Exosomal Proteins Research Articles

Extracellular vesicles as a new frontier of diagnostic biomarkers in osteosarcoma diseases: a bibliometric and visualized study.

The use of liquid biopsy in cancer research has grown exponentially, offering potential for early detection, treatment stratification, and monitoring residual disease and recurrence. Exosomes, released by cancer cells, contain tumor-derived materials and are stable in biofluids, making them valuable biomarkers for clinical evaluation. Bibliometric research on osteosarcoma (OS) and exosome-derived diagnostic biomarkers is scarce. Therefore, we aimed to conduct a bibliometric evaluation of studies on OS and exosome-derived biomarkers. Using the Web of Science Core Collection database, Microsoft Excel, the R "Bibliometrix" package, CiteSpace, and VOSviewer software, quantitative analyses of the country, author, annual publications, journals, institutions, and keywords of studies on exosome-derived biomarkers for OS from 1995 to 2023 were performed. High-quality records (average citation rate ≥ 10/year) were filtered. The corresponding authors were mainly from China, the USA, Australia, and Canada. The University of Kansas Medical Center, National Cancer Center, Japan, and University of Kansas were major institutions, with limited cooperation reported by the University of Kansas Medical Center. Keyword analysis revealed a shift from cancer progression to mesenchymal stem cells, exosome expression, biogenesis, and prognostic biomarkers. Qualitative analysis highlighted exosome cargo, including miRNAs, circRNAs, lncRNAs, and proteins, as potential diagnostic OS biomarkers. This research emphasizes the rapid enhancement of exosomes as a diagnostic frontier, offering guidance for the clinical application of exosome-based liquid biopsy in OS, contributing to the evolving landscape of cancer diagnosis.

Serum exosomal miR-3662 down-regulates the expression of RBMS3 to promote malignant progression and gemcitabine resistance of breast cancer cells.

Breast cancer (BC) is a prevalent malignancy among women worldwide. As an anticancer drug of pyrimidine nucleoside analogs, gemcitabine can be used to treat BC, but its clinical application is restricted due to drug resistance. This study investigated the effect of serum exosomal microRNA-3662 (miR-3662) on gemcitabine resistance in BC cells bytargeting RNA-Binding Motif Single-Stranded Interacting Protein 3 (RBMS3) and related molecular mechanisms. We performed the bioinformatics analyses on the differential miRNAs in BC and predicted the downstream regulators. Quantitative real-time polymerase chain reaction was conducted to determine miR-3662 and RBMS3 expression, while dual luciferase was conducted to confirme the regulatory relationship between them. Flow cytometry, cell counting kit-8, and transwell assays were applied to assess apoptosis, cell viability, invasion, and migration. The expression of marker proteins (TSG101, CD63, and CD81) in patients' serum exosomes was evaluated through western blot, and exosomes were observed using transmission electron microscopy. miR-3662 expression was significantly upregulated in BC, and miR-3662 knockdown significantly reduced BC cell viability and gemcitabine resistance. As the downstream gene of miR-3662, RBMS3 was significantly downregulated in BC, and dual luciferase assay verified the binding of RBMS3-3'UTR to miR-3662. Rescue experiments revealed that silencing RBMS3 reversed the inhibitory effect of miR-3662 knockdown on BC cells. Besides, we also found that miR-3662 expression was significantly low in serum exosome samples from BC patients and could be transmitted to tumor cells. miR-3662 was upregulated in serum exosomes and promoted BC cell progression and gemcitabine resistance by targeting RBMS3.

Comprehensive analysis of exosome gene LYPD3 and prognosis/immune cell infiltration in lung cancer.

