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.     

From tumor microenvironment communicants to biomarker discovery: Selectively packaged extracellular vesicular cargoes in pancreatic cancer

Virtually all cells release various types of vesicles into the extracellular environment. These extracellular vesicles (EVs) transport molecular cargoes, performing as communicants for information exchange both within the tumor microenvironment (TME) and to distant organs. Thus, understanding the selective packaging of EV cargoes and the mechanistic impact of those cargoes - including metabolites, lipids, proteins, and/or nucleic acids - offers an opportunity to increase our knowledge of cancer biology and identify EV cargoes that might serve as cancer biomarkers in blood, saliva, or urine samples. In this review, we collect and organize recent advances in this field with an emphasis on pancreatic cancer (pancreatic adenocarcinoma, PDAC) and the concept that cells selectively package cargo into EVs. These studies demonstrate PDAC EV cargoes signal to reprogram and remodel the TME and impact distant organs. EV cargoes identified as potential PDAC diagnostic and prognostic biomarkers are summarized.     

Extracellular vesicles – biogenesis,composition, function,uptake and therapeutic applications

Extracellular vesicles are heterogenous, nano-sized, membrane-limited structures which not only represent a specific way of intercellular communication but also endow cells with many capabilities. Apoptotic bodies created during apoptosis, microvesicles shed from the plasma membrane, and exosomes originated inside the cell and released to extracellular space upon fusion with the plasma membrane, they all belong to extracellular vesicles. Extracellular vesicles contain lipids, proteins, and nucleic acids. In this review, we describe their biogenesis, release and uptake by recipient cells, their composition, functions and potential therapeutic and diagnostic applications.     

Extracellular vesicles and pancreatitis: mechanisms,status and perspectives

Comprehensive reviews and large population-based cohort studies have played an important role in the diagnosis and treatment of pancreatitis and its sequelae. The incidence and mortality of pancreatitis have been reduced significantly due to substantial advancements in the pathophysiological mechanisms and clinically effective treatments. The study of extracellular vesicles (EVs) has the potential to identify cell-to-cell communication in diseases such as pancreatitis. Exosomes are a subset of EVs with an average diameter of 50~150 nm. Their diverse and unique constituents include nucleic acids, proteins, and lipids, which can be transferred to trigger phenotypic changes of recipient cells. In recent years, many reports have indicated the role of EVs in pancreatitis, including acute pancreatitis, chronic pancreatitis and autoimmune pancreatitis, suggesting their potential influence on the development and progression of pancreatitis. Plasma exosomes of acute pancreatitis can effectively reach the alveolar cavity and activate alveolar macrophages to cause acute lung injury. Furthermore, upregulated exosomal miRNAs can be used as biomarkers for acute pancreatitis. Here, we summarized the current understanding of EVs in pancreatitis with an emphasis on their biological roles and their potential use as diagnostic biomarkers and therapeutic agents for this disease.     

Extracellular vesicles in urologic malignancies—Implementations for future cancer care

Extracellular vesicles (EVs), a heterogeneous group of vesicles differing in size and shape, cargo content and function, are membrane‐bound and nano‐sized vesicles that could be released by nearly all variations of cells. EVs have gained considerable attention in the past decades for their functions in modulating intercellular signalling and roles as potential pools for the novel diagnostic and prognostic biomarkers, as well as therapeutic targets in several cancers including urological neoplasms. In general, human and animal cells both can release distinct types of EVs, including exosomes, microvesicles, oncosomes and large oncosomes, and apoptotic bodies, while the content of EVs can be divided into proteins, lipids and nucleic acids. However, the lack of standard methods for isolation and detection platforms rein the widespread usage in clinical applications warranted furthermore investigations in the development of reliable, specific and sensitive isolation techniques. Whether and how the EVs work has become pertinent issues. With the aid of high‐throughput proteomics or genomics methods, a fully understanding of contents contained in EVs from urogenital tumours, beyond all doubt, will improve our ability to identify the complex genomic alterations in the process of cancer and, in turn, contribute to detect potential therapeutic target and then provide personalization strategy for patient.     

