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INTRODUCTION: Recto-colic graft-versus-host disease (GVHD) is a frequent and serious complication of hematopoietic stem cell allogeneic transplantation, which is sometimes difficult to diagnose. The aim of our study was to identify histological diagnostic and prognostic criteria for recto-colic GVH. MATERIAL AND METHOD: Patients allografted at Amiens university hospital from 2012 to 2017 were retrieved. Those who had a recto-colic biopsy were included and divided into two groups (final diagnosis of GVH and non-GVH), then biopsies were reviewed by 2 pathologists. RESULTS: One hundred and nineteen patients were included. Sixty-seven were allocated to the GVH group and 52 to the non-GVH group. In the GVH group, we observed a significantly greater number of apoptotic bodies (AB) on standard HES staining and with the anti-Caspase 3 immunohistochemistry, cryptolytic AB abscesses, atrophy, regenerative glands and glands lined with eosinophilic cells (P<0.001). Anti-Caspase 3 immunohistochemistry revealed more AB than standard HES staining (P<0.005). But to differentiate GVH cases from non-GVH cases, we obtained a threshold value of 3.5 AB per 10 contiguous crypts on standard HE staining and with the anti-Caspase 3 immunohistochemistry. From 4 AB per 10 contiguous crypts, on HES staining and anti-Caspase 3 immunostaining, the diagnosis of GVH became consistent. No non-GVH case had more than 6 AB per 10 contiguous crypts. GVH patients with more than 8 AB per 10 contiguous crypts had a worse prognosis (P<0.001). CONCLUSION: We confirm the value of AB and their counting in the diagnosis of GVH, with a diagnostic threshold of 4 AB and a prognostic threshold of 8 AB. Glands lined with eosinophilic cells could be an additional diagnostic criterion in favor of GVH to be confirmed by further studies.

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Apoptotic bodies (ABs) are extracellular vesicles released during apoptosis and possess diverse biological activities. Initially, ABs were regarded as garbage bags with the main function of apoptotic cell clearance. Recent research has found that ABs carry and deliver various biological agents and are taken by surrounding and distant cells, affecting cell functions and behavior. ABs-mediated intercellular communications are involved in various physiological processes including anti-inflammation and tissue regeneration as well as the pathogenesis of a variety of diseases including cancer, cardiovascular diseases, neurodegeneration, and inflammatory diseases. ABs in biological fluids can be used as a window of altered cellular and tissue states which can be applied in the diagnosis and prognosis of various diseases. The structural and constituent versatility of ABs provides flexibility for tailoring ABs according to disease diagnostic and therapeutic needs. An in-depth understanding of ABs' constituents and biological functions is mandatory for the effective tailoring of ABs including modification of bio membrane and cargo constituents. ABs' tailoring approaches including physical, chemical, biological, and genetic have been proposed for bench-to-bed translation in disease diagnosis, prognosis, and therapy. This review summarizes the updates on ABs tailoring approaches, discusses the existing challenges, and speculates the prospects for effective diagnostic and therapeutic applications.

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Apoptosis is the most prominent mode of programmed cell death and is necessary for the maintenance of tissue homeostasis. During cell apoptosis, a distinctive population of extracellular vesicles is generated, termed apoptotic vesicles (apoVs). ApoVs inherit a variety of biological molecules such as proteins, RNAs, nuclear components, lipids, and gasotransmitters from their parent cells. ApoVs have shown promising therapeutic potential for inflammation, tumors, immune disorders, and tissue regeneration. In addition, apoVs can be used as drug carriers, vaccine development, and disease diagnosis. Recently, apoVs have been used in clinical trials to treat a variety of diseases, such as temporomandibular joint osteoarthritis and the regeneration of functional alveolar bone. Here, we review the history of apoV research, current preclinical and clinical studies, and the potential issues of apoV application.