Lung cancer (LC) is a leading cause of cancer-associated mortality worldwide, with high incidence and mortality rates. Ly6/PLAUR domain containing 3 (LYPD3) is a tumorigenic and highly glycosylated cell surface protein that has been rarely reported in LC. This study aimed to explore the prognostic role and immune cell infiltration of LYPD3 in LC. We used ExoCarta, a database of exosomal proteins and RNA, to select exosomes in LC. The Tumor Immune Estimation Resource (TIMER) and Human Protein Atlas (HPA) databases were utilized to compare the expression of LYPD3 in LC. We applied Gene Expression Profiling Interactive Analysis 2 (GEPIA2) and Kaplan-Meier (KM) plotter to evaluate the prognostic prediction performance of LYPD3. Biological processes (BPs), Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses, and gene set enrichment analysis (GSEA) analyses were performed to illustrate the possible role of LYPD3 in LC. The correlations between LYPD3 and immune cell infiltration were explored using Tumor and Immune System Interaction Database (TISIDB), GEPIA2, and TIMER. R software was used for statistical analysis and mapping. A total of 904 exosome molecules were screened in LC. Further analysis showed that the up-regulation of LYPD3 in these 904 exosome molecules was associated with poor prognosis in LC. Pan-cancer analyses revealed that the expression of LYPD3 varied in many cancers, particularly in LC. Clinical correlation analysis indicated that LYPD3 was associated with stage and T classification in LC. We observed that LYPD3 co-expression genes were associated with cell cycle, DNA replication, proteasome, and regulation of the actin cytoskeleton by GSEA. Moreover, LYPD3 was associated with immune modulators. Immunophenoscores (IPS) and IPS-CTLA4 were significantly different between the high LYPD3 group and low LYPD3 group. Additionally, the median half maximal inhibitory concentration (IC50) of bexarotene, cyclopamine, etoposide, and paclitaxel in LYPD3 high group was significantly lower than that in LYPD3 low group. LYPD3 is involved in many BPs of LC, such as regulating immune cell infiltration and affecting prognosis. Therefore, LYPD3 may have potential value as a biomarker for prognosis and immunotherapy in LC.

Exosome proteomes reveal glycolysis-related enzyme enrichment in primary canine mammary gland tumor compared to metastases

ObjectiveNumerous evidence has highlighted the differences between primary tumors and metastases. Nonetheless, the differences in exosomal proteins derived from primary tumor and metastases remain elusive. Here, we aimed to identify differentially expressed exosomal proteins from primary canine mammary gland tumor and metastases to understand how they shape their own tumor microenvironment.MethodsWe clearly distinguished primary canine mammary gland tumors (CHMp) from metastases (CHMm) and profiled the proteins within their secreted exosomes using LC–MS/MS. Moreover, the abundance of glycolysis enzymes (GPI, LDHA) in CHMp exosome was verified with Western blotting, To broaden the scope, we extended to human colorectal cancer-derived exosomes (SW480 vs. SW620) for comparison.ResultsWe identified significant differences in 87 and 65 proteins derived from CHMp and CHMm, respectively. Notably, glycolysis enzymes (GPI, LDHA, LDHB, TPI1, and ALDOA) showed specific enrichment in exosomes from the primary tumor.ConclusionWe observed significant differences in the cellular proteome between primary tumors and metastases, and intriguingly, we identified a parallel heterogeneity the protein composition of exosomes. Specifically, we reported that glycolysis enzymes were significantly enriched in CHMp exosomes compared to CHMm exosomes. We further demonstrated that this quantitative difference in glycolysis enzymes persisted across primary and metastases, extending to human colorectal cancer-derived exosomes (SW480 vs. SW620). Our findings of the specific enrichment of glycolysis enzymes in primary tumor-derived exosomes contribute to a better understanding of tumor microenvironment modulation and heterogeneity between primary tumors and metastases.