Multiplexed targeting of microRNA in stem cell-derived extracellular vesicles for regenerative medicine

Regenerative medicine is a research field that develops methods to restore damaged cell or tissue function by regeneration, repair or replacement. Stem cells are the raw material of the body that is ultimately used from the point of view of regenerative medicine, and stem cell therapy uses cells themselves or their derivatives to promote responses to diseases and dysfunctions, the ultimate goal of regenerative medicine. Stem cell-derived extracellular vesicles (EVs) are recognized as an attractive source because they can enrich exogenous microRNAs (miRNAs) by targeting pathological recipient cells for disease therapy and can overcome the obstacles faced by current cell therapy agents. However, there are some limitations that need to be addressed before using miRNA-enriched EVs derived from stem cells for multiplexed therapeutic targeting in many diseases. Here, we review various roles on miRNA-based stem cell EVs that can induce effective and stable functional improvement of stem cell-derived EVs. In addition, we introduce and review the implications of several miRNA-enriched EV therapies improved by multiplexed targeting in diseases involving the circulatory system and nervous system. This systemic review may offer potential roles for stem cell-derived therapeutics with multiplexed targeting.     

Mesenchymal stromal cell‐derived extracellular vesicles as cell‐free biologics for the ex vivo expansion of hematopoietic stem cells

Hematopoietic stem cell transplantation (HSCT) is the ultimate choice of treatment for patients with hematological diseases and cancer. The success of HSCT is critically dependent on the number and engraftment efficiency of the transplanted donor hematopoietic stem cells (HSCs). Various studies show that bone marrow‐derived mesenchymal stromal cells (MSCs) support hematopoiesis and also promote ex vivo expansion of HSCs. MSCs exert their therapeutic effect through paracrine activity, partially mediated through extracellular vesicles (EVs). Although the physiological function of EVs is not fully understood, inspiring findings indicate that MSC‐derived EVs can reiterate the hematopoiesis, supporting the ability of MSCs by transferring their cargo containing proteins, lipids, and nucleic acids to the HSCs. The activation state of the MSCs or the signaling mechanism that prevails in them also defines the composition of their EVs, thereby influencing the fate of HSCs. Modulating or preconditioning MSCs to achieve a specific composition of the EV cargo for the ex vivo expansion of HSCs is, therefore, a promising strategy that can overcome several challenges associated with the use of naïve/unprimed MSCs. This review aims to speculate upon the potential role of preconditioned/primed MSC‐derived EVs as “cell‐free biologics,” as a novel strategy for expanding HSCs in vitro.     

How does temperature play a role in the storage of extracellular vesicles?

Extracellular vesicles (EVs) contain specific proteins, lipids, and nucleic acids that can be passed to other cells as signal molecules to alter their function. However, there are many problems and challenges in the conversion and clinical application of EVs. Storage and protection of EVs is one of the issues that need further research. To adapt to potential clinical applications, this type of problem must be solved. This review summarizes the storage practices of EVs in recent years, and explains the impact of temperature on the quality and stability of EVs during storage based on current research, and explains the potential mechanisms involved in this effect as much as possible.     

Transferring intercellular signals and traits between cancer cells: extracellular vesicles as “homing pigeons”

Extracellular vesicles are cell-derived vesicles, which can transport various cargos out of cells. From their cell of origin, the content molecules (proteins, non-coding RNAs including miRNAs, DNA and others) can be delivered to neighboring or distant cells and as such extracellular vesicles can be regarded as vehicles of intercellular communication or “homing pigeons”. Extracellular vesicle shuttling is able to actively modulate the tumor microenvironment and can partake in tumor dissemination. In various diseases, including cancer, levels of extracellular vesicle secretion are altered resulting in different amounts and/or profiles of detectable vesicular cargo molecules and these distinct content profiles are currently being evaluated as biomarkers. Apart from their potential as blood-derived containers of specific biomarkers, the transfer of extracellular vesicles to surrounding cells also appears to be involved in the propagation of phenotypic traits. These interesting properties have put extracellular vesicles into the focus of many recent studies.Here we review findings on the involvement of extracellular vesicles in transferring traits of cancer cells to their surroundings and briefly discuss new data on oncosomes, a larger type of vesicle. A pressing issue in cancer treatment is rapidly evolving resistance to many initially efficient drug therapies. Studies investigating the role of extracellular vesicles in this phenomenon together with a summary of the technical challenges that this field is still facing, are also presented. Finally, emerging areas of research such as the analysis of the lipid composition on extracellular vesicles and cutting-edge techniques to visualise the trafficking of extracellular vesicles are discussed.     

