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Sample records for cells trojan exosomes

  1. HIV and mature dendritic cells: Trojan exosomes riding the Trojan horse?

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    Nuria Izquierdo-Useros

    2010-03-01

    Full Text Available Exosomes are secreted cellular vesicles that can induce specific CD4(+ T cell responses in vivo when they interact with competent antigen-presenting cells like mature dendritic cells (mDCs. The Trojan exosome hypothesis proposes that retroviruses can take advantage of the cell-encoded intercellular vesicle traffic and exosome exchange pathway, moving between cells in the absence of fusion events in search of adequate target cells. Here, we discuss recent data supporting this hypothesis, which further explains how DCs can capture and internalize retroviruses like HIV-1 in the absence of fusion events, leading to the productive infection of interacting CD4(+ T cells and contributing to viral spread through a mechanism known as trans-infection. We suggest that HIV-1 can exploit an exosome antigen-dissemination pathway intrinsic to mDCs, allowing viral internalization and final trans-infection of CD4(+ T cells. In contrast to previous reports that focus on the ability of immature DCs to capture HIV in the mucosa, this review emphasizes the outstanding role that mature DCs could have promoting trans-infection in the lymph node, underscoring a new potential viral dissemination pathway.

  2. Mesenchymal stem cell exosomes.

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    Lai, Ruenn Chai; Yeo, Ronne Wee Yeh; Lim, Sai Kiang

    2015-04-01

    MSCs are an extensively used cell type in clinical trials today. The initial rationale for their clinical testing was based on their differentiation potential. However, the lack of correlation between functional improvement and cell engraftment or differentiation at the site of injury has led to the proposal that MSCs exert their effects not through their differentiation potential but through their secreted product, more specifically, exosomes, a type of extracellular vesicle. We propose here that MSC exosomes function as an extension of MSC's biological role as tissue stromal support cells. Like their cell source, MSC exosomes help maintain tissue homeostasis for optimal tissue function. They target housekeeping biological processes that operate ubiquitously in all tissues and are critical in maintaining tissue homeostasis, enabling cells to recover critical cellular functions and begin repair and regeneration. This hypothesis provides a rationale for the therapeutic efficacy of MSCs and their secreted exosomes in a wide spectrum of diseases. Here, we give a brief introduction of the biogenesis of MSC exosomes, review their physiological functions and highlight some of their biochemical potential to illustrate how MSC exosomes could restore tissue homeostasis leading to tissue recovery and repair.

  3. Fibronectin on the Surface of Myeloma Cell-derived Exosomes Mediates Exosome-Cell Interactions.

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    Purushothaman, Anurag; Bandari, Shyam Kumar; Liu, Jian; Mobley, James A; Brown, Elizabeth E; Sanderson, Ralph D

    2016-01-22

    Exosomes regulate cell behavior by binding to and delivering their cargo to target cells; however, the mechanisms mediating exosome-cell interactions are poorly understood. Heparan sulfates on target cell surfaces can act as receptors for exosome uptake, but the ligand for heparan sulfate on exosomes has not been identified. Using exosomes isolated from myeloma cell lines and from myeloma patients, we identify exosomal fibronectin as a key heparan sulfate-binding ligand and mediator of exosome-cell interactions. We discovered that heparan sulfate plays a dual role in exosome-cell interaction; heparan sulfate on exosomes captures fibronectin, and on target cells it acts as a receptor for fibronectin. Removal of heparan sulfate from the exosome surface releases fibronectin and dramatically inhibits exosome-target cell interaction. Antibody specific for the Hep-II heparin-binding domain of fibronectin blocks exosome interaction with tumor cells or with marrow stromal cells. Regarding exosome function, fibronectin-mediated binding of exosomes to myeloma cells activated p38 and pERK signaling and expression of downstream target genes DKK1 and MMP-9, two molecules that promote myeloma progression. Antibody against fibronectin inhibited the ability of myeloma-derived exosomes to stimulate endothelial cell invasion. Heparin or heparin mimetics including Roneparstat, a modified heparin in phase I trials in myeloma patients, significantly inhibited exosome-cell interactions. These studies provide the first evidence that fibronectin binding to heparan sulfate mediates exosome-cell interactions, revealing a fundamental mechanism important for exosome-mediated cross-talk within tumor microenvironments. Moreover, these results imply that therapeutic disruption of fibronectin-heparan sulfate interactions will negatively impact myeloma tumor growth and progression.

  4. CD45 immunoaffinity depletion of vesicles from Jurkat T cells demonstrates that exosomes contain CD45: no evidence for a distinct exosome/HIV-1 budding pathway

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    Ott David E

    2008-07-01

    Full Text Available Abstract The presence of relatively high levels of cellular protein contamination in density-purified virion preparations is a confounding factor in biochemical analyses of HIV and SIV produced from hematopoietic cells. A major source of this contamination is from vesicles, either microvesicles or exosomes, that have similar physical properties as virions. Thus, these particles can not be removed by size or density fractionation. Although virions and vesicles have similar cellular protein compositions, CD45 is excluded from HIV-1 yet is present in vesicles produced from hematopoietic cells. By exploiting this finding, we have developed a CD45 immunoaffinity depletion procedure that removes vesicles from HIV-1 preparations. While this approach has been successfully applied to virion preparations from several different cell types, some groups have concluded that "exosomes" from certain T cell lines, specifically Jurkat, do not contain CD45. If this interpretation is correct, then these vesicles could not be removed by CD45 immunoaffinity depletion. Here we show that dense vesicles produced by Jurkat and SupT1/CCR5 cells contain CD45 and are efficiently removed from preparations by CD45-immunoaffinity depletion. Also, contaminating cellular proteins were removed from virion preparations produced by these lines. Previously, the absence of CD45 from both "exosomes" and virions has been used to support the so called Trojan exosome hypothesis, namely that HIV-1 is simply an exosome containing viral material. The presence of CD45 on vesicles, including exosomes, and its absence on virions argues against a specialized budding pathway that is shared by both exosomes and HIV-1.

  5. DMPD: Lipoprotein trafficking in vascular cells. Molecular Trojan horses and cellularsaboteurs. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 9287290 Lipoprotein trafficking in vascular cells. Molecular Trojan horses and cell...9287290 Title Lipoprotein trafficking in vascular cells. Molecular Trojan horses ...ularsaboteurs. Hajjar DP, Haberland ME. J Biol Chem. 1997 Sep 12;272(37):22975-8. (.png) (.svg) (.html) (.csml) Show Lipoprotein traf...ficking in vascular cells. Molecular Trojan horses and cellularsaboteurs. PubmedID

  6. Delivery of Small Interfering RNAs to Cells via Exosomes.

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    Wahlgren, Jessica; Statello, Luisa; Skogberg, Gabriel; Telemo, Esbjörn; Valadi, Hadi

    2016-01-01

    Exosomes are small membrane bound vesicles between 30 and 100 nm in diameter of endocytic origin that are secreted into the extracellular environment by many different cell types. Exosomes play a role in intercellular communication by transferring proteins, lipids, and RNAs to recipient cells.Exosomes from human cells could be used as vectors to provide cells with therapeutic RNAs. Here we describe how exogenous small interfering RNAs may successfully be introduced into various kinds of human exosomes using electroporation and subsequently delivered to recipient cells. Methods used to confirm the presence of siRNA inside exosomes and cells are presented, such as flow cytometry, confocal microscopy, and Northern blot.

  7. Exosomes released from breast cancer carcinomas stimulate cell movement.

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    Dinari A Harris

    Full Text Available For metastasis to occur cells must communicate with to their local environment to initiate growth and invasion. Exosomes have emerged as an important mediator of cell-to-cell signalling through the transfer of molecules such as mRNAs, microRNAs, and proteins between cells. Exosomes have been proposed to act as regulators of cancer progression. Here, we study the effect of exosomes on cell migration, an important step in metastasis. We performed cell migration assays, endocytosis assays, and exosome proteomic profiling on exosomes released from three breast cancer cell lines that model progressive stages of metastasis. Results from these experiments suggest: (1 exosomes promote cell migration and (2 the signal is stronger from exosomes isolated from cells with higher metastatic potentials; (3 exosomes are endocytosed at the same rate regardless of the cell type; (4 exosomes released from cells show differential enrichment of proteins with unique protein signatures of both identity and abundance. We conclude that breast cancer cells of increasing metastatic potential secrete exosomes with distinct protein signatures that proportionally increase cell movement and suggest that released exosomes could play an active role in metastasis.

  8. Exosomes released from breast cancer carcinomas stimulate cell movement.

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    Harris, Dinari A; Patel, Sajni H; Gucek, Marjan; Hendrix, An; Westbroek, Wendy; Taraska, Justin W

    2015-01-01

    For metastasis to occur cells must communicate with to their local environment to initiate growth and invasion. Exosomes have emerged as an important mediator of cell-to-cell signalling through the transfer of molecules such as mRNAs, microRNAs, and proteins between cells. Exosomes have been proposed to act as regulators of cancer progression. Here, we study the effect of exosomes on cell migration, an important step in metastasis. We performed cell migration assays, endocytosis assays, and exosome proteomic profiling on exosomes released from three breast cancer cell lines that model progressive stages of metastasis. Results from these experiments suggest: (1) exosomes promote cell migration and (2) the signal is stronger from exosomes isolated from cells with higher metastatic potentials; (3) exosomes are endocytosed at the same rate regardless of the cell type; (4) exosomes released from cells show differential enrichment of proteins with unique protein signatures of both identity and abundance. We conclude that breast cancer cells of increasing metastatic potential secrete exosomes with distinct protein signatures that proportionally increase cell movement and suggest that released exosomes could play an active role in metastasis.

  9. EXOSOMES AND TRANSFER OF (EPIGENETIC INFORMATION BY TUMOR CELLS

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    E. M. Tchevkina

    2015-01-01

    Full Text Available In this review, we will introduce the current knowledge about exosomes – vesicles that are generated in the cells and released into the extracellular space. Exosomes are forming in the cell plasma membrane and represent the spherical shapes restricted by their membrane and contained the various biomolecules including nucleic acids, proteins, lipids etc. The intent interest to exosomes is based on their ability to horizontal transfer between the cells, to permeate into vascular system reaching the different tissues and to incorporate into the recipient cells. It was shown that exosome incorporation into the cells lead to remarkable changes in the recipient cells both in genomic level (via the integration of exosomal DNA into the host DNA and in epigenomic level (via the modulation of the content and/or activity of the signaling proteins, microRNA etc.. Undoubtedly, one of the most interesting and perspective achievements in the exosome study is the demonstration of exosome ability to provide the horizontal transfer of the genetic information from cell to cell – the fact supported in the different studies with the various cell models. Here, we will discuss the recent data regarding the main characteristics and properties of exosomes, the role of exosomes in the tumorigenesis including neoplastic transformation, metastasis, multi-drug resistance. The final part of the review involves the most growing area in the exosome study – the possible usage of exosomes in the cancer treatment, in particular – as the specific drug delivery system.

  10. Biogenesis and function of T cell-derived exosomes

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    Miguel Angel Alonso

    2016-08-01

    Full Text Available Exosomes are a particular type of extracellular vesicle, characterized by their endosomal origin as intraluminal vesicles present in large endosomes with a multivesicular structure. After these endosomes fuse with the plasma membrane, exosomes are secreted into the extracellular space. The ability of exosomes to carry and selectively deliver bioactive molecules (e.g., lipids, proteins and nucleic acids confers on them the capacity to modulate the activity of receptor cells, even if these cells are located in distant tissues or organs. Since exosomal cargo depends on cell type, a detailed understanding of the mechanisms that regulate the biochemical composition of exosomes is fundamental to a comprehensive view of exosome function. Here, we review the latest advances concerning exosome function and biogenesis in T cells, with particular focus on the mechanism of protein sorting at multivesicular endosomes. Exosomes secreted by specific T-cell subsets can modulate the activity of immune cells, including other T-cell subsets. Ceramide, tetraspanins and MAL have been revealed to be important in exosome biogenesis by T cells. These molecules, therefore, constitute potential molecular targets for artificially modulating exosome production and, hence, the immune response for therapeutic purposes.

  11. Exosomes Derived from Mesenchymal Stem Cells

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    Bo Yu

    2014-03-01

    Full Text Available The functional mechanisms of mesenchymal stem cells (MSCs have become a research focus in recent years. Accumulating evidence supports the notion that MSCs act in a paracrine manner. Therefore, the biological factors in conditioned medium, including exosomes and soluble factors, derived from MSC cultures are being explored extensively. The results from most investigations show that MSC-conditioned medium or its components mediate some biological functions of MSCs. Several studies have reported that MSC-derived exosomes have functions similar to those of MSCs, such as repairing tissue damage, suppressing inflammatory responses, and modulating the immune system. However, the mechanisms are still not fully understood and the results remain controversial. Compared with cells, exosomes are more stable and reservable, have no risk of aneuploidy, a lower possibility of immune rejection following in vivo allogeneic administration, and may provide an alternative therapy for various diseases. In this review, we summarize the properties and biological functions of MSC-derived exosomes and discuss the related mechanisms.

  12. Hypoxic enhancement of exosome release by breast cancer cells

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    King Hamish W

    2012-09-01

    Full Text Available Abstract Background Exosomes are nanovesicles secreted by tumour cells which have roles in paracrine signalling during tumour progression, including tumour-stromal interactions, activation of proliferative pathways and bestowing immunosuppression. Hypoxia is an important feature of solid tumours which promotes tumour progression, angiogenesis and metastasis, potentially through exosome-mediated signalling. Methods Breast cancer cell lines were cultured under either moderate (1% O2 or severe (0.1% O2 hypoxia. Exosomes were isolated from conditioned media and quantitated by nanoparticle tracking analysis (NTA and immunoblotting for the exosomal protein CD63 in order to assess the impact of hypoxia on exosome release. Hypoxic exosome fractions were assayed for miR-210 by real-time reverse transcription polymerase chain reaction and normalised to exogenous and endogenous control genes. Statistical significance was determined using the Student T test with a P value of  Results Exposure of three different breast cancer cell lines to moderate (1% O2 and severe (0.1% O2 hypoxia resulted in significant increases in the number of exosomes present in the conditioned media as determined by NTA and CD63 immunoblotting. Activation of hypoxic signalling by dimethyloxalylglycine, a hypoxia-inducible factor (HIF hydroxylase inhibitor, resulted in significant increase in exosome release. Transfection of cells with HIF-1α siRNA prior to hypoxic exposure prevented the enhancement of exosome release by hypoxia. The hypoxically regulated miR-210 was identified to be present at elevated levels in hypoxic exosome fractions. Conclusions These data provide evidence that hypoxia promotes the release of exosomes by breast cancer cells, and that this hypoxic response may be mediated by HIF-1α. Given an emerging role for tumour cell-derived exosomes in tumour progression, this has significant implications for understanding the hypoxic tumour phenotype, whereby hypoxic

  13. Exosome-associated hepatitis C virus in cell cultures and patient plasma

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    Liu, Ziqing [Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN 46202 (United States); Zhang, Xiugen [Department of Cell Biology and Immunology, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States); Yu, Qigui [Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN 46202 (United States); He, Johnny J., E-mail: johnny.he@unthsc.edu [Department of Microbiology and Immunology, Indiana University School of Medicine, Indianapolis, IN 46202 (United States); Department of Cell Biology and Immunology, University of North Texas Health Science Center, Fort Worth, TX 76107 (United States)

    2014-12-12

    Highlights: • HCV occurs in both exosome-free and exosome-associated forms. • Exosome-associated HCV is infectious and resistant to neutralizing antibodies. • More exosome-associated HCV than exosome-free HCV is present in patient plasma. - Abstract: Hepatitis C virus (HCV) infects its target cells in the form of cell-free viruses and through cell–cell contact. Here we report that HCV is associated with exosomes. Using highly purified exosomes and transmission electron microscopic imaging, we demonstrated that HCV occurred in both exosome-free and exosome-associated forms. Exosome-associated HCV was infectious and resistant to neutralization by an anti-HCV neutralizing antibody. There were more exosome-associated HCV than exosome-free HCV detected in the plasma of HCV-infected patients. These results suggest exosome-associated HCV as an alternative form for HCV infection and transmission.

  14. Itinerant exosomes: emerging roles in cell and tissue polarity

    OpenAIRE

    2008-01-01

    Cells use secreted signals (e.g. chemokines and growth factors) and sophisticated vehicles such as argosomes, cytonemes, tunneling nanotubes and exosomes to relay important information to other cells, often over large distances. Exosomes, 30–100-nm intraluminal vesicles of multivesicular bodies (MVB) released upon exocytic fusion of the MVB with the plasma membrane, are increasingly recognized as a novel mode of cell-independent communication. Exosomes have been shown to function in antigen p...

  15. Molecular Characterization of Dendritic Cell-Derived Exosomes

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    Théry, Clotilde; Regnault, Armelle; Garin, Jérôme; Wolfers, Joseph; Zitvogel, Laurence; Ricciardi-Castagnoli, Paola; Raposo, Graça; Amigorena, Sebastian

    1999-01-01

    Exosomes are membrane vesicles secreted by hematopoietic cells upon fusion of late multivesicular endosomes with the plasma membrane. Dendritic cell (DC)-derived exosomes induce potent antitumor immune responses in mice, resulting in the regression of established tumors (Zitvogel, L., A. Regnault, A. Lozier, J. Wolfers, C. Flament, D. Tenza, P. Ricciardi-Castagnoli, G. Raposo, and S. Amigorena. 1998. Nat. Med. 4:594–600). To unravel the molecular basis of exosome-induced immune stimulation, w...

  16. Development of exosome surface display technology in living human cells.

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    Stickney, Zachary; Losacco, Joseph; McDevitt, Sophie; Zhang, Zhiwen; Lu, Biao

    2016-03-25

    Surface display technology is an emerging key player in presenting functional proteins for targeted drug delivery and therapy. Although a number of technologies exist, a desirable mammalian surface display system is lacking. Exosomes are extracellular vesicles that facilitate cell-cell communication and can be engineered as nano-shuttles for cell-specific delivery. In this study, we report the development of a novel exosome surface display technology by exploiting mammalian cell secreted nano-vesicles and their trans-membrane protein tetraspanins. By constructing a set of fluorescent reporters for both the inner and outer surface display on exosomes at two selected sites of tetraspanins, we demonstrated the successful exosomal display via gene transfection and monitoring fluorescence in vivo. We subsequently validated our system by demonstrating the expected intracellular partitioning of reporter protein into sub-cellular compartments and secretion of exosomes from human HEK293 cells. Lastly, we established the stable engineered cells to harness the ability of this robust system for continuous production, secretion, and uptake of displayed exosomes with minimal impact on human cell biology. In sum, our work paved the way for potential applications of exosome, including exosome tracking and imaging, targeted drug delivery, as well as exosome-mediated vaccine and therapy.

  17. Exosome-associated hepatitis C virus in cell cultures and patient plasma.

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    Liu, Ziqing; Zhang, Xiugen; Yu, Qigui; He, Johnny J

    2014-12-12

    Hepatitis C virus (HCV) infects its target cells in the form of cell-free viruses and through cell-cell contact. Here we report that HCV is associated with exosomes. Using highly purified exosomes and transmission electron microscopic imaging, we demonstrated that HCV occurred in both exosome-free and exosome-associated forms. Exosome-associated HCV was infectious and resistant to neutralization by an anti-HCV neutralizing antibody. There were more exosome-associated HCV than exosome-free HCV detected in the plasma of HCV-infected patients. These results suggest exosome-associated HCV as an alternative form for HCV infection and transmission.

  18. Biogenesis and function of T cell-derived exosomes

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    Miguel Angel Alonso; Leandro N. Ventimiglia

    2016-01-01

    Exosomes are a particular type of extracellular vesicle, characterized by their endosomal origin as intraluminal vesicles present in large endosomes with a multivesicular structure. After these endosomes fuse with the plasma membrane, exosomes are secreted into the extracellular space. The ability of exosomes to carry and selectively deliver bioactive molecules (e.g., lipids, proteins, and nucleic acids) confers on them the capacity to modulate the activity of receptor cells, even if these ce...

  19. Neural stem cell-derived exosomes mediate viral entry

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    Sims B

    2014-10-01

    Full Text Available Brian Sims,1,2,* Linlin Gu,3,* Alexandre Krendelchtchikov,3 Qiana L Matthews3,4 1Division of Neonatology, Department of Pediatrics, 2Department of Cell, Developmental, and Integrative Biology, 3Division of Infectious Diseases, Department of Medicine, 4Center for AIDS Research, University of Alabama at Birmingham, Birmingham, AL, USA *These authors contributed equally to this work Background: Viruses enter host cells through interactions of viral ligands with cellular receptors. Viruses can also enter cells in a receptor-independent fashion. Mechanisms regarding the receptor-independent viral entry into cells have not been fully elucidated. Exosomal trafficking between cells may offer a mechanism by which viruses can enter cells.Methods: To investigate the role of exosomes on cellular viral entry, we employed neural stem cell-derived exosomes and adenovirus type 5 (Ad5 for the proof-of-principle study. Results: Exosomes significantly enhanced Ad5 entry in Coxsackie virus and adenovirus receptor (CAR-deficient cells, in which Ad5 only had very limited entry. The exosomes were shown to contain T-cell immunoglobulin mucin protein 4 (TIM-4, which binds phosphatidylserine. Treatment with anti-TIM-4 antibody significantly blocked the exosome-mediated Ad5 entry.Conclusion: Neural stem cell-derived exosomes mediated significant cellular entry of Ad5 in a receptor-independent fashion. This mediation may be hampered by an antibody specifically targeting TIM-4 on exosomes. This set of results will benefit further elucidation of virus/exosome pathways, which would contribute to reducing natural viral infection by developing therapeutic agents or vaccines. Keywords: neural stem cell-derived exosomes, adenovirus type 5, TIM-4, viral entry, phospholipids

  20. Exosomes from B cells and Dendritic cells: mechanisms of formation, secretion and targeting

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    Buschow, S.I.

    2006-01-01

    Many cell types, including dendritic cells (DC) and B cells, secrete small vesicles called exosomes. Exosomes from immune cells are thought to have immuno-regulatory functions but their precise role remains unresolved. The aim of the studies presented in this thesis was to get more insight into the factors that determine exosome formation, composition and secretion as well as to learn more about their physiological relevance. Exosomes are equivalent to Luminal Vesicles (LV) of Multi Vesicular...

  1. Exosome Proteome of U-87MG Glioblastoma Cells

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    Sohyun Chun

    2016-12-01

    Full Text Available Exosomes are small membrane vesicles between 30 and 100 nm in diameter secreted by many cell types, and are associated with a wide range of physiological and/or pathological processes. Exosomes containing proteins, lipids, mRNA, and microRNA contribute to cell-to-cell communication and cell-to-environment regulation, however, their biological functions are not yet fully understood. In this report, exosomes in the glioblastoma cell line, U-87MG, were isolated and the proteome was investigated. In addition, exosome proteome changes in U-87MG cells exposed to a low temperature were investigated to elucidate whether the exosome proteome could respond to an external stimulus. Cell culture medium was collected, and exosomes were isolated by continuous centrifugation eliminating cell debris, nucleic acids, and other particles. The morphology of exosomes was observed by cryo-tunneling electron microscopy. According to 2-dimensional electrophoresis and matrix-assisted laser desorption ionization time-of-flight mass spectrometry, certain proteins including collagen type VI alpha 1, putative RNA-binding protein 15B chain A, substrate induced remodeling of the active site regulates HTRA1, coatomer protein complex-subunit beta 2, myosin-heavy chain 1, and keratin-type I cytoskeletal 9 showed differences between the control proteome and the low temperature-exposed proteome.

  2. Exosomes as new vesicular lipid transporters involved in cell-cell communication and various pathophysiologies.

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    Record, Michel; Carayon, Kevin; Poirot, Marc; Silvente-Poirot, Sandrine

    2014-01-01

    Exosomes are nanovesicles that have emerged as a new intercellular communication system between an intracellular compartment of a donor cell towards the periphery or an internal compartment of a recipient cell. The bioactivity of exosomes resides not only in their protein and RNA contents but also in their lipidic molecules. Exosomes display original lipids organized in a bilayer membrane and along with the lipid carriers such as fatty acid binding proteins that they contain, exosomes transport bioactive lipids. Exosomes can vectorize lipids such as eicosanoids, fatty acids, and cholesterol, and their lipid composition can be modified by in-vitro manipulation. They also contain lipid related enzymes so that they can constitute an autonomous unit of production of various bioactive lipids. Exosomes can circulate between proximal or distal cells and their fate can be regulated in part by lipidic molecules. Compared to their parental cells, exosomes are enriched in cholesterol and sphingomyelin and their accumulation in cells might modulate recipient cell homeostasis. Exosome release from cells appears to be a general biological process. They have been reported in all biological fluids from which they can be recovered and can be monitors of specific pathophysiological situations. Thus, the lipid content of circulating exosomes could be useful biomarkers of lipid related diseases. Since the first lipid analysis of exosomes ten years ago detailed knowledge of exosomal lipids has accumulated. The role of lipids in exosome fate and bioactivity and how they constitute an additional lipid transport system are considered in this review.

  3. Exosomes function in cell-cell communication during brain circuit development

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    Sharma, Pranav; Schiapparelli, Lucio; Cline, Hollis T.

    2013-01-01

    Exosomes are small extracellular vesicles that mediate intercellular signaling in the brain without requiring direct contact between cells. Although exosomes have been shown to play a role in neurological diseases and in response to nerve trauma, a role for exosome-mediated signaling in brain development and function has not yet been demonstrated. Here we review data building a case for exosome function in the brain.

  4. Ebola VP40 in exosomes can cause immune cell dysfunction

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    Michelle L Pleet

    2016-11-01

    Full Text Available Ebola virus (EBOV is an enveloped, ssRNA virus from the family Filoviridae capable of causing severe hemorrhagic fever with up to 80-90% mortality rates. The most recent outbreak of EBOV in West Africa starting in 2014 resulted in over 11,300 deaths; however, long-lasting persistence and recurrence in survivors has been documented, potentially leading to further transmission of the virus. We have previously shown that exosomes from cells infected with HIV-1, HTLV-1 and Rift Valley Fever virus are able to transfer viral proteins and non-coding RNAs to naïve recipient cells, resulting in an altered cellular activity. In the current manuscript, we examined the effect of Ebola structural proteins VP40, GP, NP and VLPs on recipient immune cells, as well as the effect of exosomes containing these proteins on naïve immune cells. We found that VP40-transfected cells packaged VP40 into exosomes, and that these exosomes were capable of inducing apoptosis in recipient immune cells. Additionally, we show that presence of VP40 within parental cells or in exosomes delivered to naïve cells could result in the regulation of RNAi machinery including Dicer, Drosha, and Ago 1, which may play a role in the induction of cell death in recipient immune cells. Exosome biogenesis was regulated by VP40 in transfected cells by increasing levels of ESCRT-II proteins EAP20 and EAP45, and exosomal marker proteins CD63 and Alix. VP40 was phosphorylated by Cdk2/Cyclin complexes at Serine 233 which could be reversed with r-Roscovitine treatment. The level of VP40-containing exosomes could also be regulated by treated cells with FDA-approved Oxytetracycline. Additionally, we utilized novel nanoparticles to safely capture VP40 and other viral proteins from Ebola VLPs spiked into human samples using SDS/reducing agents, thus minimizing the need for BSL-4 conditions for most downstream assays. Collectively, our data indicates that VP40 packaged into exosomes may be responsible

  5. Exosomes and nanotubes: Control of immune cell communication.

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    McCoy-Simandle, Kessler; Hanna, Samer J; Cox, Dianne

    2016-02-01

    Cell-cell communication is critical to coordinate the activity and behavior of a multicellular organism. The cells of the immune system not only must communicate with similar cells, but also with many other cell types in the body. Therefore, the cells of the immune system have evolved multiple ways to communicate. Exosomes and tunneling nanotubes (TNTs) are two means of communication used by immune cells that contribute to immune functions. Exosomes are small membrane vesicles secreted by most cell types that can mediate intercellular communication and in the immune system they are proposed to play a role in antigen presentation and modulation of gene expression. TNTs are membranous structures that mediate direct cell-cell contact over several cell diameters in length (and possibly longer) and facilitate the interaction and/or the transfer of signals, material and other cellular organelles between connected cells. Recent studies have revealed additional, but sometimes conflicting, structural and functional features of both exosomes and TNTs. Despite the new and exciting information in exosome and TNT composition, origin and in vitro function, biologically significant functions are still being investigated and determined. In this review, we discuss the current field regarding exosomes and TNTs in immune cells providing evaluation and perspectives of the current literature.

  6. MHC class II-associated proteins in B-cell exosomes and potential functional implications for exosome biogenesis.

    NARCIS (Netherlands)

    Buschow, S.I.; Balkom, B.W.M. van; Aalberts, M.; Heck, A.J.R. van; Wauben, M.; Stoorvogel, W.

    2010-01-01

    Professional antigen-presenting cells secrete major histocompatibility complex class II (MHC II) carrying exosomes with unclear physiological function(s). Exosomes are first generated as the intraluminal vesicles (ILVs) of a specific type of multivesicular body, and are then secreted by fusion of th

  7. Diagnostic technologies for circulating tumour cells and exosomes.

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    Shao, Huilin; Chung, Jaehoon; Issadore, David

    2015-11-24

    Circulating tumour cells (CTCs) and exosomes are promising circulating biomarkers. They exist in easily accessible blood and carry large diversity of molecular information. As such, they can be easily and repeatedly obtained for minimally invasive cancer diagnosis and monitoring. Because of their intrinsic differences in counts, size and molecular contents, CTCs and exosomes pose unique sets of technical challenges for clinical translation-CTCs are rare whereas exosomes are small. Novel technologies are underway to overcome these specific challenges to fully harness the clinical potential of these circulating biomarkers. Herein, we will overview the characteristics of CTCs and exosomes as valuable circulating biomarkers and their associated technical challenges for clinical adaptation. Specifically, we will describe emerging technologies that have been developed to address these technical obstacles and the unique clinical opportunities enabled by technological innovations.

  8. Exosomes Derived from Squamous Head and Neck Cancer Promote Cell Survival after Ionizing Radiation.

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    Mutschelknaus, Lisa; Peters, Carsten; Winkler, Klaudia; Yentrapalli, Ramesh; Heider, Theresa; Atkinson, Michael John; Moertl, Simone

    2016-01-01

    Exosomes are nanometer-sized extracellular vesicles that are believed to function as intercellular communicators. Here, we report that exosomes are able to modify the radiation response of the head and neck cancer cell lines BHY and FaDu. Exosomes were isolated from the conditioned medium of irradiated as well as non-irradiated head and neck cancer cells by serial centrifugation. Quantification using NanoSight technology indicated an increased exosome release from irradiated compared to non-irradiated cells 24 hours after treatment. To test whether the released exosomes influence the radiation response of other cells the exosomes were transferred to non-irradiated and irradiated recipient cells. We found an enhanced uptake of exosomes isolated from both irradiated and non-irradiated cells by irradiated recipient cells compared to non-irradiated recipient cells. Functional analyses by exosome transfer indicated that all exosomes (from non-irradiated and irradiated donor cells) increase the proliferation of non-irradiated recipient cells and the survival of irradiated recipient cells. The survival-promoting effects are more pronounced when exosomes isolated from irradiated compared to non-irradiated donor cells are transferred. A possible mechanism for the increased survival after irradiation could be the increase in DNA double-strand break repair monitored at 6, 8 and 10 h after the transfer of exosomes isolated from irradiated cells. This is abrogated by the destabilization of the exosomes. Our results demonstrate that radiation influences both the abundance and action of exosomes on recipient cells. Exosomes transmit prosurvival effects by promoting the proliferation and radioresistance of head and neck cancer cells. Taken together, this study indicates a functional role of exosomes in the response of tumor cells to radiation exposure within a therapeutic dose range and encourages that exosomes are useful objects of study for a better understanding of tumor

  9. Stromal-cell and cancer-cell exosomes leading the metastatic exodus for the promised niche

    OpenAIRE

    2013-01-01

    Exosomes are thought to play an important role in metastasis. Luga and colleagues have described the production of exosomes by stromal cells such as cancer-associated fibroblasts that are taken up by breast cancer cells and are then loaded with Wnt 11, which is associated with stimulation of the invasiveness and metastasis of the breast cancer cells. Previous studies have shown that exosomes produced by breast cancer cells are taken up by stromal fibroblasts and other stromal cells, suggestin...

  10. MHC class II-associated proteins in B-cell exosomes and potential functional implications for exosome biogenesis.

    Science.gov (United States)

    Buschow, Sonja I; van Balkom, Bas W M; Aalberts, Marian; Heck, Albert J R; Wauben, Marca; Stoorvogel, Willem

    2010-01-01

    Professional antigen-presenting cells secrete major histocompatibility complex class II (MHC II) carrying exosomes with unclear physiological function(s). Exosomes are first generated as the intraluminal vesicles (ILVs) of a specific type of multivesicular body, and are then secreted by fusion of this compartment with the plasma membrane. We have previously shown that in contrast to the sorting of MHC II at lysosomally targeted multivesicular bodies, sorting of MHC II into exosomes does not rely on MHC II ubiquitination. In search for proteins that drive the incorporation of MHC II into exosomes or functionally discriminate exosomal from plasma membrane MHC II, we first analyzed the total proteome of highly purified B cell-derived exosomes using sensitive and accurate mass spectrometry (MS), and identified 539 proteins, including known and not previously identified constituents. Using quantitative MS, we then identified a small subset of proteins that were specifically co-immunoprecipitated with MHC II from detergent-solubilized exosomes. These include HSC71, HSP90, 14-3-3ɛ, CD20 and pyruvate kinase type M2 (PKM2), and we speculate on the functionality of their interaction with exosomal MHC II.

  11. Quantification of plasma exosome is a potential prognostic marker for esophageal squamous cell carcinoma.

    Science.gov (United States)

    Matsumoto, Yasunori; Kano, Masayuki; Akutsu, Yasunori; Hanari, Naoyuki; Hoshino, Isamu; Murakami, Kentaro; Usui, Akihiro; Suito, Hiroshi; Takahashi, Masahiko; Otsuka, Ryota; Xin, Hu; Komatsu, Aki; Iida, Keiko; Matsubara, Hisahiro

    2016-11-01

    Exosomes play important roles in cancer progression. Although its contents (e.g., proteins and microRNAs) have been focused on in cancer research, particularly as potential diagnostic markers, the exosome behavior and methods for exosome quantification remain unclear. In the present study, we analyzed the tumor-derived exosome behavior and assessed the quantification of exosomes in patient plasma as a biomarker for esophageal squamous cell carcinoma (ESCC). A CD63-GFP expressing human ESCC cell line (TE2-CD63-GFP) was made by transfection, and mouse subcutaneous tumor models were established. Fluorescence imaging was performed on tumors and plasma exosomes harvested from mice. GFP-positive small vesicles were confirmed in the plasma obtained from TE2-CD63-GFP tumor-bearing mice. Patient plasma was collected in Chiba University Hospital (n=86). Exosomes were extracted from 100 µl of the plasma and quantified by acetylcholinesterase (AChE) activity. The relationship between exosome quantification and the patient clinical characteristics was assessed. The quantification of exosomes isolated from the patient plasma revealed that esophageal cancer patients (n=66) expressed higher exosome levels than non-malignant patients (n=20) (P=0.0002). Although there was no correlation between the tumor progression and the exosome levels, exosome number was the independent prognostic marker and low levels of exosome predicted a poor prognosis (P=0.03). In conclusion, exosome levels may be useful as an independent prognostic factor for ESCC patients.

  12. Quantification of plasma exosome is a potential prognostic marker for esophageal squamous cell carcinoma

    Science.gov (United States)

    Matsumoto, Yasunori; Kano, Masayuki; Akutsu, Yasunori; Hanari, Naoyuki; Hoshino, Isamu; Murakami, Kentaro; Usui, Akihiro; Suito, Hiroshi; Takahashi, Masahiko; Otsuka, Ryota; Xin, Hu; Komatsu, Aki; Iida, Keiko; Matsubara, Hisahiro

    2016-01-01

    Exosomes play important roles in cancer progression. Although its contents (e.g., proteins and microRNAs) have been focused on in cancer research, particularly as potential diagnostic markers, the exosome behavior and methods for exosome quantification remain unclear. In the present study, we analyzed the tumor-derived exosome behavior and assessed the quantification of exosomes in patient plasma as a biomarker for esophageal squamous cell carcinoma (ESCC). A CD63-GFP expressing human ESCC cell line (TE2-CD63-GFP) was made by transfection, and mouse subcutaneous tumor models were established. Fluorescence imaging was performed on tumors and plasma exosomes harvested from mice. GFP-positive small vesicles were confirmed in the plasma obtained from TE2-CD63-GFP tumor-bearing mice. Patient plasma was collected in Chiba University Hospital (n=86). Exosomes were extracted from 100 µl of the plasma and quantified by acetylcholinesterase (AChE) activity. The relationship between exosome quantification and the patient clinical characteristics was assessed. The quantification of exosomes isolated from the patient plasma revealed that esophageal cancer patients (n=66) expressed higher exosome levels than non-malignant patients (n=20) (P=0.0002). Although there was no correlation between the tumor progression and the exosome levels, exosome number was the independent prognostic marker and low levels of exosome predicted a poor prognosis (P=0.03). In conclusion, exosome levels may be useful as an independent prognostic factor for ESCC patients. PMID:27599779

  13. Modulation of Immune Responses by Exosomes Derived from Antigen-Presenting Cells

    Science.gov (United States)

    Shenoda, Botros B.; Ajit, Seena K.

    2016-01-01

    Exosome-mediated signaling is important in mediating the inflammatory response. To exert their biological or pathophysiological functions in the recipient cells, exosomes deliver a diverse array of biomacromolecules including long and short coding and non-coding RNAs, proteins, and lipids. Exosomes secreted by antigen-presenting cells can confer therapeutic benefits by attenuating or stimulating the immune response. Exosomes play a crucial role in carrying and presenting functional major histocompatibility peptide complexes to modulate antigen-specific T cell responses. Exosomes from Dendritic Cells (DCs) can activate T and B cells and have been explored for their immunostimulatory properties in cancer therapy. The immunosuppressive properties of exosomes derived from macrophages and DCs can reduce inflammation in animal models for several inflammatory disorders. This review focuses on the protective role of exosomes in attenuating inflammation or augmenting immune response, emphasizing studies on exosomes derived from DCs and macrophages. PMID:27660518

  14. Stromal-cell and cancer-cell exosomes leading the metastatic exodus for the promised niche.

    Science.gov (United States)

    Hoffman, Robert M

    2013-06-18

    Exosomes are thought to play an important role in metastasis. Luga and colleagues have described the production of exosomes by stromal cells such as cancer-associated fibroblasts that are taken up by breast cancer cells and are then loaded with Wnt 11, which is associated with stimulation of the invasiveness and metastasis of the breast cancer cells. Previous studies have shown that exosomes produced by breast cancer cells are taken up by stromal fibroblasts and other stromal cells, suggesting that exosomes are agents of cross-talk between cancer and stromal cells to stimulate metastasis. Imaging of exosomes by labeling with fluorescent proteins will enlighten the process by which exosomes enhance metastasis, including premetastatic niche formation.

  15. Drosophila S2 cells secrete wingless on exosome-like vesicles but the wingless gradient forms independently of exosomes.

    Science.gov (United States)

    Beckett, Karen; Monier, Solange; Palmer, Lucy; Alexandre, Cyrille; Green, Hannah; Bonneil, Eric; Raposo, Graca; Thibault, Pierre; Le Borgne, Roland; Vincent, Jean-Paul

    2013-01-01

    Wingless acts as a morphogen in Drosophila wing discs, where it specifies cell fates and controls growth several cell diameters away from its site of expression. Thus, despite being acylated and membrane associated, Wingless spreads in the extracellular space. Recent studies have focussed on identifying the route that Wingless follows in the secretory pathway and determining how it is packaged for release. We have found that, in medium conditioned by Wingless-expressing Drosophila S2 cells, Wingless is present on exosome-like vesicles and that this fraction activates signal transduction. Proteomic analysis shows that Wingless-containing exosome-like structures contain many Drosophila proteins that are homologous to mammalian exosome proteins. In addition, Evi, a multipass transmembrane protein, is also present on exosome-like vesicles. Using these exosome markers and a cell-based RNAi assay, we found that the small GTPase Rab11 contributes significantly to exosome production. This finding allows us to conclude from in vivo Rab11 knockdown experiments, that exosomes are unlikely to contribute to Wingless secretion and gradient formation in wing discs. Consistent with this conclusion, extracellularly tagged Evi expressed from a Bacterial Artificial Chromosome is not released from imaginal disc Wingless-expressing cells.

  16. Heparanase regulates secretion, composition, and function of tumor cell-derived exosomes.

    Science.gov (United States)

    Thompson, Camilla A; Purushothaman, Anurag; Ramani, Vishnu C; Vlodavsky, Israel; Sanderson, Ralph D

    2013-04-05

    Emerging evidence indicates that exosomes play a key role in tumor-host cross-talk and that exosome secretion, composition, and functional capacity are altered as tumors progress to an aggressive phenotype. However, little is known regarding the mechanisms that regulate these changes. Heparanase is an enzyme whose expression is up-regulated as tumors become more aggressive and is associated with enhanced tumor growth, angiogenesis, and metastasis. We have discovered that in human cancer cells (myeloma, lymphoblastoid, and breast cancer), when expression of heparanase is enhanced or when tumor cells are exposed to exogenous heparanase, exosome secretion is dramatically increased. Heparanase enzyme activity is required for robust enhancement of exosome secretion because enzymatically inactive forms of heparanase, even when present in high amounts, do not dramatically increase exosome secretion. Heparanase also impacts exosome protein cargo as reflected by higher levels of syndecan-1, VEGF, and hepatocyte growth factor in exosomes secreted by heparanase-high expressing cells as compared with heparanase-low expressing cells. In functional assays, exosomes from heparanase-high cells stimulated spreading of tumor cells on fibronectin and invasion of endothelial cells through extracellular matrix better than did exosomes secreted by heparanase-low cells. These studies reveal that heparanase helps drive exosome secretion, alters exosome composition, and facilitates production of exosomes that impact both tumor and host cell behavior, thereby promoting tumor progression.

  17. Immature dendritic cell-derived exosomes: a promise subcellular vaccine for autoimmunity.

    Science.gov (United States)

    Yin, Weifan; Ouyang, Song; Li, Yi; Xiao, Bo; Yang, Huan

    2013-02-01

    Exosomes, 60-90-nm-sized vesicles, are produced by a large number of cell types, including tumor cells, neurons, astrocytes, hemocytes, intestinal epithelial cells, and so on. Dendritic cell (DC), the most potent professional antigen-presenting cell in the immune system, produces exosomes in the course of maturation. Mature DCs produce exosomes with the ability to elicit potent immunoactivation, resulting in tumor eradication and bacterial or virus elimination. Given the notion that exosomes are stable and easy to be modified artificially, autologous mature DC-derived exosomes have been vaccinated into patients with malignant diseases. In clinical trials utilizing exosomes as therapeutic approaches, researchers observed considerable curative effect with little side effect. However, immature or suppressive DC-derived exosomes harbor anti-inflammatory properties distinct from mature DC-derived exosomes. In murine models of autoimmune disease and transplantation, immature DC-derived exosomes reduced T cell-dependent immunoactivation, relieved clinical manifestation of autoimmune disease, and prolonged survival time of transplantation. Although the exact mechanism of how immature DC-derived exosomes function in vivo is still unclear, and there are no clinical trials regarding application of exosome vaccine into patients with autoimmune disease, we will analyze the promise of immature DC-derived exosomes as a subcellular vaccine in autoimmunity in this review.

  18. Exosomes from B cells and Dendritic cells: mechanisms of formation, secretion and targeting

    NARCIS (Netherlands)

    Buschow, S.I.

    2006-01-01

    Many cell types, including dendritic cells (DC) and B cells, secrete small vesicles called exosomes. Exosomes from immune cells are thought to have immuno-regulatory functions but their precise role remains unresolved. The aim of the studies presented in this thesis was to get more insight into the

  19. Exosomes released by melanoma cells prepare sentinel lymph nodes for tumor metastasis.

    Science.gov (United States)

    Hood, Joshua L; San, Roman Susana; Wickline, Samuel A

    2011-06-01

    Exosomes are naturally occurring biological nanovesicles utilized by tumors to communicate signals to local and remote cells and tissues. Melanoma exosomes can incite a proangiogenic signaling program capable of remodeling tissue matrices. In this study, we show exosome-mediated conditioning of lymph nodes and define microanatomic responses that license metastasis of melanoma cells. Homing of melanoma exosomes to sentinel lymph nodes imposes synchronized molecular signals that effect melanoma cell recruitment, extracellular matrix deposition, and vascular proliferation in the lymph nodes. Our findings highlight the pathophysiologic role and mechanisms of an exosome-mediated process of microanatomic niche preparation that facilitates lymphatic metastasis by cancer cells.

  20. Melanoma cell-derived exosomes alter macrophage and dendritic cell functions in vitro.

    Science.gov (United States)

    Marton, Annamaria; Vizler, Csaba; Kusz, Erzsebet; Temesfoi, Viktoria; Szathmary, Zsuzsa; Nagy, Krisztina; Szegletes, Zsolt; Varo, Gyorgy; Siklos, Laszlo; Katona, Robert L; Tubak, Vilmos; Howard, O M Zack; Duda, Erno; Minarovits, Janos; Nagy, Katalin; Buzas, Krisztina

    2012-01-01

    To clarify controversies in the literature of the field, we have purified and characterized B16F1 melanoma cell derived exosomes (mcd-exosomes) then we attempted to dissect their immunological activities. We tested how mcd-exosomes influence CD4+ T cell proliferation induced by bone marrow derived dendritic cells; we quantified NF-κB activation in mature macrophages stimulated with mcd-exosomes, and we compared the cytokine profile of LPS-stimulated, IL-4 induced, and mcd-exosome treated macrophages. We observed that mcd-exosomes helped the maturation of dendritic cells, enhancing T cell proliferation induced by the treated dendritic cells. The exosomes also activated macrophages, as measured by NF-κB activation. The cytokine and chemokine profile of macrophages treated with tumor cell derived exosomes showed marked differences from those induced by either LPS or IL-4, and it suggested that exosomes may play a role in the tumor progression and metastasis formation through supporting tumor immune escape mechanisms.

  1. Molecular lipidomics of exosomes released by PC-3 prostate cancer cells

    DEFF Research Database (Denmark)

    Llorente, A.; Skotland, T.; Sylvanne, T.

    2013-01-01

    The molecular lipid composition of exosomes is largely unknown. In this study, sophisticated shotgun and targeted molecular lipidomic assays were performed for in-depth analysis of the lipidomes of the metastatic prostate cancer cell line, PC-3, and their released exosomes. This study, based...... in the quantification of approximately 280 molecular lipid species, provides the most extensive lipid analysis of cells and exosomes to date. Interestingly, major differences were found in the lipid composition of exosomes compared to parent cells. Exosomes show a remarkable enrichment of distinct lipids, demonstrating...

  2. Breast Cancer Exosome-like Microvesicles and Salivary Gland Cells Interplay Alters Salivary Gland Cell-Derived Exosome-like Microvesicles In Vitro

    OpenAIRE

    Lau, Chang S.; Wong, David T. W.

    2012-01-01

    Saliva is a useful biofluid for the early detection of disease, but how distal tumors communicate with the oral cavity and create disease-specific salivary biomarkers remains unclear. Using an in vitro breast cancer model, we demonstrated that breast cancer-derived exosome-like microvesicles are capable of interacting with salivary gland cells, altering the composition of their secreted exosome-like microvesicles. We found that the salivary gland cells secreted exosome-like microvesicles enca...

  3. High-resolution proteomic and lipidomic analysis of exosomes and microvesicles from different cell sources

    Directory of Open Access Journals (Sweden)

    Reka A. Haraszti

    2016-11-01

    Full Text Available Extracellular vesicles (EVs, including exosomes and microvesicles (MVs, are explored for use in diagnostics, therapeutics and drug delivery. However, little is known about the relationship of protein and lipid composition of EVs and their source cells. Here, we report high-resolution lipidomic and proteomic analyses of exosomes and MVs derived by differential ultracentrifugation from 3 different cell types: U87 glioblastoma cells, Huh7 hepatocellular carcinoma cells and human bone marrow-derived mesenchymal stem cells (MSCs. We identified 3,532 proteins and 1,961 lipid species in the screen. Exosomes differed from MVs in several different areas: (a The protein patterns of exosomes were more likely different from their cells of origin than were the protein patterns of MVs; (b The proteomes of U87 and Huh7 exosomes were similar to each other but different from the proteomes of MSC exosomes, whereas the lipidomes of Huh7 and MSC exosomes were similar to each other but different from the lipidomes of U87 exosomes; (c exosomes exhibited proteins of extracellular matrix, heparin-binding, receptors, immune response and cell adhesion functions, whereas MVs were enriched in endoplasmic reticulum, proteasome and mitochondrial proteins. Exosomes and MVs also differed in their types of lipid contents. Enrichment in glycolipids and free fatty acids characterized exosomes, whereas enrichment in ceramides and sphingomyelins characterized MVs. Furthermore, Huh7 and MSC exosomes were specifically enriched in cardiolipins; U87 exosomes were enriched in sphingomyelins. This study comprehensively analyses the protein and lipid composition of exosomes, MVs and source cells in 3 different cell types.

  4. Exosomes and other extracellular vesicles in neural cells and neurodegenerative diseases.

    Science.gov (United States)

    Janas, Anna M; Sapoń, Karolina; Janas, Teresa; Stowell, Michael H B; Janas, Tadeusz

    2016-06-01

    The function of human nervous system is critically dependent on proper interneuronal communication. Exosomes and other extracellular vesicles are emerging as a novel form of information exchange within the nervous system. Intraluminal vesicles within multivesicular bodies (MVBs) can be transported in neural cells anterogradely or retrogradely in order to be released into the extracellular space as exosomes. RNA loading into exosomes can be either via an interaction between RNA and the raft-like region of the MVB limiting membrane, or via an interaction between an RNA-binding protein-RNA complex with this raft-like region. Outflow of exosomes from neural cells and inflow of exosomes into neural cells presumably take place on a continuous basis. Exosomes can play both neuro-protective and neuro-toxic roles. In this review, we characterize the role of exosomes and microvesicles in normal nervous system function, and summarize evidence for defective signaling of these vesicles in disease pathogenesis of some neurodegenerative diseases.

  5. Exosomes derived from human mesenchymal stem cells confer drug resistance in gastric cancer.

    Science.gov (United States)

    Ji, Runbi; Zhang, Bin; Zhang, Xu; Xue, Jianguo; Yuan, Xiao; Yan, Yongmin; Wang, Mei; Zhu, Wei; Qian, Hui; Xu, Wenrong

    2015-08-01

    Mesenchymal stem cells (MSCs) play an important role in chemoresistance. Exosomes have been reported to modify cellular phenotype and function by mediating cell-cell communication. In this study, we aimed to investigate whether exosomes derived from MSCs (MSC-exosomes) are involved in mediating the resistance to chemotherapy in gastric cancer and to explore the underlying molecular mechanism. We found that MSC-exosomes significantly induced the resistance of gastric cancer cells to 5-fluorouracil both in vivo and ex vivo. MSC-exosomes antagonized 5-fluorouracil-induced apoptosis and enhanced the expression of multi-drug resistance associated proteins, including MDR, MRP and LRP. Mechanistically, MSC-exosomes triggered the activation of calcium/calmodulin-dependent protein kinases (CaM-Ks) and Raf/MEK/ERK kinase cascade in gastric cancer cells. Blocking the CaM-Ks/Raf/MEK/ERK pathway inhibited the promoting role of MSC-exosomes in chemoresistance. Collectively, MSC-exosomes could induce drug resistance in gastric cancer cells by activating CaM-Ks/Raf/MEK/ERK pathway. Our findings suggest that MSC-exosomes have profound effects on modifying gastric cancer cells in the development of drug resistance. Targeting the interaction between MSC-exosomes and cancer cells may help improve the efficacy of chemotherapy in gastric cancer.

  6. Exosomes Derived from Human Umbilical Cord Mesenchymal Stem Cells Relieve Acute Myocardial Ischemic Injury

    Directory of Open Access Journals (Sweden)

    Yuanyuan Zhao

    2015-01-01

    Full Text Available This study is aimed at investigating whether human umbilical cord mesenchymal stem cell- (hucMSC- derived exosomes (hucMSC-exosomes have a protective effect on acute myocardial infarction (AMI. Exosomes were characterized under transmission electron microscopy and the particles of exosomes were further examined through nanoparticle tracking analysis. Exosomes (400 μg protein were intravenously administrated immediately following ligation of the left anterior descending (LAD coronary artery in rats. Cardiac function was evaluated by echocardiography and apoptotic cells were counted using TUNEL staining. The cardiac fibrosis was assessed using Masson’s trichrome staining. The Ki67 positive cells in ischemic myocardium were determined using immunohistochemistry. The effect of hucMSC-exosomes on blood vessel formation was evaluated through tube formation and migration of human umbilical vein endothelial cells (EA.hy926 cells. The results indicated that ligation of the LAD coronary artery reduced cardiac function and induced cardiomyocyte apoptosis. Administration of hucMSC-exosomes significantly improved cardiac systolic function and reduced cardiac fibrosis. Moreover, hucMSC-exosomes protected myocardial cells from apoptosis and promoted the tube formation and migration of EA.hy926 cells. It is concluded that hucMSC-exosomes improved cardiac systolic function by protecting myocardial cells from apoptosis and promoting angiogenesis. These effects of hucMSC-exosomes might be associated with regulating the expression of Bcl-2 family.

  7. Free Extracellular miRNA Functionally Targets Cells by Transfecting Exosomes from Their Companion Cells.

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    Krzysztof Bryniarski

    Full Text Available Lymph node and spleen cells of mice doubly immunized by epicutaneous and intravenous hapten application produce a suppressive component that inhibits the action of the effector T cells that mediate contact sensitivity reactions. We recently re-investigated this phenomenon in an immunological system. CD8+ T lymphocyte-derived exosomes transferred suppressive miR-150 to the effector T cells antigen-specifically due to exosome surface coat of antibody light chains made by B1a lymphocytes. Extracellular RNA (exRNA is protected from plasma RNases by carriage in exosomes or by chaperones. Exosome transfer of functional RNA to target cells is well described, whereas the mechanism of transfer of exRNA free of exosomes remains unclear. In the current study we describe extracellular miR-150, extracted from exosomes, yet still able to mediate antigen-specific suppression. We have determined that this was due to miR-150 association with antibody-coated exosomes produced by B1a cell companions of the effector T cells, which resulted in antigen-specific suppression of their function. Thus functional cell targeting by free exRNA can proceed by transfecting companion cell exosomes that then transfer RNA cargo to the acceptor cells. This contrasts with the classical view on release of RNA-containing exosomes from the multivesicular bodies for subsequent intercellular targeting. This new alternate pathway for transfer of exRNA between cells has distinct biological and immunological significance, and since most human blood exRNA is not in exosomes may be relevant to evaluation and treatment of diseases.

  8. Activated human T cells secrete exosomes that participate in IL-2 mediated immune response signaling.

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    Jessica Wahlgren

    Full Text Available It has previously been shown that nano-meter sized vesicles (30-100 nm, exosomes, secreted by antigen presenting cells can induce T cell responses thus showing the potential of exosomes to be used as immunological tools. Additionally, activated CD3⁺ T cells can secrete exosomes that have the ability to modulate different immunological responses. Here, we investigated what effects exosomes originating from activated CD3⁺ T cells have on resting CD3⁺ T cells by studying T cell proliferation, cytokine production and by performing T cell and exosome phenotype characterization. Human exosomes were generated in vitro following CD3⁺ T cell stimulation with anti-CD28, anti-CD3 and IL-2. Our results show that exosomes purified from stimulated CD3⁺ T cells together with IL-2 were able to generate proliferation in autologous resting CD3⁺ T cells. The CD3⁺ T cells stimulated with exosomes together with IL-2 had a higher proportion of CD8⁺ T cells and had a different cytokine profile compared to controls. These results indicate that activated CD3⁺ T cells communicate with resting autologous T cells via exosomes.

  9. Exosomal ITGA3 interferes with non-cancerous prostate cell functions and is increased in urine exosomes of metastatic prostate cancer patients

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    Irene V. Bijnsdorp

    2013-10-01

    Full Text Available Background: Cancer cells are able to change the protein expression and behavior of non-cancerous surrounding cells. Exosomes, secreted by prostate cancer (PCa cells, may have a functional role in cancer metastasis and present a promising source for protein biomarkers. The aim of the present study was to identify which proteins in exosomes can influence non-cancerous cells, and to determine whether we can use urine exosomal proteins to identify high-risk PCa patients. Method: Exosomes were isolated by ultracentrifugation. Migration and invasion were studied by the transwell (invasion assay. Proteomics was performed by LC-MS/MS and identified proteins were validated by Western blotting. Cellular uptake of fluorescent labeled PKH67-exosomes was measured by FACS. Results: Based on comparative protein profiling by mass spectrometry-based proteomics of LNCaP- and PC3-exosomes, we selected ITGA3 and ITGB1, involved in migration/invasion, for further analyses. Inhibition of exosomal ITGA3 reduced the migration and invasion of non-cancerous prostate epithelial cells (prEC almost completely. Cellular uptake of exosomes by prEC was higher with PC3-exosomes compared to LNCaP exosomes. Finally, ITGA3 and ITGB1 were more abundant in urine exosomes of metastatic patients (p<0.05, compared to benign prostate hyperplasia or PCa. Conclusion: These data indicate exosomal ITGA3 and ITGB1 may play a role in manipulating non-cancerous surrounding cells and that measurement of ITGA3 and ITGB1 in urine exosomes has the potential to identify patients with metastatic PCa in a non-invasive manner.

  10. A gestational profile of placental exosomes in maternal plasma and their effects on endothelial cell migration.

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    Carlos Salomon

    Full Text Available Studies completed to date provide persuasive evidence that placental cell-derived exosomes play a significant role in intercellular communication pathways that potentially contribute to placentation and development of materno-fetal vascular circulation. The aim of this study was to establish the gestational-age release profile and bioactivity of placental cell-derived exosome in maternal plasma. Plasma samples (n = 20 per pregnant group were obtained from non-pregnant and pregnant women in the first (FT, 6-12 weeks, second (ST, 22-24 weeks and third (TT, 32-38 weeks trimester. The number of exosomes and placental exosome contribution were determined by quantifying immunoreactive exosomal CD63 and placenta-specific marker (PLAP, respectively. The effect of exosomes isolated from FT, ST and TT on endothelial cell migration were established using a real-time, live-cell imaging system (Incucyte. Exosome plasma concentration was more than 50-fold greater in pregnant women than in non-pregnant women (p<0.001. During normal healthy pregnancy, the number of exosomes present in maternal plasma increased significantly with gestational age by more that two-fold (p<0.001. Exosomes isolated from FT, ST and TT increased endothelial cell migration by 1.9±0.1, 1.6±0.2 and 1.3±0.1-fold, respectively compared to the control. Pregnancy is associated with a dramatic increase in the number of exosomes present in plasma and maternal plasma exosomes are bioactive. While the role of placental cell-derived exosome in regulating maternal and/or fetal vascular responses remains to be elucidated, changes in exosome profile may be of clinical utility in the diagnosis of placental dysfunction.

  11. Trichomonas vaginalis exosomes deliver cargo to host cells and mediate host∶parasite interactions.

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    Olivia Twu

    Full Text Available Trichomonas vaginalis is a common sexually transmitted parasite that colonizes the human urogential tract where it remains extracellular and adheres to epithelial cells. Infections range from asymptomatic to highly inflammatory, depending on the host and the parasite strain. Here, we use a combination of methodologies including cell fractionation, immunofluorescence and electron microscopy, RNA, proteomic and cytokine analyses and cell adherence assays to examine pathogenic properties of T. vaginalis. We have found that T.vaginalis produces and secretes microvesicles with physical and biochemical properties similar to mammalian exosomes. The parasite-derived exosomes are characterized by the presence of RNA and core, conserved exosomal proteins as well as parasite-specific proteins. We demonstrate that T. vaginalis exosomes fuse with and deliver their contents to host cells and modulate host cell immune responses. Moreover, exosomes from highly adherent parasite strains increase the adherence of poorly adherent parasites to vaginal and prostate epithelial cells. In contrast, exosomes from poorly adherent strains had no measurable effect on parasite adherence. Exosomes from parasite strains that preferentially bind prostate cells increased binding of parasites to these cells relative to vaginal cells. In addition to establishing that parasite exosomes act to modulate host∶parasite interactions, these studies are the first to reveal a potential role for exosomes in promoting parasite∶parasite communication and host cell colonization.

  12. Amnion-Epithelial-Cell-Derived Exosomes Demonstrate Physiologic State of Cell under Oxidative Stress.

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    Samantha Sheller

    Full Text Available At term, the signals of fetal maturity and feto-placental tissue aging prompt uterine readiness for delivery by transitioning quiescent myometrium to an active stage. It is still unclear how the signals reach the distant myometrium. Exosomes are a specific type of extracellular vesicle (EVs that transport molecular signals between cells, and are released from a wide range of cells, including the maternal and fetal cells. In this study, we hypothesize that i exosomes act as carriers of signals in utero-placental compartments and ii exosomes reflect the physiologic status of the origin cells. The primary aims of this study were to determine exosomal contents in exosomes derived from primary amnion epithelial cells (AEC. We also determined the effect of oxidative stress on AEC derived exosomal cargo contents. AEC were isolated from amniotic membrane obtained from normal, term, not in labor placentae at delivery, and culture under standard conditions. Oxidative stress was induced using cigarette smoke extract for 48 hours. AEC-conditioned media were collected and exosomes isolated by differential centrifugations. Both growth conditions (normal and oxidative stress induced produced cup shaped exosomes of around 50 nm, expressed exosomes enriched markers, such as CD9, CD63, CD81 and HSC70, embryonic stem cell marker Nanog, and contained similar amounts of cell free AEC DNA. Using confocal microscopy, the colocalization of histone (H 3, heat shock protein (HSP 70 and activated form of pro-senescence and term parturition associated marker p38 mitogen activated protein kinase (MAPK (P-p38 MAPK co-localized with exosome enrich marker CD9. HSP70 and P-p38 MAPK were significantly higher in exosomes from AEC grown under oxidative stress conditions than standard conditions (p<0.05. Finally, mass spectrometry and bioinformatics analysis identified 221 different proteins involved in immunomodulatory response and cell-to-cell communication. This study determined

  13. Exosome Adherence and Internalization by Hepatic Stellate Cells Triggers Sphingosine 1-Phosphate-dependent Migration.

    Science.gov (United States)

    Wang, Ruisi; Ding, Qian; Yaqoob, Usman; de Assuncao, Thiago M; Verma, Vikas K; Hirsova, Petra; Cao, Sheng; Mukhopadhyay, Debabrata; Huebert, Robert C; Shah, Vijay H

    2015-12-25

    Exosomes are cell-derived extracellular vesicles thought to promote intercellular communication by delivering specific content to target cells. The aim of this study was to determine whether endothelial cell (EC)-derived exosomes could regulate the phenotype of hepatic stellate cells (HSCs). Initial microarray studies showed that fibroblast growth factor 2 induced a 2.4-fold increase in mRNA levels of sphingosine kinase 1 (SK1). Exosomes derived from an SK1-overexpressing EC line increased HSC migration 3.2-fold. Migration was not conferred by the dominant negative SK1 exosome. Incubation of HSCs with exosomes was also associated with an 8.3-fold increase in phosphorylation of AKT and 2.5-fold increase in migration. Exosomes were found to express the matrix protein and integrin ligand fibronectin (FN) by Western blot analysis and transmission electron microscopy. Blockade of the FN-integrin interaction with a CD29 neutralizing antibody or the RGD peptide attenuated exosome-induced HSC AKT phosphorylation and migration. Inhibition of endocytosis with transfection of dynamin siRNA, the dominant negative dynamin GTPase construct Dyn2K44A, or the pharmacological inhibitor Dynasore significantly attenuated exosome-induced AKT phosphorylation. SK1 levels were increased in serum exosomes derived from mice with experimental liver fibrosis, and SK1 mRNA levels were up-regulated 2.5-fold in human liver cirrhosis patient samples. Finally, S1PR2 inhibition protected mice from CCl4-induced liver fibrosis. Therefore, EC-derived SK1-containing exosomes regulate HSC signaling and migration through FN-integrin-dependent exosome adherence and dynamin-dependent exosome internalization. These findings advance our understanding of EC/HSC cross-talk and identify exosomes as a potential target to attenuate pathobiology signals.

  14. Exosomes are fingerprints of originating cells: potential biomarkers for ovarian cancer

    Directory of Open Access Journals (Sweden)

    Kobayashi M

    2015-03-01

    Full Text Available Miharu Kobayashi, Gregory E Rice, Jorge Tapia, Murray D Mitchell, Carlos Salomon Exosome Biology Laboratory, Centre for Clinical Diagnostics, University of Queensland Centre for Clinical Research, Royal Brisbane and Women's Hospital, Brisbane, QLD, Australia. Abstract: The past decade has seen an extraordinary explosion of research in the field of extracellular vesicles, especially in a specific type of extracellular vesicles originating from endosomal compartments, called exosomes. Exosomes are a specific subtype of secreted vesicles that are defined as small (~30–120 nm but very stable membrane vesicles that are released from a wide range of cells, including normal and cancer cells. As the content of exosomes is cell type specific, it is believed that they are a "fingerprint" of the releasing cell and its metabolic status. We hypothesized that the exosomes and their specific exosomal content (eg, microribonucleic acid represent a precious biomedical tool and may be used as biomarkers for the diagnosis and prognosis of malignant tumors. In addition, exosomes may modify the phenotype of the parent and/or target cell by transferring pro-oncogenic molecules to induce cancerous phenotype of recipient cells and contribute to the formation of the premetastatic niche. The mechanism involved in these phenomena remains unclear; however, inclusion of signaling mediators into exosomes or exosome release may reduce their intracellular bioavailability in the parent cell, thereby altering cell phenotype and their metastatic potential. The aim of this review therefore is to analyze the biogenesis and role of exosomes from tumor cells, focusing primarily on ovarian cancer. Ovarian cancer is the most lethal gynecologic cancer, and an effective early diagnosis has the potential to improve patient survival. Ovarian cancer currently lacks a reliable method for early detection, however, exosomes have received great attention as potential biomarkers and mediators

  15. Exosomes mediate stromal mobilization of autocrine Wnt-PCP signaling in breast cancer cell migration.

    Science.gov (United States)

    Luga, Valbona; Zhang, Liang; Viloria-Petit, Alicia M; Ogunjimi, Abiodun A; Inanlou, Mohammad R; Chiu, Elaine; Buchanan, Marguerite; Hosein, Abdel Nasser; Basik, Mark; Wrana, Jeffrey L

    2012-12-21

    Stroma in the tumor microenvironment plays a critical role in cancer progression, but how it promotes metastasis is poorly understood. Exosomes are small vesicles secreted by many cell types and enable a potent mode of intercellular communication. Here, we report that fibroblast-secreted exosomes promote breast cancer cell (BCC) protrusive activity and motility via Wnt-planar cell polarity (PCP) signaling. We show that exosome-stimulated BCC protrusions display mutually exclusive localization of the core PCP complexes, Fzd-Dvl and Vangl-Pk. In orthotopic mouse models of breast cancer, coinjection of BCCs with fibroblasts dramatically enhances metastasis that is dependent on PCP signaling in BCCs and the exosome component, Cd81 in fibroblasts. Moreover, we demonstrate that trafficking in BCCs promotes tethering of autocrine Wnt11 to fibroblast-derived exosomes. This work reveals an intercellular communication pathway whereby fibroblast exosomes mobilize autocrine Wnt-PCP signaling to drive BCC invasive behavior.

  16. Imaging exosome transfer from breast cancer cells to stroma at metastatic sites in orthotopic nude-mouse models.

    Science.gov (United States)

    Suetsugu, Atsushi; Honma, Kimi; Saji, Shigetoyo; Moriwaki, Hisataka; Ochiya, Takahiro; Hoffman, Robert M

    2013-03-01

    Exosomes play an important role in cell-to-cell communication to promote tumor metastasis. In order to image the fate of cancer-cell-derived exosomes in orthotopic nude mouse models of breast cancer, we used green fluorescent protein (GFP)-tagged CD63, which is a general marker of exosomes. Breast cancer cells transferred their own exosomes to other cancer cells and normal lung tissue cells in culture. In orthotopic nude-mouse models, breast cancer cells secreted exosomes into the tumor microenvironment. Tumor-derived exosomes were incorporated into tumor-associated cells as well as circulating in the blood of mice with breast cancer metastases. These results suggest that tumor-derived exosomes may contribute to forming a niche to promote tumor growth and metastasis. Our results demonstrate the usefulness of GFP imaging to investigate the role of exosomes in cancer metastasis.

  17. CD109 is a component of exosome secreted from cultured cells.

    Science.gov (United States)

    Sakakura, Hiroki; Mii, Shinji; Hagiwara, Sumitaka; Kato, Takuya; Yamamoto, Noriyuki; Hibi, Hideharu; Takahashi, Masahide; Murakumo, Yoshiki

    2016-01-22

    Exosomes are 50-100-nm-diameter membrane vesicles released from various types of cells. Exosomes retain proteins, mRNAs and miRNAs, which can be transported to surrounding cells. CD109 is a glycosylphosphatidylinositol-anchored glycoprotein, and is released from the cell surface to the culture medium in vitro. Recently, it was reported that secreted CD109 from the cell surface downregulates transforming growth factor-β signaling in human keratinocytes. In this study, we revealed that CD109 is a component of the exosome in conditioned medium. FLAG-tagged human CD109 (FLAG-CD109) in conditioned medium secreted from HEK293 cells expressing FLAG-CD109 (293/FLAG-CD109) was immunoprecipitated with anti-FLAG affinity gel, and the co-precipitated proteins were analyzed by mass spectrometry and western blotting. Exosomal proteins were associated with CD109. We revealed the presence of CD109 in exosome fractions from conditioned medium of 293/FLAG-CD109. Moreover, the localization of CD109 in the exosome was demonstrated using immuno-electron microscopy. When we used HEK293 cells expressing FLAG-tagged truncated CD109, which does not contain the C-terminal region, the association of truncated CD109 with exosomes was not detected in conditioned medium. These findings indicate that CD109 is an exosomal protein and that the C-terminal region of CD109 is required for its presence in the exosome.

  18. Cell type-specific and common characteristics of exosomes derived from mouse cell lines: Yield, physicochemical properties, and pharmacokinetics.

    Science.gov (United States)

    Charoenviriyakul, Chonlada; Takahashi, Yuki; Morishita, Masaki; Matsumoto, Akihiro; Nishikawa, Makiya; Takakura, Yoshinobu

    2017-01-01

    Exosomes are small membrane vesicles secreted from cells and are expected to be used as drug delivery systems. Important characteristics of exosomes, such as yield, physicochemical properties, and pharmacokinetics, may be different among different cell types. However, there is limited information about the effect of cell type on these characteristics. In the present study, we evaluated these characteristics of exosomes derived from five different types of mouse cell lines: B16BL6 murine melanoma cells, C2C12 murine myoblast cells, NIH3T3 murine fibroblasts cells, MAEC murine aortic endothelial cells, and RAW264.7 murine macrophage-like cells. Exosomes were collected using a differential ultracentrifugation method. The exosomes collected from all the cell types were negatively charged globular vesicles with a diameter of approximately 100nm. C2C12 and RAW264.7 cells produced more exosomes than the other types of cells. The exosomes were labeled with a fusion protein of Gaussia luciferase and lactadherin to evaluate their pharmacokinetics. After intravenous injection into mice, all the exosomes rapidly disappeared from the systemic circulation and mainly distributed to the liver. In conclusion, the exosome yield was significantly different among the cell types, and all the exosomes evaluated in this study showed comparable physicochemical and pharmacokinetic properties.