Tumor-derived exosomal ADAM17 promotes pre-metastatic niche formation by enhancing vascular permeability in colorectal cancer

BackgroundHematological metastasis has been recognized as a crucial factor contributing to the high rates of metastasis and mortality observed in colorectal cancer (CRC). Notably, exosomes derived from cancer cells participate in the formation of CRC pre-metastatic niches; however, the mechanisms underlying their effects are largely unknown. While our preliminary research revealed the role of exosome-derived disintegrin and metalloproteinase 17 (ADAM17) in the early stages of CRC metastasis, the role of exosomal ADAM17 in CRC hematogenous metastasis remains unclear.MethodsIn the present study, we isolated and purified exosomes using ultracentrifugation and identified exosomal proteins through quantitative mass spectrometry. In vitro, co-culture assays were conducted to evaluate the impact of exosomal ADAM17 on the permeability of the blood vessel endothelium. Vascular endothelial cell resistance, the cell index, membrane protein separation, flow cytometry, and immunofluorescence were employed to investigate the mechanisms underlying exosomal ADAM17-induced vascular permeability. Additionally, a mouse model was established to elucidate the role of exosomal ADAM17 in the modulation of blood vessel permeability and pre-metastatic niche formation in vivo.ResultsOur clinical data indicated that ADAM17 derived from the circulating exosomes of patients with CRC could serve as a blood-based biomarker for predicting metastasis. The CRC-derived exosomal ADAM17 targeted vascular endothelial cells, thus enhancing vascular permeability by influencing vascular endothelial cadherin cell membrane localization. Moreover, exosomal ADAM17 mediated the formation of a pre-metastatic niche in nude mice by inducing vascular leakage, thereby promoting CRC metastasis. Nonetheless, ADAM17 selective inhibitors effectively reduced CRC metastasis in vivo.ConclusionsOur results suggest that exosomal ADAM17 plays a pivotal role in the hematogenous metastasis of CRC. Thus, this protein may serve as a valuable blood-based biomarker and potential drug target for CRC metastasis intervention.

Pulmonary Biodistribution of Platelet-Derived Regenerative Exosomes in a Porcine Model.

The purpose of this study was to evaluate the biodistribution of a platelet-derived exosome product (PEP), previously shown to promote regeneration in the setting of wound healing, in a porcine model delivered through various approaches. Exosomes were labeled with DiR far-red lipophilic dye to track and quantify exosomes in tissue, following delivery via intravenous, pulmonary artery balloon catheter, or nebulization in sus scrofa domestic pigs. Following euthanasia, far-red dye was detected by Xenogen IVUS imaging, while exosomal protein CD63 was detected by Western blot and immunohistochemistry. Nebulization and intravenous delivery both resulted in global uptake of exosomes within the lung parenchyma. However, nebulization resulted in the greatest degree of exosome uptake. Pulmonary artery balloon catheter-guided delivery provided the further ability to localize pulmonary delivery. No off-target absorption was noted in the heart, spleen, or kidney. However, the liver demonstrated uptake primarily in nebulization-treated animals. Nebulization also resulted in uptake in the trachea, without significant absorption in the esophagus. Overall, this study demonstrated the feasibility of pulmonary delivery of exosomes using nebulization or intravenous infusion to accomplish global delivery or pulmonary artery balloon catheter-guided delivery for localized delivery.

Pathogenic Factors and Mechanisms of the Alternaria Leaf Spot Pathogen in Apple

Alternaria leaf spot seriously threatens the sustainable development of the global apple industry, causing significant losses and reducing fruit quality and yield. The causal agent Alternaria alternata f. sp. mali (Alternaria mali, ALT) produces various molecules to modulate infection, such as cell wall-degrading enzymes, toxins, and elicitor-like molecules. ALT produces the host-specific AM-toxin, an important pathogenicity factor. ALT also releases effectors into apple cells that modify host defense, but these proteins have not yet been described. Here, we identified the pathogenic fungal types responsible for early defoliation from diseased leaves of Fuji (Malus domestica cv. ‘Fuji’) apple collected from five districts in Shandong Province, China. The ALT isolates ALT2 to ALT7 were pathogenic to four apple cultivars, with ALT7 being the most aggressive. We extracted mycotoxins (AM-toxin-2 to AM-toxin-7) from each isolate and used them to treat different apple varieties, which led to leaf-spot symptoms and damaged chloroplasts and nuclear membranes, followed by cell death. AM-toxin-7 produced the most severe symptoms, but chloroplasts remained intact when the mycotoxin was inactivated. Mass spectrometry identified 134 secretory proteins in ALT7 exosomes, and three secreted proteins (AltABC, AltAO, and AltPDE) were confirmed to be involved in apple pathogenesis. Therefore, ALT secretes AM-toxin and secretory proteins as an infection strategy to promote fungal invasion and overcome the host defense system.

