Rheumatology: Current Research
Open Access

Extracellular Vesicles Biology and its Emerging Role in Osteoarthritis and Related Arthritides

Attur M, Mignatti P, Han T and Attur MG^{* *}Correspondence: Attur MG, Division of Rheumatology, Department of Medicine, NYU School of Medicine, New York, NY 10003, USA, Tel: +212-598-6578, Email:

Author info »

Markers Of Exosomes

Once exosomes are isolated from physiological fluids, it is essential to determine their purity. Exosomes contain certain universal lipids and proteins, including membrane transport proteins (RAB, GTPases), proteins used in MVB formation (Tsg 101 and Alix), heat shock proteins (hsc70), and tetraspanins (CD9, CD63, CD81, and CD82). Rab, SNARE and other membrane proteins are responsible for MVB docking and exosomal fusion with the plasma membrane [45]. These, in addition to the heat shock proteins and annexins, are integral to exosome intracellular assembly. Integrins, which mediate vesicle arget cell binding are also present on the surface of exosomes. Other proteins are specific for exosomes originated from specific cell types or tissues. In arthritis, specific biomarkers such as aggrecan, TGF-ß1 and LTBP-1 are used to identify exosomes secreted from disease-affected cells [43].

Discussion and Conclusion

However, this method was effective thanks to the inherent capacity of curcumin to alter the fluidity of lipid membranes penetrate into the exosome lumen. Despite highly promising experimental results, there are still many challenges in the development of exosome-based therapies. One of the largest hurdles is the lack of large-scale production techniques. Typically, 1 ml of culture medium will yield<1 μg of exosomal protein, while effective doses of exosomes range from 10 ug to100 ug. Certain techniques have been found to enhance exosome production, including the application of stressors such as low pH, hypoxia and anti-cancer drugs. However, it is important to note that the effects of these techniques on exosome cargo are not fully understood, and caution must be taken in their evaluation. Another problem involves the selection of isolation methods. As discussed above, different isolation techniques exploit different properties of exosomes. Recent studies have indicated that exosomes preparations containing subpopulations of vesicles with different composition and size may have variable therapeutic potential depending on the isolation method. Future studies are warranted to overcome these technical hurdles and develop novel pharmacological therapies based on exosome-mediated delivery.

Future Developments

The dynamic nature of exosomes lends itself to many clinical applications, both for diagnosis and drug delivery. Since exosomal cargo mirrors the cell of origin’s macromolecules such as lipids, proteins, and nucleic acids, exosomes present a great potential as biomarkers for a variety of diseases, ranging from autoimmune disorders to cancer. In fact, databases have been created that correlate exosomal proteins and mRNAs with the corresponding diseases [104].

As exosomes are so heavily involved in intercellular communication via the plasma membrane, it is believed that target cells have greater tolerance for them than for inorganic drug vectors. In addition to biocompatibility, exosomes have the advantage of a long life in the blood circulation, the ability to target specific cells and little to no toxicity, which makes them an ideal target for future research in drug delivery. Exosomes carry MHC peptide complexes recognized by T lymphocytes without invoking an immune response from the host cells [105]. They are also able to traverse the blood-brain barrier and synovial membranes [106]. Many potent drugs are unable to traverse the blood-brain barrier and are rapidly cleared by phagocytes; conversely, appropriate tailoring of exosomes could afford improved drug delivery to the CNS. Due to their ability to transport miRNA, proteins and viral antigens, exosomes offer several avenues to deliver anti-inflammatory molecules throughout the body. Therefore, the use of exosomes as vehicles for targeted drug or gene delivery appears very promising.

The successful use of exosomes for drug delivery depends on the efficiency of the techniques used to load them. The techniques described in the literature include electroporation, incubation, and transfection. Electroporation consists of applying an electrical field to a suspension of exosomes and the drug/cargo of choice, forming pores in the exosome membrane that allow the drug to move into its interior. Transfection is the most commonly used method for loading therapeutic drugs into exosomes. Exosome donor cells are transfected to overexpress the gene product of interest, which the cell will abundantly package into the exosomes [107]. While a rare method, incubation has also been used as a technique to load exosomes. Curcumin was effectively loaded into exosomes after a 5-minute incubation period at 22°C [108,109].

Osteoarthritis

Recent studies have highlighted the significance of exosomes in knee osteoarthritis (OA). OA pathology is derived from dysfunction in the balance between synthesis and breakdown of extracellular matrix (ECM), thought to benmediated by MMP-13 [87-89]. MMP-13 is produced by chondrocytes and fibroblast-like synoviocytes, and its expression is stimulated by IL-1ß and TNF- α [90,91]. Addition of IL-1ß treated chondrocyte EV to FLS increases MMP-13 production almost three-fold when compared to non-IL-1ß treated EV [92]. Conversely, addition of IL-1ß treated FLS EV to chondrocytes also increased MMP-13 production in addition to downregulating type II collagen [93]. Furthermore, recent studies have shown that miRNA content in exosomes is altered in OA synovial fluid relative to healthy controls and the changes are gender specific [94].

A pathological feature of OA includes the irregular hypercalcification of the tidemark, a layer of cartilage matrix found at the junction between articular cartilage and subarticular bone [95]. This is thought to lead to abnormal handling of mechanical stress at the joint, exacerbating degradation of articular cartilage [96]. Articular cartilage from the hips and knees of OA patients was shown to have more extracellular microvesicles than normal controls, and to exhibit high alkaline phosphatase activity [97]. Further studies have shown that microvesicles from OA chondrocytes contain annexin II, V and VI, all integral components of OA pathology [98].

Exosomes derived from OA synovium contain elevated levels of pro-inflammatory cytokines and chemokines, particularly IL-1ß [99]. Furthermore, exosomes from mesenchymal stem cells have been shown to promote both inflammatory and and antiinflammatory reactions via mTOR signaling and the control of mitochondrial functions [100,101]. Exosome-associated lncRNA KLF3-AS1 promotes chondrogenesis from progenitor cells [102]. Recent studies have also highlighted the role of exosomesderived from mesenchymal stem cells in promoting extracellular matrix synthesis and decreasing cartilage degeneration in animal models of OA [103].