  19. A gestational profile of placental exosomes in maternal plasma and their effects on endothelial cell migration.

    Science.gov (United States)

    Salomon, Carlos; Torres, Maria Jose; Kobayashi, Miharu; Scholz-Romero, Katherin; Sobrevia, Luis; Dobierzewska, Aneta; Illanes, Sebastian E; Mitchell, Murray D; Rice, Gregory E

    2014-01-01

    Studies completed to date provide persuasive evidence that placental cell-derived exosomes play a significant role in intercellular communication pathways that potentially contribute to placentation and development of materno-fetal vascular circulation. The aim of this study was to establish the gestational-age release profile and bioactivity of placental cell-derived exosome in maternal plasma. Plasma samples (n = 20 per pregnant group) were obtained from non-pregnant and pregnant women in the first (FT, 6-12 weeks), second (ST, 22-24 weeks) and third (TT, 32-38 weeks) trimester. The number of exosomes and placental exosome contribution were determined by quantifying immunoreactive exosomal CD63 and placenta-specific marker (PLAP), respectively. The effect of exosomes isolated from FT, ST and TT on endothelial cell migration were established using a real-time, live-cell imaging system (Incucyte). Exosome plasma concentration was more than 50-fold greater in pregnant women than in non-pregnant women (pexosomes present in maternal plasma increased significantly with gestational age by more that two-fold (pExosomes isolated from FT, ST and TT increased endothelial cell migration by 1.9±0.1, 1.6±0.2 and 1.3±0.1-fold, respectively compared to the control. Pregnancy is associated with a dramatic increase in the number of exosomes present in plasma and maternal plasma exosomes are bioactive. While the role of placental cell-derived exosome in regulating maternal and/or fetal vascular responses remains to be elucidated, changes in exosome profile may be of clinical utility in the diagnosis of placental dysfunction.

  20. The role of exosomes contents on genetic and epigenetic alterations of recipient cancer cells

    Directory of Open Access Journals (Sweden)

    Golnoush Dehbashi Behbahani

    2016-10-01

    Full Text Available Exosomes, as a mediator of cell-to-cell transfer of genetic information, act an important role in intercommunication between tumor cells and their niche including fibroblasts, endothelial cells, adipocytes and monocytes. Several studies have shown that tumor cells can influence their neighboring cells by releasing exosomes. These exosomes provide signaling cues for stimulation, activation, proliferation and differentiation of cells. Exosomes contain mRNAs, microRNAs (miRNA, and proteins that could be transferred to target cells inducing genetic and epigenetic changes. By facilitating the horizontal transfer of bioactive molecules such as proteins, RNAs and microRNAs, they are now thought to have vital roles in tumor invasion and metastases, inflammation, coagulation, and stem cell renewal and expansion. The aim of this review article is to discuss the significance of exosome-mediated intercellular communication within the tumor biology.

  1. The role of exosomes contents on genetic and epigenetic alterations of recipient cancer cells

    Science.gov (United States)

    Behbahani, Golnoush Dehbashi; Khani, Soghra; Hosseini, Hamideh Mahmoodzadeh; Abbaszadeh-Goudarzi, Kazem; Nazeri, Saeed

    2016-01-01

    Exosomes, as a mediator of cell-to-cell transfer of genetic information, act an important role in intercommunication between tumor cells and their niche including fibroblasts, endothelial cells, adipocytes and monocytes. Several studies have shown that tumor cells can influence their neighboring cells by releasing exosomes. These exosomes provide signaling cues for stimulation, activation, proliferation and differentiation of cells. Exosomes contain mRNAs, microRNAs (miRNA), and proteins that could be transferred to target cells inducing genetic and epigenetic changes. By facilitating the horizontal transfer of bioactive molecules such as proteins, RNAs and microRNAs, they are now thought to have vital roles in tumor invasion and metastases, inflammation, coagulation, and stem cell renewal and expansion. The aim of this review article is to discuss the significance of exosome-mediated intercellular communication within the tumor biology. PMID:27872698

  2. Tumor exosomes induce tunneling nanotubes in lipid raft-enriched regions of human mesothelioma cells

    Energy Technology Data Exchange (ETDEWEB)

    Thayanithy, Venugopal [Department of Medicine, Division of Hematology, Oncology and Transplantation, University of Minnesota, Minneapolis, MN 55455 (United States); Babatunde, Victor [Moore Laboratory, Department of Cell Biology, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, New York, NY 10021 (United States); Dickson, Elizabeth L. [Department of Obstetrics and Gynecology, Division of Gynecologic Oncology, University of Minnesota, Minneapolis, MN 55455 (United States); Wong, Phillip [Department of Medicine, Division of Hematology, Oncology and Transplantation, University of Minnesota, Minneapolis, MN 55455 (United States); Oh, Sanghoon; Ke, Xu; Barlas, Afsar; Fujisawa, Sho; Romin, Yevgeniy [Molecular Cytology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021 (United States); Moreira, André L. [Department of Pathology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021 (United States); Downey, Robert J. [Department of Surgery, Memorial Sloan-Kettering Cancer Center, New York, NY 10021 (United States); Steer, Clifford J. [Departments of Medicine and Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455 (United States); Subramanian, Subbaya [Department of Surgery, University of Minnesota, Minneapolis, MN 55455 (United States); Manova-Todorova, Katia [Molecular Cytology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021 (United States); Moore, Malcolm A.S. [Moore Laboratory, Department of Cell Biology, Sloan-Kettering Institute, Memorial Sloan-Kettering Cancer Center, New York, NY 10021 (United States); Lou, Emil, E-mail: emil-lou@umn.edu [Department of Medicine, Division of Hematology, Oncology and Transplantation, University of Minnesota, Minneapolis, MN 55455 (United States)

    2014-04-15

    Tunneling nanotubes (TnTs) are long, non-adherent, actin-based cellular extensions that act as conduits for transport of cellular cargo between connected cells. The mechanisms of nanotube formation and the effects of the tumor microenvironment and cellular signals on TnT formation are unknown. In the present study, we explored exosomes as potential mediators of TnT formation in mesothelioma and the potential relationship of lipid rafts to TnT formation. Mesothelioma cells co-cultured with exogenous mesothelioma-derived exosomes formed more TnTs than cells cultured without exosomes within 24–48 h; and this effect was most prominent in media conditions (low-serum, hyperglycemic medium) that support TnT formation (1.3–1.9-fold difference). Fluorescence and electron microscopy confirmed the purity of isolated exosomes and revealed that they localized predominantly at the base of and within TnTs, in addition to the extracellular environment. Time-lapse microscopic imaging demonstrated uptake of tumor exosomes by TnTs, which facilitated intercellular transfer of these exosomes between connected cells. Mesothelioma cells connected via TnTs were also significantly enriched for lipid rafts at nearly a 2-fold higher number compared with cells not connected by TnTs. Our findings provide supportive evidence of exosomes as potential chemotactic stimuli for TnT formation, and also lipid raft formation as a potential biomarker for TnT-forming cells. - Highlights: • Exosomes derived from malignant cells can stimulate an increased rate in the formation of tunneling nanotubes. • Tunneling nanotubes can serve as conduits for intercellular transfer of these exosomes. • Most notably, exosomes derived from benign mesothelial cells had no effect on nanotube formation. • Cells forming nanotubes were enriched in lipid rafts at a greater number compared with cells not forming nanotubes. • Our findings suggest causal and potentially synergistic association of exosomes and

  3. Profile of Exosomal and Intracellular microRNA in Gamma-Herpesvirus-Infected Lymphoma Cell Lines

    Science.gov (United States)

    Hoshina, Shiho; Sekizuka, Tsuyoshi; Kataoka, Michiyo; Hasegawa, Hideki; Hamada, Hiromichi; Kuroda, Makoto; Katano, Harutaka

    2016-01-01

    Exosomes are small vesicles released from cells, into which microRNAs (miRNA) are specifically sorted and accumulated. Two gamma-herpesviruses, Kaposi sarcoma-associated herpesvirus (KSHV) and Epstein—Barr virus (EBV), encode miRNAs in their genomes and express virus-encoded miRNAs in cells and exosomes. However, there is little information about the detailed distribution of virus-encoded miRNAs in cells and exosomes. In this study, we thus identified virus- and host-encoded miRNAs in exosomes released from KSHV- or EBV-infected lymphoma cell lines and compared them with intracellular miRNAs using a next-generation sequencer. Sequencing analysis demonstrated that 48% of the annotated miRNAs in the exosomes from KSHV-infected cells originated from KSHV. Human mir-10b-5p and mir-143-3p were much more highly concentrated in exosomes than in cells. Exosomes contained more nonexact mature miRNAs that did not exactly match those in miRBase than cells. Among the KSHV-encoded miRNAs, miRK12-3-5p was the most abundant exact mature miRNA in both cells and exosomes that exactly matched those in miRBase. Recently identified EXOmotifs, nucleotide motifs that control the loading of miRNAs into exosomes were frequently found within the sequences of KSHV-encoded miRNAs, and the presence of the EXOmotif CCCT or CCCG was associated with the localization of miRNA in exosomes in KSHV-infected cells. These observations suggest that specific virus-encoded miRNAs are sorted by EXOmotifs and accumulate in exosomes in virus-infected cells. PMID:27611973

  4. Exosomes as potent cell-free peptide-based vaccine. II. Exosomes in CpG adjuvants efficiently prime naive Tc1 lymphocytes leading to tumor rejection.

    NARCIS (Netherlands)

    Chaput, N.; Schartz, N.E.; Andre, F.; Taieb, J.; Novault, S.; Bonnaventure, P.; Aubert, N.; Bernard, J.; Lemonnier, F.; Merad, M.; Adema, G.J.; Adams, M.; Ferrantini, M.; Carpentier, A.F.; Escudier, B.; Tursz, T.; Angevin, E.; Zitvogel, L.

    2004-01-01

    Ideal vaccines should be stable, safe, molecularly defined, and out-of-shelf reagents efficient at triggering effector and memory Ag-specific T cell-based immune responses. Dendritic cell-derived exosomes could be considered as novel peptide-based vaccines because exosomes harbor a discrete set of p

  5. Rab27A regulates exosome secretion from lung adenocarcinoma cells A549: involvement of EPI64.

    Science.gov (United States)

    Li, Wenhai; Hu, Yunsheng; Jiang, Tao; Han, Yong; Han, Guoliang; Chen, Jiakuan; Li, Xiaofei

    2014-11-01

    Exosomes are small membrane vesicles secreted into the extracellular compartment by exocytosis. The unique composition of exosomes can be transported to other cells which allow cells to exert biological functions at distant sites. However, in lung cancer, the regulation of exosome secretion was poorly understood. In this study, we employed human lung adenocarcinoma A549 cells to determine the exosome secretion and involved regulation mechanism. We found that Rab27A was expressed in A549 cells and the reduction of Rab27A by Rab27A-specific shRNA could significantly decrease the secretion of exosome by A549 cells. EPI64, a candidate GAP that is specific for Rab27, was also detected in A549 cells. By pull-down assay, we found that EPI64 participated in the exosome secretion of A549 cells by acting as a specific GAP for Rab27A, not Rab27B. Overexpression of EPI64 enhanced exosome secretion. Taken together, in A549 cells, EPI64 could regulate the exosome secretion by functioning as a GAP specific for Rab27A.

  6. Exosomal proteins as prognostic biomarkers in non-small cell lung cancer

    DEFF Research Database (Denmark)

    Paulsen, Birgitte Sandfeld; Aggerholm-Pedersen, N; Bæk, R

    2016-01-01

    BACKGROUND: Use of exosomes as biomarkers in non-small cell lung cancer (NSCLC) is an intriguing approach in the liquid-biopsy era. Exosomes are nano-sized vesicles with membrane-bound proteins that reflect their originating cell. Prognostic biomarkers are needed to improve patient selection...

  7. Activated T cell exosomes promote tumor invasion via Fas signaling pathway.

    Science.gov (United States)

    Cai, Zhijian; Yang, Fei; Yu, Lei; Yu, Zhou; Jiang, Lingling; Wang, Qingqing; Yang, Yunshan; Wang, Lie; Cao, Xuetao; Wang, Jianli

    2012-06-15

    Activated T cells release bioactive Fas ligand (FasL) in exosomes, which subsequently induce self-apoptosis of T cells. However, their potential effects on cell apoptosis in tumors are still unknown. In this study, we purified exosomes expressing FasL from activated CD8(+) T cell from OT-I mice and found that activated T cell exosomes had little effect on apoptosis and proliferation of tumor cells but promoted the invasion of B16 and 3LL cancer cells in vitro via the Fas/FasL pathway. Activated T cell exosomes increased the amount of cellular FLICE inhibitory proteins and subsequently activated the ERK and NF-κB pathways, which subsequently increased MMP9 expression in the B16 murine melanoma cells. In a tumor-invasive model in vivo, we observed that the activated T cell exosomes promoted the migration of B16 tumor cells to lung. Interestingly, pretreatment with FasL mAb significantly reduced the migration of B16 tumor cells to lung. Furthermore, CD8 and FasL double-positive exosomes from tumor mice, but not normal mice, also increased the expression of MMP9 and promoted the invasive ability of B16 murine melanoma and 3LL lung cancer cells. In conclusion, our results indicate that activated T cell exosomes promote melanoma and lung cancer cell metastasis by increasing the expression of MMP9 via Fas signaling, revealing a new mechanism of tumor immune escape.

  8. Stem cell-derived exosomes as a therapeutic tool for cardiovascular disease

    Science.gov (United States)

    Suzuki, Etsu; Fujita, Daishi; Takahashi, Masao; Oba, Shigeyoshi; Nishimatsu, Hiroaki

    2016-01-01

    Mesenchymal stem cells (MSCs) have been used to treat patients suffering from acute myocardial infarction (AMI) and subsequent heart failure. Although it was originally assumed that MSCs differentiated into heart cells such as cardiomyocytes, recent evidence suggests that the differentiation capacity of MSCs is minimal and that injected MSCs restore cardiac function via the secretion of paracrine factors. MSCs secrete paracrine factors in not only naked forms but also membrane vesicles including exosomes containing bioactive substances such as proteins, messenger RNAs, and microRNAs. Although the details remain unclear, these bioactive molecules are selectively sorted in exosomes that are then released from donor cells in a regulated manner. Furthermore, exosomes are specifically internalized by recipient cells via ligand-receptor interactions. Thus, exosomes are promising natural vehicles that stably and specifically transport bioactive molecules to recipient cells. Indeed, stem cell-derived exosomes have been successfully used to treat cardiovascular disease (CVD), such as AMI, stroke, and pulmonary hypertension, in animal models, and their efficacy has been demonstrated. Therefore, exosome administration may be a promising strategy for the treatment of CVD. Furthermore, modifications of exosomal contents may enhance their therapeutic effects. Future clinical studies are required to confirm the efficacy of exosome treatment for CVD. PMID:27679686

  9. Characterization of Uptake and Internalization of Exosomes by Bladder Cancer Cells

    Directory of Open Access Journals (Sweden)

    Carrie A. Franzen

    2014-01-01

    Full Text Available Bladder tumors represent a special therapeutic challenge as they have a high recurrence rate requiring repeated interventions and may progress to invasive or metastatic disease. Exosomes carry proteins implicated in bladder cancer progression and have been implicated in bladder cancer cell survival. Here, we characterized exosome uptake and internalization by human bladder cancer cells using Amnis ImageStreamX, an image cytometer. Exosomes were isolated by ultracentrifugation from bladder cancer culture conditioned supernatant, labeled with PKH-26, and analyzed on the ImageStreamX with an internal standard added to determine concentration. Exosomes were cocultured with bladder cancer cells and analyzed for internalization. Using the IDEAS software, we determined exosome uptake based on the number of PKH-26+ spots and overall PKH-26 fluorescence intensity. Using unlabeled beads of a known concentration and size, we were able to determine concentrations of exosomes isolated from bladder cancer cells. We measured exosome uptake by recipient bladder cancer cells, and we demonstrated that uptake is dose and time dependent. Finally, we found that uptake is active and specific, which can be partially blocked by heparin treatment. The characterization of cellular uptake and internalization by bladder cancer cells may shed light on the role of exosomes on bladder cancer recurrence and progression.

  10. Mast cell synapses and exosomes: membrane contacts for information exchange

    Directory of Open Access Journals (Sweden)

    Amanda eCarroll-Portillo

    2012-03-01

    Full Text Available In addition to their central role in allergy, mast cells are involved in a wide variety of cellular interactions during homeostasis and disease. In this review, we discuss the ability of mast cells to extend their mechanisms for intercellular communication beyond the release of soluble mediators. These include formation of mast cell synapses on antigen presenting surfaces, as well as cell-cell contacts with dendritic cells and T cells. Release of membrane-bound exosomes also provide for the transfer of antigen, mast cell proteins and RNA to other leukocytes. With the recognition of the extended role mast cells have during immune modulation, further investigation of the processes in which mast cells are involved is necessary. This reopens mast cell research to exciting possibilities, demonstrating it to be an immunological frontier.

  11. Cell Infectivity in relation to bovine leukemia virus gp51 and p24 in bovine milk exosomes.

    Directory of Open Access Journals (Sweden)

    Tetsuya Yamada

    Full Text Available Exosomes are small membranous microvesicles (40-100 nm in diameter and are extracellularly released from a wide variety of cells. Exosomes contain microRNA, mRNA, and cellular proteins, which are delivered into recipient cells via these exosomes, and play a role in intercellular communication. In bovine leukemia virus (BLV infection of cattle, although it is thought to be a minor route of infection, BLV can be transmitted to calves via milk. Here, we investigated the association between exosomes and BLV in bovine milk. BLV structural proteins, gp51 (Env and p24 (Gag, were detected in bovine milk exosomes from BLV-infected cattle by Western blot analysis. In cells inoculated with these milk exosomes, BLV DNA was not detected during three serial passages by nested PCR. Purification of exosomes from persistently BLV-infected cells was achieved by immuno-magnetic separation using an antibody against exosomes coupled to magnetic beads. Consistently, BLV gp51 and p24 proteins were detected in purified exosomes. Moreover, reverse transcriptase activity was observed in purified exosomes, meaning that exosomes also contain viral enzyme. However, BLV DNA was not detected in serially passaged cells after inoculation of purified exosomes, indicating that exosomes carrying BLV proteins appeared to be not infectious. These results suggest that BLV proteins are released with milk exosomes and could be transferred into recipient cells of calves via milk exosomes as an alternative route not requiring virus infection. Moreover it is also possible that bovine milk exosomes play a role in clearance of BLV proteins from infected cells.

  12. Proteomic analysis of exosomes from nasopharyngeal carcinoma cell identifies intercellular transfer of angiogenic proteins

    KAUST Repository

    Chan, Yuk-kit

    2015-04-01

    Exosomes, a group of secreted extracellular nanovesicles containing genetic materials and signaling molecules, play a critical role in intercellular communication. During tumorigenesis, exosomes have been demonstrated to promote tumor angiogenesis and metastasis while their biological functions in nasopharyngeal carcinoma (NPC) are poorly understood. In this study, we focused on the role of NPC-derived exosomes on angiogenesis. Exosomes derived from the NPC C666-1 cells and immortalized nasopharyngeal epithelial cells (NP69 and NP460) were isolated using ultracentrifugation. The molecular profile and biophysical characteristics of exosomes were verified by Western blotting, sucrose density gradient, and electron microscopy. We showed that the C666-1 exosomes (10 and 20 μg/ml) could significantly increase the tubulogenesis, migration and invasion of human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner. Subsequently, an iTRAQ-based quantitative proteomics was used to identify the differentially expressed proteins in C666-1 exosomes. Among the 640 identified proteins, 51 and 89 proteins were considered as up- and down-regulated (≥ 1.5-fold variations) in C666-1 exosomes compared to the normal counterparts, respectively. As expected, pro-angiogenic proteins including intercellular adhesion molecule-1 (ICAM-1) and CD44 variant isoform 5 (CD44v5) are among the up-regulated proteins, whereas angio-suppressive protein, thrombospondin-1 (TSP-1) was down-regulated in C666-1 exosomes. Further confocal microscopic study and Western blotting clearly demonstrated that the alteration of ICAM-1, and TSP-1 expressions in recipient HUVECs are due to internalization of exosomes. Taken together, these data strongly indicated the critical roles of identified angiogenic proteins in the involvement of exosomes-induced angiogenesis, which could potentially be developed as therapeutic targets in future. This article is protected by copyright. All rights reserved.

  13. Proteomic analysis of exosomes from nasopharyngeal carcinoma cell identifies intercellular transfer of angiogenic proteins.

    Science.gov (United States)

    Chan, Yuk-Kit; Zhang, Huoming; Liu, Pei; Tsao, Sai-Wah; Lung, Maria Li; Mak, Nai-Ki; Ngok-Shun Wong, Ricky; Ying-Kit Yue, Patrick

    2015-10-15

    Exosomes, a group of secreted extracellular nanovesicles containing genetic materials and signaling molecules, play a critical role in intercellular communication. During tumorigenesis, exosomes have been demonstrated to promote tumor angiogenesis and metastasis while their biological functions in nasopharyngeal carcinoma (NPC) are poorly understood. In this study, we focused on the role of NPC-derived exosomes on angiogenesis. Exosomes derived from the NPC C666-1 cells and immortalized nasopharyngeal epithelial cells (NP69 and NP460) were isolated using ultracentrifugation. The molecular profile and biophysical characteristics of exosomes were verified by Western blotting, sucrose density gradient and electron microscopy. We showed that the C666-1 exosomes (10 and 20 μg/ml) could significantly increase the tubulogenesis, migration and invasion of human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner. Subsequently, an iTRAQ-based quantitative proteomics was used to identify the differentially expressed proteins in C666-1 exosomes. Among the 640 identified proteins, 51 and 89 proteins were considered as up- and down-regulated (≥ 1.5-fold variations) in C666-1 exosomes compared to the normal counterparts, respectively. As expected, pro-angiogenic proteins including intercellular adhesion molecule-1 (ICAM-1) and CD44 variant isoform 5 (CD44v5) are among the up-regulated proteins, whereas angio-suppressive protein, thrombospondin-1 (TSP-1) was down-regulated in C666-1 exosomes. Further confocal microscopic study and Western blotting clearly demonstrated that the alteration of ICAM-1 and TSP-1 expressions in recipient HUVECs are due to internalization of exosomes. Taken together, these data strongly indicated the critical roles of identified angiogenic proteins in the involvement of exosomes-induced angiogenesis, which could potentially be developed as therapeutic targets in future.

  14. Morphologic and proteomic characterization of exosomes released by cultured extravillous trophoblast cells

    Energy Technology Data Exchange (ETDEWEB)

    Atay, Safinur [Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, KY (United States); Gercel-Taylor, Cicek [Obstetrics, Gynecology and Women' s Health, University of Louisville School of Medicine, Louisville, KY (United States); Kesimer, Mehmet [Department of Biochemistry and Biophysics, University of North Carolina at Chapel Hill, Chapel Hill, NC (United States); Taylor, Douglas D., E-mail: ddtaylor@louisville.edu [Department of Microbiology and Immunology, University of Louisville School of Medicine, Louisville, KY (United States); Obstetrics, Gynecology and Women' s Health, University of Louisville School of Medicine, Louisville, KY (United States)

    2011-05-01

    Exosomes represent an important intercellular communication vehicle, mediating events essential for the decidual microenvironment. While we have demonstrated exosome induction of pro-inflammatory cytokines, to date, no extensive characterization of trophoblast-derived exosomes has been provided. Our objective was to provide a morphologic and proteomic characterization of these exosomes. Exosomes were isolated from the conditioned media of Swan71 human trophoblast cells by ultrafiltration and ultracentrifugation. These were analyzed for density (sucrose density gradient centrifugation), morphology (electron microscopy), size (dynamic light scattering) and protein composition (Ion Trap mass spectrometry and western immunoblotting). Based on density gradient centrifugation, microvesicles from Sw71 cells exhibit a density between 1.134 and 1.173 g/ml. Electron microscopy demonstrated that microvesicles from Sw71 cells exhibit the characteristic cup-shaped morphology of exosomes. Dynamic light scattering showed a bell-shaped curve, indicating a homogeneous population with a mean size of 165 nm {+-} 0.5 nm. Ion Trap mass spectrometry demonstrated the presence of exosome marker proteins (including CD81, Alix, cytoskeleton related proteins, and Rab family). The MS results were confirmed by western immunoblotting. Based on morphology, density, size and protein composition, we defined the release of exosomes from extravillous trophoblast cells and provide their first extensive characterization. This characterization is essential in furthering our understanding of 'normal' early pregnancy.

  15. Wnt5b-associated exosomes promote cancer cell migration and proliferation.

    Science.gov (United States)

    Harada, Takeshi; Yamamoto, Hideki; Kishida, Shosei; Kishida, Michiko; Awada, Chihiro; Takao, Toshifumi; Kikuchi, Akira

    2017-01-01

    Wnt5b is a member of the same family of proteins as Wnt5a, the overexpression of which is associated with cancer aggressiveness. Wnt5b is also suggested to be involved in cancer progression, however, details remain unclarified. We analyzed the biochemical properties of purified Wnt5b and the mode of secretion of Wnt5b by cancer cells. Wnt5b was glycosylated at three asparagine residues and lipidated at one serine residue, and these post-translational modifications of Wnt5b were essential for secretion. Purified Wnt5b showed Dvl2 phosphorylation and Rac activation abilities to a similar extent as Wnt5a. In cultured-cell conditioned medium, Wnt5b was detected in supernatant or precipitation fractions that were separated by centrifugation at 100 000 g. In PANC-1 pancreatic cancer cells, 55% of secreted endogenous Wnt5b was associated with exosomes. Exosomes from wild-type PANC-1 cells, but not those from Wnt5b-knockout PANC-1 cells, activated Wnt5b signaling in CHO cells and stimulated migration and proliferation of A549 lung adenocarcinoma cells, suggesting that endogenous, Wnt5b-associated exosomes are active. The exosomes were taken up by CHO cells and immunoelectron microscopy revealed that Wnt5b is indeed associated with exosomes. In Caco-2 colon cancer cells, most Wnt5b was recovered in precipitation fractions when Wnt5b was ectopically expressed (Caco-2/Wnt5b cells). Knockdown of TSG101, an exosome marker, decreased the secretion of Wnt5b-associated exosomes from Caco-2/Wnt5b cells and inhibited Wnt5b-dependent cell proliferation. Exosomes secreted from Caco-2/Wnt5b cells stimulated migration and proliferation of A549 cells. These results suggest that Wnt5b-associated exosomes promote cancer cell migration and proliferation in a paracrine manner.

  16. MicroRNA and protein profiling of brain metastasis competent cell-derived exosomes.

    Directory of Open Access Journals (Sweden)

    Laura Camacho

    Full Text Available Exosomes are small membrane vesicles released by most cell types including tumor cells. The intercellular exchange of proteins and genetic material via exosomes is a potentially effective approach for cell-to-cell communication and it may perform multiple functions aiding to tumor survival and metastasis. We investigated microRNA and protein profiles of brain metastatic (BM versus non-brain metastatic (non-BM cell-derived exosomes. We studied the cargo of exosomes isolated from brain-tropic 70W, MDA-MB-231BR, and circulating tumor cell brain metastasis-selected markers (CTC1BMSM variants, and compared them with parental non-BM MeWo, MDA-MB-231P and CTC1P cells, respectively. By performing microRNA PCR array we identified one up-regulated (miR-210 and two down-regulated miRNAs (miR-19a and miR-29c in BM versus non-BM exosomes. Second, we analyzed the proteomic content of cells and exosomes isolated from these six cell lines, and detected high expression of proteins implicated in cell communication, cell cycle, and in key cancer invasion and metastasis pathways. Third, we show that BM cell-derived exosomes can be internalized by non-BM cells and that they effectively transport their cargo into cells, resulting in increased cell adhesive and invasive potencies. These results provide a strong rationale for additional investigations of exosomal proteins and miRNAs towards more profound understandings of exosome roles in brain metastasis biogenesis, and for the discovery and application of non-invasive biomarkers for new therapies combating brain metastasis.

  17. MicroRNA and protein profiling of brain metastasis competent cell-derived exosomes.

    Science.gov (United States)

    Camacho, Laura; Guerrero, Paola; Marchetti, Dario

    2013-01-01

    Exosomes are small membrane vesicles released by most cell types including tumor cells. The intercellular exchange of proteins and genetic material via exosomes is a potentially effective approach for cell-to-cell communication and it may perform multiple functions aiding to tumor survival and metastasis. We investigated microRNA and protein profiles of brain metastatic (BM) versus non-brain metastatic (non-BM) cell-derived exosomes. We studied the cargo of exosomes isolated from brain-tropic 70W, MDA-MB-231BR, and circulating tumor cell brain metastasis-selected markers (CTC1BMSM) variants, and compared them with parental non-BM MeWo, MDA-MB-231P and CTC1P cells, respectively. By performing microRNA PCR array we identified one up-regulated (miR-210) and two down-regulated miRNAs (miR-19a and miR-29c) in BM versus non-BM exosomes. Second, we analyzed the proteomic content of cells and exosomes isolated from these six cell lines, and detected high expression of proteins implicated in cell communication, cell cycle, and in key cancer invasion and metastasis pathways. Third, we show that BM cell-derived exosomes can be internalized by non-BM cells and that they effectively transport their cargo into cells, resulting in increased cell adhesive and invasive potencies. These results provide a strong rationale for additional investigations of exosomal proteins and miRNAs towards more profound understandings of exosome roles in brain metastasis biogenesis, and for the discovery and application of non-invasive biomarkers for new therapies combating brain metastasis.

  18. More Than Tiny Sacks: Stem Cell Exosomes as Cell-Free Modality for Cardiac Repair.

    Science.gov (United States)

    Kishore, Raj; Khan, Mohsin

    2016-01-22

    Stem cell therapy provides immense hope for regenerating the pathological heart, yet has been marred by issues surrounding the effectiveness, unclear mechanisms, and survival of the donated cell population in the ischemic myocardial milieu. Poor survival and engraftment coupled to inadequate cardiac commitment of the adoptively transferred stem cells compromises the improvement in cardiac function. Various alternative approaches to enhance the efficacy of stem cell therapies and to overcome issues with cell therapy have been used with varied success. Cell-free components, such as exosomes enriched in proteins, messenger RNAs, and miRs characteristic of parental stem cells, represent a potential approach for treating cardiovascular diseases. Recently, exosomes from different kinds of stem cells have been effectively used to promote cardiac function in the pathological heart. The aim of this review is to summarize current research efforts on stem cell exosomes, including their potential benefits and limitations to develop a potentially viable therapy for cardiovascular problems.

  19. Exosomes derived from mineralizing osteoblasts promote ST2 cell osteogenic differentiation by alteration of microRNA expression.

    Science.gov (United States)

    Cui, Yazhou; Luan, Jing; Li, Haiying; Zhou, Xiaoyan; Han, Jinxiang

    2016-01-01

    Mineralizing osteoblasts (MOBs) can release exosomes, although the functional significance remains unclear. In the present study, we demonstrate that exosomes derived from mineralizing pre-osteoblast MC3T3-E1 cells can promote bone marrow stromal cell (ST2) differentiation to osteoblasts. We reveal that MOB-derived exosomes significantly influence miRNA profiles in recipient ST2 cells, and these changes tend to activate the Wnt signaling pathway by inhibiting Axin1 expression and increasing β-catenin expression. We also suggest that MOB derived-exosomes partly induce the variation in miRNA expression in recipient ST2 cells by exosomal miRNA transfer. These findings suggest an exosome-mediated mode of cell-to-cell communication in the osteogenic microenvironment, and also indicate the potential of MOB exosomes in bone tissue engineering.

  20. Exosomes of BV-2 cells induced by alpha-synuclein: important mediator of neurodegeneration in PD.

    Science.gov (United States)

    Chang, Chongwang; Lang, Hongjuan; Geng, Ning; Wang, Jing; Li, Nan; Wang, Xuelian

    2013-08-26

    Parkinson's disease (PD) is a progressive neurodegenerative disease. Alpha-synuclein aggregation, which can activate microglia to enhance its dopaminergic neurotoxicity, plays a central role in the progression of PD. However the mechanism is still unclear. To investigate how alpha-synuclein affects the neuron, exosomes were derived from alpha-synuclein treated mouse microglia cell line BV-2 cells by differential centrifugation and ultracentrifugation. We found that alpha-synuclein can induce an increase of exosomal secretion by microglia. These activated exosomes expressed a high level of MHC class II molecules and membrane TNF-α. In addition, the activated exosomes cause increased apoptosis. Exosomes secreted from activated microglias might be important mediator of alpha-synuclein-induced neurodegeneration in PD.

  1. β-Elemene Reverses Chemoresistance of Breast Cancer Cells by Reducing Resistance Transmission via Exosomes

    Directory of Open Access Journals (Sweden)

    Jun Zhang

    2015-07-01

    Full Text Available Background: Currently, exosomes that act as mediators of intercellular communication are being researched extensively. Our previous studies confirmed that these exosomes contain microRNAs (miRNAs that could alter chemo-susceptibility, which is partly attributed to the successful intercellular transfer of multidrug resistance (MDR-specific miRNAs. We also confirmed that β-elemene could influence MDR-related miRNA expression and regulate the expression of the target genes PTEN and Pgp, which may lead to the reversal of the chemoresistant breast cancer (BCA cells. We are the first to report these findings, and we propose the following logical hypothesis: β-elemene can mediate MDR-related miRNA expression in cells, thereby affecting the exosome contents, reducing chemoresistance transmission via exosomes, and reversing the drug resistance of breast cancer cells. Methods: MTT-cytotoxic, miRNA microarray, real-time quantitative PCR, Dual Luciferase Activity Assay, and Western blot analysis were performed to investigate the impact of β-elemene on the expression of chemoresistance specific miRNA and PTEN as well as Pgp in chemoresistant BCA exosomes. Results: Drug resistance can be reversed by β-elemene related to exosomes. There were 104 differentially expressed miRNAs in the exosomes of two chemoresistant BCA cells: adriacin (Adr - resistant MCF-7 cells (MCF-7/Adr and docetaxel (Doc - resistant MCF-7 cells (MCF-7/Doc that underwent treatment. Of these, 31 miRNAs were correlated with the constant changes in the MDR. The expression of miR-34a and miR-452 can lead to changes in the characteristics of two chemoresistant BCA exosomes: MCF-7/Adr exosomes (A/exo and MCF-7/Doc exosomes (D/exo. The PTEN expression affected by β-elemene was significantly increased, and the Pgp expression affected by β-elemene was significantly decreased in both cells and exosomes. β-elemene induced a significant increase in the apoptosis rate in both MCF-7/Doc and MCF-7

  2. Exosomes derived from human macrophages suppress endothelial cell migration by controlling integrin trafficking.

    Science.gov (United States)

    Lee, Hee Doo; Kim, Yeon Hyang; Kim, Doo-Sik

    2014-04-01

    Integrin trafficking, including internalization, recycling, and lysosomal degradation, is crucial for the regulation of cellular functions. Exosomes, nano-sized extracellular vesicles, are believed to play important roles in intercellular communications. This study demonstrates that exosomes released from human macrophages negatively regulate endothelial cell migration through control of integrin trafficking. Macrophage-derived exosomes promote internalization of integrin β1 in primary HUVECs. The internalized integrin β1 persistently accumulates in the perinuclear region and is not recycled back to the plasma membrane. Experimental results indicate that macrophage-derived exosomes stimulate trafficking of internalized integrin β1 to lysosomal compartments with a corresponding decrease in the integrin destined for recycling endosomes, resulting in proteolytic degradation of the integrin. Moreover, ubiquitination of HUVEC integrin β1 is enhanced by the exosomes, and exosome-mediated integrin degradation is blocked by bafilomycin A, a lysosomal degradation inhibitor. Macrophage-derived exosomes were also shown to effectively suppress collagen-induced activation of the mitogen-activated protein kinase/extracellular signal-regulated kinase signaling pathway and HUVEC migration, which are both dependent on integrin β1. These observations provide new insight into the functional significance of exosomes in the regulation of integrin trafficking.

  3. Cancer cell exosomes depend on cell-surface heparan sulfate proteoglycans for their internalization and functional activity.

    Science.gov (United States)

    Christianson, Helena C; Svensson, Katrin J; van Kuppevelt, Toin H; Li, Jin-Ping; Belting, Mattias

    2013-10-22

    Extracellular vesicle (EV)-mediated intercellular transfer of signaling proteins and nucleic acids has recently been implicated in the development of cancer and other pathological conditions; however, the mechanism of EV uptake and how this may be targeted remain as important questions. Here, we provide evidence that heparan sulfate (HS) proteoglycans (PGs; HSPGs) function as internalizing receptors of cancer cell-derived EVs with exosome-like characteristics. Internalized exosomes colocalized with cell-surface HSPGs of the syndecan and glypican type, and exosome uptake was specifically inhibited by free HS chains, whereas closely related chondroitin sulfate had no effect. By using several cell mutants, we provide genetic evidence of a receptor function of HSPG in exosome uptake, which was dependent on intact HS, specifically on the 2-O and N-sulfation groups. Further, enzymatic depletion of cell-surface HSPG or pharmacological inhibition of endogenous PG biosynthesis by xyloside significantly attenuated exosome uptake. We provide biochemical evidence that HSPGs are sorted to and associate with exosomes; however, exosome-associated HSPGs appear to have no direct role in exosome internalization. On a functional level, exosome-induced ERK1/2 signaling activation was attenuated in PG-deficient mutant cells as well as in WT cells treated with xyloside. Importantly, exosome-mediated stimulation of cancer cell migration was significantly reduced in PG-deficient mutant cells, or by treatment of WT cells with heparin or xyloside. We conclude that cancer cell-derived exosomes use HSPGs for their internalization and functional activity, which significantly extends the emerging role of HSPGs as key receptors of macromolecular cargo.

  4. Exosomes as potent cell-free peptide-based vaccine. II. Exosomes in CpG adjuvants efficiently prime naive Tc1 lymphocytes leading to tumor rejection.

    Science.gov (United States)

    Chaput, Nathalie; Schartz, Nöel E C; André, Fabrice; Taïeb, Julien; Novault, Sophie; Bonnaventure, Pierre; Aubert, Nathalie; Bernard, Jacky; Lemonnier, François; Merad, Miriam; Adema, Gosse; Adams, Malcolm; Ferrantini, Maria; Carpentier, Antoine F; Escudier, Bernard; Tursz, Thomas; Angevin, Eric; Zitvogel, Laurence

    2004-02-15

    Ideal vaccines should be stable, safe, molecularly defined, and out-of-shelf reagents efficient at triggering effector and memory Ag-specific T cell-based immune responses. Dendritic cell-derived exosomes could be considered as novel peptide-based vaccines because exosomes harbor a discrete set of proteins, bear functional MHC class I and II molecules that can be loaded with synthetic peptides of choice, and are stable reagents that were safely used in pioneering phase I studies. However, we showed in part I that exosomes are efficient to promote primary MHC class I-restricted effector CD8(+) T cell responses only when transferred onto mature DC in vivo. In this work, we bring evidence that among the clinically available reagents, Toll-like receptor 3 and 9 ligands are elective adjuvants capable of triggering efficient MHC-restricted CD8(+) T cell responses when combined to exosomes. Exosome immunogenicity across species allowed to verify the efficacy of good manufactory procedures-manufactured human exosomes admixed with CpG oligonucleotides in prophylactic and therapeutic settings of melanoma in HLA-A2 transgenic mice. CpG adjuvants appear to be ideal adjuvants for exosome-based cancer vaccines.

  5. Application of a Persistent Heparin Treatment Inhibits the Malignant Potential of Oral Squamous Carcinoma Cells Induced by Tumor Cell-Derived Exosomes.

    Science.gov (United States)

    Sento, Shinya; Sasabe, Eri; Yamamoto, Tetsuya

    2016-01-01

    Exosomes are 30-100 nm-sized membranous vesicles, secreted from a variety of cell types into their surrounding extracellular space. Various exosome components including lipids, proteins, and nucleic acids are transferred to recipient cells and affect their function and activity. Numerous studies have showed that tumor cell-derived exosomes play important roles in tumor growth and progression. However, the effect of exosomes released from oral squamous cell carcinoma (OSCC) into the tumor microenvironment remains unclear. In the present study, we isolated exosomes from OSCC cells and investigated the influence of OSCC cell-derived exosomes on the tumor cell behavior associated with tumor development. We demonstrated that OSCC cell-derived exosomes were taken up by OSCC cells themselves and significantly promoted proliferation, migration, and invasion through the activation of the PI3K/Akt, MAPK/ERK, and JNK-1/2 pathways in vitro. These effects of OSCC cell-derived exosomes were obviously attenuated by treatment with PI3K, ERK-1/2, and JNK-1/2 pharmacological inhibitors. Furthermore, the growth rate of tumor xenografts implanted into nude mice was promoted by treatment with OSCC cell-derived exosomes. The uptake of exosomes by OSCC cells and subsequent tumor progression was abrogated in the presence of heparin. Taken together, these data suggest that OSCC cell-derived exosomes might be a novel therapeutic target and the use of heparin to inhibit the uptake of OSCC-derived exosomes by OSCC cells may be useful for treatment.

  6. Isolation and Characterization of Exosomes Derived from Tumor Cells Genetically Expressing Model Antigen

    Institute of Scientific and Technical Information of China (English)

    修方明; 杨云山; 蔡志坚; 王建莉; 曹雪涛

    2004-01-01

    Tumor cell-derived exosomes have been proposed as non-cellular nanomeric vaccine which could induce potent antitumor immune response in mice. In order to develop the protocols to prepare tumor cell-derived exosomes for basic research and clinical trail, we isolated exosomes from ovalbumin (OVA)-expressing thymoma cells EG. 7-OVA by various preparation methods. We demonstrate the non-sedimentation method is simple, rapid, efficient with higher yield and purity of exosomes. EG. 7-OVA-derived exosomes are 40-100 nm in diameter sequestered by lipid bi-layer, and contain rich heat shock protein (HSP) and OVA. The result of the size distribution determination is consistent with the calculation by the visual microscopic inspection, with 90.4% particles at the range of 50-90 nm. Moreover, as a model antigen of the EG. 7 cells, OVA concentration in EG.7-derived exosomes can be regarded as a good quality control parameter. Therefore, we have established a platform to efficiently prepare exosomes for tumor immunotherapy.

  7. Exosome-based tumor antigens-adjuvant co-delivery utilizing genetically engineered tumor cell-derived exosomes with immunostimulatory CpG DNA.

    Science.gov (United States)

    Morishita, Masaki; Takahashi, Yuki; Matsumoto, Akihiro; Nishikawa, Makiya; Takakura, Yoshinobu

    2016-12-01

    For cancer immunotherapy via tumor antigen vaccination in combination with an adjuvant, major challenges include the identification of a particular tumor antigen and efficient delivery of the antigen as well as adjuvant to antigen-presenting cells. In this study, we proposed an efficient exosome-based tumor antigens-adjuvant co-delivery system using genetically engineered tumor cell-derived exosomes containing endogenous tumor antigens and immunostimulatory CpG DNA. Murine melanoma B16BL6 cells were transfected with a plasmid vector encoding a fusion streptavidin (SAV; a protein that binds to biotin with high affinity)-lactadherin (LA; an exosome-tropic protein) protein, yielding genetically engineered SAV-LA-expressing exosomes (SAV-exo). SAV-exo were combined with biotinylated CpG DNA to prepare CpG DNA-modified exosomes (CpG-SAV-exo). Fluorescent microscopic observation revealed the successful modification of exosomes with CpG DNA by SAV-biotin interaction. CpG-SAV-exo showed efficient and simultaneous delivery of exosomes with CpG DNA to murine dendritic DC2.4 cells in culture. Treatment with CpG-SAV-exo effectively activated DC2.4 cells and enhanced tumor antigen presentation capacity. Immunization with CpG-SAV-exo exhibited stronger in vivo antitumor effects in B16BL6 tumor-bearing mice than simple co-administration of exosomes and CpG DNA. Thus, genetically engineered CpG-SAV-exo is an effective exosome-based tumor antigens-adjuvant co-delivery system that will be useful for cancer immunotherapy.

  8. Exosomes released by chronic lymphocytic leukemia cells induce the transition of stromal cells into cancer-associated fibroblasts.

    Science.gov (United States)

    Paggetti, Jerome; Haderk, Franziska; Seiffert, Martina; Janji, Bassam; Distler, Ute; Ammerlaan, Wim; Kim, Yeoun Jin; Adam, Julien; Lichter, Peter; Solary, Eric; Berchem, Guy; Moussay, Etienne

    2015-08-27

    Exosomes derived from solid tumor cells are involved in immune suppression, angiogenesis, and metastasis, but the role of leukemia-derived exosomes has been less investigated. The pathogenesis of chronic lymphocytic leukemia (CLL) is stringently associated with a tumor-supportive microenvironment and a dysfunctional immune system. Here, we explore the role of CLL-derived exosomes in the cellular and molecular mechanisms by which malignant cells create this favorable surrounding. We show that CLL-derived exosomes are actively incorporated by endothelial and mesenchymal stem cells ex vivo and in vivo and that the transfer of exosomal protein and microRNA induces an inflammatory phenotype in the target cells, which resembles the phenotype of cancer-associated fibroblasts (CAFs). As a result, stromal cells show enhanced proliferation, migration, and secretion of inflammatory cytokines, contributing to a tumor-supportive microenvironment. Exosome uptake by endothelial cells increased angiogenesis ex vivo and in vivo, and coinjection of CLL-derived exosomes and CLL cells promoted tumor growth in immunodeficient mice. Finally, we detected α-smooth actin-positive stromal cells in lymph nodes of CLL patients. These findings demonstrate that CLL-derived exosomes actively promote disease progression by modulating several functions of surrounding stromal cells that acquire features of cancer-associated fibroblasts.

  9. Exosomes secreted by human placenta carry functional Fas ligand and TRAIL molecules and convey apoptosis in activated immune cells, suggesting exosome-mediated immune privilege of the fetus.

    Science.gov (United States)

    Stenqvist, Ann-Christin; Nagaeva, Olga; Baranov, Vladimir; Mincheva-Nilsson, Lucia

    2013-12-01

    Apoptosis is crucially important in mediating immune privilege of the fetus during pregnancy. We investigated the expression and in vitro apoptotic activity of two physiologically relevant death messengers, the TNF family members Fas ligand (FasL) and TRAIL in human early and term placentas. Both molecules were intracellularly expressed, confined to the late endosomal compartment of the syncytiotrophoblast, and tightly associated to the generation and secretion of placental exosomes. Using immunoelectron microscopy, we show that FasL and TRAIL are expressed on the limiting membrane of multivesicular bodies where, by membrane invagination, intraluminal microvesicles carrying membranal bioactive FasL and TRAIL are formed and released in the extracellular space as exosomes. Analyzing exosomes secreted from placental explant cultures, to our knowledge, we demonstrate for the first time that FasL and TRAIL are clustered on the exosomal membrane as oligomerized aggregates ready to form death-inducing signaling complex. Consistently, placental FasL- and TRAIL-carrying exosomes triggered apoptosis in Jurkat T cells and activated PBMC in a dose-dependent manner. Limiting the expression of functional FasL and TRAIL to exosomes comprise a dual benefit: 1) storage of exosomal FasL and TRAIL in multivesicular bodies is protected from proteolytic cleavage and 2) upon secretion, delivery of preformed membranal death molecules by exosomes rapidly triggers apoptosis. Our results suggest that bioactive FasL- and TRAIL-carrying exosomes, able to convey apoptosis, are secreted by the placenta and tie up the immunomodulatory and protective role of human placenta to its exosome-secreting ability.

  10. Human T-lymphotropic virus type 1-infected cells secrete exosomes that contain Tax protein.

    Science.gov (United States)

    Jaworski, Elizabeth; Narayanan, Aarthi; Van Duyne, Rachel; Shabbeer-Meyering, Shabana; Iordanskiy, Sergey; Saifuddin, Mohammed; Das, Ravi; Afonso, Philippe V; Sampey, Gavin C; Chung, Myung; Popratiloff, Anastas; Shrestha, Bindesh; Sehgal, Mohit; Jain, Pooja; Vertes, Akos; Mahieux, Renaud; Kashanchi, Fatah

    2014-08-08

    Human T-lymphotropic virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis. The HTLV-1 transactivator protein Tax controls many critical cellular pathways, including host cell DNA damage response mechanisms, cell cycle progression, and apoptosis. Extracellular vesicles called exosomes play critical roles during pathogenic viral infections as delivery vehicles for host and viral components, including proteins, mRNA, and microRNA. We hypothesized that exosomes derived from HTLV-1-infected cells contain unique host and viral proteins that may contribute to HTLV-1-induced pathogenesis. We found exosomes derived from infected cells to contain Tax protein and proinflammatory mediators as well as viral mRNA transcripts, including Tax, HBZ, and Env. Furthermore, we observed that exosomes released from HTLV-1-infected Tax-expressing cells contributed to enhanced survival of exosome-recipient cells when treated with Fas antibody. This survival was cFLIP-dependent, with Tax showing induction of NF-κB in exosome-recipient cells. Finally, IL-2-dependent CTLL-2 cells that received Tax-containing exosomes were protected from apoptosis through activation of AKT. Similar experiments with primary cultures showed protection and survival of peripheral blood mononuclear cells even in the absence of phytohemagglutinin/IL-2. Surviving cells contained more phosphorylated Rb, consistent with the role of Tax in regulation of the cell cycle. Collectively, these results suggest that exosomes may play an important role in extracellular delivery of functional HTLV-1 proteins and mRNA to recipient cells.

  11. Human T-lymphotropic Virus Type 1-infected Cells Secrete Exosomes That Contain Tax Protein*

    Science.gov (United States)

    Jaworski, Elizabeth; Narayanan, Aarthi; Van Duyne, Rachel; Shabbeer-Meyering, Shabana; Iordanskiy, Sergey; Saifuddin, Mohammed; Das, Ravi; Afonso, Philippe V.; Sampey, Gavin C.; Chung, Myung; Popratiloff, Anastas; Shrestha, Bindesh; Sehgal, Mohit; Jain, Pooja; Vertes, Akos; Mahieux, Renaud; Kashanchi, Fatah

    2014-01-01

    Human T-lymphotropic virus type 1 (HTLV-1) is the causative agent of adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis. The HTLV-1 transactivator protein Tax controls many critical cellular pathways, including host cell DNA damage response mechanisms, cell cycle progression, and apoptosis. Extracellular vesicles called exosomes play critical roles during pathogenic viral infections as delivery vehicles for host and viral components, including proteins, mRNA, and microRNA. We hypothesized that exosomes derived from HTLV-1-infected cells contain unique host and viral proteins that may contribute to HTLV-1-induced pathogenesis. We found exosomes derived from infected cells to contain Tax protein and proinflammatory mediators as well as viral mRNA transcripts, including Tax, HBZ, and Env. Furthermore, we observed that exosomes released from HTLV-1-infected Tax-expressing cells contributed to enhanced survival of exosome-recipient cells when treated with Fas antibody. This survival was cFLIP-dependent, with Tax showing induction of NF-κB in exosome-recipient cells. Finally, IL-2-dependent CTLL-2 cells that received Tax-containing exosomes were protected from apoptosis through activation of AKT. Similar experiments with primary cultures showed protection and survival of peripheral blood mononuclear cells even in the absence of phytohemagglutinin/IL-2. Surviving cells contained more phosphorylated Rb, consistent with the role of Tax in regulation of the cell cycle. Collectively, these results suggest that exosomes may play an important role in extracellular delivery of functional HTLV-1 proteins and mRNA to recipient cells. PMID:24939845

  12. Outsmart tumor exosomes to steal the cancer initiating cell its niche.

    Science.gov (United States)

    Thuma, Florian; Zöller, Margot

    2014-10-01

    Exosomes are small vesicles that derive from endosomes and are delivered by many cells, including tumor cells that are a particular rich source of exosomes. Exosomes are suggested to be the most potent intercellular communicators. Being recovered in all body fluids, they can communicate with neighboring as well as distant cells. The latter was first described for dendritic cell exosomes that can initiate T cell activation. However, tumor exosomes (TEX) may impede this crosstalk. Besides with hematopoietic cells, TEX communicate with the tumor cell itself, but also with host stroma cells and endothelial cells. This crosstalk received much attention as there is strong evidence that TEX account for angiogenesis and premetastatic niche formation, which may proceed directly via binding and uptake of TEX by cells in the premetastatic organ or indirectly via TEX being taken up by hematopoietic progenitors in the bone marrow (BM), which mature toward lineages with immunosuppressive features or are forced toward premature release from the BM and homing into premetastatic organs. Knowing these deleterious activities of TEX, it becomes demanding to search for modes of therapeutic interference. I here introduce our hypothesis that metastasis formation may be hampered by tailored exosomes that outsmart TEX. The essential prerequisites are an in depth knowledge on TEX binding, uptake, binding-initiated signal transduction and uptake-promoted target cell reprogramming.

  13. Role of exosomes released by dendritic cells and/or by tumor targets: Regulation of NK cell plasticity.

    Directory of Open Access Journals (Sweden)

    Katrin S. Reiners

    2014-03-01

    Full Text Available Exosomes are endosomal-derived nanovesicles released by normal and tumor cells, which transfer functionally active proteins, lipids and nucleic acids between cells. They are important mediators of intercellular communication and act on the adjacent stroma as well as in the periphery. Recently, exosomes have been recognized to play a pathophysiological role in various diseases such as cancer or infectious diseases. Tumor cell-derived exosomes (Tex have been shown to act as tumor promotors by educating non-malignant cells to provide a tumor supporting microenvironment, which helps to circumvent immune detection by the host and supports metastasis. However, Tex with anti-tumor, immune-activating properties were also described reflecting the complexity of exosomes.Here, we assess the role of extracellular microvesicles/exosomes as messengers affecting NK cell function in health and disease and discuss the molecular basis for the differential impact of exosomes on NK cell activity. The molecular composition/load of exosomes and the mechanisms regulating their release remain unclear and need to be further analyzed to facilitate the development of new treatment options targeting the exosomal machinery.

  14. Functional prostate-specific membrane antigen is enriched in exosomes from prostate cancer cells.

    Science.gov (United States)

    Liu, Tiancheng; Mendes, Desiree E; Berkman, Clifford E

    2014-03-01

    Developing simple and effective approaches to detect tumor markers will be critical for early diagnosis or prognostic evaluation of prostate cancer treatment. Prostate‑specific membrane antigen (PSMA) has been validated as an important tumor marker for prostate cancer progression including angiogenesis and metastasis. As a type II membrane protein, PSMA can be constitutively internalized from the cell surface into endosomes. Early endosomes can fuse with multivesicular bodies (MVB) to form and secrete exosomes (40-100 nm) into the extracellular environment. Herein, we tested whether some of the endosomal PSMA could be transferred to exosomes as an extracellular resource for PSMA. Using PSMA-positive LNCaP cells, the secreted exosomes were collected and isolated from the cultured media. The vesicular structures of exosomes were identified by electron microscopy, and exosomal marker protein CD9 and tumor susceptibility gene (TSG 101) were confirmed by western blot analysis. Our present data demonstrate that PSMA can be enriched in exosomes, exhibiting a higher content of glycosylation and partial proteolysis in comparison to cellular PSMA. An in vitro enzyme assay further confirmed that exosomal PSMA retains functional enzymatic activity. Therefore, our data may suggest a new role for PSMA in prostate cancer progression, and provide opportunities for developing non-invasive approaches for diagnosis or prognosis of prostate cancer.

  15. High LIN28A Expressing Ovarian Cancer Cells Secrete Exosomes That Induce Invasion and Migration in HEK293 Cells.

    Science.gov (United States)

    Enriquez, Vanessa A; Cleys, Ellane R; Da Silveira, Juliano C; Spillman, Monique A; Winger, Quinton A; Bouma, Gerrit J

    2015-01-01

    Epithelial ovarian cancer is the most aggressive and deadly form of ovarian cancer and is the most lethal gynecological malignancy worldwide; therefore, efforts to elucidate the molecular factors that lead to epithelial ovarian cancer are essential to better understand this disease. Recent studies reveal that tumor cells release cell-secreted vesicles called exosomes and these exosomes can transfer RNAs and miRNAs to distant sites, leading to cell transformation and tumor development. The RNA-binding protein LIN28 is a known marker of stem cells and when expressed in cancer, it is associated with poor tumor outcome. We hypothesized that high LIN28 expressing ovarian cancer cells secrete exosomes that can be taken up by nontumor cells and cause changes in gene expression and cell behavior associated with tumor development. IGROV1 cells were found to contain high LIN28A and secrete exosomes that were taken up by HEK293 cells. Moreover, exposure to these IGROV1 secreted exosomes led to significant increases in genes involved in Epithelial-to-Mesenchymal Transition (EMT), induced HEK293 cell invasion and migration. These changes were not observed with exosomes secreted by OV420 cells, which contain no detectable amounts of LIN28A or LIN28B. No evidence was found of LIN28A transfer from IGROV1 exosomes to HEK293 cells.

  16. Activated T cells recruit exosomes secreted by dendritic cells via LFA-1.

    NARCIS (Netherlands)

    Nolte-'t Hoen, E.N.; Buschow, S.I.; Anderton, S.M.; Stoorvogel, W.; Wauben, M.H.M.

    2009-01-01

    Dendritic cells (DCs) are known to secrete exosomes that transfer membrane proteins, like major histocompatibility complex class II, to other DCs. Intercellular transfer of membrane proteins is also observed during cognate interactions between DCs and CD4(+) T cells. The acquired proteins are functi

  17. Identification and Characterization of 293T Cell-Derived Exosomes by Profiling the Protein, mRNA and MicroRNA Components

    Science.gov (United States)

    Li, Dameng; Wang, Jifeng; Hou, Dongxia; Jiang, Xiaohong; Zhang, Junfeng; Wang, Jin; Zen, Ke; Yang, Fuquan; Zhang, Chen-Yu

    2016-01-01

    Cell-derived exosomes are leading candidates for in vivo drug delivery carriers. In particular, exosomes derived from 293T cells are used most frequently, although exosome dosing has varied greatly among studies. Considering their biological origin, it is crucial to characterize the molecular composition of exosomes if large doses are to be administered in clinical settings. In this study, we present the first comprehensive analysis of the protein, messenger RNA and microRNA profiles of 293T cell-derived exosomes; then, we characterized these data using Gene Ontology annotation and Kyoto Encyclopedia for Genes and Genomes pathway analysis. Our study will provide the basis for the selection of 293T cell-derived exosome drug delivery systems. Profiling the exosomal signatures of 293T cells will lead to a better understanding of 293T exosome biology and will aid in the identification of any harmful factors in exosomes that could cause adverse clinical effects. PMID:27649079

  18. Glucose Starvation in Cardiomyocytes Enhances Exosome Secretion and Promotes Angiogenesis in Endothelial Cells.

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    Nahuel A Garcia

    Full Text Available Cardiomyocytes (CMs and endothelial cells (ECs have an intimate anatomical relationship that is essential for maintaining normal development and function in the heart. Little is known about the mechanisms that regulate cardiac and endothelial crosstalk, particularly in situations of acute stress when local active processes are required to regulate endothelial function. We examined whether CM-derived exosomes could modulate endothelial function. Under conditions of glucose deprivation, immortalized H9C2 cardiomyocytes increase their secretion of exosomes. CM-derived exosomes are loaded with a broad repertoire of miRNA and proteins in a glucose availability-dependent manner. Gene Ontology (GO analysis of exosome cargo molecules identified an enrichment of biological process that could alter EC activity. We observed that addition of CM-derived exosomes to ECs induced changes in transcriptional activity of pro-angiogenic genes. Finally, we demonstrated that incubation of H9C2-derived exosomes with ECs induced proliferation and angiogenesis in the latter. Thus, exosome-mediated communication between CM and EC establishes a functional relationship that could have potential implications for the induction of local neovascularization during acute situations such as cardiac injury.

  19. Characterization of RNA in exosomes secreted by human breast cancer cell lines using next-generation sequencing

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    Piroon Jenjaroenpun

    2013-11-01

    Full Text Available Exosomes are nanosized (30–100 nm membrane vesicles secreted by most cell types. Exosomes have been found to contain various RNA species including miRNA, mRNA and long non-protein coding RNAs. A number of cancer cells produce elevated levels of exosomes. Because exosomes have been isolated from most body fluids they may provide a source for non-invasive cancer diagnostics. Transcriptome profiling that uses deep-sequencing technologies (RNA-Seq offers enormous amount of data that can be used for biomarkers discovery, however, in case of exosomes this approach was applied only for the analysis of small RNAs. In this study, we utilized RNA-Seq technology to analyze RNAs present in microvesicles secreted by human breast cancer cell lines.Exosomes were isolated from the media conditioned by two human breast cancer cell lines, MDA-MB-231 and MDA-MB-436. Exosomal RNA was profiled using the Ion Torrent semiconductor chip-based technology. Exosomes were found to contain various classes of RNA with the major class represented by fragmented ribosomal RNA (rRNA, in particular 28S and 18S rRNA subunits. Analysis of exosomal RNA content revealed that it reflects RNA content of the donor cells. Although exosomes produced by the two cancer cell lines shared most of the RNA species, there was a number of non-coding transcripts unique to MDA-MB-231 and MDA-MB-436 cells. This suggests that RNA analysis might distinguish exosomes produced by low metastatic breast cancer cell line (MDA-MB-436 from that produced by highly metastatic breast cancer cell line (MDA-MB-231. The analysis of gene ontologies (GOs associated with the most abundant transcripts present in exosomes revealed significant enrichment in genes encoding proteins involved in translation and rRNA and ncRNA processing. These GO terms indicate most expressed genes for both, cellular and exosomal RNA.For the first time, using RNA-seq, we examined the transcriptomes of exosomes secreted by human breast

  20. Exosomes in Cancer Diagnostics

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    Young Hwa Soung

    2017-01-01

    Full Text Available Exosomes are endosome derived extracellular vesicles of 30–120 nm size ranges. Exosomes have been identified as mediators of cell-to-cell communication by transferring bioactive molecules such as nucleic acids, proteins and lipids into recipient cells. While exosomes are secreted by multiple cell types, cancer derived exosomes not only influence the invasive potentials of proximally located cells, but also affect distantly located tissues. Based on their ability to alter tumor microenvironment by regulating immunity, angiogenesis and metastasis, there has been growing interest in defining the clinical relevance of exosomes in cancers. In particular, exosomes are valuable sources for biomarkers due to selective cargo loading and resemblance to their parental cells. In this review, we summarize the recent findings to utilize exosomes as cancer biomarkers for early detection, diagnosis and therapy selection.

  1. Exosomes/miRNAs as mediating cell-based therapy of stroke

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    Hongqi eXin

    2014-11-01

    Full Text Available Cell-based therapy, e.g., multipotent mesenchymal stromal cell (MSC treatment, shows promise for the treatment of various diseases. The strong paracrine capacity of these cells and not their differentiation capacity, is the principal mechanism of therapeutic action. MSCs robustly release exosomes, membrane vesicles (~30-100nm originally derived in endosomes as intraluminal vesicles, which contain various molecular constituents including proteins and RNAs from maternal cells. Contained among these constituents, are small non-coding RNA molecules, microRNAs (miRNAs, which play a key role in mediating biological function due to their prominent role in gene regulation. The release as well as the content of the MSC generated exosomes are modified by environmental conditions. Via exosomes, MSCs transfer their therapeutic factors, especially miRNAs, to recipient cells, and therein alter gene expression and thereby promote therapeutic response. The present review focuses on the paracrine mechanism of MSC exosomes, and the regulation and transfer of exosome content, especially the packaging and transfer of miRNAs which enhance tissue repair and functional recovery. Perspectives on the developing role of MSC mediated transfer of exosomes as a therapeutic approach will also be discussed.

  2. Exosomes from human mesenchymal stem cells conduct aerobic metabolism in term and preterm newborn infants.

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    Panfoli, Isabella; Ravera, Silvia; Podestà, Marina; Cossu, Claudia; Santucci, Laura; Bartolucci, Martina; Bruschi, Maurizio; Calzia, Daniela; Sabatini, Federica; Bruschettini, Matteo; Ramenghi, Luca Antonio; Romantsik, Olga; Marimpietri, Danilo; Pistoia, Vito; Ghiggeri, Gianmarco; Frassoni, Francesco; Candiano, Giovanni

    2016-04-01

    Exosomes are secreted nanovesicles that are able to transfer RNA and proteins to target cells. The emerging role of mesenchymal stem cell (MSC) exosomes as promoters of aerobic ATP synthesis restoration in damaged cells, prompted us to assess whether they contain an extramitochondrial aerobic respiration capacity. Exosomes were isolated from culture medium of human MSCs from umbilical cord of ≥37-wk-old newborns or between 28- to 30-wk-old newborns (i.e.,term or preterm infants). Characterization of samples was conducted by cytofluorometry. Oxidative phosphorylation capacity was assessed by Western blot analysis, oximetry, and luminometric and fluorometric analyses. MSC exosomes express functional respiratory complexes I, IV, and V, consuming oxygen. ATP synthesis was only detectable in exosomes from term newborns, suggestive of a specific mechanism that is not completed at an early gestational age. Activities are outward facing and comparable to those detected in mitochondria isolated from term MSCs. MSC exosomes display an unsuspected aerobic respiratory ability independent of whole mitochondria. This may be relevant for their ability to rescue cell bioenergetics. The differential oxidative metabolism of pretermvs.term exosomes sheds new light on the preterm newborn's clinical vulnerability. A reduced ability to repair damaged tissue and an increased capability to cope with anoxic environment for preterm infants can be envisaged.-Panfoli, I., Ravera, S., Podestà, M., Cossu, C., Santucci, L., Bartolucci, M., Bruschi, M., Calzia, D., Sabatini, F., Bruschettini, M., Ramenghi, L. A., Romantsik, O., Marimpietri, D., Pistoia, V., Ghiggeri, G., Frassoni, F., Candiano, G. Exosomes from human mesenchymal stem cells conduct aerobic metabolism in term and preterm newborn infants.

  3. Schwann cell-derived exosomes enhance axonal regeneration in the peripheral nervous system.

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    Lopez-Verrilli, María Alejandra; Picou, Frederic; Court, Felipe A

    2013-11-01

    Axonal regeneration in the peripheral nervous system is greatly supported by Schwann cells (SCs). After nerve injury, SCs dedifferentiate to a progenitor-like state and efficiently guide axons to their original target tissues. Contact and soluble factors participate in the crosstalk between SCs and axons during axonal regeneration. Here we show that dedifferentiated SCs secrete nano-vesicles known as exosomes which are specifically internalized by axons. Surprisingly, SC-derived exosomes markedly increase axonal regeneration in vitro and enhance regeneration after sciatic nerve injury in vivo. Exosomes shift the growth cone morphology to a pro-regenerating phenotype and decrease the activity of the GTPase RhoA, involved in growth cone collapse and axon retraction. Altogether, our work identifies a novel mechanism by which SCs communicate with neighboring axons during regenerative processes. We propose that SC exosomes represent an important mechanism by which these cells locally support axonal maintenance and regeneration after nerve damage.

  4. Ultrastructural Evidence of Exosome Secretion by Progenitor Cells in Adult Mouse Myocardium and Adult Human Cardiospheres

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    Lucio Barile

    2012-01-01

    Full Text Available The demonstration of beneficial effects of cell therapy despite the persistence of only few transplanted cells in vivo suggests secreted factors may be the active component of this treatment. This so-called paracrine hypothesis is supported by observations that culture media conditioned by progenitor cells contain growth factors that mediate proangiogenic and cytoprotective effects. Cardiac progenitor cells in semi-suspension culture form spherical clusters (cardiospheres that deliver paracrine signals to neighboring cells. A key component of paracrine secretion is exosomes, membrane vesicles that are stored intracellularly in endosomal compartments and are secreted when these structures fuse with the cell plasma membrane. Exosomes have been identified as the active component of proangiogenic effects of bone marrow CD34+ stem cells in mice and the regenerative effects of embryonic mesenchymal stem cells in infarcted hearts in pigs and mice. Here, we provide electron microscopic evidence of exosome secretion by progenitor cells in mouse myocardium and human cardiospheres. Exosomes are emerging as an attractive vector of paracrine signals delivered by progenitor cells. They can be stored as an “off-the-shelf” product. As such, exosomes have the potential for circumventing many of the limitations of viable cells for therapeutic applications in regenerative medicine.

  5. Surface Glycoproteins of Exosomes Shed by Myeloid-Derived Suppressor Cells Contribute to Function.

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    Chauhan, Sitara; Danielson, Steven; Clements, Virginia; Edwards, Nathan; Ostrand-Rosenberg, Suzanne; Fenselau, Catherine

    2017-01-06

    In this report, we use a proteomic strategy to identify glycoproteins on the surface of exosomes derived from myeloid-derived suppressor cells (MDSCs), and then test if selected glycoproteins contribute to exosome-mediated chemotaxis and migration of MDSCs. We report successful modification of a surface chemistry method for use with exosomes and identify 21 surface N-glycoproteins on exosomes released by mouse mammary carcinoma-induced MDSCs. These glycoprotein identities and functionalities are compared with 93 N-linked glycoproteins identified on the surface of the parental cells. As with the lysate proteomes examined previously, the exosome surface N-glycoproteins are primarily a subset of the glycoproteins on the surface of the suppressor cells that released them, with related functions and related potential as therapeutic targets. The "don't eat me" molecule CD47 and its binding partners thrombospondin-1 (TSP1) and signal regulatory protein α (SIRPα) were among the surface N-glycoproteins detected. Functional bioassays using antibodies to these three molecules demonstrated that CD47, TSP1, and to a lesser extent SIRPα facilitate exosome-mediated MDSC chemotaxis and migration.

  6. Exosomes: Mechanisms of Uptake

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    Kelly J. McKelvey

    2015-07-01

    Full Text Available Exosomes are 30–100 nm microvesicles which contain complex cellular signals of RNA, protein and lipids. Because of this, exosomes are implicated as having limitless therapeutic potential for the treatment of cancer, pregnancy complications, infections, and autoimmune diseases. To date we know a considerable amount about exosome biogenesis and secretion, but there is a paucity of data regarding the uptake of exosomes by immune and non- immune cell types (e.g., cancer cells and the internal signalling pathways by which these exosomes elicit a cellular response. Answering these questions is of para‐ mount importance.

  7. Proteomic Analysis of Exosomes and Exosome-Free Conditioned Media From Human Osteosarcoma Cell Lines Reveals Secretion of Proteins Related to Tumor Progression.

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    Jerez, Sofía; Araya, Héctor; Thaler, Roman; Charlesworth, M Cristine; López-Solís, Remigio; Kalergis, Alexis M; Céspedes, Pablo F; Dudakovic, Amel; Stein, Gary S; van Wijnen, Andre J; Galindo, Mario

    2017-02-01

    Osteosarcomas are the most prevalent bone tumors in pediatric patients, but can also occur later in life. Bone tumors have the potential to metastasize to lung and occasionally other vital organs. To understand how osteosarcoma cells interact with their micro-environment to support bone tumor progression and metastasis, we analyzed secreted proteins and exosomes from three human osteosarcoma cell lines. Exosome isolation was validated by transmission electron microscopy (TEM) and immuno-blotting for characteristic biomarkers (CD63, CD9, and CD81). Exosomal and soluble proteins (less than 100 kDa) were identified by mass spectrometry analysis using nanoLC-MS/MS and classified by functional gene ontology clustering. We identified a secretome set of >3,000 proteins for both fractions, and detected proteins that are either common or unique among the three osteosarcoma cell lines. Protein ontology comparison of proteomes from exosomes and exosome-free fractions revealed differences in the enrichment of functional categories associated with different biological processes, including those related to tumor progression (i.e., angiogenesis, cell adhesion, and cell migration). The secretome characteristics of osteosarcoma cells are consistent with the pathological properties of tumor cells with metastatic potential. J. Cell. Biochem. 118: 351-360, 2017. © 2016 Wiley Periodicals, Inc.

  8. Dynamics of exosome internalization and trafficking.

    Science.gov (United States)

    Tian, Tian; Zhu, Yan-Liang; Hu, Fei-Hu; Wang, Yuan-Yuan; Huang, Ning-Ping; Xiao, Zhong-Dang

    2013-07-01

    Cells release exosomes into extracellular medium. Although the important roles of exosomes in many physiological and pathological processes are being revealed, the mechanism of exosome-cell interaction remains unclear. In this article, employing real-time fluorescence microscopy, the motion of exosomes on the plasma membrane or in the cytoplasm of recipient PC12 cells was observed directly. In addition, several motion modes of exosomes were revealed by single particle tracking (SPT). The changes between motion modes were also detected, presenting the dynamic courses of exosome attachment onto plasma membrane and exosome uptake. Octadecyl rhodamine B chloride (R18) was found to be useful to distinguish endocytosis from fusion during exosome uptake. Colocalization with organelle markers showed exosomes were sorted to acidic vesicles after internalization. The results provide new sight into the exosome-cell interaction mode and the intercellular trafficking of exosomes. This study will help to understand the roles of exosomes at cell level.

  9. Exosomes from differentiating human skeletal muscle cells trigger myogenesis of stem cells and provide biochemical cues for skeletal muscle regeneration.