Human adenovirus type 3 restores pharmacologically inhibited exosomal cargo in lung carcinoma cells.

Introduction: Drug repurposing is fast growing and becoming an attractive approach for identifying novel targets, such as exosomes for cancer and antiviral therapy. Exosomes are a specialized class of extracellular vesicles that serve as functional mediators in intercellular communication and signaling that are important in normal physiological functions. A continuously growing body of evidence has established a correlation between the abnormal release of exosomes with various viral disease pathologies including cancer. Cells that are virus-infected release exosomes known to influence the process via the loading and transfer of viral components, such as miRNA, small (s) RNA, DNA, and proteins. Inhibition of exosome release may abate the spread and severity of viral infection, thus making exosomes an attractive target for antiviral therapies. We previously demonstrated the pharmacological inhibition of exosomes. Methods: Herein, we used a cell-based assay to determine the effect of Human adenovirus type 3 (HAdV3) on the exosome inhibition process by azole and Heparin derivatives. HAdV3-infected cells were treated with two concentrations of each inhibitor at different time points. Results: HAdV3 activities led to increased total sRNA, DNA, and exosome particle concentrations via particle tracking in the presence of Climbazole and Heparin relative to uninfected exosomes. In addition, there was an increased expression of classical markers such as ALG-2 interacting protein X (ALIX), and tetraspanin (CD63), (p < 0.05) and upregulated transcription factor interferon regulatory factor (IRF) 8 in the presence of HAdV3 after 24hours (h) of treatment. Whereas higher concentrations of Climbazole and Heparin sodium salt were found to inhibit total exosome protein (p < 0.001) and exo-RNA (p < 0.01) content even in the presence of HAdV3 relative to infected exosomes only. Activities of HAdV3 in the presence of selected inhibitors resulted in the positive regulation of exosome related DNA damage/repair signaling proteins. Blocking exosome secretion partially obstructed viral entry. Immunological studies revealed that HAdV3 fiber protein levels in A549 cells were reduced at all concentrations of Climbazole and Heparin and both multiplicities of infections (p < 0.001). Discussion: Our findings suggest that while HAdV may bolster inhibited exosome content and release when modulating certain activities of the endosomal pathway mediators, HAdV entry might be constrained by the activities of these pharmacological agents.

Elucidating the EXOSC3-IRE1α interaction: a convergent study incorporating computational, in vitro and in vivo studies

The Unfolded protein response (UPR) is an adaptive signalling pathway which is triggered by accumulation of unfolded/misfolded protein in ER lumen. The UPR consist of three transmembrane proteins-IRE1α, PERK and ATF6 that sense ER stress which leads to activation and downstream signaling from ER lumen to cytosol to restore homeostasis. IRE1α is an evolutionary conserved arm of UPR and acts as an interaction platform for many potential proteins that become activated under ER stress conditions. We investigated potential partners of IRE1 α through MS studies and found EXOSC3 as one of the binding partner of IRE1α. Exosomal complex proteins have 3′5′ exonuclease properties (EXOSC3) that play an important role in mRNA surveillance. This property of exosomal proteins coincides with IRE1α ribonuclease activities and its mechanism of action is similar to that of IRE1α-RIDD pathway which degrades any unstable mRNA that disrupts cellular homeostasis. At the same time, studies have shown that knockdown of EXOSC3 causes ER stress in human cells, so we speculated that there might be a functional crosstalk between IRE1α and EXOSC3 under ER stress conditions. Therefore, we employed computational tools to predict and explore the stability and dynamics of the IRE1α-EXOSC3 complex. The analysis indicates that IRE1α and EXOSC3 exhibit potential interaction with the involvement of ScanNet, predicting binding pockets between the two proteins. Further, the interaction was validated via co-immunoprecipitation and yeast two-hybrid assays, thus suggesting EXOSC3 as a component of the UPRosome complex. Hence, this functional crosstalk might influence the dynamic functional output of IRE1α. Communicated by Ramaswamy H. Sarma

Exosomes Derived from Meningitic Escherichia coli-Infected Brain Microvascular Endothelial Cells Facilitate Astrocyte Activation.