Oncogenic Regulation of Extracellular Vesicle Proteome and Heterogeneity

Mutational and epigenetic driver events profoundly alter intercellular communication pathways in cancer. This effect includes deregulated release, molecular composition, and biological activity of extracellular vesicles (EVs), membranous cellular fragments ranging from a few microns to less than 100 nm in diameter and filled with bioactive molecular cargo (proteins, lipids, and nucleic acids). While EVs are usually classified on the basis of their physical properties and biogenetic mechanisms, recent analyses of their proteome suggest a larger than expected molecular diversity, a notion that is also supported by multicolour nano‐flow cytometry and other emerging technology platforms designed to analyze single EVs. Both protein composition and EV diversity are markedly altered by oncogenic transformation, epithelial to mesenchymal transition, and differentiation of cancer stem cells. Interestingly, only a subset of EVs released from mutant cells may carry oncogenic proteins (e.g., EGFRvIII), hence, these EVs are often referred to as “oncosomes”. Indeed, oncogenic transformation alters the repertoire of EV‐associated proteins, increases the presence of pro‐invasive cargo, and alters the composition of distinct EV populations. Molecular profiling of single EVs may reveal a more intricate effect of transforming events on the architecture of EV populations in cancer and shed new light on their biological role and diagnostic utility.     

Extracellular heat shock proteins and cancer: New perspectives

Heat shock proteins (HSPs) are a large family of molecular chaperones aberrantly expressed in cancer. The expression of HSPs in tumor cells has been shown to be implicated in the regulation of apoptosis, immune responses, angiogenesis and metastasis. Given that extracellular vesicles (EVs) can serve as potential source for the discovery of clinically useful biomarkers and therapeutic targets, it is of particular interest to study proteomic profiling of HSPs in EVs derived from various biological fluids of cancer patients. Furthermore, a divergent expression of circulating microRNAs (miRNAs) in patient samples has opened new opportunities in exploiting miRNAs as diagnostic tools. Herein, we address the current literature on the expression of extracellular HSPs with particular interest in HSPs in EVs derived from various biological fluids of cancer patients and different types of immune cells as promising targets for identification of clinical biomarkers of cancer. We also discuss the emerging role of miRNAs in HSP regulation for the discovery of blood-based biomarkers of cancer. We outline the importance of understanding relationships between various HSP networks and co-chaperones and propose the model for identification of HSP signatures in cancer. Elucidating the role of HSPs in EVs from the proteomic and miRNAs perspectives may provide new opportunities for the discovery of novel biomarkers of cancer.     

Large extracellular vesicles: Size matters in tumor progression

Extracellular Vesicles (EVs) represent a heterogeneous population of particles naturally released from all cells, delimited by a lipid bilayer and able to horizontally transfer their cargos to recipient cells. These features imply the growing interest on EVs in cancer biology as biomarkers and therapeutic targets. In this review, we will highlight the specific process related to biogenesis and release of large EVs (L-EVs) derived from the plasma membrane (PM) compared to the small and well described exosomes, generated through the classical endosome-multivesicular body (MVB) pathway. The control of PM rigidity by cells depends on lipid/protein composition, cytoskeleton dynamics, cytoplasmic viscosity, ions balance, metabolic reprogramming and specific intracellular signaling pathways, all critical determinants of L-EVs biogenesis. We will focus in details on a specific class of L-EVs, named Large Oncosomes (LO), exclusively shed by cancer cells and with a size ranging from 1 μm up to 10 μm. We will examine LO specific cargos, either proteins or nucleic acids (i.e. mRNA, microRNAs, single/double-stranded DNA), as well as their functional role in cancer development and progression, also discussing the mechanisms of L-EVs internalization by recipient cells. Overall we will highlight the potential of LO as specific diagnostic/prognostic cancer biomarkers discussing the associated challenges.     

