Osteoarthritis (OA) is a common degenerative disease. Previous studies have shown that the small molecule drug 5Z-7-Oxozeaenol (5Z-7), an inhibitor of TAK1, has a good effect on OA. 5Z-7 has been directly injected into the joint cavity frequently, and the therapeutic effect of the drug is impaired. In this study, exosomes were isolated and purified from the cell supernatant of the ATDC5 cell line, which is an ideal chondrogenic model in vitro. In rat chondrocytes induced by proinflammatory cytokines, 5Z-7-loaded exosomes can promote the expression of anabolism-related genes Col2a1 and Sox9, and inhibit the expression of catabolism-related genes Adamts5 and Mmp13. In this study, the OA model was established in 8-week-old male mice by anterior cruciate ligament transection (ACLT). In order to explore the therapeutic effect of exosomes, paraffin sections of knee joints were taken for histological evaluation after being treated with intra-articular injection of exosomes or 5Z-7-loaded exosomes. The results showed that drug-loaded exosomes could alleviate the pathological phenotypes of post-traumatic OA models. Combined with the results of Micro-CT, the treatment improved the loss of subchondral bone trabeculae, reduced osteophytes, and rendered the articular surface smoother after ACLT. This study confirmed that the ATDC5-derived and 5Z-7-loaded exosomes could alleviate the osteoarthritis phenotype in vivo and in vitro. The delivery of 5Z-7 in exosomes reduces the dosage and frequency of administration, and the loading of 5Z-7 can be superimposed onto the therapeutic effect of exosomes.

As a natural drug delivery vehicle, exosome (EXO) has numerous advantages, such as blood-brain barrier penetration and cell targeting. Paeonol (PAE) has a potential in mitochondrial protection. However, whether exosome-paeonol (EXO-PAE) can ameliorate neuronal mitochondrial dysfunction in cerebral ischemia-reperfusion injury remains unclear. The aim of this study was to investigate the effect of EXO-PAE on mitochondrial function in injured neurons. In this study, a transient middle cerebral artery occlusion model (tMCAO) in SD rats was established by using the thread-tying method. The rats showed ipsilateral Horner’s syndrome and crawling to the opposite side after wakefulness, which indicated that the model was successfully constructed. Microglia-derived exosomes were isolated using ultracentrifugation, and the transmission electron microscope and Western blotting results showed that the specific markers Alix and CD63 were expressed in EXO-PAE, but not in EXO-PAE. In vitro cell experiments showed that EXO-PAE could effectively reduce intracellular ROS levels (P<0.01), increase ATP expression (P<0.05) and mitochondrial membrane potential (P<0.01), suggesting that EXO-PAE ameliorate neuronal mitochondrial dysfunction. The results of Tunnel and MTT tests showed that EXO-PAE reduced the apoptosis of neurons (P<0.01) and Western blotting results showed that EXO-PAE restored the expression of mitochondrial fusion protein Mfn2 (P<0.01), Opa1 (P<0.05) and the mitotic protein Drp1 (P<0.01), thus it might be an important mechanism to improve mitochondrial dysfunction and decrease neuronal apoptosis. At the animal level, the results of pathological examination by TTC staining showed that EXO-PAE could effectively reduce the area of cerebral infarction (P<0.001). In conclusion, the EXO-PAE was successfully constructed and could improve mitochondrial dysfunction and play a role in the treatment of ischemic stroke.

This study aimed to investigate the effect of human papillomavirus 16 (HPV16) on the β-catenin and claudin-1 protein expression and proliferation, migration and invasion of cervical cancer cells. RNA interfere technology was utilized to establish the cell model of HPV 16 E6 knockdown (shE6 group), followed by the assays of CCK8 kit, flow cytometry, scratch and transwell tests to detect the cell proliferation, cell cycle, apoptosis, and migration. We found that in contrast with NC group, the cell cycle was arrested in the transition phase from G₀ / G₁ phase to the S phase, and the cellular ability of proliferation, migration and invasion was decreased significantly in shE6 group. The exosomes were extracted from cell lysate supernatant and the expression of β-catenin and claudin-1 was measured by Western blotting. We found that in shE6 group, the expression of cellular β-catenin was lower, but the corresponding expression in exosomes was increased when compared to NC group. And the expression of claudin-1 protein was increased both in the cells and exosomes. Our results suggested that HPV16 E6 can promote the malignant phenotype of cells, which may be related to the fact that E6 protein inhibited the release of β-catenin via exosomes, thereby increased its cellular accumulation and inhibited the accumulation of claudin-1 in cells and exosomes.

Along with the social structure of population aging, aging and age-related diseases have become an urgent issue In recent years, researchers have found that stem cells can inhibit inflammation, regulate immune responses, prevent apoptosis of target cells, replace and promote restoration of the damaged parts. Accumulating evidence suggests that the beneficial effects of stem cell therapy are predominantly stem cell-derived exosomes. Exosomes are nano-sized vesicles derived from endocytic membranes and contain biomolecules such as proteins, lipids, RNAs, and DNAs regulating cell-to-cell communications. Exosomes may transfer bioactive molecules to target cells, which in turn cause phenotypic changes and then modulate repair and regenerative programs of various organs. These phenotypic changes arise from several mechanisms, ranging from prevention of apoptosis in recipient cells, induction of target cell proliferation, stimulation of immunomodulatory responses, and reduction of oxidative stresses in target cells to the enhancement of oxygen supply. This review gives a brief introduction to biogenesis, release and signaling of exosomes, and focuses on analysis of the effects of exosomes from different stem cells on skin aging and aging-related diseases (such as cardiovascular diseases, osteoarthritis, osteoporosis, and Alzheimer's disease) in basic research and clinical applications. Although many problems with exosomes in the clinical setting exist, the prospects of basic research to clinical applications are worthy of exploration, which is of great significance for anti-ageing treatment.