Psoriatic Arthritis

The study of exosomes in psoriatic arthritis (PsA) is a very new and rapidly developing area of research. A recent study in China has begun studying exosomes as potential biomarkers for PsA. Twoexosome-associated long non-coding RNAs (lncRNA) - lnc- RP11-701H24.7 and lnc-RNU12 - were found to be increased in PsA patients compared to healthy controls, to be positively associated with clinical disease state [85]. Another recent study found five commonly expressed plasma exosomal microRNAs in patients with PsA, RA, gouty arthritis and psoriasis vulgaris, indicating similar inflammatory mechanism among these disorders [86].

Systemic Lupus Erythematosus

Systemic Lupus Erythematosus (SLE) is characterized as dysfunction in the immune system leading to abnormal activation of immune cells, with production of autoantibodies that lead to tissue inflammation and damage [80]. Serum exosomes from SLE patients have been shown to induce high cytokine production in normal peripheral blood mononuclear cells. Mechanical disruption of the exosomes causes the inflammatory response to dissipate [81]. Additionally, SLE exosomes induce higher levels of IFN-α, TNF-α and IL-1ß when compared to healthy control exosomes with no difference in IL--6 production. Interestingly, the same study showed that exosomes from RA patients increase IL-6 but not IFN-α production, possibly because type 1 interferons are a key cytokine in SLE pathogenesis [81]. Further studies, aimed to elucidate the nature of the micro vesicles present in SLE patients, found unique vesicle populations containing annexin V and apoptosis-modified chromatin that activated bloodderived myeloid and plasmacytoid dendritic cells enhancing formation of neutrophil traps [82]. SLE TNF-α type patients were also found to have high levels of microvesicles from platelets, monocytes and T-lymphocytes [83]. In SLE patients the extracellular microparticle-associated chromatin could be a potential self-antigen. Extracellular chromatin is cleared by secreted deoxyribonuclease -1 like 3 (DNASE1L3) under normal conditions and DNASE1L3 deficiency could lead to loss of this tolerance mechanism and contribute to SLE [84].

Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a systemic autoimmune disorder characterized by production of auto antibodies, inflammation of synovium, and destruction of joint cartilage and bone [68]. EV are thought to play a major role in RA pathology. Higher levels of platelet-derived microvesicles have been detected in RA patients than healthy individuals, and their circulating levels correlated with disease severity [69]. Elevated microvesicles levels were found in inflamed joints [63,64]. Microvesicles released from granulocytes, monocytes and platelets have been found in the synovial fluid of RA patients [70] and shown to stimulate synoviocytes via IL-1ß, thus activating inflammation and promoting joint destruction. Synovial fluid microvesicles have been shown to induce coagulation [70], which may be contribute to cardiovascular comorbidities in RA patients. Exosomes derived from fibroblast-like synoviocytes (FLS) have been reported to contain citrullinated proteins and a membrane form of TNF alpha [62,71]. Osteoclast-derived exosomes containing microRNA have been reported to play an inhibitory role in osteoblast activity, exacerbating bone resorption [72] and paracrine signaling by vesicles in osteoblasts [73].

Interestingly, it has been shown that disease-modifying antirheumatic drug (DMARD) treatments decrease the relative amounts of plasma microvesicles from generated from platelets, endothelial cells, monocytes and leukocytes following 4 weeks of treatment with methotrexate and prednisone. Similar changes were noted in urine, indicating a correlation between disease activity and plasma levels of microvesicles [74]. However, other studies have reported no changes in microvesicle levels after 8 weeks of DMARD treatment, despite reduction in disease activity [75].

Current pharmacological treatments of RA mostly consist of medications that inhibit tumor necrosis factor-α (TNF-α) and multiple other targets including JAK kinase inhibitor, IL-6 receptor, CD20, CD80/86 co-stimulatory molecule and methotrexate [76]. Exosomes derived from immunosuppressive dendritic cells or body fluids of immunized organisms exert antiinflammatory and immunosuppressive effects in diseased tissues [77]; they could therefore offer a safer approach to direct the differentiation of naive immune cells towards an immunosuppressive phenotype and to activate the suppressor cells. Furthermore, immunosuppressive DC-derived exosomes and blood plasma- or serum-derived exosomes have shown potent therapeutic effects in animal models of inflammatory and autoimmune disease including RA. Engineered myelogenous leukemia cell line (K562 cells) expressing triple-costimulatory signals (CD80) with HLA-A2 as an antigen presenting cells promotes long-term propagation of CD8+ T cells, and the addition of other co-stimulatory molecules such as CD80 and CD83 support the expansion of naive T cell subsets [78]. Engineered Exosome- derived from engineered immune cells provide immunosuppression and offer several advantages. RA patients treated with exosomes isolated from autologous conditioned serum (ACS) containing anti-inflammatory cytokines have shown decreased joint pain and decreased levels of circulating inflammatory markers [73]. The use of exosomes derived from stem cells has great regenerative potential since microRNAs responsible for reduction of joint inflammation and damage can be incorporated into exosomes for therapy [79].

Exosome Functions

Exosomes are believed to be mainly involved in intercellular communication, a combination of paracrine and juxtracrine signaling through the exchange of proteins, lipids, mRNA, miRNA, and DNA between cells. The cargo carried by exosomes is specific for the cell type of origin. For example, exosomes derived from dendritic cells and B cells are involved in immunity, and their dissemination pathways can be exploited by retroviruses such as HIV, which possess the multi-vesicular body (MVB) machinery for propagation [52]. Exosomes derived from tumors contain oncogenes and onco-miRNAs that contribute to tumorigenesis, metastasis, and angiogenesis. Cancer cells release larger amounts of exosomes which allows fusion of exosome with other cells, contributing to the signaling mechanisms involved in tumorigenesis and metastasis [53]. Exosomes released by malignant breast cancer cells are taken up by other cells within the tumor, promoting its metastatic potential [54].

Exosomes not only serve as signaling vesicles; they also have significant effects on the microenvironment. Tumor-derived exosomes contain cytokines and growth factors that control cell functions in a variety of cells. Similarly, exosome-associated tetraspanins contribute to activating endothelial cells, and promote angiogenesis [55].

Certain exosomal proteins, such as the MET oncoprotein, mutant KRAS, and Tissue Factor, are directly linked with tumor cell proliferation and coagulation [22,56]. Prions and viruses can be transferred to host cells via exosomes in neurodegenerative diseases, including Alzheimer’s disease (AD). It has been proposed that retroviruses exploit exosomes both for the generation of viral particles and as a mode of infection. Based on this “Trojan exosome hypothesis” exosome uptake affords virus cells targeting by viruses through a mechanism independent of specific cell membrane receptor-viral envelop protein interaction [57].

