficking could be a promising strategy for overcoming CDDP chemoresistance in OvCa.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 6","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-05-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.157","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141187633","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":"Mesenchymal stem cell-derived extracellular vesicles: Recent therapeutics and targeted drug delivery advances","authors":"Anjali Bhat,&nbsp;Anshu Malik,&nbsp;Poonam Yadav,&nbsp;Wend-Pingda Jessica Ware,&nbsp;Pratiksha Kakalij,&nbsp;Subhash Chand","doi":"10.1002/jex2.156","DOIUrl":"https://doi.org/10.1002/jex2.156","url":null,"abstract":"<p>The targeted drug delivery field is rapidly advancing, focusing on developing biocompatible nanoparticles that meet rigorous criteria of non-toxicity, biocompatibility, and efficient release of encapsulated molecules. Conventional synthetic nanoparticles (SNPs) face complications such as elevated immune responses, complex synthesis methods, and toxicity, which restrict their utility in therapeutics and drug delivery. Extracellular vesicles (EVs) have emerged as promising substitutes for SNPs, leveraging their ability to cross biological barriers, biocompatibility, reduced toxicity, and natural origin. Notably, mesenchymal stem cell-derived EVs (MSC-EVs) have garnered much curiosity due to their potential in therapeutics and drug delivery. Studies suggest that MSC-EVs, the central paracrine contributors of MSCs, replicate the therapeutic effects of MSCs. This review explores the characteristics of MSC-EVs, emphasizing their potential in therapeutics and drug delivery for various diseases, including CRISPR/Cas9 delivery for gene editing. It also delves into the obstacles and challenges of MSC-EVs in clinical applications and provides insights into strategies to overcome the limitations of biodistribution and target delivery.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 5","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-05-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.156","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140952716","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":"Comparison of cervicovaginal fluid extracellular vesicles isolated from paired cervical brushes and vaginal swabs","authors":"Emily Sarah Jane Paterson,&nbsp;Simon Scheck,&nbsp;Simon McDowell,&nbsp;Nick Bedford,&nbsp;Jane Eleanor Girling,&nbsp;Claire Elizabeth Henry","doi":"10.1002/jex2.153","DOIUrl":"https://doi.org/10.1002/jex2.153","url":null,"abstract":"<p>Endometriosis is a common gynaecological condition, with a long diagnostic delay. Surgery is required to confirm a diagnosis, highlighting the need for a non-invasive biomarker. Extracellular vesicles (EVs) may have a role in endometriosis pathogenesis, yet there is limited EV biomarker literature available. This study aimed to investigate the feasibility of isolating cervico-vaginal fluid EVs sampled using cervical brushes and vaginal swabs and to compare these methods. After providing informed consent, patients undergoing surgery for suspected endometriosis had cervical brush and vaginal swab samples collected under general anaesthetic. Isolated EVs were characterised through negative stain transmission electron microscopy (TEM), Western blotting (TSG101, CD63, Calnexin, ApoB, Albumin), tunable resistive pulse sensing (TRPS), microBCA assays and RT-qPCR of miRNAs. PCR was performed on samples prior to EV isolation to assess bacteria present in samples. Cervical brush and vaginal swab EVs were intact vesicles with limited co-isolated contaminants. Cervical brushes had higher concentrations of particles compared to match vaginal swabs, although both samples had low concentrations. Protein and miRNA yield were similar between matched samples. PCR demonstrated only a small amount DNA within samples was bacterial (&gt;0.5%). Cervico-vaginal fluids EVs were successfully isolated from cervical brushes and vaginal swabs, demonstrating a new method of sampling reproductive EVs. EV yield from both sample types was low. Similar protein and miRNA levels suggest either sampling method may be suitable for biomarker studies.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 5","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.153","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140820720","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":"Food-derived extracellular vesicles in the human gastrointestinal tract: Opportunities for personalised nutrition and targeted therapeutics","authors":"Natalie P. Turner","doi":"10.1002/jex2.154","DOIUrl":"https://doi.org/10.1002/jex2.154","url":null,"abstract":"<p>Food-derived extracellular vesicles (FDEVs) such as those found in mammalian milk and plants are of great interest for both their health benefits and ability to act as biological nanocarriers. While the extracellular vesicle (EV) field is expanding rapidly to perform characterisation studies on FDEVs from plants, yeasts and bacteria, species-specific differences in EV uptake and function in the human gastrointestinal (GI) tract are poorly understood. Moreover, the effects of food processing on the EV surfaceome and intraluminal content also raises questions surrounding biological viability once consumed. Here, I present a case for increasing community-wide focus on understanding the cellular uptake of FDEVs from different animal, plant, yeast, and bacterial species and how this may impact their function in the human, which will have implications for human health and therapeutic strategies alike.