Numerous studies have shown that exosomes play a regulatory role in a variety of biological processes as well as in disease development and progression. However, exosome-mediated intercellular communication between brain microvascular endothelial cells (BMECs) and astrocytes during meningitic Escherichia coli (E. coli)-induced neuroinflammation remains largely unknown. Here, by using in vivo and in vitro models, we demonstrate that exosomes derived from meningitic E. coli-infected BMECs can activate the inflammatory response of astrocytes. A label-free quantitation approach coupled with LC-MS/MS was used to compare the exosome proteomic profiles of human BMECs (hBMECs) in response to meningitic E. coli infection. A total of 57 proteins exhibited significant differences in BMEC-derived exosomes during the infection. Among these proteins, growth differentiation factor 15 (GDF15) was significantly increased in BMEC-derived exosomes during the infection, which triggered the Erk1/2 signaling pathway and promoted the activation of astrocytes. The identification and characterization of exosome protein profiles in BMECs during meningitic E. coli infection will contribute to the understanding of the underlying pathogenic mechanisms from the perspective of intercellular communication between BMECs and astrocytes, and provide new insights for future prevention and treatment of E. coli meningitis.

Dual-modal aptasensor based on multifunctional acridone derivative for rapid detection of exosomal proteins and cancer diagnosis

Molecular profiling of exosomes is a promising non-invasive approach for cancer diagnosis. However, the rapid, robust, and sensitive detection of exosomal proteins remains challenging. In this study, we develop a dual-modal aptamer sensor (aptasensor) using reversible exosomes isolation and 2-sulfo-acridone (2-SA) mediated signal amplification for the colorimetric and fluorescent detection of exosomal proteins. We use aptamers to specifically recognize and capture exosomal transmembrane proteins, and the captured exosomes load a large amount of 2-SA. Then the captured exosomes are released efficiently by disulfide bond-modified complementary DNA strands. The exosomes are lysed, and loaded 2-SA is released. The released 2-SA oxidizes 3,3′,5,5′-tetramethylbenzidine (TMB) under 365 nm LED light irradiation at physiological pH condition, changing the solution from colorless to blue, which enables the rapid screening and identification of samples as negative or positive, especially in point-of-care testing (POCT) settings. Furthermore, 2-SA is able to emit a bright fluorescence signal, which can enable the precise quantitative detection of exosomes with a limit of detection of 1.3×105 particles mL−1. Finally, the disulfide bond was reduced by glutathione, resulting in the dissociation between aptamers and their complementary DNA strands and enabling the next cycle of exosomes capture and release. We evaluate the ability of the aptasensor to accurately classify cancers using machine learning algorithms by analyzing different exosomal transmembrane proteins from blood samples. We believe this aptasensor will be a promising tool in the field of cancer diagnosis because it provides a dual-modal readout signal to achieve rapid qualitative screening and quantitative detection.

Bifunctional immunoaffinity magnetic nanoparticles for high-efficiency separation of exosomes based on host-guest interaction