In recent years, it has been found that microRNA (miRNA) carried by exosomes play an important role in the development of tissue and organ fibrosis. However, studies on the relationship between radiation-induced cardiac fibrosis (RICF) and exosomes and their miRNA are very limited. The purpose of this study is to analyze the possible role of radiation-induced exosome miRNA in the development of RICF by bioinformatics. Myocardial fibroblasts (CFs) are the main effector cells of RICF. The CFs were irradiated with 2 Gy X-rays, and radiation-induced exosomes (X-exo) and unirradiated exosomes of CFs (Exo) were extracted by overspeed centrifugation. Then, exosomes were observed and identified with its morphology, NTA concentration, and exosome surface marker proteins such as CD9, CD63, and CD81. Subsequently, RNA-seq technology was used to detect the miRNA expression profiles of Exo and X-exo, then the differentially expressed miRNAs were screened and their potential target genes and enrichment analysis were analyzed. The results showed that, compared with the control group (Exo), 9 miRNAs were up-regulated in the X-exo (|log₂ Fold Change|>1, P<0.05), of these, 8 are the ones with |log₂ Fold Change|>2; 19 miRNAs were down-regulated in the X-exo (|log₂ Fold Change|>1, P<0.05), of these, 12 are the ones with |log₂ Fold Change|>2. TargetScan, miRWalk and miRNADB were used to predict the target genes of differentially expressed miRNAs, and GO and KEGG enrichment analysis were performed. GO enrichment results showed that target genes regulated by differentially expressed miRNAs were mainly involved in biological processes such as protein phosphorylation and cell signal transduction. Those differentially expressed miRNAs were enriched in cytoplasm, cell membrane and other cellular components, playing molecular functions such as protein kinase binding and protein binding. KEGG enrichment results showed that target genes with differentially expressed miRNAs were mainly enriched in PI3K-Akt, MAPK, mTOR, ECM-receptor interaction, cAMP, Wnt, TGF-β and Notch related signaling pathways. Combing with literatures, it is suggested that differentially expressed exosomal miRNA may promote the development of RICF by regulating the target genes and related signaling pathways. Because the activation regulation of signaling pathways such as MAPK, PI3K/AKT, and mTOR are mainly dependent on the phosphorylation of key molecules, but not on the regulation of the transcriptional level. The detailed mechanism of "Exosomes miRNA mediate radiation bystander effects to promote RICF" will be investigated in future studies, and the signaling pathways such as ECM, cAMP, TGF-β, Wnt, Notch, Ras, and Rap1 should be the focus.

In this study, high-throughput sequencing technology was used to analyze the differentially expressed microRNA (miRNA) of astrocyte-derived exosomes in control group and oxygen and glucose deprivation/reoxygenation (OGD/R) group. Ultracentrifugation was used to extract exosomes from the supernatant of astrocyte medium in the control group and OGD/R group. Transmission electron microscopy showed that exosomes had a typicalvesicle shape with intact membrane and low electron content density. Nanoparticle tracking technology (NTA) detected astrocyte exosomes with a size of 100.5±31.1 nm, accounting for 96.8%. Western blot detection showed that the exosome contained exosome-specific proteins tumor-susceptibility protein (TSG101), heat shock protein 60 (Hsp60), ALG-2-interacting protein X (ALIX). Compared with the control group, 41 miRNAs in the exosomes of the OGD/R group were significantly changed, of which 20 miRNAs were increased and 21 miRNAs were decreased significantly (P<0.05). Gene ontology function (GO) analysis showed that significantly differentially target genes were mainly involved in protein glycosylation, lipid metabolism, phosphorylation, Golgi apparatus, endoplasmic reticulum, endosome, cytoplasmic vesicles and cell protrusions , etc. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis found that differential miRNAs were mainly related to metabolic pathways and signaling pathways such as butyrate metabolism, β-alanine metabolism, fatty acid degradation, mitophagy and P53 signaling pathway. Sequencing analysis of the exosomal miRNAs derived from control and OGD/R astrocytes and target gene function enrichment analysis can be useful for the mechanism study of astrocyte exosomes in response to oxygen and glucose deprivation reperfusion.

The study demonstrates a quick approach for isolating exosomes with substantial concentration. To achieve quality and substantial concentration and purification of exosomes, the target tissue was mechanically chopped and introduced to tissue digesting enzyme for tissue digestion and filtration. The tissue cell solution was then subjected to differential centrifugation, ultra-separation, SEC exclusion, and ultrafiltration. The protein contents of enzymatic treatment of dissociated tissue were higher as compared to protein elution of exosome tissue dissociation method. The nanoparticles were traced and seen using a transmission electron microscope after enrichment of exudate to mouse heart, liver, kidney, human colon cancer, human breast cancer, and atherosclerotic tissues. The finding of the study demonstrated that the diameter of the exocrine body was within 30-150 nm, and the structure was obvious and distinct. Western blots analysis showed that CD9, Alix, and CD63 expression were high, whereas, the TSG101 expression was low but calnexin was negative. However, in comparison to other ways, whole process takes only 4-5 hours and saves time for quantification and functional analysis of exosomes. The isolated exosomes has higher purification with less soluble heteroprotein contamination. Advances in isolation exosomes from micro tissue samples can be employed for nanoparticle size tracking, Western blotting, transmission electron microscopy, and transcriptomic analysis for future studies.