The key role of extracellular vesicles in the metastatic process

Extracellular vesicles (EVs), including exosomes, have a key role in the paracrine communication between organs and compartments. EVs shuttle virtually all types of biomolecules such as proteins, lipids, nucleic acids, metabolites and even pharmacological compounds. Their ability to transfer their biomolecular cargo into target cells enables EVs to play a key role in intercellular communication that can regulate cellular functions such as proliferation, apoptosis and migration. This has led to the emergence of EVs as a key player in tumor growth and metastasis through the formation of “tumor niches” in target organs. Recent data have also been shown that EVs may transform the microenvironment of primary tumors thus favoring the selection of cancer cells with a metastatic behavior. The release of EVs from resident non-malignant cells may contribute to the metastatic processes as well. However, cancer EVs may induce malignant transformation in resident mesenchymal stem cells, suggesting that the metastatic process is not exclusively due to circulating tumor cells. In this review, we outline and discuss evidence-based roles of EVs in actively regulating multiple steps of the metastatic process and how we can leverage EVs to impair metastasis.     

Exploring the utility of extracellular vesicles in ameliorating viral infection-associated inflammation,cytokine storm and tissue damage

Extracellular vesicles (EVs) have emerged as potential mediators of intercellular communication. EVs are nano-sized, lipid membrane–bound vesicles that contains biological information in the form of proteins, metabolites and/or nucleic acids. EVs are key regulators of tissue repair mechanisms, such as in the context of lung injuries. Recent studies suggest that EVs have the ability to repair COVID19-associated acute lung damage. EVs hold great promise for therapeutic treatments, particularly in treating a potentially fatal autoimmune response and attenuate inflammation. They are known to boost lung immunity and are involved in the pathogenesis of various lung diseases, including viral infection. EV-based immunization technology has been proven to elicit robust immune responses in many models of infectious disease, including COVID-19. The field of EV research has tremendous potential in advancing our understanding about viral infection pathogenesis, and can be translated into anti-viral therapeutic strategies.     

Breast cancer-derived exosomes: Tumor progression and therapeutic agents

Tumor cells secrete extracellular vesicles (EVs) for intercellular communication. EVs by transporting different proteins, nucleic acids, and lipids contribute to affect target cell function and fate. ‎EVs which originate directly from multivesicular bodies so-called exosomes have dramatically fascinated the attention of researchers owing to their ‎pivotal roles in the tumorigenesis. Breast cancer, arising from milk-producing cells, is the most identified cancer among women and has become the leading cause of cancer-related death in women globally. Although different therapies are applied to eliminate breast tumor cells, however, the efficient therapy and survival rate of patients remain challenges. Growing evidence ‎shows exosomes from breast cancer cells contribute to proliferation, metastasis, angiogenesis, chemoresistance, and also radioresistance and, thus carcinogenesis. Additionally, these exosomes may serve as a cancer treatment tool because they are a good candidate for cancer diagnosis (as biomarker) and therapy (as drug-carrier). Despite recent development in the biology of tumor-derived exosomes, the detailed mechanism of tumorigenesis, and exosome-based cancer-therapy remain still indefinable. Here, we discuss the key function of breast cancer-derived exosomes in tumorgenesis and shed light on the possible clinical application of these exosomes in breast cancer treatment.     

