Pub Date : 2022-01-01Epub Date: 2022-01-21DOI: 10.20517/evcna.2021.22
Zhou Zhang, Chang Zeng, Wei Zhang
Previous studies have suggested that aberrant 5-hydroxymethylcytosines (5hmC) modifications are related to cancer pathobiology. Genome-wide profiling 5hmC in circulating cell-free DNA (cfDNA) using the highly sensitive chemical labeling-based 5hmC-Seal technique has been demonstrated to have the potential to be a robust epigenomic tool for cancer biomarker discovery. Prior studies have mostly focused on cfDNA-derived 5hmC-Seal data summarized in well-annotated genic features (e.g., gene bodies) or unbiased bins. Zhou et al. recently proposed long non-coding RNAs (lncRNAs) as an alternative molecular target for biomarker discovery using publicly available 5hmC-Seal data. Considering its potential clinical impact, we would like to comment on Zhou et al. and advocate more serious consideration of critical issues such as the availability of clinical information and technical variables, especially when performing secondary analysis using publicly available data, with the aim of improving data transparency and translatability.
{"title":"Considerations before the application of 5-hydroxymethylation levels of long non-coding RNAs for non-invasive cancer diagnosis.","authors":"Zhou Zhang, Chang Zeng, Wei Zhang","doi":"10.20517/evcna.2021.22","DOIUrl":"https://doi.org/10.20517/evcna.2021.22","url":null,"abstract":"<p><p>Previous studies have suggested that aberrant 5-hydroxymethylcytosines (5hmC) modifications are related to cancer pathobiology. Genome-wide profiling 5hmC in circulating cell-free DNA (cfDNA) using the highly sensitive chemical labeling-based 5hmC-Seal technique has been demonstrated to have the potential to be a robust epigenomic tool for cancer biomarker discovery. Prior studies have mostly focused on cfDNA-derived 5hmC-Seal data summarized in well-annotated genic features (e.g., gene bodies) or unbiased bins. Zhou et al. recently proposed long non-coding RNAs (lncRNAs) as an alternative molecular target for biomarker discovery using publicly available 5hmC-Seal data. Considering its potential clinical impact, we would like to comment on Zhou <i>et al</i>. and advocate more serious consideration of critical issues such as the availability of clinical information and technical variables, especially when performing secondary analysis using publicly available data, with the aim of improving data transparency and translatability.</p>","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"3 1","pages":"10-13"},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8797161/pdf/nihms-1771817.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"39873577","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}
Extracellular vesicles (EVs) have been regarded as influential intracellular delivering parcels that possess tremendous potential because of their strict and complex secretion regulation processes. However, traditional detection methods cannot monitor the secretion of EVs due to their small particle diameters. Inspired by their peculiar diverse appearances and lipid membranes ingredients, we developed an innovative strategy to detect EVs in any kind of fluids by using rationally designed peptide probes that particularly recognize the highly curved surface of EVs. These peptide probes also serve as novel tools to selectively target cancerous cells with specific lipid compositions and distributions. With this strategy, we discovered a series of EV-secreting regulation mechanisms and identified their roles within physiological processes. Recently, we found that the transportation of oligodeoxynucleotides and cell division control protein 42 homolog from TLR9-activated macrophages to naïve cells via EVs exerts synergetic effects in the propagation of the intracellular immune response, which suggests a general mechanism for EV-mediated uptake of pathogen-associated molecular patterns.