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Alcohol ingestion is a widespread habituation that evolved along with a growing population, altering physiological conditions through immunomodulatory function. There is much research that has reported that consumption of alcohol at low and heavy levels causes different biological impacts, including cellular injury, leading to systemic dysfunction and increased inflammatory markers. In the fate of professional phagocytic cells, efferocytosis is an inevitable mechanism activated by the apoptotic cells, thus eliminating them and preventing the accumulation of cell corpses/debris in the microenvironment. Subsequently, it promotes the tissue repair mechanism and maintains cellular homeostasis. Unfortunately, defective efferocytosis is widely found in several inflammatory and age-related diseases such as atherosclerosis, autoimmune diseases, lung injury, fatty liver disease, and neurodegenerative diseases. Alcohol abuse is one of the factors that provoke an immune response that increases the rate of morbidity and mortality in parallel in systemic disease patients. Information regarding the emergence of immunomodulation during alcoholic pathogenesis and its association with efferocytosis impairment remain elusive. Hence, here in this review, we discussed the mechanism of efferocytosis, the role of defective efferocytosis in inflammatory diseases, and the role of alcohol on efferocytosis impairment.

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Evidence demonstrating the involvement of apoptosis in the death of the potentially salvageable area (penumbra zone) in patients during stroke remains limited. Our aim was to investigate whether apoptotic processes occur in penumbral brain tissue by analyzing circulating neuron- and glia-derived apoptotic bodies (CNS-ApBs), which are vesicles released into the bloodstream during the late stage of apoptosis. We have also assessed the clinical utility of plasma neuronal and glial apoptotic bodies in predicting early neurological evolution and functional outcome. The study included a total of 71 patients with acute hemispheric ischemic stroke (73 ± 10 years; 30 women). Blood samples were collected from these patients immediately upon arrival at the hospital (within 9 h) and at 24 and 72 h after symptom onset. Subsequently, isolation, quantification, and phenotypic characterization of CNS-ApBs during the first 72 h post-stroke were performed using centrifugation and flow cytometry techniques. We found a correlation between infarct growth and final infarct size with the amount of plasma CNS-ApBs detected in the first 72 h after stroke. In addition, patients with neurological worsening (progressive ischemic stroke) had higher plasma levels of CNS-ApBs at 24 h after symptom onset than those with a stable or improving course. Circulating CNS-ApB concentration was further associated with patients' functional prognosis. In conclusion, apoptosis may play an important role in the growth of the cerebral infarct area and plasma CNS-ApB quantification could be used as a predictive marker of penumbra death, neurological deterioration, and functional outcome in patients with ischemic stroke.

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In the realm of large-area trauma flap transplantation, averting ischaemic necrosis emerges as a pivotal concern. Several key mechanisms, including the promotion of angiogenesis, the inhibition of oxidative stress, the suppression of cell death, and the mitigation of inflammation, are crucial for enhancing skin flap survival. Apoptotic bodies (ABs), arising from cell apoptosis, have recently emerged as significant contributors to these functions. This study engineered three-dimensional (3D)-ABs using tissue-like mouse adipose-derived stem cells (mADSCs) cultured in a 3D environment to compare their superior biological effects against 2D-ABs in bolstering skin flap survival. The findings reveal that 3D-ABs (85.74 ± 4.51) % outperform 2D-ABs (76.48 ± 5.04) % in enhancing the survival rate of ischaemic skin flaps (60.45 ± 8.95) % (all p < 0.05). Mechanistically, they stimulated angiogenesis, mitigated oxidative stress, suppressed apoptosis, and facilitated the transition of macrophages from M1 to M2 polarization (all p < 0.05). A comparative analysis of microRNA (miRNA) profiles in 3D- and 2D-ABs identified several specific miRNAs (miR-423-5p-up, miR30b-5p-down, etc.) with pertinent roles. In summary, ABs derived from mADSCs cultured in a 3D spheroid-like arrangement exhibit heightened biological activity compared to those from 2D-cultured mADSCs and are more effective in promoting ischaemic skin flap survival. These effects are attributed to their influence on specific miRNAs.