As an important member of the non-coding RNA family, circRNA is a kind of single-stranded RNA with a covalently closed loop structure without a polyadenylated acid tail and 5′-3′ end, showing high stability, abundance and conservation across species characteristics. Recent studies have shown that circRNA plays an important role in many biological processes, including chemotherapy resistance and malignant progression. Exosomes are small extracellular phospholipid bilayer vesicles with a diameter of 30-150 nm that are secreted by living cells. They can be used as carriers to encapsulate and transfer functional molecules. Exosomes are important mediators of communication between tumor cells and stromal cells. They can play a role in the transmission of chemoresistance by transferring circRNA. As chemotherapy resistance is still a huge obstacle to the prognosis of cancer, the research of exosomal circRNA-mediated tumor chemotherapy resistance is at the forefront of academic research, which is a blue ocean with important significance. In this paper, we summarized the latest research progress in the aspects of exosome delivery of circRNA, the mechanism of exosome sorting non-coding RNA cargo, circRNA-mediated tumor chemotherapy resistance, exosome delivery of circRNA-mediated tumor chemotherapy resistance and its potential clinical application, which may provide a reference for the research of tumor chemotherapy resistance.

Exosomes are small vesicles with double lipid membranes secreted by cells into the extracellular space and body fluids. Exosomes contain genetic materials, proteins, lipids and so on, and thus unveil a novel mechanism of intercellular communication. CircRNAs, which function as “microRNA sponges”, are emerging regulator for gene expression. CircRNAs exist in a variety of cells, thereby widely affect the life process of cells through different regulatory networks at the pre- and post-transcriptional levels. The critical role of exosomal circRNAs in the physiological and pathological processes has also attracted increasing attention. Aging process is driven by various contributing factors, which inevitably impairs tissue function and increases susceptibility to disease and death. It is demonstrated that the presence and progressive accumulation of senescent cells contributes to overall organism aging. Senescent cells aggregated in aging tissues have been considered as a causal factor for aging-related disorders. Recent studies have suggested that exosomal circRNAs might act as the senescence-associated secretory phenotype (SASP)of senescent cells and play an important role in the aging process and the occurrence of aging-related diseases. This review introduced the occurrence of exosomal circRNAs in detail, and summarized the research advances of exosomal circRNAs in aging and aging-related diseases. This study may provide references for the further research and clinical application of exosomal circRNAs.

Exosomes are cell-secreted nano-scale vesicles which are rich in a variety of biologically active substances and function as the important mediators in cell-to-cell communication. Long non-coding RNAs (lncRNAs) have impacts on the tumorigenesis and tumor progression from various aspects, and are specifically sorted into exosomes. Tumor microenvironment (TME), an internal environment composed of tumor cells, non-tumor cells (such as endothelial cells, immune cells, fibroblasts, etc.) and extracellular matrix, plays a vital role in the tumorigenesis and tumor progression. Tumor cells secrete large amounts of exosomes into TME. This article summarizes the roles of tumor-derived exosomal lncRNAs in TME from the perspective of how they regulate receptor cells, such as promoting tumor metastasis, drug resistance, cell metabolic reprogramming, tumor stemness, epithelial to mesenchymal transition (EMT), tumor angiogenesis and lymphangiogenesis, and immunosuppression. An in-depth understanding of the role of tumor-derived exosomal lncRNA in TME will help to provide potential new clinical biomarkers and therapeutic targets for cancer.

Abstract: Wound healing is an important physiological process, and it is also one of the problems to be solved in clinical medicine. The exocrine from plants is green, safe and efficient, which is expected to provide a new solution for wound healing. In this paper, the exosome of Panax ginseng was extracted and purified for the first time, and labeled with fluorescent dye, the uptake of exosome of skin cells was observed, the effect of exosome on cell proliferation was detected by CCK-8 method, the gene expression level in cells was detected by real-time quantitative PCR, and the protein expression level in cells was detected by Western blotting analysis. The repair effect of ginseng exosome on skin injury in mice was verified by skin injury experiment. The results showed that the exocrine body of Panax ginseng could be separated by differential centrifugation, and the identification was in accordance with the microscopic identification characteristics of the exocrine body. CCK-8 assay showed that the exocrine body of Panax ginseng could promote the proliferation of skin cells at 24 h and 48 h (P < 0.05). The results of real-time quantitative PCR showed that ginseng exocrine could increase the gene expression of COL1A1 (human type 1 collagen α 1 chain) and fibronectin-1 (human fibronectin-1) in skin cells. Western blotting analysis showed that ginseng exocrine could regulate the increase of TGF- β 1 (transforming growth factor-β Ⅰ) and Vimentin in skin cells, and ki67 increased within 24 hours. The results of skin injury test in mice showed that ginseng exocrine body could accelerate the recovery of skin injury and reduce the expression of inflammatory factors NF- κ B (nuclear factor κ B), INOS (nitric oxide synthase) and COX-2 (cyclooxygenase). This study shows that ginseng exocrine can be extracted and purified by ultracentrifugation combined with density gradient centrifugation. It is proved that ginseng exocrine can regulate skin cell cycle, activate TGF- β pathway, enhance cytoskeleton structure, promote skin cell proliferation, accelerate the recovery of injured skin, and reduce the expression of inflammatory factors.