{"title":"Summary of Prof. Yin’s CSEMV-EVCNA award lecture 2021","authors":"Ying Zhang, Hang Yin","doi":"10.20517/evcna.2022.16","DOIUrl":"https://doi.org/10.20517/evcna.2022.16","url":null,"abstract":"Extracellular vesicles (EVs) have been regarded as influential intracellular delivering parcels that possess tremendous potential because of their strict and complex secretion regulation processes. However, traditional detection methods cannot monitor the secretion of EVs due to their small particle diameters. Inspired by their peculiar diverse appearances and lipid membranes ingredients, we developed an innovative strategy to detect EVs in any kind of fluids by using rationally designed peptide probes that particularly recognize the highly curved surface of EVs. These peptide probes also serve as novel tools to selectively target cancerous cells with specific lipid compositions and distributions. With this strategy, we discovered a series of EV-secreting regulation mechanisms and identified their roles within physiological processes. Recently, we found that the transportation of oligodeoxynucleotides and cell division control protein 42 homolog from TLR9-activated macrophages to naïve cells via EVs exerts synergetic effects in the propagation of the intracellular immune response, which suggests a general mechanism for EV-mediated uptake of pathogen-associated molecular patterns.","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"55 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"84490349","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
S. A. U. Shinge, Yin Xiao, Jiang Xia, Yujie Liang, Lijuan Duan
Plant exosome-like nanovesicles (PELNVs) are membrane-encapsulated nanostructures released from cells into their surroundings. PELNVs have an important role in intercellular and interspecies communication in all three domains of life. They act as protective compartments for the long-distance transit of signal molecules like proteins, nucleic acids, lipids, and other metabolites. A range of plants and vegetables can emit PELNVs. The importance of PELNVs in interspecies communication stems from their concentration in biomolecules (lipids, proteins, and miRNAs), lack of toxicity, ease of internalization by cells, and anti-inflammatory, immune-modulatory, and regenerative characteristics. PELNVs derived from numerous fruits and vegetables are biocompatible, biodegradable, and abundant in various plant species. Moreover, their convincing physicochemical characteristics underpin their modulative role in physiological and pathological processes, all of which have fueled speculation that these nanovesicles could be particularly adept at developing future-generation bio-therapeutic platforms. The goal of this review was not only to present an overview of the identified roles of PELNVs in physiology and pathology, but also to provide new insight toward their engineering for effective therapeutics and drug delivery nanoplatforms, a clue for future direction to the ongoing research gaps.
{"title":"New insights of engineering plant exosome-like nanovesicles as a nanoplatform for therapeutics and drug delivery","authors":"S. A. U. Shinge, Yin Xiao, Jiang Xia, Yujie Liang, Lijuan Duan","doi":"10.20517/evcna.2021.25","DOIUrl":"https://doi.org/10.20517/evcna.2021.25","url":null,"abstract":"Plant exosome-like nanovesicles (PELNVs) are membrane-encapsulated nanostructures released from cells into their surroundings. PELNVs have an important role in intercellular and interspecies communication in all three domains of life. They act as protective compartments for the long-distance transit of signal molecules like proteins, nucleic acids, lipids, and other metabolites. A range of plants and vegetables can emit PELNVs. The importance of PELNVs in interspecies communication stems from their concentration in biomolecules (lipids, proteins, and miRNAs), lack of toxicity, ease of internalization by cells, and anti-inflammatory, immune-modulatory, and regenerative characteristics. PELNVs derived from numerous fruits and vegetables are biocompatible, biodegradable, and abundant in various plant species. Moreover, their convincing physicochemical characteristics underpin their modulative role in physiological and pathological processes, all of which have fueled speculation that these nanovesicles could be particularly adept at developing future-generation bio-therapeutic platforms. The goal of this review was not only to present an overview of the identified roles of PELNVs in physiology and pathology, but also to provide new insight toward their engineering for effective therapeutics and drug delivery nanoplatforms, a clue for future direction to the ongoing research gaps.","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"75 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"83405570","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Extracellular vesicles (EVs), which are nanocarriers with phospholipid bilayer structures released by most cells, play a key role in regulating physiological and pathological processes. EVs have been investigated due to their loading capacity, low toxicity, immunogenicity, and biofunctions. Although EVs have shown good potential as therapeutic vehicles, natural EVs have a poor targeting ability, which substantially reduces the therapeutic effect. Through the addition of a targeting unit into the membrane surface of EVs or inside EVs by engineering technology, the therapeutic agent can accumulate in specific cells and tissues. Here, we focus on mammalian EVs (MEVs) and bacterial EVs (BEVs), which are the two most common types of EVs in the biomedical field. In this review, we describe engineered MEVs and BEVs as promising nanocarriers for targeted therapy and summarize the biogenesis, isolation, and characterization of MEVs and BEVs. We then describe engineering techniques for enhancement of the targeting ability of EVs. Moreover, we focus on the applications of engineered MEVs and BEVs in targeted therapy, including the treatment of cancer and brain and bone disease. We believe that this review will help improve the understanding of engineered MEVs and BEVs, thereby promoting their application and clinical translation.