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Apoptosis releases numerous apoptotic vesicles that regulate processes such as cell proliferation, immunity, and tissue regeneration and repair. Now, it has also emerged as an attractive candidate for biotherapeutics. However, apoptotic vesicles encompass a diverse range of subtypes, and it remains unclear which specific subtypes play a pivotal role. In this study, we successfully isolated different apoptotic vesicle subtypes based on their sizes and characterized them using NTA and TEM techniques, respectively. We compared the functional variances among the distinct subtypes of apoptotic vesicles in terms of stem cell proliferation, migration, and differentiation, as well as for endothelial cell and macrophage function, effectively identifying subtypes that exhibit discernible functional differences. ApoSEV (with diameter <1000 nm) promoted stem cell proliferation, migration, and multi-potent differentiation, and accelerated skin wound healing of diabetes mouse model, while apoBD (with diameter >1000 nm) played the opposite effect on cell function and tissue regeneration. Lastly, employing protein analysis and gene sequencing techniques, we elucidated the intrinsic mechanisms underlying these differences between different subtypes of apoEVs. Collectively, this study identified that apoptotic vesicle subtypes possessed distinct bio-functions in regulating stem cell function and behaviour and modulating tissue regeneration, which primarily attribute to the distinct profiling of protein and mRNA in different subtypes. This comprehensive analysis of specific subtypes of apoEVs would provide novel insights for potential therapeutic applications in cell biology and tissue regeneration.

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Preventing and treating avascular necrosis at the distal end of the flaps are critical to surgery success, but current treatments are not ideal. A recent study shows that apoptotic bodies (ABs) generated near the site of apoptosis can be taken up and promote cell proliferation. The study reveals that ABs derived from fibroblast-like cells in the subcutaneous connective tissue (FSCT cells) of skin flaps promoted ischaemic flap survival. It is also found that ABs inhibited cell death and oxidative stress and promoted M1-to-M2 polarization in macrophages. Transcriptome sequencing and protein level testing demonstrated that ABs promoted ischaemic flap survival in endothelial cells and macrophages by inhibiting ferroptosis via the KEAP1-Nrf2 axis. Furthermore, microRNA (miR) sequencing data and in vitro and in vivo experiments demonstrated that ABs inhibited KEAP1 by delivering miR-339-5p to exert therapeutic effects. In conclusion, FSCT cell-derived ABs inhibited ferroptosis, promoted the macrophage M1-to-M2 transition via the miR-339-5p/KEAP1/Nrf2 axis and promoted ischaemic flap survival. These results provide a potential therapeutic strategy to promote ischaemic flap survival by administering ABs.

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From the moment a cell is on the path to malignant transformation, its interaction with other cells from the microenvironment becomes altered. The flow of molecular information is at the heart of the cellular and systemic fate in tumors, and various processes participate in conveying key molecular information from or to certain cancer cells. For instance, the loss of tight junction molecules is part of the signal sent to cancer cells so that they are no longer bound to the primary tumors and are thus free to travel and metastasize. Upon the targeting of a single cell by a therapeutic drug, gap junctions are able to communicate death information to by-standing cells. The discovery of the importance of novel modes of cell-cell communication such as different types of extracellular vesicles or tunneling nanotubes is changing the way scientists look at these processes. However, are they all actively involved in different contexts at the same time or are they recruited to fulfill specific tasks? What does the multiplicity of modes mean for the overall progression of the disease? Here, we extend an open invitation to think about the overall significance of these questions, rather than engage in an elusive attempt at a systematic repertory of the mechanisms at play.

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BACKGROUND: Adipose tissue-derived stem cell-derived apoptotic bodies (ADSC-ABs) have shown great potential for immunomodulation and regeneration, particularly in diabetic wound therapy. However, their local application has been limited by unclear regulatory mechanisms, rapid clearance, and short tissue retention times. METHODS: We analyzed the key role molecules and regulatory pathways of ADSC-ABs in regulating inflammatory macrophages by mRNA sequencing and microRNA (miRNA) sequencing and then verified them by gene knockdown. To prevent rapid clearance, we employed microfluidics technology to prepare methacrylate-anhydride gelatin (GelMA) microspheres (GMS) for controlled release of ABs. Finally, we evaluated the effectiveness of ADSC-AB-laden GMSs (ABs@GMSs) in a diabetic rat wound model. FINDINGS: Our results demonstrated that ADSC-ABs effectively balanced macrophage inflammatory polarization through the janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway, mediated by miR-20a-5p. Furthermore, we showed that AB@GMSs had good biocompatibility, significantly delayed local clearance of ABs, and ameliorated diabetic wound inflammation and promoted vascularization, thus facilitating its healing. CONCLUSIONS: Our study reveals the regulatory mechanism of ADSC-ABs in balancing macrophage inflammatory polarization and highlightsthe importance of delaying their local clearance by GMSs. These findings have important implications for the development of novel therapies for diabetic wound healing. FUNDING: This research was supported by the National Key Research and Development Program of China (2020YFA0908200), National Natural Science Foundation of China (82272263, 82002053, 32000937, and 82202467), Shanghai "Rising Stars of Medical Talents" Youth Development Program (22MC1940300), Shanghai Municipal Health Commission (20204Y0354), and Shanghai Science and Technology Development Funds (22YF1421400).

