Potential application of biomimetic exosomes in cardiovascular disease: focused on ischemic heart disease

Cardiovascular disease, especially ischemic heart disease, is a major cause of mortality worldwide. Cardiac repair is one of the most promising strategies to address advanced cardiovascular diseases. Despite moderate improvement in heart function via stem cell therapy, there is no evidence of significant improvement in mortality and morbidity beyond standard therapy. The most salutary effect of stem cell therapy are attributed to the paracrine effects and the stem cell-derived exosomes are known as a major contributor. Hence, exosomes are emerging as a promising therapeutic agent and potent biomarkers of cardiovascular disease. Furthermore, they play a role as cellular cargo and facilitate intercellular communication. However, the clinical use of exosomes is hindered by the absence of a standard operating procedures for exosome isolation and characterization, problems related to yield, and heterogeneity. In addition, the successful clinical application of exosomes requires strategies to optimize cargo, improve targeted delivery, and reduce the elimination of exosomes. In this review, we discuss the basic concept of exosomes and stem cell-derived exosomes in cardiovascular disease, and introduce current efforts to overcome the limitations and maximize the benefit of exosomes including engineered biomimetic exosomes.     

Exosomes for angiogenesis induction in ischemic disorders

Ischaemic disorders are leading causes of morbidity and mortality worldwide. While the current therapeutic approaches have improved life expectancy and quality of life, they are unable to “cure” ischemic diseases and instate regeneration of damaged tissues. Exosomes are a class of extracellular vesicles with an average size of 100–150 nm, secreted by many cell types and considered a potent factor of cells for paracrine effects. Since exosomes contain multiple bioactive components such as growth factors, molecular intermediates of different intracellular pathways, microRNAs and nucleic acids, they are considered as cell-free therapeutics. Besides, exosomes do not rise cell therapy concerns such as teratoma formation, alloreactivity and thrombotic events. In addition, exosomes are stored and utilized more convenient. Interestingly, exosomes could be an ideal complementary therapeutic tool for ischemic disorders. In this review, we discussed therapeutic functions of exosomes in ischemic disorders including angiogenesis induction through various mechanisms with specific attention to vascular endothelial growth factor pathway. Furthermore, different delivery routes of exosomes and different modification strategies including cell preconditioning, gene modification and bioconjugation, were highlighted. Finally, pre-clinical and clinical investigations in which exosomes were used were discussed.     

Exosomes in carcinogenesis: molecular palkis carry signals for the regulation of cancer progression and metastasis

Exosomes, which act as biological cargo vessels, are cell-released, phospholipid-enclosed vesicles. In eukaryotic cells, exosomes carry and exchange biological materials or signals for the benefit or detriment to the cells. Thereby, we consider exosomes to be molecular Palkis (carriers). Although exosomes are currently one of the most popularly researched cellular entities, they have remained largely enigmatic and warrant continued investigation into their structure and functions. These membraned vesicles are between 30 and 150 nm in diameter and are actively secreted by all cell types. While initially considered cellular “trash bags,” recent years have revealed exosomes to be dynamic and multi-functional vesicles that may play a crucial role in cancer development, progression and metastasis. Thereby, they have the potential to be used in development of therapeutic modalities for cancer and other diseases. As more research studies emerge, it’s becoming evident that exosomes are released by cells with a purpose and are representatives of certain cell types and disease conditions. Hence, they may also be used as biomarkers for the detection of cancer initiation, progression and organotropic metastatic growth of cancer cells. This review will focus on the recent developments achieved in identifying the role of exosomes in cancer development and progression as well as therapeutic implications. The review will also discuss the pitfalls of methodologies used for the extraction of exosomes.     

