Exosomes: Small vesicles participating in intercellular communication

Co-culture methods to study neuronal function and disease.

Astroglia play active and diverse roles in modulating neuronal/synaptic functions in the CNS. How these astroglial functions are regulated, especially by neuronal signals, remains largely unknown. Exosomes, a major type of extracellular vesicles (EVs) that originate from endosomal intraluminal vesicles (ILVs), have emerged as a new intercellular communication process. By generating cell-type-specific ILVs/exosome reporter (CD63-GFPf/f) mice and immuno-EM/confocal image analysis, we found that neuronal CD63-GFP+ ILVs are primarily localized in soma and dendrites, but not in axonal terminals in vitro and in vivo. Secreted neuronal exosomes contain a subset of microRNAs (miRs) that is distinct from the miR profile of neurons. These miRs, especially the neuron-specific miR-124-3p, are potentially internalized into astrocytes. MiR-124-3p further up-regulates the predominant glutamate transporter GLT1 by suppressing GLT1-inhibiting miRs. Our findings suggest a previously undescribed neuronal exosomal miR-mediated genetic regulation of astrocyte functions, potentially opening a new frontier in understanding CNS intercellular communication.

Background: Rheumatoid arthritis (RA) is a chronic, invasive autoimmune disease characterized by symmetrical polyarthritis involving synovial inflammation. Epidemiological studies indicate that the incidence of RA continues to rise, yet the pathogenesis of this disease remains not fully understood. A significant infiltration of macrophages is observed in the synovium of RA patients. It can be inferred that macrophages likely play a crucial role in the onset and progression of RA. Summary: This review aims to summarize the research progress on the mechanisms by which macrophages and their associated structures contribute to RA, as well as potential therapeutic approaches, aiming to provide new insights into the study of RA pathogenesis and its clinical treatment. Key Messages: During the course of RA, besides the inherent roles of macrophages, these cells respond to microenvironmental changes such as pathogen invasion or tissue damage by undergoing polarization, pyroptosis, or forming macrophage extracellular traps (METs), all of which influence inflammatory responses and immune homeostasis, thereby mediating the occurrence and development of RA. Additionally, macrophages secrete exosomes, which participate in intercellular communication and signal transduction processes, thus contributing to the progression of RA. Therefore, it is critical to elucidate how macrophages and their related structures function in RA. Background: Rheumatoid arthritis (RA) is a chronic, invasive autoimmune disease characterized by symmetrical polyarthritis involving synovial inflammation. Epidemiological studies indicate that the incidence of RA continues to rise, yet the pathogenesis of this disease remains not fully understood. A significant infiltration of macrophages is observed in the synovium of RA patients. It can be inferred that macrophages likely play a crucial role in the onset and progression of RA. Summary: This review aims to summarize the research progress on the mechanisms by which macrophages and their associated structures contribute to RA, as well as potential therapeutic approaches, aiming to provide new insights into the study of RA pathogenesis and its clinical treatment. Key Messages: During the course of RA, besides the inherent roles of macrophages, these cells respond to microenvironmental changes such as pathogen invasion or tissue damage by undergoing polarization, pyroptosis, or forming macrophage extracellular traps (METs), all of which influence inflammatory responses and immune homeostasis, thereby mediating the occurrence and development of RA. Additionally, macrophages secrete exosomes, which participate in intercellular communication and signal transduction processes, thus contributing to the progression of RA. Therefore, it is critical to elucidate how macrophages and their related structures function in RA. Background: Rheumatoid arthritis (RA) is a chronic, invasive autoimmune disease characterized by symmetrical polyarthritis involving synovial inflammation. Epidemiological studies indicate that the incidence of RA continues to rise, yet the pathogenesis of this disease remains not fully understood. A significant infiltration of macrophages is observed in the synovium of RA patients. It can be inferred that macrophages likely play a crucial role in the onset and progression of RA. Summary: This review aims to summarize the research progress on the mechanisms by which macrophages and their associated structures contribute to RA, as well as potential therapeutic approaches, aiming to provide new insights into the study of RA pathogenesis and its clinical treatment. Key Messages: During the course of RA, besides the inherent roles of macrophages, these cells respond to microenvironmental changes such as pathogen invasion or tissue damage by undergoing polarization, pyroptosis, or forming macrophage extracellular traps (METs), all of which influence inflammatory responses and immune homeostasis, thereby mediating the occurrence and development of RA. Additionally, macrophages secrete exosomes, which participate in intercellular communication and signal transduction processes, thus contributing to the progression of RA. Therefore, it is critical to elucidate how macrophages and their related structures function in RA. Background: Rheumatoid arthritis (RA) is a chronic, invasive autoimmune disease characterized by symmetrical polyarthritis involving synovial inflammation. Epidemiological studies indicate that the incidence of RA continues to rise, yet the pathogenesis of this disease remains not fully understood. A significant infiltration of macrophages is observed in the synovium of RA patients. It can be inferred that macrophages likely play a crucial role in the onset and progression of RA. Summary: This review aims to summarize the research progress on the mechanisms by which macrophages and their associated structures contribute to RA, as well as potential therapeutic approaches, aiming to provide new insights into the study of RA pathogenesis and its clinical treatment. Key Messages: During the course of RA, besides the inherent roles of macrophages, these cells respond to microenvironmental changes such as pathogen invasion or tissue damage by undergoing polarization, pyroptosis, or forming macrophage extracellular traps (METs), all of which influence inflammatory responses and immune homeostasis, thereby mediating the occurrence and development of RA. Additionally, macrophages secrete exosomes, which participate in intercellular communication and signal transduction processes, thus contributing to the progression of RA. Therefore, it is critical to elucidate how macrophages and their related structures function in RA.