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    Choi, Ji Suk; Yoon, Hwa In; Lee, Kyoung Soo; Choi, Young Chan; Yang, Seong Hyun; Kim, In-San; Cho, Yong Woo

    2016-01-28

    Exosomes released from skeletal muscle cells play important roles in myogenesis and muscle development via the transfer of specific signal molecules. In this study, we investigated whether exosomes secreted during myotube differentiation from human skeletal myoblasts (HSkM) could induce a cellular response from human adipose-derived stem cells (HASCs) and enhance muscle regeneration in a muscle laceration mouse model. The exosomes contained various signal molecules including myogenic growth factors related to muscle development, such as insulin-like growth factors (IGFs), hepatocyte growth factor (HGF), fibroblast growth factor-2 (FGF2), and platelet-derived growth factor-AA (PDGF-AA). Interestingly, exosome-treated HASCs fused with neighboring cells at early time points and exhibited a myotube-like phenotype with increased expression of myogenic proteins (myosin heavy chain and desmin). On day 21, mRNAs of terminal myogenic genes were also up-regulated in exosome-treated HASCs. Moreover, in vivo studies demonstrated that exosomes from differentiating HSkM reduced the fibrotic area and increased the number of regenerated myofibers in the injury site, resulting in significant improvement of skeletal muscle regeneration. Our findings suggest that exosomes act as a biochemical cue directing stem cell differentiation and provide a cell-free therapeutic approach for muscle regeneration.

  10. Do Neural Cells Communicate with Endothelial Cells via Secretory Exosomes and Microvesicles?

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    Neil R. Smalheiser

    2009-01-01

    Full Text Available Neurons, glial, cells, and brain tumor cells tissues release small vesicles (secretory exosomes and microvesicles, which may represent a novel mechanism by which neuronal activity could influence angiogenesis within the embryonic and mature brain. If CNS-derived vesicles can enter the bloodstream as well, they may communicate with endothelial cells in the peripheral circulation and with cells concerned with immune surveillance.

  11. Exosomes from hypoxic endothelial cells have increased collagen crosslinking activity through up-regulation of lysyl oxidase-like 2.

    Science.gov (United States)

    de Jong, Olivier G; van Balkom, Bas W M; Gremmels, Hendrik; Verhaar, Marianne C

    2016-02-01

    Exosomes are important mediators of intercellular communication. Additionally, they contain a variety of components capable of interacting with the extracellular matrix (ECM), including integrins, matrix metalloproteinases and members of the immunoglobin superfamily. Despite these observations, research on exosome-ECM interactions is limited. Here, we investigate whether the exosome-associated lysyl oxidase family member lysyl oxidase-like 2 (LOXL2) is involved in ECM remodelling. We found that LOXL2 is present on the exterior of endothelial cell (EC)-derived exosomes, placing it in direct vicinity of the ECM. It is up-regulated twofold in EC-derived exosomes cultured under hypoxic conditions. Intact exosomes from hypoxic EC and LOXL2 overexpressing EC show increased activity in a fluorometric lysyl oxidase enzymatic activity assay as well as in a collagen gel contraction assay. Concordantly, knockdown of LOXL2 in exosome-producing EC in both normal and hypoxic conditions reduces activity of exosomes in both assays. Our findings show for the first time that ECM crosslinking by EC-derived exosomes is mediated by LOXL2 under the regulation of hypoxia, and implicate a role for exosomes in hypoxia-regulated focal ECM remodelling, a key process in both fibrosis and wound healing.

  12. Dendritic Cell-Derived Exosomes Stimulate Stronger CD8+ CTL Responses and Antitumor Immunity than Tumor Cell-Derived Exosomes

    Institute of Scientific and Technical Information of China (English)

    Siguo Hao; Ou Bai; Jinying Yuan; Mabood Qureshi; Jim Xiang

    2006-01-01

    Exosomes (EXO) derived from dendritic cells (DC) and tumor cells have been used to stimulate antitumor immune responses in animal models and in clinical trials. However, there has been no side-by-side comparison of the stimulatory efficiency of the antitumor immune responses induced by these two commonly used EXO vaccines. In this study, we selected to study the phenotype characteristics of EXO derived from a transfected EG7 tumor cells expressing ovalbumin (OVA) and OVA-pulsed DC by flow cytometry. We compared the stimulatory effect in induction of OVA-specific immune responses between these two types of EXO. We found that OVA protein-pulsed DCovA-derived EXO (EXODC) can more efficiently stimulate naive OVA-specific CD8+ T cell proliferation and differentiation into cytotoxic T lymphocytes in vivo, and induce more efficient antitumor immunity than EG7 tumor cell-derived EXO (EXOEG7). In addition, we elucidated the important role of the host DC in EXO vaccines that the stimulatory effect of EXO is delivered to T cell responses by the host DC. Therefore, DC-derived EXO may represent a more effective EXO-based vaccine in induction of antitumor immunity.

  13. Urinary Exosomes

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    Irena Dimov

    2009-01-01

    Full Text Available Exosomes are nanovesicles of endocytic origin that are secreted into the extracellular space or body fluids when a multivesicular body (MVB fuses with the cell membrane. Interest in exosomes intensified after their description in antigen-presenting cells and the observation that they can significantly moderate immune responses in vivo. In the past few years, several groups have reported on the secretion of exosomes by almost all cell types in an organism. In addition to a common set of membrane and cytosolic molecules, exosomes harbor unique subsets of proteins, reflecting their cellular source. Major research efforts were put into their surprisingly various biological functions and in translating knowledge into clinical practice. Urine provides an exciting noninvasive alternative to blood or tissue samples as a potential source of disease biomarkers. Urinary exosomes (UE became the subject of serious studies just a few years ago. A recent large-scale proteomics-based study of normal UE revealed a myriad of proteins, including disease-related gene products. Thus, UE have valuable potential as a source of biomarkers for early detection of various types of diseases, monitoring the disease evolution and/or response to therapy. As a relatively new field of research, it still faces many challenges, but UE have already shown some straightforward potential.

  14. Quantitative and qualitative analysis of small RNAs in human endothelial cells and exosomes provides insights into localized RNA processing, degradation and sorting

    NARCIS (Netherlands)

    van Balkom, Bas W M; Eisele, Almut S; Pegtel, D Michiel; Bervoets, Sander; Verhaar, Marianne C

    2015-01-01

    Exosomes are small vesicles that mediate cell-cell communication. They contain proteins, lipids and RNA, and evidence is accumulating that these molecules are specifically sorted for release via exosomes. We recently showed that endothelial-cell-produced exosomes promote angiogenesis in vivo in a sm

  15. Exosomes secreted from human colon cancer cells influence the adhesion of neighboring metastatic cells: Role of microRNA-210

    Science.gov (United States)

    Bigagli, Elisabetta; Luceri, Cristina; Guasti, Daniele; Cinci, Lorenzo

    2016-01-01

    ABSTRACT Cancer-secreted exosomes influence tumor microenvironment and support cancer growth and metastasis. MiR-210 is frequently up-regulated in colorectal cancer tissues and correlates with metastatic disease. We investigated whether exosomes are actively released by HCT-8 colon cancer cells, the role of exosomal miR-210 in the cross-talk between primary cancer cells and neighboring metastatic cells and its contribution in regulating epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET). After 7 d of culture, a subpopulation of viable HCT-8 cells detached the monolayer and started to grow in suspension, suggesting anoikis resistance and a metastatic potential. The expression of key proteins of EMT revealed that these cells were E-cadherin negative and vimentin positive further confirming their metastatic phenotype and the acquisition of anoikis resistance. Metastatic cells, in the presence of adherently growing HCT-8, continued to grow in suspension whereas only if seeded in cell-free wells, were able to adhere again and to form E-cadherin positive and vimentin negative new colonies, suggesting the occurrence of MET. The chemosensitivity to 5 fluorouracil and to FOLFOX-like treatment of metastatic cells was significantly diminished compared to adherent HCT-8 cells. Of note, adherent new colonies undergoing MET, were insensitive to both chemotherapeutic strategies. Electron microscopy analysis demonstrated that adherently growing HCT-8, actually secreted exosomes and that exosomes in turn were taken up by metastatic cells. When exosomes secreted by adherently growing HCT-8 were administered to metastatic cells, MET was significantly inhibited. miR-210 was significantly upregulated in exosomes compared to its intracellular levels in adherently growing HCT-8 cells and correlated to anoikis resistance and EMT markers. Exosomes containing miR-210 might be considered as EMT promoting signals that preserve the local cancer

  16. [Establishment and identification of the near-infrared fluorescence labeled exosomes in breast cancer cell lines].

    Science.gov (United States)

    Li, Taiming; Lan, Wenjun; Huang, Can; Zhang, Chun; Liu, Xiaomei

    2016-05-01

    Exosomes, a population of extracellular membrane vesicles of 30-100 nm in diameter, play important roles in cell biological functions, intercellular signal transduction and especially in cancer diagnosis and therapy. To better apply exosomes in mechanistic study of breast cancer signal transduction, we constructed recombinant eukaryotic expression vector expressing the near-infrared fluorescence protein and CD63 fusion protein through cloning iRFP682 gene and exosomal marker protein CD63 gene into plasmid containing the ITR of AAV. The constructed plasmids were co-transfected with helper plasmid in AAV-293 cell lines and were packaged into rAAV. After titer measurement, the recombinant plasmids were transfected into breast cancer cell lines. The cell lines that stably expressing near-infrared fluorescence protein were selected by fluorescence. Through isolation, purification and identification, we finally obtained a new biomarker: iRFP682 labeled exosomes secreted by breast cancer cell lines, which could be used in further studies of the distribution and signal transduction of exosomes in breast cancer microenvironment.

  17. Exosomes from Human Dental Pulp Stem Cells Suppress Carrageenan-Induced Acute Inflammation in Mice.

    Science.gov (United States)

    Pivoraitė, Ugnė; Jarmalavičiūtė, Akvilė; Tunaitis, Virginijus; Ramanauskaitė, Giedrė; Vaitkuvienė, Aida; Kašėta, Vytautas; Biziulevičienė, Genė; Venalis, Algirdas; Pivoriūnas, Augustas

    2015-10-01

    The primary goal of this study was to examine the effects of human dental pulp stem cell-derived exosomes on the carrageenan-induced acute inflammation in mice. Exosomes were purified by differential ultracentrifugation from the supernatants of stem cells derived from the dental pulp of human exfoliated deciduous teeth (SHEDs) cultivated in serum-free medium. At 1 h post-carrageenan injection, exosomes derived from supernatants of 2 × 10(6) SHEDs were administered by intraplantar injection to BALB/c mice; 30 mg/kg of prednisolone and phosphate-buffered saline (PBS) were used as positive and negative controls, respectively. Edema was measured at 6, 24, and 48 h after carrageenan injection. For the in vivo imaging experiments, AngioSPARK750, Cat B 750 FAST, and MMPSense 750 FAST were administered into the mouse tail vein 2 h post-carrageenan injection. Fluorescence images were acquired at 6, 24, and 48 h after edema induction by IVIS Spectrum in vivo imaging system. Exosomes significantly reduced the carrageenan-induced edema at all the time points studied (by 39.5, 41.6, and 25.6% at 6, 24, and 48 h after injection, respectively), to similar levels seen with the positive control (prednisolone). In vivo imaging experiments revealed that, both exosomes and prednisolone suppress activities of cathepsin B and matrix metalloproteinases (MMPs) at the site of carrageenan-induced acute inflammation, showing more prominent effects of prednisolone at the early stages, while exosomes exerted their suppressive effects gradually and at later time points. Our study demonstrates for the first time that exosomes derived from human dental pulp stem cells suppress carrageenan-induced acute inflammation in mice.

  18. Investigation of Content, Stoichiometry and Transfer of miRNA from Human Neural Stem Cell Line Derived Exosomes.

    Science.gov (United States)

    Stevanato, Lara; Thanabalasundaram, Lavaniya; Vysokov, Nickolai; Sinden, John D

    2016-01-01

    Exosomes are small (30-100 nm) membrane vesicles secreted by a variety of cell types and only recently have emerged as a new avenue for cell-to-cell communication. They are natural shuttles of RNA and protein cargo, making them attractive as potential therapeutic delivery vehicles. MicroRNAs (miRNAs) are short non-coding RNAs which regulate biological processes and can be found in exosomes. Here we characterized the miRNA contents of exosomes derived from human neural stem cells (hNSCs). Our investigated hNSC line is a clonal, conditionally immortalized cell line, compliant with good manufacturing practice (GMP), and in clinical trials for stroke and critical limb ischemia in the UK (clinicaltrials.gov: NCT01151124, NCT02117635, and NCT01916369). By using next generation sequencing (NGS) technology we identified the presence of a variety of miRNAs in both exosomal and cellular preparations. Many of these miRNAs were enriched in exosomes indicating that cells specifically sort them for extracellular release. Although exosomes have been proven to contain miRNAs, the copy number quantification per exosome of a given miRNA remains unclear. Herein we quantified by real-time PCR a highly shuttled exosomal miRNA subtype (hsa-miR-1246) in order to assess its stoichiometry per exosome. Furthermore, we utilized an in vitro system to confirm its functional transfer by measuring the reduction in luciferase expression using a 3' untranslated region dual luciferase reporter assay. In summary, NGS analysis allowed the identification of a unique set of hNSC derived exosomal miRNAs. Stoichiometry and functional transfer analysis of one of the most abundant identified miRNA, hsa-miR-1246, were measured to support biological relevance of exosomal miRNA delivery.

  19. Investigation of Content, Stoichiometry and Transfer of miRNA from Human Neural Stem Cell Line Derived Exosomes.

    Directory of Open Access Journals (Sweden)

    Lara Stevanato

    Full Text Available Exosomes are small (30-100 nm membrane vesicles secreted by a variety of cell types and only recently have emerged as a new avenue for cell-to-cell communication. They are natural shuttles of RNA and protein cargo, making them attractive as potential therapeutic delivery vehicles. MicroRNAs (miRNAs are short non-coding RNAs which regulate biological processes and can be found in exosomes. Here we characterized the miRNA contents of exosomes derived from human neural stem cells (hNSCs. Our investigated hNSC line is a clonal, conditionally immortalized cell line, compliant with good manufacturing practice (GMP, and in clinical trials for stroke and critical limb ischemia in the UK (clinicaltrials.gov: NCT01151124, NCT02117635, and NCT01916369. By using next generation sequencing (NGS technology we identified the presence of a variety of miRNAs in both exosomal and cellular preparations. Many of these miRNAs were enriched in exosomes indicating that cells specifically sort them for extracellular release. Although exosomes have been proven to contain miRNAs, the copy number quantification per exosome of a given miRNA remains unclear. Herein we quantified by real-time PCR a highly shuttled exosomal miRNA subtype (hsa-miR-1246 in order to assess its stoichiometry per exosome. Furthermore, we utilized an in vitro system to confirm its functional transfer by measuring the reduction in luciferase expression using a 3' untranslated region dual luciferase reporter assay. In summary, NGS analysis allowed the identification of a unique set of hNSC derived exosomal miRNAs. Stoichiometry and functional transfer analysis of one of the most abundant identified miRNA, hsa-miR-1246, were measured to support biological relevance of exosomal miRNA delivery.

  20. Exosomal signaling during hypoxia mediates microvascular endothelial cell migration and vasculogenesis.

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    Carlos Salomon

    Full Text Available Vasculogenesis and angiogenesis are critical processes in fetal circulation and placental vasculature development. Placental mesenchymal stem cells (pMSC are known to release paracrine factors (some of which are contained within exosomes that promote angiogenesis and cell migration. The aims of this study were: to determine the effects of oxygen tension on the release of exosomes from pMSC; and to establish the effects of pMSC-derived exosomes on the migration and angiogenic tube formation of placental microvascular endothelial cells (hPMEC. pMSC were isolated from placental villi (8-12 weeks of gestation, n = 6 and cultured under an atmosphere of 1%, 3% or 8% O2. Cell-conditioned media were collected and exosomes (exo-pMSC isolated by differential and buoyant density centrifugation. The dose effect (5-20 µg exosomal protein/ml of pMSC-derived exosomes on hPMEC migration and tube formation were established using a real-time, live-cell imaging system (Incucyte™. The exosome pellet was resuspended in PBS and protein content was established by mass spectrometry (MS. Protein function and canonical pathways were identified using the PANTHER program and Ingenuity Pathway Analysis, respectively. Exo-pMSC were identified, by electron microscopy, as spherical vesicles, with a typical cup-shape and diameters around of 100 nm and positive for exosome markers: CD63, CD9 and CD81. Under hypoxic conditions (1% and 3% O2 exo-pMSC released increased by 3.3 and 6.7 folds, respectively, when compared to the controls (8% O2; p<0.01. Exo-pMSC increased hPMEC migration by 1.6 fold compared to the control (p<0.05 and increased hPMEC tube formation by 7.2 fold (p<0.05. MS analysis identified 390 different proteins involved in cytoskeleton organization, development, immunomodulatory, and cell-to-cell communication. The data obtained support the hypothesis that pMSC-derived exosomes may contribute to placental vascular adaptation to low oxygen tension under both

  1. The bone marrow microenvironment enhances multiple myeloma progression by exosome-mediated activation of myeloid-derived suppressor cells.

    Science.gov (United States)

    Wang, Jinheng; De Veirman, Kim; De Beule, Nathan; Maes, Ken; De Bruyne, Elke; Van Valckenborgh, Els; Vanderkerken, Karin; Menu, Eline

    2015-12-22

    Exosomes, extracellular nanovesicles secreted by various cell types, modulate the bone marrow (BM) microenvironment by regulating angiogenesis, cytokine release, immune response, inflammation, and metastasis. Interactions between bone marrow stromal cells (BMSCs) and multiple myeloma (MM) cells play crucial roles in MM development. We previously reported that BMSC-derived exosomes directly promote MM cell growth, whereas the other possible mechanisms for supporting MM progression by these exosomes are still not clear. Here, we investigated the effect of BMSC-derived exosomes on the MM BM cells with specific emphasis on myeloid-derived suppressor cells (MDSCs). BMSC-derived exosomes were able to be taken up by MM MDSCs and induced their expansion in vitro. Moreover, these exosomes directly induced the survival of MDSCs through activating STAT3 and STAT1 pathways and increasing the anti-apoptotic proteins Bcl-xL and Mcl-1. Inhibition of these pathways blocked the enhancement of MDSC survival. Furthermore, these exosomes increased the nitric oxide release from MM MDSCs and enhanced their suppressive activity on T cells. Taken together, our results demonstrate that BMSC-derived exosomes activate MDSCs in the BM through STAT3 and STAT1 pathways, leading to increased immunosuppression which favors MM progression.

  2. Cells release subpopulations of exosomes with distinct molecular and biological properties

    NARCIS (Netherlands)

    Willms, Eduard; Johansson, Henrik J; Mäger, Imre; Lee, Yi; Blomberg, K Emelie M; Sadik, Mariam; Alaarg, Amr; Smith, C I Edvard; Lehtiö, Janne; El Andaloussi, Samir; Wood, Matthew J A; Vader, Pieter

    2016-01-01

    Cells release nano-sized membrane vesicles that are involved in intercellular communication by transferring biological information between cells. It is generally accepted that cells release at least three types of extracellular vesicles (EVs): apoptotic bodies, microvesicles and exosomes. While a wi

  3. The crosstalk of telomere dysfunction and inflammation through cell-free TERRA containing exosomes.

    Science.gov (United States)

    Wang, Zhuo; Lieberman, Paul M

    2016-08-01

    Telomeric repeats-containing RNA (TERRA) are telomere-derived non-coding RNAs that contribute to telomere function in protecting chromosome ends. We recently identified a cell-free form of TERRA (cfTERRA) enriched in extracellular exosomes. These cfTERRA-containing exosomes stimulate inflammatory cytokines when incubated with immune responsive cells. Here, we report that cfTERRA levels were increased in exosomes during telomere dysfunction induced by the expression of the dominant negative TRF2. The exosomes from these damaged cells also enriched with DNA damage marker γH2AX and fragmented telomere repeat DNA. Purified cfTERRA stimulated inflammatory cytokines, but the intact membrane-associated nucleoprotein complexes produced a more robust cytokine activation. Therefore, we propose cfTERRA-containing exosomes transport a telomere-associated molecular pattern (TAMP) and telomere-specific alarmin from dysfunctional telomeres to the extracellular environment to elicit an inflammatory response. Since cfTERRA can be readily detected in human serum it may provide a useful biomarker for the detection of telomere dysfunction in the early stage of cancers and aging-associated inflammatory disease.

  4. HPV-E7 Delivered by Engineered Exosomes Elicits a Protective CD8+ T Cell-Mediated Immune Response

    Directory of Open Access Journals (Sweden)

    Paola Di Bonito

    2015-03-01

    Full Text Available We developed an innovative strategy to induce a cytotoxic T cell (CTL immune response against protein antigens of choice. It relies on the production of exosomes, i.e., nanovesicles spontaneously released by all cell types. We engineered the upload of huge amounts of protein antigens upon fusion with an anchoring protein (i.e., HIV-1 Nefmut, which is an inactive protein incorporating in exosomes at high levels also when fused with foreign proteins. We compared the immunogenicity of engineered exosomes uploading human papillomavirus (HPV-E7 with that of lentiviral virus-like particles (VLPs incorporating equivalent amounts of the same antigen. These exosomes, whose limiting membrane was decorated with VSV-G, i.e., an envelope protein inducing pH-dependent endosomal fusion, proved to be as immunogenic as the cognate VLPs. It is noteworthy that the immunogenicity of the engineered exosomes remained unaltered in the absence of VSV-G. Most important, we provide evidence that the inoculation in mouse of exosomes uploading HPV-E7 induces production of anti-HPV E7 CTLs, blocks the growth of syngeneic tumor cells inoculated after immunization, and controls the development of tumor cells inoculated before the exosome challenge. These results represent the proof-of-concept about both feasibility and efficacy of the Nefmut-based exosome platform for the induction of CD8+ T cell immunity.

  5. Modeling putative therapeutic implications of exosome exchange between tumor and immune cells.

    Science.gov (United States)

    Lu, Mingyang; Huang, Bin; Hanash, Samir M; Onuchic, José N; Ben-Jacob, Eshel

    2014-10-07

    Development of effective strategies to mobilize the immune system as a therapeutic modality in cancer necessitates a better understanding of the contribution of the tumor microenvironment to the complex interplay between cancer cells and the immune response. Recently, effort has been directed at unraveling the functional role of exosomes and their cargo of messengers in this interplay. Exosomes are small vesicles (30-200 nm) that mediate local and long-range communication through the horizontal transfer of information, such as combinations of proteins, mRNAs and microRNAs. Here, we develop a tractable theoretical framework to study the putative role of exosome-mediated cell-cell communication in the cancer-immunity interplay. We reduce the complex interplay into a generic model whose three components are cancer cells, dendritic cells (consisting of precursor, immature, and mature types), and killer cells (consisting of cytotoxic T cells, helper T cells, effector B cells, and natural killer cells). The framework also incorporates the effects of exosome exchange on enhancement/reduction of cell maturation, proliferation, apoptosis, immune recognition, and activation/inhibition. We reveal tristability-possible existence of three cancer states: a low cancer load with intermediate immune level state, an intermediate cancer load with high immune level state, and a high cancer load with low immune-level state, and establish the corresponding effective landscape for the cancer-immunity network. We illustrate how the framework can contribute to the design and assessments of combination therapies.

  6. Exosomal proteins as potential diagnostic markers in advanced non-small cell lung carcinoma

    DEFF Research Database (Denmark)

    Jakobsen, Kristine Raaby; Paulsen, Birgitte Sandfeld; Bæk, Rikke

    2015-01-01

    Background: Lung cancer is one of the leading causes of cancer-related death. At the time of diagnosis, more than half of the patients will have disseminated disease and, yet, diagnosing can be challenging. New methods are desired to improve the diagnostic work-up. Exosomes are cell-derived vesic......Background: Lung cancer is one of the leading causes of cancer-related death. At the time of diagnosis, more than half of the patients will have disseminated disease and, yet, diagnosing can be challenging. New methods are desired to improve the diagnostic work-up. Exosomes are cell...... control subjects based on the differential display of exosomal protein markers. Methods: Plasma was isolated from 109 NSCLC patients with advanced stage (IIIa–IV) disease and 110 matched control subjects initially suspected of having cancer, but diagnosed to be cancer free. The Extracellular Vesicle Array...... (EV Array) was used to phenotype exosomes directly from the plasma samples. The array contained 37 antibodies targeting lung cancer-related proteins and was used to capture exosomes, which were visualised with a cocktail of biotin-conjugated CD9, CD63 and CD81 antibodies. Results: The EV Array...

  7. Exosomal proteins as potential diagnostic markers in advanced non-small cell lung carcinoma

    DEFF Research Database (Denmark)

    Jakobsen, Kristine R; Paulsen, Birgitte S; Bæk, Rikke

    2015-01-01

    BACKGROUND: Lung cancer is one of the leading causes of cancer-related death. At the time of diagnosis, more than half of the patients will have disseminated disease and, yet, diagnosing can be challenging. New methods are desired to improve the diagnostic work-up. Exosomes are cell-derived vesic......BACKGROUND: Lung cancer is one of the leading causes of cancer-related death. At the time of diagnosis, more than half of the patients will have disseminated disease and, yet, diagnosing can be challenging. New methods are desired to improve the diagnostic work-up. Exosomes are cell...... control subjects based on the differential display of exosomal protein markers. METHODS: Plasma was isolated from 109 NSCLC patients with advanced stage (IIIa-IV) disease and 110 matched control subjects initially suspected of having cancer, but diagnosed to be cancer free. The Extracellular Vesicle Array...... (EV Array) was used to phenotype exosomes directly from the plasma samples. The array contained 37 antibodies targeting lung cancer-related proteins and was used to capture exosomes, which were visualised with a cocktail of biotin-conjugated CD9, CD63 and CD81 antibodies. RESULTS: The EV Array...

  8. Tetraspanin-3 regulates protective immunity against Eimera tenella infection following immunization with dendritic cell-derived exosomes

    Science.gov (United States)

    The effects of immunization with dendritic cell (DC) exosomes, which had been incubated or non-incubated with an anti-tetraspanin-3 (Tspan-3) blocking antibody (Ab), were studied using an experimental model of Eimeria tenella avian coccidiosis. Purified exosomes from cecal tonsil and splenic DCs exp...

  9. Dendritic cells release HLA-B-associated transcript-3 positive exosomes to regulate natural killer function.

    Directory of Open Access Journals (Sweden)

    Venkateswara Rao Simhadri

    Full Text Available NKp30, a natural cytotoxicity receptor expressed on NK cells is critically involved in direct cytotoxicity against various tumor cells and directs both maturation and selective killing of dendritic cells. Recently the intracellular protein BAT3, which is involved in DNA damage induced apoptosis, was identified as a ligand for NKp30. However, the mechanisms underlying the exposure of the intracellular ligand BAT3 to surface NKp30 and its role in NK-DC cross talk remained elusive. Electron microscopy and flow cytometry demonstrate that exosomes released from 293T cells and iDCs express BAT3 on the surface and are recognized by NKp30-Ig. Overexpression and depletion of BAT3 in 293T cells directly correlates with the exosomal expression level and the activation of NK cell-mediated cytokine release. Furthermore, the NKp30-mediated NK/DC cross talk resulting either in iDC killing or maturation was BAT3-dependent. Taken together this puts forward a new model for the activation of NK cells through intracellular signals that are released via exosomes from accessory cells. The manipulation of the exosomal regulation may offer a novel strategy to induce tumor immunity or inhibit autoimmune diseases caused by NK cell-activation.

  10. A comparative analysis of lncRNAs in prostate cancer exosomes and their parental cell lines

    Directory of Open Access Journals (Sweden)

    Alireza Ahadi

    2016-09-01

    Full Text Available Prostate cancer is the second leading cancer in men world-wide. Due to its heterogeneous nature, a considerable amount of research effort has been dedicated in identifying effective clinical biomarkers with a focus on proteins, messenger RNA and microRNAs [1]. However, there is limited data on the role and expression of long noncoding RNAs (lncRNAs in prostate cancer exosomes [2]. This array dataset which is linked to our publication describes the profiling of human lncRNAs in prostate cancer and their exosomes from five different cell lines [3]. From this dataset, we identified a list of statistically significant prostate cancer lncRNAs which are differentially expressed in the exosomes compared to their parent cell lines. This dataset has been deposited into Gene Expression Omnibus (GSE81034.

  11. Exosomes in human semen restrict HIV-1 transmission by vaginal cells and block intravaginal replication of LP-BM5 murine AIDS virus complex.

    Science.gov (United States)

    Madison, Marisa N; Jones, Philip H; Okeoma, Chioma M

    2015-08-01

    Exosomes are membranous extracellular nanovesicles secreted by diverse cell types. Exosomes from healthy human semen have been shown to inhibit HIV-1 replication and to impair progeny virus infectivity. In this study, we examined the ability of healthy human semen exosomes to restrict HIV-1 and LP-BM5 murine AIDS virus transmission in three different model systems. We show that vaginal cells internalize exosomes with concomitant transfer of functional mRNA. Semen exosomes blocked the spread of HIV-1 from vaginal epithelial cells to target cells in our cell-to-cell infection model and suppressed transmission of HIV-1 across the vaginal epithelial barrier in our trans-well model. Our in vivo model shows that human semen exosomes restrict intravaginal transmission and propagation of murine AIDS virus. Our study highlights an antiretroviral role for semen exosomes that may be harnessed for the development of novel therapeutic strategies to combat HIV-1 transmission.

  12. Exosomes derived from SW480 colorectal cancer cells promote cell migration in HepG2 hepatocellular cancer cells via the mitogen-activated protein kinase pathway.

    Science.gov (United States)

    Chiba, Mitsuru; Watanabe, Narumi; Watanabe, Miki; Sakamoto, Maki; Sato, Akika; Fujisaki, Mizuki; Kubota, Shiori; Monzen, Satoru; Maruyama, Atsushi; Nanashima, Naoki; Kashiwakura, Ikuo; Nakamura, Toshiya

    2016-01-01

    Exosomes are membrane-derived extracellular vesicles that have recently been recognized as important mediators of intercellular communication. In the present study, we investigated the effects of exosomes derived from SW480 colorectal cancer cells in recipient HepG2 hepatocellular cancer cells. We demonstrated that SW480-derived exosomes were taken up by the recipient HepG2 cells via dynamin-dependent endocytosis and were localized to the HepG2 lysosomes. In addition, SW480-derived exosomes induced the phosphorylation of extracellular signal-regulated kinase (ERK)1/2 following their uptake into HepG2 cells. Of note, these changes occurred during the early phase after exosome treatment. Furthermore, SW480-derived exosomes promoted the migration of recipient HepG2 cells in a wound-healing assay, which was suppressed by pretreatment with U0126, an upstream inhibitor of ERK1/2. These results indicated that SW480-derived exosomes activated a classical mitogen-activated protein kinase pathway in recipient HepG2 cells via dynamin-dependent endocytosis and subsequently enhanced cell migration by ERK1/2 activation. Our results provide new insights into the regulation of cellular functions by exosomes.

  13. Mesenchymal Stem Cell-Derived Exosomes: Immunomodulatory Evaluation in an Antigen-Induced Synovitis Porcine Model

    Science.gov (United States)

    Casado, Javier G.; Blázquez, Rebeca; Vela, Francisco Javier; Álvarez, Verónica; Tarazona, Raquel; Sánchez-Margallo, Francisco Miguel

    2017-01-01

    Synovitis is an inflammatory process associated with pain, disability, and discomfort, which is usually treated with anti-inflammatory drugs or biological agents. Mesenchymal stem cells (MSCs) have been also successfully used in the treatment of inflammatory-related diseases such as synovitis or arthritis. In the last years, the exosomes derived from MSCs have become a promising tool for the treatment of inflammatory-related diseases and their therapeutic effect is thought to be mediated (at least in part) by their immunomodulatory potential. In this work, we aimed to evaluate the anti-inflammatory effect of these exosomes in an antigen-induced synovitis animal model. To our knowledge, this is the first report where exosomes derived from MSCs have been evaluated in an animal model of synovitis. Our results demonstrated a decrease of synovial lymphocytes together with a downregulation of TNF-α transcripts in those exosome-treated joints. These results support the immunomodulatory effect of these exosomes and point out that they may represent a promising therapeutic option for the treatment of synovitis.

  14. Quantitative Analysis of Exosomes From Murine Lung Cancer Cells by Flow Cytometry

    Science.gov (United States)

    Rim, Kyung-Taek; Kim, Soo-Jin

    2016-01-01

    In vivo studies regarding biochemical, molecular biological, and histopathological changes in cancer tissues have been widely performed by the administration of carcinogens in rodents. In these established methods, dissection of the animal following sacrifice must be carried out. Exosomes are cell-derived vesicles that are present in all body fluids and these vesicles have specific roles within cells. Thus, much attention is given to the clinical application of exosomes that can possibly be used for prediction and therapy and as biomarkers related to cancer. To develop a new tool for monitoring in vivo genetic alterations, as a result of carcinogenesis, without the need for frequent euthanasia, we performed quantitative measurement of exosomes in Mlg2908 murine lung fibroblasts and LA-4 and KLN 205 murine lung cancer cells using fluorescence-activated cell sorting. We detected an increase in CD63-specific exosomes in LA-4 lung cancer cells. This result is able to be applied to the classification of cancer-specific proteins and miRNA as diagnostic markers. PMID:27722146

  15. Dendritic cells as Achilles' heel and Trojan horse during varicella zoster virus infection

    Directory of Open Access Journals (Sweden)

    Günther eSchönrich

    2015-05-01

    Full Text Available Varicella zoster virus (VZV, a human alphaherpesvirus, causes varicella and subsequently estab-lishes latency within sensory nerve ganglia. Later in life VZV can reactivate to cause herpes zoster. A reduced frequency of VZV-specific T cells is strongly associated with herpes zoster illustrating that these immune cells are central to control latency. Dendritic cells (DCs are required for the generation of VZV-specific T cells. However, DCs can also be infected in vitro and in vivo allowing VZV to evade the antiviral immune response. Thus, DCs represent the immune systems’ Achilles heel. Uniquely among the human herpesviruses, VZV infects both DCs and T cells, and exploits both as Trojan horses. During primary infection VZV-infected DCs traffic to the draining lymph nodes and tonsils, where the virus is transferred to T cells. VZV-infected T cells subsequently spread infection throughout the body to give the typical varicella skin rash. The delicate interplay between VZV and DCs and its consequences for viral immune evasion and viral dissemination will be discussed in this article.

  16. Exosome-shuttling microRNA-21 promotes cell migration and invasion-targeting PDCD4 in esophageal cancer.

    Science.gov (United States)

    Liao, Juan; Liu, Ran; Shi, Ya-Juan; Yin, Li-Hong; Pu, Yue-Pu

    2016-06-01

    Recent evidence indicates that exosomes can mediate certain microRNAs (miRNAs) involved in a series of biological functions in tumor occurrence and development. Our previous studies showed that microRNA-21 (miR-21) was abundant in both esophageal cancer cells and their corresponding exosomes. The present study explored the function of exosome-shuttling miR-21 involved in esophageal cancer progression. We found that exosomes could be internalized from the extracellular space to the cytoplasm. The exosome-derived Cy3-labeled miR-21 mimics could be transported into recipient cells in a neutral sphingomyelinase 2 (nSMase2)-dependent manner. miR-21 overexpression from donor cells significantly promoted the migration and invasion of recipient cells by targeting programmed cell death 4 (PDCD4) and activating its downstream c-Jun N-terminal kinase (JNK) signaling pathway after co-cultivation. Our population plasma sample analysis indicated that miR-21 was upregulated significantly in plasma from esophageal cancer patients and showed a significant risk association for esophageal cancer. Our data demonstrated that a close correlation existed between exosome-shuttling miR-21 and esophageal cancer recurrence and distant metastasis. Thus, exosome-shuttling miR-21 may become a potential biomarker for prognosis among esophageal cancer patients.

  17. Trojan-Like Internalization of Anatase Titanium Dioxide Nanoparticles by Human Osteoblast Cells

    Science.gov (United States)

    Ribeiro, A. R.; Gemini-Piperni, S.; Travassos, R.; Lemgruber, L.; C. Silva, R.; Rossi, A. L.; Farina, M.; Anselme, K.; Shokuhfar, T.; Shahbazian-Yassar, R.; Borojevic, R.; Rocha, L. A.; Werckmann, J.; Granjeiro, J. M.

    2016-03-01

    Dentistry and orthopedics are undergoing a revolution in order to provide more reliable, comfortable and long-lasting implants to patients. Titanium (Ti) and titanium alloys have been used in dental implants and total hip arthroplasty due to their excellent biocompatibility. However, Ti-based implants in human body suffer surface degradation (corrosion and wear) resulting in the release of metallic ions and solid wear debris (mainly titanium dioxide) leading to peri-implant inflammatory reactions. Unfortunately, our current understanding of the biological interactions with titanium dioxide nanoparticles is still very limited. Taking this into consideration, this study focuses on the internalization of titanium dioxide nanoparticles on primary bone cells, exploring the events occurring at the nano-bio interface. For the first time, we report the selective binding of calcium (Ca), phosphorous (P) and proteins from cell culture medium to anatase nanoparticles that are extremely important for nanoparticle internalization and bone cells survival. In the intricate biological environment, anatase nanoparticles form bio-complexes (mixture of proteins and ions) which act as a kind of ‘Trojan-horse’ internalization by cells. Furthermore, anatase nanoparticles-induced modifications on cell behavior (viability and internalization) could be understand in detail. The results presented in this report can inspire new strategies for the use of titanium dioxide nanoparticles in several regeneration therapies.

  18. Trojan-Like Internalization of Anatase Titanium Dioxide Nanoparticles by Human Osteoblast Cells.

    Science.gov (United States)

    Ribeiro, A R; Gemini-Piperni, S; Travassos, R; Lemgruber, L; Silva, R C; Rossi, A L; Farina, M; Anselme, K; Shokuhfar, T; Shahbazian-Yassar, R; Borojevic, R; Rocha, L A; Werckmann, J; Granjeiro, J M

    2016-03-29

    Dentistry and orthopedics are undergoing a revolution in order to provide more reliable, comfortable and long-lasting implants to patients. Titanium (Ti) and titanium alloys have been used in dental implants and total hip arthroplasty due to their excellent biocompatibility. However, Ti-based implants in human body suffer surface degradation (corrosion and wear) resulting in the release of metallic ions and solid wear debris (mainly titanium dioxide) leading to peri-implant inflammatory reactions. Unfortunately, our current understanding of the biological interactions with titanium dioxide nanoparticles is still very limited. Taking this into consideration, this study focuses on the internalization of titanium dioxide nanoparticles on primary bone cells, exploring the events occurring at the nano-bio interface. For the first time, we report the selective binding of calcium (Ca), phosphorous (P) and proteins from cell culture medium to anatase nanoparticles that are extremely important for nanoparticle internalization and bone cells survival. In the intricate biological environment, anatase nanoparticles form bio-complexes (mixture of proteins and ions) which act as a kind of 'Trojan-horse' internalization by cells. Furthermore, anatase nanoparticles-induced modifications on cell behavior (viability and internalization) could be understand in detail. The results presented in this report can inspire new strategies for the use of titanium dioxide nanoparticles in several regeneration therapies.

  19. Trojan-Like Internalization of Anatase Titanium Dioxide Nanoparticles by Human Osteoblast Cells

    Science.gov (United States)

    Ribeiro, A. R.; Gemini-Piperni, S.; Travassos, R.; Lemgruber, L.; C. Silva, R.; Rossi, A. L.; Farina, M.; Anselme, K.; Shokuhfar, T.; Shahbazian-Yassar, R.; Borojevic, R.; Rocha, L. A.; Werckmann, J.; Granjeiro, J. M.

    2016-01-01

    Dentistry and orthopedics are undergoing a revolution in order to provide more reliable, comfortable and long-lasting implants to patients. Titanium (Ti) and titanium alloys have been used in dental implants and total hip arthroplasty due to their excellent biocompatibility. However, Ti-based implants in human body suffer surface degradation (corrosion and wear) resulting in the release of metallic ions and solid wear debris (mainly titanium dioxide) leading to peri-implant inflammatory reactions. Unfortunately, our current understanding of the biological interactions with titanium dioxide nanoparticles is still very limited. Taking this into consideration, this study focuses on the internalization of titanium dioxide nanoparticles on primary bone cells, exploring the events occurring at the nano-bio interface. For the first time, we report the selective binding of calcium (Ca), phosphorous (P) and proteins from cell culture medium to anatase nanoparticles that are extremely important for nanoparticle internalization and bone cells survival. In the intricate biological environment, anatase nanoparticles form bio-complexes (mixture of proteins and ions) which act as a kind of ‘Trojan-horse’ internalization by cells. Furthermore, anatase nanoparticles-induced modifications on cell behavior (viability and internalization) could be understand in detail. The results presented in this report can inspire new strategies for the use of titanium dioxide nanoparticles in several regeneration therapies. PMID:27021687

  20. Extraction and identification of exosomes from drug-resistant breast cancer cells and their potential role in cell-to-cell drug-resistance transfer

    Institute of Scientific and Technical Information of China (English)

    许金金

    2014-01-01

    Objective To explore whether docetaxel-resistant cells(MCF-7/Doc)and doxorubicin-resistant cells(MCF-7/ADM)can secrete Exosomes and their potential role in cell-cell drug-resistance transfer.Methods Exosomes were extracted from the cell culture supernatants of MCF-7/Doc and MCF-7/ADM cells by fractionation ultracentrifugation,and were identified by transmission

  1. Pharmacokinetics of Exosomes-an Important Factor for Elucidating the Biological Roles of Exosomes and for the Development of Exosome-Based Therapeutics.

    Science.gov (United States)

    Morishita, Masaki; Takahashi, Yuki; Nishikawa, Makiya; Takakura, Yoshinobu

    2017-03-07

    Exosomes are small membrane vesicles containing lipids, proteins, and nucleic acids. Recently, researchers have uncovered that exosomes are involved in various biological events, such as tumor growth, metastasis, and the immune response, by delivering their cargos to exosome-receiving cells. Moreover, exosomes are expected to be employed in therapeutic treatments, such as tissue regeneration therapy and antitumor immunotherapy, since exosomes are effective delivery vehicles for proteins, nucleic acids, and other bioactive compounds. To elucidate the biological functions of exosomes, and for the development of exosome-based therapeutics, the pharmacokinetics of exosomes is important. In this review, we aim to summarize current knowledge about the pharmacokinetics and biodistribution of exosomes. The pharmacokinetics of exogenously administered exosomes is discussed based on the tissue distribution, types of cells taking up exosomes, and key molecules in the pharmacokinetics of exosomes. In addition, recent progress in the methods to control the pharmacokinetics of exosomes is reviewed.

  2. Exosomes secreted by nematode parasites transfer small RNAs to mammalian cells and modulate innate immunity.

    Science.gov (United States)

    Buck, Amy H; Coakley, Gillian; Simbari, Fabio; McSorley, Henry J; Quintana, Juan F; Le Bihan, Thierry; Kumar, Sujai; Abreu-Goodger, Cei; Lear, Marissa; Harcus, Yvonne; Ceroni, Alessandro; Babayan, Simon A; Blaxter, Mark; Ivens, Alasdair; Maizels, Rick M

    2014-11-25

    In mammalian systems RNA can move between cells via vesicles. Here we demonstrate that the gastrointestinal nematode Heligmosomoides polygyrus, which infects mice, secretes vesicles containing microRNAs (miRNAs) and Y RNAs as well as a nematode Argonaute protein. These vesicles are of intestinal origin and are enriched for homologues of mammalian exosome proteins. Administration of the nematode exosomes to mice suppresses Type 2 innate responses and eosinophilia induced by the allergen Alternaria. Microarray analysis of mouse cells incubated with nematode exosomes in vitro identifies Il33r and Dusp1 as suppressed genes, and Dusp1 can be repressed by nematode miRNAs based on a reporter assay. We further identify miRNAs from the filarial nematode Litomosoides sigmodontis in the serum of infected mice, suggesting that miRNA secretion into host tissues is conserved among parasitic nematodes. These results reveal exosomes as another mechanism by which helminths manipulate their hosts and provide a mechanistic framework for RNA transfer between animal species.

  3. Inhibition of myocardial ischemia/reperfusion injury by exosomes secreted from mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Heng; XIANG Meng; MENG Dan; SUN Ning; CHEN Si-feng

    2016-01-01

    Exosomes secreted by mesenchymal stem cells have shown great therapeutic potential in regenerative medicine .In this study, we performed meta-analysis to assess the clinical effectiveness of using exosomes in ischemia /reperfusion injury based on the reports pub-lished between January 2000 and September 2015 and indexed in the PubMed and Web of Science databases .The effect of exosomes on heart function was evaluated according to the following parameters:the area at risk as a percentage of the left ventricle , infarct size as a percentage of the area at risk , infarct size as a percentage of the left ventricle , left ventricular ejection fraction , left ventricular frac-tion shortening , end-diastolic volume , and end-systolic volume .Our analysis indicated that the currently available evidence confirmed the therapeutic potential of mesenchymal stem cell-secreted exosomes in the improvement of heart function .However , further mechanis-tic studies, therapeutic safety and clinical trials are required for optimization and validation of this approach to cardiac regeneration after ischemia/reperfusion injury .

  4. Exosome-Mediated Intercellular Communication between Hepatitis C Virus-Infected Hepatocytes and Hepatic Stellate Cells.

    Science.gov (United States)

    Devhare, Pradip B; Sasaki, Reina; Shrivastava, Shubham; Di Bisceglie, Adrian M; Ray, Ranjit; Ray, Ratna B

    2017-03-15

    Fibrogenic pathways in the liver are principally regulated by activation of hepatic stellate cells (HSC). Fibrosis is associated with chronic hepatitis C virus (HCV) infection, although the mechanism is poorly understood. HSC comprise the major population of nonparenchymal cells in the liver. Since HCV does not replicate in HSC, we hypothesized that exosomes secreted from HCV-infected hepatocytes activate HSC. Primary or immortalized human hepatic stellate (LX2) cells were exposed to exosomes derived from HCV-infected hepatocytes (HCV-exo), and the expression of fibrosis-related genes was examined. Our results demonstrated that HCV-exo internalized to HSC and increased the expression of profibrotic markers. Further analysis suggested that HCV-exo carry miR-19a and target SOCS3 in HSC, which in turn activates the STAT3-mediated transforming growth factor β (TGF-β) signaling pathway and enhances fibrosis marker genes. The higher expression of miR-19a in exosomes was also observed from HCV-infected hepatocytes and in sera of chronic HCV patients with fibrosis compared to healthy volunteers and non-HCV-related liver disease patients with fibrosis. Together, our results demonstrated that miR-19a carried through the exosomes from HCV-infected hepatocytes activates HSC by modulating the SOCS-STAT3 axis. Our results implicated a novel mechanism of exosome-mediated intercellular communication in the activation of HSC for liver fibrosis in HCV infection.IMPORTANCE HCV-associated liver fibrosis is a critical step for end-stage liver disease progression. However, the molecular mechanisms for hepatic stellate-cell activation by HCV-infected hepatocytes are underexplored. Here, we provide a role for miR-19a carried through the exosomes in intercellular communication between HCV-infected hepatocytes and HSC in fibrogenic activation. Furthermore, we demonstrate the role of exosomal miR-19a in activation of the STAT3-TGF-β pathway in HSC. This study contributes to the

  5. Porcine milk-derived exosomes promote proliferation of intestinal epithelial cells

    Science.gov (United States)

    Chen, Ting; Xie, Mei-Ying; Sun, Jia-Jie; Ye, Rui-Song; Cheng, Xiao; Sun, Rui-Ping; Wei, Li-Min; Li, Meng; Lin, De-Lin; Jiang, Qing-Yan; Xi, Qian-Yun; Zhang, Yong-Liang

    2016-01-01

    Milk-derived exosomes were identified as a novel mechanism of mother-to-child transmission of regulatory molecules, but their functions in intestinal tissues of neonates are not well-studied. Here, we characterized potential roles of porcine milk-derived exosomes in the intestinal tract. In vitro, treatment with milk-derived exosomes (27 ± 3 ng and 55 ± 5 ng total RNA) significantly promoted IPEC-J2 cell proliferation by MTT, CCK8, EdU fluorescence and EdU flow cytometry assays. The qRT-PCR and Western blot analyses indicated milk-derived exosomes (0.27 ± 0.03 μg total RNA) significantly promoted expression of CDX2, IGF-1R and PCNA, and inhibited p53 gene expression involved in intestinal proliferation. Additionally, six detected miRNAs were significantly increased in IPEC-J2 cell, while FAS and SERPINE were significantly down-regulated relative to that in control. In vivo, treated groups (0.125 μg and 0.25 μg total RNA) significantly raised mice’ villus height, crypt depth and ratio of villus length to crypt depth of intestinal tissues, significantly increased CDX2, PCNA and IGF-1R’ expression and significantly inhibited p53′ expression. Our study demonstrated that milk-derived exosomes can facilitate intestinal cell proliferation and intestinal tract development, thus giving a new insight for milk nutrition and newborn development and health. PMID:27646050

  6. Targeting exosomes from preadipocytes inhibits preadipocyte to cancer stem cell signaling in early-stage breast cancer.

    Science.gov (United States)

    Gernapudi, Ramkishore; Yao, Yuan; Zhang, Yongshu; Wolfson, Benjamin; Roy, Sanchita; Duru, Nadire; Eades, Gabriel; Yang, Peixin; Zhou, Qun

    2015-04-01

    The tumor microenvironment plays a critical role in regulating breast tumor progression. Signaling between preadipocytes and breast cancer cells has been found to promote breast tumor formation and metastasis. Exosomes secreted from preadipocytes are important components of the cancer stem cell niche. Mouse preadipocytes (3T3L1) are treated with the natural antitumor compound shikonin (SK) and exosomes derived from mouse preadipocytes are co-cultured with MCF10DCIS cells. We examine how preadipocyte-derived exosomes can regulate early-stage breast cancer via regulating stem cell renewal, cell migration, and tumor formation. We identify a critical miR-140/SOX2/SOX9 axis that regulates differentiation, stemness, and migration in the tumor microenvironment. Next, we find that the natural antitumor compound SK can inhibit preadipocyte signaling inhibiting nearby ductal carcinoma in situ (DCIS) cells. Through co-culture experiments, we find that SK-treated preadipocytes secrete exosomes with high levels of miR-140, which can impact nearby DCIS cells through targeting SOX9 signaling. Finally, we find that preadipocyte-derived exosomes promote tumorigenesis in vivo, providing strong support for the importance of exosomal signaling in the tumor microenvironment. Our data also show that targeting the tumor microenvironment may assist in blocking tumor progression.

  7. Sedentary lifestyle related exosomal release of Hotair from gluteal-femoral fat promotes intestinal cell proliferation

    Science.gov (United States)

    Lu, Xiaozhao; Bai, Danna; Liu, Xiangwei; Zhou, Chen; Yang, Guodong

    2017-01-01

    Pioneering epidemiological work has established strong association of sedentary lifestyle and obesity with the risk of colorectal cancer, while the detailed underlying mechanism remains unknown. Here we show that Hotair (HOX transcript antisense RNA) is a pro-adipogenic long non-coding RNA highly expressed in gluteal-femoral fat over other fat depots. Hotair knockout in adipose tissue results in gluteal-femoral fat defect. Squeeze of the gluteal-femoral fat induces intestinal proliferation in wildtype mice, while not in Hotair knockout mice. Mechanistically, squeeze of the gluteal-femoral fat induces exosomal Hotair secretion mainly by transcriptional upregulation of Hotair via NFκB. And increased exosomal Hotair in turn circulates in the blood and is partially endocytosed by the intestine, finally promoting the stemness and proliferation of intestinal stem/progenitor cells via Wnt activation. Clinically, obese subjects with sedentary lifestyle have much higher exosomal HOTAIR expression in the serum. These findings establish that sedentary lifestyle promotes exosomal Hotair release from the gluteal-femoral fat, which in turn facilitates intestinal stem and/or progenitor proliferation, raising a possible link between sedentary lifestyle with colorectal tumorigenesis. PMID:28361920

  8. Exosomal proteins as potential diagnostic markers in advanced non-small cell lung carcinoma

    Directory of Open Access Journals (Sweden)

    Kristine R. Jakobsen

    2015-03-01

    Full Text Available Background: Lung cancer is one of the leading causes of cancer-related death. At the time of diagnosis, more than half of the patients will have disseminated disease and, yet, diagnosing can be challenging. New methods are desired to improve the diagnostic work-up. Exosomes are cell-derived vesicles displaying various proteins on their membrane surfaces. In addition, they are readily available in blood samples where they constitute potential biomarkers of human diseases, such as cancer. Here, we examine the potential of distinguishing non-small cell lung carcinoma (NSCLC patients from control subjects based on the differential display of exosomal protein markers. Methods: Plasma was isolated from 109 NSCLC patients with advanced stage (IIIa–IV disease and 110 matched control subjects initially suspected of having cancer, but diagnosed to be cancer free. The Extracellular Vesicle Array (EV Array was used to phenotype exosomes directly from the plasma samples. The array contained 37 antibodies targeting lung cancer-related proteins and was used to capture exosomes, which were visualised with a cocktail of biotin-conjugated CD9, CD63 and CD81 antibodies. Results: The EV Array analysis was capable of detecting and phenotyping exosomes in all samples from only 10 µL of unpurified plasma. Multivariate analysis using the Random Forests method produced a combined 30-marker model separating the two patient groups with an area under the curve of 0.83, CI: 0.77–0.90. The 30-marker model has a sensitivity of 0.75 and a specificity of 0.76, and it classifies patients with 75.3% accuracy. Conclusion: The EV Array technique is a simple, minimal-invasive tool with potential to identify lung cancer patients.

  9. Oxygen tension regulates the miRNA profile and bioactivity of exosomes released from extravillous trophoblast cells - Liquid biopsies for monitoring complications of pregnancy.

    Science.gov (United States)

    Truong, Grace; Guanzon, Dominic; Kinhal, Vyjayanthi; Elfeky, Omar; Lai, Andrew; Longo, Sherri; Nuzhat, Zarin; Palma, Carlos; Scholz-Romero, Katherin; Menon, Ramkumar; Mol, Ben W; Rice, Gregory E; Salomon, Carlos

    2017-01-01

    Our understanding of how cells communicate has undergone a paradigm shift since the recent recognition of the role of exosomes in intercellular signaling. In this study, we investigated whether oxygen tension alters the exosome release and miRNA profile from extravillous trophoblast (EVT) cells, modifying their bioactivity on endothelial cells (EC). Furthermore, we have established the exosomal miRNA profile at early gestation in women who develop pre-eclampsia (PE) and spontaneous preterm birth (SPTB). HTR-8/SVneo cells were used as an EVT model. The effect of oxygen tension (i.e. 8% and 1% oxygen) on exosome release was quantified using nanocrystals (Qdot®) coupled to CD63 by fluorescence NTA. A real-time, live-cell imaging system (Incucyte™) was used to establish the effect of exosomes on EC. Plasma samples were obtained at early gestation (<18 weeks) and classified according to pregnancy outcomes. An Illumina TrueSeq Small RNA kit was used to construct a small RNA library from exosomal RNA obtained from EVT and plasma samples. The number of exosomes was significantly higher in EVT cultured under 1% compared to 8% oxygen. In total, 741 miRNA were identified in exosomes from EVT. Bioinformatic analysis revealed that these miRNA were associated with cell migration and cytokine production. Interestingly, exosomes isolated from EVT cultured at 8% oxygen increased EC migration, whilst exosomes cultured at 1% oxygen decreased EC migration. These changes were inversely proportional to TNF-α released from EC. Finally, we have identified a set of unique miRNAs in exosomes from EVT cultured at 1% oxygen and exosomes isolated from the circulation of mothers at early gestation, who later developed PE and SPTB. We suggest that aberrant exosomal signalling by placental cells is a common aetiological factor in pregnancy complications characterised by incomplete SpA remodeling and is therefore a clinically relevant biomarker of pregnancy complications.

  10. Oxygen tension regulates the miRNA profile and bioactivity of exosomes released from extravillous trophoblast cells – Liquid biopsies for monitoring complications of pregnancy

    Science.gov (United States)

    Truong, Grace; Guanzon, Dominic; Kinhal, Vyjayanthi; Elfeky, Omar; Lai, Andrew; Longo, Sherri; Nuzhat, Zarin; Palma, Carlos; Scholz-Romero, Katherin; Menon, Ramkumar; Mol, Ben W.; Rice, Gregory E.; Salomon, Carlos

    2017-01-01

    Our understanding of how cells communicate has undergone a paradigm shift since the recent recognition of the role of exosomes in intercellular signaling. In this study, we investigated whether oxygen tension alters the exosome release and miRNA profile from extravillous trophoblast (EVT) cells, modifying their bioactivity on endothelial cells (EC). Furthermore, we have established the exosomal miRNA profile at early gestation in women who develop pre-eclampsia (PE) and spontaneous preterm birth (SPTB). HTR-8/SVneo cells were used as an EVT model. The effect of oxygen tension (i.e. 8% and 1% oxygen) on exosome release was quantified using nanocrystals (Qdot®) coupled to CD63 by fluorescence NTA. A real-time, live-cell imaging system (Incucyte™) was used to establish the effect of exosomes on EC. Plasma samples were obtained at early gestation (<18 weeks) and classified according to pregnancy outcomes. An Illumina TrueSeq Small RNA kit was used to construct a small RNA library from exosomal RNA obtained from EVT and plasma samples. The number of exosomes was significantly higher in EVT cultured under 1% compared to 8% oxygen. In total, 741 miRNA were identified in exosomes from EVT. Bioinformatic analysis revealed that these miRNA were associated with cell migration and cytokine production. Interestingly, exosomes isolated from EVT cultured at 8% oxygen increased EC migration, whilst exosomes cultured at 1% oxygen decreased EC migration. These changes were inversely proportional to TNF-α released from EC. Finally, we have identified a set of unique miRNAs in exosomes from EVT cultured at 1% oxygen and exosomes isolated from the circulation of mothers at early gestation, who later developed PE and SPTB. We suggest that aberrant exosomal signalling by placental cells is a common aetiological factor in pregnancy complications characterised by incomplete SpA remodeling and is therefore a clinically relevant biomarker of pregnancy complications. PMID:28350871

  11. Mesenchymal Stem Cell-Derived Exosomes: New Opportunity in Cell-Free Therapy

    Science.gov (United States)

    Pashoutan Sarvar, Davod; Shamsasenjan, Karim; Akbarzadehlaleh, Parvin

    2016-01-01

    Mesenchymal stromal/stem cells (MSCs) are involved in tissue homeostasis through direct cell-to-cell interaction, as well as secretion of soluble factors. Exosomes are the sort of soluble biological mediators that obtained from MSCs cultured media in vitro. MSC-derived exosomes (MSC-DEs) which produced under physiological or pathological conditions are central mediators of intercellular communications by conveying proteins, lipids, mRNAs, siRNA, ribosomal RNAs and miRNAs to the neighbor or distant cells. MSC-DEs have been tested in various disease models, and the results have revealed that their functions are similar to those of MSCs. They have the supportive functions in organisms such as repairing tissue damages, suppressing inflammatory responses, and modulating the immune system. MSC-DEs are of great interest in the scope of regenerative medicine because of their unique capacity to the regeneration of the damaged tissues, and the present paper aims to introduce MSC-DEs as a novel hope in cell-free therapy.

  12. Potential Therapies by Stem Cell-Derived Exosomes in CNS Diseases: Focusing on the Neurogenic Niche

    Science.gov (United States)

    Luarte, Alejandro; Bátiz, Luis Federico; Wyneken, Ursula; Lafourcade, Carlos

    2016-01-01

    Neurodegenerative disorders are one of the leading causes of death and disability and one of the biggest burdens on health care systems. Novel approaches using various types of stem cells have been proposed to treat common neurodegenerative disorders such as Alzheimer's Disease, Parkinson's Disease, or stroke. Moreover, as the secretome of these cells appears to be of greater benefit compared to the cells themselves, the extracellular components responsible for its therapeutic benefit have been explored. Stem cells, as well as most cells, release extracellular vesicles such as exosomes, which are nanovesicles able to target specific cell types and thus to modify their function by delivering proteins, lipids, and nucleic acids. Exosomes have recently been tested in vivo and in vitro as therapeutic conveyors for the treatment of diseases. As such, they could be engineered to target specific populations of cells within the CNS. Considering the fact that many degenerative brain diseases have an impact on adult neurogenesis, we discuss how the modulation of the adult neurogenic niches may be a therapeutic target of stem cell-derived exosomes. These novel approaches should be examined in cellular and animal models to provide better, more effective, and specific therapeutic tools in the future. PMID:27195011

  13. Potential Therapies by Stem Cell-Derived Exosomes in CNS Diseases: Focusing on the Neurogenic Niche

    Directory of Open Access Journals (Sweden)

    Alejandro Luarte

    2016-01-01

    Full Text Available Neurodegenerative disorders are one of the leading causes of death and disability and one of the biggest burdens on health care systems. Novel approaches using various types of stem cells have been proposed to treat common neurodegenerative disorders such as Alzheimer’s Disease, Parkinson’s Disease, or stroke. Moreover, as the secretome of these cells appears to be of greater benefit compared to the cells themselves, the extracellular components responsible for its therapeutic benefit have been explored. Stem cells, as well as most cells, release extracellular vesicles such as exosomes, which are nanovesicles able to target specific cell types and thus to modify their function by delivering proteins, lipids, and nucleic acids. Exosomes have recently been tested in vivo and in vitro as therapeutic conveyors for the treatment of diseases. As such, they could be engineered to target specific populations of cells within the CNS. Considering the fact that many degenerative brain diseases have an impact on adult neurogenesis, we discuss how the modulation of the adult neurogenic niches may be a therapeutic target of stem cell-derived exosomes. These novel approaches should be examined in cellular and animal models to provide better, more effective, and specific therapeutic tools in the future.

  14. Carcinogenic activity of PbS quantum dots screened using exosomal biomarkers secreted from HEK293 cells.

    Science.gov (United States)

    Kim, Jung-Hee; Kim, Hye-Rim; Lee, Bo-Ram; Choi, Eun-Sook; In, Su-Il; Kim, Eunjoo

    2015-01-01

    Lead sulfide (PbS) quantum dots (QDs) have been applied in the biomedical area because they offer an excellent platform for theragnostic applications. In order to comprehensively evaluate the biocompatibility of PbS QDs in human cells, we analyzed the exosomes secreted from cells because exosomes are released during cellular stress to convey signals to other cells and serve as a reservoir of enriched biomarkers. PbS QDs were synthesized and coated with 3-mercaptopropionic acid (MPA) to allow the particles to disperse in water. Exosomes were isolated from HEK293 cells treated with PbS-MPA at concentrations of 0 µg/mL, 5 µg/mL, and 50 µg/mL, and the exosomal expression levels of miRNAs and proteins were analyzed. As a result, five miRNAs and two proteins were proposed as specific exosomal biomarkers for the exposure of HEK293 cells to PbS-MPA. Based on the pathway analysis, the molecular signature of the exosomes suggested that PbS-MPA QDs had carcinogenic activity. The comet assay and expression of molecular markers, such as p53, interleukin (IL)-8, and C-X-C motif chemokine 5, indicated that DNA damage occurred in HEK293 cells following PbS-MPA exposure, which supported the carcinogenic activity of the particles. In addition, there was obvious intensification of miRNA expression signals in the exosomes compared with that of the parent cells, which suggested that exosomal biomarkers could be detected more sensitively than those of whole cellular extracts.

  15. Exosome: A Novel and Safer Therapeutic Refinement of Mesenchymal Stem Cell

    OpenAIRE

    Yeo, Ronne Wee Yeh; Lai, Ruenn Chai; Tan, Kok Hian; Lim, Sai Kiang

    2013-01-01

    Mesenchymal stem cell (MSC) has just been approved as the first “off-the-shelf” stem cell pharmaceutical drug with an anticipation of more approvals following completion of numerous rigorous clinical trials. Despite this progress, the rationale for MSC therapeutic efficacy remains tenuous and is increasingly rationalized on a secretion rather than differentiation mechanism. Recent studies identifying exosome as the secreted agent mediating MSC therapeutic efficacy coul...

  16. Exosomes derived from M. Bovis BCG infected macrophages activate antigen-specific CD4+ and CD8+ T cells in vitro and in vivo.

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    Pramod K Giri

    Full Text Available Activation of both CD4(+ and CD8(+ T cells is required for an effective immune response to an M. tuberculosis infection. However, infected macrophages are poor antigen presenting cells and may be spatially separated from recruited T cells, thus limiting antigen presentation within a granuloma. Our previous studies showed that infected macrophages release from cells small membrane-bound vesicles called exosomes which contain mycobacterial lipid components and showed that these exosomes could stimulate a pro-inflammatory response in naïve macrophages. In the present study we demonstrate that exosomes stimulate both CD4(+ and CD8(+ splenic T cells isolated from mycobacteria-sensitized mice. Although the exosomes contain MHC I and II as well as costimulatory molecules, maximum stimulation of T cells required prior incubation of exosomes with antigen presenting cells. Exosomes isolated from M. bovis and M. tuberculosis infected macrophages also stimulated activation and maturation of mouse bone marrow-derived dendritic cells. Interestingly, intranasal administration of mice with exosomes isolated from M. bovis BCG infected macrophages induce the generation of memory CD4(+ and CD8(+ T cells. The isolated T cells also produced IFN-gamma upon restimulation with BCG antigens. The release of exosomes from infected macrophages may overcome some of the defects in antigen presentation associated with mycobacterial infections and we suggest that exosomes may be a promising M. tuberculosis vaccine candidate.

  17. Retinal pigment epithelium cell-derived exosomes: Possible relevance to CNV in wet-age related macular degeneration.

    Science.gov (United States)

    Tong, Yao; Zhou, Ya-Li; Wang, Yi-Xiao; Zhao, Pei-Quan; Wang, Zhao-Yang

    2016-12-01

    Exosomes are small vesicles that are released by almost every cell type and play a crucial role in many physiological and pathological processes associated with different diseases. Specifically, they promote angiogenesis in the pathogenesis of some diseases. According to previous research, the proteins of exosomes taken from the aqueous humor (AH) of patients with wet-age related macular degeneration (AMD) may function as a new diagnostic biomarker of AMD, suggesting that exosomes may play an important role in the occurrence and development of choroidal neovascularization (CNV). Moreover, additional research has revealed that the levels of some protein makers of exosomes are up-regulated in aged retinal pigment epithelium (RPE) and that drusen and oxidative stress may promote the secretion of exosomes derived from RPE cells. Consequently, we hypothesize that RPE cell-derived exosomes may be relevant to CNV in wet AMD. If this hypothesis is proven correct, future studies based on this link may also help to elucidate the molecular mechanisms of wet AMD and to find new therapeutic targets for the treatment of AMD.

  18. Comprehensive Proteomic Analysis of Mesenchymal Stem Cell Exosomes Reveals Modulation of Angiogenesis via Nuclear Factor-KappaB Signaling.

    Science.gov (United States)

    Anderson, Johnathon D; Johansson, Henrik J; Graham, Calvin S; Vesterlund, Mattias; Pham, Missy T; Bramlett, Charles S; Montgomery, Elizabeth N; Mellema, Matt S; Bardini, Renee L; Contreras, Zelenia; Hoon, Madeline; Bauer, Gerhard; Fink, Kyle D; Fury, Brian; Hendrix, Kyle J; Chedin, Frederic; El-Andaloussi, Samir; Hwang, Billie; Mulligan, Michael S; Lehtiö, Janne; Nolta, Jan A

    2016-03-01

    Mesenchymal stem cells (MSC) are known to facilitate healing of ischemic tissue related diseases through proangiogenic secretory proteins. Recent studies further show that MSC derived exosomes function as paracrine effectors of angiogenesis, however, the identity of which components of the exosome proteome responsible for this effect remains elusive. To address this we used high-resolution isoelectric focusing coupled liquid chromatography tandem mass spectrometry, an unbiased high throughput proteomics approach to comprehensively characterize the proteinaceous contents of MSCs and MSC derived exosomes. We probed the proteome of MSCs and MSC derived exosomes from cells cultured under expansion conditions and under ischemic tissue simulated conditions to elucidate key angiogenic paracrine effectors present and potentially differentially expressed in these conditions. In total, 6,342 proteins were identified in MSCs and 1,927 proteins in MSC derived exosomes, representing to our knowledge the first time these proteomes have been probed comprehensively. Multilayered analyses identified several putative paracrine effectors of angiogenesis present in MSC exosomes and increased in expression in MSCs exposed to ischemic tissue-simulated conditions; these include platelet derived growth factor, epidermal growth factor, fibroblast growth factor, and most notably nuclear factor-kappaB (NFkB) signaling pathway proteins. NFkB signaling was identified as a key mediator of MSC exosome induced angiogenesis in endothelial cells by functional in vitro validation using a specific inhibitor. Collectively, the results of our proteomic analysis show that MSC derived exosomes contain a robust profile of angiogenic paracrine effectors, which have potential for the treatment of ischemic tissue-related diseases.

  19. Inhibition of the Expression of the Small Heat Shock Protein αB-Crystallin Inhibits Exosome Secretion in Human Retinal Pigment Epithelial Cells in Culture.

    Science.gov (United States)

    Gangalum, Rajendra K; Bhat, Ankur M; Kohan, Sirus A; Bhat, Suraj P

    2016-06-17

    Exosomes carry cell type-specific molecular cargo to extracellular destinations and therefore act as lateral vectors of intercellular communication and transfer of genetic information from one cell to the other. We have shown previously that the small heat shock protein αB-crystallin (αB) is exported out of the adult human retinal pigment epithelial cells (ARPE19) packaged in exosomes. Here, we demonstrate that inhibition of the expression of αB via shRNA inhibits exosome secretion from ARPE19 cells indicating that exosomal cargo may have a role in exosome biogenesis (synthesis and/or secretion). Sucrose density gradient fractionation of the culture medium and cellular extracts suggests continued synthesis of exosomes but an inhibition of exosome secretion. In cells where αB expression was inhibited, the distribution of CD63 (LAMP3), an exosome marker, is markedly altered from the normal dispersed pattern to a stacked perinuclear presence. Interestingly, the total anti-CD63(LAMP3) immunofluorescence in the native and αB-inhibited cells remains unchanged suggesting continued exosome synthesis under conditions of impaired exosome secretion. Importantly, inhibition of the expression of αB results in a phenotype of the RPE cell that contains an increased number of vacuoles and enlarged (fused) vesicles that show increased presence of CD63(LAMP3) and LAMP1 indicating enhancement of the endolysosomal compartment. This is further corroborated by increased Rab7 labeling of this compartment (RabGTPase 7 is known to be associated with late endosome maturation). These data collectively point to a regulatory role for αB in exosome biogenesis possibly via its involvement at a branch point in the endocytic pathway that facilitates secretion of exosomes.

  20. Exosomal microRNA miR-1246 induces cell motility and invasion through the regulation of DENND2D in oral squamous cell carcinoma

    Science.gov (United States)

    Sakha, Sujata; Muramatsu, Tomoki; Ueda, Koji; Inazawa, Johji

    2016-01-01

    Metastasis is associated with poor prognosis in cancers. Exosomes, which are packed with RNA and proteins and are released in all biological fluids, are emerging as an important mediator of intercellular communication. However, the function of exosomes remains poorly understood in cancer metastasis. Here, we demonstrate that exosomes isolated by size-exclusion chromatography from a highly metastatic human oral cancer cell line, HOC313-LM, induced cell growth through the activation of ERK and AKT as well as promoted cell motility of the poorly metastatic cancer cell line HOC313-P. MicroRNA (miRNA) array analysis identified two oncogenic miRNAs, miR-342–3p and miR-1246, that were highly expressed in exosomes. These miRNAs were transferred to poorly metastatic cells by exosomes, which resulted in increased cell motility and invasive ability. Moreover, miR-1246 increased cell motility by directly targeting DENN/MADD Domain Containing 2D (DENND2D). Taken together, our findings support the metastatic role of exosomes and exosomal miRNAs, which highlights their potential for applications in miRNA-based therapeutics. PMID:27929118

  1. Exosomes in cancer: small particle, big player.

    Science.gov (United States)

    Zhang, Xu; Yuan, Xiao; Shi, Hui; Wu, Lijun; Qian, Hui; Xu, Wenrong

    2015-07-10

    Exosomes have emerged as a novel mode of intercellular communication. Exosomes can shuttle bioactive molecules including proteins, DNA, mRNA, as well as non-coding RNAs from one cell to another, leading to the exchange of genetic information and reprogramming of the recipient cells. Increasing evidence suggests that tumor cells release excessive amount of exosomes, which may influence tumor initiation, growth, progression, metastasis, and drug resistance. In addition, exosomes transfer message from tumor cells to immune cells and stromal cells, contributing to the escape from immune surveillance and the formation of tumor niche. In this review, we highlight the recent advances in the biology of exosomes as cancer communicasomes. We review the multifaceted roles of exosomes, the small secreted particles, in communicating with other cells within tumor microenvironment. Given that exosomes are cell type specific, stable, and accessible from body fluids, exosomes may provide promising biomarkers for cancer diagnosis and represent new targets for cancer therapy.

  2. Therapeutic application of mesenchymal stem cell-derived exosomes: A promising cell-free therapeutic strategy in regenerative medicine.

