Extracellular vesicles and circulating nucleic acids最新文献

Host-bacteria and bacteria-bacteria interactions can be facilitated by extracellular vesicles (EVs) secreted by both human and bacterial cells. Human and bacterial EVs (BEVs) propagate and transfer immunogenic cargos that may elicit immune responses in nearby or distant recipient cells/tissues. Hence, direct colonization of tissues by bacterial cells is not required for immunogenic stimulation. This phenomenon is important in the feto-maternal interface, where optimum tolerance between the mother and fetus is required for a successful pregnancy. Though the intrauterine cavity is widely considered sterile, BEVs from diverse sources have been identified in the placenta and amniotic cavity. These BEVs can be internalized by human cells, which may help them evade host immune surveillance. Though it appears logical, whether bacterial cells internalize human EVs or human EV cargo is yet to be determined. However, the presence of BEVs in placental tissues or amniotic cavity is believed to trigger a low-grade immune response that primes the fetal immune system for ex-utero survival, but is insufficient to disrupt the progression of pregnancy or cause immune intolerance required for adverse pregnancy events. Nevertheless, the exchange of bioactive cargos between human and BEVs, and the mechanical underpinnings and health implications of such interactions, especially during pregnancy, are still understudied. Therefore, while focusing on the feto-maternal interface, we discussed how human cells take up BEVs and whether bacterial cells take up human EVs or their cargo, the exchange of cargos between human and BEVs, host cell (feto-maternal) inflammatory responses to BEV immunogenic stimulation, and associations of these interactions with fetal immune priming and adverse reproductive outcomes such as preeclampsia and preterm birth.

Extracellular vesicles (EVs) contribute to the development of cancer in various ways. Non-Hodgkin lymphoma (NHL) is a cancer of mature lymphocytes and the most common hematological malignancy globally. The most common form of NHL, diffuse large B-cell lymphoma (DLBCL), is primarily treated with chemotherapy, autologous stem cell transplantation (ASCT), and/or chimeric antigen receptor T-cell (CAR-T) therapy. With NHL disease progression and its treatment, extracellular vesicles play remarkable roles in influencing outcomes. This finding can be utilized for therapeutic intervention to improve patient outcomes for NHL. This review focuses on the multifaceted roles of EVs with NHL and its potential for guiding patient care.

Aim: The objective of the present study was to determine the variability of platelet lysate-derived extracellular vesicles (pEV), in terms of characteristics and functionality through wound healing assays, when isolated either from platelet concentrates (PC, obtained from 5 donors) or from multiple PC (MPC, that is 50 donors). Methods: pEV were isolated under GMP-like conditions in a clean room using Size Exclusion Chromatography (SEC). The differential characteristics between pEV obtained from PC (PC-EV) or MPC (MPC-EV) were evaluated by means of protein concentration, Nanoparticle Tracking Analysis (NTA), Transmission Electron Microscopy (TEM), and flow cytometry using the MACSPlex™ arrays for surface analysis profiling of EV. The functionality of the isolated pEV was determined in cell culture by metabolic activity and LDH activity determination and through a wound healing assay after 24 h treatment. Results: No significant differences were observed in the pEV characteristics evaluated, whether isolated from PC or MPC. As regards functionality, a higher wound closure percentage was obtained in those pEV pools isolated from PC (5 donors). No differences in the coefficient of variation (CV) were found when comparing all the evaluated variables of pEV derived either from PC (5 donors) or from MPC (50 donors). Conclusion: Our findings challenge the necessity of a larger donor pool for pEV isolation, revealing no significant variations in the analyzed variables of MPC-EV and PC-EV. Notably, our results suggest that, unlike platelet concentrates, a high number of donors is not required to reduce the variability of pEV, showing that the pool of only 5 donors can provide a consistent and reliable therapeutic product.

This review article presents a detailed examination of the integral role that electrochemical detection of extracellular vesicles (EVs) plays, particularly focusing on the potential application for early disease diagnostics through EVs biomarker analysis. Through an exploration of the benefits and challenges presented by electrochemical detection vetted for protein, lipid, and nucleic acid biomarker analysis, we underscore the significance of these techniques. Evidence from recent studies renders this detection modality imperative in identifying diverse biomarkers from EVs, leading to early diagnosis of diseases such as cancer and neurodegenerative disorders. Recent advancements that have led to enhanced sensitivity, specificity and point-of-care testing (POCT) potential are elucidated, along with equipment deployed for electrochemical detection. The review concludes with a contemplation of future perspectives, recognizing the potential shifts in disease diagnostics and prognosis, necessary advances for broad adoption, and potential areas of ongoing research. The objective is to propel further investigation into this rapidly burgeoning field, thereby facilitating a potential paradigm shift in disease detection, monitoring, and treatment toward human health management.

Aims: Analysis of miRNA (18-23nt) encapsulated in small extracellular vesicles (sEVs) (diameter ~30-200 nm) is critical in understanding the diagnostic and therapeutic value of sEV miRNA. However, various sEV enrichment techniques yield different quantities and qualities of sEV miRNA. Here, we compare the efficacy of three sEV isolation techniques in four combinations for miRNA next-generation sequencing.