Exosomes contribute to spreading pathological proteins (such as phosphorylated tau, or α-synuclein) across the nervous system in neurological conditions [58]. Exosome levels of total tau proteins (pT181-tau and pS396-tau) have been shown to be higher in AD patients than in case-controls, even 1–10 years before the diagnosis, suggesting that tau levels associated with blood exosomes can predict AD development before clinical onset [59]. In addition, the presence of exosome-associated tau phosphorylated at Thr-181 (AT270) in human cerebrospinal fluid suggests exosomes’ involevement in the pathogenesis of AD [60]. Exosomes also play an important role in immune surveillance through antigen presentation and the ability to modulate immune responses [19]. Exosomal long non-coding RNAs have also been implicated in aging and aging-related diseases, including atherosclerosis, Type 2 diabetes, osteoporosis, and osteoarthritis, by promoting cell senescence and apoptosis [61].

Exosomes contain autoantigens in patients with autoimmune diseases. Citrullinated proteins consisting of fibrinogen peptides and the CD5 precursor antigen are autoantigens involved in the pro-inflammatory response of rheumatoid arthritis (RA) [62]. Platelet-derived proinflammatory exosomes contain cytokines that induce synovial fibroblast secretion of IL-8, which causes joint inflammation and contributes to the irreversible process of cartilage erosion [1]. Long noncoding RNAs (lncRNA) such as Hotair are also found in the exosomes of RA patients. Hotair expression induces macrophage activation and matrix metalloproteinase (MMP) synthesis. Exosome-associated Hothair could therefore be used as a biomarker for rheumatoid arthritis [63]. Studies have also shown that IL-10 treated dendritic cells release exosomes that can prevent collagen-induced arthritis in rabbit and mouse models [64].

Due to their roles in intercellular communication, exosomes have been linked to viruses. Exosomes and viruses are similar in both size (under 300 nm in diameter) and composition. Similar to exosomes, viruses also contain proteins such as tetraspanins, cytoplasmic proteins, and heat shock proteins for proliferation. For these reasons, it can be very difficult to separate exosomes from the viruses themselves [65]. Exosomes can mediate both infection and immunity as they can transport viral content but also activate lymphocyte responses. In addition, exosomes transport to target cells antiviral response components, such as Toll-like receptor (TLR) ligands that alert neighboring cells to mount an immune response against the virus. Antigen presentation by exosomes induces immune cell activation. Inversely, exosomes can impair immune cell functions by transmitting viral content to target cells, facilitating virus binding to host cells, and inducing immune cell apoptosis [66]. Exosomes released from HIV-infected cells contain virusassociated proteins and genetic material that facilitate prolongation of the infection. By mediating delivery of virulent factors such as HIV Negative Regulatory Factor, exosomes can actually induce T cell apoptosis during the early stages of HIV infection [67]. Other viruses have also been shown to use exosomal pathways of infection. Epstein-Barr virus infects Bcells, causing release of EBV miRNA via exosomes. In addition, human intestinal epithelial cells infected with rotaviruses release exosomes containing various viral proteins as well as immunomodulators.

Exosomes as Biomarkers

Due to their essential roles as transport vehicles for proteins, microRNA, and lipids, exosomes are being increasingly seen as important effectors of important physiological and pathological functions. Exosomes from different parts of the body have different functions - those derived from platelets are involved in the inflammatory response, while those secreted by antigenpresenting cells can carry MHC class I or II molecules, participating in immune responses [13].

Exosomes are associated with numerous diseases, ranging from cancer to arthritis. They can be isolated from a variety of biofluids, including saliva, urine, synovial tissue, plasma, and serum, which makes them convenient for diagnostic purposes lends great specificity and sensitivity for diagnostics. Exosomal cargo, sorted and highly concentrated by the ESCRT complex, consists of genetic material in the form of DNA, messenger RNA and miRNA, which can provide insight into the pathology of many different types of cancer. Several exosomal proteins serve as biomarkers, including tetraspanins and heat shock proteins for melanoma, epidermal growth factor (EGF) for bladder cancer, and Fetuin-A for kidney disease [46]. Exosomes transport misfolded proteins in patients with Parkinson’s disease [47], and exosomes with particularly high concentrations of tetraspanins, such as CD63, are possible indicators of melanoma [48] and other malignant tumors, as tetraspanin levels are elevated in variety of cancer cells [49].

Recent studies indicate that exosomes have a highly specific tropism for their cell of origin, which can be used to target them to diseased tissues and/or organs. For example, cancer cellderived exosomes have unique characteristics that can be of interest for drug transport system design [50]. Urinary exosomes could provide diagnostic information for liver and kidney conditions, as well as bladder and prostate cancer [51]. Because RNase present in biological fluids cannot degrade miRNAs contained in exosomes, the use of exosomal biomarkers provides a high level of stability for accurate and sensitive bio-diagnostics. Currently, alternative sources of biofluids, such as saliva and amniotic fluid, are being investigated along with blood and urine in order to devevop novel, more sensitive methods of diagnosis. Amniotic fluid, in particular, could provide fetal exosomal markers that can allow the early diagnosis of congenital conditions.

Introduction

Multicellular organisms depend on intercellular communication to control a variety of cell functions and maintain homeostasis. While the mechanisms behind cell-cell contact and molecular signaling have been extensively studied, the importance of extracellular vesicles (EV) in intercellular communication has only recently been appreciated [1-3].

The first observation of EV is believed to have been made in 1946 by Chargaff and West, who wrote of procoagulant plateletderived particles found in normal plasma [4]. In the 1970s there followed an increasing number of independent EV observations, from the detection of vesicles in seminal plasma [5] to observations of tumor-originating membrane fragments [6]. The next large steps in EV research occurred in the 1980s, when studies of reticulocytes found that vesicles, in addition to being released from the cell surface, could also be released via fusion of multi-vesicular bodies (MVB) to the plasma membrane [7,8].

Immunological Separation

Lastly, numerous immunological separation methods can be used to isolate exosomes from biological fluids, including ELISA-based techniques or magnetic beads coated with specific antibodies. This method affords separating cell type-specific exosomes; however, it may alter EV’s biological activities and is not convenient to use with large sample volumes [43]. Nowadays, commercial reagents such as Exoquick (Takara Bio USA, Inc) are used to isolate exosomes from serum by a single overnight incubation, without the multiple long periods of differential centrifugation.