    Science.gov (United States)

    Motavaf, M; Pakravan, K; Babashah, S; Malekvandfard, F; Masoumi, M; Sadeghizadeh, M

    2016-06-30

    Mesenchymal stem cells have emerged as promising therapeutic candidates in regenerative medicine. The mechanisms underlying mesenchymal stem cells regenerative properties were initially attributed to their engraftment in injured tissues and their subsequent transdifferentiation to repair and replace damaged cells. However, studies in animal models and patients indicated that the low number of transplanted mesenchymal stem cells localize to the target tissue and transdifferentiate to appropriate cell lineage. Instead the regenerative potential of mesenchymal stem cells has been found - at least in part - to be mediated via their paracrine actions. Recently, a secreted group of vesicles, called "exosome" has been identified as major mediator of mesenchymal stem cells therapeutic efficacy. In this review, we will summarize the current literature on administration of exosomes released by mesenchymal stem cells in regenerative medicine and suggest how they could help to improve tissue regeneration following injury.

  3. Exosomes in Cancer Disease.

    Science.gov (United States)

    Zöller, Margot

    2016-01-01

    Cancer diagnosis and therapy is steadily improving. Still, diagnosis is frequently late and diagnosis and follow-up procedures mostly are time-consuming and expensive. Searching for tumor-derived exosomes (TEX) in body fluids may provide an alternative, minimally invasive, yet highly reliable diagnostic tool. Beyond this, there is strong evidence that TEX could become a potent therapeutics. Exosomes, small vesicles delivered by many cells of the organism, are found in all body fluids. Exosomes are characterized by lipid composition, common and donor cell specific proteins, mRNA, small non-coding RNA including miRNA and DNA. Particularly the protein and miRNA markers received much attention as they may allow for highly specific diagnosis and can provide hints toward tumor aggressiveness and progression, where exosome-based diagnosis and follow-up is greatly facilitated by the recovery of exosomes in body fluids, particularly the peripheral blood. Beyond this, exosomes are the most important intercellular communicators that modulate, instruct, and reprogram their surrounding as well as distant organs. In concern about TEX this includes message transfer from tumor cells toward the tumor stroma, the premetastatic niche, the hematopoietic system and, last but not least, the instruction of non-cancer stem cells by cancer-initiating cells (CIC). Taking this into account, it becomes obvious that "tailored" exosomes offer themselves as potent therapeutic delivery system. In brief, during the last 4-5 years there is an ever-increasing, overwhelming interest in exosome research. This boom appears fully justified provided the content of the exosomes becomes most thoroughly analyzed and their mode of intercellular interaction can be unraveled in detail as this knowledge will open new doors toward cancer diagnosis and therapy including immunotherapy and CIC reprogramming.

  4. Altered microRNA expression profile in exosomes during osteogenic differentiation of human bone marrow-derived mesenchymal stem cells.

    Directory of Open Access Journals (Sweden)

    Ji-Feng Xu

    Full Text Available The physiological role of microRNAs (miRNAs in osteoblast differentiation remains elusive. Exosomal miRNAs isolated from human bone marrow-derived mesenchymal stem cells (BMSCs culture were profiled using miRNA arrays containing probes for 894 human matured miRNAs. Seventy-nine miRNAs (∼8.84% could be detected in exosomes isolated from BMSC culture supernatants when normalized to endogenous control genes RNU44. Among them, nine exosomal miRNAs were up regulated and 4 miRNAs were under regulated significantly (Relative fold>2, p<0.05 when compared with the values at 0 day with maximum changes at 1 to 7 days. Five miRNAs (miR-199b, miR-218, miR-148a, miR-135b, and miR-221 were further validated and differentially expressed in the individual exosomal samples from hBMSCs cultured at different time points. Bioinformatic analysis by DIANA-mirPath demonstrated that RNA degradation, mRNA surveillance pathway, Wnt signaling pathway, RNA transport were the most prominent pathways enriched in quantiles with differential exosomal miRNA patterns related to osteogenic differentiation. These data demonstrated exosomal miRNA is a regulator of osteoblast differentiation.

  5. Altered MicroRNA Expression Profile in Exosomes during Osteogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells

    Science.gov (United States)

    Zhang, Shui-Jun; Zhao, Chen; Qiu, Bin-Song; Gu, Hai-Feng; Hong, Jian-Fei; Cao, Li; Chen, Yu; Xia, Bing; Bi, Qin; Wang, Ya-Ping

    2014-01-01

    The physiological role of microRNAs (miRNAs) in osteoblast differentiation remains elusive. Exosomal miRNAs isolated from human bone marrow-derived mesenchymal stem cells (BMSCs) culture were profiled using miRNA arrays containing probes for 894 human matured miRNAs. Seventy-nine miRNAs (∼8.84%) could be detected in exosomes isolated from BMSC culture supernatants when normalized to endogenous control genes RNU44. Among them, nine exosomal miRNAs were up regulated and 4 miRNAs were under regulated significantly (Relative fold>2, p<0.05) when compared with the values at 0 day with maximum changes at 1 to 7 days. Five miRNAs (miR-199b, miR-218, miR-148a, miR-135b, and miR-221) were further validated and differentially expressed in the individual exosomal samples from hBMSCs cultured at different time points. Bioinformatic analysis by DIANA-mirPath demonstrated that RNA degradation, mRNA surveillance pathway, Wnt signaling pathway, RNA transport were the most prominent pathways enriched in quantiles with differential exosomal miRNA patterns related to osteogenic differentiation. These data demonstrated exosomal miRNA is a regulator of osteoblast differentiation. PMID:25503309

  6. Biomolecular characterization of exosomes released from cancer stem cells: Possible implications for biomarker and treatment of cancer.

    Science.gov (United States)

    Kumar, Dhruv; Gupta, Dwijendra; Shankar, Sharmila; Srivastava, Rakesh K

    2015-02-20

    Cancer recognized as one of the leading irrepressible health issues is contributing to increasing mortality-rate day-by-day. The tumor microenvironment is an important field of cancer to understand the detection, treatment and prevention of cancer. Recently, cancer stem cell (CSC) research has shown promising results aiming towards cancer diagnostics and treatment. Here, we found that prostate and breast cancer stem cells secreted vesicles of endosomal origin, called exosomes showed strong connection between autophagy and exosomes released from CSCs. Exosomes may serve as vesicles to communicate with neoplastic cells (autocrine and paracrine manner) and normal cells (paracrine and endocrine manner) and thereby suppress immune systems and regulate neoplastic growth, and metastasis. They can also be used as biomarkers for various cancers. We detected tetraspanin proteins (CD9, CD63, CD81), Alix and tumor susceptibility gene-101 (TSG101) of exosomal markers from rotenone treated CSCs. We have also detected the induction of autophagy genes, Atg7 and conversion of autophagy marker (LC3-I to LC3-II), and tetraspanin proteins (CD9, CD63, CD81) in rotenone treated CSCs by western blotting. The mRNA expression of CD9, CD63, CD81 and TSG101 analyzed by qRT-PCR showed that the rotenone induced the expression of CD9, CD63, CD81 and TSG101 in CSCs. Electron microscopy of rotenone treated CSCs showed the mitochondrial damage of CSCs as confirmed by the release of exosomes from CSCs. The constituents of exosomes may be useful to understand the mechanism of exosomes formation, release and function, and also serve as a useful biomarker and provide novel therapeutic strategies for the treatment and prevention of cancer.

  7. Macrophages play an essential role in antigen-specific immune suppression mediated by T CD8⁺ cell-derived exosomes.

    Science.gov (United States)

    Nazimek, Katarzyna; Ptak, Wlodzimierz; Nowak, Bernadeta; Ptak, Maria; Askenase, Philip W; Bryniarski, Krzysztof

    2015-09-01

    Murine contact sensitivity (CS) reaction could be antigen-specifically regulated by T CD8(+) suppressor (Ts) lymphocytes releasing microRNA-150 in antibody light-chain-coated exosomes that were formerly suggested to suppress CS through action on macrophages (Mφ). The present studies investigated the role of Mφ in Ts cell-exosome-mediated antigen-specific suppression as well as modulation of Mφ antigen-presenting function in humoral and cellular immunity by suppressive exosomes. Mice depleted of Mφ by clodronate liposomes could not be tolerized and did not produce suppressive exosomes. Moreover, isolated T effector lymphocytes transferring CS were suppressed by exosomes only in the presence of Mφ, demonstrating the substantial role of Mφ in the generation and action of Ts cell regulatory exosomes. Further, significant decrease of number of splenic B cells producing trinitrophenyl (TNP) -specific antibodies with the alteration of the ratio of serum titres of IgM to IgG was observed in recipients of exosome-treated, antigen-pulsed Mφ and the significant suppression of CS was demonstrated in recipients of exosome-treated, TNP-conjugated Mφ. Additionally, exosome-pulsed, TNP-conjugated Mφ mediated suppression of CS in mice pre-treated with a low-dose of cyclophosphamide, suggesting de novo induction of T regulatory (Treg) lymphocytes. Treg cell involvement in the effector phase of the studied suppression mechanism was proved by unsuccessful tolerization of DEREG mice depleted of Treg lymphocytes. Furthermore, the inhibition of proliferation of CS effector cells cultured with exosome-treated Mφ in a transmembrane manner was observed. Our results demonstrated the essential role of Mφ in antigen-specific immune suppression mediated by Ts cell-derived exosomes and realized by induction of Treg lymphocytes and inhibition of T effector cell proliferation.

  8. Proteolytic factors in exosomes.

    Science.gov (United States)

    Shimoda, Masayuki; Khokha, Rama

    2013-05-01

    Exosomes are small microvesicles secreted from the late endosomal compartment of cells. Although an increasing body of evidence indicates that they play a pivotal role in cell-to-cell communication, the biological functions of exosomes are far from fully understood. Recent work has revealed detailed proteomic profiles of exosomes from cell lines and body fluids, which may provide clues to understanding their biological significance and general importance in human diseases. Metalloproteinases include the cell surface-anchored sheddases a disintegrin and metalloproteinases, as well as cell surface-bound and soluble matrix metalloproteinases and these extracellular proteases have been detected in exosomes by proteomic analyses. Exosomes play a key role in the transfer of proteins to other cells and metalloproteinases may provide a novel platform where ectodomain shedding by these membrane proteases alters the makeup of the recipient cell's surface. This review aims to address some of the facets of exosome biology with particular emphasis on the proteolytic factors and we discuss their potential involvement in human diseases, especially tumor biology.

  9. Tumor-stroma interaction: Revealing fibroblast-secreted exosomes as potent regulators of Wnt-planar cell polarity signaling in cancer metastasis.

    Science.gov (United States)

    Luga, Valbona; Wrana, Jeffrey L

    2013-12-01

    Cancer-associated fibroblasts (CAF) regulate tumor progression, but their role in cancer metastasis remains largely unexplored. Exosomes are secreted microvesicles that are emerging as potent mediators of cell-cell communication that are of particular importance in tumor-stroma interactions. The Wnt-planar cell polarity (PCP) pathway is the primary regulator of convergent extension cell movements during vertebrate development, but the role of this signaling pathway in cancer cell migration and metastasis has been unclear. Recently, we revealed that fibroblasts secrete exosomes that promote breast cancer cell (BCC) protrusive activity, motility, and metastasis by activating autocrine Wnt-PCP signaling in BCCs. Moreover, we showed that Wnt ligands produced by BCCs tether to fibroblast exosomes upon trafficking of exosomes in BCCs. These findings have several implications that motivate promising future research in the fields of tumor-stroma communication, exosome function, and Wnt-PCP signaling in cancer metastasis.

  10. 外泌体(Exosome)及其在肿瘤调控中的作用%Exosome and Its Roles in Regulation of Tumor Cell

    Institute of Scientific and Technical Information of China (English)

    元小宁; 朱运峰

    2013-01-01

    Exosome is a kind of small membranous vesicles secreted by numerous of cells including reticulocytes,cytotoxic T lymphocytes,B lymphocytes,dendritic cells,mast cells,platelets,epithelial cells and tumor cells.It can be isolated from the media of cultured cell or bodily fluids such as urine and plasma.Its biogenesis includes the inward budding of endosomes,which form multivesicles bodies (MVB),and releasing of the vesicle into the extracellular environment by fusing with the plasma membrane.As a inter-cell commumicator,exosome is considered to participate not only in physiological regulation such as cell-to-cell communication and genetic reprogramming of their target cells,but also in pathological regulation such as metastasis of cancer.Notably,among the contents in exosome,some long non-coding RNAs (LncRNAs) were identified,which gives a clue that LncRNAs protected by exosome from degradation may conduct some important functions for cell regulation.%外泌体是多种活细胞分泌的直径约为40~l00nm的小囊泡体,分布于外周血、尿液、唾液、腹水、羊水等体液中.外泌体的来源多样,可以来自各种类型细胞,其内容物含蛋白、脂质和核酸等,特别是在其中发现的Long non-coding RNA使其更具有意义,其内容物种类和数量与低氧和酸碱微环境相关.外泌体在免疫监视、炎症反应及癌症发生发展等许多生理和病理过程中有重要的功能,尤其在细胞间交流、对靶细胞中的基因重编码以及肿瘤的发生发展和侵袭转移中具有重要作用.从外泌体的起源、种类及其在肿瘤发生发展和转移相关的研究方面进行综述,对其深入研究可为治疗肿瘤提供新的思路.

  11. Exosomes in cancer: small particle, big player

    OpenAIRE

    2015-01-01

    Exosomes have emerged as a novel mode of intercellular communication. Exosomes can shuttle bioactive molecules including proteins, DNA, mRNA, as well as non-coding RNAs from one cell to another, leading to the exchange of genetic information and reprogramming of the recipient cells. Increasing evidence suggests that tumor cells release excessive amount of exosomes, which may influence tumor initiation, growth, progression, metastasis, and drug resistance. In addition, exosomes transfer messag...

  12. Effect of exosome isolation methods on physicochemical properties of exosomes and clearance of exosomes from the blood circulation.

    Science.gov (United States)

    Yamashita, Takuma; Takahashi, Yuki; Nishikawa, Makiya; Takakura, Yoshinobu

    2016-01-01

    Exosomes, which are expected to be delivery systems for biomolecules such as nucleic acids, are collected by several methods. However, the effect of exosome isolation methods on the characteristics of exosomes as drug carriers, such as recovery efficiency after sterile filtration and pharmacokinetics, has not been investigated despite the importance of these characteristics for the development of exosome-based delivery systems. In the present study, exosomes collected from murine melanoma B16-BL6 cells by several methods were compared with respect to dispersibility, recovery rate after filtering, and clearance from the blood circulation in mice. The exosomes were collected by three ultracentrifugation-based methods: simple ultracentrifugation/pelleting (pelleting method), ultracentrifugation with an iodixanol cushion (cushion method), and ultracentrifugation on an iodixanol density gradient (gradient method). The isolation methods had little effect on the particle number of exosomes. In contrast, transmission electron microscopy observation and size distribution measurement using tunable resistive pulse sensing indicated that the exosomes of the gradient method were more dispersed than the others. The exosomes were labeled with Gaussia luciferase and intravenously injected into mice. Clearance of injected exosomes from the blood circulation did not significantly change with isolation methods. When the exosomes were filtered using a 0.2-μm filter, the recovery rate was 82% for the exosomes of the gradient method, whereas it was less than 50% for the others. These results indicate that the exosome isolation method markedly affects the dispersibility and filtration efficiency of the exosomes.

  13. [Proteomic analysis of urinary exosomes].

    Science.gov (United States)

    Nakayama, Aki

    2014-07-01

    Exosomes are 40-100-nm membrane vesicles secreted into the extracellular space by various types of cell in many biological fluids, including serum, saliva, breast milk, amniotic fluid, and urine. Exosomes, which contain several key proteins, lipids, mRNAs, and microRNAs, were considered as an alternative secretion pathway. In addition, recent findings suggest that the exosome itself is a functional biomolecule involved in intracellular communication; thus, its components can be transferred to recipient cells by fusion, changing the function of the target cell. Recently, urinary exosomes have attracted much attention because some of their proteins have been identified as biomarkers related to certain physiological events and disease-related metabolism of the kidney. This review provides an overview of urinary exosomes, including methods of isolation and associated problems, and focuses on urinary exosomes as protein biomarker sources involved in numerous physiological and pathophysiological processes.

  14. Vaccination of metastatic melanoma patients with autologous dendritic cell (DC derived-exosomes: results of thefirst phase I clinical trial

    Directory of Open Access Journals (Sweden)

    Piperno Sophie

    2005-03-01

    Full Text Available Abstract Background DC derived-exosomes are nanomeric vesicles harboring functional MHC/peptide complexes capable of promoting T cell immune responses and tumor rejection. Here we report the feasability and safety of the first Phase I clinical trial using autologous exosomes pulsed with MAGE 3 peptides for the immunization of stage III/IV melanoma patients. Secondary endpoints were the monitoring of T cell responses and the clinical outcome. Patients and methods Exosomes were purified from day 7 autologous monocyte derived-DC cultures. Fifteen patients fullfilling the inclusion criteria (stage IIIB and IV, HLA-A1+, or -B35+ and HLA-DPO4+ leukocyte phenotype, tumor expressing MAGE3 antigen were enrolled from 2000 to 2002 and received four exosome vaccinations. Two dose levels of either MHC class II molecules (0.13 versus 0.40 × 1014 molecules or peptides (10 versus 100 μg/ml were tested. Evaluations were performed before and 2 weeks after immunization. A continuation treatment was performed in 4 cases of non progression. Results The GMP process allowed to harvest about 5 × 1014 exosomal MHC class II molecules allowing inclusion of all 15 patients. There was no grade II toxicity and the maximal tolerated dose was not achieved. One patient exhibited a partial response according to the RECIST criteria. This HLA-B35+/A2+ patient vaccinated with A1/B35 defined CTL epitopes developed halo of depigmentation around naevi, a MART1-specific HLA-A2 restricted T cell response in the tumor bed associated with progressive loss of HLA-A2 and HLA-BC molecules on tumor cells during therapy with exosomes. In addition, one minor, two stable and one mixed responses were observed in skin and lymph node sites. MAGE3 specific CD4+ and CD8+ T cell responses could not be detected in peripheral blood. Conclusion The first exosome Phase I trial highlighted the feasibility of large scale exosome production and the safety of exosome administration.

  15. Exosomes contribute to the transmission of anti-HIV activity from TLR3-activated brain microvascular endothelial cells to macrophages

    Science.gov (United States)

    Sun, Li; Wang, Xu; Zhou, Yu; Zhou, Run-Hong; Ho, Wen-Zhe; Li, Jie-Liang

    2017-01-01

    Human brain microvascular endothelial cells (HBMECs), the major cell type in the blood-brain barrier (BBB), play a key role in maintaining brain homeostasis. However, their role in the BBB innate immunity against HIV invasion of the central nervous system (CNS) remains to be determined. Our early work showed that TLR3 signaling of HBMECs could produce the antiviral factors that inhibit HIV replication in macrophages. The present study examined whether exosomes from TLR3-activated HBMECs mediate the intercellular transfer of antiviral factors to macrophages. Primary human macrophages could take up exosomes from TLR3-activated HBMECs. HBMECs-derived exosomes contained multiple antiviral factors, including several key IFN-stimulated genes (ISGs; ISG15, ISG56, and Mx2) at mRNA and protein levels. The depletion of exosomes from TLR3-activated HBMECs culture supernatant diminished HBMECs-mediated anti-HIV activity in macrophages. In conclusion, we demonstrate that exosomes shed by HBMECs are able to transport the antiviral molecules to macrophages. This finding suggests the possibility that HIV nonpermissive BBB cells (HBMECs) can help to restore the antiviral state in HIV-infected macrophages, which may be a defense mechanism against HIV neuroinvasion. PMID:27496004

  16. Quantitative proteomics of fractionated membrane and lumen exosome proteins from isogenic metastatic and nonmetastatic bladder cancer cells reveal differential expression of EMT factors

    DEFF Research Database (Denmark)

    Jeppesen, Dennis Kjølhede; Nawrocki, Arkadiusz; Jensen, Steffen Grann

    2014-01-01

    Cancer cells secrete soluble factors and various extracellular vesicles, including exosomes, into their tissue microenvironment. The secretion of exosomes is speculated to facilitate local invasion and metastatic spread. Here, we used an in vivo metastasis model of human bladder carcinoma cell line...... T24 without metastatic capacity and its two isogenic derivate cell lines SLT4 and FL3, which form metastases in the lungs and liver of mice, respectively. Cultivation in CLAD1000 bioreactors rather than conventional culture flasks resulted in a 13-16-fold increased exosome yield and facilitated...... quantitative proteomics of fractionated exosomes. Exosomes from T24, SLT4, and FL3 cells were partitioned into membrane and luminal fractions and changes in protein abundance related to the gain of metastatic capacity were identified by quantitative iTRAQ- proteomics. We identified several proteins linked...

  17. Quantitative proteomics of fractionated membrane and lumen exosome proteins from isogenic metastatic and nonmetastatic bladder cancer cells reveal differential expression of EMT factors.

    Science.gov (United States)

    Jeppesen, Dennis Kjølhede; Nawrocki, Arkadiusz; Jensen, Steffen Grann; Thorsen, Kasper; Whitehead, Bradley; Howard, Kenneth A; Dyrskjøt, Lars; Ørntoft, Torben Falck; Larsen, Martin R; Ostenfeld, Marie Stampe

    2014-03-01

    Cancer cells secrete soluble factors and various extracellular vesicles, including exosomes, into their tissue microenvironment. The secretion of exosomes is speculated to facilitate local invasion and metastatic spread. Here, we used an in vivo metastasis model of human bladder carcinoma cell line T24 without metastatic capacity and its two isogenic derivate cell lines SLT4 and FL3, which form metastases in the lungs and liver of mice, respectively. Cultivation in CLAD1000 bioreactors rather than conventional culture flasks resulted in a 13- to 16-fold increased exosome yield and facilitated quantitative proteomics of fractionated exosomes. Exosomes from T24, SLT4, and FL3 cells were partitioned into membrane and luminal fractions and changes in protein abundance related to the gain of metastatic capacity were identified by quantitative iTRAQ proteomics. We identified several proteins linked to epithelial-mesenchymal transition, including increased abundance of vimentin and hepatoma-derived growth factor in the membrane, and casein kinase II α and annexin A2 in the lumen of exosomes, respectively, from metastatic cells. The change in exosome protein abundance correlated little, although significant for FL3 versus T24, with changes in cellular mRNA expression. Our proteomic approach may help identification of proteins in the membrane and lumen of exosomes potentially involved in the metastatic process.

  18. Exosomes released from M. tuberculosis infected cells can suppress IFN-γ mediated activation of naive macrophages.

    Directory of Open Access Journals (Sweden)

    Prachi P Singh

    Full Text Available BACKGROUND: Macrophages infected with Mycobacterium tuberculosis (M.tb are known to be refractory to IFN-γ stimulation. Previous studies have shown that M.tb express components such as the 19-kDa lipoprotein and peptidoglycan that can bind to macrophage receptors including the Toll-like receptor 2 resulting in the loss in IFN-γ responsiveness. However, it is unclear whether this effect is limited to infected macrophages. We have previously shown that M.tb-infected macrophages release exosomes which are 30-100 nm membrane bound vesicles of endosomal origin that function in intercellular communication. These exosomes contain mycobacterial components including the 19-kDa lipoprotein and therefore we hypothesized that macrophages exposed to exosomes may show limited response to IFN-γ stimulation. METHODOLOGY/PRINCIPAL FINDINGS: Exosomes were isolated from resting as well as M.tb-infected RAW264.7 macrophages. Mouse bone marrow-derived macrophages (BMMØ were treated with exosomes +/- IFN-γ. Cells were harvested and analyzed for suppression of IFN-γ responsive genes by flow cytometry and real time PCR. We found that exosomes derived from M.tb H37Rv-infected but not from uninfected macrophages inhibited IFN-γ induced MHC class II and CD64 expression on BMMØ. This inhibition was only partially dependent on the presence of lipoproteins but completely dependent on TLR2 and MyD88. The exosomes isolated from infected cells did not inhibit STAT1 Tyrosine phosphorylation but down-regulated IFN-γ induced expression of the class II major histocompatibility complex transactivator; a key regulator of class II MHC expression. Microarray studies showed that subsets of genes induced by IFN-γ were inhibited by exosomes from H37Rv-infected cells including genes involved in antigen presentation. Moreover, this set of genes partially overlapped with the IFN-γ-induced genes inhibited by H37Rv infection. CONCLUSIONS: Our study suggests that exosomes, as

  19. Nano-plasmonic exosome diagnostics

    OpenAIRE

    Im, Hyungsoon; Shao, Huilin; Weissleder, Ralph; Castro, Cesar M.; Lee, Hakho

    2015-01-01

    Exosomes have emerged as a promising biomarker. These vesicles abound in biofluids and harbor molecular constituents from their parent cells, thereby offering a minimally-invasive avenue for molecular analyses. Despite such clinical potential, routine exosomal analysis, particularly the protein assay, remains challenging, due to requirements for large sample volumes and extensive processing. We have been developing miniaturized systems to facilitate clinical exosome studies. These systems can...

  20. Exosomes as therapeutics: The implications of molecular composition and exosomal heterogeneity.

    Science.gov (United States)

    Ferguson, Scott W; Nguyen, Juliane

    2016-04-28

    Harnessing exosomes as therapeutic drug delivery vehicles requires a better understanding of exosomal composition and their mode of action. A full appreciation of all the exosomal components (proteins, lipids, and RNA content) will be important for the design of effective exosome-based or exosome-mimicking drug carriers. In this review we describe the presence of rarely studied, non-coding RNAs that exist in high numbers in exosomes. We discuss the implications of the molecular composition and heterogeneity of exosomes on their biological and therapeutic effects. Finally, we highlight outstanding questions with regard to RNA loading into exosomes, analytical methods to sort exosomes and their sub-populations, and the effects of exosomal proteins and lipids on recipient cells. Investigations into these facets of exosome biology will further advance the field, could lead to the clinical translation of exosome-based therapeutics, and aid in the reverse-engineering of synthetic exosomes. Although synthetic exosomes are still an underexplored area, they could offer researchers a way to manufacture exosomes with highly defined structure, composition, and function.

  1. Interactions between exosomes from breast cancer cells and primary mammary epithelial cells leads to generation of reactive oxygen species which induce DNA damage response, stabilization of p53 and autophagy in epithelial cells.

    Directory of Open Access Journals (Sweden)

    Sujoy Dutta

    Full Text Available Exosomes are nanovesicles originating from multivesicular bodies and are released by all cell types. They contain proteins, lipids, microRNAs, mRNAs and DNA fragments, which act as mediators of intercellular communications by inducing phenotypic changes in recipient cells. Tumor-derived exosomes have been shown to play critical roles in different stages of tumor development and metastasis of almost all types of cancer. One of the ways by which exosomes affect tumorigenesis is to manipulate the tumor microenvironments to create tumor permissive "niches". Whether breast cancer cell secreted exosomes manipulate epithelial cells of the mammary duct to facilitate tumor development is not known. To address whether and how breast cancer cell secreted exosomes manipulate ductal epithelial cells we studied the interactions between exosomes isolated from conditioned media of 3 different breast cancer cell lines (MDA-MB-231, T47DA18 and MCF7, representing three different types of breast carcinomas, and normal human primary mammary epithelial cells (HMECs. Our studies show that exosomes released by breast cancer cell lines are taken up by HMECs, resulting in the induction of reactive oxygen species (ROS and autophagy. Inhibition of ROS by N-acetyl-L-cysteine (NAC led to abrogation of autophagy. HMEC-exosome interactions also induced the phosphorylation of ATM, H2AX and Chk1 indicating the induction of DNA damage repair (DDR responses. Under these conditions, phosphorylation of p53 at serine 15 was also observed. Both DDR responses and phosphorylation of p53 induced by HMEC-exosome interactions were also inhibited by NAC. Furthermore, exosome induced autophagic HMECs were found to release breast cancer cell growth promoting factors. Taken together, our results suggest novel mechanisms by which breast cancer cell secreted exosomes manipulate HMECs to create a tumor permissive microenvironment.

  2. Interactions between exosomes from breast cancer cells and primary mammary epithelial cells leads to generation of reactive oxygen species which induce DNA damage response, stabilization of p53 and autophagy in epithelial cells.

    Science.gov (United States)

    Dutta, Sujoy; Warshall, Case; Bandyopadhyay, Chirosree; Dutta, Dipanjan; Chandran, Bala

    2014-01-01

    Exosomes are nanovesicles originating from multivesicular bodies and are released by all cell types. They contain proteins, lipids, microRNAs, mRNAs and DNA fragments, which act as mediators of intercellular communications by inducing phenotypic changes in recipient cells. Tumor-derived exosomes have been shown to play critical roles in different stages of tumor development and metastasis of almost all types of cancer. One of the ways by which exosomes affect tumorigenesis is to manipulate the tumor microenvironments to create tumor permissive "niches". Whether breast cancer cell secreted exosomes manipulate epithelial cells of the mammary duct to facilitate tumor development is not known. To address whether and how breast cancer cell secreted exosomes manipulate ductal epithelial cells we studied the interactions between exosomes isolated from conditioned media of 3 different breast cancer cell lines (MDA-MB-231, T47DA18 and MCF7), representing three different types of breast carcinomas, and normal human primary mammary epithelial cells (HMECs). Our studies show that exosomes released by breast cancer cell lines are taken up by HMECs, resulting in the induction of reactive oxygen species (ROS) and autophagy. Inhibition of ROS by N-acetyl-L-cysteine (NAC) led to abrogation of autophagy. HMEC-exosome interactions also induced the phosphorylation of ATM, H2AX and Chk1 indicating the induction of DNA damage repair (DDR) responses. Under these conditions, phosphorylation of p53 at serine 15 was also observed. Both DDR responses and phosphorylation of p53 induced by HMEC-exosome interactions were also inhibited by NAC. Furthermore, exosome induced autophagic HMECs were found to release breast cancer cell growth promoting factors. Taken together, our results suggest novel mechanisms by which breast cancer cell secreted exosomes manipulate HMECs to create a tumor permissive microenvironment.

  3. Quantitative proteomic analysis of exosome protein content changes induced by hepatitis B virus in Huh-7 cells using SILAC labeling and LC-MS/MS.

    Science.gov (United States)

    Zhao, Xue; Wu, Yanxin; Duan, Jinlin; Ma, Yanchun; Shen, Zhongliang; Wei, Lili; Cui, Xiaoxian; Zhang, Junqi; Xie, Youhua; Liu, Jing

    2014-12-05

    Hepatitis B virus (HBV) infection could cause hepatitis, liver cirrhosis, and hepatocellular carcinoma. HBV-mediated pathogenesis is only partially understood, but X protein (HBx) reportedly possesses oncogenic potential. Exosomes are small membrane vesicles with diverse functions released by various cells including hepatocytes, and HBV harnesses cellular exosome biogenesis and export machineries for virion morphogenesis and secretion. Therefore, HBV infection might cause changes in exosome contents with functional implications for both virus and host. In this work, exosome protein content changes induced by HBV and HBx were quantitatively analyzed by SILAC/LC-MS/MS. Exosomes prepared from SILAC-labeled hepatoma cell line Huh-7 transfected with HBx, wildtype, or HBx-null HBV replicon plasmids were analyzed by LC-MS/MS. Systematic analyses of MS data and confirmatory immunoblotting showed that HBx overexpression and HBV, with or without HBx, replication in Huh-7 cells indeed caused marked and specific changes in exosome protein contents. Furthermore, specific changes in protein contents were also detected in exosomes purified from HBV-infected patients' sera compared with control sera negative for HBV markers. These results illustrate a new aspect of interactions between HBV and the host and provide the foundation for future research into roles played by exosomes in HBV infection and pathogenesis.

  4. Nano-plasmonic exosome diagnostics.

    Science.gov (United States)

    Im, Hyungsoon; Shao, Huilin; Weissleder, Ralph; Castro, Cesar M; Lee, Hakho

    2015-06-01

    Exosomes have emerged as a promising biomarker. These vesicles abound in biofluids and harbor molecular constituents from their parent cells, thereby offering a minimally-invasive avenue for molecular analyses. Despite such clinical potential, routine exosomal analysis, particularly the protein assay, remains challenging, due to requirements for large sample volumes and extensive processing. We have been developing miniaturized systems to facilitate clinical exosome studies. These systems can be categorized into two components: microfluidics for sample preparation and analytical tools for protein analyses. In this report, we review a new assay platform, nano-plasmonic exosome, in which sensing is based on surface plasmon resonance to achieve label-free exosome detection. Looking forward, we also discuss some potential challenges and improvements in exosome studies.

  5. Exosomes in development, metastasis and drug resistance of breast cancer

    OpenAIRE

    2015-01-01

    Transport through the cell membrane can be divided into active, passive and vesicular types (exosomes). Exosomes are nano-sized vesicles released by a variety of cells. Emerging evidence shows that exosomes play a critical role in cancers. Exosomes mediate communication between stroma and cancer cells through the transfer of nucleic acid and proteins. It is demonstrated that the contents and the quantity of exosomes will change after occurrence of cancers. Over the last decade, growing attent...

  6. Comparative proteomics of exosomes secreted by tumoral Jurkat T cells and normal human T cell blasts unravels a potential tumorigenic role for valosin-containing protein

    Science.gov (United States)

    Sanclemente, Manuel; Iturralde, María; Naval, Javier; Alava, María Angeles; Martínez-Lostao, Luis; Thierse, Hermann-Josef; Anel, Alberto

    2016-01-01

    We have previously characterized that FasL and Apo2L/TRAIL are stored in their bioactive form inside human T cell blasts in intraluminal vesicles present in multivesicular bodies. These vesicles are rapidly released to the supernatant in the form of exosomes upon re-activation of T cells. In this study we have compared for the first time proteomics of exosomes produced by normal human T cell blasts with those produced by tumoral Jurkat cells, with the objective of identify proteins associated with tumoral exosomes that could have a previously unrecognized role in malignancy. We have identified 359 and 418 proteins in exosomes from T cell blasts and Jurkat cells, respectively. Interestingly, only 145 (around a 40%) are common. The major proteins in both cases are actin and tubulin isoforms and the common interaction nodes correspond to these cytoskeleton and related proteins, as well as to ribosomal and mRNA granule proteins. We detected 14 membrane proteins that were especially enriched in exosomes from Jurkat cells as compared with T cell blasts. The most abundant of these proteins was valosin-containing protein (VCP), a membrane ATPase involved in ER homeostasis and ubiquitination. In this work, we also show that leukemic cells are more sensitive to cell death induced by the VCP inhibitor DBeQ than normal T cells. Furthermore, VCP inhibition prevents functional exosome secretion only in Jurkat cells, but not in T cell blasts. These results suggest VCP targeting as a new selective pathway to exploit in cancer treatment to prevent tumoral exosome secretion. PMID:27086912

  7. Noncytotoxic suppression of human immunodeficiency virus type 1 transcription by exosomes secreted from CD8+ T cells.

    Science.gov (United States)

    Tumne, Ashwin; Prasad, Varsha Shridhar; Chen, Yue; Stolz, Donna B; Saha, Kunal; Ratner, Deena M; Ding, Ming; Watkins, Simon C; Gupta, Phalguni

    2009-05-01

    CD8(+) T cells display a noncytotoxic activity that suppresses transcription of human immunodeficiency virus type 1 (HIV-1) in an antigen-independent and major histocompatibility complex-unrestricted manner. To date, the precise cellular and molecular factors mediating this CD8(+) T-cell effector function remain unsolved. Despite evidence indicating the dependence of the activity on cell-cell contact, the possibility of a membrane-mediated activity that represses transcription from the viral promoter remains unexplored. We therefore investigated whether this inhibition of HIV-1 transcription might be elicited by a membrane-bound determinant. Using a CD8(+) T-cell line displaying potent noncytotoxic HIV-1 suppression activity, we have identified a membrane-localized HIV-1-suppressing activity that is concomitantly secreted as 30- to 100-nm endosome-derived tetraspanin-rich vesicles known as exosomes. Purified exosomes from CD8(+) T-cell culture supernatant noncytotoxically suppressed CCR5-tropic (R5) and CXCR4-tropic (X4) replication of HIV-1 in vitro through a protein moiety. Similar antiviral activity was also found in exosomes isolated from two HIV-1-infected subjects. The antiviral exosomes specifically inhibited HIV-1 transcription in both acute and chronic models of infection. Our results, for the first time, indicate the existence of an antiviral membrane-bound factor consistent with the hallmarks defining noncytotoxic CD8(+) T-cell suppression of HIV-1.

  8. Dendritic cells derived exosomes migration to spleen and induction of inflammation are regulated by CCR7

    Science.gov (United States)

    Wei, Gao; Jie, Yuan; Haibo, Liu; Chaoneng, Wu; Dong, Huang; Jianbing, Zhu; Junjie, Guo; Leilei, Ma; Hongtao, Shi; Yunzeng, Zou; Junbo, Ge

    2017-01-01

    Mature dendritic cells (DCs) home to secondary lymphoid organs through CC chemokine receptor 7 (CCR7). Exosomes derived from DCs (DC-exos) are reported to migrate to spleen and induce inflammation in vivo. In this study, we demonstrated that mature bone marrow DC-exos can activate immature DC and T cells in vitro. Then we intravenously injected DC-exos into C57BL/6 mice, observing that mature DC-exos accumulated more in spleen than immature DC-exos. These DC-exos in spleen could be uptaken by splenetic DCs and T cells and induce an inflammatory response. We further showed that the increased accumulation of mature DC-exos in spleen was regulated by CCR7, whose reduction led to a decrease of accumulation in spleen and attenuated inflammatory response in serum. These data provide us a new perspective to comprehensively understand exosomes, which might inherit some special functions from their parent cells and exert these functions in vivo. PMID:28223684

  9. Exosome and Exosomal MicroRNA:Trafficking, Sorting, and Function

    Institute of Scientific and Technical Information of China (English)

    Jian Zhang; Sha Li; Lu Li; Meng Li; Chongye Guo; Jun Yao; Shuangli Mi

    2015-01-01

    Exosomes are 40–100 nm nano-sized vesicles that are released from many cell types into the extracellular space. Such vesicles are widely distributed in various body fluids. Recently, mRNAs and microRNAs (miRNAs) have been identified in exosomes, which can be taken up by neighboring or distant cells and subsequently modulate recipient cells. This suggests an active sort-ing mechanism of exosomal miRNAs, since the miRNA profiles of exosomes may differ from those of the parent cells. Exosomal miRNAs play an important role in disease progression, and can stimu-late angiogenesis and facilitate metastasis in cancers. In this review, we will introduce the origin and the trafficking of exosomes between cells, display current research on the sorting mechanism of exo-somal miRNAs, and briefly describe how exosomes and their miRNAs function in recipient cells. Finally, we will discuss the potential applications of these miRNA-containing vesicles in clinical settings.

  10. Exosome and exosomal microRNA: trafficking, sorting, and function.

    Science.gov (United States)

    Zhang, Jian; Li, Sha; Li, Lu; Li, Meng; Guo, Chongye; Yao, Jun; Mi, Shuangli

    2015-02-01

    Exosomes are 40-100 nm nano-sized vesicles that are released from many cell types into the extracellular space. Such vesicles are widely distributed in various body fluids. Recently, mRNAs and microRNAs (miRNAs) have been identified in exosomes, which can be taken up by neighboring or distant cells and subsequently modulate recipient cells. This suggests an active sorting mechanism of exosomal miRNAs, since the miRNA profiles of exosomes may differ from those of the parent cells. Exosomal miRNAs play an important role in disease progression, and can stimulate angiogenesis and facilitate metastasis in cancers. In this review, we will introduce the origin and the trafficking of exosomes between cells, display current research on the sorting mechanism of exosomal miRNAs, and briefly describe how exosomes and their miRNAs function in recipient cells. Finally, we will discuss the potential applications of these miRNA-containing vesicles in clinical settings.

  11. Exosome and Exosomal MicroRNA: Trafficking, Sorting, and Function

    Directory of Open Access Journals (Sweden)

    Jian Zhang

    2015-02-01

    Full Text Available Exosomes are 40–100 nm nano-sized vesicles that are released from many cell types into the extracellular space. Such vesicles are widely distributed in various body fluids. Recently, mRNAs and microRNAs (miRNAs have been identified in exosomes, which can be taken up by neighboring or distant cells and subsequently modulate recipient cells. This suggests an active sorting mechanism of exosomal miRNAs, since the miRNA profiles of exosomes may differ from those of the parent cells. Exosomal miRNAs play an important role in disease progression, and can stimulate angiogenesis and facilitate metastasis in cancers. In this review, we will introduce the origin and the trafficking of exosomes between cells, display current research on the sorting mechanism of exosomal miRNAs, and briefly describe how exosomes and their miRNAs function in recipient cells. Finally, we will discuss the potential applications of these miRNA-containing vesicles in clinical settings.

  12. Paracrine Induction of Endothelium by Tumor Exosomes

    OpenAIRE

    2009-01-01

    Cancers utilize a nanoscale messenger system known as exosomes to communicate with surrounding tissues and immune cells. However, the functional relationship between tumor exosomes, endothelial signaling, angiogenesis, and metastasis is poorly understood. Herein, we describe a standardized approach for defining the angiogenic potential of isolated exosomes. We created a powerful technique to rapidly and efficiently isolate and track exosomes for study using dynamic light scattering in conjunc...

  13. Secretion of extracellular hsp90α via exosomes increases cancer cell motility: a role for plasminogen activation

    Directory of Open Access Journals (Sweden)

    Chan Doug

    2010-06-01

    Full Text Available Abstract Background Metastasis is a multi-step process that is responsible for the majority of deaths in cancer patients. Current treatments are not effective in targeting metastasis. The molecular chaperone hsp90α is secreted from invasive cancer cells and activates MMP-2 to enhance invasiveness, required for the first step in metastasis. Methods We analyzed the morphology and motility of invasive cancer cells that were treated with exogenous exosomes in the presence or absence of hsp90α. We performed mass spectrometry and immunoprecipitation to identify plasminogen as a potential client protein of extracellular hsp90α. Plasmin activation assays and migration assays were performed to test if plasminogen is activated by extracellular hsp90α and has a role in migration. Results We found that hsp90α is secreted in exosomes in invasive cancer cells and it contributes to their invasive nature. We identified a novel interaction between hsp90α and tissue plasminogen activator that together with annexin II, also found in exosomes, activates plasmin. Extracellular hsp90α promotes plasmin activation as well as increases plasmin dependent cell motility. Conclusions Our data indicate that hsp90α is released by invasive cancer cells via exosomes and implicates hsp90α in activating plasmin, a second protease that acts in cancer cell invasion.

  14. Epstein-Barr virus-encoded small RNAs (EBERs) are present in fractions related to exosomes released by EBV-transformed cells.

    Science.gov (United States)

    Ahmed, Waqar; Philip, Pretty S; Tariq, Saeed; Khan, Gulfaraz

    2014-01-01

    Epstein-Barr virus (EBV) is an oncogenic herpesvirus associated with a number of human malignancies of epithelial and lymphoid origin. However, the mechanism of oncogenesis is unclear. A number of viral products, including EBV latent proteins and non-protein coding RNAs have been implicated. Recently it was reported that EBV-encoded small RNAs (EBERs) are released from EBV infected cells and they can induce biological changes in cells via signaling from toll-like receptor 3. Here, we investigated if these abundantly expressed non-protein coding EBV RNAs (EBER-1 and EBER-2) are excreted from infected cells in exosomal fractions. Using differential ultracentrifugation we isolated exosomes from three EBV positive cell lines (B95-8, EBV-LCL, BL30-B95-8), one EBER-1 transfected cell line (293T-pHEBo-E1) and two EBV-negative cell lines (BL30, 293T-pHEBo). The identity of purified exosomes was determined by electron microscopy and western blotting for CD63. The presence of EBERs in cells, culture supernatants and purified exosomal fractions was determined using RT-PCR and confirmed by sequencing. Purified exosomal fractions were also tested for the presence of the EBER-1-binding protein La, using western blotting. Both EBER-1 and EBER-2 were found to be present not only in the culture supernatants, but also in the purified exosome fractions of all EBV-infected cell lines. EBER-1 could also be detected in exosomal fractions from EBER-1 transfected 293T cells whilst the fractions from vector only transfectants were clearly negative. Furthermore, purified exosomal fractions also contained the EBER-binding protein (La), supporting the notion that EBERs are most probably released from EBV infected cells in the form of EBER-La complex in exosomes.

  15. Epstein-Barr virus-encoded small RNAs (EBERs are present in fractions related to exosomes released by EBV-transformed cells.

    Directory of Open Access Journals (Sweden)

    Waqar Ahmed

    Full Text Available Epstein-Barr virus (EBV is an oncogenic herpesvirus associated with a number of human malignancies of epithelial and lymphoid origin. However, the mechanism of oncogenesis is unclear. A number of viral products, including EBV latent proteins and non-protein coding RNAs have been implicated. Recently it was reported that EBV-encoded small RNAs (EBERs are released from EBV infected cells and they can induce biological changes in cells via signaling from toll-like receptor 3. Here, we investigated if these abundantly expressed non-protein coding EBV RNAs (EBER-1 and EBER-2 are excreted from infected cells in exosomal fractions. Using differential ultracentrifugation we isolated exosomes from three EBV positive cell lines (B95-8, EBV-LCL, BL30-B95-8, one EBER-1 transfected cell line (293T-pHEBo-E1 and two EBV-negative cell lines (BL30, 293T-pHEBo. The identity of purified exosomes was determined by electron microscopy and western blotting for CD63. The presence of EBERs in cells, culture supernatants and purified exosomal fractions was determined using RT-PCR and confirmed by sequencing. Purified exosomal fractions were also tested for the presence of the EBER-1-binding protein La, using western blotting. Both EBER-1 and EBER-2 were found to be present not only in the culture supernatants, but also in the purified exosome fractions of all EBV-infected cell lines. EBER-1 could also be detected in exosomal fractions from EBER-1 transfected 293T cells whilst the fractions from vector only transfectants were clearly negative. Furthermore, purified exosomal fractions also contained the EBER-binding protein (La, supporting the notion that EBERs are most probably released from EBV infected cells in the form of EBER-La complex in exosomes.

  16. Exosomes in the Immune Response and Tolerance

    Institute of Scientific and Technical Information of China (English)

    修方明; 曹雪涛

    2004-01-01

    Exosomes, secreted by many live cells, are small non-cell vesicles with nanoparticle-grade size. In addition to the original function of discarding the uselessful membrane molecules, exosomes are involved in a range of immunoregulatory functions. Dendritic cell-derived exosomes and tumor-derived exosomes are the best characterized vesicles with potent antitumor effect by efficienfly inducing immune response. Down-regtdation of immune response or induction of immune tolerance is another interesting function of exosomes, Further functional studies of the exosomes will shed light on the application of exosomes。

  17. Exosome Derived From Human Umbilical Cord Mesenchymal Stem Cell Mediates MiR-181c Attenuating Burn-induced Excessive Inflammation

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    Xiao Li

    2016-06-01

    Full Text Available Mesenchymal stem cell (MSC-derived exosomes have diverse functions in regulating wound healing and inflammation; however, the molecular mechanism of human umbilical cord MSC (hUCMSC-derived exosomes in regulating burn-induced inflammation is not well understood. We found that burn injury significantly increased the inflammatory reaction of rats or macrophages exposed to lipopolysaccharide (LPS, increased tumor necrosis factor α (TNF-α and interleukin-1β (IL-1β levels and decreased IL-10 levels. hUCMSC-exosome administration successfully reversed this reaction. Further studies showed that miR-181c in the exosomes played a pivotal role in regulating inflammation. Compared to control hUCMSC-exosomes, hUCMSC-exosomes overexpressing miR-181c more effectively suppressed the TLR4 signaling pathway and alleviated inflammation in burned rats. Administration of miR-181c-expressing hUCMSC-exosomes or TLR4 knockdown significantly reduced LPS-induced TLR4 expression by macrophages and the inflammatory reaction. In summary, miR-181c expression in hUCMSC-exosomes reduces burn-induced inflammation by downregulating the TLR4 signaling pathway.

  18. Exosome Derived From Human Umbilical Cord Mesenchymal Stem Cell Mediates MiR-181c Attenuating Burn-induced Excessive Inflammation.

    Science.gov (United States)

    Li, Xiao; Liu, Lingying; Yang, Jing; Yu, Yonghui; Chai, Jiake; Wang, Lingyan; Ma, Li; Yin, Huinan

    2016-06-01

    Mesenchymal stem cell (MSC)-derived exosomes have diverse functions in regulating wound healing and inflammation; however, the molecular mechanism of human umbilical cord MSC (hUCMSC)-derived exosomes in regulating burn-induced inflammation is not well understood. We found that burn injury significantly increased the inflammatory reaction of rats or macrophages exposed to lipopolysaccharide (LPS), increased tumor necrosis factor α (TNF-α) and interleukin-1β (IL-1β) levels and decreased IL-10 levels. hUCMSC-exosome administration successfully reversed this reaction. Further studies showed that miR-181c in the exosomes played a pivotal role in regulating inflammation. Compared to control hUCMSC-exosomes, hUCMSC-exosomes overexpressing miR-181c more effectively suppressed the TLR4 signaling pathway and alleviated inflammation in burned rats. Administration of miR-181c-expressing hUCMSC-exosomes or TLR4 knockdown significantly reduced LPS-induced TLR4 expression by macrophages and the inflammatory reaction. In summary, miR-181c expression in hUCMSC-exosomes reduces burn-induced inflammation by downregulating the TLR4 signaling pathway.

  19. Effects of exosomes derived from MDA-MB-231 on proliferation of endothelial cells and the role of MAPK/ERK and PI3K/Akt pathways

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    Shuang LONG

    2012-11-01

    Full Text Available Objective  To investigate the effects of exosomes derived from breast cancer cell line MDA-MB-231 on proliferation of human umbilical cord vein endothelial cells (HUVECs, and evaluate the role of MAPK/ERK and PI3K/Akt signal transduction pathway during the process. Methods  Exosomes were derived and purified from MDA-MB-231 by cryogenic ultracentrifugation and density gradient centrifugation. MTT assay was carried out for measurement of cell proliferation in HUVECs with exosome of 50, 100, 200 and 400μg/ml. The states of cell cycle of HUVECs co-cultured with 200μg/ml exosomes were detected by flow cytometry. The effects of 200μg/ml exosomes on the expression of ERK, Akt and phosphorylated ERK, Akt in HUVECs were detected with Western blotting. Results  Exosomes derived from MDA-MB-231 significantly promoted HUVECs proliferation in a classical time-and dose-dependent manner. Flow cytometry revealed that, co-cultured with 200μg/ml exosomes for 24h, S-phase cells in HUVECs increased, while G1/S phase cells in HUVECs decreased. Western blotting showed that, cocultured with 200μg/ml exosomes for 24h, 48h and 72h, the expressions of phosphorylated ERK and Akt were up-regulated in a time-dependent manner. Conclusion  Exosomes derived from breast cancer cell line MDA-MB-231 may promote HUVECs proliferation, the changes in cell cycle and the continuous activation of the MAPK/ERK and PI3K/Akt signal transduction pathways may be the underlying mechanism.

  20. Low pH increases the yield of exosome isolation.

    Science.gov (United States)

    Ban, Jae-Jun; Lee, Mijung; Im, Wooseok; Kim, Manho

    2015-05-22

    Exosomes are the extracellular vesicles secreted by various cells. Exosomes mediate intercellular communication by delivering a variety of molecules between cells. Cancer cell derived exosomes seem to be related with tumor progression and metastasis. Tumor microenvironment is thought to be acidic and this low pH controls exosome physiology, leading to tumor progression. Despite the importance of microenvironmental pH on exosome, most of exosome studies have been performed without regard to pH. Therefore, the difference of exosome stability and yield of isolation by different pH need to be studied. In this research, we investigated the yield of total exosomal protein and RNA after incubation in acidic, neutral and alkaline conditioned medium. Representative exosome markers were investigated by western blot after incubation of exosomes in different pH. As a result, the concentrations of exosomal protein and nucleic acid were significantly increased after incubation in the acidic medium compared with neutral medium. The higher levels of exosome markers including CD9, CD63 and HSP70 were observed after incubation in an acidic environment. On the other hand, no exosomal protein, exosomal RNA and exosome markers have been detected after incubation in an alkaline condition. In summary, our results indicate that the acidic condition is the favorable environment for existence and isolation of exosomes.

  1. 喉癌 Hep -2细胞来源的 exosomes 的发现和鉴定%Discovery and isolation of exosomes derived from laryngocarcinoma Hep - 2 cells

    Institute of Scientific and Technical Information of China (English)

    吉晓滨; 梁俊毅; 刘启才; 谢景华

    2015-01-01

    Objective:To observe whether laryngocarcinoma Hep - 2 cells can secrete exosomes,and to identify exosomes morphologically. Methods:A large number of laryngocarcinoma Hep - 2 cells were cultivated,the yield of exosomes increased by hot shock,cell culture supernatant was gathered. Firstly,the culture supernatant was pretreat-ment by clarification through a 3 / 0. 8μm small filter element for deep filter to remove particles and impurities with larger diameter. Secondly,exosome isolation kit was used to isolate and extract exosomes. Cells culture supernatant 4ml was gathered,the solutions of the kit were added into the supernatant in proper sequence,then filtered by the special column,the concentrated fluid was obtained. The exosomes were observed under transmission electron microscopy. Re-sults:Exosomes could be isolated and extracted from culture supernatant of laryngocarcinoma Hep - 2 cells,and it present circular or elliptical vesicle with bilayer membrane,high density,well - distribution,and with range from 20 to 100nm of diameter. Conclusion:Exosomes can be secreted from laryngocarcinoma Hep - 2 cells was first discovered, which provide a new research to laryngocarcinoma immunotherapy.%目的:观察喉癌 Hep -2细胞可否分泌 exosomes,并从形态学角度鉴定。方法:大量培养喉癌 Hep -2细胞,热休克处理,收集培养上清。先通过3/0.8μm 深层过滤小型滤芯对上清进行预处理,去除直径较大的颗粒和杂质。采用 Exosome Isolation Kit(商品化试剂盒)收集培养上清液4ml,依次加入 Exosome Isolation Kit内试剂,通过 exosomes 提取专用过滤柱,收集浓缩液。用高倍透射电子显微镜对 exosomes 做鉴定。结果:成功从喉癌 Hep -2细胞培养上清中分离提取出 exosomes,电镜观察见 exosomes 呈圆形或椭圆形双层膜的囊泡状结构,直径约20~100nm,密度较高,分散均匀。结论:首次发现喉癌细胞自身能分泌 exosomes,为喉

  2. Resolving sorting mechanisms into exosomes

    NARCIS (Netherlands)

    Stoorvogel, Willem

    2015-01-01

    The complexity of mechanisms driving protein sorting into exosomes is only beginning to emerge. In a paper recently published in Cell Research, Roucourt et al. report that trimming of heparan sulfate side chains of syndecans by endosomal heparanase facilitates sorting into exosomes by the formation

  3. Induction of protective immunity against Eimeria tenella, Eimeria maxima, and Eimeria acervulina infections using dendritic cell-derived exosomes.

    Science.gov (United States)

    del Cacho, Emilio; Gallego, Margarita; Lee, Sung Hyen; Lillehoj, Hyun Soon; Quilez, Joaquin; Lillehoj, Erik P; Sánchez-Acedo, Caridad

    2012-05-01

    This study describes a novel immunization strategy against avian coccidiosis using exosomes derived from Eimeria parasite antigen (Ag)-loaded dendritic cells (DCs). Chicken intestinal DCs were isolated and pulsed in vitro with a mixture of sporozoite-extracted Ags from Eimeria tenella, E. maxima, and E. acervulina, and the cell-derived exosomes were isolated. Chickens were nonimmunized or immunized intramuscularly with exosomes and subsequently noninfected or coinfected with E. tenella, E. maxima, and E. acervulina oocysts. Immune parameters compared among the nonimmunized/noninfected, nonimmunized/infected, and immunized/infected groups were the numbers of cells secreting T(h)1 cytokines, T(h)2 cytokines, interleukin-16 (IL-16), and Ag-reactive antibodies in vitro and in vivo readouts of protective immunity against Eimeria infection. Cecal tonsils, Peyer's patches, and spleens of immunized and infected chickens had increased numbers of cells secreting the IL-16 and the T(h)1 cytokines IL-2 and gamma interferon, greater Ag-stimulated proliferative responses, and higher numbers of Ag-reactive IgG- and IgA-producing cells following in vitro stimulation with the sporozoite Ags compared with the nonimmunized/noninfected and nonimmunized/infected controls. In contrast, the numbers of cells secreting the T(h)2 cytokines IL-4 and IL-10 were diminished in immunized and infected chickens compared with the nonimmunized/noninfected and the nonimmunized/infected controls. Chickens immunized with Ag-loaded exosomes and infected in vivo with Eimeria oocysts had increased body weight gains, reduced feed conversion ratios, diminished fecal oocyst shedding, lessened intestinal lesion scores, and reduced mortality compared with the nonimmunized/infected controls. These results suggest that successful field vaccination against avian coccidiosis using exosomes derived from DCs incubated with Ags isolated from Eimeria species may be possible.

  4. Melanoma cell-derived exosomes promote epithelial-mesenchymal transition in primary melanocytes through paracrine/autocrine signaling in the tumor microenvironment.

    Science.gov (United States)

    Xiao, Deyi; Barry, Samantha; Kmetz, Daniel; Egger, Michael; Pan, Jianmin; Rai, Shesh N; Qu, Jifu; McMasters, Kelly M; Hao, Hongying

    2016-07-01

    The tumor microenvironment is abundant with exosomes that are secreted by the cancer cells themselves. Exosomes are nanosized, organelle-like membranous structures that are increasingly being recognized as major contributors in the progression of malignant neoplasms. A critical element in melanoma progression is its propensity to metastasize, but little is known about how melanoma cell-derived exosomes modulate the microenvironment to optimize conditions for tumor progression and metastasis. Here, we provide evidence that melanoma cell-derived exosomes promote phenotype switching in primary melanocytes through paracrine/autocrine signaling. We found that the mitogen-activated protein kinase (MAPK) signaling pathway was activated during the exosome-mediated epithelial-to-mesenchymal transition (EMT)-resembling process, which promotes metastasis. Let-7i, an miRNA modulator of EMT, was also involved in this process. We further defined two other miRNA modulators of EMT (miR-191 and let-7a) in serum exosomes for differentiating stage I melanoma patients from non-melanoma subjects. These results provide the first strong molecular evidence that melanoma cell-derived exosomes promote the EMT-resembling process in the tumor microenvironment. Thus, novel strategies targeting EMT and modulating the tumor microenvironment may emerge as important approaches for the treatment of metastatic melanoma.

  5. Exosomes derived from human adipose mensenchymal stem cells accelerates cutaneous wound healing via optimizing the characteristics of fibroblasts

    Science.gov (United States)

    Hu, Li; Wang, Juan; Zhou, Xin; Xiong, Zehuan; Zhao, Jiajia; Yu, Ran; Huang, Fang; Zhang, Handong; Chen, Lili

    2016-01-01

    Prolonged healing and scar formation are two major challenges in the treatment of soft tissue trauma. Adipose mesenchymal stem cells (ASCs) play an important role in tissue regeneration, and recent studies have suggested that exosomes secreted by stem cells may contribute to paracrine signaling. In this study, we investigated the roles of ASCs-derived exosomes (ASCs-Exos) in cutaneous wound healing. We found that ASCs-Exos could be taken up and internalized by fibroblasts to stimulate cell migration, proliferation and collagen synthesis in a dose-dependent manner, with increased genes expression of N-cadherin, cyclin-1, PCNA and collagen I, III. In vivo tracing experiments demonstrated that ASCs-Exos can be recruited to soft tissue wound area in a mouse skin incision model and significantly accelerated cutaneous wound healing. Histological analysis showed increased collagen I and III production by systemic administration of exosomes in the early stage of wound healing, while in the late stage, exosomes might inhibit collagen expression to reduce scar formation. Collectively, our findings indicate that ASCs-Exos can facilitate cutaneous wound healing via optimizing the characteristics of fibroblasts. Our results provide a new perspective and therapeutic strategy for the use of ASCs-Exos in soft tissue repair. PMID:27615560

  6. Intravenous Cardiac Stem Cell-Derived Exosomes Ameliorate Cardiac Dysfunction in Doxorubicin Induced Dilated Cardiomyopathy

    Directory of Open Access Journals (Sweden)

    Adam C. Vandergriff

    2015-01-01

    Full Text Available Despite the efficacy of cardiac stem cells (CSCs for treatment of cardiomyopathies, there are many limitations to stem cell therapies. CSC-derived exosomes (CSC-XOs have been shown to be responsible for a large portion of the regenerative effects of CSCs. Using a mouse model of doxorubicin induced dilated cardiomyopathy, we study the effects of systemic delivery of human CSC-XOs in mice. Mice receiving CSC-XOs showed improved heart function via echocardiography, as well as decreased apoptosis and fibrosis. In spite of using immunocompetent mice and human CSC-XOs, mice showed no adverse immune reaction. The use of CSC-XOs holds promise for overcoming the limitations of stem cells and improving cardiac therapies.

  7. Mast cell synapses and exosomes: membrane contacts for information exchange.

    NARCIS (Netherlands)

    Carroll-Portillo, A.; Surviladze, Z.; Cambi, A.; Lidke, D.S.; Wilson, B.S.

    2012-01-01

    In addition to their central role in allergy, mast cells are involved in a wide variety of cellular interactions during homeostasis and disease. In this review, we discuss the ability of mast cells to extend their mechanisms for intercellular communication beyond the release of soluble mediators. Th

  8. Exosome Function: From Tumor Immunology to Pathogen Biology

    OpenAIRE

    Schorey, Jeffrey S; Bhatnagar, Sanchita

    2008-01-01

    Exosomes are the newest family member of ‘bioactive vesicles’ that function to promote intercellular communication. Exosomes are derived from the fusion of multi-vesicular bodies with the plasma membrane and extracellular release of the intraluminal vesicles. Recent studies have focused on the biogenesis and composition of exosomes as well as regulation of exosome release. Exosomes have been shown to be released by cells of hematopoietic and non-hematopoietic origin, yet their function remain...

  9. Neisseria gonorrhoeae Modulates Cell Death in Human Endocervical Epithelial Cells through Export of Exosome-Associated cIAP2.

    Science.gov (United States)

    Nudel, Kathleen; Massari, Paola; Genco, Caroline A

    2015-09-01

    Several bacterial pathogens persist and survive in the host by modulating host cell death pathways. We previously demonstrated that Neisseria gonorrhoeae, a Gram-negative pathogen responsible for the sexually transmitted infection gonorrhea, protects against exogenous induction of apoptosis in human cervical epithelial cells. However, induction of cell death by N. gonorrhoeae has also been reported in other cell types. The mechanisms by which N. gonorrhoeae modulates cell death are not clear, although a role for the inhibitor of apoptosis-2 (cIAP2) has been proposed. In this study, we confirmed that N. gonorrhoeae induces production of cIAP2 in human cervical epithelial cells. High levels of intracellular cIAP2 were detected early after N. gonorrhoeae stimulation, which was followed by a marked decrease at 24 h. At this time point, we observed increased levels of extracellular cIAP2 associated with exosomes and an overall increase in production of exosomes. Inhibition of cIAP2 in N. gonorrhoeae-stimulated epithelial cells resulted in increased cell death and interleukin-1β (IL-1β) production. Collectively these results indicate that N. gonorrhoeae stimulation of human endocervical epithelial cells induces the release of cIAP2, an essential regulator of cell death and immune signaling.

  10. Exosomes from HIV-1-infected Cells Stimulate Production of Pro-inflammatory Cytokines through Trans-activating Response (TAR) RNA.

    Science.gov (United States)

    Sampey, Gavin C; Saifuddin, Mohammed; Schwab, Angela; Barclay, Robert; Punya, Shreya; Chung, Myung-Chul; Hakami, Ramin M; Zadeh, Mohammad Asad; Lepene, Benjamin; Klase, Zachary A; El-Hage, Nazira; Young, Mary; Iordanskiy, Sergey; Kashanchi, Fatah

    2016-01-15

    HIV-1 infection results in a chronic illness because long-term highly active antiretroviral therapy can lower viral titers to an undetectable level. However, discontinuation of therapy rapidly increases virus burden. Moreover, patients under highly active antiretroviral therapy frequently develop various metabolic disorders, neurocognitive abnormalities, and cardiovascular diseases. We have previously shown that exosomes containing trans-activating response (TAR) element RNA enhance susceptibility of undifferentiated naive cells to HIV-1 infection. This study indicates that exosomes from HIV-1-infected primary cells are highly abundant with TAR RNA as detected by RT-real time PCR. Interestingly, up to a million copies of TAR RNA/μl were also detected in the serum from HIV-1-infected humanized mice suggesting that TAR RNA may be stable in vivo. Incubation of exosomes from HIV-1-infected cells with primary macrophages resulted in a dramatic increase of proinflammatory cytokines, IL-6 and TNF-β, indicating that exosomes containing TAR RNA could play a direct role in control of cytokine gene expression. The intact TAR molecule was able to bind to PKR and TLR3 effectively, whereas the 5' and 3' stems (TAR microRNAs) bound best to TLR7 and -8 and none to PKR. Binding of TAR to PKR did not result in its phosphorylation, and therefore, TAR may be a dominant negative decoy molecule in cells. The TLR binding through either TAR RNA or TAR microRNA potentially can activate the NF-κB pathway and regulate cytokine expression. Collectively, these results imply that exosomes containing TAR RNA could directly affect the proinflammatory cytokine gene expression and may explain a possible mechanism of inflammation observed in HIV-1-infected patients under cART.

  11. Proteomics of MUC1-containing lipid rafts from plasma membranes and exosomes of human breast carcinoma cells MCF-7.

    Science.gov (United States)

    Staubach, Simon; Razawi, Hanieh; Hanisch, Franz-Georg

    2009-05-01

    Apically expressed human MUC1 is known to become endocytosed and either to re-enter the secretory pathway for recycling to the plasma membrane or to be exported by the cells via the formation of multi-vesicular bodies and the release of exosomes. By using recombinant fusion-tagged MUC1 as a bait protein we followed an anti-myc affinity-based approach for isolating subpopulations of lipid rafts from the plasma membranes and exosomes of MCF-7 breast cancer cells. MUC1(+) lipid rafts were not only found to contain genuine raft proteins (flotillin-1, prohibitin, G protein, annexin A2), but also raft-associated proteins linking these to the cytoskeleton (ezrin/villin-2, profilin II, HSP27, gamma-actin, beta-actin) or proteins in complexes with raft proteins, including the bait protein (HSP60, HSP70). Major overlaps were revealed for the subproteomes of plasma membranous and exosomal lipid raft preparations, indicating that MUC1 is sorted into subpopulations of rafts for its trafficking via flotillin-dependent pathways and export via exosomes.

  12. Development of a rapid lateral flow immunoassay test for detection of exosomes previously enriched from cell culture medium and body fluids

    Directory of Open Access Journals (Sweden)

    Myriam Oliveira-Rodríguez

    2016-08-01

    Full Text Available Exosomes are cell-secreted nanovesicles (40–200 nm that represent a rich source of novel biomarkers in the diagnosis and prognosis of certain diseases. Despite the increasingly recognized relevance of these vesicles as biomarkers, their detection has been limited due in part to current technical challenges in the rapid isolation and analysis of exosomes. The complexity of the development of analytical platforms relies on the heterogeneous composition of the exosome membrane. One of the most attractive tests is the inmunochromatographic strips, which allow rapid detection by unskilled operators. We have successfully developed a novel lateral flow immunoassay (LFIA for the detection of exosomes based on the use of tetraspanins as targets. We have applied this platform for the detection of exosomes purified from different sources: cell culture supernatants, human plasma and urine. As proof of concept, we explored the analytical potential of this LFIA platform to accurately quantify exosomes purified from a human metastatic melanoma cell line. The one-step assay can be completed in 15 min, with a limit of detection of 8.54×105 exosomes/µL when a blend of anti-CD9 and anti-CD81 were selected as capture antibodies and anti-CD63 labelled with gold nanoparticles as detection antibody. Based on our results, this platform could be well suited to be used as a rapid exosome quantification tool, with promising diagnostic applications, bearing in mind that the detection of exosomes from different sources may require adaptation of the analytical settings to their specific composition.

  13. Tumor microenvironment derived exosomes pleiotropically modulate cancer cell metabolism

    Science.gov (United States)

    Cancer-associated fibroblasts (CAFs) are a major cellular component of tumor microenvironment in most solid cancers. Altered cellular metabolism is a hallmark of cancer, and much of the published literature has focused on neoplastic cell-autonomous processes for these adaptations. We demonstrate tha...

  14. Cardiac myocyte exosomes: stability, HSP60, and proteomics.

    Science.gov (United States)

    Malik, Z A; Kott, K S; Poe, A J; Kuo, T; Chen, L; Ferrara, K W; Knowlton, A A

    2013-04-01

    Exosomes, which are 50- to 100-nm-diameter lipid vesicles, have been implicated in intercellular communication, including transmitting malignancy, and as a way for viral particles to evade detection while spreading to new cells. Previously, we demonstrated that adult cardiac myocytes release heat shock protein (HSP)60 in exosomes. Extracellular HSP60, when not in exosomes, causes cardiac myocyte apoptosis via the activation of Toll-like receptor 4. Thus, release of HSP60 from exosomes would be damaging to the surrounding cardiac myocytes. We hypothesized that 1) pathological changes in the environment, such as fever, change in pH, or ethanol consumption, would increase exosome permeability; 2) different exosome inducers would result in different exosomal protein content; 3) ethanol at "physiological" concentrations would cause exosome release; and 4) ROS production is an underlying mechanism of increased exosome production. We found the following: first, exosomes retained their protein cargo under different physiological/pathological conditions, based on Western blot analyses. Second, mass spectrometry demonstrated that the protein content of cardiac exosomes differed significantly from other types of exosomes in the literature and contained cytosolic, sarcomeric, and mitochondrial proteins. Third, ethanol did not affect exosome stability but greatly increased the production of exosomes by cardiac myocytes. Fourth, ethanol- and hypoxia/reoxygenation-derived exosomes had different protein content. Finally, ROS inhibition reduced exosome production but did not completely inhibit it. In conclusion, exosomal protein content is influenced by the cell source and stimulus for exosome formation. ROS stimulate exosome production. The functions of exosomes remain to be fully elucidated.

  15. Distinct Dasatinib-Induced Mechanisms of Apoptotic Response and Exosome Release in Imatinib-Resistant Human Chronic Myeloid Leukemia Cells

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    Juan Liu

    2016-04-01

    Full Text Available Although dasatinib is effective in most imatinib mesylate (IMT-resistant chronic myeloid leukemia (CML patients, the underlying mechanism of its effectiveness in eliminating imatinib-resistant cells is only partially understood. This study investigated the effects of dasatinib on signaling mechanisms driving-resistance in imatinib-resistant CML cell line K562 (K562RIMT. Compared with K562 control cells, exsomal release, the phosphoinositide 3-kinase (PI3K/protein kinase B (Akt/ mammalian target of rapamycin (mTOR signaling and autophagic activity were increased significantly in K562RIMT cells and mTOR-independent beclin-1/Vps34 signaling was shown to be involved in exosomal release in these cells. We found that Notch1 activation-mediated reduction of phosphatase and tensin homolog (PTEN was responsible for the increased Akt/mTOR activities in K562RIMT cells and treatment with Notch1 γ-secretase inhibitor prevented activation of Akt/mTOR. In addition, suppression of mTOR activity by rapamycin decreased the level of activity of p70S6K, induced upregulation of p53 and caspase 3, and led to increase of apoptosis in K562RIMT cells. Inhibition of autophagy by spautin-1 or beclin-1 knockdown decreased exosomal release, but did not affect apoptosis in K562RIMT cells. In summary, in K562RIMT cells dasatinib promoted apoptosis through downregulation of Akt/mTOR activities, while preventing exosomal release and inhibiting autophagy by downregulating expression of beclin-1 and Vps34. Our findings reveal distinct dasatinib-induced mechanisms of apoptotic response and exosomal release in imatinib-resistant CML cells.

  16. Exosomes released in vitro from Epstein-Barr virus (EBV)-infected cells contain EBV-encoded latent phase mRNAs.

    Science.gov (United States)

    Canitano, Andrea; Venturi, Giulietta; Borghi, Martina; Ammendolia, Maria Grazia; Fais, Stefano

    2013-09-01

    EBV is a human herpesvirus associated with a number of malignancies. Both lymphoblastoid cell lines (LCLs), and EBV-infected nasopharyngeal carcinoma (NPC) cells have been demonstrated to release exosomes containing the EBV-encoded latent membrane protein 1 (LMP1), and mature micro-RNAs (EBV-miRNAs). Here we analyze the EBV protein and nucleic acid content of exosomes from different EBV-infected cells (LCL, 721 and Daudi) and we show for the first time that exosomes released from LCLs and 721 also contain EBV-encoded latent phase mRNAs. This confirms and strengthens exosomes pathogenetic potential, and might provide insights for development of novel diagnostic and therapeutic strategies.