{"title":"Engineered mammalian and bacterial extracellular vesicles as promising nanocarriers for targeted therapy","authors":"Han Liu, Zhen Geng, Jiacan Su","doi":"10.20517/evcna.2022.04","DOIUrl":"https://doi.org/10.20517/evcna.2022.04","url":null,"abstract":"Extracellular vesicles (EVs), which are nanocarriers with phospholipid bilayer structures released by most cells, play a key role in regulating physiological and pathological processes. EVs have been investigated due to their loading capacity, low toxicity, immunogenicity, and biofunctions. Although EVs have shown good potential as therapeutic vehicles, natural EVs have a poor targeting ability, which substantially reduces the therapeutic effect. Through the addition of a targeting unit into the membrane surface of EVs or inside EVs by engineering technology, the therapeutic agent can accumulate in specific cells and tissues. Here, we focus on mammalian EVs (MEVs) and bacterial EVs (BEVs), which are the two most common types of EVs in the biomedical field. In this review, we describe engineered MEVs and BEVs as promising nanocarriers for targeted therapy and summarize the biogenesis, isolation, and characterization of MEVs and BEVs. We then describe engineering techniques for enhancement of the targeting ability of EVs. Moreover, we focus on the applications of engineered MEVs and BEVs in targeted therapy, including the treatment of cancer and brain and bone disease. We believe that this review will help improve the understanding of engineered MEVs and BEVs, thereby promoting their application and clinical translation.","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"43 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"88062365","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Obituary for Prof. Dr. Johng Sik Rhim","authors":"","doi":"10.20517/evcna.2022.15","DOIUrl":"https://doi.org/10.20517/evcna.2022.15","url":null,"abstract":"","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"8 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"88910623","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Bing Sun, Scott Kitchen, Norina Tang, Andreas Garza, Sheela Jacob, Lynn Pulliam
Aim: A peripheral inflammatory response can drive neuroinflammation in a number of infections including human immunodeficiency virus (HIV). Monocyte/macrophage (M/Mφ) activation is a hallmark of acute HIV infection and a source of chronic inflammation in a subset of HIV-infected individuals. We sought to decrease peripheral inflammation and M/Mφ transmigration after HIV infection by engineering extracellular vesicles (EV) to antagonize a microRNA (miR) associated with inflammation. We hypothesized that induced pluripotent stem cell (iPSC)-derived monocyte EVs (mEVs), engineered to contain an antagomir to miR-155 (αmiR mEV) would target monocyte inflammation and influence neuroinflammation in an HIV-infected humanized mice.
Methods: mEVs were characterized by tetraspanins, nanoparticle tracking analysis, electron microscopy, and their preferential entry into circulating monocytes as well as testing for endogenous selected miRNAs. HIV-infected humanized mice were treated with control or antagomir155 mEVs. Plasma viral load was measured plus activation markers on lymphocytes and monocytes and the number of macrophages in the brain was quantified.
Results: mEVs preferentially entered peripheral monocytes. HIV infection increased C-C chemokine receptor type 5 (CCR5) and major histocompatibility complex, class II, DR (HLA-DR) expression on T cells and monocytes. Treatments with mEVs did not decrease plasma HIV viral load; however, mEVs alone resulted in a decrease in %CCR5+ and %HLA-DR+ on T cells and an increase in %CCR5+ monocytes. αmiR mEVs decreased %CCR5 on M/Mφ. The mEV-treated HIV-infected mice did not show an increase in macrophage transmigration to the brain.
Conclusion: mEVs alone caused an unexpected decrease in lymphocyte activation and increase in monocyte %CCR5; however, this did not translate to an increase in macrophage transmigration to the brain.