Once isolated from biological fluids, EV can be further analyzed by electron microscopy and various biochemical and biological approaches including genomics, proteomics and metabolomics. The characterization of the isolated vesicles is essential to ensure that the purified material contains the appropriate extracellular vesicle. The identification of biomarkers affords understanding exosome biological functions, and the characterization of their proteins, lipids and nucleic acids can provide indications of their tissue of origin. While there are universal proteins on all exosomes, the presence and quantitity of a variety of protein components depend on the specific cell the exosomes are secreted from [44]. Western blotting and ELISA can be used to identify these macromolecular components of EV.

Precipitation

Polymer-based precipitation consists of the addition of a large polymer (usually polythrilenglycol, or PEG) to a biological fluid with the resulting precipitation of large molecular complexes such as EV. However, the polymer may irreversibly denature EV components, affecting their biological activities and therefore hampering functional studies of the purified EV. Furthermore, the isolation is often incomplete, and results in EV with a low grade of purification [42].

Chromatography

Size-exclusion chromatography separates components of biological fluids based on their size. While this method is precise, and the structure of EV is not affected by shearing force, the process is time-consuming and quantitatively inefficient, which can limit the amount and biological activity of the purified material. Ultrafiltration membranes can also be used as a way to eliminate larger components from the exosomes; however, exosome adhesion to the filtration membrane results in reduced amounts of pure exosomes recovered by this method [42].

Centrifugation

Differential centrifugation is the most common method for isolating EV [39]. By this method, the sample undergoes several centrifugation cycles to remove intact cells or debris. The supernatant is then processed through ultracentrifugation to obtain pure exosomes. While differential centrifugation is most commonly used, it is more effective with serum than plasma samples due to the higher viscosity of plasma. Important limitations of this technique include co-precipitation of apoptotic bodies and various nucleosomal fragments, as well as the need for low viscous biofluids. Longer centrifugation times and higher centrifugal force are needed to compensate for both limitations [40]. Sucrose density gradient centrifugation separates vesicles based on differences in flotation densities [41]. Density gradient centrifugation is best used for low-density exosome purification but is very sensitive to alterations in centrifugation time [19].

Methods Of Isolation

A number of techniques have been developed to isolate and analyze EV taking advantage of their structural and biochemical features. These techniques include differential or density gradient centrifugation, chromatography, filtration, polymerbased precipitation, and immunological separation.

Biogenesis

Extracellular vesicles (EV) are membrane-contained vesicles whose release from cells has been conserved through evolution, from prokaryotes to unicellular and multicellular eukaryotes [17]. Heterogenous in structure, EV carry a wide variety of molecules, ranging from proteins to multiple RNA species, DNA, and lipids [18-21]. EV release often occurs into bodily fluids such as serum, plasma, and urine. To elicit their functional effects, EV bind to the plasma membrane of target cells, inducing phenotypic changes in the cell via cell surface signal initiation or vesicle internalization [22]. Extracellular vesicles are categorized into three main categories-microvesicles, apoptotic bodies, and exosomes, based on intracellular origin and size.

Microvesicles (also termed microparticles, particles or ectosomes in the literature) are a heterogeneous population of vesicles primarily formed through the direct outward budding and fission of the plasma membrane, followed by release into the extracellular space [23]. Microvesicles range in size from 100 nm to 1,000 nm [3]. The blebbing of the plasma membrane is accompanied by localized shifts in plasma membrane components, with a resulting local enrichment of microvesicle cargo components as well as changes in membrane curvature and rigidity [24]. Microvesicle budding utilizes the ARF6 (ADPribosylation factor 6, a RAS-related GTPase) and the ESCRT (endosomal sorting complexes required for transport) system [25]. Studies have identified ARF6 as integral to the regulation of selective protein recruitment into microvesicles, and of microvesicle shedding through its function in cell invasion, peripheral actin remodeling, and endocytic trafficking [26,27]. Unlike other sub-classes of EV, microvesicles can vary greatly in composition and morphology depending on cell type (s) and health or disease conditions [28]. They contribute to various physiological and pathological processes, including tumor invasion and metastasis, and are believed to play a role in inflammation, coagulation, stem-cell renewal and expansion, evasion from the immune response, and bone mineralization [29]. For example, microvesicles secreted by skeletal cells have been found to play a role in initiating bone mineralization, while microvesicles secreted from endothelial cells or adipocytes have been implicated in angiogenesis and glucose metabolism, respectively [30-32].

Apoptotic bodies, or apoptosomes, are extracellular structures released from cells undergoing apoptosis (programmed cell death). A cell undergoing apoptosis goes through multiple stages, starting with fragmentation of nuclear chromatin and progressing to membrane blebbing, with formation of membrane-enclosed vesicles containing disintegrated cell components [33]. After release, apoptotic bodies are phagocytosed by local macrophages through interactions between the vesicle cell membrane and the phagocytic receptors [34]. While apoptotic bodies range in size from 500 to 4,000 nm, oftentimes during apoptosis vesicles in the 50-500 nm range are also released [33-35]. It is currently unclear whether they are formed through the same processes as the larger vesicles released. It has been suggested that uptake of apoptotic bodies can result in transfer of genetic material from a dying cell to a viable cell. For example, apoptotic bodies from human c-myctransfected cells confer loss of contact inhibition on murine fibroblasts in vitro [36].

Unlike apoptotic bodies and microvesicles, exosomes originate in the endosomal system, and are typically smaller than 100 nm [3]. Early endosomes mature into late endosomes, or MVB. As this process occurs, the ESCRT machinery mediates the invagination of the endosomal membranes to form intraluminal vesicles (ILV), which are released as exosomes when the MVB fuse with the plasma membrane [37]. Exosomes have a multitude of functions involved in cell-cell communication and transport, depending on the cell type from which they are secreted. They can transfer proteins, lipids, and nucleic acids into, and alter gene expression in target cells. Like the plasma membrane they originate from, exosomes consist of a lipid bilayer with higher cholesterol and phosphatidylserine levels, which may facilitate internalization through endocytosis, phagocytosis, and fusion [38]. Exosomes are the most studied among EV. We will therefore focus our review on this type of EV.

Review Of The History

In the 1990s, studies began showing an increasingly clear connection between the release of EV and the immune response, such as the capacity of EV derived from Epstein-Barr virus-transformed B to induce a T-cell response [9] Multiple studies have shown that EV contain multiple RNA species, including microRNAs (miRNA), making them a potentially important player in cell-cell communication [10-15]. The number of publications on EV has skyrocketed in the past 10 years (Figure 1), and the increased interest has led to a deeper characterization of the various types of EV, as well as the standardization of the nomenclature, also thanks to the creation of the International Society of Extracellular Vesicles (ISEV) and the NIH program, Extracellular RNA Communication Consortium (ERCC) [16].