Abstract Exosomes are involved in invasion, migration and angiogenesis of tumor cells, and invasion is the main cause of death in glioma patients. Studies have shown that the exosomes secreted by tumor cells can carry miRNA into the receptor cells and regulate the biological functions of the receptor cells, such as proliferation, migration and invasion. However, whether the exosomes derived from glioma cells express miR-574-5p and its role in the growth, invasion and migration of glioma cells have not been reported. Therefore, this study will explore the mechanism of miR-574-5p in the process of cell proliferation, migration and invasion. The exosomes were characterized by electron microscopy and nanoparticle size tracking. the internalization of exosomes was detected by immunofluorescence assay. the differential exosomes miRNA miR-574-5p and its target LATS2 were identified by bioinformatics and online data. qRT-PCR was used to detect the expression of miR-574-5P and LATS2 in glioma cell lines LN229. NC mimics (miR-NC group), miR-574-5P mimics (miR-574-5P group) were transfected respectively into LN229 cells by liposome method. The proliferation of the cells was detected by CCK-8 method. The migration and invasion of the cells were detected by Transwell chamber assay. The protein expression of LATS2 in the cells was detected by Western blot. The fluorescence activity of the cells was detected by the dual luciferase reporter assay. The results showed that, compared with the control group, the expression of mir-574-5p was significantly up-regulated and the expression of LATS2 was significantly downregulated in the glioma cells incubated with exosomes (P < 0.05). Overexpression of miR-574-5p can promote the proliferation, migration and invasion of LN229 cells. miR-574-5p can inhibit the fluorescence activity of wild type LATS2, negatively regulate the expression of LATS2, and activate LATS2 / YAP signaling pathway. In conclusion, the exosomal miR-574-5p can promote the proliferation, migration and invasion of glioma cells by downregulating LATS2 and activating LATS2 / YAP signaling pathway, which can provide a scientific and reliable target for the diagnosis and treatment of glioma.

Wound healing is an important physiological process as well as one of the problems to be solved in clinical medicine. Plant-derived exosomes are green, safe and efficient, and are expected to provide a new solution for wound healing. In this paper, ginseng exosomes were extracted and purified by hypervelocity centrifugation, and were identified to be consistent with the microscopic identification characteristics of exosomes. Fluorescence dye was used to label exosomes, which showed that exosomes could be ingested by skin cells. CCK-8 methods indicated that ginseng exosomes could promote the proliferation of skin cells at 24 h and 48 h (P<0.05). Panax ginseng exosomes detected by real-time quantitative PCR increased the transcription levels of human collagen type 1 α1 chain (COL1A1) and fibronectin-1 (human fibronecin-1) genes in skin cells (P<0.05).Western blotting analysis showed that ginseng exosomes could increase the protein levels of TGF-β1 (transforming growth factor-β Ⅰ), Vimentin and Ki67 in skin cells (P<0.05). The skin injury experiment in mice confirmed that ginseng exosomes could accelerate recovery after skin injury. Moreover, the inflammatory cytokines such as nuclear factor kappa B (NF-κB ), INOS (nitric oxide synthase) and cycoxidase ( COX-2 ) could be decreased (P<0.05). In this study, ginseng exosomes were successfully extracted and purified by hypervelocity centrifugation combined with density gradient centrifugation, which suggested that ginseng exosomes can regulate the skin cell cycle, activate the TGF-β pathway, promote the proliferation of skin cells, accelerate the recovery of damaged skin, and reduce the level of inflammatory factors.

Exosomes are involved in invasion, migration and angiogenesis of tumor cells, and invasion is the main cause of death in glioma patients. Studies have shown that the exosomes secreted by tumor cells can carry miRNA into the receptor cells and regulate the biological functions of the receptor cells, such as proliferation, migration and invasion. miR-574-5p plays a key role in the occurrence and development of a variety of tumors. However, whether the exosomes derived from glioma cells express miR-574-5p and its role in the growth, invasion and migration of glioma cells have not been reported. This study investigated the mechanism of the exosomal miR-574-5p secreted from glioma cells in the process of cell proliferation, migration and invasion. The exosomes were characterized by electron microscopy, nanoparticle size tracking and Western blot. The results displayed that the extracted exosomes were round particles with a diameter of 30 ~ 100 nm. The internalization of exosomes was detected by immunofluorescence assay. The results showed that exosomes were internalized into LN229 cells; Bioinformatics and online data were used to screen the differentially secreted miRNA between LN229 and H4 glioma cells. The results showed that the differentially secreted miRNA was miR-574-5p, and large tumor suppressor 2 (LATS2) was predicted to be the target gene of miR-574-5p; Duel luciferase reporter assay confirmed that miR-574-5p was complementary to the 3'UTR region of LATS2; The transfection assay, qRT-PCR and Western blot was conducted to measure the relationship between miR-574-5p and LATS2. The results demonstrated that there was no significant difference in LATS2 mRNA levels between the control group and the group with miR-574-5P overexpression (P > 0.05), suggesting the regulatory effect of miR-574-5P on LATS2 was achieved by inhibiting its translation (P < 0.05). CCK-8, Transwell migration and invasion assays were conducted to explore the effect of miR-574-5p on proliferation, migration and invasion of LN229 cells. The results showed that overexpression of miR-574-5p could significantly promote the ability of proliferation, migration and invasion of LN229 cells (P < 0.05). In addition, Western blot was performed to measure the expression of kinase proteins involved in the LATS2/YAP signaling pathway,and the influence of the exosomes on this signaling pathway. The results revealed that the exosomes down-regulated the protein expression level of LATS2 and reduced p-YAP phosphorylation. In conclusion, the exosomal miR-574-5p can promote the proliferation, migration and invasion of glioma cells by down-regulating LATS2 and activating LATS2/YAP signaling pathway, which may provide a potential biomarker for the diagnosis and target for the treatment of glioma.