{"title":"Engineered induced-pluripotent stem cell derived monocyte extracellular vesicles alter inflammation in HIV humanized mice.","authors":"Bing Sun, Scott Kitchen, Norina Tang, Andreas Garza, Sheela Jacob, Lynn Pulliam","doi":"10.20517/evcna.2022.11","DOIUrl":"https://doi.org/10.20517/evcna.2022.11","url":null,"abstract":"<p><strong>Aim: </strong>A peripheral inflammatory response can drive neuroinflammation in a number of infections including human immunodeficiency virus (HIV). Monocyte/macrophage (M/Mφ) activation is a hallmark of acute HIV infection and a source of chronic inflammation in a subset of HIV-infected individuals. We sought to decrease peripheral inflammation and M/Mφ transmigration after HIV infection by engineering extracellular vesicles (EV) to antagonize a microRNA (miR) associated with inflammation. We hypothesized that induced pluripotent stem cell (iPSC)-derived monocyte EVs (mEVs), engineered to contain an antagomir to miR-155 (αmiR mEV) would target monocyte inflammation and influence neuroinflammation in an HIV-infected humanized mice.</p><p><strong>Methods: </strong>mEVs were characterized by tetraspanins, nanoparticle tracking analysis, electron microscopy, and their preferential entry into circulating monocytes as well as testing for endogenous selected miRNAs. HIV-infected humanized mice were treated with control or antagomir155 mEVs. Plasma viral load was measured plus activation markers on lymphocytes and monocytes and the number of macrophages in the brain was quantified.</p><p><strong>Results: </strong>mEVs preferentially entered peripheral monocytes. HIV infection increased C-C chemokine receptor type 5 (CCR5) and major histocompatibility complex, class II, DR (HLA-DR) expression on T cells and monocytes. Treatments with mEVs did not decrease plasma HIV viral load; however, mEVs alone resulted in a decrease in %CCR5+ and %HLA-DR+ on T cells and an increase in %CCR5+ monocytes. αmiR mEVs decreased %CCR5 on M/Mφ. The mEV-treated HIV-infected mice did not show an increase in macrophage transmigration to the brain.</p><p><strong>Conclusion: </strong>mEVs alone caused an unexpected decrease in lymphocyte activation and increase in monocyte %CCR5; however, this did not translate to an increase in macrophage transmigration to the brain.</p>","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"3 2","pages":"118-132"},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10104589/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9324444","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}
Chunping Liu, Dongyue He, Huan Cen, Huiqi Chen, Longmei Li, G. Nie, Zixue Zhong, Qingfeng He, Xiaofei Yang, Sien Guo, Lei Wang, Zhijin Fan
Extracellular vesicles (EVs), as natural carriers, are regarded as a new star in nanomedicine due to their excellent biocompatibility, fascinating physicochemical properties, and unique biological regulatory functions. However, there are still some challenges to using natural EVs, including poor targeting ability and the clearance from circulation, which may limit their further development and clinical use. Nucleic acid has the functions of programmability, targeting, gene therapy, and immune regulation. Owing to the engineering design and modification by integrating functional nucleic acid, EVs offer excellent performances as a therapeutic system in vivo. This review briefly introduces the function and mechanism of nucleic acid in the diagnosis and treatment of diseases. Then, the strategies of nucleic acid-functionalized EVs are summarized and the latest progress of nucleic acid-functionalized EVs in nanomedicine is highlighted. Finally, the challenges and prospects of nucleic acid-functionalized EVs as a promising diagnostic system are proposed.
{"title":"Nucleic acid functionalized extracellular vesicles as promising therapeutic systems for nanomedicine","authors":"Chunping Liu, Dongyue He, Huan Cen, Huiqi Chen, Longmei Li, G. Nie, Zixue Zhong, Qingfeng He, Xiaofei Yang, Sien Guo, Lei Wang, Zhijin Fan","doi":"10.20517/evcna.2021.21","DOIUrl":"https://doi.org/10.20517/evcna.2021.21","url":null,"abstract":"Extracellular vesicles (EVs), as natural carriers, are regarded as a new star in nanomedicine due to their excellent biocompatibility, fascinating physicochemical properties, and unique biological regulatory functions. However, there are still some challenges to using natural EVs, including poor targeting ability and the clearance from circulation, which may limit their further development and clinical use. Nucleic acid has the functions of programmability, targeting, gene therapy, and immune regulation. Owing to the engineering design and modification by integrating functional nucleic acid, EVs offer excellent performances as a therapeutic system in vivo. This review briefly introduces the function and mechanism of nucleic acid in the diagnosis and treatment of diseases. Then, the strategies of nucleic acid-functionalized EVs are summarized and the latest progress of nucleic acid-functionalized EVs in nanomedicine is highlighted. Finally, the challenges and prospects of nucleic acid-functionalized EVs as a promising diagnostic system are proposed.","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"12 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"78397901","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