Figure 1. Number of publications containing the word “microvesicles” or “exosomes” in the title or abstract.

References

1. Raposo G, Stahl PD. Extracellular vesicles: a new communication paradigm?. Nat Rev Mol Cell Biol. 2019;20(9):509-510.
2. van Niel G, D'Angelo G, Raposo G. Shedding light on the cell biology of extracellular vesicles. Nat Rev Mol Cell Biol. 2018;19(4):213-228.
3. Raposo G, Stoorvogel W. Extracellular vesicles: exosomes, microvesicles, and friends. J Cell Biol. 2013;200(4):373-383.
4. Chargaff E, West R. The biological significance of the thromboplastic protein of blood. J Biol Chem. 1946;166(1):189-197.
5. Ronquist G, Brody I, Gottfries A, Stegmayr B. An Mg2+ and Ca2+-stimulated adenosine triphosphatase in human prostatic fluid: part I. Andrologia. 1978;10(4):261-272.
6. Taylor DD, Homesley HD, Doellgast GJ. Binding of specific peroxidase-labeled antibody to placental-type phosphatase on tumor-derived membrane fragments. Cancer Res.  1980;40(11):4064-4069.
7. Harding C, Heuser J, Stahl P. Endocytosis and intracellular processing of transferrin and colloidal gold-transferrin in rat reticulocytes: demonstration of a pathway for receptor shedding. Eur J Cell Biol. 1984;35(2):256-263.
8. Johnstone RM, Adam M, Hammond JR, Orr L, Turbide C. Vesicle formation during reticulocyte maturation. Association of plasma membrane activities with released vesicles (exosomes). J Biol Chem. 1987;262(19):9412-9420.
9. Raposo G, Nijman HW, Stoorvogel W, Liejendekker R, Harding CV, Melief CJ, et al. B lymphocytes secrete antigen-presenting vesicles. J Exp Med. 1996;183(3):1161-1172.
10. Baj-Krzyworzeka M, Szatanek R, Weglarczyk K, Baran J, Urbanowicz B, Branski P, et al. Tumour-derived microvesicles carry several surface determinants and mRNA of tumour cells and transfer some of these determinants to monocytes. Cancer Immunol Immunother. 2006;55(7):808-818.
11. Ratajczak J, Wysoczynski M, Hayek F, Janowska-Wieczorek A, Ratajczak MZ. Membrane-derived microvesicles: important and underappreciated mediators of cell-to-cell communication. Leukemia. 2006;20(9):1487-1495.
12. Aliotta JM, Sanchez-Guijo FM, Dooner GJ, Bearer EL, Passero MA, Adedi M, et al. Alteration of marrow cell gene expression, protein production, and engraftment into lung by lung-derived microvesicles: a novel mechanism for phenotype modulation. Stem Cells. 2007;25(9):2245-2256.
13. Valadi H, Ekstrom K, Bossios A, Sjostrand M, Lee JJ, Lotvall JO. Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells. Nat Cell Biol. 2007;9(6):654-659.
14. Skog J, Wurdinger T, van Rijn S, Carter RS, Krichevsky AM, Breakefield XO, et al. Glioblastoma microvesicles transport RNA and proteins that promote tumour growth and provide diagnostic biomarkers. Nat Cell Biol. 2008;10(12):1470-1476.
15. Pegtel DM, Cosmopoulos K, Thorley-Lawson DA, Lindenberg JL, de Gruijl TD, Würdinger T, et al. Functional delivery of viral miRNAs via exosomes. Proc Natl Acad Sci US A. 2010;107(14):6328-6333.
16. Leslie M. Cell Biology. NIH effort gambles on mysterious extracellular RNAs. Science. 2013;341(6149):947.
17. Schorey JS, Cheng Y, Singh PP, Smith VL. Exosomes and other extracellular vesicles in host-pathogen interactions. EMBO Rep. 2015;16(1):24-43.
18. Simpson RJ, Lim JW, Moritz RL, Mathivanan S. Exosomes: proteomic insights and diagnostic potential. Expert Rev Proteomics. 2009;6(3):267-283.
19. Colombo M, Raposo G, Thery C. Biogenesis, secretion, and intercellular interactions of exosomes and other extracellular vesicles. Annu Rev Cell Dev Biol. 2014;30(1):255-289.
20. Christianson HC, Svensson KJ, Belting M. Exosome and microvesicle mediated phene transfer in mammalian cells. Semin Cancer Biol. 2014;28(3):31-38.
21. Rajendran L, Bali J, Barr MM, van Niel G, Wang J, Breakefield XO, et al. Emerging roles of extracellular vesicles in the nervous system. J Neurosci. 2014;34(46):15482-15489.
22. French KC, Antonyak MA, Cerione RA. Extracellular vesicle docking at the cellular port: Extracellular vesicle binding and uptake. Semin Cell Dev Biol. 2017;67(8):48-55.
23. Muralidharan-Chari V, Clancy JW, Sedgwick A, D'Souza-Schorey C. Microvesicles: mediators of extracellular communication during cancer progression. J Cell Sci. 2010;123(Pt 10):1603-1611.
24. Kozlov MM, Campelo F, Liska N, Chernomordik LV, Marrink SJ, McMahon HT. Mechanisms shaping cell membranes. Curr Opin Cell Biol. 2014;29(8):53-60.
25. Nabhan JF, Hu R, Oh RS, Cohen SN, Lu Q. Formation and release of arrestin domain-containing protein 1-mediated microvesicles (ARMMs) at plasma membrane by recruitment of TSG101 protein. Proc Natl Acad Sci USA. 2012;109(11):4146-4151.
26. Pellon-Cardenas O, Clancy J, Uwimpuhwe H, D'Souza-Schorey C. ARF6-regulated endocytosis of growth factor receptors links cadherin-based adhesion to canonical Wnt signaling in epithelia. Mol Cell Biol. 2013;33(15):2963-2975.
27. D'Souza-Schorey C, Chavrier P. ARF proteins: roles in membrane traffic and beyond. Nat Rev Mol Cell Biol. 2006;7(5):347-358.
28. Yanez-Mo M, Siljander PR, Andreu Z, Stukelj R, Van der Grein SG, Vasconcelos MH, et al. Biological properties of extracellular vesicles and their physiological functions. J Extracell Vesicles. 2015;4:27066.