Exosome is a kind of vesicular body which can be secreted by most cells and can communicate information between cells through the transfer of specific signal molecules. More and more studies have shown that exosomes are widely involved in the occurrence and development of cardiovascular diseases, such as hypertension, heart failure and myocardial infarction. In recent years, studies have shown that exercise has a great impact on the biological function of body fluids and blood circulation exosomes. Different exercise modes can promote the release of exosomes and affect the expression of miRNA and proteins. At present, studies have found that exercise promotion of exosome release may be related to the laminar shear force induced by blood flow, the increase of Ca⁺ levels, the signal transduction between various cells during exercise and the activation of body physiological states, but the mechanism of exercise promotion of exosome release still needs to be improved. In addition, the release of exosomes and changes in their contents are also considered to be markers of adaptive changes in the body. Recent studies have shown that exercise promotes the release of exosomes and regulates the expression of miRNAs and proteins, thus playing key roles in promoting angiogenesis, inhibiting myocardial apoptosis, inhibiting myocardial fibrosis and protecting endothelial cells, providing a theoretical basis for the prevention and treatment of cardiovascular diseases. Herein the role of exosomes from different cell sources in cardiovascular diseases is reviewed in this paper. We analyze the regulation of exercise exosomes and the molecular mechanism of exercise regulation exosomes in cardiovascular protection, aiming to provide new ideas for the prevention and treatment of cardiovascular diseases.

Skin wound healing is one of the difficult problems in clinical medicine. The long-term wound healing has a great impact on patients and causes huge social and economic burden. Hydrogels have good biocompatibility, biodegradability and plasticity, and have excellent properties of hemostasis, antibacterial and water retention. They are widely used as skin wound dressing. The exosomes derived from stem cells contain the unique contents and membrane components of stem cells. These exosomes have the similar repair function as stem cells, and avoid the safety problems of teratoma and graft immune rejection caused by stem cell therapy, and therefore has become the research hotspot of cell-free repair. However, exosomes in the treatment process will face high clearance rate, short half-life and difficulty in large-scale preparation, which limits the therapeutic effect of exosomes. Hydrogel loaded stem cell exosomes can not only prevent exosome from being rapidly removed, but also play the role of exosome and hydrogel in promoting wound healing, skin repair synergy, and thereafter has potential application value in wound repair research. In this paper, production and characteristics of exosomes, commonly used hydrogel materials, stem cell exosomes from different sources, exosomes extraction and identification methods have been summarized. Furthermore, stem cell exosomes related mechanisms of wound repair, protective and sustained release effects of hydrogel on exosomes, and synergistic effects of hydrogel and exosome are also reviewed. This paper provides a reference for the future research in the field of skin repair.

Exosomes are phospholipid bilayer extracellular vesicles secreted by cells, and they act as carriers for material transfer and information exchange between cells. Exosomes exist in many different biofluids and have good application prospects in disease diagnosis and drug delivery. Due to the nano-scale size and heterogeneity of exosomes, as well as the complex composition of biofluids, the separation of exosomes from biofluids is particularly difficult. At present, there are six commonly used methods forisolating exosomes from biofluids: ultracentrifugation, precipitation, size exclusion chromatography, density gradient centrifugation, ultrafiltration and immuno-affinity membrane. Each method has its own advantages and disadvantages. It is difficult to separate and obtain high-quality exosomes when used alone. There are also differences in the selection of methods for separating exosomes from different biofluids. With the continuous emergence of some new technologies such as microfluidic technology and the continuous improvement of classical methods, the exosome separation technology is improved, which is beneficial to the application of exosomes in clinical and scientific research. This review compares and evaluates the isolation and identification methods of exosomes from different biofluid sources, summarizes the selection and differences of exosome isolation methods from different biofluid sources, and provides a reference for subsequent research on exosomes from biofluids.

Cervical cancer, as the second malignant female cancer, remains a major public health problem worldwide. Exosomes, nanometer-sized extracellular vesicles enveloped in a lipid bilayer membrane, are released by living cells and carry a variety of proteins, lipids, DNA, RNA(including mRNA, miRNA, lncRNA and circRNA) and other biologically active substances. As a novel intercellular communication molecule, exosomes are not only involved in biological processes such as signaling and substance exchange, but also play an important role in the process of the occurrence and development of cervical cancer such as regulating the tumor microenvironment to participate in HPV infection and the immune escape, promoting tumor cell proliferation and blood vessel formation, and regulating the invasion and metastasis of tumor. Exosomes with stability structure, are secreted by cervical cells of different lesion degrees, are ubiquitous in various biological fluids and can be enriched in cervicovaginal lavage sample fluid and plasma, thus it is expected to be a new type of liquid biopsy marker for the early diagnosis of cervical cancer. In addition, exosomes have the characteristics of low immunogenicity, good stability and strong penetration, which can overcome many shortcomings such as low bioavailability and enhance the targeting and drug effect of drugs while reducing nontargeted cytotoxicity and immunogenicity, thus it has the potential to be a new generation of drugs or drug carriers for tumor targeted therapy. This article reviews the latest research on the role of exosomes in the development of cervical cancer and the relevant clinical applications, aiming to provide the basis for novel biomarker for the clinical diagnosis and treatment of cervical cancer.