The investigation of exosomes holds substantial importance in the field of disease diagnosis and prognosis, where in the rapid and low-loss isolation of exosomes emerges as a crucial step owing to their exceedingly low concentration in bodily fluids. Herein, bifunctional immunoaffinity magnetic nanoparticles (BI-NPs) were successfully constructed by binding Ti4+ and antibody to magnetic nanoparticles through host-guest interaction. Exosomes in the cell culture supernatant could be effectively captured by the BI-NPs and then gently eluted with α-CD and 10% ammonia solution. Following the elution process, the content of captured exosome protein was determined to be up to 97 μg/mL. A total of 2822 protein groups were identified in the exosomes isolated by BI-NPs, 1060 protein groups were derived from exosomes. At the same time, the mass of exosome protein obtained by BI-NPs was more than 2 times that of UC isolation, and the above results indicated that BI-NPs had high purity enrichment performance. This is attributed to the fact that BI-NPs combine the dual affinity of Ti4+ and antibody to achieve efficient enrichment of exosomes, in addition to the mild elution property of BI-NPs due to the presence of the host-guest system. BI-NPs offer a novel approach for the isolation of exosomes, in order to further promote the application of exosomes in the required fields.

The core exosome proteome of Trichomonas vaginalis

BackgroundTrichomonas vaginalis is parasitic protozoan that causes human urogenital infections. Accumulated reports indicated that exosomes released by this parasite play a crucial role in transmitting information and substances between cells during host-parasite interactions. Current knowledge on the protein contents in T. vaginalis exosome is mainly generated from three previous studies that used different T. vaginalis isolates as an experimental model. Whether T. vaginalis exosomes comprise a common set of proteins (core exosome proteome) is still unclear. MethodsTo explore the core exosome proteome in T. vaginalis, we used liquid chromatography-tandem mass spectrometry (LC-MS/MS) to identify the contents of sucrose ultracentrifugation-enriched exosome and supernatant fractions isolated from six isolates. ResultsTransmission electron microscopy (TEM) confirmed the presence of exosomes in the enriched fraction. Proteomic analysis identified a total of 1870 proteins from exosomal extracts. There were 1207 exosomal-specific proteins after excluding 436 ‘non-core exosomal proteins’. Among these, 72 common exosomal-specific proteins were expressed in all six isolates. Compared with three published T. vaginalis exosome proteome datasets, we identified 16 core exosomal-specific proteins. These core exosomal-specific proteins included tetraspanin (TvTSP1), the classical exosome marker, and proteins mainly involved in catalytic activity and binding such as ribosomal proteins, ras-associated binding (Rab) proteins, and heterotrimeric G proteins. ConclusionsOur study highlighted the importance of using supernatant fraction from exosomal extract as a control to eliminate ‘non-core exosomal proteins’. We compiled a reference core exosome proteome of T. vaginalis, which is essential for developing a fundamental understanding of exosome-mediated cell communication and host-parasite interaction.

Downregulation of the CD151 protects the cardiac function by the crosstalk between the endothelial cells and cardiomyocytes via exosomes.

Heart failure (HF) is the last stage in the progression of various cardiovascular diseases. Although it is documented that CD151 contributes to regulate the myocardial infarction, the function of CD151 on HF and involved mechanisms are still unclear. In the present study, we found that the recombinant adeno-associated virus (rAAV)-mediated endothelial cell-specific knockdown of CD151-transfected mice improved transverse aortic constriction (TAC)-induced cardiac function, attenuated myocardial hypertrophy and fibrosis, and increased coronary perfusion, whereas overexpression of the CD151 protein aggravated cardiac dysfunction and showed the opposite effects. In vitro, the cardiomyocytes hypertrophy induced by PE were significantly improved, while the proliferation and migration of cardiac fibroblasts (CFs) were significantly reduced, when co-cultured with the CD151-silenced endothelial cells (ECs). To further explore the mechanisms, the exosomes from the CD151-silenced ECs were taken by cardiomyocyte (CMs) and CFs, verified the intercellular communication. And the protective effects of CD151-silenced ECs were inhibited when exosome inhibitor (GW4869) was added. Additionally, a quantitative proteomics method was used to identify potential proteins in CD151-silenced EC exosomes. We found that the suppression of CD151 could regulate the PPAR signaling pathway via exosomes. Our observations suggest that the downregulation of CD151 is an important positive regulator of cardiac function of heart failure, which can regulate exosome-stored proteins to play a role in the cellular interaction on the CMs and CFs. Modulating the exosome levels of ECs by reducing CD151 expression may offer novel therapeutic strategies and targets for HF treatment.