Methods: Blood plasma from four Holstein-Friesian dairy cows (Bos taurus) (n = 4) with similar genetic traits and physical characteristics were pooled to isolate sEV. Ultracentrifugation (UC) (100,000 × g, 2 h at 4 °C), size-exclusion chromatography (SEC) and ultrafiltration (UF) were used to design four groups of sEV isolations (UC+SEC, SEC+UC, SEC+UF and UC+SEC+UF). sEV miRNAs were isolated using a combination of TRIzol, Chloroform and miRNeasy mini kit (n = 4/each), later sequenced utilizing Novaseq S1 platform (single-end 100 bp sequencing).

Results: All four sEV methods yielded > 1,700 miRNAs and sEV miRNAs demonstrated a clear separation from control blood plasma circulating miRNA (PCA analysis). MiR-381-3p, miR-23-3p, and miR-18b-3p are among the 25 miRNAs unique to sEV, indicating potential sEV-specific miRNA markers. Further, those 25 miRNAs mostly regulate immune-related functions, indicating the value of sEV miRNA cargo in immunology.

Conclusion: The four sEV miRNA isolation methods employed in this study are valid techniques. The choice of method depends on the research question and study design. If purity is of concern, the UC+SEC method resulted in the best particles/µg protein ratio, which is often used as an indication of sample purity. These results could eventually establish sEV miRNAs as effective diagnostic and therapeutic tools of immunology.

Background: Vascular cell adhesion molecule-1 (VCAM-1⁺) endothelial cell-derived extracellular vesicles (EC-EVs) are augmented in cardiovascular disease, where they can signal the deployment of immune cells from the splenic reserve. Endothelial cells in culture activated with pro-inflammatory tumor necrosis factor-α (TNF-a) also release VCAM-1⁺ EC-EVs. However, isolating VCAM-1⁺ EC-EVs from conditioned cell culture media for subsequent in-depth analysis remains challenging. Aim: We utilized the extracellular vesicles (EV) microfluidics herringbone chip (^EVHB-Chip), coated with anti-VCAM-1 antibodies, for selective capture of VCAM-1⁺ cells and EC-EVs. Methods and Results: Engineered EA.hy926 endothelial cells overexpressing VCAM-1 (P < 0.001 versus control) showed increased binding to the VCAM-1- ^EVHB-Chip versus an IgG device. TNF-α-stimulated human umbilical cord vein endothelial cells (HUVECs) exhibited elevated VCAM-1 protein levels (P < 0.001) and preferential binding to the VCAM-1- ^EVHB-Chip versus the IgG device. HUVECs stimulated with TNF-α showed differential gene expression of intercellular adhesion molecule-1 (ICAM-1) (P < 0.001) and VCAM-1 (P < 0.001) by digital droplet PCR versus control cells. HUVEC-derived EC-EVs were positive for CD9, CD63, HSP70, and ALIX and had a modal size of 83.5 nm. Control and TNF-α-stimulated HUVEC-derived EC-EV cultures were captured on the VCAM-1- ^EVHB-Chip, demonstrating selective capture. VCAM-1⁺ EC-EV were significantly enriched for ICAM-1 (P < 0.001) mRNA transcripts. Conclusion: This study presents a novel approach using the ^EVHB-Chip, coated with anti-VCAM-1 antibodies and digital droplet PCR for the study of VCAM-1⁺ EC-EVs. Isolation of VCAM-1⁺ EC-EV from heterogeneous sources such as conditioned cell culture media holds promise for subsequent detailed characterization, and may facilitate the study of VCAM-1⁺ EC-EVs in cardiovascular and metabolic diseases, for disease monitoring and therapeutic insights.

Stem cell therapy is a novel approach for treating various severe and intractable diseases, including autoimmune disorders, organ transplants, tumors, and neurodegenerative diseases. Nevertheless, the extensive utilization of stem cells is constrained by potential tumorigenicity, challenges in precise differentiation, rejection concerns, and ethical considerations. Extracellular vesicles possess the ability to carry diverse bioactive factors from stem cells and deliver them to specific target cells or tissues. Moreover, they offer the advantage of low immunogenicity. Consequently, they have the potential to facilitate the therapeutic potential of stem cells, mitigating the risks associated with direct stem cell application. Therefore, the use of stem cell extracellular vesicles in clinical diseases has received increasing attention. This review summarizes advances in the use of extracellular vesicles from mesenchymal stem cells (MSC). MSC extracellular vesicles are used in the treatment of inflammatory diseases such as rheumatoid arthritis, liver injury, COVID-19, and allergies; in the repair of tissue damage in heart disease, kidney injury, and osteoarthritic diseases; as a carrier in the treatment of tumors; and as a regenerative agent in neurodegenerative disorders such as Alzheimer's and Parkinson's.

Cell-to-cell communication is believed to be facilitated by membrane-bound vesicles called extracellular vesicles (EVs), which are released by cells. Protein, lipids, and nucleic acids are major cargo of EVs and are transported in these vesicles. Depending on the parent and recipient cell types, they can affect a wide variety of biological processes in the tissues to which they are delivered. EVs are essential for embryo implantation and endometriosis, and they are located in the uterine cavities of different species, where they promote blastocyst and endometrial preparation for implantation. This review focuses on what is currently understood regarding pathologic and diagnostic characteristics, and the potential therapeutic value of EVs in the context of endometriosis, where they can be used for drug delivery and targeted therapy due to their ability to carry bioactive molecules to specific cells or tissues. The findings of this review highlight the potential for a wide range of clinical applications that involve endometrial EVs in the areas of treatment, such as surgical and pharmacological, diagnostic biomarker development, and drug delivery systems, all with the ultimate goal of improving pregnancy success rates.