Exosomes are a type of nano-sized extracellular vesicles, which play essential roles in many cellular processes, such as signal transduction, immune response and antigen presentation, and exist in diverse body fluids, including serum, saliva, urine, cerebrospinal fluid and milk. As the main source of nutrition for mammalian offsprings after birth, breast milk contains various nutrients and bioactive ingredients, which are crucial for animals’ immune system and intestinal development. As active molecules of milk, milk-derived exosomes are involved in complex secretory mechanisms and contain multiple nucleic acids, such as mRNAs and non-coding RNAs. Bioinformatics predicted that these exosomal nucleic acids may participate in immunoregulatory pathways. Moreover, milk-derived exosomal proteins are abundant and may enter target cells to exert regulatory functions, while the lipid components contribute significantly to maintaining the stability and uptake of exosomes. It has been published that milk-derived exosomes could resist animal gastrointestinal digestion in the physiological state, and could be further absorbed by the intestinal tract and participate in regulating the processes of intestinal cell proliferation, intestinal inflammation and diversity of gut microbiota, which are beneficial to animal intestines and have become novel regulatory factor for intestinal health in recent years. This article summarizes the formation, composition, and intestinal fate of milk-derived exosomes, and emphasizes their special functions on intestinal health in the animal field, which may provide a theoretical basis for the study of milk-derived exosomes.

Adipose tissue is an important energy metabolism organ in human body, which maintains heat production and metabolic homeostasis by storing and consuming energy. Meanwhile, it can also act as a secretion organ, secreting a series of adipokines, such as leptin and adiponectin, which plays a role in its own tissues and other metabolic organs. In addition to the secretion of adipokines, more and more studies have focused on the vesicles secreted by adipose tissue, that is, exosomes, which act on tissues and organs throughout the body through autocrine, paracrine and endocrine ways to maintain the crosstalk between organs. In this process, non-coding RNA—microRNA (miRNA), circular RNA (circRNA), long non-coding RNA (lncRNA), proteins, and lipids, etc, can be wrapped by exosomes and transported to target cells, regulating the transcription of target genes and protein modification, and affecting the physiological and pathological processes of the human body. Recent studies have shown that brown adipose tissue can secrete a large number of exosomal miRNA, which can be transported to the liver by endocrine and regulate lipid metabolism. White adipose tissue can induce insulin resistance and tumor development through exosomal miRNA. Perivascular adipose tissue can regulate vascular remodeling. In addition, adipose tissue can also regulate the functions of liver, muscle, blood vessels and other organs through circRNA, lncRNA and proteins. This review will summarize the role of different cargoes in adipose tissue exosomes in metabolic diseases such as diabetes, cardiovascular disease, and cancer, and further explore the mechanism of non-coding RNA in exosomes, in order to understand the indispensable role of adipose tissue exosomes in our life activities.

Exosome, as an important mediator of cell paracrine, plays an important role in promoting angiogenesis. In our preliminary studies, we have successfully isolated and identified exosomes from olfactory mucosa mesenchymal stem cells (OM-MSCs). However, the effect of OM-MSCs exosomes (OM-MSC-Exos) on angiogenesis is still unknown. This study was to evaluate OM-MSC-Exos angiogenesis promotion in vitro and in vivo. After co-culture of PKH67 fluorescently labeled OM-MSC-Exos with human brain microvessel endothelial cells (HBMECs), fluorescence microscopy had been used to observe whether OM-MSCs-Exos could absorbed by HBMECs. The proliferation, migration and effect of tubular structure formation of endothelial cells were detected by cell counting kit 8 (CCK-8), transwell and tube formation assays. Effect of OM-MSCs derived exosomes on angiogenesis in vivo was detected by the Matrigel plug assay and CD31 immunofluorescence. The same amount of PBS was used as control in the above experiments. The results showed that OM-MSCsExos could be absorbed by HBMECs. The CCK-8 analysis showed that experimental group could promote HBMECs proliferation in processing 1, 2, 3, 4, 5 d each time point, when compared with the control group (1.32±0.14 vs. 0.98±0.04, 1.36±0.14 vs.1.04±0.06, 1.75±0.18 vs.1.33±0.11, 2.16±0.11 vs.1.50±0.19, 2.71±0.11 vs. 1.81±0.20, P<0.05). The result of Transwell assay showed that the absorbance value of transmembrane migration cells in the experimental group were significantly higher than that in the control group (1.12±0.05 vs.0.02±0.02, P<0.05). In vitro, Matrigel tube-forming experiment showed that the number of the nodes, junctions, meshes, branches and total length in the experimental group was significantly higher than that of the control group (374.33±127.74 vs. 193.33±44.79, 104.56±33.07 vs. 54.33±11.65, 20.11±11.20 vs. 7.56±3.64, 81.67±19.07 vs. 57.00±13.02, 11466.22±2781.03 vs. 8544.00±1848.61, P <0.05). In vivo, the percentage of CD31+ cells in the experimental group was significantly higher than that of the control group (85.00±5.57 vs.8.00±2.08, P <0.05). The above results indicated that the exosomes from OM-MSCs can induce the angiogenesis performance of endothelial cells.