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 5","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.154","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140820718","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":"Microvesicles derived from dermal myofibroblasts modify the integrity of the blood and lymphatic barriers using distinct endocytosis pathways","authors":"Syrine Arif,&nbsp;Megan Richer,&nbsp;Sébastien Larochelle,&nbsp;Véronique J. Moulin","doi":"10.1002/jex2.151","DOIUrl":"https://doi.org/10.1002/jex2.151","url":null,"abstract":"<p>Microvesicles (MVs) are a subtype of extracellular vesicles that can transfer biological information from their producer cells to target cells. This communication can in turn affect both normal and pathological processes. Mounting evidence has revealed that dermal wound myofibroblasts (Wmyo) produce MVs, which can transfer biomolecules impacting receptor cells such as human dermal microvascular endothelial cells (HDMECs). While the effects of MVs on HDMECs are generally well described in the literature, little is known about the transport of MVs across the HDMEC barrier, and their potential effect on the barrier integrity remains unknown. Here, we investigated these roles of Wmyo-derived MVs on two sub-populations of HDMECs, blood endothelial cells (BECs) and lymphatic endothelial cells (LECs). Using an in vitro model to mimic the endothelial barrier, we showed that MVs crossed the LEC barrier but not the BEC barrier. In addition, we demonstrated that MVs were able to influence the cell-cell junctions of HDMECs. Specifically, we observed that after internalization via the predominantly caveolin-dependent pathway, MVs induced the opening of junctions in BECs. Conversely, in LECs, MVs mainly use the macropinocytosis pathway and induce closure of these junctions. Moreover, proteins in the MV membrane were responsible for this effect, but not specifically those belonging to the VEGF family. Finally, we found that once the LEC barrier permeability was reduced by MV stimuli, MVs ceased to cross the barrier. Conversely, when the BEC barrier was rendered permeable following stimulation with MVs, they were subsequently able to cross the barrier via the paracellular pathway. Taken together, these results suggest that the study of Wmyo-derived MVs offers valuable insights into their interaction with the HDMEC barrier in the context of wound healing. They highlight the potential significance of these MVs in the overall process.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 5","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-05-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.151","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140820719","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":"Comparative analysis of the effects of different purification methods on the yield and purity of cow milk extracellular vesicles","authors":"Santeri Kankaanpää,&nbsp;Markus Nurmi,&nbsp;Markus Lampimäki,&nbsp;Heidi Leskinen,&nbsp;Anni Nieminen,&nbsp;Anatoliy Samoylenko,&nbsp;Seppo J. Vainio,&nbsp;Sari Mäkinen,&nbsp;Lauri Ahonen,&nbsp;Juha Kangasluoma,&nbsp;Tuukka Petäjä,&nbsp;Sirja Viitala","doi":"10.1002/jex2.149","DOIUrl":"https://doi.org/10.1002/jex2.149","url":null,"abstract":"<p>Isolation of extracellular vesicles (EV) has been developing rapidly in parallel with the interest in EVs. However, commonly utilized protocols may not suit more challenging sample matrixes and could potentially yield suboptimal results. Knowing and assessing the pitfalls of isolation procedure to be used, should be involved to some extent for EV analytics. EVs in cow milk are of great interest due to their abundancy and large-scale availability as well as their cross-species bioavailability and possible use as drug carriers. However, the characteristics of milk EVs overlap with those of other milk components. This makes it difficult to isolate and study EVs individually. There exists also a lack of consensus for isolation methods. In this study, we demonstrated the differences between various differential centrifugation-based approaches for isolation of large quantities of EVs from cow milk. Samples were further purified with gradient centrifugation and size exclusion chromatography (SEC) and differences were analyzed. Quality measurements were conducted on multiple independent platforms. Particle analysis, electron microscopy and RNA analysis were used, to comprehensively characterize the isolated samples and to identify the limitations and possible sources of contamination in the EV isolation protocols. Vesicle concentration to protein ratio and RNA to protein ratios were observed to increase as samples were purified, suggesting co-isolation with major milk proteins in direct differential centrifugation protocols. We demonstrated a novel size assessment of vesicles using a particle mobility analyzer that matched the sizing using electron microscopy in contrast to commonly utilized nanoparticle tracking analysis. Based on the standards of the International Society for Extracellular Vesicles and the quick checklist of EV-Track.org for EV isolation, we emphasize the need for complete characterization and validation of the isolation protocol with all EV-related work to ensure the accuracy of results and allow further analytics and experiments.</p>","PeriodicalId":73747,"journal":{"name":"Journal of extracellular biology","volume":"3 4","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/jex2.149","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140632004","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}