Exosome-derived lncRNA A1BG-AS1 attenuates the progression of prostate cancer depending on ZC3H13-mediated m6A modification

BackgroundExosome-derived long non-coding RNAs (lncRNAs) and N6-methyladenosine (m6A) modifications of lncRNAs have been shown crucial functions in prostate cancer (PCa). Herein, we aim to investigate the detailed mechanism of exosome-derived lncRNA A1BG-AS1 in PCa process.MethodsPCa cell exosomes were extracted, exosomal marker proteins (CD63, CD9) were detected utilizing western blotting, and exosomes with overexpressing A1BG-AS1 were co-cultured with targeted PCa cells. qRT-PCR was used to detect A1BG-AS1 expression and m6A methyltransferase ZC3H13 in PCa. Transwell, colony formation and CCK-8 assays were utilized to assess the invasion, migration, and proliferation ability of PCa cells. Then, we performed actinomycin D and MeRIP assays to analyze the regulatory effect of ZC3H13 on A1BG-AS1 mRNA stability and m6A modification level.ResultsWe observed that A1BG-AS1 and ZC3H13 expression was restricted in PCa tumors. The invasion, proliferation and migratory capacities of PCa cells could be inhibited by up-regulating A1BG-AS1 or by co-culturing with exosomes that up-regulate A1BG-AS1. Additionally, ZC3H13 promoted stable A1BG-AS1 expression by regulating the m6A level of A1BG-AS1.ConclusionExosomal A1BG-AS1 was m6A-modified by the m6A methyltransferase ZC3H13 to stabilize expression and thus prevent PCa cell malignancy. These findings offer a possible target for clinical therapy of PCa.

The Uridylyl Transferase TUT7-Mediated Accumulation of Exosomal miR-1246 Reprograms TAMs to Support CRC Progression.

Tumor-associated macrophages (TAMs) play a crucial role in promoting tumor growth and dissemination, motivating a search for key targets to interfere with the activation of TAMs or reprogram TAMs into the tumor-suppressive type. To gain insight into the mechanisms of macrophage polarization, a designed co-culture system is established, allowing for the education of macrophages in a manner that closely mimics the intricacies of TAMs in the tumor immune microenvironment (TIME). Through database mining, exosomal miR-1246 is identified and is then validated. Exosomal miR-1246-driven polarization of TAMs disrupts the infiltration and function of CD8+ T cells. Mechanically, the amassment of exosomal miR-1246 stems from TUT7-mediated degradation of small noncoding RNA, a process stabilized by SNRPB, but not the precursor of miR-1246. Moreover, an Exo-motif is present in the exosomal miR-1246 sequence, enabling it to bind with the exosomal sorting protein hnRNPA2B1. RNA-seq analysis reveals that exogenous miR-1246 modulates the polarization of TAMs at a post-transcriptional level, emphasizing the pivotal role of the NLRP3 in macrophage polarization. In conclusion, the findings underscore the importance of exosomal miR-1246 as a trigger of macrophage reprogramming and uncover a novel mechanism for its enhanced presence in the TIME.

Aptamer-bivalent-cholesterol-mediated proximity entropy-driven exosomal protein reporter for tumor diagnosis

Exosomal proteins from the parental cells are considered to be promising biomarker sets for precise tumor diagnostics and monitoring. However, the accurate quantitative analysis of low-abundance exosomal proteins remains challenging due to the heterogeneity of clinical samples. Here, we standardized the exosomal concentration with a fluorogenic membrane probe and developed an aptamer-bivalent-cholesterol-mediated Proximity Entropy-driven Exosomal Protein Reporter (PEEPR). The proposed PEEPR enables the in-situ analysis of multiple exosomal proteins by integrating bivalent cholesterol anchor (exosomal lipid bilayer) and aptamer (exosomal proteins) with a proximity entropy-driven circuit. Based on this strategy, we successfully achieved detection limits of 3.9 pg/mL exosomal GPC-3 and 3.4 pg/mL exosomal PD-L1. Notably, the standardization of exosome concentrations is designed to avoid errors due to biological heterogeneity. The results showed that evaluating the levels of exosomal GPC-3 and PD-L1 in clinical samples via this strategy could accurately differentiate healthy individuals, hepatitis B patients, and hepatocellular carcinoma patients. In summary, PEEPR is a promising clinical diagnostic strategy for the quantitative analysis of a variety of tumor-associated exosomal proteins for the precise diagnosis and personalized treatment monitoring of tumors.