Chronic benzene exposure damages the hematopoietic system and causes aplastic anemia and even leukemia. Exosomes are nanoscale vesicles secreted by cells and play an important role in many physiological and pathological processes. However, the effects of benzene and its metabolites on secretion of exosomes is unclear. The aim of this study was to investigate whether 1,4-benzoquinone (1,4-BQ), an active metabolite of benzene, can cause changes of secretion of exosomes in HL-60 cells and the role of exosome release in 1,4-BQinduced apoptosis. The cells were treated with different concentrations of 1,4-BQ for 24 hours, and the exosomes were extracted from the cell culture medium by low-temperature ultra-high speed centrifugation. The results showed that 1,4-BQ promotes the secretion of exosomes in a dose-response relationship. Furthermore, the exosome inhibitor GW4869 was used to investigate the effect of exosome secretion in 1,4-BQinduced apoptosis. The apoptosis ratios were analyzed by flow cytometry and expression of apoptosis-related proteins were detected by Western blotting. The apoptosis ratio and the expression of cleaved caspase-9, cleaved caspase-3 and Bcl-2 were increased compared with the control group (P<0.05), and during the co-treatment of 1,4-BQ and GW4869, the apoptotic ratio and expression of apoptosis-related proteins were significantly higher than that in the 1,4-BQ group (P<0.05), indicating that inhibiting exosome secretion could increase 1,4-BQ-induced apoptosis. In summary, 1,4-BQ increases exosomes secretion which plays a protective role in 1,4-BQ-induced apoptosis. This study provides new evidence for understanding benzene’s toxic effects and mechanisms.

Exosomes released into the extracellular microenvironment are nanoscale membrane vesicles with a diameter of 30-150 nm．As a kind of newly discovered mesenchymal stem cells, olfactory mucosa mesenchymal stem cells (OM-MSCs) are potential therapeutic targets for various diseases, and the mechanism of action is postulated to be mediated by secretion of paracrine factors-exosomes. However, the studies for isolation, identification and biological characterization of OM-MSC exosomes remain unreported. In this study, ultracentrifugation was used to isolate exosomes from conditioned medium of OM-MSCs. After the OM-MSCs were isolated and cultured, we used flow cytometry and immunofluorescence for identification. Transmission electron microscopy, nanoparticle size analysis and Western blotting were used to analyze and identify the shape, particle size and specific surface molecular markers of exosomes, respectively. CCK8 proliferation tests, Western blotting and scratch tests were used to analyze the effects of exosomes on the proliferation and migration of human brain microvessel endothelial cells (HBMECs). Our results showed that the OM-MSC exosome morphology was mostly round with a diameter of 40-150 nm, and they expressed exosome markers CD63 and CD81. Moreover, OM-MSC exosomes could significantly promote HBMEC proliferation and migration. This study demonstrates that exosomes can be successfully isolated and purified from the supernatant of OM-MSCs by ultracentrifugation and these exosomes can significantly promote the proliferation and migration of HBMECs.

Exosomes are biologically active vesicles secreted by cells with a diameter of 30-150 nm. Some exosomes from cancer cells can polarize macrophages (Mφ) into M2 subtypes. However, the role of prostate cancer-derived exosomes in macrophage polarization remains unknown. In this study, ultrafiltration was used to extract exosomes (PCa-exo) from conditioned medium of prostate cancer cells PC-3M-2B4 and PC-3M-IE8. Transmission electron microscopy, nanoparticle size analysis and Western blotting were used to analyze and identify the shape, particle size and specific surface molecular markers of exosomes respectively. Exosomes were labeled with PKH67 to observe whether PCa-exo could be absorbed by macrophages. The differential expression of CD206 on M2 macrophages was detected by immunofluorescence. The expression of cytokines such as IL-10 and IL-1β in macrophages induced by PCa-exo was observed by q-PCR. Our results showed that the PCa-exo morphology was mostly round and the diameter was about 40-150 nm. PCa-exo could be absorbed by macrophages in large quantities. After PCa-exo induction, the expression of CD206 in macrophages was significantly increased, and the expression levels of inflammatory factors such as IL-10, IL-1β and IL-Exosomes are biologically active vesicles secreted by cells with a diameter of 30-150 nm. Some exosomes from cancer cells can polarize macrophages (Mφ) into M2 subtypes. However, the role of prostate cancer-derived exosomes in macrophage polarization remains unknown. In this study, ultrafiltration was used to extract exosomes (PCa-exo) from conditioned medium of prostate cancer cells PC-3M-2B4 and PC-3M-IE8. Transmission electron microscopy, nanoparticle size analysis and Western blotting were used to analyze and identify the shape, particle size and specific surface molecular markers of exosomes respectively. Exosomes were labeled with PKH67 to observe whether PCa-exo could be absorbed by macrophages. The differential expression of CD206 on M2 macrophages was detected by immunofluorescence. The expression of cytokines such as IL-10 and IL-1β in macrophages induced by PCa-exo was observed by q-PCR. Our results showed that the PCa-exo morphology was mostly round and the diameter was about 40-150 nm. PCa-exo could be absorbed by macrophages in large quantities. After PCa-exo induction, the expression of CD206 in macrophages was significantly increased, and the expression levels of inflammatory factors such as IL-10, IL-1β and IL-12 were consistent with those of M2/TAM subtype macrophages. This study demonstrates that exosome from prostate cancer cells can induce macrophage polarization to the M2 phenotype.
12 were consistent with those of M2/TAM subtype macrophages. This study demonstrates that exosome from prostate cancer cells can induce macrophage polarization to the M2 phenotype.