an excellent and well-presented overview of the complexity of extracellular vesicles. the second lecture. The speaker addressed basic questions regarding the role of tumor-released EVs in mediating the pro-metastatic effects of chemotherapy. He showed advanced and elegant research in his laboratory, providing evidence that two classes of cytotoxic drugs are broadly employed in preoperative (neoadjuvant) breast cancer therapy, elicit tumor-derived EVs with enhanced pro-metastatic capacity. Mechanistically he showed that chemotherapy-elicited EVs are enriched in annexin A6 (ANXA6), a Ca 2+ -dependent protein that promotes NF- κ B-dependent endothelial cell activation, Ccl2 induction, and Ly6C+CCR2+ monocyte expansion in the pulmonary pre-metastatic niche to facilitate the establishment of lung metastasis [3] . Genetic inactivation of Anxa6 in cancer cells or Ccr2 in host cells blunts the pro-metastatic effects of chemotherapy-elicited EVs. ANXA6 is detected and potentially enriched, in the circulating EVs of breast cancer patients undergoing neoadjuvant chemotherapy. the application of of the
{"title":"Meeting report of the 2nd Lugano ExoDay: extracellular vesicles as next-generation clinical biomarkers and therapeutic agents","authors":"C. Balbi, M. Cretich, L. Barile","doi":"10.20517/evcna.2022.17","DOIUrl":"https://doi.org/10.20517/evcna.2022.17","url":null,"abstract":"an excellent and well-presented overview of the complexity of extracellular vesicles. the second lecture. The speaker addressed basic questions regarding the role of tumor-released EVs in mediating the pro-metastatic effects of chemotherapy. He showed advanced and elegant research in his laboratory, providing evidence that two classes of cytotoxic drugs are broadly employed in preoperative (neoadjuvant) breast cancer therapy, elicit tumor-derived EVs with enhanced pro-metastatic capacity. Mechanistically he showed that chemotherapy-elicited EVs are enriched in annexin A6 (ANXA6), a Ca 2+ -dependent protein that promotes NF- κ B-dependent endothelial cell activation, Ccl2 induction, and Ly6C+CCR2+ monocyte expansion in the pulmonary pre-metastatic niche to facilitate the establishment of lung metastasis [3] . Genetic inactivation of Anxa6 in cancer cells or Ccr2 in host cells blunts the pro-metastatic effects of chemotherapy-elicited EVs. ANXA6 is detected and potentially enriched, in the circulating EVs of breast cancer patients undergoing neoadjuvant chemotherapy. the application of of the","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"25 1","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"75023253","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
S. Staubach, T. Tertel, Bernd Walkenfort, Dominik Buschmann, M. Pfaffl, G. Weber, B. Giebel
Aim: Despite intensive research during the last decade, it remains challenging to prepare extracellular vesicles (EVs) of high purity, especially from primary body liquids or protein-rich conditioned media. For now, time-consuming combinations of at least two orthogonal methods, e.g., density and size separation, are required to enrich EVs to high purity, often at the expense of processing time. Therefore, novel technologies are required that allow EV preparation in acceptable time intervals and to fair purities. Free-flow electrophoresis (FFE) constitutes a well-established semi-preparative method to separate and prepare analytes, e.g., by inherent differences in their electric charges. FFE combines a flow-driven longitudinal transport of sample material with vertical electrophoresis and allows the separation of sample components into up to 96 different fractions. It was our aim to evaluate the potential of FFE for the separation of EVs from other sample components of EV-containing protein-rich conditioned cell culture media. Methods: Exemplarily, conditioned media of mesenchymal stem/stromal cells raised in the presence of EV-containing 10% human platelet lysate were processed. We analyzed the obtained fractions by different technologies, including imaging flow cytometry, western blot and nanoparticle tracking analysis. Results: We demonstrate that FFE quickly and reproducibly separates EVs from a huge proportion of molecules included in the original sample. Conclusion: Our results qualify FFE as a feasible, quick and reproducible technology for the preparation of bona fide EVs.