  17. Pancreatic cancer-derived exosomes transfer miRNAs to dendritic cells and inhibit RFXAP expression via miR-212-3p.

    Science.gov (United States)

    Ding, Guoping; Zhou, Liangjing; Qian, Yingming; Fu, Mingnian; Chen, Jian; Chen, Jionghuang; Xiang, Jianyang; Wu, Zhengrong; Jiang, Guixing; Cao, Liping

    2015-10-06

    It has been reported tumor-derived exosomes can transfer miRNAs to recipient cells in the tumor microenvironment, promoting tumor invasion and metastasis. The present research aimed to explore how pancreatic cancer (PC) derived exosomal miRNAs inhibited mRNA expression of dendritic cells and induced immune tolerance. Our study revealed that 9 PC-related miRNAs were increased and 208 mRNAs were inhibited in exosome-stimulated dendritic cells (exo-iDCs) compared to immature dendritic cells (iDCs). A target prediction between the 9 miRNAs and 208 mRNAs was performed by bioinformatics database analysis. From the target prediction, it was predicted and validated that regulatory factor X-associated protein (RFXAP), an important transcription factor for MHC II, was inhibited by miR-212-3p transferred from PC-secreted exosomes, resulting in decreased MHC II expression. Moreover, a clinical study showed a negative correlation between miR-212-3p and RFXAP in PC tissue. From these data, we concluded that PC-related miRNAs can be transferred to dendritic cells via exosome and inhibit target mRNA expression. More importantly, PC-derived exosomes inhibit RFXAP expression via miR-212-3p, which decrease MHC II expression and induce immune tolerance of dendritic cells. RFXAP deficiency has never been reported in solid tumors. The functions and mechanisms of RFXAP in tumors deserve future explorations.

  18. Sorting it out: regulation of exosome loading.

    Science.gov (United States)

    Villarroya-Beltri, Carolina; Baixauli, Francesc; Gutiérrez-Vázquez, Cristina; Sánchez-Madrid, Francisco; Mittelbrunn, María

    2014-10-01

    Extracellular vesicles (EVs), a term that includes both exosomes of endocytic origin and vesicles derived from plasma membranes, are continuously secreted by cells to the extracellular environment, and represent a novel vehicle for cell-cell communication. Exosomes contain specific repertoires of proteins and RNAs, indicating the existence of mechanisms that control the sorting of molecules into them. Although the molecular mechanisms that regulate the loading of proteins into exosomes have been studied for years, the sorting of RNA has been elusive until recently. Here we review the molecular mechanisms that control the sorting of molecules into exosomes, with special attention to the sorting of RNA. We also discuss how the cellular context affects the composition of exosomes, and thus the outcome of the communication between the exosome-producer and recipient cells, with particular focus on the communication between tumor cells and with cells of the tumor microenvironment.

  19. Transcriptome analysis of exosome-compromised human cells using high-density tiling arrays

    DEFF Research Database (Denmark)

    Jensen, Torben Heick

    The extent of RNA degradation in the nucleus has traditionally been underestimated. However, all major RNA species are synthesized, processed and can be degraded in this compartment and consequently an enormous amount of nucleosides are turned over and recycled. The RNA exosome, a multisubunit...... complex of 3’-5’ exoribonucleases, is a key player in these processive/degradative pathways. The exosome is highly conserved between yeast and man, and exists in a cytoplasmic and a nuclear form; the 3’-5’ exoribonuclease Rrp6 (human homologue PM/Scl100) is a specific component of the nuclear exosome.......Studies in yeast using exosome-mutant strains has revealed specific functions of the nuclear exosome: (i) processing or degradation of small nuclear/nucleolar RNAs (snRNAs, snoRNAs), (ii) surveillance and degradation of malformed mRNAs and (iii) processing or degradation of ribosomal precursor RNA to mature r...

  20. Stabilization of exosome-targeting peptides via engineered glycosylation.

    Science.gov (United States)

    Hung, Michelle E; Leonard, Joshua N

    2015-03-27

    Exosomes are secreted extracellular vesicles that mediate intercellular transfer of cellular contents and are attractive vehicles for therapeutic delivery of bimolecular cargo such as nucleic acids, proteins, and even drugs. Efficient exosome-mediated delivery in vivo requires targeting vesicles for uptake by specific recipient cells. Although exosomes have been successfully targeted to several cellular receptors by displaying peptides on the surface of the exosomes, identifying effective exosome-targeting peptides for other receptors has proven challenging. Furthermore, the biophysical rules governing targeting peptide success remain poorly understood. To evaluate one factor potentially limiting exosome delivery, we investigated whether peptides displayed on the exosome surface are degraded during exosome biogenesis, for example by endosomal proteases. Indeed, peptides fused to the N terminus of exosome-associated transmembrane protein Lamp2b were cleaved in samples derived from both cells and exosomes. To suppress peptide loss, we engineered targeting peptide-Lamp2b fusion proteins to include a glycosylation motif at various positions. Introduction of this glycosylation motif both protected the peptide from degradation and led to an increase in overall Lamp2b fusion protein expression in both cells and exosomes. Moreover, glycosylation-stabilized peptides enhanced targeted delivery of exosomes to neuroblastoma cells, demonstrating that such glycosylation does not ablate peptide-target interactions. Thus, we have identified a strategy for achieving robust display of targeting peptides on the surface of exosomes, which should facilitate the evaluation and development of new exosome-based therapeutics.

  1. CLL Exosomes Modulate the Transcriptome and Behaviour of Recipient Stromal Cells and Are Selectively Enriched in miR-202-3p.

    Directory of Open Access Journals (Sweden)

    Mosavar Farahani

    Full Text Available Bi-directional communication with the microenvironment is essential for homing and survival of cancer cells with implications for disease biology and behaviour. In chronic lymphocytic leukemia (CLL, the role of the microenvironment on malignant cell behaviour is well described. However, how CLL cells engage and recruit nurturing cells is poorly characterised. Here we demonstrate that CLL cells secrete exosomes that are nanovesicles originating from the fusion of multivesicular bodies with the plasma membrane, to shuttle proteins, lipids, microRNAs (miR and mRNAs to recipient cells. We characterise and confirm the size (50-100 nm and identity of the CLL-derived exosomes by Electron microscopy (EM, Atomic force microscopy (AFM, flow cytometry and western blotting using both exosome- and CLL-specific markers. Incubation of CLL-exosomes, derived either from cell culture supernatants or from patient plasma, with human stromal cells shows that they are readily taken up into endosomes, and induce expression of genes such as c-fos and ATM as well as enhance proliferation of recipient HS-5 cells. Furthermore, we show that CLL exosomes encapsulate abundant small RNAs and are enriched in certain miRs and specifically hsa-miR-202-3p. We suggest that such specific packaging of miR-202-3p into exosomes results in enhanced expression of 'suppressor of fused' (Sufu, a Hedgehog (Hh signalling intermediate, in the parental CLL cells. Thus, our data show that CLL cells secrete exosomes that alter the transcriptome and behaviour of recipient cells. Such communication with microenvironment is likely to have an important role in CLL disease biology.

  2. [Analysis and Control of in Vivo Kinetics of Exosomes for the Development of Exosome-based DDS].

    Science.gov (United States)

    Takahashi, Yuki; Nishikawa, Makiya; Takakura, Yoshinobu

    2016-01-01

      Exosomes are secretory membrane vesicles containing lipids, proteins, and nucleic acids. They act as intercellular transporters by delivering their components to exosome recipient cells. Based on their endogenous delivery system properties, exosomes are expected to become drug delivery systems (DDS) for various molecules such as nucleic acid-based drugs. Important factors such as drug loading to exosomes, production, and pharmacokinetics of exosomes need to be considered for the development of exosome-based DDS. Of these, the pharmacokinetics of exosomes have rarely been studied, probably because of the lack of quantitative evaluation methods of in vivo exosomal pharmacokinetics. We selected lactadherin as an exosome tropic protein and developed it as a fusion protein with Gaussia luciferase to label exosomes for in vivo imaging. In addition, a fusion protein of lactadherin and streptavidin was developed, and the tissue distribution of exosomes was quantitatively evaluated by radiolabeling the exosomes using (125)I-labeled biotin. Using labeled exosomes, we found that intravenously injected exosomes were rapidly cleared from the systemic circulation by macrophages. In addition, the exosomes were mainly distributed to the liver, lung, and spleen. We also examined the effect of exosome isolation methods on their physicochemical and pharmacokinetic properties. We found that exosomes collected by the ultracentrifugation-based density-gradient method were more dispersed than exosomes collected by other methods, including the ultracentrifugation-based pelleting method. The gradient method is more time-consuming than others; therefore the development of a more efficient method for exosome isolation will advance the development of exosome-based DDS.

  3. Exosome mediated communication within the tumor microenvironment.

    Science.gov (United States)

    Milane, Lara; Singh, Amit; Mattheolabakis, George; Suresh, Megha; Amiji, Mansoor M

    2015-12-10

    It is clear that exosomes (endosome derived vesicles) serve important roles in cellular communication both locally and distally and that the exosomal process is abnormal in cancer. Cancer cells are not malicious cells; they are cells that represent 'survival of the fittest' at its finest. All of the mutations, abnormalities, and phenomenal adaptations to a hostile microenvironment, such as hypoxia and nutrient depletion, represent the astute ability of cancer cells to adapt to their environment and to intracellular changes to achieve a single goal - survival. The aberrant exosomal process in cancer represents yet another adaptation that promotes survival of cancer. Cancer cells can secrete more exosomes than healthy cells, but more importantly, the content of cancer cells is distinct. An illustrative distinction is that exosomes derived from cancer cells contain more microRNA than healthy cells and unlike exosomes released from healthy cells, this microRNA can be associated with the RNA-induced silencing complex (RISC) which is required for processing mature and biologically active microRNA. Cancer derived exosomes have the ability to transfer metastatic potential to a recipient cell and cancer exosomes function in the physical process of invasion. In this review we conceptualize the aberrant exosomal process (formation, content selection, loading, trafficking, and release) in cancer as being partially attributed to cancer specific differences in the endocytotic process of receptor recycling/degradation and plasma membrane remodeling and the function of the endosome as a signaling entity. We discuss this concept and, to advance comprehension of exosomal function in cancer as mediators of communication, we detail and discuss exosome biology, formation, and communication in health and cancer; exosomal content in cancer; exosomal biomarkers in cancer; exosome mediated communication in cancer metastasis, drug resistance, and interfacing with the immune system; and

  4. Tetherin is an exosomal tether

    Science.gov (United States)

    Edgar, James R; Manna, Paul T; Nishimura, Shinichi; Banting, George; Robinson, Margaret S

    2016-01-01

    Exosomes are extracellular vesicles that are released when endosomes fuse with the plasma membrane. They have been implicated in various functions in both health and disease, including intercellular communication, antigen presentation, prion transmission, and tumour cell metastasis. Here we show that inactivating the vacuolar ATPase in HeLa cells causes a dramatic increase in the production of exosomes, which display endocytosed tracers, cholesterol, and CD63. The exosomes remain clustered on the cell surface, similar to retroviruses, which are attached to the plasma membrane by tetherin. To determine whether tetherin also attaches exosomes, we knocked it out and found a 4-fold reduction in plasma membrane-associated exosomes, with a concomitant increase in exosomes discharged into the medium. This phenotype could be rescued by wild-type tetherin but not tetherin lacking its GPI anchor. We propose that tetherin may play a key role in exosome fate, determining whether they participate in long-range or short-range interactions. DOI: http://dx.doi.org/10.7554/eLife.17180.001 PMID:27657169

  5. Differential Distribution of Exosome Subunits at the Nuclear Lamina and in Cytoplasmic FociD⃞V⃞

    OpenAIRE

    Amy C Graham; Kiss, Daniel L.; Andrulis, Erik D.

    2006-01-01

    The exosome complex plays important roles in RNA processing and turnover. Despite significant mechanistic insight into exosome function, we still lack a basic understanding of the subcellular locales where exosome complex biogenesis and function occurs. Here, we employ a panel of Drosophila S2 stable cell lines expressing epitope-tagged exosome subunits to examine the subcellular distribution of exosome complex components. We show that tagged Drosophila exosome subunits incorporate into compl...

  6. Proteomic profiling of human plasma exosomes identifies PPARgamma as an exosome-associated protein.

    Science.gov (United States)

    Looze, Christopher; Yui, David; Leung, Lester; Ingham, Matthew; Kaler, Maryann; Yao, Xianglan; Wu, Wells W; Shen, Rong-Fong; Daniels, Mathew P; Levine, Stewart J

    2009-01-16

    Exosomes are nanovesicles that are released from cells as a mechanism of cell-free intercellular communication. Only a limited number of proteins have been identified from the plasma exosome proteome. Here, we developed a multi-step fractionation scheme incorporating gel exclusion chromatography, rate zonal centrifugation through continuous sucrose gradients, and high-speed centrifugation to purify exosomes from human plasma. Exosome-associated proteins were separated by SDS-PAGE and 66 proteins were identified by LC-MS/MS, which included both cellular and extracellular proteins. Furthermore, we identified and characterized peroxisome proliferator-activated receptor-gamma (PPARgamma), a nuclear receptor that regulates adipocyte differentiation and proliferation, as well as immune and inflammatory cell functions, as a novel component of plasma-derived exosomes. Given the important role of exosomes as intercellular messengers, the discovery of PPARgamma as a component of human plasma exosomes identifies a potential new pathway for the paracrine transfer of nuclear receptors.

  7. Exosomes released by granulocytic myeloid-derived suppressor cells attenuate DSS-induced colitis in mice.

    Science.gov (United States)

    Wang, Yungang; Tian, Jie; Tang, Xinyi; Rui, Ke; Tian, Xinyu; Ma, Jie; Ma, Bin; Xu, Huaxi; Lu, Liwei; Wang, Shengjun

    2016-03-29

    Myeloid-derived suppressor cells (MDSC) have been described in inflammatory bowel disease (IBD), but their role in the disease remains controversial. We sought to define the effect of granulocytic MDSC-derived exosomes (G-MDSC exo) in dextran sulphate sodium (DSS)-induced murine colitis. G-MDSC exo-treated mice showed greater resistance to colitis, as reflected by lower disease activity index, decreased inflammatory cell infiltration damage. There was a decrease in the proportion of Th1 cells and an increase in the proportion of regulatory T cells (Tregs) in mesenteric lymph nodes (MLNs) from G-MDSC exo-treated colitis mice. Moreover, lower serum levels of interferon (IFN)-γ and tumor necrosis factor (TNF)-α were detected in G-MDSC exo-treated colitis mice. Interestingly, inhibition of arginase (Arg)-1 activity in G-MDSC exo partially abrogated the spontaneous improvement of colitis. In addition, G-MDSC exo could suppress CD4+ T cell proliferation and IFN-γ secretion in vitro and inhibit the delayed-type hypersensitivity (DTH) response, and these abilities were associated with Arg-1 activity. Moreover, G-MDSC exo promoted the expansion of Tregs in vitro. Taken together, these results suggest that G-MDSC exo attenuate DSS-induced colitis through inhibiting Th1 cells proliferation and promoting Tregs expansion.

  8. Physiological and pathological impact of exosomes of adipose tissue.

    Science.gov (United States)

    Zhang, Yan; Yu, Mei; Tian, Weidong

    2016-02-01

    Exosomes are nanovesicles that have emerged as a new intercellular communication system for transporting proteins and RNAs; recent studies have shown that they play a role in many physiological and pathological processes such as immune regulation, cell differentiation, infection and cancer. By transferring proteins, mRNAs and microRNAs, exosomes act as information vehicles that alter the behavior of recipient cells. Compared to direct cell-cell contact or secreted factors, exosomes can affect recipient cells in more efficient ways. In whole adipose tissues, it has been shown that exosomes exist in supernatants of adipocytes and adipose stromal cells (ADSCs). Adipocyte exosomes are linked to lipid metabolism and obesity-related insulin resistance and exosomes secreted by ADSCs are involved in angiogenesis, immunomodulation and tumor development. This review introduces characteristics of exosomes in adipose tissue, summarizes their functions in different physiological and pathological processes and provides the further insight into potential application of exosomes to disease diagnosis and treatment.

  9. Exosomes: Implications in HIV-1 Pathogenesis.

    Science.gov (United States)

    Madison, Marisa N; Okeoma, Chioma M

    2015-07-20

    Exosomes are membranous nanovesicles of endocytic origin that carry host and pathogen derived genomic, proteomic, and lipid cargos. Exosomes are secreted by most cell types into the extracellular milieu and are subsequently internalized by recipient cells. Upon internalization, exosomes condition recipient cells by donating their cargos and/or activating various signal transduction pathways, consequently regulating physiological and pathophysiological processes. The role of exosomes in viral pathogenesis, especially human immunodeficiency virus type 1 [HIV-1] is beginning to unravel. Recent research reports suggest that exosomes from various sources play important but different roles in the pathogenesis of HIV-1. From these reports, it appears that the source of exosomes is the defining factor for the exosomal effect on HIV-1. In this review, we will describe how HIV-1 infection is modulated by exosomes and in turn how exosomes are targeted by HIV-1 factors. Finally, we will discuss potentially emerging therapeutic options based on exosomal cargos that may have promise in preventing HIV-1 transmission.

  10. Exosomes: Implications in HIV-1 Pathogenesis

    Directory of Open Access Journals (Sweden)

    Marisa N. Madison

    2015-07-01

    Full Text Available Exosomes are membranous nanovesicles of endocytic origin that carry host and pathogen derived genomic, proteomic, and lipid cargos. Exosomes are secreted by most cell types into the extracellular milieu and are subsequently internalized by recipient cells. Upon internalization, exosomes condition recipient cells by donating their cargos and/or activating various signal transduction pathways, consequently regulating physiological and pathophysiological processes. The role of exosomes in viral pathogenesis, especially human immunodeficiency virus type 1 [HIV-1] is beginning to unravel. Recent research reports suggest that exosomes from various sources play important but different roles in the pathogenesis of HIV-1. From these reports, it appears that the source of exosomes is the defining factor for the exosomal effect on HIV-1. In this review, we will describe how HIV-1 infection is modulated by exosomes and in turn how exosomes are targeted by HIV-1 factors. Finally, we will discuss potentially emerging therapeutic options based on exosomal cargos that may have promise in preventing HIV-1 transmission.

  11. Tumour exosome integrins determine organotropic metastasis.

    Science.gov (United States)

    Hoshino, Ayuko; Costa-Silva, Bruno; Shen, Tang-Long; Rodrigues, Goncalo; Hashimoto, Ayako; Tesic Mark, Milica; Molina, Henrik; Kohsaka, Shinji; Di Giannatale, Angela; Ceder, Sophia; Singh, Swarnima; Williams, Caitlin; Soplop, Nadine; Uryu, Kunihiro; Pharmer, Lindsay; King, Tari; Bojmar, Linda; Davies, Alexander E; Ararso, Yonathan; Zhang, Tuo; Zhang, Haiying; Hernandez, Jonathan; Weiss, Joshua M; Dumont-Cole, Vanessa D; Kramer, Kimberly; Wexler, Leonard H; Narendran, Aru; Schwartz, Gary K; Healey, John H; Sandstrom, Per; Labori, Knut Jørgen; Kure, Elin H; Grandgenett, Paul M; Hollingsworth, Michael A; de Sousa, Maria; Kaur, Sukhwinder; Jain, Maneesh; Mallya, Kavita; Batra, Surinder K; Jarnagin, William R; Brady, Mary S; Fodstad, Oystein; Muller, Volkmar; Pantel, Klaus; Minn, Andy J; Bissell, Mina J; Garcia, Benjamin A; Kang, Yibin; Rajasekhar, Vinagolu K; Ghajar, Cyrus M; Matei, Irina; Peinado, Hector; Bromberg, Jacqueline; Lyden, David

    2015-11-19

    Ever since Stephen Paget's 1889 hypothesis, metastatic organotropism has remained one of cancer's greatest mysteries. Here we demonstrate that exosomes from mouse and human lung-, liver- and brain-tropic tumour cells fuse preferentially with resident cells at their predicted destination, namely lung fibroblasts and epithelial cells, liver Kupffer cells and brain endothelial cells. We show that tumour-derived exosomes uptaken by organ-specific cells prepare the pre-metastatic niche. Treatment with exosomes from lung-tropic models redirected the metastasis of bone-tropic tumour cells. Exosome proteomics revealed distinct integrin expression patterns, in which the exosomal integrins α6β4 and α6β1 were associated with lung metastasis, while exosomal integrin αvβ5 was linked to liver metastasis. Targeting the integrins α6β4 and αvβ5 decreased exosome uptake, as well as lung and liver metastasis, respectively. We demonstrate that exosome integrin uptake by resident cells activates Src phosphorylation and pro-inflammatory S100 gene expression. Finally, our clinical data indicate that exosomal integrins could be used to predict organ-specific metastasis.

  12. Exo-MFA - A 13C metabolic flux analysis framework to dissect tumor microenvironment-secreted exosome contributions towards cancer cell metabolism.

    Science.gov (United States)

    Achreja, Abhinav; Zhao, Hongyun; Yang, Lifeng; Yun, Tae Hyun; Marini, Juan; Nagrath, Deepak

    2017-01-11

    Dissecting the pleiotropic roles of tumor micro-environment (TME) on cancer progression has been brought to the foreground of research on cancer pathology. Extracellular vesicles such as exosomes, transport proteins, lipids, and nucleic acids, to mediate intercellular communication between TME components and have emerged as candidates for anti-cancer therapy. We previously reported that cancer-associated fibroblast (CAF) derived exosomes (CDEs) contain metabolites in their cargo that are utilized by cancer cells for central carbon metabolism and promote cancer growth. However, the metabolic fluxes involved in donor cells towards packaging of metabolites in extracellular vesicles and exosome-mediated metabolite flux upregulation in recipient cells are still not known. Here, we have developed a novel empirical and computational technique, exosome-mediated metabolic flux analysis (Exo-MFA) to quantify flow of cargo from source cells to recipient cells via vesicular transport. Our algorithm, which is based on (13)C metabolic flux analysis, successfully predicts packaging fluxes to metabolite cargo in CAFs, dynamic changes in rate of exosome internalization by cancer cells, and flux of cargo release over time. We find that cancer cells internalize exosomes rapidly leading to depletion of extracellular exosomes within 24h. However, metabolite cargo significantly alters intracellular metabolism over the course of 24h by regulating glycolysis pathway fluxes via lactate supply. Furthermore, it can supply up to 35% of the TCA cycle fluxes by providing TCA intermediates and glutamine. Our algorithm will help gain insight into (i) metabolic interactions in multicellular systems (ii) biogenesis of extracellular vesicles and their differential packaging of cargo under changing environments, and (iii) regulation of cancer cell metabolism by its microenvironment.

  13. Organtropic Metastatic Secretomes and Exosomes in Breast Cancer

    Science.gov (United States)

    2014-10-01

    1. Analysis of exosome secretion in organ -tropic breast cancer models. The graph on the left represents exosome production per million BC cells...fluorescent dyes (i.e. CellVue). Figure 2. Breast cancer exosomes are efficiently uptaken by stromal cells. Pɘ.05 * Exosome production per...derived from parental MDA-MB-231 and organ -tropic variants, and that they can be efficiently labeled with fluorescent and near-infrared dyes . • We have

  14. Exosomes: mediators of communication in eukaryotes.

    Science.gov (United States)

    Lopez-Verrilli, María A; Court, Felipe A

    2013-01-01

    In addition to the established mechanisms of intercellular signaling, a new way of communication has gained much attention in the last decade: communication mediated by exosomes. Exosomes are nanovesicles (with a diameter of 40-120 nm) secreted into the extracellular space by the multivesicular endosome after its outer membrane fuses with the plasma membrane. Once released, exosomes modulate the response of the recipient cells that recognize them. This indicates that exosomes operate in a specific manner and participate in the regulation of the target cell. Remarkably, exosomes occur from unicellular organisms to mammals, suggesting an evolutionarily conserved mechanism of communication. In this review we describe the cascade of exosome formation, intracellular traffic, secretion, and internalization by recipient cells, and review their most relevant effects. We also highlight important steps that are still poorly understood.

  15. Exosome-bound WD repeat protein Monad inhibits breast cancer cell invasion by degrading amphiregulin mRNA.

    Directory of Open Access Journals (Sweden)

    Makio Saeki

    Full Text Available Increased stabilization of mRNA coding for key cancer genes can contribute to invasiveness. This is achieved by down-regulation of exosome cofactors, which bind to 3'-UTR in cancer-related genes. Here, we identified amphiregulin, an EGFR ligand, as a target of WD repeat protein Monad, a component of R2TP/prefoldin-like complex, in MDA-MB-231 breast cancer cells. Monad specifically interacted with both the 3'-UTR of amphiregulin mRNA and the RNA degrading exosome, and enhanced decay of amphiregulin transcripts. Knockdown of Monad increased invasion and this effect was abolished with anti-amphiregulin neutralizing antibody. These results suggest that Monad could prevent amphiregulin-mediated invasion by degrading amphiregulin mRNA.

  16. Functional Roles and Therapeutic Applications of Exosomes in Hepatocellular Carcinoma.

    Science.gov (United States)

    Santangelo, Laura; Battistelli, Cecilia; Montaldo, Claudia; Citarella, Franca; Strippoli, Raffaele; Cicchini, Carla

    2017-01-01

    Exosomes are important in intercellular communication. They assure the horizontal transfer of specific functional contents (i.e., proteins, lipids, RNA molecules, and circulating DNA) from donor to recipient cells. Notably, tumor-derived exosomes (TDEs) appear to be an important vehicle of specific signals in cancer, impacting on tumor growth and metastasis. Recent researches point to the characterization of exosomes in Hepatocellular Carcinoma (HCC), the major adult liver malignancy. In this review, we summarize current findings on HCC exosomes, focusing on the identification of noncoding RNAs as exosome-enriched functional regulators and new potential biomarkers. The great potential of exosomes in future HCC diagnostic and therapeutic approaches is underlined.

  17. Exosome mediated growth effect on the non-growing pre-B acute lymphoblastic leukemia cells at low starting cell density

    Science.gov (United States)

    Patel, Sapan J; Darie, Costel C; Clarkson, Bayard D

    2016-01-01

    Tumors contain heterogeneous cell populations and achieve dominance by functioning as collective systems. The mechanisms underlying the aberrant growth and interactions between cells are not very well understood. The pre-B acute lymphoblastic leukemia cells we studied were obtained directly from a patient with Ph+ ALL. A new Ph+ ALL cell line (ALL3) was established from the leukemic cells growing as ascitic cells in his pleural fluid. The patient died of his disease shortly after the cells were obtained. ALL3 cells grow well at high cell densities (HD), but not at low cell densities. ALL3 cells are very sensitive to potent tyrosine kinase inhibitors (TKIs) such as Dasatinib and PD166325, but less sensitive to AMN 107, Imatinib, and BMS 214662 (a farnesyl transferase inhibitor). Here, we show that the growth of the LD ALL3 cells can be stimulated to grow in the presence of diffusible, soluble factors secreted by ALL3 cells themselves growing at high density. We also show that exosomes, part of the secretome components, are also able to stimulate the growth of the non-growing LD ALL3 cells and modulate their proliferative behavior. Characterization of the exosome particles also showed that the HD ALL3 cells are able to secret them in large quantities and that they are capable of inducing the growth of the LD ALL3 cells without which they will not survive. Direct stimulation of non-growing LD ALL3 cells using purified exosomes shows that the ALL3 cells can also communicate with each other by means of exchange of exosomes independently of direct cell-cell contacts or diffusible soluble stimulatory factors secreted by HD ALL3 cells. PMID:27725845

  18. The p75 neurotrophin receptor evades the endolysosomal route in neuronal cells, favouring multivesicular bodies specialised for exosomal release

    Science.gov (United States)

    Escudero, Claudia A.; Lazo, Oscal M.; Galleguillos, Carolina; Parraguez, Jose I.; Lopez-Verrilli, Maria A.; Cabeza, Carolina; Leon, Luisa; Saeed, Uzma; Retamal, Claudio; Gonzalez, Alfonso; Marzolo, Maria-Paz; Carter, Bruce D.; Court, Felipe A.; Bronfman, Francisca C.

    2014-01-01

    ABSTRACT The p75 neurotrophin receptor (p75, also known as NGFR) is a multifaceted signalling receptor that regulates neuronal physiology, including neurite outgrowth, and survival and death decisions. A key cellular aspect regulating neurotrophin signalling is the intracellular trafficking of their receptors; however, the post-endocytic trafficking of p75 is poorly defined. We used sympathetic neurons and rat PC12 cells to study the mechanism of internalisation and post-endocytic trafficking of p75. We found that p75 internalisation depended on the clathrin adaptor protein AP2 and on dynamin. More surprisingly, p75 evaded the lysosomal route at the level of the early endosome, instead accumulating in two different types of endosomes, Rab11-positive endosomes and multivesicular bodies (MVBs) positive for CD63, a marker of the exosomal pathway. Consistently, depolarisation by KCl induced the liberation of previously endocytosed full-length p75 into the extracellular medium in exosomes. Thus, p75 defines a subpopulation of MVBs that does not mature to lysosomes and is available for exosomal release by neuronal cells. PMID:24569882

  19. Mesenchymal stem cell therapy ameliorates diabetic nephropathy via the paracrine effect of renal trophic factors including exosomes

    Science.gov (United States)

    Nagaishi, Kanna; Mizue, Yuka; Chikenji, Takako; Otani, Miho; Nakano, Masako; Konari, Naoto; Fujimiya, Mineko

    2016-01-01

    Bone marrow-derived mesenchymal stem cells (MSCs) have contributed to the improvement of diabetic nephropathy (DN); however, the actual mediator of this effect and its role has not been characterized thoroughly. We investigated the effects of MSC therapy on DN, focusing on the paracrine effect of renal trophic factors, including exosomes secreted by MSCs. MSCs and MSC-conditioned medium (MSC-CM) as renal trophic factors were administered in parallel to high-fat diet (HFD)-induced type 2 diabetic mice and streptozotocin (STZ)-induced insulin-deficient diabetic mice. Both therapies showed approximately equivalent curative effects, as each inhibited the exacerbation of albuminuria. They also suppressed the excessive infiltration of BMDCs into the kidney by regulating the expression of the adhesion molecule ICAM-1. Proinflammatory cytokine expression (e.g., TNF-α) and fibrosis in tubular interstitium were inhibited. TGF-β1 expression was down-regulated and tight junction protein expression (e.g., ZO-1) was maintained, which sequentially suppressed the epithelial-to-mesenchymal transition of tubular epithelial cells (TECs). Exosomes purified from MSC-CM exerted an anti-apoptotic effect and protected tight junction structure in TECs. The increase of glomerular mesangium substrate was inhibited in HFD-diabetic mice. MSC therapy is a promising tool to prevent DN via the paracrine effect of renal trophic factors including exosomes due to its multifactorial action. PMID:27721418

  20. Exosomes from bulk and stem cells from human prostate cancer have a differential microRNA content that contributes cooperatively over local and pre-metastatic niche.

    Science.gov (United States)

    Sánchez, Catherine A; Andahur, Eliana I; Valenzuela, Rodrigo; Castellón, Enrique A; Fullá, Juan A; Ramos, Christian G; Triviño, Juan C

    2016-01-26

    The different prostate cancer (PCa) cell populations (bulk and cancer stem cells, CSCs) release exosomes that contain miRNAs that could modify the local or premetastatic niche. The analysis of the differential expression of miRNAs in exosomes allows evaluating the differential biological effect of both populations on the niche, and the identification of potential biomarkers and therapeutic targets. Five PCa primary cell cultures were established to originate bulk and CSCs cultures. From them, exosomes were purified by precipitation for miRNAs extraction to perform a comparative profile of miRNAs by next generation sequencing in an Illumina platform. 1839 miRNAs were identified in the exosomes. Of these 990 were known miRNAs, from which only 19 were significantly differentially expressed: 6 were overexpressed in CSCs and 13 in bulk cells exosomes. miR-100-5p and miR-21-5p were the most abundant miRNAs. Bioinformatics analysis indicated that differentially expressed miRNAs are highly related with PCa carcinogenesis, fibroblast proliferation, differentiation and migration, and angiogenesis. Besides, miRNAs from bulk cells affects osteoblast differentiation. Later, their effect was evaluated in normal prostate fibroblasts (WPMY-1) where transfection with miR-100-5p, miR-21-5p and miR-139-5p increased the expression of metalloproteinases (MMPs) -2, -9 and -13 and RANKL and fibroblast migration. The higher effect was achieved with miR21 transfection. As conclusion, miRNAs have a differential pattern between PCa bulk and CSCs exosomes that act collaboratively in PCa progression and metastasis. The most abundant miRNAs in PCa exosomes are interesting potential biomarkers and therapeutic targets.

  1. An intestinal Trojan horse for gene delivery

    Science.gov (United States)

    Peng, Haisheng; Wang, Chao; Xu, Xiaoyang; Yu, Chenxu; Wang, Qun

    2015-02-01

    The intestinal epithelium forms an essential element of the mucosal barrier and plays a critical role in the pathophysiological response to different enteric disorders and diseases. As a major enteric dysfunction of the intestinal tract, inflammatory bowel disease is a genetic disease which results from the inappropriate and exaggerated mucosal immune response to the normal constituents in the mucosal microbiota environment. An intestine targeted drug delivery system has unique advantages in the treatment of inflammatory bowel disease. As a new concept in drug delivery, the Trojan horse system with the synergy of nanotechnology and host cells can achieve better therapeutic efficacy in specific diseases. Here, we demonstrated the feasibility of encapsulating DNA-functionalized gold nanoparticles into primary isolated intestinal stem cells to form an intestinal Trojan horse for gene regulation therapy of inflammatory bowel disease. This proof-of-concept intestinal Trojan horse will have a wide variety of applications in the diagnosis and therapy of enteric disorders and diseases.

  2. Micro RNA in Exosomes from HIV-Infected Macrophages

    Directory of Open Access Journals (Sweden)

    William W. Roth

    2015-12-01

    Full Text Available Exosomes are small membrane-bound vesicles secreted by cells that function to shuttle RNA and proteins between cells. To examine the role of exosomal micro RNA (miRNA during the early stage of HIV-1 infection we characterized miRNA in exosomes from HIV-infected macrophages, compared with exosomes from non-infected macrophages. Primary human monocytes from uninfected donors were differentiated to macrophages (MDM which were either mock-infected or infected with the macrophage-tropic HIV-1 BaL strain. Exosomes were recovered from culture media and separated from virus particles by centrifugation on iodixanol density gradients. The low molecular weight RNA fraction was prepared from purified exosomes. After pre-amplification, RNA was hybridized to microarrays containing probes for 1200 miRNA species of known and unknown function. We observed 48 miRNA species in both infected and uninfected MDM exosomes. Additionally, 38 miRNAs were present in infected-cell exosomes but not uninfected-cell exosomes. Of these, 13 miRNAs were upregulated in exosomes from HIV-infected cells, including 4 miRNA species that were increased by more than 10-fold. Though numerous miRNA species have been identified in HIV-infected cells, relatively little is known about miRNA content in exosomes from these cells. In the future, we plan to investigate whether the upregulated miRNA species we identified are increased in exosomes from HIV-1-positive patients.

  3. Micro RNA in Exosomes from HIV-Infected Macrophages.

    Science.gov (United States)

    Roth, William W; Huang, Ming Bo; Addae Konadu, Kateena; Powell, Michael D; Bond, Vincent C

    2015-12-22

    Exosomes are small membrane-bound vesicles secreted by cells that function to shuttle RNA and proteins between cells. To examine the role of exosomal micro RNA (miRNA) during the early stage of HIV-1 infection we characterized miRNA in exosomes from HIV-infected macrophages, compared with exosomes from non-infected macrophages. Primary human monocytes from uninfected donors were differentiated to macrophages (MDM) which were either mock-infected or infected with the macrophage-tropic HIV-1 BaL strain. Exosomes were recovered from culture media and separated from virus particles by centrifugation on iodixanol density gradients. The low molecular weight RNA fraction was prepared from purified exosomes. After pre-amplification, RNA was hybridized to microarrays containing probes for 1200 miRNA species of known and unknown function. We observed 48 miRNA species in both infected and uninfected MDM exosomes. Additionally, 38 miRNAs were present in infected-cell exosomes but not uninfected-cell exosomes. Of these, 13 miRNAs were upregulated in exosomes from HIV-infected cells, including 4 miRNA species that were increased by more than 10-fold. Though numerous miRNA species have been identified in HIV-infected cells, relatively little is known about miRNA content in exosomes from these cells. In the future, we plan to investigate whether the upregulated miRNA species we identified are increased in exosomes from HIV-1-positive patients.

  4. Registered report: Melanoma exosomes educate bone marrow progenitor cells toward a pro-metastatic phenotype through MET.

    Science.gov (United States)

    Lesnik, Jake; Antes, Travis; Kim, Jeewon; Griner, Erin; Pedro, Luisa

    2016-01-29

    The Reproducibility Project: Cancer Biology seeks to address growing concerns about reproducibility in scientific research by conducting replications of selected experiments from a number of high-profile papers in the field of cancer biology. The papers, which were published between 2010 and 2012, were selected on the basis of citations and Altmetric scores (Errington et al., 2014). This Registered Report describes the proposed replication plan of key experiments from "Melanoma exosomes educate bone marrow progenitor cells toward a pro-metastatic phenotype through MET" by Peinado and colleagues, published in Nature Medicine in 2012 (Peinado et al., 2012). The key experiments being replicated are from Figures 4E, as well as Supplementary Figures 1C and 5A. In these experiments, Peinado and colleagues show tumor exosomes enhance metastasis to bones and lungs, which is diminished by reducing Met expression in exosomes (Peinado et al., 2012). The Reproducibility Project: Cancer Biology is a collaboration between the Center for Open Science and Science Exchange and the results of the replications will be published in eLife.

  5. Keystone Symposia "ncRNAs in Development and Cancer", Vancouver, Canada: Increased release of exosomes and export of invasion-modulating miRNAs miR921, -23b, -and -224 from metastatic urothelial carcinoma cells

    DEFF Research Database (Denmark)

    Ostenfeld, Marie Stampe; Jeppesen, Dennis Kjølhede; Laurberg, Jens Reumert

    2013-01-01

    Cancer cells secrete soluble factors and various extracellular vesicles, including exosomes, into their tissue microenvironment. The secretion of exosomes is speculated to facilitate local invasion and increase the propensity of tumors to form distant metastases. Here we present a characterization...... of exosome vesicles from isogenic urothelial carcinoma cell lines, with different metastatic propensity by western blotting, electron microscopy, nanoparticle tracking analysis, dynamic light scattering, and profiling of 671 miRNAs by qRT-PCR. An increase in the number of multivesicular bodies and exosomes...... was observed for metastatic FL3 cells compared to isogenic non-metastatic T24 cells. The release was significantly inhibited by knockdown of Rab27b and pharmacological inhibition of nsmase2 by GW4869. miRNA profiling was conducted on parental cells and their secreted exosomes. Here, selective export of miR921...

  6. Characterization and proteomic analysis of ovarian cancer-derived exosomes.

    Science.gov (United States)

    Liang, Bing; Peng, Peng; Chen, She; Li, Lin; Zhang, Meijun; Cao, Dongyan; Yang, Jiaxin; Li, Haixia; Gui, Ting; Li, Xialu; Shen, Keng

    2013-03-27

    Ovarian cancer is the most lethal type of cancer among all frequent gynecologic malignancies, because most patients present with advanced disease at diagnosis. Exosomes are important intercellular communication vehicles, released by various cell types. Here we presented firstly the protein profile of highly purified exosomes derived from two ovarian cancer cell lines, OVCAR-3 and IGROV1. The exosomes derived from ovarian cancer cell lines were round and mostly 30-100 nm in diameter when viewed under an electron microscope. The exosomal marker proteins TSG101 and Alix were detected in exosome preparations. The range of density was between 1.09 g/ml and 1.15 g/ml. A total of 2230 proteins were identified from two ovarian cell-derived exosomes. Among them, 1017 proteins were identified in both exosomes including all of the major exosomal protein markers. There were 380 proteins that are not reported in the ExoCarta database. In addition to common proteins from exosomes of various origins, our results showed that ovarian cancer-derived exosomes also carried tissue specific proteins associated with tumorigenesis and metastasis, especially in ovarian carcinoma. Based on the known roles of exosomes in cellular communication, these data indicate that exosomes released by ovarian cancer cells may play important roles in ovarian cancer progression and provide a potential source of blood-based protein biomarkers.

  7. Exosomes in tumor microenvironment influence cancer progression and metastasis.

    Science.gov (United States)

    Kahlert, Christoph; Kalluri, Raghu

    2013-04-01

    Exosomes are small membrane vesicles of endocytic origin with a size of 50-100 nm. They can contain microRNAs, mRNAs, DNA fragments, and proteins, which are shuttled from a donor cell to recipient cells. Many different cell types including immune cells, mesenchymal cells, and cancer cells release exosomes. There is emerging evidence that cancer-derived exosomes contribute to the recruitment and reprogramming of constituents associated with tumor environment. Here, we discuss different mechanisms associated with biogenesis, payload, and transport of exosomes. We highlight the functional relevance of exosomes in cancer, as related to tumor microenvironment, tumor immunology, angiogenesis, and metastasis. Exosomes may exert an immunosuppressive function as well as trigger an anti-tumor response by presenting tumor antigens to dendritic cells. Exosomes may serve as cancer biomarkers and aid in the treatment of cancer.

  8. Let-7 microRNA family is selectively secreted into the extracellular environment via exosomes in a metastatic gastric cancer cell line.

    Directory of Open Access Journals (Sweden)

    Keiichi Ohshima

    Full Text Available BACKGROUND: Exosomes play a major role in cell-to-cell communication, targeting cells to transfer exosomal molecules including proteins, mRNAs, and microRNAs (miRNAs by an endocytosis-like pathway. miRNAs are small noncoding RNA molecules on average 22 nucleotides in length that regulate numerous biological processes including cancer pathogenesis and mediate gene down-regulation by targeting mRNAs to induce RNA degradation and/or interfering with translation. Recent reports imply that miRNAs can be stably detected in circulating plasma and serum since miRNAs are packaged by exosomes to be protected from RNA degradation. Thus, profiling exosomal miRNAs are in need to clarify intercellular signaling and discover a novel disease marker as well. METHODOLOGY/PRINCIPAL FINDINGS: Exosomes were isolated from cultured cancer cell lines and their quality was validated by analyses of transmission electron microscopy and western blotting. One of the cell lines tested, a metastatic gastric cancer cell line, AZ-P7a, showed the highest RNA yield in the released exosomes and distinctive shape in morphology. In addition, RNAs were isolated from cells and culture media, and profiles of these three miRNA fractions were obtained using microarray analysis. By comparing signal intensities of microarray data and the following validation using RT-PCR analysis, we found that let-7 miRNA family was abundant in both the intracellular and extracellular fractions from AZ-P7a cells, while low metastatic AZ-521, the parental cell line of AZ-P7a, as well as other cancer cell lines showed no such propensity. CONCLUSIONS/SIGNIFICANCE: The enrichment of let-7 miRNA family in the extracellular fractions, particularly, in the exosomes from AZ-P7a cells may reflect their oncogenic characteristics including tumorigenesis and metastasis. Since let-7 miRNAs generally play a tumor-suppressive role as targeting oncogenes such as RAS and HMGA2, our results suggest that AZ-P7a cells

  9. Hypoxic tumor cell modulates its microenvironment to enhance angiogenic and metastatic potential by secretion of proteins and exosomes.

    Science.gov (United States)

    Park, Jung Eun; Tan, Hon Sen; Datta, Arnab; Lai, Ruenn Chai; Zhang, Huoming; Meng, Wei; Lim, Sai Kiang; Sze, Siu Kwan

    2010-06-01

    Under hypoxia, tumor cells produce a secretion that modulates their microenvironment to facilitate tumor angiogenesis and metastasis. Here, we observed that hypoxic or reoxygenated A431 carcinoma cells exhibited enhanced angiogenic and metastatic potential such as reduced cell-cell and cell-extracellular matrix adhesion, increased invasiveness, and production of a secretion with increased chorioallantoic membrane angiogenic activity. Consistent with these observations, quantitative proteomics revealed that under hypoxia the tumor cells secreted proteins involved in angiogenesis, focal adhesion, extracellular matrix-receptor interaction, and immune cell recruitment. Unexpectedly, the secreted proteins were predominantly cytoplasmic and membrane proteins. Ultracentrifugation at 100,000 x g precipitated 54% of the secreted proteins and enriched for many exosome-associated proteins such as the tetraspanins and Alix and also proteins with the potential to facilitate angiogenesis and metastasis. Two tetraspanins, CD9 and CD81, co-immunoprecipitated. Together, these data suggested that tumor cells secrete proteins and exosomes with the potential to modulate their microenvironment and facilitate angiogenesis and metastasis.

  10. Exosome removal as a therapeutic adjuvant in cancer

    OpenAIRE

    Marleau Annette M; Chen Chien-Shing; Joyce James A; Tullis Richard H

    2012-01-01

    Abstract Exosome secretion is a notable feature of malignancy owing to the roles of these nanoparticles in cancer growth, immune suppression, tumor angiogenesis and therapeutic resistance. Exosomes are 30–100 nm membrane vesicles released by many cells types during normal physiological processes. Tumors aberrantly secrete large quantities of exosomes that transport oncoproteins and immune suppressive molecules to support tumor growth and metastasis. The role of exosomes in intercellular signa...

  11. Exosomes in Development and Therapy of Malignant Mesothelioma

    Science.gov (United States)

    2015-09-01

    1 AWARD NUMBER: W81XWH-14-1-0199 TITLE: Exosomes in Development and Therapy of Malignant Mesothelioma PRINCIPAL INVESTIGATOR: Arti Shukla...TO THE ABOVE ADDRESS. 1. REPORT DATE September 2015 2. REPORT TYPE Annual 3. DATES COVERED 1Sep2014 - 31Aug2015 4. TITLE AND SUBTITLE Exosomes in...SUPPLEMENTARY NOTES 14. ABSTRACT Exosomes are tiny vesicles that carry information from one body cell type to another. We proposed that exosomes

  12. Curb challenges of the "Trojan Horse" approach: smart strategies in achieving effective yet safe cell-penetrating peptide-based drug delivery.

    Science.gov (United States)

    Huang, Yongzhuo; Jiang, Yifan; Wang, Huiyuan; Wang, Jianxin; Shin, Meong Cheol; Byun, Youngro; He, Huining; Liang, Yanqin; Yang, Victor C

    2013-10-01

    Cell-penetrating peptide (CPP)-mediated intracellular drug delivery system, often specifically termed as "the Trojan horse approach", has become the "holy grail" in achieving effective delivery of macromolecular compounds such as proteins, DNA, siRNAs, and drug carriers. It is characterized by the unique cell- (or receptor-), temperature-, and payload-independent mechanisms, therefore offering potent means to improve poor cellular uptake of a variety of macromolecular drugs. Nevertheless, this "Trojan horse" approach also acts like a double-edged sword, causing serious safety and toxicity concerns to normal tissues or organs for in vivo application, due to lack of target selectivity of the powerful cell penetrating activity. To overcome this problem of potent yet non-selective penetration vs. targeting delivery, a number of "smart" strategies have been developed in recent years, including controllable CPP-based drug delivery systems based on various stimuli-responsive mechanisms. This review article provides a fundamental understanding of these smart systems, as well as a discussion of their real-time in vivo applicability.

  13. Research progress of exosomes in stem cells%外泌体及其在干细胞中的研究进展

    Institute of Scientific and Technical Information of China (English)

    胡国文; 汪泱; 沈晓黎; 邓志锋

    2013-01-01

    Exosomes are small secreted vesicles,which play key roles in cell-to-cell communication.Exosomes can be released by different types of cells,and help to exchange membrane and transfer proteins,mRNA,microRNA and organelles between cells.Exosomes modulate normal physiological processes by transforming gene regulatory network or epigenetic reorganization.Recent studies have found that exosomes secreted by stem cells can transport mRNA,microRNA and proteins effectively,and play an important role in regulating tissue regeneration.The generation of exosomes and the research progress of exosomes in stem cells are reviewed in this paper.%外泌体是在细胞间交流中起重要作用的小分泌囊泡,可由多种类型的细胞分泌,在细胞间起着质膜交换及转运蛋白质、mRNA、microRNA和细胞器等生命活性物质的作用.外泌体可通过改变基因调控网络或表观遗传重组来调控正常的生理过程.最近的研究发现,干细胞分泌的外泌体可以有效转运mRNA、microRNA和蛋白质,在调控组织再生方面发挥重要作用.该文对外泌体的生成及其在干细胞中的研究进展进行综述.

  14. DC/CIKs细胞通过无 miRNA 的 exosome 蛋白刺激后能增强对胰腺癌细胞的免疫作用%Increasing the immune activity of exosomes:the effect of miRNA-depleted exosome proteins on activating dendritic cell/cytokine-induced killer cells against pancreatic cancer

    Institute of Scientific and Technical Information of China (English)

    Ri-sheng QUE; Cheng LIN; Guo-ping DING; Zheng-rong WU; Li-ping CAO

    2016-01-01

    Background: Tumor-derived exosomes were considered to be potential candidates for tumor vaccines because they are abundant in immune-regulating proteins, whereas tumor exosomal miRNAs may induce immune tolerance, thereby having an opposite immune function. Objective: This study was designed to separate exosomal protein and depleted exosomal microRNAs (miRNAs), increasing the immune activity of exosomes for activating dendritic cell/cytokine-induced kil er cel s (DC/CIKs) against pancreatic cancer (PC). Methods:PC-derived exosomes (PEs) were extracted from cultured PANC-1 cel supernatants and then ruptured; this was fol owed by ultrafiltered exosome lysates (UELs). DCs were stimulated with lipopolysaccharide (LPS), PE, and UEL, fol owed by co-culture with CIKs. The anti-tumor effects of DC/CIKs against PC were evaluated by proliferation and kil ing rates, tumor ne-crosis factor-α(TNF-α) and perforin secretion. Exosomal miRNAs were depleted after lysis and ultrafiltration, while 128 proteins were retained, including several immune-activating proteins. Results: UEL-stimulated DC/CIKs showed a higher killing rate than LPS- and PE-stimulated DC/CIKs. Conclusions: miRNA-depleted exosome proteins may be promising agonists for specifical y activating DC/CIKs against PC.%目的:本文通过分离提取无小 RNA(miRNA)的外来体(exosome)刺激树突细胞/细胞因子活化杀伤细胞(DC/CIKs),激活其对于胰腺癌细胞的免疫杀伤作用。  创新点:无 miRNA的 exosome超速离心裂解产物可以通过激活 DC/CIKs 细胞增强其对肿瘤细胞的杀伤作用。  方法:通过收集PANC-1细胞的上清并超速离心提取其中的exosome。提取的DC细胞分别通过脂多糖、肿瘤来源exosome及无miRNA的exosome刺激后,与CIK细胞共培养。通过计算增值与杀伤效率,肿瘤坏死因子-α(TNF-α)及穿孔素的分泌,比较各组间CIK细胞对胰腺癌细胞的杀伤作用。  结论:经

  15. Adiponectin is partially associated with exosomes in mouse serum.

    Science.gov (United States)

    Phoonsawat, Worrawalan; Aoki-Yoshida, Ayako; Tsuruta, Takeshi; Sonoyama, Kei

    2014-06-06

    Exosomes are membrane vesicles 30-120 nm in diameter that are released by many cell types and carry a cargo of proteins, lipids, mRNA, and microRNA. Cultured adipocytes reportedly release exosomes that may play a role in cell-to-cell communication during the development of metabolic diseases. However, the characteristics and function of exosomes released from adipocytes in vivo remain to be elucidated. Clearly, adipocyte-derived exosomes could exist in the circulation and may be associated with adipocyte-specific proteins such as adipocytokines. We isolated exosomes from serum of mice by differential centrifugation and analyzed adiponectin, leptin, and resistin in the exosome fraction. Western blotting detected adiponectin but no leptin and only trace amounts of resistin in the exosome fraction. The adiponectin signal in the exosome fraction was decreased by proteinase K treatment and completely quenched by a combination of proteinase K and Triton X-100. Quantitative ELISA showed that the exosome fraction contains considerable amounts of adiponectin, but not leptin or resistin. The concentration of adiponectin in the serum and the ratio of adiponectin to total protein in the exosome fraction were lower in obese mice than in lean mice. These results suggest that a portion of adiponectin exists as a transmembrane protein in the exosomes in mouse serum. We propose adiponectin as a marker of exosomes released from adipocytes in vivo.

  16. Using exosomes, naturally-equipped nanocarriers, for drug delivery.

    Science.gov (United States)

    Batrakova, Elena V; Kim, Myung Soo

    2015-12-10

    Exosomes offer distinct advantages that uniquely position them as highly effective drug carriers. Comprised of cellular membranes with multiple adhesive proteins on their surface, exosomes are known to specialize in cell-cell communications and provide an exclusive approach for the delivery of various therapeutic agents to target cells. In addition, exosomes can be amended through their parental cells to express a targeting moiety on their surface, or supplemented with desired biological activity. Development and validation of exosome-based drug delivery systems are the focus of this review. Different techniques of exosome isolation, characterization, drug loading, and applications in experimental disease models and clinic are discussed. Exosome-based drug formulations may be applied to a wide variety of disorders such as cancer, various infectious, cardiovascular, and neurodegenerative disorders. Overall, exosomes combine benefits of both synthetic nanocarriers and cell-mediated drug delivery systems while avoiding their limitations.

  17. The biology and function of exosomes in cancer.

    Science.gov (United States)

    Kalluri, Raghu

    2016-04-01

    Humans circulate quadrillions of exosomes at all times. Exosomes are a class of extracellular vesicles released by all cells, with a size range of 40-150 nm and a lipid bilayer membrane. Exosomes contain DNA, RNA, and proteins. Exosomes likely remove excess and/or unnecessary constituents from the cells, functioning like garbage bags, although their precise physiological role remains unknown. Additionally, exosomes may mediate specific cell-to-cell communication and activate signaling pathways in cells they fuse or interact with. Exosomes are detected in the tumor microenvironment, and emerging evidence suggests that they play a role in facilitating tumorigenesis by regulating angiogenesis, immunity, and metastasis. Circulating exosomes can be used as liquid biopsies and noninvasive biomarkers for early detection, diagnosis, and treatment of cancer patients.

  18. Research progress in dendritic cell-derived exosomes%树突状细胞来源的exosomes研究进展

    Institute of Scientific and Technical Information of China (English)

    朱伟国; 朱建华

    2009-01-01

    Exosomes ale small vesicles that form within late endocytic compartments by various cell types.Exosomes from different cellular origins have different properties which ale functionally relevant to their distinct proteins derived from the producing cell and the microenvironment around.Dendritic cell-derived exosomes (Dex) which richly contain various bioactive molecules such as MHC-I/MHC-II and costimulatory molecules were shown to be able to induee immune response or immune tolerance in vivo and in vitro,which is similar to that induced by the parent dendritic cells:The immunogenic potential of Dex as cell-free vaccines has been highlighted widespreadly these years,exceptionally for their immunostimulatory properties in anticancer immunotherapy and their potential tolerogenesis in reducing transplantation rejections and autoimmune diseases.%Exosomes 是多种活细胞晚期内体分泌的小囊泡体,不同来源的 exosomes 其特异性功能与它所含的特异性蛋白质以及它所处的微环境密切相关.树突状细胞来源的 exosomes(Dex) 富含树突状细胞的MHC-Ⅰ/Ⅱ类分子、协同刺激分子等多种生物活性分子,在体内、外实验中显示出与树突状细胞相似的功能,可诱发机体免疫应答或诱导免疫耐受.作为一种新型的非细胞疫苗,exosomes在抗肿瘤免疫治疗以及抑制移植免疫排斥和自身免疫性疾病治疗等各方面的应用前景受到极大的关注.

  19. Exosomal HIF1α supports invasive potential of nasopharyngeal carcinoma-associated LMP1-positive exosomes.

    Science.gov (United States)

    Aga, M; Bentz, G L; Raffa, S; Torrisi, M R; Kondo, S; Wakisaka, N; Yoshizaki, T; Pagano, J S; Shackelford, J

    2014-09-11

    It has emerged recently that exosomes are potential carriers of pro-tumorigenic factors that participate in oncogenesis. However, whether oncogenic transcription factors are transduced by exosomes is unknown. Hypoxia-inducible factor-1α (HIF1α) transcriptionally regulates numerous key aspects of tumor development and progression by promoting a more aggressive tumor phenotype, characterized by increased proliferation and invasiveness coupled with neoangiogenesis. It has been shown that the principal oncoprotein of Epstein-Barr virus (EBV), latent membrane protein 1 (LMP1), drives oncogenic processes and tumor progression of the highly invasive EBV malignancy, nasopharyngeal carcinoma (NPC). We now demonstrate that endogenous HIF1α is detectable in exosomes and that LMP1 significantly increases levels of HIF1α in exosomes. HIF1 recovered from exosomes retains DNA-binding activity and is transcriptionally active in recipient cells after exosome uptake. We also show that treatment of EBV-negative cells with LMP1-exosomes increases migration and invasiveness of NP cell lines in functional assays, which correlates with the phenotype associated with epithelial-mesenchymal transition (EMT). In addition, we provide evidence that HIF1α itself participates in exosome-mediated pro-metastatic effects in recipient cells, as exosome-mediated delivery of active and inactive forms of HIF1α results in reciprocal changes in the expression of E- and N-cadherins associated with EMT. Further, immunohistochemical analysis of NPC tumor tissues revealed direct correlation between protein levels of LMP1 and of the endosome/exosome marker tetraspanin, CD63, which suggests an increase in exosome formation in this EBV-positive malignancy. We hypothesize that exosome-mediated transfer of functional pro-metastatic factors by LMP1-positive NPC cells to surrounding tumor cells promotes cancer progression.

  20. Cancer Cell-derived Exosomes Induce Mitogen-activated Protein Kinase-dependent Monocyte Survival by Transport of Functional Receptor Tyrosine Kinases.

    Science.gov (United States)

    Song, Xiao; Ding, Yanping; Liu, Gang; Yang, Xiao; Zhao, Ruifang; Zhang, Yinlong; Zhao, Xiao; Anderson, Gregory J; Nie, Guangjun

    2016-04-15

    Tumor-associated macrophages (TAM) play pivotal roles in cancer initiation and progression. Monocytes, the precursors of TAMs, normally undergo spontaneous apoptosis within 2 days, but can subsist in the inflammatory tumor microenvironment for continuous survival and generation of sufficient TAMs. The mechanisms underlying tumor-driving monocyte survival remain obscure. Here we report that cancer cell-derived exosomes were crucial mediators for monocyte survival in the inflammatory niche. Analysis of the survival-promoting molecules in monocytes revealed that cancer cell-derived exosomes activated Ras and extracellular signal-regulated kinases in the mitogen-activated protein kinase (MAPK) pathway, resulting in the prevention of caspase cleavage. Phosphorylated receptor tyrosine kinases (RTKs), such as phosphorylated epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor 2 (HER-2), were abundantly expressed in cancer cell-derived exosomes. Knock-out of EGFR or/and HER-2, or alternatively, inhibitors against their phosphorylation significantly disturbed the exosome-mediated activation of the MAPK pathway, inhibition of caspase cleavage, and increase in survival rate in monocytes. Moreover, the deprived survival-stimulating activity of exosomes due to null expression of EGFR and HER-2 could be restored by activation of another RTK, insulin receptor. Overall, our study uncovered a mechanism of tumor-associated monocyte survival and demonstrated that cancer cell-derived exosomes can stimulate the MAPK pathway in monocytes through transport of functional RTKs, leading to inactivation of apoptosis-related caspases. This work provides insights into the long sought question on monocyte survival prior to formation of plentiful TAMs in the tumor microenvironment.

  1. Biochemistry and Function of the RNA Exosomes

    DEFF Research Database (Denmark)

    Lubas, Michal Szymon; Chlebowski, Aleksander; Dziembowski, Andrzej

    2012-01-01

    Discovery of the evolutionary conserved RNA exosome was a milestone in RNA biology. First identified as an activity essential for the processing of ribosomal RNA, the exosome has since proved to be central for RNA processing and degradation in both the nucleus and the cytoplasm of eukaryotic cell...

  2. Epidermal Growth Factor Receptor in Prostate Cancer Derived Exosomes.

    Directory of Open Access Journals (Sweden)

    Geetanjali Kharmate

    Full Text Available Exosomes proteins and microRNAs have gained much attention as diagnostic tools and biomarker potential in various malignancies including prostate cancer (PCa. However, the role of exosomes and membrane-associated receptors, particularly epidermal growth factor receptor (EGFR as mediators of cell proliferation and invasion in PCa progression remains unexplored. EGFR is frequently overexpressed and has been associated with aggressive forms of PCa. While PCa cells and tissues express EGFR, it is unknown whether exosomes derived from PCa cells or PCa patient serum contains EGFR. The aim of this study was to detect and characterize EGFR in exosomes derived from PCa cells, LNCaP xenograft and PCa patient serum. Exosomes were isolated from conditioned media of different PCa cell lines; LNCaP xenograft serum as well as patient plasma/serum by differential centrifugation and ultracentrifugation on a sucrose density gradient. Exosomes were confirmed by electron microscopy, expression of exosomal markers and NanoSight™ analysis. EGFR expression was determined by western blot analysis and ELISA. This study demonstrates that exosomes may easily be derived from PCa cell lines, serum obtained from PCa xenograft bearing mice and clinical samples derived from PCa patients. Presence of exosomal EGFR in PCa patient exosomes may present a novel approach for measuring of the disease state. Our work will allow to build on this finding for future understanding of PCa exosomes and their potential role in PCa progression and as minimal invasive biomarkers for PCa.

  3. Exosomes and Exosomal miRNA in Respiratory Diseases

    Science.gov (United States)

    Alipoor, Shamila D.; Garssen, Johan; Movassaghi, Masoud

    2016-01-01

    Exosomes are nanosized vesicles released from every cell in the body including those in the respiratory tract and lungs. They are found in most body fluids and contain a number of different biomolecules including proteins, lipids, and both mRNA and noncoding RNAs. Since they can release their contents, particularly miRNAs, to both neighboring and distal cells, they are considered important in cell-cell communication. Recent evidence has shown their possible importance in the pathogenesis of several pulmonary diseases. The differential expression of exosomes and of exosomal miRNAs in disease has driven their promise as biomarkers of disease enabling noninvasive clinical diagnosis in addition to their use as therapeutic tools. In this review, we summarize recent advances in this area as applicable to pulmonary diseases.

  4. Exosomes and Exosomal miRNA in Respiratory Diseases

    Directory of Open Access Journals (Sweden)

    Shamila D. Alipoor

    2016-01-01

    Full Text Available Exosomes are nanosized vesicles released from every cell in the body including those in the respiratory tract and lungs. They are found in most body fluids and contain a number of different biomolecules including proteins, lipids, and both mRNA and noncoding RNAs. Since they can release their contents, particularly miRNAs, to both neighboring and distal cells, they are considered important in cell-cell communication. Recent evidence has shown their possible importance in the pathogenesis of several pulmonary diseases. The differential expression of exosomes and of exosomal miRNAs in disease has driven their promise as biomarkers of disease enabling noninvasive clinical diagnosis in addition to their use as therapeutic tools. In this review, we summarize recent advances in this area as applicable to pulmonary diseases.

  5. Exosomes in the pathogenesis, diagnostics and therapeutics of liver diseases.

    Science.gov (United States)

    Masyuk, Anatoliy I; Masyuk, Tatyana V; Larusso, Nicholas F

    2013-09-01

    Exosomes are small (30-100 nm in diameter) extracellular membrane-enclosed vesicles released by different cell types into the extracellular space or into biological fluids by exocytosis as a result of fusion of intracellular multivesicular bodies with the plasma membrane. The primary function of exosomes is intercellular communication with both beneficial (physiological) and harmful (pathological) potential outcomes. Liver cells are exosome-releasing cells as well as targets for endogenous exosomes and exosomes derived from cells of other organs. Despite limited studies on liver exosomes, initial observations suggest that these vesicles are important in liver physiology and pathophysiology. In this review, we briefly summarize the recent findings on liver exosomes, their functions and significance for novel diagnostic and therapeutic approaches.

  6. Organizing polarized delivery of exosomes at synapses.

    Science.gov (United States)

    Mittelbrunn, Maria; Vicente-Manzanares, Miguel; Sánchez-Madrid, Francisco

    2015-04-01

    Exosomes are extracellular vesicles that transport different molecules between cells. They are formed and stored inside multivesicular bodies (MVB) until they are released to the extracellular environment. MVB fuse along the plasma membrane, driving non-polarized secretion of exosomes. However, polarized signaling potentially directs MVBs to a specific point in the plasma membrane to mediate a focal delivery of exosomes. MVB polarization occurs across a broad set of cellular situations, e.g. in immune and neuronal synapses, cell migration and in epithelial sheets. In this review, we summarize the current state of the art of polarized MVB docking and the specification of secretory sites at the plasma membrane. The current view is that MVB positioning and subsequent exosome delivery requires a polarizing, cytoskeletal dependent-trafficking mechanism. In this context, we propose scenarios in which biochemical and mechanical signals could drive the polarized delivery of exosomes in highly polarized cells, such as lymphocytes, neurons and epithelia.

  7. New Insights into Regulatory T Cells: Exosome- and Non-Coding RNA-Mediated Regulation of Homeostasis and Resident Treg Cells

    Science.gov (United States)

    Li, Peiyao; Liu, Changhong; Yu, Zhibin; Wu, Minghua

    2016-01-01

    Regulatory T (Treg) cells are a group of cells that are heterogeneous in origin and in functional activity. Treg cells comprise a necessary balance to adaptive immune responses. As key regulators of self-tolerance, Treg cells have been involved in a series of pathologic processes and considered as therapeutic targets. Here, we summarize recent research regarding Treg cell origins and their functional classification, highlight the role of exosomes and non-coding RNA in modulating Treg cell homeostasis, and discuss the current understanding of resident Treg cells. PMID:27999575

  8. New Insights into Regulatory T Cells: Exosome- and Non-Coding RNA-Mediated Regulation of Homeostasis and Resident Treg Cells.

    Science.gov (United States)

    Li, Peiyao; Liu, Changhong; Yu, Zhibin; Wu, Minghua

    2016-01-01

    Regulatory T (Treg) cells are a group of cells that are heterogeneous in origin and in functional activity. Treg cells comprise a necessary balance to adaptive immune responses. As key regulators of self-tolerance, Treg cells have been involved in a series of pathologic processes and considered as therapeutic targets. Here, we summarize recent research regarding Treg cell origins and their functional classification, highlight the role of exosomes and non-coding RNA in modulating Treg cell homeostasis, and discuss the current understanding of resident Treg cells.

  9. Exosomes released by EBV-infected nasopharyngeal carcinoma cells convey the viral Latent Membrane Protein 1 and the immunomodulatory protein galectin 9

    Directory of Open Access Journals (Sweden)

    Hirashima Mitsuomi

    2006-12-01

    Full Text Available Abstract Background Nasopharyngeal carcinomas (NPC are consistently associated with the Epstein-Barr virus (EBV. Their malignant epithelial cells contain the viral genome and express several antigenic viral proteins. However, the mechanisms of immune escape in NPCs are still poorly understood. EBV-transformed B-cells have been reported to release exosomes carrying the EBV-encoded latent membrane protein 1 (LMP1 which has T-cell inhibitory activity. Although this report suggested that NPC cells could also produce exosomes carrying immunosuppressive proteins, this hypothesis has remained so far untested. Methods Malignant epithelial cells derived from NPC xenografts – LMP1-positive (C15 or negative (C17 – were used to prepare conditioned culture medium. Various microparticles and vesicles released in the culture medium were collected and fractionated by differential centrifugation. Exosomes collected in the last centrifugation step were further purified by immunomagnetic capture on beads carrying antibody directed to HLA class II molecules. Purified exosomes were visualized by electron microscopy and analysed by western blotting. The T-cell inhibitory activities of recombinant LMP1 and galectin 9 were assessed on peripheral blood mononuclear cells activated by CD3/CD28 cross-linking. Results HLA-class II-positive exosomes purified from C15 and C17 cell supernatants were containing either LMP1 and galectin 9 (C15 or galectin 9 only (C17. Recombinant LMP1 induced a strong inhibition of T-cell proliferation (IC50 = 0.17 nM. In contrast recombinant galectin 9 had a weaker inhibitory effect (IC50 = 46 nM with no synergy with LMP1. Conclusion This study provides the proof of concept that NPC cells can release HLA class-II positive exosomes containing galectin 9 and/or LMP1. It confirms that the LMP1 molecule has intrinsic T-cell inhibitory activity. These findings will encourage investigations of tumor exosomes in the blood of NPC patients and

  10. Exosomes in Tumor Microenvironment Influence Cancer Progression and Metastasis

    OpenAIRE

    2013-01-01

    Exosomes are small membrane vesicles of endocytic origin with a size of 50 – 100 nm. They can contain microRNAs, mRNAs, DNA fragments and proteins, which are shuttled from a donar cell to recipient cells. Many different cell types including immune cells, mesenchymal cells and cancer cells release exosomes. There is emerging evidence that cancer-derived exosomes contribute to the recruitment and reprogramming of constituents associated with tumor environment. Here, we discuss different mechani...

  11. AB059. Mutant KRAS cells release tumor specific exosomes that create an immunosuppressive phenotype in lung tumor microenvironment

    Science.gov (United States)

    Petanidis, Savvas; Domvri, Kalliopi; Nikolaos, Zogas; Zarogoulidis, Paul; Kioseoglou, Efrosini; Anestakis, Doxakis; Zarogoulidis, Konstantinos; Salifoglou, Athanasios

    2016-01-01

    Background To investigate whether KRAS-secreted exosomal RNAs contribute to metastatic related-EMT phenotype, and their association with the hypoxic immunosuppressive phenotype in lung carcinoma. Methods Lung cancer patient tissues and mice were used for exosome isolation. Exosomal molecular characterization was performed using Immunofluorescence microscopy, microRNA analysis and immunohistochemistry. Results The findings reveal that mutant KRAS secreted exosomes contain high expression of oncogenic miR-31/miR-182/miR-205 which are associated with HIF-2a-related hypoxia. Furthermore, exosomal secretion in lung tumor microenvironment caused accumulation of IL-10, CTL-4 and IDO suggesting KRAS-dependent exosomal immunosuppression that drives EMT-related lung metastasis. Conclusions These findings suggest that KRAS secreted exosomal miRNAs can modulate the hypoxic immunosuppressive mechanism in lung tumor microenvironment and trigger the EMT-related metastatic niche.

  12. Tumor exosomes: cellular postmen of cancer diagnosis and personalized therapy.

    Science.gov (United States)

    Sharma, Aman; Khatun, Zamila; Shiras, Anjali

    2016-02-01

    Nanosized (30-150 nm) extracellular vesicles 'exosomes' are secreted by cells for intercellular communication during normal and pathological conditions. Exosomes carry biomacromolecules from cell-of-origin and, therefore, represent molecular bioprint of the cell. Tumor-derived exosomes or TDEx modulate tumor microenvironment by transfer of macromolecules locally as well as at distant metastatic sites. Due to their biological stability, TDEx are rich source of biomarkers in cancer patients. TDEx focused cancer diagnosis allows liquid biopsy-based tumor typing and may facilitate therapy response monitoring by developing novel exosomes diagnostics. Therefore, efficient and specific capturing of exosomes for subsequent amplification of the biomessages; for example, DNA, RNA, miRNA can reinvent cancer diagnosis. Here, in this review, we discuss advancements in exosomes isolation strategies, presence of exosomes biomarkers and importance of TDEx in gauging tumor heterogeneity for their potential use in cancer diagnosis, therapy.

  13. Cartography for Martian Trojans

    CERN Document Server

    Tabachnik, S A

    1999-01-01

    The last few months have seen the discovery of a second Martian Trojan (1998 VF31), as well as two further possible candidates (1998 QH56 and 1998 SD4). Together with the previously discovered Martian satellite 5261 Eureka, these are the only known possible solar system Trojan asteroids not associated with Jupiter. Here, maps of the locations of the stable Trojan trajectories of Mars are presented. These are constructed by integrating an ensemble of in-plane and inclined orbits in the vicinity of the Martian Lagrange points for between 25 million and 60 million years. The survivors occupy a band of inclinations between 15 degrees and 40 degrees and longitudes between 240 degrees and 330 degrees at the L5 Lagrange point. Around the L4 point, stable Trojans inhabit two bands of inclinations (15 degrees < i < 30 degrees and 32 degrees < i < 40 degrees) with longitudes restricted between 25 degrees and 120 degrees. Both 5261 Eureka and 1998 VF31 lie deep within one of the stable zones, which suggests ...