Exosomes are the cellderived small vesicles with a diameter of 30-150 nm, which contain mRNA, microRNA and long noncoding RNA (lncRNA). It has been known that lncRNA H19 in exosomes plays an important role in regulating cell proliferation, migration and invasion in tumor. However, the detailed biological function of lncRNA is not well understood. In order to evaluate the effects of lncRNA H19 hepatocarcinoma cells on other tumor cells, we screened the cell lines of hepatocellular carcinoma with different lncRNA H19 expression levels. HCCLM3 cells showed high expression of lncRNA H19, while Hep3B and HepG2 cells showed low expression of lncRNA H19. The lncRNAH19 was knocked down by siRNA in HCCLM3 cells. The results showed that the expression of lncRNA H19 was significantly reduced. Both Hep3B and HepG2 cells were transfected with exosomes derived from HCCLM3 cells with high expression of lncRNA H19 or HCCLM3 cells with the lncRNA H19 knockdown. MTs, colony formation, and transwell assays were performed to investigate the behavior of Hep3B and HepG2 cells. The results showed that HCCLM3derived exosomes could be released from cells in culture and endocytosed by the Hep3B and HepG2 cells. Compared with the control cells, the depletion of lncRNA H19 in exosomes inhibited cells proliferation, migration and invasion in Hep3B and HepG2. The data also showed that the PI3K/AKT/mTOR pathway was activated by lncRNA H19 overexpression to promote cell proliferation and migration. The above results indicated that lncRNA H19 in exosomes plays an important role in regulating cell proliferation, migration and invasion in hepatocarcinoma, and promotes the carcinogenesis and development of cancer cells.

As a kind of nanoscale vesicles, related research on exosomes has become a hot topic in recent years. Exosomes are derived from multi-vesicular endosomes (MVBs) and are released into extracellular space after the MVBs fused with the cell membrane. Because exosomes from specific cell types contain a variety of specific proteins and microRNAs, they can be widely used as innovative biomarkers for disease diagnosis and prognosis. Compared with other exogenous drug carriers, exosomes have lower immunogenicity and can target the diseased cells. The application potential of exosomes in clinics has been continuously expanded, such as that exosomes produced naturally or artificially by cells may be potentially developed as a new drug carrier. In addition, rapid progress has been made on exosomes-based diagnostic methods and drug development for major diseases such as cancer, diabetes, cardiovascular, cerebrovascular, and neurodegenerative diseases. This review focuses on the role of exosomes as biomarkers in disease diagnosis and prognosis, as well as the advantages and problems of exosomes as emerging drug delivery vehicles.

Exosomes are nano-vesicular structures that are actively secreted to the external environment by cells. In common, the diameter of the exosomes is below 100 nm. It is a new type of carriers for material exchange and information communication between original cells and target cells. Exosomes can be taken up by the surrounding cells directly or through the blood circulation to reach everywhere of the body. Exosomes play a regulatory role in both normal physiological processes and disease occurrence and progression. As a kind of paracrine mediator, exosomes derived from mesenchymal stem cells (MSC-exosomes) could play physiological roles similar to that of stem cells. Bioactive proteins, lipids and bioactive substances, such as DNA, mRNA and non-coding RNA carried by exosomes may have the same functions as the therapeutic mechanisms of MSCs. In this review, we mainly summarize the origin and release, extraction and identification, biological functions and future prospect of MSC-exosomes. This review provides a comprehensive summary of MSC-exosomes, which would be helpful for further research in this field.

Urinary exosomes are virus-sized vesicles or sacs including the physiological and pathological information originated from the kidney and urogenital tracts. The approach of polyethylene glycol precipitation (PGP) has been successfully and effectively utilized in the isolation and enrichment of exosomes from serum samples. However, there is a shortage of detailed reports focused on the isolation of urinary exosomes by PGP. In this study, we isolated and enriched the urinary exosomes by PGP. Urine samples of ten healthy volunteers were recruited. Results of electron microscope observation displayed the cystoid vesicles wrapped in double layer membranes in diameters from 30 to 100 nm, and with a high electron density area. CD63, CD9, TSG101 and ADAM10 as the biomarkers of urinary exosomes were detected by Western blot analysis. Two peaks, at 25.37 and 95.07 nm respectively, were observed at all isolated urinary exosomes. Representative exosomal RNAs (SnRNA-U6 and β-actin) were detected by qRT-PCR in all samples. Our results suggest that PGP method is an effective, economical and convenient way for the isolation of urinary exosomes, which might accelerate the application of noninvasive liquid biopsy, especially suitable for kidney and urogenital tract diseases.

Exosomes are a type of extracellular vesicles that are released upon fusion of multivesicular bodies with the cellular plasma membrane. Exosomes are shown to deliver functional substances, including proteins, lipids and RNAs to recipient cells, mediating cell-to-cell communications and hence affecting the physiological and pathological functions of cells. In recent years, increased studies have found that exosomes play an important role in the pathogenesis of microbial infections. In chronic infection, pathogens have evolved strategies to spread infectious proteins and viral RNAs via exosomes to change the functions of uninfected cells. At the same time, proteins with high immunogenicity can deliver the pathogenic information to the immune system and hence activate anti-infectious immunity. This article reviews the progress of exosomes in microbial infections. Investigating these mechanisms may inspire future applications for the diagnosis and treatment of chronic infection.