外泌体作为疾病诊断标志物及药物载体的应用前景

作为一种纳米级别的囊泡,外泌体的相关研究近年来逐渐成为热点。外泌体来源于细胞内的多囊泡胞内体,经由细胞膜释放到细胞外。由于来自特定细胞类型的外泌体含有多种特异性的蛋白质和microRNA,使其成为了可以广泛用于疾病诊断及预后的新型生物标志物。相较于其他外源性药物载体,外泌体具有更低的免疫原性,并能够靶向作用于病变细胞。这使得由细胞天然产生或经过人工改造的外泌体作为一种新兴的药物载体具有良好的发展前景。特别是近几年,外泌体在临床应用领域的发展潜力不断获得拓展,针对肿瘤、糖尿病、心脑血管疾病、神经退行性病变等重大疾病,以外泌体为基础的疾病诊断和药物的研发都取得了快速的进步。本篇综述重点介绍了外泌体作为一种生物标志物在疾病诊断和预后中的应用,同时阐述了外泌体作为一种新兴的药物载体所具有的优势以及存在的问题。     

外泌体在淋巴瘤中的研究进展

摘要：病毒感染和环境污染等使得淋巴瘤的发病率逐年升高,早期诊断和精准治疗具有十分重要的临床意义。外泌体是一种脂质双层膜结构的微小囊泡,介导了细胞间交流和信息交换。近几年许多研究证实外泌体是淋巴瘤的发生、进展和耐药的重要机制。外泌体内核酸和小分子可用于淋巴瘤的早期诊断和预测患者预后。材料学修饰可显著增强外泌体治疗的靶向性和治疗效能。本文总结了外泌体生物学特性、分离和鉴定方法、与肿瘤相关性、及其在淋巴瘤中的研究进展,为淋巴瘤的预警和治疗提供参考。     

Detachment of breast tumor cells induces rapid secretion of exosomes which subsequently mediate cellular adhesion and spreading

Exosomes are nano-vesicles secreted by a wide range of mammalian cell types. These vesicles are abundant in serum and other extracellular fluids and contain a large repertoire of proteins, mRNA and microRNA. Exosomes have been implicated in cell to cell communication, the transfer of infectious agents, and neurodegenerative diseases as well as tumor progression. However, the precise mechanisms by which they are internalized and/or secreted remain poorly understood. In order to follow their release and uptake in breast tumor cells in real time, cell-derived exosomes were tagged with green fluorescent protein (GFP)-CD63 while human serum exosomes were rhodamine isothiocynate-labeled. We show that detachment of adherent cells from various substrata induces a rapid and substantial secretion of exosomes, which then concentrate on the cell surfaces and mediate adhesion to various extracellular matrix proteins. We also demonstrate that disruption of lipid rafts with methyl-beta-cyclodextrin (MβCD) inhibits the internalization of exosomes and that annexins are essential for the exosomal uptake mechanisms. Taken together, these data suggest that cellular detachment is accompanied by significant release of exosomes while cellular adhesion and spreading are enhanced by rapid uptake and disposition of exosomes on the cell surface.     

Retrovirus infection strongly enhances scrapie infectivity release in cell culture

Prion diseases are neurodegenerative disorders associated in most cases with the accumulation in the central nervous system of PrPSc (conformationally altered isoform of cellular prion protein (PrPC); Sc for scrapie), a partially protease-resistant isoform of the PrPC. PrPSc is thought to be the causative agent of transmissible spongiform encephalopathies. The mechanisms involved in the intercellular transfer of PrPSc are still enigmatic. Recently, small cellular vesicles of endosomal origin called exosomes have been proposed to contribute to the spread of prions in cell culture models. Retroviruses such as murine leukemia virus (MuLV) or human immunodeficiency virus type 1 (HIV-1) have been shown to assemble and bud into detergent-resistant microdomains and into intracellular compartments such as late endosomes/multivesicular bodies. Here we report that moloney murine leukemia virus (MoMuLV) infection strongly enhances the release of scrapie infectivity in the supernatant of coinfected cells. Under these conditions, we found that PrPC, PrPSc and scrapie infectivity are recruited by both MuLV virions and exosomes. We propose that retroviruses can be important cofactors involved in the spread of the pathological prion agent.     