Exosomes are small vesicles with a diameter of approximately 50-100 nm that are secreted by a number of different cells and function in a multitude of intercellular signaling processes. Exosomes are thought to derive from release of intraluminal vesicles (ILVs) in multivesicular bodies (MVBs) after the fusion of MVBs with the plasma membrane.To understand the little explored molecular details of how cargoes are sorted to these subsets of ILVs in MVBs and secreted with exosomes, the transport of proteolipid protein (PLP) in Oli-neu cells, an oligodendroglial precursor cell line, was studied. We found that PLP was transported to distinct subdomains of the endosomal membrane and that the transfer of PLP into the lumen of endosomes did not depend on the function of the endosomal sorting complex required for transport (ESCRT) machinery but required ceramide formation. The purified exosomes were enriched with ceramide and inhibition of neutral sphingomyelinase activity reduced the release of exosomes. Sphingomyelin hydrolysis on the outer leaflet of phase separated giant unilamellar vesicles resulted in intraluminal budding from the lipid ordered phase. These results identified a novel lipid-dependent pathway for cargo transport to the ILVs in MVBs for exosomes formation.Rab family guanosine triphosphatases (GTPases) are regulators involved in intracellular membrane traffcking. A screen for the effects of Rab GTPase activating proteins (GAPs) on exosome release was performed. Expression of TBC1D10B, RN-tre, TBC1D10A, TBC1D10C and TBC1D15 reduced exosome secretion in a catalytic activity dependent manner. Rab35 was identified to be the target of TBC1D10A-C. Inhibition of Rab35 function impaired exosome secretion, induced accumulation of PLP containing endosomes and increased the mobility of endosomes closed to the plasma membrane, suggesting a function in MVB tethering. Furthermore, Rab35 was biochemically purified together with non-compact myelin, where MVBs were observed, providing a basis for understanding the biogenesis and functions of exosomes in the central nervous system.

Atherosclerotic cardiovascular disease is the leading cause of morbidity and mortality all over the world. Extracellular vesicles (EVs), small lipid-bilayer membrane vesicles released by most cellular types, exert pivotal and multifaceted roles in physiology and disease. Emerging evidence emphasizes the importance of EVs in intercellular communication processes with key effects on cell survival, endothelial homeostasis, inflammation, neoangiogenesis, and thrombosis. This review focuses on EVs as effective signaling molecules able to both derail vascular homeostasis and induce vascular dysfunction, inflammation, plaque progression, and thrombus formation as well as drive anti-inflammation, vascular repair, and atheroprotection. We provide a comprehensive and updated summary of the role of EVs in the development or regression of atherosclerotic lesions, highlighting the link between thrombosis and inflammation. Importantly, we also critically describe their potential clinical use as disease biomarkers or therapeutic agents in atherothrombosis.