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Angiogenesis, the process of new blood vessels formation from existing vasculature, plays a vital role in development, wound healing, and various pathophysiological conditions. In recent years, extracellular vesicles (EVs) have emerged as crucial mediators in intercellular communication and have gained significant attention for their role in modulating angiogenic processes. This review explores the multifaceted role of EVs in angiogenesis and their capacity to modulate angiogenic signaling pathways. Through comprehensive analysis of a vast body of literature, this review highlights the potential of utilizing EVs as therapeutic tools to modulate angiogenesis for both physiological and pathological purposes. A good understanding of these concepts holds promise for the development of novel therapeutic interventions targeting angiogenesis-related disorders.

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BACKGROUND: Medullary thyroid carcinoma (MTC) is a rare cancer of parafollicular C-cell origin. The International MTC Grading System (IMTCGS) incorporates mitotic activity, the presence of necrosis, and the Ki67 proliferation rate (PR) to classify MTCs as low or high grade. The ability to predict IMTCGS grade in cytology was assessed. METHODS: MTCs with cytology and subsequent surgical follow-up were reviewed. Cytology slides were reviewed for mitotic figures, apoptoses, and necrosis, and a Ki67 PR was calculated when possible. Findings were correlated with final IMTCGS grade. RESULTS: Twenty-five MTC fine-needle aspirations (FNAs) were identified, with nine identified as high grade (36%). By using a PR cutoff of 5%, Ki67 on FNA material (Ki67FNA) showed 92% concordance (n = 22 of 24) with surgical Ki67 and a correlation coefficient (R2) of 0.72. Sensitivity and specificity of Ki67FNA for predicting high-grade MTC were 38% and 100%, respectively. Multiple mitotic figures were present in a single slide of 43% (n = 3 of 7) of evaluable high-grade MTCs, whereas only one of 16 low-grade MTCs showed a single mitotic figure. Definitive apoptoses were present in five of seven high-grade MTC FNAs but were absent in 16 low-grade MTCs. The sensitivity and specificity of apoptoses/necrosis on cytology for high-grade MTCs were 71% and 88%, respectively. CONCLUSIONS: Ki67FNA ≥5% shows low sensitivity but high specificity for predicting high-grade MTC. The presence of multiple mitotic figures in a single slide or definitive apoptotic bodies are both highly suggestive of high-grade MTC, and should warrant a close examination for necrosis and a careful Ki67 PR count.

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COVID-19 is heterogeneous; therefore, it is crucial to identify early biomarkers for adverse outcomes. Extracellular vesicles (EV) are involved in the pathophysiology of COVID-19 and have both negative and positive effects. The objective of this study was to identify the potential role of EV in the prognostic stratification of COVID-19 patients. A total of 146 patients with severe or critical COVID-19 were enrolled. Demographic and comorbidity characteristics were collected, together with routine haematology, blood chemistry and lymphocyte subpopulation data. Flow cytometric characterization of the dimensional and antigenic properties of COVID-19 patients' plasma EVs was conducted. Elastic net logistic regression with cross-validation was employed to identify the best model for classifying critically ill patients. Features of smaller EVs (i.e. the fraction of EVs smaller than 200 nm expressing either cluster of differentiation [CD] 31, CD 140b or CD 42b), albuminemia and the percentage of monocytes expressing human leukocyte antigen DR (HLA-DR) were associated with a better outcome. Conversely, the proportion of larger EVs expressing N-cadherin, CD 34, CD 56, CD31 or CD 45, interleukin 6, red cell width distribution (RDW), N-terminal pro-brain natriuretic peptide (NT-proBNP), age, procalcitonin, Charlson Comorbidity Index and pro-adrenomedullin were associated with disease severity. Therefore, the simultaneous assessment of EV dimensions and their antigenic properties complements laboratory workup and helps in patient stratification.