  14. Exosomes and the kidney: blaming the messenger.

    Science.gov (United States)

    Fang, Doreen Yp; King, Hamish W; Li, Jordan Yz; Gleadle, Jonathan M

    2013-01-01

    Exosomes are membrane-bound vesicles of endosomal origin, present in a wide range of biological fluids, including blood and urine. They range between 30 and 100 nm in diameter, and consist of a limiting lipid bilayer, transmembrane proteins and a hydrophilic core containing proteins, mRNAs and microRNAs (miRNA). Exosomes can act as extracellular vehicles by which cells communicate, through the delivery of their functional cargo to recipient cells, with many important biological, physiological and pathological implications. The exosome release pathway contributes towards protein secretion, antigen presentation, pathogen transfer and cancer progression. Exosomes and exosome-mediated signalling have been implicated in disease processes such as atherosclerosis, calcification and kidney diseases. Circulating levels of exosomes and extracellular vesicles can be influenced by the progression of renal disease. Advances in methods for purification and analysis of exosomes are leading to potential diagnostic and therapeutic avenues for kidney diseases. This review will focus on biophysical properties and biogenesis of exosomes, their pathophysiological roles and their potential as biomarkers and therapeutics in kidney diseases.

  15. 3D plasmonic nanobowl platform for the study of exosomes in solution

    Science.gov (United States)

    Lee, Changwon; Carney, Randy P.; Hazari, Sidhartha; Smith, Zachary J.; Knudson, Alisha; Robertson, Christopher S.; Lam, Kit S.; Wachsmann-Hogiu, Sebastian

    2015-05-01

    Thin silver film coated nanobowl Surface Enhanced Raman Spectroscopy (SERS) substrates are used to capture exosomes in solution for SERS measurements that can provide biochemical analysis of intact and ruptured exosomes. Exosomes derived via Total Exosome Isolation Reagent (TEIR) as well as ultracentrifugation (UC) from the SKOV3 cell line were analyzed. Spectra of exosomes derived via TEIR are dominated by a signal characteristic for the TEIR kit that needs to be subtracted for all measurements. Differences in SERS spectra recorded at different times during the drying of the exosome solution are statistically analyzed with Principal Component Analysis (PCA). At the beginning of the drying process, SERS spectra of exosomes exhibit peaks characteristic for both lipids and proteins. Later on during the drying process, new SERS peaks develop, suggesting that the initially intact exosome ruptures over time. This time-dependent evolution of SERS peaks enables analysis of exosomal membrane contents and the contents inside the exosomes.

  16. Genetic engineering of mesenchymal stromal cells for cancer therapy: turning partners in crime into Trojan horses

    Directory of Open Access Journals (Sweden)

    Niess Hanno

    2016-09-01

    Full Text Available Mesenchymal stromal cells (MSCs are adult progenitor cells with a high migratory and differentiation potential, which influence a broad range of biological functions in almost every tissue of the body. Among other mechanisms, MSCs do so by the secretion of molecular cues, differentiation toward more specialized cell types, or influence on the immune system. Expanding tumors also depend on the contribution of MSCs to building a supporting stroma, but the effects of MSCs appear to go beyond the mere supply of connective tissues. MSCs show targeted “homing” toward growing tumors, which is then followed by exerting direct and indirect effects on cancer cells. Several research groups have developed novel strategies that make use of the tumor tropism of MSCs by engineering them to express a transgene that enables an attack on cancer growth. This review aims to familiarize the reader with the current knowledge about MSC biology, the existing evidence for MSC contribution to tumor growth with its underlying mechanisms, and the strategies that have been developed using MSCs to deploy an anticancer therapy.

  17. A Study of Jupiter Trojans

    OpenAIRE

    Karlsson, Ola

    2012-01-01

    Jupiter Trojan asteroid dynamics have been studied for a long time but it is only within the last decades that the known population has become large enough to make other studies meaningful. In four articles I have been scratching the surface of the unknown Trojan knowledge space. Paper I presents photometric observations confirming a larger variety in surface redness for the smaller Trojans compared to the larger ones, in line with the groups in the outer main asteroid belt. However, the larg...

  18. Comparative proteomics evaluation of plasma exosome isolation techniques and assessment of the stability of exosomes in normal human blood plasma.

    Science.gov (United States)

    Kalra, Hina; Adda, Christopher G; Liem, Michael; Ang, Ching-Seng; Mechler, Adam; Simpson, Richard J; Hulett, Mark D; Mathivanan, Suresh

    2013-11-01

    Exosomes are nanovesicles released by a variety of cells and are detected in body fluids including blood. Recent studies have highlighted the critical application of exosomes as personalized targeted drug delivery vehicles and as reservoirs of disease biomarkers. While these research applications have created significant interest and can be translated into practice, the stability of exosomes needs to be assessed and exosome isolation protocols from blood plasma need to be optimized. To optimize methods to isolate exosomes from blood plasma, we performed a comparative evaluation of three exosome isolation techniques (differential centrifugation coupled with ultracentrifugation, epithelial cell adhesion molecule immunoaffinity pull-down, and OptiPrep(TM) density gradient separation) using normal human plasma. Based on MS, Western blotting and microscopy results, we found that the OptiPrep(TM) density gradient method was superior in isolating pure exosomal populations, devoid of highly abundant plasma proteins. In addition, we assessed the stability of exosomes in plasma over 90 days under various storage conditions. Western blotting analysis using the exosomal marker, TSG101, revealed that exosomes are stable for 90 days. Interestingly, in the context of cellular uptake, the isolated exosomes were able to fuse with target cells revealing that they were indeed biologically active.

  19. Exosomes as nanocarriers for immunotherapy of cancer and inflammatory diseases.

    Science.gov (United States)

    Tran, Thanh-Huyen; Mattheolabakis, George; Aldawsari, Hibah; Amiji, Mansoor

    2015-09-01

    Cell secreted exosomes (30-100nm vesicles) play a major role in intercellular communication due to their ability to transfer proteins and nucleic acids from one cell to another. Depending on the originating cell type and the cargo, exosomes can have immunosuppressive or immunostimulatory effects, which have potential application as immunotherapies for cancer and autoimmune diseases. Cellular components shed from tumor cells or antigen presenting cells (APCs), such as dendritic cells, macrophages and B cells, have been shown to be efficiently packaged in exosomes. In this review, we focus on the application of exosomes as nanocarriers and immunological agents for cancer and autoimmune immunotherapy. APC-derived exosomes demonstrate effective therapeutic efficacy for the treatment of cancer and experimental autoimmune diseases such as rheumatoid arthritis, inflammatory bowel disease, and multiple sclerosis. In addition to their intrinsic immunomodulating activity, exosomes have many advantages over conventional nanocarriers for drug and gene delivery.

  20. Information transfer by exosomes: A new frontier in hematologic malignancies.

    Science.gov (United States)

    Boyiadzis, Michael; Whiteside, Theresa L

    2015-09-01

    Exosomes are small (30-150 mm) vesicles secreted by all cell types and present in all body fluids. They are emerging as vehicles for delivery of membrane-tethered signaling molecules and membrane enclosed genes to target cells. Exosome-mediated information transfer allows for crosstalk of cells within the hematopoietic system and for interactions between hematopoietic cells and local or distant tissue cells. Exosomes carry physiological signals essential for health and participate in pathological processes, including malignant transformation. In hematologic malignancies, exosomes reprogram the bone marrow microenvironment, creating a niche for abnormal cells and favoring their expansion. The molecular and genetic mechanisms exosomes utilize to shuttle information between cells are currently being examined as are the potential roles exosomes play as biomarkers of disease or future therapeutic targets.

  1. Designer exosomes as next-generation cancer immunotherapy.

    Science.gov (United States)

    Bell, Brandon M; Kirk, Isabel D; Hiltbrunner, Stefanie; Gabrielsson, Susanne; Bultema, Jarred J

    2016-01-01

    Exosomes are small 40-120 nm vesicles secreted by nearly all cells and are an important form of intercellular communication. Exosomes are abundant, stable, and highly bioavailable to tissues in vivo. Increasingly, exosomes are being recognized as potential therapeutics as they have the ability to elicit potent cellular responses in vitro and in vivo. Patient-derived exosomes have been employed as a novel cancer immunotherapy in several clinical trials, but at this point lack sufficient efficacy. Still other researchers have focused on modifying the content and function of exosomes in various ways, toward the end-goal of specialized therapeutic exosomes. Here we highlight major advances in the use of exosomes for cancer immunotherapy and exosome bioengineering followed by a discussion of focus areas for future research to generate potent therapeutic exosomes. From the Clinical Editor: Exosomes are small vesicles used by cells for intercellular communication. In this short article, the authors described the current status and the potential use of exosomes in the clinical setting.

  2. Exosomes as miRNA Carriers: Formation-Function-Future.

    Science.gov (United States)

    Yu, Xiaojie; Odenthal, Margarete; Fries, Jochen W U

    2016-12-02

    Exosomes, which are one of the smallest extracellular vesicles released from cells, have been shown to carry different nucleic acids, including microRNAs (miRNAs). miRNAs significantly regulate cell growth and metabolism by posttranscriptional inhibition of gene expression. The rapidly changing understanding of exosomes' formation and function in delivering miRNAs from cell to cell has prompted us to review current knowledge in exosomal miRNA secretion mechanisms as well as possible therapeutic applications for personalized medicine.

  3. A niche role for cancer exosomes in metastasis.

    Science.gov (United States)

    Zhang, Yun; Wang, Xiao-Fan

    2015-06-01

    Cancer cells are known to secrete exosomes with pro-metastatic effects. Pancreatic-cancer-derived exosomes are now shown to promote liver metastasis by eliciting pre-metastatic niche formation through a multi-step process. This involves uptake of exosome-derived factors by liver Kupffer cells and hepatic stellate cell activation to generate a fibrotic microenvironment with immune cell infiltrates that favours metastasis.

  4. Exosomes and exosomal miRNAs in cardiovascular protection and repair.

    Science.gov (United States)

    Emanueli, Costanza; Shearn, Andrew I U; Angelini, Gianni D; Sahoo, Susmita

    2015-08-01

    Cell-cell communication between cardiac and vascular cells and from stem and progenitor cells to differentiated cardiovascular cells is both an important and complex process, achieved through a diversity of mechanisms that have an impact on cardiovascular biology, disease and therapeutics. In recent years, evidence has accumulated suggesting that extracellular vesicles (EVs) are a new system of intercellular communication. EVs of different sizes are produced via different biogenesis pathways and have been shown to be released and taken up by most of known cell types, including heart and vascular cells, and stem and progenitor cells. This review will focus on exosomes, the smallest EVs (up to 100nm in diameter) identified so far. Cells can package cargoes consisting of selective lipids, proteins and RNA in exosomes and such cargoes can be shipped to recipient cells, inducing expressional and functional changes. This review focuses on exosomes and microRNAs in the context of cardiovascular disease and repair. We will describe exosome biogenesis and cargo formation and discuss the available information on in vitro and in vivo exosomes-based cell-to-cell communication relevant to cardiovascular science. The methods used in exosome research will be also described. Finally, we will address the promise of exosomes as clinical biomarkers and their impact as a biomedical tool in stem cell-based cardiovascular therapeutics.

  5. Exosomes: From Garbage Bins to Promising Therapeutic Targets.

    Science.gov (United States)

    H Rashed, Mohammed; Bayraktar, Emine; K Helal, Gouda; Abd-Ellah, Mohamed F; Amero, Paola; Chavez-Reyes, Arturo; Rodriguez-Aguayo, Cristian

    2017-03-02

    Intercellular communication via cell-released vesicles is a very important process for both normal and tumor cells. Cell communication may involve exosomes, small vesicles of endocytic origin that are released by all types of cells and are found in abundance in body fluids, including blood, saliva, urine, and breast milk. Exosomes have been shown to carry lipids, proteins, mRNAs, non-coding RNAs, and even DNA out of cells. They are more than simply molecular garbage bins, however, in that the molecules they carry can be taken up by other cells. Thus, exosomes transfer biological information to neighboring cells and through this cell-to-cell communication are involved not only in physiological functions such as cell-to-cell communication, but also in the pathogenesis of some diseases, including tumors and neurodegenerative conditions. Our increasing understanding of why cells release exosomes and their role in intercellular communication has revealed the very complex and sophisticated contribution of exosomes to health and disease. The aim of this review is to reveal the emerging roles of exosomes in normal and pathological conditions and describe the controversial biological role of exosomes, as it is now understood, in carcinogenesis. We also summarize what is known about exosome biogenesis, composition, functions, and pathways and discuss the potential clinical applications of exosomes, especially as biomarkers and novel therapeutic agents.

  6. Exosomes: From Garbage Bins to Promising Therapeutic Targets

    Directory of Open Access Journals (Sweden)

    Mohammed H. Rashed

    2017-03-01

    Full Text Available Intercellular communication via cell-released vesicles is a very important process for both normal and tumor cells. Cell communication may involve exosomes, small vesicles of endocytic origin that are released by all types of cells and are found in abundance in body fluids, including blood, saliva, urine, and breast milk. Exosomes have been shown to carry lipids, proteins, mRNAs, non-coding RNAs, and even DNA out of cells. They are more than simply molecular garbage bins, however, in that the molecules they carry can be taken up by other cells. Thus, exosomes transfer biological information to neighboring cells and through this cell-to-cell communication are involved not only in physiological functions such as cell-to-cell communication, but also in the pathogenesis of some diseases, including tumors and neurodegenerative conditions. Our increasing understanding of why cells release exosomes and their role in intercellular communication has revealed the very complex and sophisticated contribution of exosomes to health and disease. The aim of this review is to reveal the emerging roles of exosomes in normal and pathological conditions and describe the controversial biological role of exosomes, as it is now understood, in carcinogenesis. We also summarize what is known about exosome biogenesis, composition, functions, and pathways and discuss the potential clinical applications of exosomes, especially as biomarkers and novel therapeutic agents.

  7. Exosomes from Human Synovial-Derived Mesenchymal Stem Cells Prevent Glucocorticoid-Induced Osteonecrosis of the Femoral Head in the Rat

    Science.gov (United States)

    Guo, Shang-Chun; Tao, Shi-Cong; Yin, Wen-Jing; Qi, Xin; Sheng, Jia-Gen; Zhang, Chang-Qing

    2016-01-01

    Osteonecrosis of the femoral head (ONFH) represents a debilitating complication following glucocorticoid (GC)-based therapy. Synovial-derived mesenchymal stem cells (SMSCs) can exert protective effect in the animal model of GC-induced ONFH by inducing cell proliferation and preventing cell apoptosis. Recent studies indicate the transplanted cells exert therapeutic effects primarily via a paracrine mechanism and exosomes are an important paracrine factor that can be directly used as therapeutic agents for tissue engineering. Herein, we provided the first demonstration that the early treatment of exosomes secreted by human synovial-derived mesenchymal stem cells (SMSC-Exos) could prevent GC-induced ONFH in the rat model. Using a series of in vitro functional assays, we found that SMSC-Exos could be internalized into bone marrow derived stromal cells (BMSCs) and enhance their proliferation and have anti-apoptotic abilities. Finally, SMSC-Exos may be promising for preventing GC-induced ONFH.

  8. A Troop of Trojans: Photometry of 24 Jovian Trojan Asteroids

    Science.gov (United States)

    French, Linda M.; Stephens, R. D.; Coley, D.; Wasserman, L. H.; La Rocca, D.; Vilas, F.

    2014-01-01

    Because of their greater distance from the Sun, the Jovian Trojans have been less studied than main belt asteroids. Although they are numerous (nearly 6000 have well determined orbits as of July 2013), the Trojans remain mysterious in many ways. Their spectra are unlike those of any meteorites in terrestrial collections. The spectra and the low albedos of Trojans, however, bear a strong resemblance to those of cometary nuclei (Abell et al. 2005; Fornasier et al. 2007; Emery et al. 2011). The Nice Model (Morbidelli et al. 2005; 2009) predicts that the Trojans may well be objects that originated with today's Kuiper Belt Objects. The rotation of asteroids larger than ~50 km in diameter seems to be determined largely by collisions, while that of smaller bodies is shaped primarily by YORP forces and torques (Pravec et al. 2008). We are surveying the rotation properties of Trojans to see whether similar trends are present. We find an abundance of slow rotators, including the first documented tumbler among the Trojans. We present 24 new Trojan lightcurves (French et al. 2013), mostly from objects ranging from 30-50 km in diameter, as well as more recent observations. We also discuss observations of five sub-20 km Trojans, whose rotation properties are consistent with cometary densities.

  9. Detection of tumor cell-specific mRNA and protein in exosome-like microvesicles from blood and saliva.

    Science.gov (United States)

    Yang, Jieping; Wei, Fang; Schafer, Christopher; Wong, David T W

    2014-01-01

    The discovery of disease-specific biomarkers in oral fluids has revealed a new dimension in molecular diagnostics. Recent studies have reported the mechanistic involvement of tumor cells derived mediators, such as exosomes, in the development of saliva-based mRNA biomarkers. To further our understanding of the origins of disease-induced salivary biomarkers, we here evaluated the hypothesis that tumor-shed secretory lipidic vesicles called exosome-like microvesicles (ELMs) that serve as protective carriers of tissue-specific information, mRNAs, and proteins, throughout the vasculature and bodily fluids. RNA content was analyzed in cell free-saliva and ELM-enriched fractions of saliva. Our data confirmed that the majority of extracellular RNAs (exRNAs) in saliva were encapsulated within ELMs. Nude mice implanted with human lung cancer H460 cells expressing hCD63-GFP were used to follow the circulation of tumor cell specific protein and mRNA in the form of ELMs in vivo. We were able to identify human GAPDH mRNA in ELMs of blood and saliva of tumor bearing mice using nested RT-qPCR. ELMs positive for hCD63-GFP were detected in the saliva and blood of tumor bearing mice as well as using electric field-induced release and measurement (EFIRM). Altogether, our results demonstrate that ELMs carry tumor cell-specific mRNA and protein from blood to saliva in a xenografted mouse model of human lung cancer. These results therefore strengthen the link between distal tumor progression and the biomarker discovery of saliva through the ELMs.

  10. The carrying pigeons of the cell: exosomes and their role in infectious diseases caused by human pathogens.

    Science.gov (United States)

    Fleming, Adam; Sampey, Gavin; Chung, Myung-Chul; Bailey, Charles; van Hoek, Monique L; Kashanchi, Fatah; Hakami, Ramin M

    2014-07-01

    Exosomes have recently been classified as the newest family members of 'bioactive vesicles' that function to promote intercellular communication. Long ignored and thought to be only a mechanism by which cellular waste is removed, exosomes have garnered a huge amount of interest in recent years as their critical functions in maintaining homeostasis through intercellular communication and also in different types of diseases have been demonstrated. Many groundbreaking studies of exosome functions have been performed in the cancer field and the infectious disease areas of study, revealing the importance and also the fascinating complexity of exosomal packaging, targeting, and functions. Selective packaging of exosomes in response to the type of infection, exosomal modulation of the immune response and host signaling pathways, exosomal regulation of pathogen spread, and effects of exosomes on the degree of pathogenesis have all been well documented. In this review, we provide a synthesis of the current understanding of the role of exosomes during infections caused by human pathogens and discuss the implications of these findings for a better understanding of pathogenic mechanisms and future therapeutic and diagnostic applications.

  11. Emerging roles of exosomes in neuron-glia communication

    Directory of Open Access Journals (Sweden)

    Carsten eFrühbeis

    2012-04-01

    Full Text Available Brain function depends on coordinated interactions between neurons and glial cells. Recent evidence indicates that these cells release endosome-derived microvesicles termed exosomes, which are 50-100 nm in size and carry specific protein and RNA cargo. Exosomes can interact with neighboring cells raising the concept that exosomes may mediate signaling between brain cells and facilitate the delivery of bioactive molecules. Oligodendrocytes myelinate axons and furthermore maintain axonal integrity by an yet uncharacterized pathway of trophic support. Here, we highlight the role of exosomes in nervous system cell communication with particular focus on exosomes released by oligodendrocytes and their potential implications in axon-glia interaction and myelin disease, such as multiple sclerosis. These secreted vesicles may contribute to eliminate overproduced myelin membrane or to transfer antigens facilitating immune surveillance of the brain. Furthermore, there is emerging evidence that exosomes participate in axon-glia communication.

  12. Digital Detection of Exosomes by Interferometric Imaging

    OpenAIRE

    2016-01-01

    Exosomes, which are membranous nanovesicles, are actively released by cells and have been attributed to roles in cell-cell communication, cancer metastasis, and early disease diagnostics. The small size (30–100 nm) along with low refractive index contrast of exosomes makes direct characterization and phenotypical classification very difficult. In this work we present a method based on Single Particle Interferometric Reflectance Imaging Sensor (SP-IRIS) that allows multiplexed phenotyping and ...

  13. Exosomes in development, metastasis and drug resistance of breast cancer.

    Science.gov (United States)

    Yu, Dan-dan; Wu, Ying; Shen, Hong-yu; Lv, Meng-meng; Chen, Wei-xian; Zhang, Xiao-hui; Zhong, Shan-liang; Tang, Jin-hai; Zhao, Jian-hua

    2015-08-01

    Transport through the cell membrane can be divided into active, passive and vesicular types (exosomes). Exosomes are nano-sized vesicles released by a variety of cells. Emerging evidence shows that exosomes play a critical role in cancers. Exosomes mediate communication between stroma and cancer cells through the transfer of nucleic acid and proteins. It is demonstrated that the contents and the quantity of exosomes will change after occurrence of cancers. Over the last decade, growing attention has been paid to the role of exosomes in the development of breast cancer, the most life-threatening cancer in women. Breast cancer could induce salivary glands to secret specific exosomes, which could be used as biomarkers in the diagnosis of early breast cancer. Exosome-delivered nucleic acid and proteins partly facilitate the tumorigenesis, metastasis and resistance of breast cancer. Exosomes could also transmit anti-cancer drugs outside breast cancer cells, therefore leading to drug resistance. However, exosomes are effective tools for transportation of anti-cancer drugs with lower immunogenicity and toxicity. This is a promising way to establish a drug delivery system.

  14. Exosomes: Fundamental Biology and Roles in Cardiovascular Physiology.

    Science.gov (United States)

    Ibrahim, Ahmed; Marbán, Eduardo

    2016-01-01

    Exosomes are nanosized membrane particles that are secreted by cells that transmit information from cell to cell. The information within exosomes prominently includes their protein and RNA payloads. Exosomal microRNAs in particular can potently and fundamentally alter the transcriptome of recipient cells. Here we summarize what is known about exosome biogenesis, content, and transmission, with a focus on cardiovascular physiology and pathophysiology. We also highlight some of the questions currently under active investigation regarding these extracellular membrane vesicles and their potential in diagnostic and therapeutic applications.

  15. The human urinary exosome as a potential metabolic effector cargo.

    Science.gov (United States)

    Bruschi, Maurizio; Ravera, Silvia; Santucci, Laura; Candiano, Giovanni; Bartolucci, Martina; Calzia, Daniela; Lavarello, Chiara; Inglese, Elvira; Petretto, Andrea; Ghiggeri, Gianmarco; Panfoli, Isabella

    2015-08-01

    Exosomes are nanovesicles, derived from the endocytic pathway, released by most cell types and found in many body fluids, including urine. A variety of exosomal functions have been reported, including transfer of RNA, cell communication, control of apoptosis and protein lifespan. Exosomes from mesenchymal stem cells can rescue bioenergetics of injured cells. Here the urinary exosome proteome, non-urinary exosome proteome and urinome are compared. A consistent number of identified proteins cluster to metabolic functions. Cytoscape software analysis based on biological processes gene ontology database shows that metabolic pathways such as aerobic glycolysis and oxidative phosphorylation have a high probability (p ≤ 0.05) of being expressed and therefore functional. A metabolic function appears to be associated with human urinary exosomes, whose relevance experimental studies can assess.

  16. Placental exosomes in normal and complicated pregnancy.

    Science.gov (United States)

    Mitchell, Murray D; Peiris, Hassendrini N; Kobayashi, Miharu; Koh, Yong Q; Duncombe, Gregory; Illanes, Sebastian E; Rice, Gregory E; Salomon, Carlos

    2015-10-01

    While there is considerable contemporary interest in elucidating the role of placenta-derived extracellular vesicles in normal and complicated pregnancies and their utility as biomarkers and therapeutic interventions, progress in the field is hindered by a lack of standardized extracellular vesicle taxonomy and isolation protocols. The term "extracellular vesicle" is nonspecific and refers to all membrane-bound vesicles from nanometer to micrometer diameters and of different biogenic origins. To meaningfully ascribe biological function and/or diagnostic and therapeutic utility to extracellular vesicles, and in particular exosomes, greater specificity and vesicle characterization is required. The current literature relating to exosome biology must be interpreted in this context. Exosomes are a subtype of extracellular vesicle that are specifically defined by an endosomal biogenesis and particle size (40-120 nm) and density (1.13-1.19 g/mL(-1)). Exosomes are specifically package with signaling molecules (including protein, messenger RNA, microRNA, and noncoding RNA) and are released by exocytosis into biofluid compartments. Exosomes regulate the activity of both proximal and distal target cells, including translational activity, angiogenesis, proliferation, metabolism, and apoptosis. As such, exosomal signaling represents an integral pathway mediating intercellular communication. During pregnancy, the placenta releases exosomes into the maternal circulation from as early as 6 weeks of gestation. Release is regulated by factors that include both oxygen tension and glucose concentration and correlates with placental mass and perfusion. The concentration of placenta-derived exosomes in maternal plasma increases progressively during gestation. Exosomes isolated from maternal plasma are bioactive in vitro and are incorporated into target cells by endocytosis. While the functional significance of placental exosomes in pregnancy remains to be fully elucidated, available

  17. Isolation of biologically-active exosomes from human plasma.

    Science.gov (United States)

    Muller, Laurent; Hong, Chang-Sook; Stolz, Donna B; Watkins, Simon C; Whiteside, Theresa L

    2014-09-01

    Effects of exosomes present in human plasma on immune cells have not been examined in detail. Immunological studies with plasma-derived exosomes require their isolation by procedures involving ultracentrifugation. These procedures were largely developed using supernatants of cultured cells. To test biologic activities of plasma-derived exosomes, methods are necessary that ensure adequate recovery of exosome fractions free of contaminating larger vesicles, cell fragments and protein/nucleic acid aggregates. Here, an optimized method for exosome isolation from human plasma/serum specimens of normal controls (NC) or cancer patients and its advantages and pitfalls are described. To remove undesirable plasma-contaminating components, ultrafiltration of differentially-centrifuged plasma/serum followed by size-exclusion chromatography prior to ultracentrifugation facilitated the removal of contaminants. Plasma or serum was equally acceptable as a source of exosomes based on the recovered protein levels (in μg protein/mL plasma) and TEM image quality. Centrifugation on sucrose density gradients led to large exosome losses. Fresh plasma was the best source of morphologically-intact exosomes, while the use of frozen/thawed plasma decreased exosome purity but not their biologic activity. Treatments of frozen plasma with DNAse, RNAse or hyaluronidase did not improve exosome purity and are not recommended. Cancer patients' plasma consistently yielded more isolated exosomes than did NCs' plasma. Cancer patients' exosomes also mediated higher immune suppression as evidenced by decreased CD69 expression on responder CD4+ T effector cells. Thus, the described procedure yields biologically-active, morphologically-intact exosomes that have reasonably good purity without large protein losses and can be used for immunological, biomarker and other studies.

  18. Organotropic metastasis: role of tumor exosomes.

    Science.gov (United States)

    Liu, Yang; Cao, Xuetao

    2016-02-01

    A recent paper in Nature shows that tumor exosomes expressing unique integrins can determine organotropic metastasis by preparing pre-metastatic niche through their integrins-mediated fusion with and fertilization of organ-specific resident cells.

  19. Exosomes as drug delivery vehicles for Parkinson's disease therapy.

    Science.gov (United States)

    Haney, Matthew J; Klyachko, Natalia L; Zhao, Yuling; Gupta, Richa; Plotnikova, Evgeniya G; He, Zhijian; Patel, Tejash; Piroyan, Aleksandr; Sokolsky, Marina; Kabanov, Alexander V; Batrakova, Elena V

    2015-06-10

    Exosomes are naturally occurring nanosized vesicles that have attracted considerable attention as drug delivery vehicles in the past few years. Exosomes are comprised of natural lipid bilayers with the abundance of adhesive proteins that readily interact with cellular membranes. We posit that exosomes secreted by monocytes and macrophages can provide an unprecedented opportunity to avoid entrapment in mononuclear phagocytes (as a part of the host immune system), and at the same time enhance delivery of incorporated drugs to target cells ultimately increasing drug therapeutic efficacy. In light of this, we developed a new exosomal-based delivery system for a potent antioxidant, catalase, to treat Parkinson's disease (PD). Catalase was loaded into exosomes ex vivo using different methods: the incubation at room temperature, permeabilization with saponin, freeze-thaw cycles, sonication, or extrusion. The size of the obtained catalase-loaded exosomes (exoCAT) was in the range of 100-200nm. A reformation of exosomes upon sonication and extrusion, or permeabilization with saponin resulted in high loading efficiency, sustained release, and catalase preservation against proteases degradation. Exosomes were readily taken up by neuronal cells in vitro. A considerable amount of exosomes was detected in PD mouse brain following intranasal administration. ExoCAT provided significant neuroprotective effects in in vitro and in vivo models of PD. Overall, exosome-based catalase formulations have a potential to be a versatile strategy to treat inflammatory and neurodegenerative disorders.

  20. Cancer Exosomes as Mediators of Drug Resistance.

    Science.gov (United States)

    André, Maria do Rosário; Pedro, Ana; Lyden, David

    2016-01-01

    In the last decades, several studies demonstrated that the tumor microenvironment is a critical determinant not only of tumor progression and metastasis, but also of resistance to therapy. Exosomes are small membrane vesicles of endocytic origin, which contain mRNAs, DNA fragments, and proteins, and are released by many different cell types, including cancer cells. Mounting evidence has shown that cancer-derived exosomes contribute to the recruitment and reprogramming of constituents associated with the tumor microenvironment. Understanding how exosomes and the tumor microenvironment impact drug resistance will allow novel and better strategies to overcome drug resistance and treat cancer. Here, we describe a technique for exosome purification from cell culture, and fresh and frozen plasma, and further analysis by electron microscopy, NanoSight microscope, and Western blot.

  1. Regulation of exosome release by glycosphingolipids and flotillins.

    Science.gov (United States)

    Phuyal, Santosh; Hessvik, Nina P; Skotland, Tore; Sandvig, Kirsten; Llorente, Alicia

    2014-05-01

    Exosomes are released by cells after fusion of multivesicular bodies with the plasma membrane. The molecular mechanism of this process is still unclear. We investigated the role of sphingolipids and flotillins, which constitute a raft-associated family of proteins, in the release of exosomes. Interestingly, our results show that dl-threo-1-phenyl-2-decanoylamino-3-morpholino-1-propanol, an inhibitor of glucosylceramide synthase, seemed to affect the composition of exosomes released from PC-3 cells. However, the inhibition of ceramide formation from the de novo pathway by fumonisin B1 did not affect exosome secretion. Moreover, in contrast to findings obtained with other cell lines published so far, inhibition of neutral sphingomyelinase 2, an enzyme that catalyzes the formation of ceramide from sphingomyelin, did not inhibit the secretion of exosomes in PC-3 cells. Finally, small interfering RNA-mediated downregulation of flotillin-1 and flotillin-2 did not significantly change the levels of released exosomes as such, but seemed to affect the composition of exosomes. In conclusion, our results reveal the involvement of glycosphingolipids and flotillins in the release of exosomes from PC-3 cells, and indicate that the role of ceramide in exosome formation may be cell-dependent.

  2. Emerging potential of exosomes for treatment of traumatic brain injury

    Directory of Open Access Journals (Sweden)

    Ye Xiong

    2017-01-01

    Full Text Available Traumatic brain injury (TBI is one of the major causes of death and disability worldwide. No effective treatment has been identified from clinical trials. Compelling evidence exists that treatment with mesenchymal stem cells (MSCs exerts a substantial therapeutic effect after experimental brain injury. In addition to their soluble factors, therapeutic effects of MSCs may be attributed to their generation and release of exosomes. Exosomes are endosomal origin small-membrane nano-sized vesicles generated by almost all cell types. Exosomes play a pivotal role in intercellular communication. Intravenous delivery of MSC-derived exosomes improves functional recovery and promotes neuroplasticity in rats after TBI. Therapeutic effects of exosomes derive from the exosome content, especially microRNAs (miRNAs. miRNAs are small non-coding regulatory RNAs and play an important role in posttranscriptional regulation of genes. Compared with their parent cells, exosomes are more stable and can cross the blood-brain barrier. They have reduced the safety risks inherent in administering viable cells such as the risk of occlusion in microvasculature or unregulated growth of transplanted cells. Developing a cell-free exosome-based therapy may open up a novel approach to enhancing multifaceted aspects of neuroplasticity and to amplifying neurological recovery, potentially for a variety of neural injuries and neurodegenerative diseases. This review discusses the most recent knowledge of exosome therapies for TBI, their associated challenges and opportunities.

  3. Emerging potential of exosomes for treatment of traumatic brain injury

    Science.gov (United States)

    Xiong, Ye; Mahmood, Asim; Chopp, Michael

    2017-01-01

    Traumatic brain injury (TBI) is one of the major causes of death and disability worldwide. No effective treatment has been identified from clinical trials. Compelling evidence exists that treatment with mesenchymal stem cells (MSCs) exerts a substantial therapeutic effect after experimental brain injury. In addition to their soluble factors, therapeutic effects of MSCs may be attributed to their generation and release of exosomes. Exosomes are endosomal origin small-membrane nano-sized vesicles generated by almost all cell types. Exosomes play a pivotal role in intercellular communication. Intravenous delivery of MSC-derived exosomes improves functional recovery and promotes neuroplasticity in rats after TBI. Therapeutic effects of exosomes derive from the exosome content, especially microRNAs (miRNAs). miRNAs are small non-coding regulatory RNAs and play an important role in posttranscriptional regulation of genes. Compared with their parent cells, exosomes are more stable and can cross the blood-brain barrier. They have reduced the safety risks inherent in administering viable cells such as the risk of occlusion in microvasculature or unregulated growth of transplanted cells. Developing a cell-free exosome-based therapy may open up a novel approach to enhancing multifaceted aspects of neuroplasticity and to amplifying neurological recovery, potentially for a variety of neural injuries and neurodegenerative diseases. This review discusses the most recent knowledge of exosome therapies for TBI, their associated challenges and opportunities.

  4. New insights into Regulatory T cells:exosome and non-coding RNA mediated regulation of homeostasis, and resident regulatory T cells

    Directory of Open Access Journals (Sweden)

    Peiyao Li

    2016-12-01

    Full Text Available Regulatory T (Treg cells are a population of cells that are heterogeneous in origin and in functional activity. Treg cells constitute an essential counterbalance to adaptive immune responses. As key regulators of self-tolerance, Treg cells have been implicated in a number of pathologic processes and considered as therapeutic targets. Here, we summarize recent research regarding Treg cell origins and their functional classification, highlight the role of exosomes and non-coding RNA in modulating Treg cell homeostasis, and discuss the current understanding of resident Treg cells.

  5. Trojan twin planets

    Science.gov (United States)

    Dvorak, R.; Loibnegger, B.; Schwarz, R.

    2017-03-01

    The Trojan asteroids are moving in the vicinity of the stable Lagrange points L_4 and L_5 of the gas giants Jupiter, Uranus and Neptune. Their motion can be described and understood with the aid of the restricted three-body problem. As an extension of this problem we investigate how stable motion close to the Lagrange points of two massive bodies can exist. This configuration can be described as the Trojan Twin Problem when we regard the two additional bodies as having a mass significantly smaller than the the two primary bodies: a star in the center (m_1) and an additional Jupiter-like mass (m_2). Using this 4-body problem we have undertaken numerical investigations concerning possible stable "twin orbits". However, these two bodies (m_3 and m_4) in Trojan-like orbits may have quite different masses. We decided to choose 6 different scenaria for this problem: as primary body, m2, we have taken a Jupiter-like planet, a Saturn-like one, and a super-Earth with 10 Earthmasses (m_{Earth}) respectively. As quasi twin planets, we have used different mass ratios namely objects for m3 and m4 from 10m_{Earth} to Moon like ones. We found different stable configurations depending on the involved masses and the initial distances between the twins (always close to the Lagrange point). Although the formation of such a configuration seems to be not very probable we should not exclude that it exists regarding the huge number of planets even in our own galaxy. This model is of special interest when the most massive planet (m_2) is moving on an orbit in the habitable zone around a main sequence star. One can use our results of stable orbits of Trojan Twin Planets (or asteroids) for extrasolar systems having as second primary a Jupiter-like, a Saturn-like or a super-Earth like planet around a star similar to our Sun.

  6. Isolation and immunologic characteristics of exosomes derived from colon carcinoma cells%结肠癌源性exosomes的分离及其相关免疫学性质

    Institute of Scientific and Technical Information of China (English)

    冯业童; 刘朋飞; 吴昊昱; 刘迪; 董超; 吴璇; 周余来; 孙波

    2012-01-01

    目的 分离结肠癌细胞株的exosomes,并分析其在致敏抗原呈递细胞及激活相关效应细胞过程中的作用.方法 差速离心法分离体外培养的正常exosomes和经热休克处理的sw1116细胞(Heat shocked sw1116,HS-sw1116)分泌的exosomes (Heat shocked exosomes,HS-Exo),并在电子显微镜下观察exosomes和HS-Exo的形态结构;SDS-PAGE初步分析exosomes和HS-Exo的蛋白组分,CCK-8法检测其促外周血单个核细胞(Peripheral blood monouclear cells,PBMCs)增殖的能力.结果 电子显微镜观察,exosomes和HS-Exo的形态学结构无明显差异,其平均直径约为150 nm;exosomes和HS-Exo的蛋白条带分布情况基本相同,在高相对分子质量区域蛋白分布较多;exosomes比sw1116细胞更易引起PBMCs的增殖反应,HS-sw1116细胞和HS-Exo促PBMCs增殖的作用比sw1116细胞和exosomes更明显(P<0.05).结论 结肠癌sw1116细胞株可分泌exosomes,其比肿瘤细胞更易引起PBMCs的增殖,热休克处理可进一步增强细胞和exosomes的促PBMCs增殖的能力,exosomes在结肠癌免疫治疗方面具有重要的应用价值.%Objective To isolate exosomes from colon carcinoma cell strain and analyze its role in sensitization of antigen-presenting cells (APCs) and activation of effecter cells. Methods Normal exosomes cultured in vitro and heat shocked exosomes (HS-Exo) were isolated by differential centrifugation and observed for morphology by electron microscopy. The protein components of exosomes and HS-Exo were preliminarily analyzed by SDS-PAGE, and their abilities in promoting the proliferation of peripheral blood monouclear cells(PBMCs) by CCK-8 method. Results Normal exosomes and HS-Exo showed no significant difference in morphology under electron microscope, of which the mean diameter was about 150 nm. The distributions of protein bands of exosomes and HS-Exo were similar, which were mainly in the zones with high relative molecular masses. Compared with sw1116 cells, exosomes

  7. The Capture of Jupiter Trojans

    Science.gov (United States)

    Morbidelli, A.; Nesvorny, D.; Vokrouhlicky, D.

    2013-09-01

    The origin of Jupiter Trojans remained mysterious for decades. Particularly, it was difficult to explain the excitation of the inclinations of the Trojan population [1]. In 2005, Morbidelli et al. [2] proposed a scenario of capture from the trans-Neptunian disk, in the framework of the so-called "Nice model" [3,4]. This scenario explained in a natural way the observed orbital distribution of Trojans. The Nice model, however, evolved in the years, in order to satisfy an increasingly large number of constraints. It now appears that the dynamical evolution of the giant planets was different from that envisioned in [2]. Here, we assess again the process of capture of Trojans within this new evolution. We show that (6-8)×10 - 7 of the original trans-Neptunian planetesimals are captured in the Trojan region, with an orbital distribution consistent with the one observed. Relative to [2], the new capture mechanism has the potential of explaining the asymmetry between the L4 and L5 populations. Moreover, the resulting population of Trojans is consistent with that of the Irregular Satellites of Jupiter, which are captured in the same process; a few bodies from the main asteroid belt could also be captured in the Trojan cloud.

  8. Formation and role of exosomes in cancer.

    Science.gov (United States)

    Brinton, Lindsey T; Sloane, Hillary S; Kester, Mark; Kelly, Kimberly A

    2015-02-01

    Exosomes offer new insight into cancer biology with both diagnostic and therapeutic implications. Because of their cell-to-cell communication, exosomes influence tumor progression, metastasis, and therapeutic efficacy. They can be isolated from blood and other bodily fluids to reveal disease processes occurring within the body, including cancerous growth. In addition to being a reservoir of cancer biomarkers, they can be re-engineered to reinstate tumor immunity. Tumor exosomes interact with various cells of the microenvironment to confer tumor-advantageous changes that are responsible for stromal activation, induction of the angiogenic switch, increased vascular permeability, and immune escape. Exosomes also contribute to metastasis by aiding in the epithelial-to-mesenchymal transition and formation of the pre-metastatic niche. Furthermore, exosomes protect tumor cells from the cytotoxic effects of chemotherapy drugs and transfer chemoresistance properties to nearby cells. Thus, exosomes are essential to many lethal elements of cancer and it is important to understand their biogenesis and role in cancer.

  9. Characteristics and Roles of Exosomes in Cardiovascular Disease.

    Science.gov (United States)

    Zhang, Yuan; Hu, Yan-Wei; Zheng, Lei; Wang, Qian

    2017-03-01

    Exosomes are nano-sized biological membrane-enclosed vesicles that contain a cell-specific cargo of proteins, lipids, and nucleic acids that are released and taken up by most cell types, thereby inducing expression and functional changes via horizontal transfer of cargos between cells. Thus, exosomes present a largely unknown "cell-to-cell" communication system, which is now increasingly being investigated for diagnostic and therapeutic use in cardiovascular disease (CVD). The purpose of this review is to summarize recent findings on the properties and roles of exosomes in a variety of physiological and pathological settings related to CVD. We focus on available information on exosome-mediated intercellular communication relevant to myocardial injury, repair, and regeneration. Finally, we address the promise of exosomes as valuable diagnostic and prognostic biomarkers, and their potential use as therapeutic tools in CVD. Exosomes remain largely unexplored for therapeutic use in the field of cardiovascular diagnosis and medicine. A more detailed characterization of cardiac exosomes shed by different components of the heart will be of fundamental importance to address specific changes in the profile of exosomal microRNAs and proteins, which will enable the clinical use of exosomes as minimally invasive diagnostic tools and vehicles for delivery of targeted therapies for CVD.

  10. Effects of subtoxic concentrations of TiO{sub 2} and ZnO nanoparticles on human lymphocytes, dendritic cells and exosome production

    Energy Technology Data Exchange (ETDEWEB)

    Andersson-Willman, Britta; Gehrmann, Ulf; Cansu, Zekiye [Translational Immunology Unit, Department of Medicine Solna, Karolinska Institutet, Stockholm (Sweden); Buerki-Thurnherr, Tina; Krug, Harald F. [Laboratory for Materials — Biology Interactions, Swiss Federal Laboratories of Materials Testing and Research, St. Gallen (Switzerland); Gabrielsson, Susanne [Translational Immunology Unit, Department of Medicine Solna, Karolinska Institutet, Stockholm (Sweden); Scheynius, Annika, E-mail: annika.scheynius@ki.se [Translational Immunology Unit, Department of Medicine Solna, Karolinska Institutet, Stockholm (Sweden)

    2012-10-01

    Metal oxide nanoparticles are widely used in the paint and coating industry as well as in cosmetics, but the knowledge of their possible interactions with the immune system is very limited. Our aims were to investigate if commercially available TiO{sub 2} and ZnO nanoparticles may affect different human immune cells and their production of exosomes, nano-sized vesicles that have a role in cell to cell communication. We found that the TiO{sub 2} or ZnO nanoparticles at concentrations from 1 to 100 μg/mL did not affect the viability of primary human peripheral blood mononuclear cells (PBMC). In contrast, monocyte-derived dendritic cells (MDDC) reacted with a dose dependent increase in cell death and caspase activity to ZnO but not to TiO{sub 2} nanoparticles. Non-toxic exposure, 10 μg/mL, to TiO{sub 2} and ZnO nanoparticles did not significantly alter the phenotype of MDDC. Interestingly, ZnO but not TiO{sub 2} nanoparticles induced a down regulation of FcγRIII (CD16) expression on NK-cells in the PBMC population, suggesting that subtoxic concentrations of ZnO nanoparticles might have an effect on FcγR-mediated immune responses. The phenotype and size of exosomes produced by PBMC or MDDC exposed to the nanoparticles were similar to that of exosomes harvested from control cultures. TiO{sub 2} or ZnO nanoparticles could not be detected within or associated to exosomes as analyzed with TEM. We conclude that TiO{sub 2} and ZnO nanoparticles differently affect immune cells and that evaluations of nanoparticles should be performed even at subtoxic concentrations on different primary human immune cells when investigating potential effects on immune functions. -- Highlights: ► ZnO nanoparticles induce cell death of MDDC but not of PBMC. ► ZnO nanoparticles induce caspase activation and DNA fragmentation in MDDC. ► TiO{sub 2} nanoparticles are taken up by MDDC but have no effect on their phenotype. ► ZnO nanoparticles induce a significant reduction of CD16

  11. Exosomes as a Nanodelivery System: a Key to the Future of Neuromedicine?

    Science.gov (United States)

    Aryani, Arian; Denecke, Bernd

    2016-03-01

    Since the beginning of the last decade, exosomes have been of increased interest in the science community. Exosomes represent a new kind of long distance transfer of biological molecules among cells. This review provides a comprehensive overview about the construction of exosomes, their targeting and their fusion mechanisms to the recipient cells. Complementarily, the current state of research regarding the cargo of exosomes is discussed. A particular focus was placed on the role of exosomes in the central nervous system. An increasing number of physiological processes in the brain could be associated with exosomes. In this context, it is becoming more apparent that exosomes are involved in several neurological and specifically neurodegenerative diseases. The treatment of these kinds of diseases is often difficult not least because of the blood-brain barrier. Exosomes are very stable, can pass the blood-brain barrier and, therefore, reveal bright perspectives towards diagnosis and therapeutic treatments. A prerequisite for clinical applications is a standardised approach. Features necessary for a standardised diagnosis using exosomes are discussed. In therapeutic terms, exosomes represent a promising drug delivery system able to pass the blood-brain barrier. One option to overcome the disadvantages potentially associated with the use of endogenous exosomes is the design of artificial exosomes. The artificial exosomes with a clearly defined therapeutic active cargo and surface marker ensuring the specific targeting to the recipient cells is proposed as a promising approach.

  12. Digital Detection of Exosomes by Interferometric Imaging

    Science.gov (United States)

    Daaboul, George G.; Gagni, Paola; Benussi, Luisa; Bettotti, Paolo; Ciani, Miriam; Cretich, Marina; Freedman, David S.; Ghidoni, Roberta; Ozkumur, Ayca Yalcin; Piotto, Chiara; Prosperi, Davide; Santini, Benedetta; Ünlü, M. Selim; Chiari, Marcella

    2016-01-01

    Exosomes, which are membranous nanovesicles, are actively released by cells and have been attributed to roles in cell-cell communication, cancer metastasis, and early disease diagnostics. The small size (30–100 nm) along with low refractive index contrast of exosomes makes direct characterization and phenotypical classification very difficult. In this work we present a method based on Single Particle Interferometric Reflectance Imaging Sensor (SP-IRIS) that allows multiplexed phenotyping and digital counting of various populations of individual exosomes (>50 nm) captured on a microarray-based solid phase chip. We demonstrate these characterization concepts using purified exosomes from a HEK 293 cell culture. As a demonstration of clinical utility, we characterize exosomes directly from human cerebrospinal fluid (hCSF). Our interferometric imaging method could capture, from a very small hCSF volume (20 uL), nanoparticles that have a size compatible with exosomes, using antibodies directed against tetraspanins. With this unprecedented capability, we foresee revolutionary implications in the clinical field with improvements in diagnosis and stratification of patients affected by different disorders. PMID:27853258

  13. 大鼠T细胞源性exosome提取方法比较%The comparison among extraction methods of exosomes from T cells of rats

    Institute of Scientific and Technical Information of China (English)

    孙祯; 黄赤兵; 宋亚军; 陈益荣; 李传贵

    2013-01-01

    目的:采用3种方法从大鼠T细胞培养上清中提取纯化exosome ,以获取高质量的exosome。方法分别采用Exo-Quick Precipitation提取法、超滤密度梯度离心法、差速离心法提取 T 细胞培养上清中的exosome。利用透射电镜进行形态学观察,2,2-联喹啉-4,4-二甲酸二钠(BCA )法进行蛋白定量,十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE )分析蛋白表达的差异,Western blotting检测白细胞介素2(IL-2)的表达。结果3种方法均可提取出exosome;ExoQuick Precipitation 提取法、超滤密度梯度离心法所得样本浓度显著高于差速离心法所得样本浓度(P<0.05);SDS-PAGE结果显示3种方法所得样本具有蛋白表达强度的差异;Western blotting显示3种方法所得样本均表达IL-2。结论 ExoQuick Precipitation提取法、超滤密度梯度离心法可获得高纯度、无蛋白丢失的exosome样本。%Objective The purification methods of the exosomes derived form T cells were established in order to get high quan-tity exosomes .Methods Exosomes from T cells culture supernatants were purified by ExoQuick Precipitation ,ultrafiltration and sucrose gradient centrifugation ,differential ultracentrifugation ,and confirmed via using transmission electron microscopy .The pro-tein expression of the exosomes were analyzed by SDS-PAGE electrophoresis .Western blotting was used to test the expression of IL-2 .Results The protein concentration of the exosomes purified through ExoQuick Precipitation ,ultrafiltration and sucrose gradi-ent centrifugation were higher than through differential ultracentrifugation (P<0 .05) .SDS-PAGE displayed the difference among the exosome purified by three methods .Three kinds of exosomes all expressed IL-2 .Conclusion ExoQuick Precipitation ,ultrafiltra-tion and sucrose gradient centrifugation technique can obtain high purity and complete exosome sample .

  14. Bovine milk-derived exosomes for drug delivery.

    Science.gov (United States)

    Munagala, Radha; Aqil, Farrukh; Jeyabalan, Jeyaprakash; Gupta, Ramesh C

    2016-02-01

    Exosomes are biological nanovesicles that are involved in cell-cell communication via the functionally-active cargo (such as miRNA, mRNA, DNA and proteins). Because of their nanosize, exosomes are explored as nanodevices for the development of new therapeutic applications. However, bulk, safe and cost-effective production of exosomes is not available. Here, we show that bovine milk can serve as a scalable source of exosomes that can act as a carrier for chemotherapeutic/chemopreventive agents. Drug-loaded exosomes showed significantly higher efficacy compared to free drug in cell culture studies and against lung tumor xenografts in vivo. Moreover, tumor targeting ligands such as folate increased cancer-cell targeting of the exosomes resulting in enhanced tumor reduction. Milk exosomes exhibited cross-species tolerance with no adverse immune and inflammatory response. Thus, we show the versatility of milk exosomes with respect to the cargo it can carry and ability to achieve tumor targetability. This is the first report to identify a biocompatible and cost-effective means of exosomes to enhance oral bioavailability, improve efficacy and safety of drugs.

  15. Exosome secretion affects social motility in Trypanosoma brucei

    Science.gov (United States)

    Shaked, Hadassa; Arvatz, Gil; Tkacz, Itai Dov; Binder, Lior; Waldman Ben-Asher, Hiba; Okalang, Uthman; Chikne, Vaibhav; Cohen-Chalamish, Smadar; Michaeli, Shulamit

    2017-01-01

    Extracellular vesicles (EV) secreted by pathogens function in a variety of biological processes. Here, we demonstrate that in the protozoan parasite Trypanosoma brucei, exosome secretion is induced by stress that affects trans-splicing. Following perturbations in biogenesis of spliced leader RNA, which donates its spliced leader (SL) exon to all mRNAs, or after heat-shock, the SL RNA is exported to the cytoplasm and forms distinct granules, which are then secreted by exosomes. The exosomes are formed in multivesicular bodies (MVB) utilizing the endosomal sorting complexes required for transport (ESCRT), through a mechanism similar to microRNA secretion in mammalian cells. Silencing of the ESCRT factor, Vps36, compromised exosome secretion but not the secretion of vesicles derived from nanotubes. The exosomes enter recipient trypanosome cells. Time-lapse microscopy demonstrated that cells secreting exosomes or purified intact exosomes affect social motility (SoMo). This study demonstrates that exosomes are delivered to trypanosome cells and can change their migration. Exosomes are used to transmit stress signals for communication between parasites. PMID:28257521

  16. Exosomes: Origins and Therapeutic Potential for Neurodegenerative Disease

    Science.gov (United States)

    Sarko, Diana K.; McKinney, Cindy E.

    2017-01-01

    Exosomes, small lipid bilayer vesicles, are part of the transportable cell secretome that can be taken up by nearby recipient cells or can travel through the bloodstream to cells in distant organs. Selected cellular cytoplasm containing proteins, RNAs, and other macromolecules is packaged into secreted exosomes. This cargo has the potential to affect cellular function in either healthy or pathological ways. Exosomal content has been increasingly shown to assist in promoting pathways of neurodegeneration such as β-amyloid peptide (Aβ) accumulation forming amyloid plaques in the brains of patients with Alzheimer's disease, and pathological aggregates of proteins containing α-synuclein in Parkinson's disease transferred to the central nervous system via exosomes. In attempting to address such debilitating neuropathologies, one promising utility of exosomes lies in the development of methodology to use exosomes as natural delivery vehicles for therapeutics. Because exosomes are capable of penetrating the blood-brain barrier, they can be strategically engineered to carry drugs or other treatments, and possess a suitable half-life and stability for this purpose. Overall, analyses of the roles that exosomes play between diverse cellular sites will refine our understanding of how cells communicate. This mini-review introduces the origin and biogenesis of exosomes, their roles in neurodegenerative processes in the central nervous system, and their potential utility to deliver therapeutic drugs to cellular sites. PMID:28289371

  17. Exosomes: A Promising Factor Involved in Cancer Hypoxic Microenvironments.

    Science.gov (United States)

    Yang, Y; Yang, X; Yang, Y; Zhu, H; Chen, X; Zhang, H; Wang, F; Qin, Q; Cheng, H; Sun, X

    2015-01-01

    As a significant tumor feature, hypoxia can trigger cancer adaptive processes, induce malignant phenotype development, and promote drug resistance. Previous studies demonstrated that exosomes are critical during these procedures. Exosomes are small vesicles formed in vesicular bodies in the endosomal network. These small vesicles are mainly involved in the transport of bioactive molecules between cells. Exosomes are also involved in the mediation of some cellular communications depending on derived donor cells; thus, recipient cells undergo phenotypic changes. Furthermore, hypoxia can remarkably stimulate exosomal secretion; for instance, nucleic acids and proteins as transmission signals in exosomes in a tumor microenvironment are involved in various functions, such as inducing intratumoral heterogeneity, altering immunological responses, producing cancer-associated fibroblasts, and promoting angiogenesis and metastasis. Moreover, exosome contents resemble those of a donor cell; this finding indicates that exosomes may also be regarded as suitable biomarkers of hypoxia status. Therefore, exosomes can be used to facilitate diagnosis and prognosis with minimal invasive procedures. Further studies on exosomes in cancer may provide new therapeutic strategies.

  18. ExoCarta as a resource for exosomal research

    OpenAIRE

    Simpson, Richard J; Kalra, Hina; Mathivanan, Suresh

    2012-01-01

    Exosomes are a class of extracellular vesicles that are secreted by various cell types. Unlike other extracellular vesicles (ectosomes and apoptotic blebs), exosomes are of endocytic origin. The roles of exosomes in vaccine/ drug delivery, intercellular communication and as a possible source of disease biomarkers have sparked immense interest in them, resulting in a plethora of studies. Whilst multidimensional datasets are continuously generated, it is difficult to harness the true potential ...

  19. Interrogating Circulating Microsomes and Exosomes Using Metal Nanoparticles.

    Science.gov (United States)

    Zhou, Yi-Ge; Mohamadi, Reza M; Poudineh, Mahla; Kermanshah, Leyla; Ahmed, Sharif; Safaei, Tina Saberi; Stojcic, Jessica; Nam, Robert K; Sargent, Edward H; Kelley, Shana O

    2016-02-10

    A chip-based approach for electrochemical characterization and detection of microsomes and exosomes based on direct electro-oxidation of metal nanoparticles (MNPs) that specifically recognize surface markers of these vesicles is reported. It is found that exosomes and microsomes derived from prostate cancer cells can be identified by their surface proteins EpCAM and PSMA, suggesting the potential of exosomes and microsomes for use as diagnostic biomarkers.

  20. Exosomes derived from miR-140-5p-overexpressing human synovial mesenchymal stem cells enhance cartilage tissue regeneration and prevent osteoarthritis of the knee in a rat model

    Science.gov (United States)

    Tao, Shi-Cong; Yuan, Ting; Zhang, Yue-Lei; Yin, Wen-Jing; Guo, Shang-Chun; Zhang, Chang-Qing

    2017-01-01

    OBJECTIVES: Osteoarthritis (OA) is the most common joint disease throughout the world. Exosomes derived from miR-140-5p-overexpressing synovial mesenchymal stem cells (SMSC-140s) may be effective in treating OA. We hypothesized that exosomes derived from SMSC-140 (SMSC-140-Exos) would enhance the proliferation and migration abilities of articular chondrocytes (ACs) without harming extracellular matrix (ECM) secretion. METHODS: SMSCs were transfected with or without miR-140-5p. Exosomes derived from SMSCs or SMSC-140s (SMSC-Exos or SMSC-140-Exos) were isolated and identified. Proliferation, migration and ECM secretion were measured in vitro and compared between groups. The mechanism involving alternative Wnt signalling and activation of Yes-associated protein (YAP) was investigated using lentivirus, oligonucleotides or chemical drugs. The preventative effect of exosomes in vivo was measured using Safranin-O and Fast green staining and immunohistochemical staining. RESULTS: Wnt5a and Wnt5b carried by exosomes activated YAP via the alternative Wnt signalling pathway and enhanced proliferation and migration of chondrocytes with the side-effect of significantly decreasing ECM secretion. Highly-expressed miR-140-5p blocked this side-effect via RalA. SMSC-140-Exos enhanced the proliferation and migration of ACs without damaging ECM secretion in vitro, while in vivo, SMSC-140-Exos successfully prevented OA in a rat model. CONCLUSIONS: These findings highlight the promising potential of SMSC-140-Exos in preventing OA. We first found a potential source of exosomes and studied their merits and shortcomings. Based on our understanding of the molecular mechanism, we overcame the shortcomings by modifying the exosomes. Such exosomes derived from modified cells hold potential as future therapeutic strategies. PMID:28042326

  1. The odyssey of Hsp60 from tumor cells to other destinations includes plasma membrane-associated stages and Golgi and exosomal protein-trafficking modalities.

    Directory of Open Access Journals (Sweden)

    Claudia Campanella

    Full Text Available BACKGROUND: In a previous work we showed for the first time that human tumor cells secrete Hsp60 via exosomes, which are considered immunologically active microvesicles involved in tumor progression. This finding raised questions concerning the route followed by Hsp60 to reach the exosomes, its location in them, and whether Hsp60 can be secreted also via other mechanisms, e.g., by the Golgi. We addressed these issues in the work presented here. PRINCIPAL FINDINGS: We found that Hsp60 localizes in the tumor cell plasma membrane, is associated with lipid rafts, and ends up in the exosomal membrane. We also found evidence that Hsp60 localizes in the Golgi apparatus and its secretion is prevented by an inhibitor of this organelle. CONCLUSIONS/SIGNIFICANCE: We propose a multistage process for the translocation of Hsp60 from the inside to the outside of the cell that includes a combination of protein traffic pathways and, ultimately, presence of the chaperonin in the circulating blood. The new information presented should help in designing future strategies for research and for developing diagnostic-monitoring means useful in clinical oncology.

  2. Exosomes and their roles in immune regulation and cancer.

    Science.gov (United States)

    Greening, David W; Gopal, Shashi K; Xu, Rong; Simpson, Richard J; Chen, Weisan

    2015-04-01

    Exosomes, a subset of extracellular vesicles (EVs), function as a mode of intercellular communication and molecular transfer. Exosomes facilitate the direct extracellular transfer of proteins, lipids, and miRNA/mRNA/DNAs between cells in vitro and in vivo. The immunological activities of exosomes affect immunoregulation mechanisms including modulating antigen presentation, immune activation, immune suppression, immune surveillance, and intercellular communication. Besides immune cells, cancer cells secrete immunologically active exosomes that influence both physiological and pathological processes. The observation that exosomes isolated from immune cells such as dendritic cells (DCs) modulate the immune response has enforced the way these membranous vesicles are being considered as potential immunotherapeutic reagents. Indeed, tumour- and immune cell-derived exosomes have been shown to carry tumour antigens and promote immunity, leading to eradication of established tumours by CD8(+) T cells and CD4(+) T cells, as well as directly suppressing tumour growth and resistance to malignant tumour development. Further understanding of these areas of exosome biology, and especially of molecular mechanisms involved in immune cell targeting, interaction and manipulation, is likely to provide significant insights into immunorecognition and therapeutic intervention. Here, we review the emerging roles of exosomes in immune regulation and the therapeutic potential in cancer.

  3. Exosomes as mediators of intercellular communication: clinical implications.

    Science.gov (United States)

    Nazimek, Katarzyna; Bryniarski, Krzysztof; Santocki, Michał; Ptak, Włodzimierz

    2015-01-01

    Cells of multicellular organisms exchange informative signals by diverse mechanisms. Recent findings uncovered the special role of extracellular vesicles, especially exosomes, in intercellular communication. Exosomes, present in all tested human bodily fluids, carry various functional compounds including proteins, lipids, and diverse RNA molecules. The composition of exosome cargo in vivo is likely formed by a regulated selection of specific components and can express the current status of the exosome-secreting cell. Therefore, particular emphasis is now placed on the extremely high potential of exosomes as essentially noninvasive prognostic and diagnostic biomarkers, but also as therapeutic nanocarriers, especially after the discovery that their cargo as well as cell-targeting specificity could be shaped in vitro. In addition, targeting the exosomes mediating pathological intercellular communication may also express high therapeutic potential. Hence, numerous studies are conducted to explore the profile and function of exosomes and their cargo in health and disease and to shape their properties to facilitate their clinical application. The present review summarizes the current knowledge on the role of exosomes in different physiological and pathological mechanisms of intercellular communication with a particular focus on the use of exosomes in the diagnosis and treatment of various inflammatory, cardiovascular, metabolic, and neurodegenerative disorders as well as malignant neoplasms.

  4. The non-targeted effects of radiation are perpetuated by exosomes.

    Science.gov (United States)

    Al-Mayah, Ammar; Bright, Scott; Chapman, Kim; Irons, Sarah; Luo, Ping; Carter, David; Goodwin, Edwin; Kadhim, Munira

    2015-02-01

    Exosomes contain cargo material from endosomes, cytosol, plasma membrane and microRNA molecules, they are released by a number of non-cancer and cancer cells into both the extracellular microenvironment and body fluids such as blood plasma. Recently we demonstrated radiation-induced non-targeted effects [NTE: genomic instability (GI) and bystander effects (BE)] are partially mediated by exosomes, particularly the RNA content. However the mechanistic role of exosomes in NTE is yet to be fully understood. The present study used MCF7 cells to characterise the longevity of exosome-induced activity in the progeny of irradiated and unirradiated bystander cells. Exosomes extracted from conditioned media of irradiated and bystander progeny were added to unirradiated cells. Analysis was carried out at 1 and 20/24 population doublings following medium/exosome transfer for DNA/chromosomal damage. Results confirmed exosomes play a significant role in mediating NTE of ionising radiation (IR). This effect was remarkably persistent, observed >20 doublings post-irradiation in the progeny of bystander cells. Additionally, cell progeny undergoing a BE were themselves capable of inducing BE in other cells via exosomes they released. Furthermore we investigated the role of exosome cargo. Culture media from cells exposed to 2 Gy X-rays was subjected to ultracentrifugation and four inoculants prepared, (a) supernatants with exosomes removed, and pellets with (b) exosome proteins denatured, (c) RNA degraded, and (d) a combination of protein-RNA inactivation. These were added to separate populations of unirradiated cells. The BE was partially inhibited when either exosome protein or exosome RNA were inactivated separately, whilst combined RNA-protein inhibition significantly reduced or eliminated the BE. These results demonstrate that exosomes are associated with long-lived signalling of the NTE of IR. Both RNA and protein molecules of exosomes work in a synergistic manner to initiate NTE

  5. Functional Roles and Therapeutic Applications of Exosomes in Hepatocellular Carcinoma

    Directory of Open Access Journals (Sweden)

    Laura Santangelo

    2017-01-01

    Full Text Available Exosomes are important in intercellular communication. They assure the horizontal transfer of specific functional contents (i.e., proteins, lipids, RNA molecules, and circulating DNA from donor to recipient cells. Notably, tumor-derived exosomes (TDEs appear to be an important vehicle of specific signals in cancer, impacting on tumor growth and metastasis. Recent researches point to the characterization of exosomes in Hepatocellular Carcinoma (HCC, the major adult liver malignancy. In this review, we summarize current findings on HCC exosomes, focusing on the identification of noncoding RNAs as exosome-enriched functional regulators and new potential biomarkers. The great potential of exosomes in future HCC diagnostic and therapeutic approaches is underlined.

  6. Imaging and Intracellular Tracking of Cancer-Derived Exosomes Using Single-Molecule Localization-Based Super-Resolution Microscope.

    Science.gov (United States)

    Chen, Chen; Zong, Shenfei; Wang, Zhuyuan; Lu, Ju; Zhu, Dan; Zhang, Yizhi; Cui, Yiping

    2016-10-05

    Exosomes are small membrane vesicles secreted by cells and enriched with plenty of proteins. Considering their significant roles in different physical activities and potential value for diagnostic drug delivery, researchers have put great efforts in in vitro tracking and content analysis of exosomes. Recently, the emergence of different kinds of super-resolution microscopy provides powerful tools for exosome study. Here, we demonstrate the application of single-molecule localization based super-resolution imaging technique in the imaging and tracking of cancer-derived exosomes. In the experiment, first, cancer-derived exosomes are extracted from the culture media of tumor cells. Then the exosome membrane receptors are labeled with photoswitchable probes, which allow super-resolution imaging of these membrane receptors via photoactivated localization microscopy (PALM) or stochastic optical reconstruction microscopy (STORM). By using human breast cancer cell-derived exosomes, we demonstrated simultaneous dual-color PALM/STORM imaging of two kinds of membrane receptors on the exosome membrane. Moreover, the successful labeling and imaging of exosomes make it possible to observe the interaction between cancer-derived exosomes and normal cells. Meanwhile, we realized the colocalization of cancer-derived exosomes and lysosomes in recipient cells with PALM/STORM imaging. Since exosomes play a vital role in intercellular communications, we anticipate that the presented PALM/STORM-based imaging and tracking of exosomes holds a great potential in the investigation of the mechanism of exosome-mediated cancer metastasis.

  7. Immunoregulatory Role of Dendritic Cell-derived Exosomes%树突状细胞来源的Exosomes的免疫调节作用

    Institute of Scientific and Technical Information of China (English)

    刘袁媛; 范华骅; 陈亮

    2007-01-01

    Exosomes是多种细胞经晚期内体形成的一种膜性小囊泡,最初认为其功能仅为降解内吞物质,但研究发现exosomes的特异功能与其来源细胞相关,尤其是抗原提呈细胞(APCs)--树突状细胞来源的exosomes(dendritic cell-derived exosomes,DEXs)集MHC-I/MHC-Ⅱ、共刺激分子、黏附分子、热休克蛋白于一身,在体内外免疫调节中起非常重要的作用.现对DEXs诱导抗肿瘤免疫应答和诱导免疫耐受两方面的功能及可能的免疫调节机制进行综述.

  8. Smart blood cell and microvesicle-based Trojan horse drug delivery: Merging expertise in blood transfusion and biomedical engineering in the field of nanomedicine.

    Science.gov (United States)

    Wu, Yu-Wen; Goubran, Hadi; Seghatchian, Jerard; Burnouf, Thierry

    2016-04-01

    Therapeutic and diagnostic applications of nanomedicine are playing increasingly important roles in human health. Various types of synthetic nanoparticles, including liposomes, micelles, and other nanotherapeutic platforms and conjugates, are being engineered to encapsulate or carry drugs for treating diseases such as cancer, cardiovascular disorders, neurodegeneration, and inflammations. Nanocarriers are designed to increase the half-life of drugs, decrease their toxicity and, ideally, target pathological sites. Developing smart carriers with the capacity to deliver drugs specifically to the microenvironment of diseased cells with minimum systemic toxicity is the goal. Blood cells, and potentially also the liposome-like micro- and nano-vesicles they generate, may be regarded as ideally suited to perform such specific targeting with minimum immunogenic risks. Blood cell membranes are "decorated" with complex physiological receptors capable of targeting and communicating with other cells and tissues and delivering their content to the surrounding pathological microenvironment. Blood cells, such as erythrocytes, have been developed as permeable carriers to release drugs to diseased tissues or act as biofactory allowing enzymatic degradation of a pathological substrate. Interestingly, attempts are also being made to improve the targeting capacity of synthetic nanoparticles by "decorating" their surface with blood cell membrane receptor-like biochemical structures. Research is needed to further explore the benefits that blood cell-derived microvesicles, as a Trojan horse delivery systems, can bring to the arsenal of therapeutic micro- and nanotechnologies. This short review focuses on the therapeutic roles that red blood cells and platelets can play as smart drug-delivery systems, and highlights the benefits that blood transfusion expertise can bring to this exciting and novel biomedical engineering field.

  9. Natural CD8{sup +}25{sup +} regulatory T cell-secreted exosomes capable of suppressing cytotoxic T lymphocyte-mediated immunity against B16 melanoma

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Yufeng; Zhang, Xueshu; Zhao, Tuo; Li, Wei; Xiang, Jim, E-mail: jim.xiang@saskcancer.ca

    2013-08-16

    Highlights: •CD8{sup +}25{sup +} regulatory T cells secrete tolerogenic exosomes. •CD8{sup +}25{sup +} regulatory T cell-derived exosomes exhibit immunosuppressive effect. •CD8{sup +}25{sup +} regulatory T cell-derived exosomes inhibit antitumor immunity. -- Abstract: Natural CD4{sup +}25{sup +} and CD8{sup +}25{sup +} regulatory T (Tr) cells have been shown to inhibit autoimmune diseases. Immune cells secrete exosomes (EXOs), which are crucial for immune regulation. However, immunomodulatory effect of natural Tr cell-secreted EXOs is unknown. In this study, we purified natural CD8{sup +}25{sup +} Tr cells from C57BL/6 mouse naive CD8{sup +} T cells, and in vitro amplified them with CD3/CD28 beads. EXOs (EXO{sub Tr}) were purified from Tr cell’s culture supernatants by differential ultracentrifugation and analyzed by electron microscopy, Western blot and flow cytometry. Our data showed that EXO{sub Tr} had a “saucer” or round shape with 50–100 nm in diameter, contained EXO-associated markers LAMP-1 and CD9, and expressed natural Tr cell markers CD25 and GITR. To assess immunomodulatory effect, we i.v. immunized C57BL/6 mice with ovalbumin (OVA)-pulsed DCs (DC{sub OVA}) plus Tr cells or EXO{sub Tr}, and then assessed OVA-specific CD8{sup +} T cell responses using PE-H-2K{sup b}/OVA tetramer and FITC-anti-CD8 antibody staining by flow cytometry and antitumor immunity in immunized mice with challenge of OVA-expressing BL6–10{sub OVA} melanoma cells. We demonstrated that DC{sub OVA}-stimulated CD8{sup +} T cell responses and protective antitumor immunity significantly dropped from 2.52% to 1.08% and 1.81% (p < 0.05), and from 8/8 to 2/8 and 5/8 mice DC{sub OVA} (p < 0.05) in immunized mice with co-injection of Tr cells and EXO{sub Tr}, respectively. Our results indicate that natural CD8{sup +}25{sup +} Tr cell-released EXOs, alike CD8{sup +}25{sup +} Tr cells, can inhibit CD8{sup +} T cell responses and antitumor immunity. Therefore, EXOs derived from

  10. Exosomes from hypoxic endothelial cells have increased collagen crosslinking activity through up-regulation of lysyl oxidase-like 2

    NARCIS (Netherlands)

    de Jong, Olivier G.; van Balkom, Bas W M; Gremmels, Hendrik; Verhaar, Marianne C.