Normal and malignant cells release a variety of different vesicles into their extracellular environment. The most prominent vesicles are the microvesicles (MVs, 100-1 000 nm in diameter), which are shed of the plasma membrane, and the exosomes (70-120 nm in diameter), derivates of the endosomal system. MVs have been associated with intercellular communication processes and transport numerous proteins, lipids and RNAs. As essential component of immune-escape mechanisms tumor-derived MVs suppress immune responses. Additionally, tumor-derived MVs have been found to promote metastasis, tumor-stroma interactions and angiogenesis. Since members of the carcinoembryonic antigen related cell adhesion molecule (CEACAM)-family have been associated with similar processes, we studied the distribution and function of CEACAMs in MV fractions of different human epithelial tumor cells and of human and murine endothelial cells. Here we demonstrate that in association to their cell surface phenotype, MVs released from different human epithelial tumor cells contain CEACAM1, CEACAM5 and CEACAM6, while human and murine endothelial cells were positive for CEACAM1 only. Furthermore, MVs derived from CEACAM1 transfected CHO cells carried CEACAM1. In terms of their secretion kinetics, we show that MVs are permanently released in low doses, which are extensively increased upon cellular starvation stress. Although CEACAM1 did not transmit signals into MVs it served as ligand for CEACAM expressing cell types. We gained evidence that CEACAM1-positive MVs significantly increase the CD3 and CD3/CD28-induced T-cell proliferation. All together, our data demonstrate that MV-bound forms of CEACAMs play important roles in intercellular communication processes, which can modulate immune response, tumor progression, metastasis and angiogenesis.

Atherosclerosis is a chronic vascular inflammatory disorder, and the cardiovascular disease resulting from it is the leading cause of death worldwide. Extracellular vesicles (EVs) are small membrane vesicles secreted by cells, playing a crucial role in regulating intercellular communication, signal transduction, and various pathophysiological processes. EVs have been shown to play a significant role in the progression and regression of atherosclerotic cardiovascular disease. Therefore, a comprehensive understanding of the role of EVs in the pathogenesis of atherosclerosis is essential for advancing knowledge in this field and driving the development of innovation of novel diagnostic and therapeutic strategies. This review summarizes the pathological roles of EVs in atherosclerosis, including their involvement in endothelial cell inflammation, the phenotypic transformation of vascular smooth muscle cells, macrophage activation, foam cell formation, and plaque thrombosis and rupture. Furthermore, it discusses innovative EV-based strategies for diagnosing and treating atherosclerosis. Finally, the challenges and prospects of translational research on EVs in atherosclerosis are discussed. This work aims to provide an EV-centered perspective on the diagnosis and treatment of atherosclerosis.

Cancer is a complex organ whose behavior is not only influenced by genetic and epigenetic changes in cancer cells but also by stromal cells, local extracellular matrix and specific tissue architecture. Intercellular communications within the cancer microenvironment are critical to coordinate the assembly of multiple cell types for an amalgamated form and function of a cancer. Exosomes are small membrane vesicles with an endosome origin that are released by cells into the extracellular environment. They carry a cargo of proteins, lipids, and nucleic acids and transfer their cargo to recipient cells and altering the recipient cells' biochemical composition, signaling pathways, and gene regulation. Exosomes can thus serve as extracellular messengers mediating cell-cell communication. Both cancer cells and stromal cells release exosomes not only into the cancer microenvironment but also into the circulation. In this review, we summarize the research done so far on cancer-derived exosomes and assess their roles as extracellular messengers facilitating cancer progression and metastasis.

In the last decades, new evidence on brain structure and function has been acquired by morphological investigations based on synergic interactions between biochemical anatomy approaches, new techniques in microscopy and brain imaging, and quantitative analysis of the obtained images. This effort produced an expanded view on brain architecture, illustrating the central nervous system as a huge network of cells and regions in which intercellular communication processes, involving not only neurons but also other cell populations, virtually determine all aspects of the integrative function performed by the system. The main features of these processes are described. They include the two basic modes of intercellular communication identified (i.e., wiring and volume transmission) and mechanisms modulating the intercellular signaling, such as cotransmission and allosteric receptor–receptor interactions. These features may also open new possibilities for the development of novel pharmacological approaches to address central nervous system diseases. This aspect, with a potential major impact on molecular medicine, will be also briefly discussed.