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OBJECTIVES: Periodontitis is induced by the imbalance between osteoblast and osteoclast activity, which leads to periodontal tissue destruction. Macrophages play a vital role in periodontitis. However, the hypoxic periodontal environment will also induce macrophage apoptosis within a short time. Apoptotic bodies (ABs) are the major products generated from apoptotic cells, but whether macrophage-derived ABs play a regulatory role as their mother cells in periodontitis remains unknown. In the present study, we aimed to investigate the effects of ABs on osteoblasts. METHOD: ABs derived from hypoxia-induced macrophages were co-cultured with osteoblasts and the impact of ABs on osteoblast differentiation in vitro was assessed. In vivo, periodontitis model was established and macrophages-derived ABs were injected into the gingival sulcus. The effects of ABs on periodontal bone resorption were determined. RESULTS: The results showed that ABs significantly inhibit osteoblast differentiation and promoted alveolar bone resorption in periodontitis. MicroRNA (miRNAs) array analysis was performed and revealed that miR-483-5p is the key miRNA in ABs. Dual luciferase reporter assays were performed and confirmed that miR-483-5p targeted Col1A1 mRNA and attenuated its expression. CONCLUSION: Macrophage-derived ABs inhibit osteoblast differentiation via the transfer of miR-483-5p, which downregulates Col1A1 expression and finally suppresses osteogenic activity.

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African swine fever virus (ASFV), a devastating pathogen to the worldwide swine industry, mainly targets macrophage/monocyte lineage, but how the virus enters host cells has remained unclear. Here, we report that ASFV utilizes apoptotic bodies (ApoBDs) for infection and cell-cell transmission. We show that ASFV induces cell apoptosis of primary porcine alveolar macrophages (PAMs) at the late stage of infection to productively shed ApoBDs that are subsequently swallowed by neighboring PAMs to initiate a secondary infection as evidenced by electron microscopy and live-cell imaging. Interestingly, the virions loaded within ApoBDs are exclusively single-enveloped particles that are devoid of the outer layer of membrane and represent a predominant form produced during late infection. The in vitro purified ApoBD vesicles are capable of mediating virus infection of naive PAMs, but the transmission can be significantly inhibited by blocking the "eat-me" signal phosphatidyserine on the surface of ApoBDs via Annexin V or the efferocytosis receptor TIM4 on the recipient PAMs via anti-TIM4 antibody, whereas overexpression of TIM4 enhances virus infection. The same treatment however did not affect the infection by intracellular viruses. Importantly, the swine sera to ASFV exert no effect on the ApoBD-mediated transmission but can partially act on the virions lacking the outer layer of membrane. Thus, ASFV has evolved to hijack a normal cellular pathway for cell-cell spread to evade host responses.

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Background: In recent years, extracellular vesicles have been recognized as important mediators of intercellular communication through the transfer of active biomolecules (proteins, lipids, and nucleic acids) across the plant and animal kingdoms and have considerable roles in several physiological and pathological mechanisms, showing great promise as new therapeutic strategies for a variety of pathologies. Methods: In this study, we carefully reviewed the numerous articles published over the last few decades on the general knowledge of extracellular vesicles, their application in the therapy of various pathologies, and their prospects as an approach for the future. Results: The recent discovery and characterization of extracellular vesicles (EVs) of diverse origins and biogenesis have altered the current paradigm of intercellular communication, opening up new diagnostic and therapeutic perspectives. Research into these EVs released by plant and mammalian cells has revealed their involvement in a number of physiological and pathological mechanisms, such as embryonic development, immune response, tissue regeneration, and cancer. They are also being studied as potential biomarkers for disease diagnosis and vectors for drug delivery. Conclusion: Nanovesicles represent powerful tools for intercellular communication and the transfer of bioactive molecules. Their molecular composition and functions can vary according to their origin (plant and mammalian), so their formation, composition, and biological roles open the way to therapeutic applications in a variety of pathologies, which is arousing growing interest in the scientific community. Clinical Trial Registration: ClinicalTrials.gov identifier: NCT03608631.