29. Gangoda L, Boukouris S, Liem M, Kalra H, Mathivanan S. Extracellular vesicles including exosomes are mediators of signal transduction: are they protective or pathogenic? Proteomics. 2015;15(2-3):260-271.
30. Nawaz M, Shah N, Zanetti BR, Silvestre RN, Fatima F, Neder, et al. Extracellular vesicles and matrix remodeling enzymes: The emerging roles in extracellular matrix remodeling, progression of diseases and tissue repair. Cells. 2018;7(10). E167.
31. Hromada C, Muhleder S, Grillari J, Redl H, Holnthoner W. Endothelial extracellular vesicles-promises and challenges. Front Physiol. 2017;8:275.
32. Wu Q, Li J, Li Z, Wang C, Wu L, Zhu S, et al. Exosomes from the tumour-adipocyte interplay stimulate beige/brown differentiation and reprogram metabolism in stromal adipocytes to promote tumour progression. J Exp Clin Cancer Res. 2019;38(1):223.
33. Kerr JF, Wyllie AH, Currie AR. Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br J Cancer. 1972;26(4):239-257.
34. Takizawa F, Tsuji S, Nagasawa S. Enhancement of macrophage phagocytosis upon iC3b deposition on apoptotic cells. FEBS Lett. 1996;397(2-3):269-272.
35. Ihara T, Yamamoto T, Sugamata M, Okumura H, Ueno Y. The process of ultrastructural changes from nuclei to apoptotic body. Virchows Arch. 1998;433(5):443-447.
36. Bergsmedh A, Szeles A, Henriksson M, Bratt A, Folkman MJ, Spetz AL, et al. Horizontal transfer of oncogenes by uptake of apoptotic bodies. Proc Natl Acad Sci USA. 2001;98(11):6407-6411.
37. Huotari J, Helenius A. Endosome maturation. EMBO J. 2011;30(17):3481-3500.
38. van Meer G, Voelker DR, Feigenson GW. Membrane lipids: where they are and how they behave. Nat Rev Mol Cell Biol. 2008;9(2):112-124.
39. Livshits MA, Khomyakova E, Evtushenko EG. Isolation of exosomes by differential centrifugation: Theoretical analysis of a commonly used protocol. Sci Rep. 2015;5:17319.
40. Momen-Heravi F, Balaj L, Alian S, Semina SE, Generozov EV, Govorun VM, et al. Current methods for the isolation of extracellular vesicles. Biol Chem. 2013;394(10):1253-1262.
41. Cantin R, Diou J, Belanger D, Tremblay AM, Gilbert C. Discrimination between exosomes and HIV-1: purification of both vesicles from cell-free supernatants. J Immunol Methods. 2008;338(1-2):21-30.
42. Konoshenko MY, Lekchnov EA, Vlassov AV, Laktionov PP. Isolation of extracellular vesicles: general methodologies and latest trends. Biomed Res Int. 2018;2018:8545347.
43. Buzas EI, Gyorgy B, Nagy G, Falus A, Gay S. Emerging role of extracellular vesicles in inflammatory diseases. Nat Rev Rheumatol. 2014;10(6):356-364.
44. Zhang Y, Liu Y, Liu H, Tang WH. Exosomes: biogenesis, biologic function and clinical potential. Cell Biosci. 2019;9:19.
45. Ghidoni R, Benussi L, Binetti G. Exosomes: the Trojan horses of neurodegeneration. Med Hypotheses. 2008;70(6):1226-1227.
46. Lachenal G, Pernet-Gallay K, Chivet M, Blot B, Haase G, Goldberg Y, et al. Release of exosomes from differentiated neurons and its regulation by synaptic glutamatergic activity. Mol Cell Neurosci. 2011;46(2):409-418.
47. Ghidoni R, Paterlini A, Albertini V, Benussi L, Levy E, Binetti G, et al. Cystatin C is released in association with exosomes: a new tool of neuronal communication which is unbalanced in Alzheimer's disease. Neurobiol Aging. 2011;32(8):1435-1442.
48. Logozzi M, De Milito A, Lugini L, Maio M, Rivoltini L, Fais S, et al. High levels of exosomes expressing CD63 and caveolin-1 in plasma of melanoma patients. PLoS One. 2009;4(4):e5219.
49. Detchokul S, Williams ED, Parker MW, Frauman AG. Tetraspanins as regulators of the tumour microenvironment: implications for metastasis and therapeutic strategies. Br J Pharmacol. 2014;171(24):5462-5490.
50. Batrakova EV, Kim MS. Using exosomes, naturally-equipped nanocarriers, for drug delivery. J Control Release. 2015;219:396-405.
51. van Balkom BW, Pisitkun T, Verhaar MC, Knepper MA. Exosomes and the kidney: prospects for diagnosis and therapy of renal diseases. Kidney Int. 2011;80(11):1138-1145.
52. Wolfson B, Yu JE, Zhou Q. Exosomes may play a crucial role in HIV dendritic cell immunotherapy. Ann Transl Med. 2017;5(16):337.
53. Kharaziha P, Ceder S, Li Q, Panaretakis T. Tumor cell-derived exosomes: a message in a bottle. Biochim Biophys Acta. 2012;1826(1):103-111.
54. Le MT, Hamar P, Guo C, Basar E, Balaj L, Lieberman J, et al. miR-200-containing extracellular vesicles promote breast cancer cell metastasis. J Clin Invest. 2014;124(12):5109-5128.
55. Nazarenko I, Rana S, Baumann A, Trendelenburg M, Lochnit G, Preissner KT, et al. Cell surface tetraspanin Tspan8 contributes to molecular pathways of exosome-induced endothelial cell activation. Cancer Res. 2010;70(4):1668-1678.
56. Beckler MD, Higginbotham JN, Franklin JL, Halvey PJ, Imasuen IE, Whitwell C, et al. Proteomic analysis of exosomes from mutant KRAS colon cancer cells identifies intercellular transfer of mutant KRAS. Mol Cell Proteomics. 2013;12(2):343-355.
57. Gould SJ, Booth AM, Hildreth JE. The Trojan exosome hypothesis. Proc Natl Acad Sci USA. 2003;100(19):10592-10597.
58. Chivet M, Javalet C, Hemming F, Laulagnier K, Fraboulet S, Sadoul R, et al. Exosomes as a novel way of interneuronal communication. Biochem Soc Trans. 2013;41(1):241-244.