Accurate Cancer Screening and Prediction of PD-L1-Guided Immunotherapy Efficacy Using Quantum Dot Nanosphere Self-Assembly and Machine Learning.

Accurate quantification of exosomal PD-L1 protein in tumors is closely linked to the response to immunotherapy, but robust methods to achieve high-precision quantitative detection of PD-L1 expression on the surface of circulating exosomes are still lacking. In this work, we developed a signal amplification approach based on aptamer recognition and DNA scaffold hybridization-triggered assembly of quantum dot nanospheres, which enables bicolor phenotyping of exosomes to accurately screen for cancers and predict PD-L1-guided immunotherapeutic effects through machine learning. Through DNA-mediated assembly, we utilized two aptamers for simultaneous ultrasensitive detection of exosomal antigens, which have synergistic roles in tumor diagnosis and treatment prediction, and thus, we achieved better sample classification and prediction through machine-learning algorithms. With a drop of blood, we can distinguish between different cancer patients and healthy individuals and predict the outcome of immunotherapy. This approach provides valuable insights into the development of personalized diagnostics and precision medicine.

Miniaturized Laser Probe for Exosome-Based Cancer Liquid Biopsy.

Exosomes have been established as a valuable tool for clinical applications for the purpose of liquid biopsy and therapy. However, the clinical practice of exosomes as cancer biopsy markers is still to a very low extent. Active mode optical microcavity with microlaser emission has aroused as a versatile approach for chemical and biological sensing due to its benefits of larger photon population, increased effective Q-factor, decreased line width, and improved sensitivity. Herein, we report a label-free and precise quantification of exosome vesicles and surface protein profiling of breast cancer exosomes using functionalized active whispering gallery mode (WGM) microlaser probes. A detection limit of 40 exosomes per microresonator was achieved. The proposed system enabled a pilot assay of quantitative exosome analysis in cancer patients' blood with only a few microliters of sample consumption, holding good potential for large-scale cancer liquid biopsy. Multiplexed functionalization of the optical microresonator allowed us to profile cancer exosomal surface markers and distinct subclasses of breast cancer-associated exosomes and monitor drug treatment outcomes. Our findings speak volumes about the advantages of the WGM microlaser sensor, including very small sample consumption, low detection limit, high specificity, and ease of operation, offering a promising means for precious clinical sample analysis.

Integrated analysis of RNA-seq datasets reveals novel targets and regulators of COVID-19 severity.

During the COVID-19 pandemic, RNA-seq datasets were produced to investigate the virus-host relationship. However, much of these data remains underexplored. To improve the search for molecular targets and biomarkers, we performed an integrated analysis of multiple RNA-seq datasets, expanding the cohort and including patients from different countries, encompassing severe and mild COVID-19 patients. Our analysis revealed that severe COVID-19 patients exhibit overexpression of genes coding for proteins of extracellular exosomes, endomembrane system, and neutrophil granules (e.g., S100A9, LY96, and RAB1B), which may play an essential role in the cellular response to infection. Concurrently, these patients exhibit down-regulation of genes encoding components of the T cell receptor complex and nucleolus, including TP53, IL2RB, and NCL Finally, SPI1 may emerge as a central transcriptional factor associated with the up-regulated genes, whereas TP53, MYC, and MAX were associated with the down-regulated genes during COVID-19. This study identified targets and transcriptional factors, lighting on the molecular pathophysiology of syndrome coronavirus 2 infection.