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Efferocytosis is the physiological process of phagocytic clearance of apoptotic cells by both professional phagocytic cells, such as macrophages, and non-professional phagocytic cells, such as epithelial cells. This process is crucial for maintaining tissue homeostasis in normal physiology. Any defects in efferocytosis can lead to pathological consequences and result in inflammatory diseases. Extracellular vesicles (EVs), including exosomes, microvesicles (MVs), and apoptotic vesicles (ApoVs), play a crucial role in proper efferocytosis. These EVs can significantly impact efferocytosis by affecting the polarization of macrophages and impacting calreticulin (CRT), TAM receptors, and MFG-E8. With further knowledge of these effects, new treatment strategies can be proposed for many inflammatory diseases caused by efferocytosis disorders. This review article aims to investigate the role of EVs during efferocytosis and its potential clinical applications in inflammatory diseases.

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Systemic lupus erythematosus (SLE) is an autoimmune disease caused by environmental factors and loss of key proteins, including the endonuclease Dnase1L3. Dnase1L3 absence causes pediatric-onset lupus in humans, while reduced activity occurs in adult-onset SLE. The amount of Dnase1L3 that prevents lupus remains unknown. To genetically reduce Dnase1L3 levels, we developed a mouse model lacking Dnase1L3 in macrophages (conditional knockout [cKO]). Serum Dnase1L3 levels were reduced 67%, though Dnase1 activity remained constant. Homogeneous and peripheral antinuclear antibodies were detected in the sera by immunofluorescence, consistent with anti-double-stranded DNA (anti-dsDNA) antibodies. Total immunoglobulin M, total immunoglobulin G, and anti-dsDNA antibody levels increased in cKO mice with age. The cKO mice developed anti-Dnase1L3 antibodies. In contrast to global Dnase1L3-/- mice, anti-dsDNA antibodies were not elevated early in life. The cKO mice had minimal kidney pathology. Therefore, we conclude that an intermediate reduction in serum Dnase1L3 causes mild lupus phenotypes, and macrophage-derived DnaselL3 helps limit lupus.

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Homeostasis disruption is visible at the molecular and cellular levels and may often lead to cell death. This vital process allows us to maintain the more extensive system's integrity by keeping the different features (genetic, metabolic, physiologic, and individual) intact. Interestingly, while cells can die in different manners, dying cells still communicate with their environment. This communication was, for a long time, perceived as only driven by the release of soluble factors. However, it has now been reconsidered with the increasing interest in extracellular vesicles (EVs), which are discovered to be released during different regulated cell death programs, with the observation of specific effects. EVs are game changers in the paradigm of cell-cell communication with tremendous implications in fundamental research with regard to noncell autonomous functions, as well as in biomarkers research, all of which are geared toward diagnostic and therapeutic purposes. This review is composed of two main parts. The first is a comprehensive presentation of the state of the art of the EV field at large. In the second part, we focus on EVs discovered to be released during different regulated cell death programs, also known as cell death EVs (cdEVs), and EV-associated specific effects on recipient cells in the context of cell death and inflammation/inflammatory responses.

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Immunotherapy, including immunostimulation and immunosuppression, has seen significant development in the last 10 years. Immunostimulation has been verified as effective in anti-cancer treatment, while immunosuppression is used in the treatment of autoimmune disease and inflammation. Currently, with the update of newly-invented simplified isolation methods and the findings of potent triggered immune responses, extracellular vesicle-based immunotherapy is very eye-catching. However, the research on three main types of extracellular vesicles, exosomes, microvesicles and apoptotic bodies, needs to be more balanced. These three subtypes share a certain level of similarity, and at the same time, they have their own properties caused by the different methods of biogensis. Herein, we summarized respectively the status of immunotherapy based on each kind of vesicle and discuss the possible involved mechanisms. In conclusion, we highlighted that the effect of the apoptotic body is clear and strong. Apoptotic bodies have an excellent potential in immunosuppressive and anti-inflammatory therapies .

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