59. Fiandaca MS, Kapogiannis D, Mapstone M, Federoff HJ, Miller BL, Goetzl EJ, et al. Identification of preclinical Alzheimer's disease by a profile of pathogenic proteins in neurally derived blood exosomes: A case-control study. Alzheimers Dement. 2015;11(6):600-607.e1.
60. Saman S, Kim W, Raya M, Alvarez VE, Lee CY, Hall GF, et al. Exosome-associated tau is secreted in tauopathy models and is selectively phosphorylated in cerebrospinal fluid in early Alzheimer disease. J Biol Chem. 2012;287(6):3842-3849.
61. Cao Q, Guo Z, Yan Y, Wu J, Song C. Exosomal long noncoding RNAs in aging and age-related diseases. IUBMB Life. 2019.
62. Skriner K, Adolph K, Jungblut PR, Burmester GR. Association of citrullinated proteins with synovial exosomes. Arthritis Rheum. 2006;54(12):3809-3814.
63. Song J, Kim D, Han J, Kim Y, Lee M, Jin EJ. PBMC and exosome-derived Hotair is a critical regulator and potent marker for rheumatoid arthritis. Clin Exp Med. 2015;15(1):121-126.
64. Kim SH, Lechman ER, Bianco N, Watkins SC, Gambotto A, Robbins PD, et al. Exosomes derived from IL-10-treated dendritic cells can suppress inflammation and collagen-induced arthritis. J Immunol. 2005;174(10):6440-6448.
65. Zhang Y, Bergelson JM. Adenovirus receptors. J Virol. 2005;79(19):12125-12131.
66. Chahar HS, Bao X, Casola A. Exosomes and their role in the life cycle and pathogenesis of RNA viruses. Viruses. 2015;7(6):3204-3225.
67. Lenassi M, Cagney G, Liao M, Krogan NJ, Plemenitas A, Peterlin BM, et al. HIV Nef is secreted in exosomes and triggers apoptosis in bystander CD4+ T cells. Traffic. 2010;11(1):110-122.
68. McInnes IB, Schett G. The pathogenesis of rheumatoid arthritis. N Engl J Med. 2011;365(23):2205-2219.
69. Knijff-Dutmer EA, Koerts J, Nieuwland R, Kalsbeek-Batenburg EM, van de Laar MA. Elevated levels of platelet microparticles are associated with disease activity in rheumatoid arthritis. Arthritis Rheum. 2002;46(6):1498-1503.
70. Berckmans RJ, Nieuwland R, Tak PP, Breedveld FC, Hack CE, Sturk A, et al. Cell-derived microparticles in synovial fluid from inflamed arthritic joints support coagulation exclusively via a factor VII-dependent mechanism. Arthritis Rheum. 2002;46(11):2857-2866.
71. Zhang HG, Liu C, Su K, Cao X, Grizzle W, Kimberly RP, et al. A membrane form of TNF-alpha presented by exosomes delays T cell activation-induced cell death. J Immunol. 2006;176(12):7385-7393.
72. Sun W, Zhao C, Li Y, Ling S, Chang YZ, Li Y, et al. Osteoclast-derived microRNA-containing exosomes selectively inhibit osteoblast activity. Cell Discov. 2016;2:16015.
73. Morhayim J, Rudjito R, van Leeuwen JP, van Driel M. Paracrine Signaling by Extracellular Vesicles via Osteoblasts. Curr Mol Biol Rep. 2016;2:48-55.
74. Vinuela-Berni V, Doniz-Padilla L, Figueroa-Vega N, Abud-Mendoza C, Baranda L, González-Amaro R, et al. Proportions of several types of plasma and urine microparticles are increased in patients with rheumatoid arthritis with active disease. Clin Exp Immunol. 2015;180(3):442-451.
75. van Eijk IC, Tushuizen ME, Sturk A, Wolbink GJ, Nieuwland R, Nurmohamed MT, et al. Circulating microparticles remain associated with complement activation despite intensive anti-inflammatory therapy in early rheumatoid arthritis. Ann Rheum Dis. 2010;69(7):1378-1382.
76. Aletaha D, Smolen JS. Diagnosis and management of rheumatoid arthritis: A Review. JAMA. 2018;320(13):1360-1372.
77. De Toro J, Herschlik L, Waldner C, Mongini C. Emerging roles of exosomes in normal and pathological conditions: new insights for diagnosis and therapeutic applications. Front Immunol. 2015;6:203.
78. Kim S, Sohn HJ, Lee HJ, Hyun SJ, Cho HI, Kim TG, et al. Use of engineered exosomes expressing HLA and costimulatory molecules to generate antigen-specific CD8+ T cells for adoptive cell therapy. J Immunother. 2017;40(3):83-93.
79. Chen Z, Wang H, Xia Y, Yan F, Lu Y. Therapeutic potential of mesenchymal cell-derived MIRNA-150-5p-expressing exosomes in rheumatoid arthritis mediated by the modulation of MMP14 and VEGF. J Immunol. 2018;201(8):2472-2482.
80. Rahman A, Isenberg DA. Systemic lupus erythematosus. N Engl J Med. 2008;358(9):929-939.
81. Lee JY, Park JK, Lee EY, Lee EB, Song YW. Circulating exosomes from patients with systemic lupus erythematosus induce an proinflammatory immune response. Arthritis Res Ther. 2016;18(1):264.
82. Dieker J, Tel J, Pieterse E, de Vries JM, Hilbrands LB, van der Vlag J, et al. Circulating Apoptotic microparticles in systemic lupus erythematosus patients drive the activation of dendritic cell subsets and prime neutrophils for NETosis. Arthritis Rheumatol. 2016;68(2):462-472.
83. Lopez P, Rodriguez-Carrio J, Martinez-Zapico A, Caminal-Montero L, Suarez A. Circulating microparticle subpopulations in systemic lupus erythematosus are affected by disease activity. Int J Cardiol. 2017;236:138-144.
84. Sisirak V, Sally B, D'Agati V, Clancy RM, Buyon JP, Reizis B, et al. Digestion of chromatin in apoptotic cell microparticles prevents autoimmunity. Cell. 2016;166(1):88-101.
85. Yue T, Ji M, Qu H, Wang W, Gong X, Zhang Z, et al. Comprehensive analyses of long non-coding RNA expression profiles by RNA sequencing and exploration of their potency as biomarkers in psoriatic arthritis patients. BMC Immunol. 2019;20(1):28.