Physiological and pathological impact of exosomes of adipose tissue

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.     

The role of exosomes in hepatitis,liver cirrhosis and hepatocellular carcinoma

Exosomes are small vesicles that were initially thought to be a mechanism for discarding unneeded membrane proteins from reticulocytes. Their mediation of intercellular communication appears to be associated with several biological functions. Current studies have shown that most mammalian cells undergo the process of exosome formation and utilize exosome‐mediated cell communication. Exosomes contain various microRNAs, mRNAs and proteins. They have been reported to mediate multiple functions, such as antigen presentation, immune escape and tumour progression. This concise review highlights the findings regarding the roles of exosomes in liver diseases, particularly hepatitis B, hepatitis C, liver cirrhosis and hepatocellular carcinoma. However, further elucidation of the contributions of exosomes to intercellular information transmission is needed. The potential medical applications of exosomes in liver diseases seem practical and will depend on the ingenuity of future investigators and their insights into exosome‐mediated biological processes.     

Exosomes as Tools to Suppress Primary Brain Tumor

Exosomes are small microvesicles released by cells that efficiently transfer their molecular cargo to other cells, including tumor. Exosomes may pass the blood–brain barrier and have been demonstrated to deliver RNAs contained within to brain. As they are non-viable, the risk profile of exosomes is thought to be less than that of cellular therapies. Exosomes can be manufactured at scale in culture, and exosomes can be engineered to incorporate therapeutic miRNAs, siRNAs, or chemotherapeutic molecules. As natural biological delivery vehicles, interest in the use of exosomes as therapeutic delivery agents is growing. We previously demonstrated a novel treatment whereby mesenchymal stromal cells were employed to package tumor-suppressing miR-146b into exosomes, which were then used to reduce malignant glioma growth in rat. The use of exosomes to raise the immune system against tumor is also drawing interest. Exosomes from dendritic cells which are antigen-presenting, and have been used for treatment of brain tumor may be divided into three categories: (1) exosomes for immunomodulation-based therapy, (2) exosomes as delivery vehicles for anti-tumor nucleotides, and (3) exosomes as drug delivery vehicles. Here, we will provide an overview of these three applications of exosomes to treat brain tumor, and examine their prospects on the long road to clinical use.     

缺氧条件下中枢神经系统外泌体作用机制

外泌体是来源于细胞内吞噬作用的细胞外囊泡(extracellular vesicles,EVs),其含有特定的蛋白质、脂质、RNA和DNA,能将信号传递给受体细胞,从而介导细胞通讯过程.缺氧作为一种严重的细胞应激,是脑部疾病的重要特征,可以诱导外泌体的释放并影响其内容物.越来越多的证据显示,外泌体携带的生物活性物质可以...     

Exosomes as a novel cell-free therapeutic approach in gastrointestinal diseases

Cell communication through extracellular vesicles (EVs) has been defined for many years and it is not limited only to neighboring cells, but also distant ones in organisms receive these signals. These vesicles are secreted from the variety of cells and are composed of a distinctive component such as proteins, lipids, and nucleic acids. EVs have different classified subgroups regarding their cell origin, in this context, exosomes are the most appealing particles in cell biology, especially clinical in recent years and are represented as novel therapeutic agents with numerous advantages alongside and/or over cell therapy. However, cell therapy had a hopeful outcome in gastrointestinal diseases which have minimal alternatives in their treatments. Inflammatory bowel disease (IBD), liver fibrosis, gastrointestinal cancers are the examples that cell therapy and immunotherapy were applied in their treatment, therefore, the cell products like exosomes are the beneficial option in their treatment even cancers with promising results in animal models. In this review, we consider the main defined biogenesis, function, and component of secreted exosomes in different cells with a specific focus on the potential application of these exosomes as a cell-free therapeutic approach in gastrointestinal diseases like IBD, gastric cancer, and colon cancer. Additionally, exosomes role as therapeutic reagents mainly mesenchymal stem cells and dendritic cell-derived exosomes in different studies have been under intense investigation and even they are being studied in different clinical trials. Therefore, all these striking functions described for secretome implies the importance of these biocarriers.     