{"title":"Free flow electrophoresis allows quick and reproducible preparation of extracellular vesicles from conditioned cell culture media","authors":"S. Staubach, T. Tertel, Bernd Walkenfort, Dominik Buschmann, M. Pfaffl, G. Weber, B. Giebel","doi":"10.20517/evcna.2021.26","DOIUrl":"https://doi.org/10.20517/evcna.2021.26","url":null,"abstract":"Aim: Despite intensive research during the last decade, it remains challenging to prepare extracellular vesicles (EVs) of high purity, especially from primary body liquids or protein-rich conditioned media. For now, time-consuming combinations of at least two orthogonal methods, e.g., density and size separation, are required to enrich EVs to high purity, often at the expense of processing time. Therefore, novel technologies are required that allow EV preparation in acceptable time intervals and to fair purities. Free-flow electrophoresis (FFE) constitutes a well-established semi-preparative method to separate and prepare analytes, e.g., by inherent differences in their electric charges. FFE combines a flow-driven longitudinal transport of sample material with vertical electrophoresis and allows the separation of sample components into up to 96 different fractions. It was our aim to evaluate the potential of FFE for the separation of EVs from other sample components of EV-containing protein-rich conditioned cell culture media. Methods: Exemplarily, conditioned media of mesenchymal stem/stromal cells raised in the presence of EV-containing 10% human platelet lysate were processed. We analyzed the obtained fractions by different technologies, including imaging flow cytometry, western blot and nanoparticle tracking analysis. Results: We demonstrate that FFE quickly and reproducibly separates EVs from a huge proportion of molecules included in the original sample. Conclusion: Our results qualify FFE as a feasible, quick and reproducible technology for the preparation of bona fide EVs.","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":" 8","pages":""},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"91414110","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Aim: Targeting the modes of pathogen shedding/transmission via exosomes or extracellular vesicles has been envisioned as the best approach to control vector-borne diseases. This study is focused on altering exosomes stability to affect the pathogen transmission from infected to naïve recipient cells.
Methods: In this study, neuronal or arthropod exosomes were treated at different temperatures or with different salts or pH conditions to analyze their ability and efficiency in the transmission of tick-borne Langat virus (LGTV) from infected to naïve recipient cells.
Results: Quantitative real-time PCR (qRT-PCR) and immunoblotting analyses revealed that treatment of neuronal or tick exosomes at warmer temperatures of 37 °C or 23 °C, respectively, or with sulfate salts such as Magnesium or Ammonium sulfates or with highly alkaline pH of 9 or 11.5, dramatically reduced transmission of LGTV via infectious exosomes (human or tick cells-derived) to human neuronal (SH-SY5Y) cells or skin keratinocytes (HaCaT cells), respectively.
Conclusion: Overall, this study suggests that exosome-mediated viral transmission of vector-borne pathogens to the vertebrate host or the viral dissemination and replication within or between the mammalian host can be reduced by altering the ability of exosomes with basic changes in temperatures, salts or pH conditions.
{"title":"Alterations in arthropod and neuronal exosomes reduce virus transmission and replication in recipient cells.","authors":"Kehinde Damilare Fasae, Girish Neelakanta, Hameeda Sultana","doi":"10.20517/evcna.2022.30","DOIUrl":"https://doi.org/10.20517/evcna.2022.30","url":null,"abstract":"<p><strong>Aim: </strong>Targeting the modes of pathogen shedding/transmission via exosomes or extracellular vesicles has been envisioned as the best approach to control vector-borne diseases. This study is focused on altering exosomes stability to affect the pathogen transmission from infected to naïve recipient cells.</p><p><strong>Methods: </strong>In this study, neuronal or arthropod exosomes were treated at different temperatures or with different salts or pH conditions to analyze their ability and efficiency in the transmission of tick-borne Langat virus (LGTV) from infected to naïve recipient cells.</p><p><strong>Results: </strong>Quantitative real-time PCR (qRT-PCR) and immunoblotting analyses revealed that treatment of neuronal or tick exosomes at warmer temperatures of 37 °C or 23 °C, respectively, or with sulfate salts such as Magnesium or Ammonium sulfates or with highly alkaline pH of 9 or 11.5, dramatically reduced transmission of LGTV via infectious exosomes (human or tick cells-derived) to human neuronal (SH-SY5Y) cells or skin keratinocytes (HaCaT cells), respectively.</p><p><strong>Conclusion: </strong>Overall, this study suggests that exosome-mediated viral transmission of vector-borne pathogens to the vertebrate host or the viral dissemination and replication within or between the mammalian host can be reduced by altering the ability of exosomes with basic changes in temperatures, salts or pH conditions.</p>","PeriodicalId":73008,"journal":{"name":"Extracellular vesicles and circulating nucleic acids","volume":"3 3","pages":"247-279"},"PeriodicalIF":0.0,"publicationDate":"2022-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10018778/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9153814","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}
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