    2016-01-01

    Exosomes are important mediators of intercellular communication. Additionally, they contain a variety of components capable of interacting with the extracellular matrix (ECM), including integrins, matrix metalloproteinases and members of the immunoglobin superfamily. Despite these observations, rese

  11. Exosome release from infected dendritic cells: a clue for a fast spread of prions in the periphery?

    Science.gov (United States)

    Klöhn, Peter-Christian; Castro-Seoane, Rocio; Collinge, John

    2013-11-01

    Prion diseases are incurable transmissible neurological disorders. In many natural and experimental prion diseases, infectious prions can be detected in the lymphoreticular system (LRS) long before they reach the brain where they cause a fatal rapidly progressive degeneration. Although major cell types that contribute to prion accumulation have been identified, the mode of prion dissemination in the LRS remains elusive. Recent evidence of a remarkably fast splenic prion accumulation after peripheral infection of mice, resulting in high prion titers in dendritic cells (DCs) and a release of prions from infected DCs via exosomes suggest that intercellular dissemination may contribute to rapid prion colonization in the LRS. A vast body of evidence from retroviral infections shows that DCs and other antigen-presenting cells (APCs) share viral antigens by intercellular transfer to warrant immunity against viruses if APCs remain uninfected. Evolved to adapt the immune response to evading pathogens, these pathways may constitute a portal for unimpeded prion dissemination owing to the tolerance of the immune system against host-encoded prion protein. In this review we summarize current paradigms for antigen-sharing pathways which may be relevant to better understand dissemination of rogue neurotoxic proteins.

  12. Exosomes derived from dendritic cells improve cardiac function via activation of CD4(+) T lymphocytes after myocardial infarction.

    Science.gov (United States)

    Liu, Haibo; Gao, Wei; Yuan, Jie; Wu, Chaoneng; Yao, Kang; Zhang, Li; Ma, Leilei; Zhu, Jianbing; Zou, Yunzeng; Ge, Junbo

    2016-02-01

    CD4(+) T cell activation plays a key role in facilitating wound healing after myocardial infarction (MI). Exosomes (EXs) secreted from dendritic cells (DCs) can activate T cells in tumor models; however, whether DEXs (DC-EXs) can mediate CD4(+) T cell activation and improve wound healing post-MI remains unknown. This study sought to determine whether DEXs mediate CD4(+) T cell activation and improve cardiac function post-MI in mice. We used supernatants of hypoxic primary or necrotic HL-1 cardiomyocytes to simulate the post-MI cardiomyocyte microenvironment in vitro. Cultured bone marrow-derived DCs (BMDCs) from mice were stimulated with the supernatants of normal (Control group), hypoxic primary or necrotic HL-1 cardiomyocytes (MI group); a subset of BMDCs remained unstimulated (Negative group). DEXs were then isolated from the BMDC supernatants and either incubated with CD4(+) T cells or injected into mice via the tail vein. In this study, we found that the supernatants of both hypoxic primary and necrotic HL-1 cardiomyocytes upregulate DC maturation markers. After the injection of DEXs, a greater number of MI-DEXs are recruited by the mouse spleen and with greater rapidity than control- or negative-DEXs. Confocal imaging and flow cytometry revealed that MI-DEXs exhibited higher uptake by splenic CD4(+) T cells than the control- and negative-DEXs, and this increase was correlated with significantly greater increases in the expression of chemokines and the inflammatory cytokines IFN-γ and TNF by the CD4(+) T cells in vitro and in vivo. In addition, the injection of MI-DEXs improved cardiac function in mice post-MI. These results suggest that DEXs could mediate the activation of CD4(+) T cells through an endocrine mechanism and improve cardiac function post-MI. Our findings provide the basis for a novel strategy for the treatment of MI through the systemic delivery of DEXs.

  13. 'Trojan Horse" Teaching

    Directory of Open Access Journals (Sweden)

    Alexander N. Poddiakov

    2009-01-01

    Full Text Available An advanced strategic behavior, which we term, “Trojan horse” teaching (ThT, is described. In this type of counteractive behavior, a “teacher”, ostensibly helping his or her rival to learn something, really teaches the rival useless or disadvantageous things. This interaction is an object of interdisciplinary research related to the theory of human capital, the theory of agency, knowledge management, the theory of conflict, and to social and educational psychology. Examples of ThT in real life, and results of experiential studies, including the administration of a survey concerning people’s beliefs about teaching “with evil intent”, and a set of experiments with participation of adults and children, have been described. Possible directions of artificial intelligence systems development related to ThT are described. General relations between: (a counteraction to learning, and (b development in spite of the counteraction are discussed.

  14. Exosomes Secreted from Human-Induced Pluripotent Stem Cell-Derived Mesenchymal Stem Cells Prevent Osteonecrosis of the Femoral Head by Promoting Angiogenesis

    Science.gov (United States)

    Liu, Xiaolin; Li, Qing; Niu, Xin; Hu, Bin; Chen, Shengbao; Song, Wenqi; Ding, Jian; Zhang, Changqing; Wang, Yang

    2017-01-01

    Background: Local ischemia is the main pathological performance in osteonecrosis of the femoral head (ONFH). There is currently no effective therapy to promote angiogenesis in the femoral head. Recent studies revealed that exosomes secreted by induced pluripotent stem cell-derived mesenchymal stem cells (iPS-MSC-Exos) have great therapeutic potential in ischemic tissues, but whether they could promote angiogenesis in ONFH has not been reported, and little is known regarding the underlying mechanism. Methods: iPS-MSC-Exos were intravenously injected to a steroid-induced rat osteonecrosis model. Samples of the femoral head were obtained 3 weeks after all the injections. The effects were assessed by measuring local angiogenesis and bone loss through histological and immunohistochemical (IHC) staining, micro-CT and three-dimensional microangiography. The effects of exosomes on endothelial cells were studied through evaluations of proliferation, migration and tube-forming analyses. The expression levels of angiogenic related PI3K/Akt signaling pathway of endothelial cells were evaluated following stimulation of iPS-MSC-Exos. The promoting effects of exosomes were re-evaluated following blockade of PI3K/Akt. Results: The in vivo study revealed that administration of iPS-MSC-Exos significantly prevented bone loss, and increased microvessel density in the femoral head compared with control group. We found that iPS-MSC-Exos significantly enhanced the proliferation, migration and tube-forming capacities of endothelial cells in vitro. iPS-MSC-Exos could activate PI3K/Akt signaling pathway in endothelial cells. Moreover, the promoting effects of iPS-MSC-Exos were abolished after blockade of PI3K/Akt on endothelial cells. Conclusions: Our findings suggest that transplantation of iPS-MSC-Exos exerts a preventative effect on ONFH by promoting local angiogenesis and preventing bone loss. The promoting effect might be attributed to activation of the PI3K/Akt signaling pathway on

  15. Targeted therapeutic delivery using engineered exosomes and its applications in cardiovascular diseases.

    Science.gov (United States)

    Xitong, Dang; Xiaorong, Zeng

    2016-01-10

    Exosomes are 30-120 nm membrane bound vesicles secreted naturally by almost all cells and exist in all body fluids. Accumulating evidence has shown that exosomes contain proteins, lipids, DNA, mRNA, miRNA, and lncRNA that can be transferred from producer cells to recipient cells, facilitating cell-cell communication. As the natural carrier of these signal molecules, exosomes possess many other properties such as stability, biocompatibility, biological barrier permeability, low toxicity, and low immunogenicity, which make them an attractive vehicle for therapeutic delivery. How exosomes target recipient cells in vivo remains largely unknown, however, exosomes are selectively enriched in some transmembrane proteins that can be genetically engineered to display ligands/homing peptides on their surface, which confers exosome targeting capability to cells bearing cognate receptors. With the discovery of many peptides homing to diseased tissues or organs through phage display and in vivo biopanning technologies, there is ample opportunity to explore the potential use of exosome for targeted gene therapy. Here, we briefly review exosome biogenesis, mechanisms of exosome-mediated cell–cell communication, and exosome isolation and purification methods, and specifically focus on the emerging exosome targeting technologies.

  16. Pathologic function and therapeutic potential of exosomes in cardiovascular disease.

    Science.gov (United States)

    Ailawadi, Shaina; Wang, Xiaohong; Gu, Haitao; Fan, Guo-Chang

    2015-01-01

    The heart is a very complex conglomeration of organized interactions between various different cell types that all aid in facilitating myocardial function through contractility, sufficient perfusion, and cell-to-cell reception. In order to make sure that all features of the heart work effectively, it is imperative to have a well-controlled communication system among the different types of cells. One of the most important ways that the heart regulates itself is by the use of extracellular vesicles, more specifically, exosomes. Exosomes are types of nano-vesicles, naturally released from living cells. They are believed to play a critical role in intercellular communication through the means of certain mechanisms including direct cell-to-cell contact, long-range signals as well as electrical and extracellular chemical molecules. Exosomes contain many unique features like surface proteins/receptors, lipids, mRNAs, microRNAs, transcription factors and other proteins. Recent studies indicate that the exosomal contents are highly regulated by various stress and disease conditions, in turn reflective of the parent cell status. At present, exosomes are well appreciated to be involved in the process of tumor and infection disease. However, the research on cardiac exosomes is just emerging. In this review, we summarize recent findings on the pathologic effects of exosomes on cardiac remodeling under stress and disease conditions, including cardiac hypertrophy, peripartum cardiomyopathy, diabetic cardiomyopathy and sepsis-induced cardiovascular dysfunction. In addition, the cardio-protective effects of stress-preconditioned exosomes and stem cell-derived exosomes are also summarized. Finally, we discuss how to epigenetically reprogram exosome contents in host cells which makes them beneficial for the heart.

  17. αB crystallin is apically secreted within exosomes by polarized human retinal pigment epithelium and provides neuroprotection to adjacent cells.

    Directory of Open Access Journals (Sweden)

    Parameswaran G Sreekumar

    Full Text Available αB crystallin is a chaperone protein with anti-apoptotic and anti-inflammatory functions and has been identified as a biomarker in age-related macular degeneration. The purpose of this study was to determine whether αB crystallin is secreted from retinal pigment epithelial (RPE cells, the mechanism of this secretory pathway and to determine whether extracellular αB crystallin can be taken up by adjacent retinal cells and provide protection from oxidant stress. We used human RPE cells to establish that αB crystallin is secreted by a non-classical pathway that involves exosomes. Evidence for the release of exosomes by RPE and localization of αB crystallin within the exosomes was achieved by immunoblot, immunofluorescence, and electron microscopic analyses. Inhibition of lipid rafts or exosomes significantly reduced αB crystallin secretion, while inhibitors of classic secretory pathways had no effect. In highly polarized RPE monolayers, αB crystallin was selectively secreted towards the apical, photoreceptor-facing side. In support, confocal microscopy established that αB crystallin was localized predominantly in the apical compartment of RPE monolayers, where it co-localized in part with exosomal marker CD63. Severe oxidative stress resulted in barrier breakdown and release of αB crystallin to the basolateral side. In normal mouse retinal sections, αB crystallin was identified in the interphotoreceptor matrix. An increased uptake of exogenous αB crystallin and protection from apoptosis by inhibition of caspase 3 and PARP activation were observed in stressed RPE cultures. αB Crystallin was taken up by photoreceptors in mouse retinal explants exposed to oxidative stress. These results demonstrate an important role for αB crystallin in maintaining and facilitating a neuroprotective outer retinal environment and may also explain the accumulation of αB crystallin in extracellular sub-RPE deposits in the stressed microenvironment in age

  18. The Multiple Roles of Exosomes in Metastasis.

    Science.gov (United States)

    Weidle, Ulrich H; Birzele, Fabian; Kollmorgen, Gwen; Rüger, Rüdiger

    2017-01-02

    Exosomes are important contributors to cell-cell communication and their role as diagnostic markers for cancer and the pathogenesis for cancer is under intensive investigation. Here, we focus on their role in metastasis-related processes. We discuss their impact regarding promotion of invasion and migration of tumor cells, conditioning of lymph nodes, generation of premetastatic niches and organotropism of metastasis. Furthermore, we highlight interactions of exosomes with bone marrow and stromal components such as fibroblasts, endothelial cells, myeloid- and other immune-related cells in the context of metastases. For all processes as described above, we outline molecular and cellular components for therapeutic intervention with metastatic processes.

  19. Detection of Exosomal Biomarker by Electric Field-induced Release and Measurement (EFIRM)

    Science.gov (United States)

    Tu, Michael; Wei, Fang; Yang, Jieping; Wong, David

    2015-01-01

    Exosomes are microvesicular structures that play a mediating role in intercellular communication. It is of interest to study the internal cargo of exosomes to determine if they carry disease discriminatory biomarkers. For performing exosomal analysis, it is necessary to develop a method for extracting and analyzing exosomes from target biofluids without damaging the internal content. Electric field-induced release and measurement (EFIRM) is a method for specifically extracting exosomes from biofluids, unloading their cargo, and testing their internal RNA/protein content. Using an anti-human CD63 specific antibody magnetic microparticle, exosomes are first precipitated from biofluids. Following extraction, low-voltage electric cyclic square waves (CSW) are applied to disrupt the vesicular membrane and cause cargo unloading. The content of the exosome is hybridized to DNA primers or antibodies immobilized on an electrode surface for quantification of molecular content. The EFIRM method is advantageous for extraction of exosomes and unloading cargo for analysis without lysis buffer. This method is capable of performing specific detection of both RNA and protein biomarker targets in the exosome. EFIRM extracts exosomes specifically based on their surface markers as opposed to size-based techniques. Transmission electron microscopy (TEM) and assay demonstrate the functionality of the method for exosome capture and analysis. The EFIRM method was applied to exosomal analysis of 9 mice injected with human lung cancer H640 cells (a cell line transfected to express the exosome marker human CD63-GFP) in order to test their exosome profile against 11 mice receiving saline controls. Elevated levels of exosomal biomarkers (reference gene GAPDH and protein surface marker human CD63-GFP) were found for the H640 injected mice in both serum and saliva samples. Furthermore, saliva and serum samples were demonstrated to have linearity (R = 0.79). These results are suggestive for the

  20. Exosome Biogenesis, Regulation, and Function in Viral Infection.

    Science.gov (United States)

    Alenquer, Marta; Amorim, Maria João

    2015-09-17

    Exosomes are extracellular vesicles released upon fusion of multivesicular bodies(MVBs) with the cellular plasma membrane. They originate as intraluminal vesicles (ILVs) during the process of MVB formation. Exosomes were shown to contain selectively sorted functional proteins, lipids, and RNAs, mediating cell-to-cell communications and hence playing a role in the physiology of the healthy and diseased organism. Challenges in the field include the identification of mechanisms sustaining packaging of membrane-bound and soluble material to these vesicles and the understanding of the underlying processes directing MVBs for degradation or fusion with the plasma membrane. The investigation into the formation and roles of exosomes in viral infection is in its early years. Although still controversial, exosomes can, in principle, incorporate any functional factor, provided they have an appropriate sorting signal, and thus are prone to viral exploitation.This review initially focuses on the composition and biogenesis of exosomes. It then explores the regulatory mechanisms underlying their biogenesis. Exosomes are part of the endocytic system,which is tightly regulated and able to respond to several stimuli that lead to alterations in the composition of its sub-compartments. We discuss the current knowledge of how these changes affect exosomal release. We then summarize how different viruses exploit specific proteins of endocytic sub-compartments and speculate that it could interfere with exosome function, although no direct link between viral usage of the endocytic system and exosome release has yet been reported. Many recent reports have ascribed functions to exosomes released from cells infected with a variety of animal viruses, including viral spread, host immunity, and manipulation of the microenvironment, which are discussed. Given the ever-growing roles and importance of exosomes in viral infections, understanding what regulates their composition and levels, and

  1. Exosomes Isolated from Ascites of T-Cell Lymphoma-Bearing Mice Expressing Surface CD24 and HSP-90 Induce a Tumor-Specific Immune Response

    Science.gov (United States)

    Menay, Florencia; Herschlik, Leticia; De Toro, Julieta; Cocozza, Federico; Tsacalian, Rodrigo; Gravisaco, María José; Di Sciullo, María Paula; Vendrell, Alejandrina; Waldner, Claudia I.; Mongini, Claudia

    2017-01-01

    Extracellular vesicles (EVs), including endosome-derived nanovesicles (exosomes), are involved in cell–cell communication. Through transfer of their molecular contents, extracellular nanovesicles can alter the function of recipient cells. Due to these characteristics, EVs have shown potential as a new alternative for cancer immunotherapy. Tumor exosomes isolated from malignant ascites can activate dendritic cells, thereby priming the immune system to recognize and kill cancer cells. However, a suppressive role on tumor immune response has also been reported, suggesting that the neoplastic stage of carcinogenesis and the microenvironment where tumor cells grow may influence the amount of EVs released by the cell. This neoplastic stage and microenvironment may also impact EVs’ components such as proteins and miRNA, determining their biological behavior. Most T-cell lymphomas have an aggressive clinical course and poor prognosis. Consequently, complementary alternative therapies are needed to improve the survival rates achieved with conventional treatments. In this work, we have characterized EVs isolated from ascites of mice bearing a very aggressive murine T-cell lymphoma and have studied their immunogenic properties. Small EVs were isolated by differential centrifugation, ultrafiltration, and ultracentrifugation at 100,000 × g on a sucrose cushion. The EVs were defined as exosomes by their morphology and size analyzed by electron microscopy, their floating density on a sucrose gradient, as well as their expression of endosome marker proteins ALIX, TSG-101; the tetraspanins CD63, CD9, and CD81. In addition, they contain tumor antigens, the marker for malignancy CD24, the heat shock protein HSP-70, and an unusual surface expression of HSP-90 was demonstrated. The administration of EVs isolated from ascites (EVs A) into naïve-syngeneic mice induced both humoral and cellular immune responses that allowed the rejection of subsequent tumor challenges. However

  2. The roles of tumor-derived exosomes in cancer pathogenesis.

    Science.gov (United States)

    Yang, Chenjie; Robbins, Paul D

    2011-01-01

    Exosomes are endosome-derived, 30-100 nm small membrane vesicles released by most cell types including tumor cells. They are enriched in a selective repertoire of proteins and nucleic acids from parental cells and are thought to be actively involved in conferring intercellular signals. Tumor-derived exosomes have been viewed as a source of tumor antigens that can be used to induce antitumor immune responses. However, tumor-derived exosomes also have been found to possess immunosuppressive properties and are able to facilitate tumor growth, metastasis, and the development of drug resistance. These different effects of tumor-derived exosomes contribute to the pathogenesis of cancer. This review will discuss the roles of tumor-derived exosomes in cancer pathogenesis, therapy, and diagnostics.

  3. The Roles of Tumor-Derived Exosomes in Cancer Pathogenesis

    Directory of Open Access Journals (Sweden)

    Chenjie Yang

    2011-01-01

    Full Text Available Exosomes are endosome-derived, 30–100 nm small membrane vesicles released by most cell types including tumor cells. They are enriched in a selective repertoire of proteins and nucleic acids from parental cells and are thought to be actively involved in conferring intercellular signals. Tumor-derived exosomes have been viewed as a source of tumor antigens that can be used to induce antitumor immune responses. However, tumor-derived exosomes also have been found to possess immunosuppressive properties and are able to facilitate tumor growth, metastasis, and the development of drug resistance. These different effects of tumor-derived exosomes contribute to the pathogenesis of cancer. This review will discuss the roles of tumor-derived exosomes in cancer pathogenesis, therapy, and diagnostics.

  4. The Trojans' Odyssey space mission

    Science.gov (United States)

    Lamy, P.; Vernazza, P.; Groussin, O.; Poncy, J.; Martinot, V.; Hinglais, E.; Bell, J.; Cruikshank, D.; Helbert, J.; Marzari, F.; Morbidelli, A.; Rosenblatt, P.

    2011-10-01

    In our present understanding of the Solar System, small bodies (asteroids, Jupiter Trojans, comets and TNOs) are the most direct remnants of the original building blocks that formed the planets. Jupiter Trojan and Hilda asteroids are small primitive bodies located beyond the "snow line", around respectively the L4 and L5 Lagrange points of Jupiter at 5.2 AU (Trojans) and in the 2:3 mean-motion resonance with Jupiter near 3.9 AU (Hildas). They are at the crux of several outstanding and still conflicting issues regarding the formation and evolution of the Solar System. They hold the potential to unlock the answers to fundamental questions about planetary migration, the late heavy bombardment, the formation of the Jovian system, the origin and evolution of trans-neptunian objects, and the delivery of water and organics to the inner planets. The proposed Trojans' Odyssey mission is envisioned as a reconnaissance, multiple flyby mission aimed at visiting several objects, typically five Trojans and one Hilda. It will attempt exploring both large and small objects and sampling those with any known differences in photometric properties. The orbital strategy consists in a direct trajectory to one of the Trojan swarms. By carefully choosing the aphelion of the orbit (typically 5.3 AU), the trajectory will offer a long arc in the swarm thus maximizing the number of flybys. Initial gravity assists from Venus and Earth will help reducing the cruise to 7 years as well as the ?V needed for injection thus offering enough capacity to navigate among Trojans. This solution further opens the unique possibility to flyby a Hilda asteroid when leaving the Trojan swarm. During the cruise phase, a Main Belt Asteroid could be targeted if requiring a modest ?V. The specific science objectives of the mission will be best achieved with a payload that will perform high-resolution panchromatic and multispectral imaging, thermal-infrared imaging/ radiometry, near- and mid-infrared spectroscopy

  5. Benchtop isolation and characterization of functional exosomes by sequential filtration.

    Science.gov (United States)

    Heinemann, Mitja L; Ilmer, Matthias; Silva, Leslie P; Hawke, David H; Recio, Alejandro; Vorontsova, Maria A; Alt, Eckhard; Vykoukal, Jody

    2014-12-05

    Early and minimally invasive detection of malignant events or other pathologies is of utmost importance in the pursuit of improved patient care and outcomes. Recent evidence indicates that exosomes and extracellular vesicles in serum and body fluids can contain nucleic acid, protein, and other biomarkers. Accordingly, there is great interest in applying these clinically as prognostic, predictive, pharmacodynamic, and early detection indicators. Nevertheless, existing exosome isolation methods can be time-consuming, require specialized equipment, and/or present other inefficiencies regarding purity, reproducibility and assay cost. We have developed a straightforward, three-step protocol for exosome isolation of cell culture supernatants or large volumes of biofluid based on sequential steps of dead-end pre-filtration, tangential flow filtration (TFF), and low-pressure track-etched membrane filtration that we introduce here. Our approach yields exosome preparations of high purity and defined size distribution and facilitates depletion of free protein and other low-molecular-weight species, extracellular vesicles larger than 100nm, and cell debris. Samples of exosomes prepared using the approach were verified morphologically by nanoparticle tracking analysis and electron microscopy, and mass spectrometry analyses confirmed the presence of previously reported exosome-associated proteins. In addition to being easy-to-implement, sequential filtration yields exosomes of high purity and, importantly, functional integrity as a result of the relatively low-magnitude manipulation forces employed during isolation. This answers an unmet need for preparation of minimally manipulated exosomes for investigations into exosome function and basic biology. Further, the strategy is amenable to translation for clinical exosome isolations because of its speed, automatability, scalability, and specificity for isolating exosomes from complex biological samples.

  6. Finding the Exosome

    OpenAIRE

    Mitchell, Phil; Tollervey, David

    2010-01-01

    We describe the events surrounding the identification of the exosome complex and the subsequent early development of the field. Like many scientific discoveries, the initial identification and characterization of the exosome was a based on a combination of skill, good fortune - and the availability of cutting edge technology.

  7. Bovine milk exosome proteome

    Science.gov (United States)

    Exosomes are 40-100 nm membrane vesicles of endocytic origin and are found in blood, urine, amniotic fluid, bronchoalveolar lavage (BAL) fluid, as well as human and bovine milk. Exosomes are extracellular organelles important in intracellular communication/signaling, immune function, and biomarkers ...

  8. Blood Exosomes Endowed with Magnetic and Targeting Properties for Cancer Therapy.

    Science.gov (United States)

    Qi, Hongzhao; Liu, Chaoyong; Long, Lixia; Ren, Yu; Zhang, Shanshan; Chang, Xiaodan; Qian, Xiaomin; Jia, Huanhuan; Zhao, Jin; Sun, Jinjin; Hou, Xin; Yuan, Xubo; Kang, Chunsheng

    2016-03-22

    Exosomes are a class of naturally occurring nanoparticles that are secreted endogenously by mammalian cells. Clinical applications for exosomes remain a challenge because of their unsuitable donors, low scalability, and insufficient targeting ability. In this study, we developed a dual-functional exosome-based superparamagnetic nanoparticle cluster as a targeted drug delivery vehicle for cancer therapy. The resulting exosome-based drug delivery vehicle exhibits superparamagnetic behavior at room temperature, with a stronger response to an external magnetic field than individual superparamagnetic nanoparticles. These properties enable exosomes to be separated from the blood and to target diseased cells. In vivo studies using murine hepatoma 22 subcutaneous cancer cells showed that drug-loaded exosome-based vehicle delivery enhanced cancer targeting under an external magnetic field and suppressed tumor growth. Our developments overcome major barriers to the utility of exosomes for cancer application.

  9. Exploitation of Exosomes as Nanocarriers for Gene-, Chemo-, and Immune-Therapy of Cancer.

    Science.gov (United States)

    Srivastava, Akhil; Babu, Anish; Filant, Justyna; Moxley, Katherine M; Ruskin, Rachel; Dhanasekaran, Danny; Sood, Anil K; McMeekin, Scott; Ramesh, Rajagopal

    2016-06-01

    The bottleneck in current vector-based cancer therapy is the targeted and controlled release of therapeutics in tumors. Exosomes are submicron-sized vesicles that are secreted by all cell types and are involved in communication and transportation of materials between cells. Analogous in size and function to synthetic nanoparticles, exosomes offer many advantages, rendering them the most promising candidates for targeted drug or gene delivery vehicles. Patient-specific customized therapeutic strategies can be engineered using exosomes derived from the patient's own healthy cells. Therefore, exosome-based cancer therapy has the potential to become an important part of personalized medicine. Interest in exosomes as carrier organelles is relatively recent. Knowledge about exosomal biology and its applications remains limited. The present review is an attempt to describe the current status of the application of exosomes to cancer therapy and the potential challenges associated with their use.

  10. Redefining the Breast Cancer Exosome Proteome by Tandem Mass Tag Quantitative Proteomics and Multivariate Cluster Analysis.

    Science.gov (United States)

    Clark, David J; Fondrie, William E; Liao, Zhongping; Hanson, Phyllis I; Fulton, Amy; Mao, Li; Yang, Austin J

    2015-10-20

    Exosomes are microvesicles of endocytic origin constitutively released by multiple cell types into the extracellular environment. With evidence that exosomes can be detected in the blood of patients with various malignancies, the development of a platform that uses exosomes as a diagnostic tool has been proposed. However, it has been difficult to truly define the exosome proteome due to the challenge of discerning contaminant proteins that may be identified via mass spectrometry using various exosome enrichment strategies. To better define the exosome proteome in breast cancer, we incorporated a combination of Tandem-Mass-Tag (TMT) quantitative proteomics approach and Support Vector Machine (SVM) cluster analysis of three conditioned media derived fractions corresponding to a 10 000g cellular debris pellet, a 100 000g crude exosome pellet, and an Optiprep enriched exosome pellet. The quantitative analysis identified 2 179 proteins in all three fractions, with known exosomal cargo proteins displaying at least a 2-fold enrichment in the exosome fraction based on the TMT protein ratios. Employing SVM cluster analysis allowed for the classification 251 proteins as "true" exosomal cargo proteins. This study provides a robust and vigorous framework for the future development of using exosomes as a potential multiprotein marker phenotyping tool that could be useful in breast cancer diagnosis and monitoring disease progression.

  11. Quantitative and stoichiometric analysis of the microRNA content of exosomes.

    Science.gov (United States)

    Chevillet, John R; Kang, Qing; Ruf, Ingrid K; Briggs, Hilary A; Vojtech, Lucia N; Hughes, Sean M; Cheng, Heather H; Arroyo, Jason D; Meredith, Emily K; Gallichotte, Emily N; Pogosova-Agadjanyan, Era L; Morrissey, Colm; Stirewalt, Derek L; Hladik, Florian; Yu, Evan Y; Higano, Celestia S; Tewari, Muneesh

    2014-10-14

    Exosomes have been proposed as vehicles for microRNA (miRNA) -based intercellular communication and a source of miRNA biomarkers in bodily fluids. Although exosome preparations contain miRNAs, a quantitative analysis of their abundance and stoichiometry is lacking. In the course of studying cancer-associated extracellular miRNAs in patient blood samples, we found that exosome fractions contained a small minority of the miRNA content of plasma. This low yield prompted us to perform a more quantitative assessment of the relationship between miRNAs and exosomes using a stoichiometric approach. We quantified both the number of exosomes and the number of miRNA molecules in replicate samples that were isolated from five diverse sources (i.e., plasma, seminal fluid, dendritic cells, mast cells, and ovarian cancer cells). Regardless of the source, on average, there was far less than one molecule of a given miRNA per exosome, even for the most abundant miRNAs in exosome preparations (mean ± SD across six exosome sources: 0.00825 ± 0.02 miRNA molecules/exosome). Thus, if miRNAs were distributed homogenously across the exosome population, on average, over 100 exosomes would need to be examined to observe one copy of a given abundant miRNA. This stoichiometry of miRNAs and exosomes suggests that most individual exosomes in standard preparations do not carry biologically significant numbers of miRNAs and are, therefore, individually unlikely to be functional as vehicles for miRNA-based communication. We propose revised models to reconcile the exosome-mediated, miRNA-based intercellular communication hypothesis with the observed stoichiometry of miRNAs associated with exosomes.

  12. Trojan capture by terrestrial planets

    CERN Document Server

    Schwarz, Richard

    2016-01-01

    The paper is devoted to investigate the capture of asteroids by Venus, Earth and Mars into the 1:1 mean motion resonance especially into Trojan orbits. Current theoretical studies predict that Trojan asteroids are a frequent by-product of the planet formation. This is not only the case for the outer giant planets, but also for the terrestrial planets in the inner Solar System. By using numerical integrations, we investigated the capture efficiency and the stability of the captured objects. We found out that the capture efficiency is larger for the planets in the inner Solar System compared to the outer ones, but most of the captured Trojan asteroids are not long term stable. This temporary captures caused by chaotic behaviour of the objects were investigated without any dissipative forces. They show an interesting dynamical behaviour of mixing like jumping from one Lagrange point to the other one.

  13. The roles and implications of exosomes in sarcoma.

    Science.gov (United States)

    Min, Li; Shen, Jacson; Tu, Chongqi; Hornicek, Francis; Duan, Zhenfeng

    2016-09-01

    Better diagnostic biomarkers and therapeutic options are still necessary for patients with sarcomas due to the current limitations of diagnosis and treatment. Exosomes are small extracellular membrane vesicles that are released by various cells and are found in most body fluids. Tumor-derived exosomes have been proven to mediate tumorigenesis, intercellular communication, microenvironment modulation, and metastasis in different cancers, including in sarcomas. Recently, exosomes have been considered as potential biomarkers for sarcoma diagnosis and prognosis, and as possible targets for sarcoma therapy. Moreover, due to their specific cell tropism and bioavailability, exosomes can also be engineered as vehicles for drug delivery. In this review, we discuss recent advances in the roles of tumor-derived exosomes in sarcoma and their potential clinical applications.

  14. Exosomes derived form bladder transitional cell carcinoma cells induce CTL cytotoxicity in vitro%膀胱移行细胞癌来源的exosome诱导体外细胞毒性T细胞杀伤效应

    Institute of Scientific and Technical Information of China (English)

    张家模; 吴小候; 张尧; 夏雨果; 罗春丽

    2009-01-01

    目的 观察膀胱移行细胞癌T24细胞来源的exosome体外诱导细胞毒性特异性T淋巴细胞(CTL)对肿瘤细胞的杀伤效应.方法 采用超滤和蔗糖密度梯度离心法分离T24细胞释放的exosome,电镜、Western blot观察exosome的特征.将exosome和肿瘤细胞负载到人外周血分离培养的树突状细胞(Dc)上,并与T细胞体外共同培养,分为exosome致敏DC组、未致敏DC组和对照组,Alamar blue检测CTL对T24细胞的细胞毒活性.结果 T24细胞分泌的exosome为直径约30~90nm的类圆碟形小囊泡.Western blot证实,exosome表达热休克蛋白70(HSP70)、细胞间黏附分子1(ICAM-1)和人细胞角蛋白20(CK20)分子.与未致敏DC组和对照组比较,exosome致敏DC组活化的T细胞对T24细胞有更强的细胞毒活性(P<0.01).结论 T24细胞来源的exosome负载了HSP70、ICAM-1等免疫相关蛋白;exosome经DC负载后活化CTL产生抗肿瘤活性.%Objective To isolate and purify exosomes derived from human bladder transitional cell carcinoma T24 cells,analyze the morphology and protein composition,and investigate the antitumor effect of specific cytotoxic T lymphocytes induced by exosomes.Methods Exosomes were isolated and purified by ultrafihration and sucrose gradient centrifugation,and characterized by electron microscopy and Western blot.Dendritic cells were amplified and purified from peripheral blood and pulsed with exosomes.Then they were co-cultured with T cells,and divided into 3 groups:exosome-pulsed DC group,unplused DC group and control group.Alamar-Blue assay was used to evaluate the specific cytolytic activity.Results The exosomes were in size about 30~90 nm saucer-shaped membranous vesicles.HSP70,ICAM-1 and CK20 were detected by Western blot.The CTL induced by DC pulsed with exosomes had significant cytolytic activity (P<0.01).Conclusion The exosomes derived from T24 cells are loaded with immunoprotein HSP70 and ICAM-1,and DC pulsed with exosomes can promote the anti

  15. Exosome enrichment of human serum using multiple cycles of centrifugation.

    Science.gov (United States)

    Kim, Jeongkwon; Tan, Zhijing; Lubman, David M

    2015-09-01

    In this work, we compared the use of repeated cycles of centrifugation at conventional speeds for enrichment of exosomes from human serum compared to the use of ultracentrifugation (UC). After removal of cells and cell debris, a speed of 110 000 × g or 40 000 × g was used for the UC or centrifugation enrichment process, respectively. The enriched exosomes were analyzed using the bicinchoninic acid assay, 1D gel separation, transmission electron microscopy, Western blotting, and high-resolution LC-MS/MS analysis. It was found that a five-cycle repetition of UC or centrifugation is necessary for successful removal of nonexosomal proteins in the enrichment of exosomes from human serum. More significantly, 5× centrifugation enrichment was found to provide similar or better performance than 5× UC enrichment in terms of enriched exosome protein amount, Western blot band intensity for detection of CD-63, and numbers of identified exosome-related proteins and cluster of differentiation (CD) proteins. A total of 478 proteins were identified in the LC-MS/MS analyses of exosome proteins obtained from 5× UCs and 5× centrifugations including many important CD membrane proteins. The presence of previously reported exosome-related proteins including key exosome protein markers demonstrates the utility of this method for analysis of proteins in human serum.

  16. Development of an aptasensor for electrochemical detection of exosomes.

    Science.gov (United States)

    Zhou, Qing; Rahimian, Ali; Son, Kyungjin; Shin, Dong-Sik; Patel, Tushar; Revzin, Alexander

    2016-03-15

    Exosomes are small (50-100 nm in diameter) vesicles secreted from various mammalian cells. Exosomes have been correlated with tumor antigens and anti-tumor immune responses and may represent cancer biomarkers. Herein, we report on the development of an aptamer-based electrochemical biosensor for quantitative detection of exosomes. Aptamers specific to exosome transmembrane protein CD63 were immobilized onto gold electrode surfaces and incorporated into a microfluidic system. Probing strands pre-labeled with redox moieties were hybridized onto aptamer molecules anchored on the electrode surface. In the presence of exosomes these beacons released probing strands with redox reporters causing electrochemical signal to decrease. These biosensors could be used to detect as few as 1×10(6) particles/mL of exosomes, which represents 100-fold decrease in the limit of detection compared to commercial immunoassays relying on anti-CD63 antibodies. Given the importance of exosome-mediated signal transmission among cells, our study may represent an important step towards development of a simple biosensor that detects exosomes without washing or labeling steps in complex media.

  17. Perturbations in the Urinary Exosome in Transplant Rejection

    Energy Technology Data Exchange (ETDEWEB)

    Sigdel, Tara K.; NG, Yolanda; Lee, Sangho; Nicora, Carrie D.; Qian, Weijun; Smith, Richard D.; Camp, David G.; Sarwal, Minnie M.

    2015-01-05

    Background: Urine exosomes, vesicles exocytosed into urine by all renal epithelial cell types, occur under normal physiologic and disease states. Exosome contents may mirror disease-specific proteome perturbations in kidney injury. Analysis methodologies for the exosomal fraction of the urinary proteome were developed and for comparing the urinary exosomal fraction versus unfractionated proteome for biomarker discovery. Methods: Urine exosomes were isolated by centrifugal filtration from mid-stream, second morning void, urine samples collected from kidney transplant recipients with and without biopsy matched acute rejection. The proteomes of unfractionated whole urine (Uw) and urine exosomes (Uexo) underwent mass spectrometry-based quantitative proteomics analysis. The proteome data were analyzed for significant differential protein abundances in acute rejection (AR). Results: Identifications of 1018 and 349 proteins, Uw and Uexo fractions, respectively, demonstrated a 279 protein overlap between the two urinary compartments with 25%(70) of overlapping proteins unique to Uexoand represented membrane bound proteins (p=9.31e-7). Of 349 urine exosomal proteins identified in transplant patients 220 were not previously identified in the normal urine exosomal fraction. Uexo proteins (11), functioning in the inflammatory / stress response, were more abundant in patients with biopsy-confirmed acute rejection, 3 of which were exclusive to Uexo. Uexo AR-specific biomarkers (8) were also detected in Uw, but since they were observed at significantly lower abundances in Uw, they were not significant for AR in Uw. Conclusions: A rapid urinary exosome isolation method and quantitative measurement of enriched Uexo proteins was applied. Urine proteins specific to the exosomal fraction were detected either in unfractionated urine (at low abundances) or by Uexo fraction analysis. Perturbed proteins in the exosomal compartment of urine collected from kidney transplant patients were

  18. Neutrophil-Derived Exosomes: A New Mechanism Contributing to Airway Smooth Muscle Remodeling.

    Science.gov (United States)

    Vargas, Amandine; Roux-Dalvai, Florence; Droit, Arnaud; Lavoie, Jean-Pierre

    2016-09-01

    Neutrophils infiltrate the airways of patients with asthma of all severities, yet their role in the pathogenesis of asthma and their contribution to airway remodeling is largely unknown. We hypothesized that neutrophils modulate airway smooth muscle (ASM) proliferation in asthma by releasing bioactive exosomes. These newly discovered nano-sized vesicles have the capacity to modulate immune responses, cell migration, cell differentiation, and other aspects of cell-to-cell communication. The aim of the study is to determine whether bioactive exosomes are released by neutrophils, and, if so, characterize their proteomic profile and evaluate their capacity to modulate ASM cell proliferation. Exosomes were isolated from equine neutrophil supernatants by differential centrifugation and filtration methods, followed by size-exclusion chromatography. Nanovesicles were characterized using electron microscopy, particle size determination, and proteomic analyses. Exosomes were cocultured with ASM cells and analyzed for exosome internalization by confocal microscopy. ASM proliferation was measured using an impedance-based system. Neutrophils release exosomes that have characteristic size, morphology, and exosomal markers. We identified 271 proteins in exosomes from both LPS and unstimulated neutrophils, and 16 proteins that were differentially expressed, which carried proteins associated with immune response and positive regulation of cell communication. Furthermore, neutrophil-derived exosomes were rapidly internalized by ASM cells and altered their proliferative properties. Upon stimulation of LPS, neutrophil-derived exosomes can enhance the proliferation of ASM cells and could therefore play an important role in the progression of asthma and promoting airway remodeling in severe and corticosteroid-insensitive patients with asthma.

  19. FedExosomes: Engineering Therapeutic Biological Nanoparticles that Truly Deliver

    Directory of Open Access Journals (Sweden)

    Michelle E. Marcus

    2013-04-01

    Full Text Available Many aspects of intercellular communication are mediated through “sending” and “receiving” packets of information via the secretion and subsequent receptor-mediated detection of biomolecular species including cytokines, chemokines, and even metabolites. Recent evidence has now established a new modality of intercellular communication through which biomolecular species are exchanged between cells via extracellular lipid vesicles. A particularly important class of extracellular vesicles is exosomes, which is a term generally applied to biological nanovesicles ~30–200 nm in diameter. Exosomes form through invagination of endosomes to encapsulate cytoplasmic contents, and upon fusion of these multivesicular endosomes to the cell surface, exosomes are released to the extracellular space and transport mRNA, microRNA (miRNA and proteins between cells. Importantly, exosome-mediated delivery of such cargo molecules results in functional modulation of the recipient cell, and such modulation is sufficiently potent to modulate disease processes in vivo. It is possible that such functional delivery of biomolecules indicates that exosomes utilize native mechanisms (e.g., for internalization and trafficking that may be harnessed by using exosomes to deliver exogenous RNA for therapeutic applications. A complementary perspective is that understanding the mechanisms of exosome-mediated transport may provide opportunities for “reverse engineering” such mechanisms to improve the performance of synthetic delivery vehicles. In this review, we summarize recent progress in harnessing exosomes for therapeutic RNA delivery, discuss the potential for engineering exosomes to overcome delivery challenges and establish robust technology platforms, and describe both potential challenges and advantages of utilizing exosomes as RNA delivery vehicles.

  20. Electrokinetic Evaluation of Individual Exosomes by On-Chip Microcapillary Electrophoresis with Laser Dark-Field Microscopy

    Science.gov (United States)

    Kato, Kei; Kobayashi, Masashi; Hanamura, Nami; Akagi, Takanori; Kosaka, Nobuyoshi; Ochiya, Takahiro; Ichiki, Takanori

    2013-06-01

    Cell-secreted nanovesicles called exosomes are expected as a promising candidate biomarker of various diseases. Toward the future application of exosomes as a disease biomarker for low-invasive diagnostics, challenges remain in the development of sensitive and precise analysis methods for exosomes. In this study, we performed the electrokinetic evaluation of individual exosomes by the combined use of on-chip microcapillary electrophoresis and laser dark-field microscopy. We extracted exosomes from six types of human cell cultured in a serum-free medium by differential ultracentrifugation and their zeta potential (electrophoretic mobility) were evaluated. We demonstrated that the proposed electrophoresis apparatus is particularly suitable for the tracking analysis of the electrophoretic migration of individual exosomes and enables the accurate evaluation of the zeta potential distribution of exosomes, for the first time. From the experimental results, we found that there is a strong correlation between the average zeta potentials of exosomes and their cells of origin.

  1. Exosomes mediated pentose phosphate pathway in ovarian cancer metastasis: a proteomics analysis.

    Science.gov (United States)

    Yi, Huan; Zheng, Xiangqin; Song, Jianrong; Shen, Rongkai; Su, Yanzhao; Lin, Danmei

    2015-01-01

    Epithelial ovarian cancer is the most lethal gynecological malignancies for readily metastasis. Exosomes have played an influential role in carcinogenicity and cancer progression. Our aim is to discover exosome-related mechanisms in ovarian cancer progress and explore potential diagnostic biomarkers and therapeutic targets of ovarian cancer. We initially presented the proteomic profiles of exosomes derived from two late-stage ovarian cell lines, OVCA429 and HO8910PM. A total of 2940 exosomal proteins were recorded by MS. FunRich appropriately processed these exosomal proteins, manifesting some superiority in contrast to Blast2go. Moreover, we demonstrated the pentose phosphate pathway was a dominant mechanism in exosome mediated intracellular communication. Glucose-6-phosphate dehydrogenase, transketolase and transaldolase 1, three key enzymes regulated pentose phosphate pathway, were all marked in the same exosomal parts of proteins between two ovarian cell lines. Moreover, these key proteins might become diagnostic, prognostic biomarkers and therapeutic targets of ovarian cancer.

  2. Exosome release of ADAM15 and the functional implications of human macrophage-derived ADAM15 exosomes.

    Science.gov (United States)

    Lee, Hee Doo; Koo, Bon-Hun; Kim, Yeon Hyang; Jeon, Ok-Hee; Kim, Doo-Sik

    2012-07-01

    A disintegrin and metalloproteinase 15 (ADAM15), the only ADAM protein containing an Arg-Gly-Asp (RGD) motif in its disintegrin-like domain, is a widely expressed membrane protein that is involved in tumor progression and suppression. However, the underlying mechanism of ADAM15-mediated tumor suppression is not clearly understood. This study demonstrates that ADAM15 is released as an exosomal component, and ADAM15 exosomes exert tumor suppressive activities. We found that exosomal ADAM15 release is stimulated by phorbol 12-myristate 13-acetate, a typical protein kinase C activator, in various tumor cell types, and this results in a corresponding decrease in plasma membrane-associated ADAM15. Exosomes rich in ADAM15 display enhanced binding affinity for integrin αvβ3 in an RGD-dependent manner and suppress vitronectin- and fibronectin-induced cell adhesion, growth, and migration, as well as in vivo tumor growth. Exosomal ADAM15 is released from human macrophages, and macrophage-derived ADAM15 exosomes have tumor inhibitory effects. This work suggests a primary role of ADAM15 for exosome-mediated tumor suppression, as well as functional significance of exosomal ADAM protein in antitumor immunity.

  3. Exosomal miRNAs as cancer biomarkers and therapeutic targets

    Directory of Open Access Journals (Sweden)

    Arron Thind

    2016-07-01

    Full Text Available Intercommunication between cancer cells and with their surrounding and distant environments is key to the survival, progression and metastasis of the tumour. Exosomes play a role in this communication process. MicroRNA (miRNA expression is frequently dysregulated in tumour cells and can be reflected by distinct exosomal miRNA (ex-miRNA profiles isolated from the bodily fluids of cancer patients. Here, the potential of ex-miRNA as a cancer biomarker and therapeutic target is critically analysed. Exosomes are a stable source of miRNA in bodily fluids but, despite a number of methods for exosome extraction and miRNA quantification, their suitability for diagnostics in a clinical setting is questionable. Furthermore, exosomally transferred miRNAs can alter the behaviour of recipient tumour and stromal cells to promote oncogenesis, highlighting a role in cell communication in cancer. However, our incomplete understanding of exosome biogenesis and miRNA loading mechanisms means that strategies to target exosomes or their transferred miRNAs are limited and not specific to tumour cells. Therefore, if ex-miRNA is to be employed in novel non-invasive diagnostic approaches and as a therapeutic target in cancer, two further advances are necessary: in methods to isolate and detect ex-miRNA, and a better understanding of their biogenesis and functions in tumour-cell communication.

  4. Controlled exosome release from the retinal pigment epithelium in situ.

    Science.gov (United States)

    Locke, Christina J; Congrove, Nicole R; Dismuke, W Michael; Bowen, Trent J; Stamer, W Daniel; McKay, Brian S

    2014-12-01

    Retinal Pigment Epithelial cells (RPE) express both GPR143 and myocilin, which interact in a signal transduction-dependent manner. In heterologous systems, activation of GPR143 with ligand causes transient recruitment of myocilin to internalized receptors, which appears to be the entry point of myocilin to the endocytic pathway. In some but not all cells, myocilin also traffics through the multivesicular body (MVB) and is released on the surface of exosomes in a signal transduction-dependent fashion. Little is known regarding the role of exosomes in RPE, but they likely serve as a mode of communication between the RPE and the outer retina. In this study, we used posterior poles with retina removed from fresh human donor eyes as a model to test the relationship between GPR143, myocilin, and exosomes in an endogenous system. We isolated exosomes released by RPE using differential centrifugation of media conditioned by the RPE for 25 min, and then characterized the exosomes using nanoparticle tracking to determine the number and size of the exosomes. Next, we tested whether ligand stimulation of GPR143 using l-DOPA altered RPE exosome release. Finally, we investigated whether myocilin was present on the exosomes released by RPE and whether l-DOPA stimulation of GPR143 caused recruitment of myocilin to the endocytic pathway, as we have previously observed using cultured cells. Activation of GPR143 halted RPE exosome release, while simultaneously recruiting myocilin to the endocytic compartment. Together, our results indicate that GPR143 and myocilin function in a signal transduction system that can control exosome release from RPE.

  5. Exosome Biogenesis, Regulation, and Function in Viral Infection

    Directory of Open Access Journals (Sweden)

    Marta Alenquer

    2015-09-01

    Full Text Available Exosomes are extracellular vesicles released upon fusion of multivesicular bodies(MVBs with the cellular plasma membrane. They originate as intraluminal vesicles (ILVs duringthe process of MVB formation. Exosomes were shown to contain selectively sorted functionalproteins, lipids, and RNAs, mediating cell-to-cell communications and hence playing a role in thephysiology of the healthy and diseased organism. Challenges in the field include the identificationof mechanisms sustaining packaging of membrane-bound and soluble material to these vesicles andthe understanding of the underlying processes directing MVBs for degradation or fusion with theplasma membrane. The investigation into the formation and roles of exosomes in viral infection is inits early years. Although still controversial, exosomes can, in principle, incorporate any functionalfactor, provided they have an appropriate sorting signal, and thus are prone to viral exploitation.This review initially focuses on the composition and biogenesis of exosomes. It then explores theregulatory mechanisms underlying their biogenesis. Exosomes are part of the endocytic system,which is tightly regulated and able to respond to several stimuli that lead to alterations in thecomposition of its sub-compartments. We discuss the current knowledge of how these changesaffect exosomal release. We then summarize how different viruses exploit specific proteins ofendocytic sub-compartments and speculate that it could interfere with exosome function, althoughno direct link between viral usage of the endocytic system and exosome release has yet beenreported. Many recent reports have ascribed functions to exosomes released from cells infectedwith a variety of animal viruses, including viral spread, host immunity, and manipulation of themicroenvironment, which are discussed. Given the ever-growing roles and importance of exosomesin viral infections, understanding what regulates their composition and levels, and

  6. Characterization of CLL exosomes reveals a distinct microRNA signature and enhanced secretion by activation of BCR signaling.

    Science.gov (United States)

    Yeh, Yuh-Ying; Ozer, Hatice Gulcin; Lehman, Amy M; Maddocks, Kami; Yu, Lianbo; Johnson, Amy J; Byrd, John C

    2015-05-21

    Multiple studies show that chronic lymphocytic leukemia (CLL) cells are heavily dependent on their microenvironment for survival. Communication between CLL cells and the microenvironment is mediated through direct cell contact, soluble factors, and extracellular vesicles. Exosomes are small particles enclosed with lipids, proteins, and small RNAs that can convey biological materials to surrounding cells. Our data herein demonstrate that CLL cells release significant amounts of exosomes in plasma that exhibit abundant CD37, CD9, and CD63 expression. Our work also pinpoints the regulation of B-cell receptor (BCR) signaling in the release of CLL exosomes: BCR activation by α-immunoglobulin (Ig)M induces exosome secretion, whereas BCR inactivation via ibrutinib impedes α-IgM-stimulated exosome release. Moreover, analysis of serial plasma samples collected from CLL patients on an ibrutinib clinical trial revealed that exosome plasma concentration was significantly decreased following ibrutinib therapy. Furthermore, microRNA (miR) profiling of plasma-derived exosomes identified a distinct exosome microRNA signature, including miR-29 family, miR-150, miR-155, and miR-223 that have been associated with CLL disease. Interestingly, expression of exosome miR-150 and miR-155 increases with BCR activation. In all, this study successfully characterized CLL exosomes, demonstrated the control of BCR signaling in the release of CLL exosomes, and uncovered a disease-relevant exosome microRNA profile.

  7. Exosomes and Their Signiifcance in Diagnosis and Treatment of Tumors

    Institute of Scientific and Technical Information of China (English)

    WANG Jian; LI Chao; LI Wei

    2015-01-01

    In the research field of biological markers for tumor diagnosis, the appearance of exosomes has resolved the problem that RNA molecules can be easily degraded. Exosomes carry various RNAs and can protect them from being degraded. They are deifned as polymorphism vesicle-like corpuscles (diameter: 30-100 nm) derived from late endosome or multi-vesicular endosomes in cellular endocytosis system, which contain abundant biological information, including multiple lipids, proteins and nucleic acids, etc. Exosomes are extracellular nanoscale vesicae formed in a series of regulating process of cellular “endocytosis-fusion-excretion”, and they carry proteins and transport RNAs, thus playing an important role in the intercellular material and informational transduction. There are still large amount of mRNAs and miRNAs in exosomes. Exosomes can not only protect in-vitro RNA stability, but also transfer RNA to speciifc target cells as effective carriers so as to play their regulatory function. Exosomes realize their biological information exchanges and transition via endocrine, paracrine and autocrine, and regulate cellular biological activities through direct action on superficial signal molecules or extracellular release and membrane fusion of biological active ingredients. They can directly act on tumors to impact tumor progression, or improve tumor angiogenesis and metastasis by regulating immunological function. Additionally, they can also be used for tumor diagnosis. Therefore, this study mainly summarized the biological characteristics of exosomes and their application in the regulation, diagnosis and treatment of tumors, hoping to provide references for the application of exosomes in tumors.

  8. Exosome removal as a therapeutic adjuvant in cancer

    Directory of Open Access Journals (Sweden)

    Marleau Annette M

    2012-06-01

    Full Text Available Abstract Exosome secretion is a notable feature of malignancy owing to the roles of these nanoparticles in cancer growth, immune suppression, tumor angiogenesis and therapeutic resistance. Exosomes are 30–100 nm membrane vesicles released by many cells types during normal physiological processes. Tumors aberrantly secrete large quantities of exosomes that transport oncoproteins and immune suppressive molecules to support tumor growth and metastasis. The role of exosomes in intercellular signaling is exemplified by human epidermal growth factor receptor type 2 (HER2 over-expressing breast cancer, where exosomes with the HER2 oncoprotein stimulate tumor growth and interfere with the activity of the therapeutic antibody Herceptin®. Since numerous observations from experimental model systems point toward an important clinical impact of exosomes in cancer, several pharmacological strategies have been proposed for targeting their malignant activities. We also propose a novel device strategy involving extracorporeal hemofiltration of exosomes from the entire circulatory system using an affinity plasmapheresis platform known as the Aethlon ADAPT™ (adaptive dialysis-like affinity platform technology system, which would overcome the risks of toxicity and drug interactions posed by pharmacological approaches. This technology allows affinity agents, including exosome-binding lectins and antibodies, to be immobilized in the outer-capillary space of plasma filtration membranes that integrate into existing kidney dialysis systems. Device therapies that evolve from this platform allow rapid extracorporeal capture and selective retention of target particles 

  9. Exosome mimetics: a novel class of drug delivery systems.

    Science.gov (United States)

    Kooijmans, Sander A A; Vader, Pieter; van Dommelen, Susan M; van Solinge, Wouter W; Schiffelers, Raymond M

    2012-01-01

    The identification of extracellular phospholipid vesicles as conveyors of cellular information has created excitement in the field of drug delivery. Biological therapeutics, including short interfering RNA and recombinant proteins, are prone to degradation, have limited ability to cross biological membranes, and may elicit immune responses. Therefore, delivery systems for such drugs are under intensive investigation. Exploiting extracellular vesicles as carriers for biological therapeutics is a promising strategy to overcome these issues and to achieve efficient delivery to the cytosol of target cells. Exosomes are a well studied class of extracellular vesicles known to carry proteins and nucleic acids, making them especially suitable for such strategies. However, the considerable complexity and the related high chance of off-target effects of these carriers are major barriers for translation to the clinic. Given that it is well possible that not all components of exosomes are required for their proper functioning, an alternative strategy would be to mimic these vesicles synthetically. By assembly of liposomes harboring only crucial components of natural exosomes, functional exosome mimetics may be created. The low complexity and use of well characterized components strongly increase the pharmaceutical acceptability of such systems. However, exosomal components that would be required for the assembly of functional exosome mimetics remain to be identified. This review provides insights into the composition and functional properties of exosomes, and focuses on components which could be used to enhance the drug delivery properties of exosome mimetics.

  10. Exosomes and the emerging field of exosome-based gene therapy.

    Science.gov (United States)

    O'Loughlin, Aisling J; Woffindale, Caroline A; Wood, Matthew J A

    2012-08-01

    Exosomes are a subtype of membrane vesicle released from the endocytic compartment of live cells. They play an important role in endogenous cell-to-cell communication. Previously shown to be capable of traversing biological barriers and to naturally transport functional nucleic acids between cells, they potentially represent a novel and exciting drug delivery vehicle for the field of gene therapy. Existing delivery vehicles are limited by concerns regarding their safety, toxicity and efficacy. In contrast, exosomes, as a natural cell-derived nanocarrier, are immunologically inert if purified from a compatible cell source and possess an intrinsic ability to cross biological barriers. Already utilised in a number of clinical trials, exosomes appear to be well-tolerated, even following repeat administration. Recent studies have shown that exosomes may be used to encapsulate and protect exogenous oligonucleotides for delivery to target cells. They therefore may be valuable for the delivery of RNA interference and microRNA regulatory molecules in addition to other single-stranded oligonucleotides. Prior to clinical translation, this nanotechnology requires further development by refinement of isolation, purification, loading, delivery and targeting protocols. Thus, exosome-mediated nanodelivery is highly promising and may fill the void left by current delivery methods for systemic gene therapy.

  11. Exosomes serve as tumour markers for personalized diagnostics owing to their important role in cancer metastasis

    OpenAIRE

    2015-01-01

    Exosomes, membrane vesicles of 40–100 nm in diameter, are derived from endosomes in various cells. The bioactive molecules specifically packed into exosomes can be horizontally transferred into recipient cells changing their biological properties, by which tumour cells continuously modify their surrounding microenvironment and distant target cells favouring cancer metastasis. It has been suspected for a long time that exosomes participate in the whole process of tumour metastasis. Although th...

  12. Tumor-derived exosomes and their role in cancer progression

    Science.gov (United States)

    Whiteside, Theresa L

    2017-01-01

    Tumor cells actively produce, release and utilize exosomes to promote tumor growth. Mechanisms through which tumor-derived exosomes subserve the tumor are under intense investigation. These exosomes are information carriers, conveying molecular and genetic messages from tumor cells to normal or other abnormal cells residing at close or distant sites. Tumor-derived exosomes are found in all body fluids. Upon the contact with target cells, they alter phenotypic and functional attributes of recipients, reprogramming them into active contributors to angiogenesis, thrombosis, metastasis and immunosuppression. Exosomes produced by tumors carry cargos that in part mimic contents of parent cells and are of potential interest as non-invasive biomarkers of cancer. Their role in inhibiting the host antitumor responses and in mediating drug resistance is important for cancer therapy. Tumor-derived exosomes may interfere with cancer immunotherapy, but they also could serve as adjuvants and antigenic components of antitumor vaccines. Their biological roles in cancer development or progression as well as cancer therapy suggest that tumor-derived exosomes are critical components of oncogenic transformation. PMID:27117662

  13. Tumor-Derived Exosomes and Their Role in Cancer Progression.

    Science.gov (United States)

    Whiteside, Theresa L

    2016-01-01

    Tumor cells actively produce, release, and utilize exosomes to promote tumor growth. Mechanisms through which tumor-derived exosomes subserve the tumor are under intense investigation. These exosomes are information carriers, conveying molecular and genetic messages from tumor cells to normal or other abnormal cells residing at close or distant sites. Tumor-derived exosomes are found in all body fluids. Upon contact with target cells, they alter phenotypic and functional attributes of recipients, reprogramming them into active contributors to angiogenesis, thrombosis, metastasis, and immunosuppression. Exosomes produced by tumors carry cargos that in part mimic contents of parent cells and are of potential interest as noninvasive biomarkers of cancer. Their role in inhibiting the host antitumor responses and in mediating drug resistance is important for cancer therapy. Tumor-derived exosomes may interfere with cancer immunotherapy, but they also could serve as adjuvants and antigenic components of antitumor vaccines. Their biological roles in cancer development or progression as well as cancer therapy suggest that tumor-derived exosomes are critical components of oncogenic transformation.

  14. Neptune´s Trojans

    Directory of Open Access Journals (Sweden)

    Tabaré Gallardo

    2001-01-01

    Full Text Available A numerical exploration of the dynamical evolution of hypothetical bodies located at 1:1 resonance with Neptune is performed. We roughly estimate a time-scale of some 100 Myrs for the destruction of the librations, so we cannot expect to find primordial Neptune´s trojans. Temporary satellite capture s were also observed.

  15. A Trojan Horse in Birmingham

    Science.gov (United States)

    Yarker, Patrick

    2014-01-01

    "Trojan Horse" has become journalistic shorthand for an apparent attempt by a small group in East Birmingham to secure control of local non-faith schools and impose policies and practices in keeping with the very conservative (Salafist and Wahhabi) version of Islam which they hold. In this article, Pat Yarker gives an account of two…

  16. Trojan Horses or Local Allies

    DEFF Research Database (Denmark)

    Müllner, Jakob; Klopf, Patricia; Nell, Phillip Christopher

    . On the negative side, we argue that these characteristics of HCNs can potentially be used to the harm of a foreign MNC. We analyze how formal and informal institutions affect the trade-off between positive effects and the potential costs associated with HCN managers (“Local allies” vs. “Trojan horses”). We find...

  17. Radiation increases the cellular uptake of exosomes through CD29/CD81 complex formation

    Energy Technology Data Exchange (ETDEWEB)

    Hazawa, Masaharu; Tomiyama, Kenichi; Saotome-Nakamura, Ai; Obara, Chizuka; Yasuda, Takeshi; Gotoh, Takaya; Tanaka, Izumi; Yakumaru, Haruko; Ishihara, Hiroshi; Tajima, Katsushi, E-mail: tajima@nirs.go.jp

    2014-04-18

    Highlights: • Radiation increases cellular uptake of exosomes. • Radiation induces colocalization of CD29 and CD81. • Exosomes selectively bind the CD29/CD81 complex. • Radiation increases the cellular uptake of exosomes through CD29/CD81 complex formation. - Abstract: Exosomes mediate intercellular communication, and mesenchymal stem cells (MSC) or their secreted exosomes affect a number of pathophysiologic states. Clinical applications of MSC and exosomes are increasingly anticipated. Radiation therapy is the main therapeutic tool for a number of various conditions. The cellular uptake mechanisms of exosomes and the effects of radiation on exosome–cell interactions are crucial, but they are not well understood. Here we examined the basic mechanisms and effects of radiation on exosome uptake processes in MSC. Radiation increased the cellular uptake of exosomes. Radiation markedly enhanced the initial cellular attachment to exosomes and induced the colocalization of integrin CD29 and tetraspanin CD81 on the cell surface without affecting their expression levels. Exosomes dominantly bound to the CD29/CD81 complex. Knockdown of CD29 completely inhibited the radiation-induced uptake, and additional or single knockdown of CD81 inhibited basal uptake as well as the increase in radiation-induced uptake. We also examined possible exosome uptake processes affected by radiation. Radiation-induced changes did not involve dynamin2, reactive oxygen species, or their evoked p38 mitogen-activated protein kinase-dependent endocytic or pinocytic pathways. Radiation increased the cellular uptake of exosomes through CD29/CD81 complex formation. These findings provide essential basic insights for potential therapeutic applications of exosomes or MSC in combination with radiation.

  18. Combining Exosomes Derived from Immature DCs with Donor Antigen-Specific Treg Cells Induces Tolerance in a Rat Liver Allograft Model

    Science.gov (United States)

    Ma, Ben; Yang, Jing-Yue; Song, Wen-jie; Ding, Rui; Zhang, Zhuo-chao; Ji, Hong-chen; Zhang, Xuan; Wang, Jian-lin; Yang, Xi-sheng; Tao, Kai-shan; Dou, Ke-feng; Li, Xiao

    2016-01-01

    Allograft tolerance is the ultimate goal in the field of transplantation immunology. Immature dendritic cells (imDCs) play an important role in establishing tolerance but have limitations, including potential for maturation, short lifespan in vivo and short storage times in vitro. However, exosomes (generally 30–100 nm) from imDCs (imDex) retain many source cell properties and may overcome these limitations. In previous reports, imDex prolonged the survival time of heart or intestine allografts. However, tolerance or long-term survival was not achieved unless immune suppressants were used. Regulatory T cells (Tregs) can protect allografts from immune rejection, and our previous study showed that the effects of imDex were significantly associated with Tregs. Therefore, we incorporated Tregs into the treatment protocol to further reduce or avoid suppressant use. We defined the optimal exosome dose as approximately 20 μg (per treatment before, during and after transplantation) in rat liver transplantation and the antigen-specific role of Tregs in protecting liver allografts. In the co-treatment group, recipients achieved long-term survival, and tolerance was induced. Moreover, imDex amplified Tregs, which required recipient DCs and were enhanced by IL-2. Fortunately, the expanded Tregs retained their regulatory ability and donor-specificity. Thus, imDex and donor-specific Tregs can collaboratively induce graft tolerance. PMID:27640806

  19. The non-targeted effects of radiation are perpetuated by exosomes

    Energy Technology Data Exchange (ETDEWEB)

    Al-Mayah, Ammar; Bright, Scott; Chapman, Kim [Genomic Instability Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP (United Kingdom); Irons, Sarah [Insect Virus Research Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP (United Kingdom); Luo, Ping [Izon Science Ltd., The Oxford Science Park, Magdalen Centre, Robert Robinson Avenue, Oxford OX4 4GA (United Kingdom); Carter, David [Chromatin and non-coding RNA, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP (United Kingdom); Goodwin, Edwin [The New Mexico Consortium, Los Alamos, NM 87544 (United States); Kadhim, Munira, E-mail: mkadhim@brookes.ac.uk [Genomic Instability Group, Oxford Brookes University, Gipsy Lane Campus, Headington, Oxford OX3 0BP (United Kingdom)

    2015-02-15

    Highlights: • Radiation induces a DNA damaging process in bystander cells through cell–cell signalling. • Exosome RNA and protein molecules play crucial roles in bystander effects. • Cell progeny inherit the ability to secret bystander effect-inducing exosomes. • This mechanism is most likely accountable for the propagation of GI. - Abstract: Exosomes contain cargo material from endosomes, cytosol, plasma membrane and microRNA molecules, they are released by a number of non-cancer and cancer cells into both the extracellular microenvironment and body fluids such as blood plasma. Recently we demonstrated radiation-induced non-targeted effects [NTE: genomic instability (GI) and bystander effects (BE)] are partially mediated by exosomes, particularly the RNA content. However the mechanistic role of exosomes in NTE is yet to be fully understood. The present study used MCF7 cells to characterise the longevity of exosome-induced activity in the progeny of irradiated and unirradiated bystander cells. Exosomes extracted from conditioned media of irradiated and bystander progeny were added to unirradiated cells. Analysis was carried out at 1 and 20/24 population doublings following medium/exosome transfer for DNA/chromosomal damage. Results confirmed exosomes play a significant role in mediating NTE of ionising radiation (IR). This effect was remarkably persistent, observed >20 doublings post-irradiation in the progeny of bystander cells. Additionally, cell progeny undergoing a BE were themselves capable of inducing BE in other cells via exosomes they released. Furthermore we investigated the role of exosome cargo. Culture media from cells exposed to 2 Gy X-rays was subjected to ultracentrifugation and four inoculants prepared, (a) supernatants with exosomes removed, and pellets with (b) exosome proteins denatured, (c) RNA degraded, and (d) a combination of protein–RNA inactivation. These were added to separate populations of unirradiated cells. The BE was

  20. Exosomes: improved methods to characterize their morphology, RNA content, and surface protein biomarkers.

    Science.gov (United States)

    Wu, Yueting; Deng, Wentao; Klinke, David J

    2015-10-07

    As a type of secreted membrane vesicle, exosomes are an emerging mode of cell-to-cell communication. Yet as exosome samples are commonly contaminated with other extracellular vesicles, the biological roles of exosomes in regulating immunity and promoting oncogenesis remain controversial. Wondering whether existing methods could distort our view of exosome biology, we compared two direct methods for imaging extracellular vesicles and quantified the impact of different production and storage conditions on the quality of exosome samples. Scanning electron microscopy (SEM) was compared to transmission electron microscopy (TEM) as alternatives to examine the morphology of exosomes. Using SEM, we were able to distinguish exosomes from other contaminating extracellular vesicles based on the size distribution. More importantly, freezing of samples prior to SEM imaging made it more difficult to distinguish exosomes from extracellular vesicles secreted during cell death. In addition to morphology, the quality of RNA contained within the exosomes was characterized under different storage conditions, where freezing of samples also degraded RNA. Finally, we developed a new flow cytometry approach to assay transmembrane proteins on exosomes. While high-copy-number proteins could be readily detected, detecting low-copy-number proteins was improved using a lipophilic tracer that clustered exosomes. To illustrate this, we observed that exosomes derived from SKBR3 cells, a cell model for human HER2+ breast cancer, contained both HER1 and HER2 but at different levels of abundance. Collectively, these new methods will help to ensure a consistent framework to identify specific roles that exosomes play in regulating cell-to-cell communication.

  1. Functional significance of macrophage-derived exosomes in inflammation and pain.

    Science.gov (United States)

    McDonald, Marguerite K; Tian, Yuzhen; Qureshi, Rehman A; Gormley, Michael; Ertel, Adam; Gao, Ruby; Aradillas Lopez, Enrique; Alexander, Guillermo M; Sacan, Ahmet; Fortina, Paolo; Ajit, Seena K

    2014-08-01

    Exosomes, secreted microvesicles transporting microRNAs (miRNAs), mRNAs, and proteins through bodily fluids, facilitate intercellular communication and elicit immune responses. Exosomal contents vary, depending on the source and the physiological conditions of cells, and can provide insights into how cells and systems cope with physiological perturbations. Previous analysis of circulating miRNAs in patients with complex regional pain syndrome (CRPS), a debilitating chronic pain disorder, revealed a subset of miRNAs in whole blood that are altered in the disease. To determine functional consequences of alterations in exosomal biomolecules in inflammation and pain, we investigated exosome-mediated information transfer in vitro, in a rodent model of inflammatory pain, and in exosomes from patients with CRPS. Mouse macrophage cells stimulated with lipopolysaccharides secrete exosomes containing elevated levels of cytokines and miRNAs that mediate inflammation. Transcriptome sequencing of exosomal RNA revealed global alterations in both innate and adaptive immune pathways. Exosomes from lipopolysaccharide-stimulated cells were sufficient to cause nuclear factor-κB activation in naive cells, indicating functionality in recipient cells. A single injection of exosomes attenuated thermal hyperalgesia in a murine model of inflammatory pain, suggesting an immunoprotective role for macrophage-derived exosomes. Macrophage-derived exosomes carry a protective signature that is altered when secreting cells are exposed to an inflammatory stimulus. We also show that circulating miRNAs altered in patients with complex regional pain syndrome are trafficked by exosomes. With their systemic signaling capabilities, exosomes can induce pleiotropic effects potentially mediating the multifactorial pathology underlying chronic pain, and should be explored for their therapeutic utility.

  2. Tumour-derived exosomes: Tiny envelopes for big stories.

    Science.gov (United States)

    Miller, Isabella V; Grunewald, Thomas G P

    2015-09-01

    The discovery of exosomes, which are small, 30-100 nm sized extracellular vesicles that are released by virtual all cells, has initiated a rapidly expanding and vibrant research field. Current investigations are mainly directed toward the role of exosomes in intercellular communication and their potential value as biomarkers for a broad set of diseases. By horizontal transfer of molecular information such as micro RNAs, messenger RNAs or proteins, as well as by receptor-cell interactions, exosomes are capable to mediate the reprogramming of surrounding cells. Herein, we review how especially cancer cells take advantage of this mechanism to influence their microenvironment in favour of immune escape, therapy resistance, tumour growth and metastasis. Moreover, we provide a comprehensive microarray analysis (n > 1970) to study the expression patterns of genes known to be intimately involved in exosome biogenesis across 26 different cancer entities and a normal tissue atlas. Consistent with the elevated production of exosomes observed in cancer patient plasma, we found a significant overexpression especially of RAB27A, CHMP4C and SYTL4 in the corresponding cancer entities as compared to matched normal tissues. Finally, we discuss the immune-modulatory and anti-tumorigenic functions of exosomes as well as innovative approaches to specifically target the exosomal circuits in experimental cancer therapy.