Small but significant: Insights and new perspectives of exosomes in cardiovascular disease

Cardiovascular diseases (CVDs) are a major health problem worldwide, and health professionals are still actively seeking new and effective approaches for CVDs treatment. Presently, extracellular vesicles, particularly exosomes, have gained its popularity for CVDs treatment because of their function as messengers for inter‐ and extra‐cellular communications to promote cellular functions in cardiovascular system. However, as a newly developed field, researchers are still trying to fully understand the role of exosomes, and their mechanism in mediating cardiac repair process. Therefore, a comprehensive review of this topic can be timely and favourable. In this review, we summarized the basic biogenesis and characterization of exosomes and then further extended the focus on the circulating exosomes in cellular communication and stem cell‐derived exosomes in cardiac disease treatment. In addition, we covered interactions between the heart and other organs through exosomes, leading to the diagnostic characteristics of exosomes in CVDs. Future perspectives and limitations of exosomes in CVDs were also discussed with a special focus on exploring the potential delivery routes, targeting the injured tissue and engineering novel exosomes, as well as its potential as one novel target in the metabolism‐related puzzle.     

外泌体对睾丸微环境调控机制研究及应用进展

外泌体（exsomes）是一类具有生物活性的双层脂质组成的小囊泡,可携带不同类型的信息物质与受体细胞结合,通过信息传递与物质交换,促发受体细胞的表型发生变化。本文总结了在睾丸微环境中,外泌体通过调节细胞因子促进睾丸微环境中细胞增殖、调节细胞免疫维持睾丸免疫环境、调节氧化应激修复受体细胞功能、调节细胞自噬改善生精能力、调节细胞凋亡改善精子发生、调节细胞因子影响睾酮分泌等机制维持睾丸微环境稳态,并对外泌体在男科相关疾病预防、诊断、治疗中的作用进行归纳,外泌体在男性不育、勃起功能障碍、精索静脉曲张、性腺功能减退、前列腺癌等疾病诊疗中具备明显优势。外泌体作为一种新兴的应用物质,随着外泌体工程和提取工艺的不断优化,以及相关机制研究的不断深入,外泌体在男科疾病中的临床运用成为可能,有望成为治疗男科疾病的新手段。     

Proteomic analysis of melanoma-derived exosomes by two-dimensional polyacrylamide gel electrophoresis and mass spectrometry

Exosomes are 40-100 nm vesicles released by numerous cell types and are thought to have a variety of roles depending on their origin. Exosomes derived from antigen presenting cells have been shown to be capable of initiating immune responses in vivo and eradicating established tumours in murine models. Tumour-derived exosomes can be utilised as a source of tumour antigen for cross-priming to T-cells and are thus of interest for use in anti-tumour immunotherapy. Further exploration into the protein composition of exosomes may increase our understanding of their potential roles in vivo and this study has examined the proteome of exosomes purified from cell supernatants of the melanoma cell lines MeWo and SK-MEL-28. The vesicular nature and size (30-100 nm) of the purified exosomes was confirmed by electron microscopy and sucrose density gradient centrifugation. Western blotting demonstrated the absence of calnexin and cytochrome c, verifying the purity of the exosome preparations, as well as enrichment of MHC class I and the tumour-associated antigens Mart-1 and Mel-CAM. The two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) protein profiles of exosomes from the two cell lines were highly comparable and strikingly different from the profiles of the total cell lysates. Mass spectrometric sequencing identified proteins present in 49 protein spots in the exosome lysates. Several of these have been identified previously in exosomes but some are novel, including p120 catenin, radixin, and immunoglobulin superfamily member 8 (PGRL). Proteins present in whole-cell lysates that were significantly reduced or excluded from exosomes were also identified and included several mitochondrial and lysosomal proteins, again confirming the proposed endosomal origin of exosomes. This study presents a starting point for future more in-depth protein studies of tumour-derived exosomes which will aid the understanding of their biogenesis and targeting for use in anti-tumour immunotherapy protocols.     