The therapeutic potential of immune cell-derived exosomes as an alternative to adoptive cell transfer

Adoptive cell transfer (ACT), a form of cell-based immunotherapy that eliminates cancer by restoring and strengthening the body’s immune system, has revolutionized cancer treatment. ACT entails intravenous transfer of either tumor-resident or peripheral blood-modified immune cells into cancer patients to mediate anti-tumor response. Although these immune cells control and eradicate cancer via enhanced cytotoxicity against specific tumor antigens, several side effects have been frequently reported in clinical trials. Recently, exosomes, potential cell-free therapeutics, have emerged as an alternative to cell-based immunotherapies, due to their higher stability under same storage condition, lower risk of GvHD and CRS, and higher resistance to immunosuppressive tumor microenvironment. Exosomes, which are nano-sized lipid vesicles, are secreted by living cells, including immune cells. Exosomes contain proteins, lipids, and nucleic acids, and the functional role of each exosome is determined by the specific cargo derived from parental cells. Exosomes derived from cytotoxic effectors including T cells and NK cells exert anti-tumor effects via proteins such as granzyme B and FasL. In this mini-review, we describe the current understanding of the ACT and immune cell-derived exosomes and discuss the limitations of ACT and the opportunities for immune cell-derived exosomes as immune therapies.     

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

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

Extracellular vesicles-released parathyroid hormone-related protein from Lewis lung carcinoma induces lipolysis and adipose tissue browning in cancer cachexia

Cancer cachexia is a metabolic disorder characterized by skeletal muscle wasting and white adipose tissue browning. Specific functions of several hormones, growth factors, and cytokines derived from tumors can trigger cachexia. Moreover, adipose tissue lipolysis might explain weight loss that occurs owing to cachexia. Extracellular vesicles (EVs) are involved in intercellular communication. However, whether EVs participate in lipolysis induced by cancer cachexia has not been thoroughly investigated. Using Lewis lung carcinoma (LLC) cell culture, we tested whether LLC cell-derived EVs can induce lipolysis in 3T3-L1 adipocytes. EVs derived from LLC cells were isolated and characterized biochemically and biophysically. Western blotting and glycerol assay were used to study lipolysis. LLC cell-derived EVs induced lipolysis in vivo and vitro. EVs fused directly with target 3T3-L1 adipocytes and transferred parathyroid hormone-related protein (PTHrP), activating the PKA signaling pathway in 3T3-L1 adipocytes. Blocking PTHrP activity in LLC-EVs using a neutralizing antibody and by knocking down PTHR expression prevented lipolysis in adipocytes. Inhibiting the PKA signaling pathway also prevents the lipolytic effects of EVs. In vivo, suppression of LLC-EVs release by knocking down Rab27A alleviated white adipose tissue browning and lipolysis. Our data showed that LLC cell-derived EVs induced adipocyte lipolysis via the extracellular PTHrP-mediated PKA pathway. Our data demonstrate that LLC-EVs induce lipolysis in vitro and vivo by delivering PTHrP, which interacts with PTHR. The lipolytic effect of LLC-EVs was abrogated by PTHR knockdown and treatment with a neutralizing anti-PTHrP antibody. Together, these data show that LLC-EV-induced lipolysis is mediated by extracellular PTHrP. These findings suggest a novel mechanism of lipid droplet loss and identify a potential therapeutic strategy for cancer cachexia.Subject terms: Oncogenes, Mechanisms of disease     

哺乳动物精浆胞外囊泡及各组分的功能

精浆胞外囊泡是一种存在于精浆的膜性囊泡,按分泌器官分为附睾小体和前列腺小体。囊泡可与精子细胞膜发生融合,通过传递内容物或介导信号通路进而调节精子功能。它含有多种活性物质,其中蛋白质组分可影响精子活力以及顶体反应,并有清除损伤精子和促进细胞粘附的作用;脂质组分具有调节靶细胞质膜稳定性的作用;核酸组分主要参与免疫反应、跨代遗传及男性不育;离子则是多种酶的辅助因子,在调节酶活性和精浆微环境中发挥重要作用。不同组分对精子功能的影响不尽相同,本文将对此方面的研究进展进行详尽的综述,以期为该领域相关研究人员提供一定的参考。