  3. Exosomes serve as tumour markers for personalized diagnostics owing to their important role in cancer metastasis

    Directory of Open Access Journals (Sweden)

    Taixue An

    2015-06-01

    Full Text Available Exosomes, membrane vesicles of 40–100 nm in diameter, are derived from endosomes in various cells. The bioactive molecules specifically packed into exosomes can be horizontally transferred into recipient cells changing their biological properties, by which tumour cells continuously modify their surrounding microenvironment and distant target cells favouring cancer metastasis. It has been suspected for a long time that exosomes participate in the whole process of tumour metastasis. Although there is much unknown and many controversies in the role of cancer exosome, the major contribution of tumour-associated exosomes to different steps of cancer metastasis are demonstrated in this review. Mainly because these exosomes are easily accessible and capable of representing their parental cells, exosomes draw much attention as a promising biomarker for tumour screening, diagnosis and prognosis. Currently, researchers have found numerous biomarkers in exosomes with great potential to be utilized in personalized medicine. In this article, we summarize the roles of biomarkers, which are validated by clinical samples. Even though many conundrums remain, such as exosome extraction, large multicentre validation of biomarkers and data interpretation, exosomes are certain to be used in clinical practice in the near future as the field rapidly expands.

  4. Exosomes serve as tumour markers for personalized diagnostics owing to their important role in cancer metastasis.

    Science.gov (United States)

    An, Taixue; Qin, Sihua; Xu, Yong; Tang, Yueting; Huang, Yiyao; Situ, Bo; Inal, Jameel M; Zheng, Lei

    2015-01-01

    Exosomes, membrane vesicles of 40-100 nm in diameter, are derived from endosomes in various cells. The bioactive molecules specifically packed into exosomes can be horizontally transferred into recipient cells changing their biological properties, by which tumour cells continuously modify their surrounding microenvironment and distant target cells favouring cancer metastasis. It has been suspected for a long time that exosomes participate in the whole process of tumour metastasis. Although there is much unknown and many controversies in the role of cancer exosome, the major contribution of tumour-associated exosomes to different steps of cancer metastasis are demonstrated in this review. Mainly because these exosomes are easily accessible and capable of representing their parental cells, exosomes draw much attention as a promising biomarker for tumour screening, diagnosis and prognosis. Currently, researchers have found numerous biomarkers in exosomes with great potential to be utilized in personalized medicine. In this article, we summarize the roles of biomarkers, which are validated by clinical samples. Even though many conundrums remain, such as exosome extraction, large multicentre validation of biomarkers and data interpretation, exosomes are certain to be used in clinical practice in the near future as the field rapidly expands.

  5. miR150*-Loaded exosomes from marrow stromal cells could inhibit glioma cells growth%miR-150*修饰骨髓间充质干细胞来源的 exosome对胶质瘤细胞的影响

    Institute of Scientific and Technical Information of China (English)

    廖克曼; 季卫阳; 鲁晓杰

    2015-01-01

    Objective MiRNA-based therapeutics hold great promise for tumor suppression, this study was to investigate the effect of miR-150*-loaded exosomes on regulation of glioma cells proliferation and cell cycle.Methods Quantitative real-time PCR on 15 glioblastoma tissues samples and normal controls were used to confirm the miR-150* expression level.Western blotting analysis and electron microscopy were employed to test exosomal biomarkers and their morphology. Transfection assay were used to collect miR-150*-loaded exosomes from bone marrow mesenchymal stem cells (BMSCs)culture medium.CCK-8 and cell cycle assays were used to analyze miR-150*-loaded exosomes effects on glioma cells.Results Level of miR-150* expression was much lower in glioblastoma than in normal tissues.Transfection assay successfully acquired miR150*-loaded exosomes which derived from bone marrow mesenchymal stem cells (BMSCs).Furthermore,miR-150*-loaded exosomes could largely inhibited glioma cells proliferation and suppress cell cycle progression.Cell counting kit 8 (CCK-8)assays also demonstrated miR-150* delivered in exosomes was much less toxic.Conclusions This study demonstrated miR-150* is down-regulated in glioblastoma.miR-150*-loaded exosomes could suppress glioma cells and exosomes may be a potentially efficient therapeutic delivery system.%目的:研究 miR-150*修饰对骨髓间充质干细胞来源的囊泡(exosome)对胶质瘤细胞的影响。方法qRT-PCR 检测 miR-150*在胶质母细胞瘤组织与正常组织间的表达量差异。培养骨髓间充质干细胞(BMSCs),分别转染 miR-150*模拟物和阴性对照序列,上调 BMSCs 中 miR-150*表达水平,提取 BMSCs 培养基中的 exosome。Western blot 验证 exosomal 的表面标记蛋白 CD63和 flotillin-1,电镜下观察 exosome 的形态。CCK-8和细胞周期实验验证 miR-150*修饰 BMSCs 来源的 exosome 对胶质瘤细胞的影响。结果miR-150*在胶质母细胞瘤组织中表达明显

  6. Exosome removal as a therapeutic adjuvant in cancer.

    Science.gov (United States)

    Marleau, Annette M; Chen, Chien-Shing; Joyce, James A; Tullis, Richard H

    2012-06-27

    Exosome secretion is a notable feature of malignancy owing to the roles of these nanoparticles in cancer growth, immune suppression, tumor angiogenesis and therapeutic resistance. Exosomes are 30-100 nm membrane vesicles released by many cells types during normal physiological processes. Tumors aberrantly secrete large quantities of exosomes that transport oncoproteins and immune suppressive molecules to support tumor growth and metastasis. The role of exosomes in intercellular signaling is exemplified by human epidermal growth factor receptor type 2 (HER2) over-expressing breast cancer, where exosomes with the HER2 oncoprotein stimulate tumor growth and interfere with the activity of the therapeutic antibody Herceptin®. Since numerous observations from experimental model systems point toward an important clinical impact of exosomes in cancer, several pharmacological strategies have been proposed for targeting their malignant activities. We also propose a novel device strategy involving extracorporeal hemofiltration of exosomes from the entire circulatory system using an affinity plasmapheresis platform known as the Aethlon ADAPT™ (adaptive dialysis-like affinity platform technology) system, which would overcome the risks of toxicity and drug interactions posed by pharmacological approaches. This technology allows affinity agents, including exosome-binding lectins and antibodies, to be immobilized in the outer-capillary space of plasma filtration membranes that integrate into existing kidney dialysis systems. Device therapies that evolve from this platform allow rapid extracorporeal capture and selective retention of target particles exosomes. This review discusses the possible therapeutic approaches for targeting immune suppressive exosomes in cancer patients, and the anticipated significance of these strategies for reversing immune dysfunction and improving responses to standard of care treatments.

  7. Exosomes: novel effectors of human platelet lysate activity

    Directory of Open Access Journals (Sweden)

    E Torreggiani

    2014-09-01

    Full Text Available Despite the popularity of platelet-rich plasma (PRP and platelet lysate (PL in orthopaedic practice, the mechanism of action and the effectiveness of these therapeutic tools are still controversial. So far, the activity of PRP and PL has been associated with different growth factors (GF released during platelet degranulation. This study, for the first time, identifies exosomes, nanosized vesicles released in the extracellular compartment by a number of elements, including platelets, as one of the effectors of PL activity. Exosomes were isolated from human PL by differential ultracentrifugation, and analysed by electron microscopy and Western blotting. Bone marrow stromal cells (MSC treated with three different exosome concentrations (0.6 μg, 5 μg and 50 μg showed a significant, dose-dependent increase in cell proliferation and migration compared to the control. In addition, osteogenic differentiation assays demonstrated that exosome concentration differently affected the ability of MSC to deposit mineralised matrix. Finally, the analysis of exosome protein content revealed a higher amount of basic fibroblast growth factor (bFGF, vascular endothelial growth factor (VEGF, platelet-derived growth factor (PDGF-BB and transforming growth factor beta 1 (TGF-β1 as compared to PL. In regards to RNA content, an enrichment of small RNAs in exosomes as compared to donor platelets has been found. These results suggest that exosomes consistently contribute to PL activity and could represent an advantageous nanodelivery system for cell-free regeneration therapies.

  8. The role of exosomes in tumor progression and metastasis (Review).

    Science.gov (United States)

    Suchorska, Wiktoria M; Lach, Michal S

    2016-03-01

    Tumor cells have developed various mechanisms in defense against applied treatment, which prevent their total elimination from an organism. One of the underestimated mechanisms of defense is secretion of highly specialized double-membrane structures called exosomes. They play a crucial role in the control of the local microenvironment and intracellular communication. It has been shown that the exosomes can be carriers of various proteins, lipids, miRNAs and mRNAs. There are extensive data concerning the influence and participation by exosomes in metastasis and cancer progression. It has been demonstrated that exosomes are involved in multidrug resistance mechanisms, radiation-induced bystander effect and epithelial-mesenchymal transition. Furthermore, exosomes are able to form a premetastatic niche and enable the escape of cancer cells from recognition by host immune cells. Moreover, exosomes are responsible for the formation of vessels. This indicates the significance of secreted extracellular vesicles in the development and prognosis of cancer. The aim of the present review is to briefly describe the role of exosomes in tumor biology.

  9. Exosomes derived from renal cancer cells induce Jurkat T cell apoptosis in vitro%肾癌细胞来源的exosomes诱导Jurkat T细胞凋亡

    Institute of Scientific and Technical Information of China (English)

    杨林; 吴小候; 罗春丽; 王丹; 陈力学

    2013-01-01

    Objective To investigate the underlying mechanism of exosomes derived from renal cancer cell lines 786-0 to mediate tumor immune escape in vitro. Methods CCK-8 assay was used to determine the effects of exosomes on proliferation in Jurkat T cells. Morphological changes were by wright-giemsa staining;flow cytometry with Annexin V-FITC/PI double staining was used to detect the apoptosis; secretion functions of Jurkat T cell were detected by ELISA assay; effects of exosomes on apoptosis of Jurkat T cell were detected by soluble Fas block experiment; effects on the protein expression of FasL, caspase, Bax and Bcl-2 were assessed by Western blot analysis. Results Exosomes could inhibit Jurkat T cell proliferation, 10 μg/mL exosomes act on Jurkat T cell for 24 and 72 h, growth inhibition rate was (19. 64 ±0. 92)% and (36. 24 ± 1. 12)% ; while 400 μg/mL exosomes act on it for 24 h and 72 h, growth inhibition rate was (55.96 ± 1.35)% and (76.51 ± 1. 37)% respectively. Exosomes could induce Jurkat T cell apoptosis, 10 μg/mL exosomes act on Jurkat T cell for 8 h, apoptosis rate was (7. 31 ±1.32)% , extending this monitoring to 24 h, apoptosis rate was (20. 19 ± 1.47)% ; while 400μg/mL exosomes act on it for 8 and 24 h, apoptosis rate was (27. 28 ± 1. 29)% and (41.72 ±0.88)% respectively. Exosomes also suppressed IL-2, IFN-γ, IL-6 and IL-10 secretion obviously. FasL was highly expressed in exosomes, soluble Fas block could reverse Jurkat T cell apoptosis. In this course, caspase-3 , caspase-8, caspase-9 were activated, and the ratio of Bax/Bcl-2 increased. Conclusion Exosomes could inhibit the growth of Jurkat T cell and induce apoptosis. It could mediate tumor immune escape.%目的 体外研究肾癌786-0细胞来源的exosomes介导肿瘤免疫逃逸的机制.方法 采用CCK-8法检测肾癌786-0细胞来源的exosomes对Jurkat T细胞生长的影响,瑞氏-姬姆萨染色检测Jurkat T细胞形态变化,Annexin V-FITC/PI双染色流式细胞术检测Jurkat T

  10. Identification of Hardware Trojans triggering signals

    OpenAIRE

    Dupuis, Sophie; Di Natale, Giorgio; Flottes, Marie-Lise; Rouzeyre, Bruno

    2013-01-01

    International audience; Hardware Trojans are malicious alterations to a circuit. These modifications can be inserted either during the design phase or during the fabrication process. Due to the diversity of Hardware Trojans (HTs), detecting and/or locating them are challenging tasks. Numerous approaches have been proposed to address this problem. Methods based on logic testing consist in trying to activate potential Hardware Trojans in order to detect erroneous outputs during simulation. Howe...

  11. The Complex History of Trojan Asteroids

    Science.gov (United States)

    Emery, J. P.; Marzari, F.; Morbidelli, A.; French, L. M.; Grav, T.

    The Trojan asteroids, orbiting the Sun in Jupiter's stable Lagrange points, provide a unique perspective on the history of our solar system. As a large population of small bodies, they record important gravitational interactions in the dynamical evolution of the solar system. As primitive bodies, their compositions and physical properties provide windows into the conditions in the solar nebula in the region in which they formed. In the past decade, significant advances have been made in understanding their physical properties, and there has been a revolution in thinking about the origin of Trojans. The ice and organics generally presumed to be a significant part of Trojan composition have yet to be detected directly, although the low density of the binary system Patroclus (and possibly low density of the binary/moonlet system Hektor) is consistent with an interior ice component. By contrast, fine-grained silicates that appear to be similar to cometary silicates in composition have been detected, and a color bimodality may indicate distinct compositional groups among the Trojans. Whereas Trojans had traditionally been thought to have formed near 5 AU, a new paradigm has developed in which the Trojans formed in the proto-Kuiper belt, and were scattered inward and captured in the Trojan swarms as a result of resonant interactions of the giant planets. Whereas the orbital and population distributions of current Trojans are consistent with this origin scenario, there are significant differences between current physical properties of Trojans and those of Kuiper belt objects. These differences may be indicative of surface modification due to the inward migration of objects that became the Trojans, but understanding of appropriate modification mechanisms is poor and would benefit from additional laboratory studies. Many open questions about this intriguing population remain, and the future promises significant strides in our understanding of Trojans. The time is ripe for a

  12. Coronary Artery-Bypass-Graft Surgery Increases the Plasma Concentration of Exosomes Carrying a Cargo of Cardiac MicroRNAs: An Example of Exosome Trafficking Out of the Human Heart with Potential for Cardiac Biomarker Discovery.

    Directory of Open Access Journals (Sweden)

    Costanza Emanueli

    Full Text Available Exosome nanoparticles carry a composite cargo, including microRNAs (miRs. Cultured cardiovascular cells release miR-containing exosomes. The exosomal trafficking of miRNAs from the heart is largely unexplored. Working on clinical samples from coronary-artery by-pass graft (CABG surgery, we investigated if: 1 exosomes containing cardiac miRs and hence putatively released by cardiac cells increase in the circulation after surgery; 2 circulating exosomes and exosomal cardiac miRs correlate with cardiac troponin (cTn, the current "gold standard" surrogate biomarker of myocardial damage.The concentration of exosome-sized nanoparticles was determined in serial plasma samples. Cardiac-expressed (miR-1, miR-24, miR-133a/b, miR-208a/b, miR-210, non-cardiovascular (miR-122 and quality control miRs were measured in whole plasma and in plasma exosomes. Linear regression analyses were employed to establish the extent to which the circulating individual miRs, exosomes and exosomal cardiac miR correlated with cTn-I. Cardiac-expressed miRs and the nanoparticle number increased in the plasma on completion of surgery for up to 48 hours. The exosomal concentration of cardiac miRs also increased after CABG. Cardiac miRs in the whole plasma did not correlate significantly with cTn-I. By contrast cTn-I was positively correlated with the plasma exosome level and the exosomal cardiac miRs.The plasma concentrations of exosomes and their cargo of cardiac miRs increased in patients undergoing CABG and were positively correlated with hs-cTnI. These data provide evidence that CABG induces the trafficking of exosomes from the heart to the peripheral circulation. Future studies are necessary to investigate the potential of circulating exosomes as clinical biomarkers in cardiac patients.

  13. Hardware Trojan by Hot Carrier Injection

    CERN Document Server

    Shiyanovskii, Y; Papachristou, C; Weyer, D; Clay, W

    2009-01-01

    This paper discusses how hot carrier injection (HCI) can be exploited to create a trojan that will cause hardware failures. The trojan is produced not via additional logic circuitry but by controlled scenarios that maximize and accelerate the HCI effect in transistors. These scenarios range from manipulating the manufacturing process to varying the internal voltage distribution. This new type of trojan is difficult to test due to its gradual hardware degradation mechanism. This paper describes the HCI effect, detection techniques and discusses the possibility for maliciously induced HCI trojans.

  14. Exploiting Semiconductor Properties for Hardware Trojans

    CERN Document Server

    Shiyanovskii, Y; Papachristou, C; Weyer, D; Clay, W

    2009-01-01

    This paper discusses the possible introduction of hidden reliability defects during CMOS foundry fabrication processes that may lead to accelerated wearout of the devices. These hidden defects or hardware Trojans can be created by deviation from foundry design rules and processing parameters. The Trojans are produced by exploiting time-based wearing mechanisms (HCI, NBTI, TDDB and EM) and/or condition-based triggers (ESD, Latchup and Softerror). This class of latent damage is difficult to test due to its gradual degradation nature. The paper describes life-time expectancy results for various Trojan induced scenarios. Semiconductor properties, processing and design parameters critical for device reliability and Trojan creation are discussed.

  15. Proteomics Characterization of Exosome Cargo

    OpenAIRE

    Schey, Kevin L.; Luther, J. Matthew; Rose, Kristie L

    2015-01-01

    Characterization of exosomal cargo is of significant interest because this cargo can provide clues to exosome biogenesis, targeting, and cellular effects and may be a source of biomarkers for disease diagnosis, prognosis and response to treatment. With recent improvements in proteomics technologies, both qualitative and quantitative characterization of exosomal proteins is possible. Here we provide a brief review of exosome proteomics studies and provide detailed protocols for global qualitat...

  16. From structures to functions: insights into exosomes as promising drug delivery vehicles.

    Science.gov (United States)

    Ren, Jinghua; He, Wenshan; Zheng, Lifen; Duan, Hongwei

    2016-05-24

    Exosomes are small membrane vesicles secreted by most cell types, and appear ubiquitously in cell culture supernatants and body fluids. Increasing evidence supports that exosomes play important roles in intercellular communication, both locally and systemically, by transporting their contents such as proteins, lipids and RNAs between cells. Of particular interest for controlled drug delivery is that cell-derived exosomes offer the possibilities of overcoming biological barriers, thereby allowing the incorporated gene and drug to reach targeted tissue, which have been considerable challenges for synthetic carriers. Great research efforts have been dedicated to developing exosome-based drug delivery systems for the treatment of inflammatory diseases, degenerative disorders and cancer. In this review, we will describe the structural and functional properties of exosomes and emphasize current advances in the therapeutic applications of exosomes as drug delivery vehicles, followed by a discussion on current challenges and future perspectives.

  17. Comparative analysis of discrete exosome fractions obtained by differential centrifugation

    DEFF Research Database (Denmark)

    Jeppesen, Dennis Kjølhede; Hvam, Michael L; Primdahl-Bengtson, Bjarke

    2014-01-01

    BACKGROUND: Cells release a mixture of extracellular vesicles, amongst these exosomes, that differ in size, density and composition. The standard isolation method for exosomes is centrifugation of fluid samples, typically at 100,000×g or above. Knowledge of the effect of discrete...... ultracentrifugation speeds on the purification from different cell types, however, is limited. METHODS: We examined the effect of applying differential centrifugation g-forces ranging from 33,000×g to 200,000×g on exosome yield and purity, using 2 unrelated human cell lines, embryonic kidney HEK293 cells and bladder...... of phenol red and cleared by 200,000×g overnight centrifugation. The centrifugation tube fill level impacted the sedimentation efficacy. Comparative analysis by NTA, protein quantification, and detection of exosomal and contamination markers identified differences in vesicle size, concentration...

  18. Tumor-derived exosomes in oncogenic reprogramming and cancer progression.

    Science.gov (United States)

    Saleem, Sarmad N; Abdel-Mageed, Asim B

    2015-01-01

    In multicellular organisms, effective communication between cells is a crucial part of cellular and tissue homeostasis. This communication mainly involves direct cell-cell contact as well as the secretion of molecules that bind to receptors at the recipient cells. However, a more recently characterized mode of intercellular communication-the release of membrane vesicles known as exosomes-has been the subject of increasing interest and intensive research over the past decade. Following the discovery of the exosome-mediated immune activation, the pathophysiological roles of exosomes have been recognized in different diseases, including cancer. In this review, we describe the biogenesis and main physical characteristics that define exosomes as a specific population of secreted vesicles, with a special focus on their role in oncogenic transformation and cancer progression.

  19. A Comprehensive Review on Exosomes and Microvesicles as Epigenetic Factors.

    Science.gov (United States)

    Bakhshandeh, Behnaz; Kamaleddin, Mohammad Amin; Aalishah, Khadijeh

    2017-01-01

    Exosomes and microvesicles, which are released by most of the cells, play important roles in intracellular correspondence by transferring DNA, messenger RNA, micro RNA, and other types of RNA and proteins. Exosomes and microvesicles may contribute to the distribution of cancers and diseases through delivering the pathogenic agents to the non-infected cells; in cancers, they can modify the cells in the tumor niche and lead them to transformation. In addition, these vesicles can affect stem cell activity and their physiological properties. On the other hand, exosomes and microvesicles can be applied in the therapeutic strategies as they are small, non-viral, flexible and able to cross biological barriers. In this review, we focused on some details about the exosomes and microvesicles both functionally and structurally.

  20. Influence of maternal BMI on the exosomal profile during gestation and their role on maternal systemic inflammation.

    Science.gov (United States)

    Elfeky, Omar; Longo, Sherri; Lai, Andrew; Rice, Gregory E; Salomon, Carlos

    2017-02-01

    Recent studies report that 35% of women are either overweight or obese at reproductive age. The placenta continuously releases exosomes across gestation and their concentration is higher in pregnancy complications. While there is considerable interest in elucidating the role of exosomes during gestation, important questions remain to be answered: i) Does maternal BMI affect the exosomal profile across gestation? and ii) What is the contribution of placenta-derived exosomes to the total number of exosomes present in maternal plasma across gestation? Plasma samples were classified according to the maternal BMI into three groups (n = 15 per group): Lean, overweight, and obese. Total exosomes and specific placenta-derived exosomes were determined by Nanoparticle Tracking Analysis (NanoSight™) using quantum dots coupled with CD63 or PLAP antibodies. The effect of exosomes on cytokine (IL-6, IL-8, IL-10 and TNF-α) release from endothelial cells was established by cytokine array analysis (Bioplex-200). The total number of exosomes present in maternal circulation was strongly correlated with maternal BMI. Between ∼12% and ∼25% of circulating exosomes in maternal blood are of placental origin during gestation, and the contribution of placental exosomes to the total exosomal population decreases with higher maternal BMI across gestation. Exosomes increase IL-6, IL-8 and TNF-α release from endothelial cells, an effect even higher when exosomes were isolated from obese women compared to lean and overweight. This study established that maternal BMI is a factor that explains a significant component of the variation in the exosomes data. Exosomes may contribute to the maternal systemic inflammation during pregnancy.

  1. Role of tyrosine kinase Src in gastric cancer exosome mediated promotion of tumor cell proliferation%Src激酶在胃癌细胞来源的exosome促进肿瘤细胞增殖中的作用

    Institute of Scientific and Technical Information of China (English)

    曲晶磊; 曲秀娟; 刘云鹏; 赵明芳; 侯科佐; 姜又红; 杨向红

    2011-01-01

    目的:研究胃癌细胞来源的外泌体(exosome)对肿瘤细胞增殖的影响,初步探讨Src蛋白激酶在此过程中的作用.方法:采用离心超滤和蔗糖密度梯度超速离心的方法从胃癌SGC7901细胞的上清液中分离出胃癌细胞来源的exosome.透射电子显微镜下观察exosome形态.MTT法检测细胞增殖能力,Western blot检测蛋白的表达.结果:透射电子显微镜下观察胃癌SGC7901细胞来源的exosome具有特征性的盘状结构.由双层膜构成,他们的直径30-1 00 nm.Westernblot结果显示exosome表面富含CD9和TSG101分子.MTT结果显示exosome能以时间和剂量依赖性的方式促进SGC7901细胞的增殖,200 mg/L和400 mg/L的exosome处理SGC7901细胞72 h,细胞的增殖比率分别是对照组的138%(P<0.001)和144%(P<0.001),在此过程中伴随有p-Src表达的上调.结论:胃癌细胞来源的exosome能促进肿瘤细胞的增殖,其机制可能与激活Src蛋白激酶有关.%AIM: To investigate the effect of gastric cancer exosomes on tumor cell proliferation and to evaluate the role of tyrosine kinase Src in this process.METHODS: Exosomes were isolated and purified from gastric cancer SGC7901 cells by serial centrifugation and sucrose gradient ultracentrifugation and observed by electron microscopy.Cell proliferation was measured by MTT assay.Protein expression was assayed by Western blot.RESULTS: Gastric cancer exosomes had a characteristic saucer-like shape that was limited by a lipid bilayer, and their diameter ranged from 30 to 100 nm.CD9 and TSG101 were abundant on the surface of exosomes.Gastric cancer exosomes significantly increased SGC7901 cell proliferation in a time- and dose-dependent manner.Compared with control cells, the proliferation of cells treated with 200 and 400 mg/L exosomes for 72 h were increased to 138% and 144%, respectively (both P < 0.01).The expression of phosphorylated Src in SGC7901 cells was up-regulated in a time- and dose-dependent manner after

  2. A strategy of antigen incorporation into exosomes: comparing cross-presentation levels of antigens delivered by engineered exosomes and by lentiviral virus-like particles.

    Science.gov (United States)

    Lattanzi, Laura; Federico, Maurizio

    2012-11-26

    Among strategies aimed at developing new nanoparticle-based vaccines, exosomes hold much promise. They are nanovesicles released by basically all eukaryotic cell types originating from intraluminal vesicles which accumulate in multivesicular bodies. Exosomes have immunogenic properties whose strength correlates with the amounts of associated antigens. Engineering antigens to target them in exosomes represents the last frontier in terms of nanoparticle-based vaccines. Here we report a new method to incorporate protein antigens in exosomes relying on the unique properties of a mutant of the HIV-1 Nef protein, Nef(mut). This is a biologically inactive mutant we found incorporating into exosomes at high levels also when fused at its C-terminus with foreign proteins. We compared both biochemical and antigenic properties of Nef(mut) exosomes with those of previously characterized Nef(mut) -based lentiviral virus-like particles (VLPs). We found that exosomes incorporate Nef(mut) and fusion protein derivatives with similar efficiency of VLPs. When an envelope fusion protein was associated with both exosomes and VLPs to favor cross-presentation of associated antigens, Nef(mut) and its derivatives incorporated in exosomes were cross-presented at levels at least similar to what observed when the antigens were delivered by engineered VLPs. This occurred despite exosomes entered target cells with an apparent lower efficiency than VLPs. The unique properties of HIV-1 Nef(mut) in terms of exosome incorporation efficiency, carrier of foreign antigens, and lack of anti-cellular effects open the way toward the development of a flexible, safe, cost-effective exosome-based CD8(+) T cell vaccine platform.

  3. Comparative analysis of discrete exosome fractions obtained by differential centrifugation

    Directory of Open Access Journals (Sweden)

    Dennis K. Jeppesen

    2014-11-01

    Full Text Available Background: Cells release a mixture of extracellular vesicles, amongst these exosomes, that differ in size, density and composition. The standard isolation method for exosomes is centrifugation of fluid samples, typically at 100,000×g or above. Knowledge of the effect of discrete ultracentrifugation speeds on the purification from different cell types, however, is limited. Methods: We examined the effect of applying differential centrifugation g-forces ranging from 33,000×g to 200,000×g on exosome yield and purity, using 2 unrelated human cell lines, embryonic kidney HEK293 cells and bladder carcinoma FL3 cells. The fractions were evaluated by nanoparticle tracking analysis (NTA, total protein quantification and immunoblotting for CD81, TSG101, syntenin, VDAC1 and calreticulin. Results: NTA revealed the lowest background particle count in Dulbecco's Modified Eagle's Medium media devoid of phenol red and cleared by 200,000×g overnight centrifugation. The centrifugation tube fill level impacted the sedimentation efficacy. Comparative analysis by NTA, protein quantification, and detection of exosomal and contamination markers identified differences in vesicle size, concentration and composition of the obtained fractions. In addition, HEK293 and FL3 vesicles displayed marked differences in sedimentation characteristics. Exosomes were pelleted already at 33,000×g, a g-force which also removed most contaminating microsomes. Optimal vesicle-to-protein yield was obtained at 67,000×g for HEK293 cells but 100,000×g for FL3 cells. Relative expression of exosomal markers (TSG101, CD81, syntenin suggested presence of exosome subpopulations with variable sedimentation characteristics. Conclusions: Specific g-force/k factor usage during differential centrifugation greatly influences the purity and yield of exosomes. The vesicle sedimentation profile differed between the 2 cell lines.

  4. Exosome Biogenesis, Regulation, and Function in Viral Infection

    OpenAIRE

    Marta Alenquer; Maria João Amorim

    2015-01-01

    Exosomes are extracellular vesicles released upon fusion of multivesicular bodies(MVBs) with the cellular plasma membrane. They originate as intraluminal vesicles (ILVs) duringthe process of MVB formation. Exosomes were shown to contain selectively sorted functionalproteins, lipids, and RNAs, mediating cell-to-cell communications and hence playing a role in thephysiology of the healthy and diseased organism. Challenges in the field include the identificationof mechanisms sustaining packaging ...

  5. Exosomes in Prostate Cancer: Putting Together the Pieces of a Puzzle

    Energy Technology Data Exchange (ETDEWEB)

    Soekmadji, Carolina, E-mail: carolina.soekmadji@qut.edu.au; Russell, Pamela J.; Nelson, Colleen C. [Australian Prostate Cancer Research Centre-Queensland, Institute of Health and Biomedical Innovation, Queensland University of Technology, Translational Research Institute, Level 3 West, 37 Kent Street, Brisbane, Queensland 4102 (Australia)

    2013-11-11

    Exosomes have been shown to act as mediators for cell to cell communication and as a potential source of biomarkers for many diseases, including prostate cancer. Exosomes are nanosized vesicles secreted by cells and consist of proteins normally found in multivesicular bodies, RNA, DNA and lipids. As a potential source of biomarkers, exosomes have attracted considerable attention, as their protein content resembles that of their cells of origin, even though it is noted that the proteins, miRNAs and lipids found in the exosomes are not a reflective stoichiometric sampling of the contents from the parent cells. While the biogenesis of exosomes in dendritic cells and platelets has been extensively characterized, much less is known about the biogenesis of exosomes in cancer cells. An understanding of the processes involved in prostate cancer will help to further elucidate the role of exosomes and other extracellular vesicles in prostate cancer progression and metastasis. There are few methodologies available for general isolation of exosomes, however validation of those methodologies is necessary to study the role of exosomal-derived biomarkers in various diseases. In this review, we discuss “exosomes” as a member of the family of extracellular vesicles and their potential to provide candidate biomarkers for prostate cancer.

  6. Exosomes in Prostate Cancer: Putting Together the Pieces of a Puzzle

    Directory of Open Access Journals (Sweden)

    Colleen C. Nelson

    2013-11-01

    Full Text Available Exosomes have been shown to act as mediators for cell to cell communication and as a potential source of biomarkers for many diseases, including prostate cancer. Exosomes are nanosized vesicles secreted by cells and consist of proteins normally found in multivesicular bodies, RNA, DNA and lipids. As a potential source of biomarkers, exosomes have attracted considerable attention, as their protein content resembles that of their cells of origin, even though it is noted that the proteins, miRNAs and lipids found in the exosomes are not a reflective stoichiometric sampling of the contents from the parent cells. While the biogenesis of exosomes in dendritic cells and platelets has been extensively characterized, much less is known about the biogenesis of exosomes in cancer cells. An understanding of the processes involved in prostate cancer will help to further elucidate the role of exosomes and other extracellular vesicles in prostate cancer progression and metastasis. There are few methodologies available for general isolation of exosomes, however validation of those methodologies is necessary to study the role of exosomal-derived biomarkers in various diseases. In this review, we discuss “exosomes” as a member of the family of extracellular vesicles and their potential to provide candidate biomarkers for prostate cancer.

  7. The limited capacity of malignant glioma-derived exosomes to suppress peripheral immune effectors.

    Science.gov (United States)

    Iorgulescu, J Bryan; Ivan, Michael E; Safaee, Michael; Parsa, Andrew T

    2016-01-15

    Tumor-derived microvesicular exosomes permit intercellular communication both locally and systemically by delivering a snapshot of the tumor cell's constituents. We thus investigated whether exosomes mediate malignant glioma's facility for inducing peripheral immunosuppression. In Western blot and RT-PCR analyses, glioma-derived exosomes displayed exosome-specific markers, but failed to recapitulate the antigen-presentation machinery, surface co-modulatory signals, or immunosuppressive mediator status of their parent tumor cells. Treatment with glioma-derived exosomes promoted immunosuppressive HLA-DR(low) monocytic phenotypes, but failed to induce monocytic PD-L1 expression or alter the activation of cytotoxic T-cells from patients' peripheral blood by FACS and RT-PCR analyses. Our results suggest that malignant glioma-derived exosomes are restricted in their capacity to directly prime peripheral immunosuppression.

  8. Role of Exosomal Noncoding RNAs in Lung Carcinogenesis

    Directory of Open Access Journals (Sweden)

    Tao Sun

    2015-01-01

    Full Text Available Lung cancer is the major cause of cancer death worldwide. Novel, recently discovered classes of noncoding RNAs (ncRNAs have diverse functional and regulatory activities and increasing evidence suggests crucial roles for deregulated ncRNAs in the onset and progression of cancer, including lung cancer. Exosomes are small extracellular membrane vesicles of endocytic origin that are released by many cells and are found in most body fluids. Tumor-derived exosomes mediate tumorigenesis by facilitating tumor growth and metastasis. MicroRNAs (miRNAs are a subclass of ncRNAs that are present in exosomes. miRNAs are taken up by neighboring or distant cells and modulate various functions of recipient cells. Here, we review exosome-derived ncRNAs with a focus on miRNAs and their role in lung cancer biology.

  9. Anti-tumor effect induced by exosomes derived from dendritic cells loaded with lung cancer cell lysates%肺癌细胞裂解物负载对树突状细胞分泌的exosome诱导抗肿瘤作用的影响

    Institute of Scientific and Technical Information of China (English)

    张在云; 李希德; 刘叶; 王志仑; 潘祥林

    2011-01-01

    目的 为制备高效的胞外体(exosome)肿瘤疫苗提供理论依据.方法 用细胞因子诱导培养树突状细胞(DC),将肺癌细胞裂解物负载DC,提取exosome;用exosome活化T细胞(负载组),以未负载DC的exosome(未负载组)及肺癌细胞裂解物负载DC(DC组)活化的T细胞为对照,MTT法检测三组肺癌细胞的杀伤率.结果 exosome中有HSP70、HLA及CEA表达.活化T细胞/肺癌细胞为25∶1、10∶1 、5∶1时负载组杀伤率均明显高于未负载组及DC组(P均<0.05).结论 肺癌细胞裂解物负载能增强DC分泌的exosome诱导的抗肿瘤作用;本研究为制备高效的exosome肿瘤疫苗提供了理论依据.%Objective To obtain theoretical bases for making high efficacy exosome cancer vaccine. Methods Dendritic cells (DC) were induced with cytokines and then loaded with whole lung cancer cell lysates. Exosomes were isolated from supernatant of DC, and T cells activated by the exosomes (group loaded) , T cells activated by exosomes from nonloaded DC (group non-loaded) or activated by lysate-loaded DC(group DC) were taken as control. 11k activity of T cells for killing lung cancer cells were detected by MTT method. Results HSP70, HLA and CEA protein were found in exosomes. The kill rates of activated T cells in group loaded at E: T ratio 25:1, 10= 1, 5:1 were much higher than those in group non-loaded and group DC( all P <0.05). Condnsiong Lung cancer cell lysates loading can promote the anti-tumor activity induced by DC-derived exosomes; this study can provide theoretical bases for making high efficacy exosome cancer vaccine.

  10. The experimental study of effect of exosomes derived from breast cancer cell line on human umbilical vein endothelial cells%乳腺癌细胞exosomes对人脐静脉内皮作用的实验研究

    Institute of Scientific and Technical Information of China (English)

    谢莹珊; 沈宜; 隆霜; 孙迪; 姜蓉; 陈黎

    2011-01-01

    Objective:To observe the effect of exosomes derived from MDA- MB- 231 cell line on proliferation, immigration and capillary- like tube formation of Human Umbilical Vein Endothelial cells. Method: Exosomes were puried by serial ultracentrifugation and sugar density ultracentrifugation, MTT assay was used to observe the effect of exosomes on proliferation of HUVECs;HUVECs were treated with exosomes for 24 h,the change of cell migration was detected by Transwell chamber method. The capillary-like tube formations by HUVECs were observed. Result: MTT result showed that the concentration range of exosomes significantly increased HUVECs proliferation in a concentration - and time - dependent manner ( P < 0.01 ); It may significantly enhance the migration of HUVECs after treated with 200 μ/ml exosomes for 24 h ( P < 0.01 ) , and may significantly promoted the capability of capillary - like tube formation of HUVECs ( P < 0.05 ) Conclusion: Exosomes derived from MDA - MB - 231 cell line significantly increased HUVECs proliferation in a concentration - and time - dependent manner and it may promote the migration and the capability of capillary - like tube formation of HUVECs.%目的:观察人乳腺癌细胞株MDA-MB-231细胞源exosomes对人脐静脉内皮细胞株(HUVECs)增殖、迁移能力及血管样结构形成的影响.方法:超速离心及密度梯度离心法提取exosomes;MTT法检测MDA-MB-231细胞源exosomes对HUVECs增殖的影响;Transwell小室法检测HUVECs与exosomes混合培养24h后迁移能力的影响;观察HUVECs与exosomes混合培养24h后管腔样结构形成变化.结果:各浓度exosomes均具有促进HUVECs细胞增殖作用,且以时间剂量依赖性促进HUVEC细胞增殖(P

  11. Relevance of circulating tumor cells, extracellular nucleic acids, and exosomes in breast cancer

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    Friel, Anne M.; Corcoran, Claire; Crown, John; O'Driscoll, Lorraine

    2010-01-01

    Abstract Early detection of cancer is vital to improved overall survival rates. At present, evidence is accumulating for the clinical value of detecting occult tumor cells in peripheral blood, plasma, and serum specimens from cancer patients. Both molecular and cellular approaches, which differ in sensitivity and specificity, have been used for such means. Circulating tumor cells and extracellular nucleic acids have been detected within blood, plasma, and sera of cancer patients. A...

  12. Identification and proteomic analysis of osteoblast-derived exosomes.

    Science.gov (United States)

    Ge, Min; Ke, Ronghu; Cai, Tianyi; Yang, Junyi; Mu, Xiongzheng

    2015-11-01

    Exosomes are nanometer-sized vesicles with the function of intercellular communication, and they are released by various cell types. To reveal the knowledge about the exosomes from osteoblast, and explore the potential functions of osteogenesis, we isolated microvesicles from supernatants of mouse Mc3t3 by ultracentrifugation, characterized exosomes by electron microscopy and immunoblotting and presented the protein profile by proteomic analysis. The result demonstrated that microvesicles were between 30 and 100 nm in diameter, round shape with cup-like concavity and expressed exosomal marker tumor susceptibility gene (TSG) 101 and flotillin (Flot) 1. We identified a total number of 1069 proteins among which 786 proteins overlap with ExoCarta database. Gene Oncology analysis indicated that exosomes mostly derived from plasma membrane and mainly involved in protein localization and intracellular signaling. The Ingenuity Pathway Analysis showed pathways are mostly involved in exosome biogenesis, formation, uptake and osteogenesis. Among the pathways, eukaryotic initiation factor 2 pathways played an important role in osteogenesis. Our study identified osteoblast-derived exosomes, unveiled the content of them, presented potential osteogenesis-related proteins and pathways and provided a rich proteomics data resource that will be valuable for further studies of the functions of individual proteins in bone diseases.

  13. Exosomal and Non-Exosomal Transport of Extra-Cellular microRNAs in Follicular Fluid: Implications for Bovine Oocyte Developmental Competence.

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    Md Mahmodul Hasan Sohel

    Full Text Available Cell-cell communication within the follicle involves many signaling molecules, and this process may be mediated by secretion and uptake of exosomes that contain several bioactive molecules including extra-cellular miRNAs. Follicular fluid and cells from individual follicles of cattle were grouped based on Brilliant Cresyl Blue (BCB staining of the corresponding oocytes. Both Exoquick precipitation and differential ultracentrifugation were used to separate the exosome and non-exosomal fraction of follicular fluid. Following miRNA isolation from both fractions, the human miRCURY LNA™ Universal RT miRNA PCR array system was used to profile miRNA expression. This analysis found that miRNAs were present in both exosomal and non-exosomal fraction of bovine follicular fluid. We found 25 miRNAs differentially expressed (16 up and 9 down in exosomes and 30 miRNAs differentially expressed (21 up and 9 down in non-exosomal fraction of follicular fluid in comparison of BCB- versus BCB+ oocyte groups. Expression of selected miRNAs was detected in theca, granulosa and cumulus oocyte complex. To further explore the potential roles of these follicular fluid derived extra-cellular miRNAs, the potential target genes were predicted, and functional annotation and pathway analysis revealed most of these pathways are known regulators of follicular development and oocyte growth. In order to validate exosome mediated cell-cell communication within follicular microenvironment, we demonstrated uptake of exosomes and resulting increase of endogenous miRNA level and subsequent alteration of mRNA levels in follicular cells in vitro. This study demonstrates for the first time, the presence of exosome or non-exosome mediated transfer of miRNA in the bovine follicular fluid, and oocyte growth dependent variation in extra-cellular miRNA signatures in the follicular environment.

  14. Exosomal and Non-Exosomal Transport of Extra-Cellular microRNAs in Follicular Fluid: Implications for Bovine Oocyte Developmental Competence.

    Science.gov (United States)

    Sohel, Md Mahmodul Hasan; Hoelker, Michael; Noferesti, Sina Seifi; Salilew-Wondim, Dessie; Tholen, Ernst; Looft, Christian; Rings, Franca; Uddin, Muhammad Jasim; Spencer, Thomas E; Schellander, Karl; Tesfaye, Dawit

    2013-01-01

    Cell-cell communication within the follicle involves many signaling molecules, and this process may be mediated by secretion and uptake of exosomes that contain several bioactive molecules including extra-cellular miRNAs. Follicular fluid and cells from individual follicles of cattle were grouped based on Brilliant Cresyl Blue (BCB) staining of the corresponding oocytes. Both Exoquick precipitation and differential ultracentrifugation were used to separate the exosome and non-exosomal fraction of follicular fluid. Following miRNA isolation from both fractions, the human miRCURY LNA™ Universal RT miRNA PCR array system was used to profile miRNA expression. This analysis found that miRNAs were present in both exosomal and non-exosomal fraction of bovine follicular fluid. We found 25 miRNAs differentially expressed (16 up and 9 down) in exosomes and 30 miRNAs differentially expressed (21 up and 9 down) in non-exosomal fraction of follicular fluid in comparison of BCB- versus BCB+ oocyte groups. Expression of selected miRNAs was detected in theca, granulosa and cumulus oocyte complex. To further explore the potential roles of these follicular fluid derived extra-cellular miRNAs, the potential target genes were predicted, and functional annotation and pathway analysis revealed most of these pathways are known regulators of follicular development and oocyte growth. In order to validate exosome mediated cell-cell communication within follicular microenvironment, we demonstrated uptake of exosomes and resulting increase of endogenous miRNA level and subsequent alteration of mRNA levels in follicular cells in vitro. This study demonstrates for the first time, the presence of exosome or non-exosome mediated transfer of miRNA in the bovine follicular fluid, and oocyte growth dependent variation in extra-cellular miRNA signatures in the follicular environment.

  15. Exosomal transfer of functional small RNAs mediates cancer-stroma communication in human endometrium.

    Science.gov (United States)

    Maida, Yoshiko; Takakura, Masahiro; Nishiuchi, Takumi; Yoshimoto, Tanihiro; Kyo, Satoru

    2016-02-01

    Exosomes are small membrane vesicles secreted from a variety of cell types. Recent evidence indicates that human cells communicate with each other by exchanging exosomes. Cancer cells closely interact with neighboring stromal cells, and together they cooperatively promote disease via bidirectional communication. Here, we investigated whether exosomes can play roles in intercellular communication between cancer cells and neighboring fibroblasts. Endometrial fibroblasts were isolated from normal endometrial tissues and from endometrial cancer tissues, and cell-to-cell transfer of endometrial cancer cell line Ishikawa-derived exosomes was examined. The isolated fibroblasts were cultured in conditioned media from CD63-GFP-expressing Ishikawa cells, and we found that GFP-positive exosomes were transferred from Ishikawa cells to the fibroblasts. Next, we introduced a shRNA for a luciferase gene into Ishikawa cells. This shRNA was encapsulated into exosomes, was transferred to the fibroblasts, and then downregulated luciferase expression in the fibroblasts. The mature microRNAs naturally expressed in Ishikawa-derived exosomes were also transported into the endometrial fibroblasts, and they altered the microRNA expression profiles of the fibroblasts. These results indicated that endometrial cancer cells could transmit small regulatory RNAs to endometrial fibroblasts via exosomes. Our findings document a previously unknown mode of intercellular communication between cancer cells and related fibroblasts in human endometrium.

  16. Proteomics characterization of exosome cargo.

    Science.gov (United States)

    Schey, Kevin L; Luther, J Matthew; Rose, Kristie L

    2015-10-01

    Characterization of exosomal cargo is of significant interest because this cargo can provide clues to exosome biogenesis, targeting, and cellular effects and may be a source of biomarkers for disease diagnosis, prognosis and response to treatment. With recent improvements in proteomics technologies, both qualitative and quantitative characterization of exosomal proteins is possible. Here we provide a brief review of exosome proteomics studies and provide detailed protocols for global qualitative, global quantitative, and targeted quantitative analysis of exosomal proteins. In addition, we provide an example application of a standard global quantitative analysis followed by validation via a targeted quantitative analysis of urine exosome samples from human patients. Advantages and limitations of each method are discussed as well as future directions for exosome proteomics analysis.

  17. The Influence of Ionizing Radiation on Exosome Composition, Secretion and Intercellular Communication.

    Science.gov (United States)

    Jelonek, Karol; Widlak, Piotr; Pietrowska, Monika

    2016-01-01

    A large variety of vesicles is actively secreted into the extracellular space by most type of cells. The smallest nanoparticles (30-120 nm), called exosomes, are known to transport their cargo (nucleic acids, proteins and lipids) between diverse locations in the body. Specific content of exosomes and their influence on recipient cells depends primarily on the type of the secretory (donor) cell, yet several studies highlight the importance of environmental stress on which the donor cells are exposed. Ionizing radiation, which induces damage to DNA and other structures of a target cell, is one of well-recognized stress conditions influencing behavior of affected cells. A few recent studies have evidenced radiationinduced changes in composition of exosomes released from irradiated cells and their involvement in radiation-related communication between cells. Inducible pathways of exosome secretion activated in irradiated cells are regulated by TSAP6 protein (the transmembrane protein tumor suppressor-activated pathway 6), which is transcriptionally regulated by p53, hence cellular status of this major DNA damage response factor affects composition and secretion rate of exosomes released from target cells. Moreover, exosomes released from irradiated cells have been shown to mediate the radiation-induced bystander effect. Understanding radiation-related mechanisms involved in exosome formation and "makeup" of their cargo would shed light on the role of exosomes in systemic response of cells, tissues and organisms to ionizing radiation which may open new perspectives in translational medicine and anticancer-treatment.

  18. Antibody-coupled monolithic silica microtips for highthroughput molecular profiling of circulating exosomes.

    Science.gov (United States)

    Ueda, Koji; Ishikawa, Nobuhisa; Tatsuguchi, Ayako; Saichi, Naomi; Fujii, Risa; Nakagawa, Hidewaki

    2014-08-29

    Exosome-mediated signal transportation plays a variety of critical roles in cancer progression and metastasis. From the aspect of cancer diagnosis, circulating exosomes are ideal resources of biomarkers because molecular features of tumor cells are transcribed on them. However, isolating pure exosomes from body fluids is time-consuming and still major challenge to be addressed for comprehensive profiling of exosomal proteins and miRNAs. Here we constructed anti-CD9 antibody-coupled highly porous monolithic silica microtips which allowed automated rapid and reproducible exosome extraction from multiple clinical samples. We applied these tips to explore lung cancer biomarker proteins on exosomes by analyzing 46 serum samples. The mass spectrometric quantification of 1,369 exosomal proteins identified CD91 as a lung adenocarcinoma specific antigen on exosomes, which was further validated with CD9-CD91 exosome sandwich ELISA measuring 212 samples. Our simple device can promote not only biomarker discovery studies but also wide range of omics researches about exosomes.

  19. Perturbations in the Urinary Exosome in Transplant Rejection

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    Tara eSigdel

    2015-01-01

    Full Text Available Urine exosomes are small vesicles exocytosed into the urine by all renal epithelial cell types under normal physiologic and disease states. Urine exosomal proteins may mirror disease specific proteome perturbations in kidney injury. Analysis methodologies for the exosomal fraction of the urinary proteome were developed for comparing the urinary exosomal fraction versus unfractionated proteome for biomarker discovery. Urine exosomes were isolated by centrifugal filtration of urine samples collected from kidney transplant patients with and without acute rejection, which were biopsy matched. The proteomes of unfractionated whole urine (Uw and urine exosomes (Ue underwent mass spectroscopy-based quantitative proteonomics analysis. The proteome data were analyzed for significant differential protein abundances in acute rejection (AR. A total of 1018 proteins were identified in Uw and 349 proteins in Ue. 279 overlapped between the two urinary compartments and 70 proteins were unique to the Ue compartment. Of 349 exosomal proteins identified from transplant patients,220 had not been previously identified in the normal Ue fraction. 11 Ue proteins, functionally involved in an inflammatory and stress response, were more abundant in urine samples from patients with acute rejection, 3 of which are exclusive to the Ue fraction. Ue AR-specific biomarkers(8 were also detected in Uw, but since they were observed at significantly lower abundances in Uw, they were not significant for AR in Uw. A rapid urinary exosome isolation method and quantitative measurement of enriched Ue proteins was applied. Perturbed proteins in the exosomal compartment of urine collected from kidney transplant patients were specific to inflammatory responses, and were not observed in the Ue fraction from normal healthy subjects. Ue specific protein alterations in renal disease provide potential mechanistic insights and offer a unique panel of sensitive biomarkers for monitoring AR.

  20. Exosome mimetics: a novel class of drug delivery systems

    Directory of Open Access Journals (Sweden)

    Kooijmans SAA

    2012-03-01

    Full Text Available Sander AA Kooijmans, Pieter Vader, Susan M van Dommelen, Wouter W van Solinge, Raymond M SchiffelersDepartment of Clinical Chemistry and Haematology, University Medical Center Utrecht, Utrecht, The NetherlandsAbstract: The identification of extracellular phospholipid vesicles as conveyors of cellular information has created excitement in the field of drug delivery. Biological therapeutics, including short interfering RNA and recombinant proteins, are prone to degradation, have limited ability to cross biological membranes, and may elicit immune responses. Therefore, delivery systems for such drugs are under intensive investigation. Exploiting extracellular vesicles as carriers for biological therapeutics is a promising strategy to overcome these issues and to achieve efficient delivery to the cytosol of target cells. Exosomes are a well studied class of extracellular vesicles known to carry proteins and nucleic acids, making them especially suitable for such strategies. However, the considerable complexity and the related high chance of off-target effects of these carriers are major barriers for translation to the clinic. Given that it is well possible that not all components of exosomes are required for their proper functioning, an alternative strategy would be to mimic these vesicles synthetically. By assembly of liposomes harboring only crucial components of natural exosomes, functional exosome mimetics may be created. The low complexity and use of well characterized components strongly increase the pharmaceutical acceptability of such systems. However, exosomal components that would be required for the assembly of functional exosome mimetics remain to be identified. This review provides insights into the composition and functional properties of exosomes, and focuses on components which could be used to enhance the drug delivery properties of exosome mimetics.Keywords: exosomes, extracellular vesicles, liposomes, drug delivery systems

  1. Integrated Magneto-Electrochemical Sensor for Exosome Analysis.

    Science.gov (United States)

    Jeong, Sangmoo; Park, Jongmin; Pathania, Divya; Castro, Cesar M; Weissleder, Ralph; Lee, Hakho

    2016-02-23

    Extracellular vesicles, including exosomes, are nanoscale membrane particles that carry molecular information on parental cells. They are being pursued as biomarkers of cancers that are difficult to detect or serially follow. Here we present a compact sensor technology for rapid, on-site exosome screening. The sensor is based on an integrated magneto-electrochemical assay: exosomes are immunomagnetically captured from patient samples and profiled through electrochemical reaction. By combining magnetic enrichment and enzymatic amplification, the approach enables (i) highly sensitive, cell-specific exosome detection and (ii) sensor miniaturization and scale-up for high-throughput measurements. As a proof-of-concept, we implemented a portable, eight-channel device and applied it to screen extracellular vesicles in plasma samples from ovarian cancer patients. The sensor allowed for the simultaneous profiling of multiple protein markers within an hour, outperforming conventional methods in assay sensitivity and speed.

  2. Relevance of circulating tumor cells, extracellular nucleic acids, and exosomes in breast cancer.

    Science.gov (United States)

    Friel, Anne M; Corcoran, Claire; Crown, John; O'Driscoll, Lorraine

    2010-10-01

    Early detection of cancer is vital to improved overall survival rates. At present, evidence is accumulating for the clinical value of detecting occult tumor cells in peripheral blood, plasma, and serum specimens from cancer patients. Both molecular and cellular approaches, which differ in sensitivity and specificity, have been used for such means. Circulating tumor cells and extracellular nucleic acids have been detected within blood, plasma, and sera of cancer patients. As the presence of malignant tumors are clinically determined and/or confirmed upon biopsy procurement-which in itself may have detrimental effects in terms of stimulating cancer progression/metastases-minimally invasive methods would be highly advantageous to the diagnosis and prognosis of breast cancer and the subsequent tailoring of targeted treatments for individuals, if reliable panels of biomarkers suitable for such an approach exist. Herein, we review the current advances made in the detection of such circulating tumor cells and nucleic acids, with particular emphasis on extracellular nucleic acids, specifically extracellular mRNAs and discuss their clinical relevance.

  3. Antigen transfer from exosomes to dendritic cells as an explanation for the immune enhancement seen by IgE immune complexes.

    Directory of Open Access Journals (Sweden)

    Rebecca K Martin

    Full Text Available IgE antigen complexes induce increased specific T cell proliferation and increased specific IgG production. Immediately after immunization, CD23(+ B cells capture IgE antigen complexes, transport them to the spleen where, via unknown mechanisms, dendritic cells capture the antigen and present it to T cells. CD23, the low affinity IgE receptor, binds IgE antigen complexes and internalizes them. In this study, we show that these complexes are processed onto B-cell derived exosomes (bexosomes in a CD23 dependent manner. The bexosomes carry CD23, IgE and MHC II and stimulate antigen specific T-cell proliferation in vitro. When IgE antigen complex stimulated bexosomes are incubated with dendritic cells, dendritic cells induce specific T-cell proliferation in vivo, similar to IgE antigen complexes. This suggests that bexosomes can provide the essential transfer mechanism for IgE antigen complexes from B cells to dendritic cells.

  4. Cardiac progenitor-derived exosomes protect ischemic myocardium from acute ischemia/reperfusion injury

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Lijuan [Department of Cardiology, Zhongda Hospital, Medical School of Southeast University, Nanjing 210009 (China); Cardiovascular Disease, Internal Medicine, University of Cincinnati, 231 Albert Sabin Way, Cincinnati, OH 45267 (United States); Wang, Yingjie [Cardiovascular Disease, Internal Medicine, University of Cincinnati, 231 Albert Sabin Way, Cincinnati, OH 45267 (United States); Internal Medicine of Traditional Chinese Medicine, Shuguang Hospital of Shanghai University of Traditional Chinese Medicine, Shanghai 201203 (China); Pan, Yaohua; Zhang, Lan [Cardiovascular Disease, Internal Medicine, University of Cincinnati, 231 Albert Sabin Way, Cincinnati, OH 45267 (United States); Shen, Chengxing [Department of Cardiology, Xinhua Hospital, Shanghai Jiao Tong University, Shanghai (China); Qin, Gangjian [Feinberg Cardiovascular Research Institute, Northwestern University Feinberg School of Medicine, Chicago, IL 60611 (United States); Ashraf, Muhammad [Pathology and Lab Med, University of Cincinnati, 231 Albert Sabin Way, Cincinnati, OH 45267 (United States); Weintraub, Neal [Cardiovascular Disease, Internal Medicine, University of Cincinnati, 231 Albert Sabin Way, Cincinnati, OH 45267 (United States); Ma, Genshan, E-mail: magenshan@hotmail.com [Department of Cardiology, Zhongda Hospital, Medical School of Southeast University, Nanjing 210009 (China); Tang, Yaoliang, E-mail: tangyg@ucmail.uc.edu [Cardiovascular Disease, Internal Medicine, University of Cincinnati, 231 Albert Sabin Way, Cincinnati, OH 45267 (United States)

    2013-02-15

    Highlights: ► Cardiac progenitor-derived (CPC) Exosomes protect H9C2 from apoptosis in vitro. ► CPC-exosomes protect cardiomyoyctes from MI/R induced apoptosis in vivo. ► CPC-exosomes were taken up by H9C2 with high efficiency using PKH26 labeling. ► miR-451, one of GATA4-responsive miRNA cluster, is enriched in CPC-exosomes. -- Abstract: Background: Cardiac progenitors (CPC) mediate cardioprotection via paracrine effects. To date, most of studies focused on secreted paracrine proteins. Here we investigated the CPC-derived-exosomes on protecting myocardium from acute ischemia/reperfusion (MI/R) injury. Methods and results: CPC were isolated from mouse heart using two-step protocol. Exosomes were purified from conditional medium, and confirmed by electron micrograph and Western blot using CD63 as a marker. qRT-PCR shows that CPC-exosomes have high level expression of GATA4-responsive-miR-451. Exosomes were ex vivo labeled with PKH26, We observed exosomes can be uptaken by H9C2 cardiomyoblasts with high efficiency after 12 h incubation. CPC-exosomes protect H9C2 from oxidative stress by inhibiting caspase 3/7 activation invitro. In vivo delivery of CPC-exosomes in an acute mouse myocardial ischemia/reperfusion model inhibited cardiomyocyte apoptosis by about 53% in comparison with PBS control (p < 0.05). Conclusion: Our results suggest, for the first time, the CPC-exosomes can be used as a therapeutic vehicle for cardioprotection, and highlights a new perspective for using non-cell exosomes for cardiac disease.

  5. Isolation of Exosomes from the Plasma of HIV-1 Positive Individuals.

    Science.gov (United States)

    Konadu, Kateena Addae; Huang, Ming Bo; Roth, William; Armstrong, Wendy; Powell, Michael; Villinger, Francois; Bond, Vincent

    2016-01-05

    Exosomes are small vesicles ranging in size from 30 nm to 100 nm that are released both constitutively and upon stimulation from a variety of cell types. They are found in a number of biological fluids and are known to carry a variety of proteins, lipids, and nucleic acid molecules. Originally thought to be little more than reservoirs for cellular debris, the roles of exosomes regulating biological processes and in diseases are increasingly appreciated. Several methods have been described for isolating exosomes from cellular culture media and biological fluids. Due to their small size and low density, differential ultracentrifugation and/or ultrafiltration are the most commonly used techniques for exosome isolation. However, plasma of HIV-1 infected individuals contains both exosomes and HIV viral particles, which are similar in size and density. Thus, efficient separation of exosomes from HIV viral particles in human plasma has been a challenge. To address this limitation, we developed a procedure modified from Cantin et. al., 2008 for purification of exosomes from HIV particles in human plasma. Iodixanol velocity gradients were used to separate exosomes from HIV-1 particles in the plasma of HIV-1 positive individuals. Virus particles were identified by p24 ELISA. Exosomes were identified on the basis of exosome markers acetylcholinesterase (AChE), and the CD9, CD63, and CD45 antigens. Our gradient procedure yielded exosome preparations free of virus particles. The efficient purification of exosomes from human plasma enabled us to examine the content of plasma-derived exosomes and to investigate their immune modulatory potential and other biological functions.

  6. Ubiquitination as a Mechanism To Transport Soluble Mycobacterial and Eukaryotic Proteins to Exosomes.

    Science.gov (United States)

    Smith, Victoria L; Jackson, Liam; Schorey, Jeffrey S

    2015-09-15

    Exosomes are extracellular vesicles of endocytic origin that function in intercellular communication. Our previous studies indicate that exosomes released from Mycobacterium tuberculosis-infected macrophages contain soluble mycobacterial proteins. However, it was unclear how these secreted proteins were targeted to exosomes. In this study, we determined that exosome production by the murine macrophage cell line RAW264.7 requires the endosomal sorting complexes required for transport and that trafficking of mycobacterial proteins from phagocytosed bacilli to exosomes was dependent on protein ubiquitination. Moreover, soluble mycobacterial proteins, when added exogenously to RAW264.7 or human HEK293 cells, were endocytosed, ubiquitinated, and released via exosomes. This suggested that endocytosed proteins could be recycled from cells through exosomes. This hypothesis was supported using the tumor-associated protein He4, which, when endocytosed by RAW264.7 or HEK293 cells, was transported to exosomes in a ubiquitin-dependent manner. Our data suggest that ubiquitination is a modification sufficient for trafficking soluble proteins within the phagocytic/endocytic network to exosomes.

  7. The Complete Exosome Workflow Solution: From Isolation to Characterization of RNA Cargo

    Directory of Open Access Journals (Sweden)

    Jeoffrey Schageman

    2013-01-01

    Full Text Available Exosomes are small (30–150 nm vesicles containing unique RNA and protein cargo, secreted by all cell types in culture. They are also found in abundance in body fluids including blood, saliva, and urine. At the moment, the mechanism of exosome formation, the makeup of the cargo, biological pathways, and resulting functions are incompletely understood. One of their most intriguing roles is intercellular communication—exosomes function as the messengers, delivering various effector or signaling macromolecules between specific cells. There is an exponentially growing need to dissect structure and the function of exosomes and utilize them for development of minimally invasive diagnostics and therapeutics. Critical to further our understanding of exosomes is the development of reagents, tools, and protocols for their isolation, characterization, and analysis of their RNA and protein contents. Here we describe a complete exosome workflow solution, starting from fast and efficient extraction of exosomes from cell culture media and serum to isolation of RNA followed by characterization of exosomal RNA content using qRT-PCR and next-generation sequencing techniques. Effectiveness of this workflow is exemplified by analysis of the RNA content of exosomes derived from HeLa cell culture media and human serum, using Ion Torrent PGM as a sequencing platform.

  8. Exosome-mediated delivery of functionally active miRNA-155 inhibitor to macrophages.

    Science.gov (United States)

    Momen-Heravi, Fatemeh; Bala, Shashi; Bukong, Terence; Szabo, Gyongyi

    2014-10-01

    Exosomes, membranous nanovesicles, naturally carry bio-macromolecules and play pivotal roles in both physiological intercellular crosstalk and disease pathogenesis. Here, we showed that B cell-derived exosomes can function as vehicles to deliver exogenous miRNA-155 mimic or inhibitor into hepatocytes or macrophages, respectively. Stimulation of B cells significantly increased exosome production. Unlike in parental cells, baseline level of miRNA-155 was very low in exosomes derived from stimulated B cells. Exosomes loaded with a miRNA-155 mimic significantly increased miRNA-155 levels in primary mouse hepatocytes and the liver of miRNA-155 knockout mice. Treatment of RAW macrophages with miRNA-155 inhibitor loaded exosomes resulted in statistically significant reduction in LPS-induced TNFα production and partially prevented LPS-induced decrease in SOCS1 mRNA levels. Furthermore, exosome-mediated miRNA-155 inhibitor delivery resulted in functionally more efficient inhibition and less cellular toxicity compared to conventional transfection methods. Similar approaches could be useful in modification of target biomolecules in vitro and in vivo. From the clinical editor: In this study, exosome-based delivery of miRNA-155 mimicker or inhibitor was found to have significant biological response in hepatocytes and macrophages. Exosome-based approaches may be useful in the modification of other target biomolecules.

  9. Exosomes as Novel microRNA-Delivery Vehicles to Modulate Prostate Cancer Progression

    Science.gov (United States)

    2015-10-01

    AWARD NUMBER: W81XWH-14-1-0548 TITLE: Exosomes as Novel microRNA-Delivery Vehicles to Modulate Prostate Cancer Progression PRINCIPAL...Sep 2015 4. TITLE AND SUBTITLE Exosomes as Novel microRNA-Delivery Vehicles to Modulate Prostate Cancer Progression 5a. CONTRACT NUMBER 5b. GRANT...they are produced, but can also signal intercellularly to other cells and tissues at distant sites via exosomal transport. We hypothesize that miRNAs

  10. Exosomes enriched in stemness/metastatic-related mRNAS promote oncogenic potential in breast cancer

    Science.gov (United States)

    Rodríguez, Marta; Silva, Javier; Herrera, Alberto; Herrera, Mercedes; Peña, Cristina; Martín, Paloma; Gil-Calderón, Beatriz; Larriba, María Jesús; Coronado, Mª José; Soldevilla, Beatriz; Turrión, Víctor S.; Provencio, Mariano; Sánchez, Antonio; Bonilla, Félix; García-Barberán, Vanesa

    2015-01-01

    Cancer cells efficiently transfer exosome contents (essentially mRNAs and microRNAs) to other cell types, modifying immune responses, cell growth, angiogenesis and metastasis. Here we analyzed the exosomes release by breast tumor cells with different capacities of stemness/metastasis based on CXCR4 expression, and evaluated their capacity to generate oncogenic features in recipient cells. Breast cancer cells overexpressing CXCR4 showed an increase in stemness-related markers, and in proliferation, migration and invasion capacities. Furthermore, recipient cells treated with exosomes from CXCR4-cells showed increased in the same abilities. Moreover, inoculation of CXCR4-cell-derived exosomes in immunocompromised mice stimulated primary tumor growth and metastatic potential. Comparison of nucleic acids contained into exosomes isolated from patients revealed a “stemness and metastatic” signature in exosomes of patients with worse prognosis. Finally, our data supported the view that cancer cells with stem-like properties show concomitant metastatic behavior, and their exosomes stimulate tumor progression and metastasis. Exosomes-derived nucleic acids from plasma of breast cancer patients are suitable markers in the prognosis of such patients. PMID:26528758

  11. Exosomes enriched in stemness/metastatic-related mRNAS promote oncogenic potential in breast cancer.

    Science.gov (United States)

    Rodríguez, Marta; Silva, Javier; Herrera, Alberto; Herrera, Mercedes; Peña, Cristina; Martín, Paloma; Gil-Calderón, Beatriz; Larriba, María Jesús; Coronado, M Josés; Soldevilla, Beatriz; Turrión, Víctor S; Provencio, Mariano; Sánchez, Antonio; Bonilla, Félix; García-Barberán, Vanesa

    2015-12-01

    Cancer cells efficiently transfer exosome contents (essentially mRNAs and microRNAs) to other cell types, modifying immune responses, cell growth, angiogenesis and metastasis. Here we analyzed the exosomes release by breast tumor cells with different capacities of stemness/metastasis based on CXCR4 expression, and evaluated their capacity to generate oncogenic features in recipient cells. Breast cancer cells overexpressing CXCR4 showed an increase in stemness-related markers, and in proliferation, migration and invasion capacities. Furthermore, recipient cells treated with exosomes from CXCR4-cells showed increased in the same abilities. Moreover, inoculation of CXCR4-cell-derived exosomes in immunocompromised mice stimulated primary tumor growth and metastatic potential. Comparison of nucleic acids contained into exosomes isolated from patients revealed a "stemness and metastatic" signature in exosomes of patients with worse prognosis. Finally, our data supported the view that cancer cells with stem-like properties show concomitant metastatic behavior, and their exosomes stimulate tumor progression and metastasis. Exosomes-derived nucleic acids from plasma of breast cancer patients are suitable markers in the prognosis of such patients.

  12. 外质体(Exosomes)与肾脏疾病%Exosomes and kidney diseases

    Institute of Scientific and Technical Information of China (English)

    柏云

    2012-01-01

    Exosomes are nanovesicles originating from multivesicular bodies ( MVBs) and secreted into the extracellular space or body fluids when a multivesicular body {endocytic origin) fuses with the plasma membrane. Exosomes contain multiple proteins, mRNAs, microRNAs, and signaling molecules that may reflect the physiological state of their cells of origin and consequently provide potential biomarkers. At present,the studies on exosomes are mostly focused on their roles in immunology and oncology and exosorne-based immunotherapy has become a new means in cancer treatment and immune tolerance. In recent years, urinary exosomes (UE) and their roles in kidney diseases have been receiving great attention. Exosomes are secreted to the urine from all types of renal epithelial cell, including glomerular podocytes, renal tubular cells, and the cells lining the urinary drainage system. Thus, urinary exosomes have potential as a source of valuable biomarkers for early detection of kidney diseases. The present review aims to summarize their biological characteristics,and their potential uses in the diagnosis and treatment of kidney disease.%外质体( Exosomes)足起源于多泡体的微小囊泡,由细胞内吞途径中的多泡体外膜和细胞膜融合后释放到胞外环境或体液中.Exosomes含有多种蛋白、mRNAs、microRNAs、信号分子等,能够反映来源细胞的生物学状态,因而可能成为潜在的生物学标志物.目前,exosomes的研究大多集中在免疫学和肿瘤学,并已经成为一种免疫治疗的新手段,应用于肿瘤治疗和免疫耐受等方面.近年人们才关注exosomes与肾脏疾病的关系,研究表明几乎所有肾脏上皮细胞包括肾小球足细胞、肾小管上皮细胞、尿道上皮细胞均可分泌exosomes,因此尿液来源的exosomes可能成为寻找肾脏疾病早期诊断的标志物.本文着重从exosomes的生物学特性及其在肾脏疾病诊断和治疗的研究进行综述.

  13. Matrix metalloproteinase 13-containing exosomes promote nasopharyngeal carcinoma metastasis.

    Science.gov (United States)

    You, Yiwen; Shan, Ying; Chen, Jing; Yue, Huijun; You, Bo; Shi, Si; Li, Xingyu; Cao, Xiaolei

    2015-12-01

    Nasopharyngeal cancer (NPC) is an endemic type of head and neck cancer with a high rate of cervical lymph node metastasis. Metastasis is the major cause of death in NPC patients. Increasing evidence indicates that exosomes play a pivotal role in promoting cancer metastasis by enhancing angiogenesis and ECM degradation. Matrix metalloproteinase 13 is an important kind of matrix proteinase that is often overexpressed in various tumors and increases the risk of metastasis. However, little is known about the potential role of MMP13-containing exosomes in NPC. In this study, we found that MMP13 was overexpressed in NPC cells and exosomes purified from conditioned medium (CM) as well as NPC patients' plasma. Transwell analysis revealed that MMP13-containing exosomes facilitated the metastasis of NPC cells. Furthermore, siRNA inhibited the effect of MMP13-containing exosomes on tumor cells metastasis as well as angiogenesis. The current findings provided novel insight into the vital role of MMP13-containing exosomes in NPC progression which might offer unique insights for potential therapeutic strategies for NPC progressions.

  14. Urinary exosomal microRNAs in incipient diabetic nephropathy.

    Science.gov (United States)

    Barutta, Federica; Tricarico, Marinella; Corbelli, Alessandro; Annaratone, Laura; Pinach, Silvia; Grimaldi, Serena; Bruno, Graziella; Cimino, Daniela; Taverna, Daniela; Deregibus, Maria Chiara; Rastaldi, Maria Pia; Perin, Paolo Cavallo; Gruden, Gabriella

    2013-01-01

    MicroRNAs (miRNAs), a class of small non-protein-encoding RNAs, regulate gene expression via suppression of target mRNAs. MiRNAs are present in body fluids in a remarkable stable form as packaged in microvesicles of endocytic origin, named exosomes. In the present study, we have assessed miRNA expression in urinary exosomes from type 1 diabetic patients with and without incipient diabetic nephropathy. Results showed that miR-130a and miR-145 were enriched, while miR-155 and miR-424 reduced in urinary exosomes from patients with microalbuminuria. Similarly, in an animal model of early experimental diabetic nephropathy, urinary exosomal miR-145 levels were increased and this was paralleled by miR-145 overexpression within the glomeruli. Exposure of cultured mesangial cells to high glucose increased miR-145 content in both mesangial cells and mesangial cells-derived exosomes, providing a potential mechanism for diabetes-induced miR-145 overexpression. In conclusion, urinary exosomal miRNA content is altered in type 1 diabetic patients with incipient diabetic nephropathy and miR-145 may represent a novel candidate biomarker/player in the complication.

  15. Urinary exosomal microRNAs in incipient diabetic nephropathy.

    Directory of Open Access Journals (Sweden)

    Federica Barutta

    Full Text Available MicroRNAs (miRNAs, a class of small non-protein-encoding RNAs, regulate gene expression via suppression of target mRNAs. MiRNAs are present in body fluids in a remarkable stable form as packaged in microvesicles of endocytic origin, named exosomes. In the present study, we have assessed miRNA expre