Application of mesenchymal stem cell exosomes and their drug‐loading systems in acute liver failure

Stem cell exosomes are nanoscale membrane vesicles released from stem cells of various origins that can regulate signal transduction pathways between liver cells, and their functions in intercellular communication have been recognized. Due to their natural substance transport properties and excellent biocompatibility, exosomes can also be used as drug carriers to release a variety of substances, which has great prospects in the treatment of critical and incurable diseases. Different types of stem cell exosomes have been used to study liver diseases. Due to current difficulties in the treatment of acute liver failure (ALF), this review will outline the potential of stem cell exosomes for ALF treatment. Specifically, we reviewed the pathogenesis of acute liver failure and the latest progress in the use of stem cell exosomes in the treatment of ALF, including the role of exosomes in inhibiting the ALF inflammatory response and regulating signal transduction pathways, the advantages of stem cell exosomes and their use as a drug‐loading system, and their pre‐clinical application in the treatment of ALF. Finally, the clinical research status of stem cell therapy for ALF and the current challenges of exosome clinical transformation are summarized.     

Characterization of Human Thymic Exosomes

Exosomes are nanosized membrane-bound vesicles that are released by various cell types and are capable of carrying proteins, lipids and RNAs which can be delivered to recipient cells. Exosomes play a role in intercellular communication and have been described to mediate immunologic information. In this article we report the first isolation and characterization of exosomes from human thymic tissue. Using electron microscopy, particle size determination, density gradient measurement, flow cytometry, proteomic analysis and microRNA profiling we describe the morphology, size, density, protein composition and microRNA content of human thymic exosomes. The thymic exosomes share characteristics with previously described exosomes such as antigen presentation molecules, but they also exhibit thymus specific features regarding surface markers, protein content and microRNA profile. Interestingly, thymic exosomes carry proteins that have a tissue restricted expression in the periphery which may suggest a role in T cell selection and the induction of central tolerance. We speculate that thymic exosomes may provide the means for intercellular information exchange necessary for negative selection and regulatory T cell formation of the developing thymocytes within the human thymic medulla.     

Exosome target cell selection and the importance of exosomal tetraspanins: a hypothesis

Exosomes are derived from limiting membranes of MVBs (multivesicular bodies). They carry and transfer selected membrane and cytoplasmic proteins, mRNA and microRNA into target cells. It is due to this shipping of information that exosomes are considered to be the most promising therapeutic tool for multiple diseases. However, whereas knowledge on the composition of exosomes is rapidly increasing, the mode of selective recruitment into exosomes as well as target cell selection is poorly understood. We suggest that at least part of this task is taken over by tetraspanins. Tetraspanins, which are involved in morphogenesis, fission and fusion processes, are enriched in exosomes, and our previous work revealed that the recruitment of distinct tetraspanins into exosomes follows very selective routes, including a rearrangement of the tetraspanin web. Furthermore, only exosomes expressing a defined set of tetraspanins and associated molecules target endothelial cells, thereby contributing to angiogenesis and vasculogenesis. On the basis of these findings we hypothesize (i) that the protein assembly of exosomes and possibly the recruitment of microRNA will be regulated to a large extent by tetraspanins and (ii) that tetraspanins account for target cell selection and the tight interaction/uptake of exosomes by the target cell. Exosomes herald an unanticipated powerful path of cell-cell communication. An answer to how exosomes collect and transfer information will allow the use of Nature's concept to cope with malfunctions.