86. Chen XM, Zhao Y, Wu XD, Huang QC, Huang RY, Lu CJ, et al. Novel findings from determination of common expressed plasma exosomal microRNAs in patients with psoriatic arthritis, psoriasis vulgaris, rheumatoid arthritis, and gouty arthritis. Discov Med. 2019;28(151):47-68.
87. Nakamura H, Shibakawa A, Tanaka M, Kato T, Nishioka K. Effects of glucosamine hydrochloride on the production of prostaglandin E2, nitric oxide and metalloproteases by chondrocytes and synoviocytes in osteoarthritis. Clin Exp Rheumatol. 2004;22(3):293-299.
88. Little CB, Barai A, Burkhardt D, Werb Z, Shah M, Thompson EW, et al. Matrix metalloproteinase 13-deficient mice are resistant to osteoarthritic cartilage erosion but not chondrocyte hypertrophy or osteophyte development. Arthritis Rheum. 2009;60(12):3723-3733.
89. Goldring MB, Otero M, Plumb DA, Olivotto E, Borzì RM, Marcu KB, et al. Roles of inflammatory and anabolic cytokines in cartilage metabolism: signals and multiple effectors converge upon MMP-13 regulation in osteoarthritis. Eur Cell Mater. 2011;21:202-220.
90. Liacini A, Sylvester J, Li WQ, Dehnade F, Ahmad M, Zafarullah M, et al. Induction of matrix metalloproteinase-13 gene expression by TNF-alpha is mediated by MAP kinases, AP-1, and NF-kappaB transcription factors in articular chondrocytes. Exp Cell Res. 2003;288(1):208-217.
91. Mengshol JA, Vincenti MP, Coon CI, Barchowsky A, Brinckerhoff CE. Interleukin-1 induction of collagenase 3 (matrix metalloproteinase 13) gene expression in chondrocytes requires p38, c-Jun N-terminal kinase, and nuclear factor kappaB: differential regulation of collagenase 1 and collagenase 3. Arthritis Rheum. 2000;43(4):801-811.
92. Li Z, Wang Y, Xiao K, Xiang S, Li Z, Weng X. Emerging Role of Exosomes in the Joint Diseases. Cell Physiol Biochem. 2018;47(5):2008-2017.
93. Kato T, Miyaki S, Ishitobi H, Nakamura Y, Nakasa T, Ochi M, et al. Exosomes from IL-1beta stimulated synovial fibroblasts induce osteoarthritic changes in articular chondrocytes. Arthritis Res Ther. 2014;16(4):R163.
94. Kolhe R, Hunter M, Liu S, Guldberg RE, Hamrick MW, Fulzele S, et al. Gender-specific differential expression of exosomal miNA in synovial fluid of patients with osteoarthritis. Sci Rep. 2017;7(1):2029.
95. Bonde HV, Talman ML, Kofoed H. The area of the tidemark in osteoarthritis--a three-dimensional stereological study in 21 patients. APMIS. 2005;113(5):349-352.
96. Thambyah A. A hypothesis matrix for studying biomechanical factors associated with the initiation and progression of posttraumatic osteoarthritis. Med Hypotheses. 2005;64(6):1157-1161.
97. Ali SY, Griffiths S. Formation of calcium phosphate crystals in normal and osteoarthritic cartilage. Ann Rheum Dis. 1983;42 Suppl 1:45-48.
98. Kirsch T, Swoboda B, Nah H. Activation of annexin II and V expression, terminal differentiation, mineralization and apoptosis in human osteoarthritic cartilage. Osteoarthritis Cartilage. 2000;8(4):294-302.
99. Gao K, Zhu W, Li H, Wang L, Cao Y, Jiang Y, et al. Association between cytokines and exosomes in synovial fluid of individuals with knee osteoarthritis. Mod Rheumatol. 2019:1-7.
100. Wu J, Kuang L, Chen C, Ni Z, Chen L, Yang L, et al. miR-100-5p-abundant exosomes derived from infrapatellar fat pad MSCs protect articular cartilage and ameliorate gait abnormalities via inhibition of mTOR in osteoarthritis. Biomaterials. 2019;206:87-100.
101. Qi H, Liu DP, Xiao DW, Tian DC, Su YW, Jin SF. Exosomes derived from mesenchymal stem cells inhibit mitochondrial dysfunction-induced apoptosis of chondrocytes via p38, ERK, and Akt pathways. In Vitro Cell Dev Biol Anim. 2019;55(3):203-210.
102. Liu Y, Zou R, Wang Z, Wen C, Zhang F, Lin F. Exosomal KLF3-AS1 from hMSCs promoted cartilage repair and chondrocyte proliferation in osteoarthritis. Biochem J. 2018;475(22):3629-3638.
103. Wang Y, Yu D, Liu Z, Zou XH, Ouyang H, Liu H, et al. Exosomes from embryonic mesenchymal stem cells alleviate osteoarthritis through balancing synthesis and degradation of cartilage extracellular matrix. Stem Cell Res Ther. 2017;8(1):189.
104. Mathivanan S, Fahner CJ, Reid GE, Simpson RJ. ExoCarta 2012: database of exosomal proteins, RNA and lipids. Nucleic Acids Res. 2012;40(Database issue):D1241-1244.
105. Muntasell A, Berger AC, Roche PA. T cell-induced secretion of MHC class II-peptide complexes on B cell exosomes. EMBO J. 2007;26(19):4263-4272.
106. Wood MJ, O'Loughlin AJ, Samira L. Exosomes and the blood-brain barrier: implications for neurological diseases. Ther Deliv. 2011;2(9):1095-1099.
107. Bunggulawa EJ, Wang W, Yin T, Wang N, Durkan C, Wang Y, Wang G. Recent advancements in the use of exosomes as drug delivery systems. J Nanobiotechnology. 2018;16(1):81.
108. Sun D, Zhuang X, Xiang X, Grizzle W, Miller D, Zhang HG, et al. A novel nanoparticle drug delivery system: the anti-inflammatory activity of curcumin is enhanced when encapsulated in exosomes. Mol Ther. 2010;18(9):1606-1614.
109. Harmati M, Tarnai Z, Decsi G, Nagy I, Nagy K, Buzas A, et al. Stressors alter intercellular communication and exosome profile of nasopharyngeal carcinoma cells. J Oral Pathol Med. 2017;46(4):259-266.