Intercellular small molecular weight signaling molecules modulate a variety of biological functions in bacteria. One of the more complex behaviors mediated by intercellular signaling molecules is the suite of activities regulated by quorum sensing molecules. These molecules mediate a variety of population-dependent responses, including the expression of genes that regulate bioluminescence, type III secretion, siderophore production, colony morphology, biofilm formation, and metalloprotease production. Given their central role in regulating these responses, the detection and quantification of QS molecules has important practical implications. Until recently, the detection of QS molecules from Gram-negative bacteria has relied primarily on bacterial reporter systems. These bioassays though immensely useful are subject to interference by compounds that affect bacterial growth and metabolism. In addition, the reporter response is highly dependent on culture age and cell population density. To overcome such limitations, we developed an in vitro protein-based assay system for the rapid detection and quantification of the furanosyl borate diester (BAI-2) subclass of autoinducer-2 (AI-2) QS molecules. The biosensor is based on the interaction of BAI-2 with the Vibrio harveyi QS receptor LuxP. Conformation changes associated with BAI-2 binding to the LuxP receptor change the orientation of cyan and yellow variants of GFP (CFP and YFP) fused the N- and C-termini, respectively, of the LuxP receptor. LuxP-BAI2 binding induces changes in fluorescence resonance energy transfer (FRET) between CFP and YFP, whose magnitude of change is ligand concentration dependent. A set of ligand-insensitive LuxP-mutant FRET protein sensor was also developed for use as control biosensors. The FRET-based BAI-2 biosensor responds selectively to both synthetic and biologically derived BAI-2compounds. This report describes the use of the LuxP-FRET biosensor for the detection and quantification of BAI-2.

It has long been known that keratinocytes influence cutaneous immunity through secretion of soluble factors. Exosomes, small membrane vesicles of endocytotic origin, have been implicated in intercellular communication processes such as the transfer of tumor cell antigens and the activation of recipient dendritic cells (DC). However, little is known about immunomodulatory functions of keratinocyte-derived exosomes. To address this question, we analysed exosome secretion of the murine keratinocyte cell line MPEK under steady state as well as inflammatory conditions (+/- IFNγ). These exosomes were readily taken up by bone marrow-derived DC (BMDC) in vitro resulting in a matured phenotype, as evidenced by increased CD40 expression as well as by the production of large amounts of IL-6, IL-10 and IL-12. When the transfer of antigen-specific information through exosomes was investigated, it was found that keratinocytes took up antigen (ovalbumin) and transferred it to their exosomes. However, these antigen-harbouring exosomes failed to induce antigen-specific T cell responses via BMDC. Together, this novel biological function suggests that keratinocytes are able to direct unspecific immune processes but do not elicit specific immune responses.

Intercellular signalling is an indispensable part of multicellular life. Understanding the commonalities and differences in how signalling molecules function in two remote branches of the tree of life may shed light on the reasons these molecules were originally recruited for intercellular signalling. Here we review the plant function of three highly studied animal intercellular signalling molecules, namely glutamate, γ-aminobutyric acid (GABA), and melatonin. By considering both their signalling function in plants and their broader physiological function, we suggest that molecules with an original function as key metabolites or active participants in reactive ion species scavenging have a high chance of becoming intercellular signalling molecules. Naturally, the evolution of machinery to transduce a message across the plasma membrane is necessary. This fact is demonstrated by three other well-studied animal intercellular signalling molecules, namely serotonin, dopamine, and acetylcholine, for which there is currently no evidence that they act as intercellular signalling molecules in plants.

Origin and characterization of small membranous vesicles present in the venom of Crotalus durissus terrificus

decompression sickness (DCS) has long been attributed to physical forces exerted by inert gas bubbles that may form in tissues, resulting in vascular occlusion and tissue disruption. Bubble formation occurs when a decrease in ambient pressure exceeds the rate at which soluble inert gas (e.g.,

Proteomics characterization of blood and circulating material has been extensively explored for the study of pathological states. In particular, circulating small extracellular vesicles (sEV, diameter: 30-150nm) are known to play an important role in intercellular communication processes, and proteomics profiling has been explored to develop minimally invasive assays for disease monitoring and diagnosis. Due to the genetic and physiological similarities between the two species, and also on account of their shorter life span and rapid disease progression, rodent models are the most commonly used animal model for many human diseases. Such models have provided invaluable insight into the molecular mechanisms of disease progression, candidate drug efficacy, therapy monitoring, and biomarkers research.Longitudinal investigations, in which individuals are monitored over periods of time, are more able to resolve molecular changes during disease progression because they circumvent the inter-individual variation. Longitudinal investigations of rodent models are challenging because of the limited amount of blood that can be withdrawn at each time; the American Association of Veterinary Science stipulates that fortnightly sampling should be limited to a maximum of 10% of the total blood volume. For adult mice this corresponds to approximately 75μL of serum. We developed an approach for the isolation and characterization of serum sEV proteins from just 50μL of serum, for longitudinal studies of disease mouse models. This chapter describes in detail the steps and considerations involved in the sEV isolation, morphological characterization, and proteome profiling by mass spectrometry.