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Theranostics 2024Device implantation frequently triggers cardiac remodeling and fibrosis, with monocyte-driven inflammatory responses precipitating arrhythmias. This study investigates...
Device implantation frequently triggers cardiac remodeling and fibrosis, with monocyte-driven inflammatory responses precipitating arrhythmias. This study investigates the role of mA modification enzymes METTL3 and METTL14 in these responses and explores a novel therapeutic strategy targeting these modifications to mitigate cardiac remodeling and fibrosis. Peripheral blood mononuclear cells (PBMCs) were collected from patients with ventricular septal defects (VSD) who developed conduction blocks post-occluder implantation. The expression of METTL3 and METTL14 in PBMCs was measured. METTL3 and METTL14 deficiencies were induced to evaluate their effect on angiotensin II (Ang II)-induced myocardial inflammation and fibrosis. mA modifications were analyzed using methylated RNA immunoprecipitation followed by quantitative PCR. NF-κB pathway activity and levels of monocyte migration and fibrogenesis markers (CXCR2 and TGF-β1) were assessed. An erythrocyte microvesicle-based nanomedicine delivery system was developed to target activated monocytes, utilizing the METTL3 inhibitor STM2457. Cardiac function was evaluated via echocardiography. Significant upregulation of METTL3 and METTL14 was observed in PBMCs from patients with VSD occluder implantation-associated persistent conduction block. Deficiencies in METTL3 and METTL14 significantly reduced Ang II-induced myocardial inflammation and fibrosis by decreasing mA modification on and mRNAs. This disruption reduced NF-κB pathway activation, lowered CXCR2 and TGF-β1 levels, attenuated monocyte migration and fibrogenesis, and alleviated cardiac remodeling. The erythrocyte microvesicle-based nanomedicine delivery system effectively targeted inflamed cardiac tissue, reducing inflammation and fibrosis and improving cardiac function. Inhibiting METTL3 and METTL14 in monocytes disrupts the NF-κB feedback loop, decreases monocyte migration and fibrogenesis, and improves cardiac function. Targeting mA modifications of monocytes with STM2457, delivered via erythrocyte microvesicles, reduces inflammation and fibrosis, offering a promising therapeutic strategy for cardiac remodeling associated with device implantation.
Topics: Humans; Methyltransferases; Monocytes; Fibrosis; Male; Animals; NF-kappa B; Erythrocytes; Adenosine; Female; Methylation; Mice; Transforming Growth Factor beta1; Cell-Derived Microparticles; Leukocytes, Mononuclear; Angiotensin II; Receptors, Interleukin-8B; Ventricular Remodeling; Myocardium; Nanomedicine
PubMed: 38948064
DOI: 10.7150/thno.95664 -
Journal of Extracellular Biology Jul 2024Neuroinflammation is initiated through microglial activation and cytokine release which can be induced through lipopolysaccharide treatment (LPS) leading to a...
Neuroinflammation is initiated through microglial activation and cytokine release which can be induced through lipopolysaccharide treatment (LPS) leading to a transcriptional cascade culminating in the differential expression of target proteins. These differentially expressed proteins can then be packaged into extracellular vesicles (EVs), a form of cellular communication, further propagating the neuroinflammatory response over long distances. Despite this, the EV proteome in the brain, following LPS treatment, has not been investigated. Brain tissue and brain derived EVs (BDEVs) isolated from the cortex of LPS-treated mice underwent thorough characterisation to meet the minimal information for studies of extracellular vesicles guidelines before undergoing mass spectrometry analysis to identify the differentially expressed proteins. Fourteen differentially expressed proteins were identified in the LPS brain tissue samples compared to the controls and 57 were identified in the BDEVs isolated from the LPS treated mice compared to the controls. This included proteins associated with the initiation of the inflammatory response, epigenetic regulation, and metabolism. These results allude to a potential link between small EV cargo and early inflammatory signalling.
PubMed: 38947878
DOI: 10.1002/jex2.165 -
Journal of Extracellular Biology Jul 2024Previously, we showed that quantification of lymphoma-associated miRNAs miR-155-5p, -127-3p and let-7a-5p levels in plasma extracellular vesicles (EVs) report treatment...
Towards IVDR-compliance by implementing quality control steps in a quantitative extracellular vesicle-miRNA liquid biopsy assay for response monitoring in patients with classic Hodgkin lymphoma.
Previously, we showed that quantification of lymphoma-associated miRNAs miR-155-5p, -127-3p and let-7a-5p levels in plasma extracellular vesicles (EVs) report treatment response in patients with classic Hodgkin lymphoma (cHL). Prior to clinical implementation, quality control (QC) steps and validation are required to meet international regulatory standards. Most published EV-based diagnostic assays have yet to meet these requirements. In order to advance the assay towards regulatory compliance (e.g., IVDR 2017/746), we incorporated three QC steps in our experimental EV-miRNA quantitative real-time reverse-transcription PCR (q-RT-PCR) assay in an ISO-13485 certified quality-management system (QMS). Liposomes encapsulated with a synthetic (nematode-derived) miRNA spike-in controlled for EV isolation by automated size-exclusion chromatography (SEC). Additional miRNA spike-ins controlled for RNA isolation and cDNA conversion efficiency. After deciding on quality criteria, in total 107 out of 120 samples from 46 patients passed QC. Generalized linear mixed-effect modelling with bootstrapping determined the diagnostic performance of the quality-controlled data at an area under the curve (AUC) of 0.84 (confidence interval [CI]: 0.76-0.92) compared to an AUC of 0.87 (CI: 0.80-0.94) of the experimental assay. After the inclusion of QC steps, the accuracy of the assay was determined to be 78.5% in predicting active disease status in cHL patients during treatment. We demonstrate that a quality-controlled plasma EV-miRNA assay is technically robust, taking EV-miRNA as liquid biopsy assay an important step closer to clinical evaluation.
PubMed: 38947877
DOI: 10.1002/jex2.164 -
Journal of Extracellular Biology Jul 2024multiple nucleopolyhedrovirus (AcMNPV) is an enveloped DNA virus of the family. This baculovirus is widely exploited for the biological control of insect pest species...
multiple nucleopolyhedrovirus (AcMNPV) is an enveloped DNA virus of the family. This baculovirus is widely exploited for the biological control of insect pest species and as an expression platform to produce recombinant proteins in insect cells. Extracellular vesicles (EVs) are secreted by all cells and are involved in key roles in many biological processes through their cargo consisting of proteins, RNA or DNA. In viral infections, EVs have been found to transfer both viral and cellular cargo that can elicit either a pro- or antiviral response in recipient cells. Here, small EVs (sEVs) released by (Sf) insect cells were characterised for the first time. Using (SfC1B5) cells stably expressing the baculovirus , the viral envelope protein GP64 was shown to be incorporated into sEVs. Sf9 cells were also transfected with a bacmid AcMNPV genome lacking (AcΔP6.9) to prevent budded virus production. The protein content of sEVs from both mock- and AcΔP6.9-transfected cells were analysed by mass spectrometry. In addition to GP64, viral proteins Ac-F, ME-53 and viral ubiquitin were identified, as well as many host proteins including TSG101-which may be useful as a protein marker for sEVs.
PubMed: 38947876
DOI: 10.1002/jex2.163 -
Journal of Cancer 2024Bone cancer among adolescents and children exhibits varying survival outcomes based on disease state. While localized bone cancer cases have a survival rate exceeding... (Review)
Review
Bone cancer among adolescents and children exhibits varying survival outcomes based on disease state. While localized bone cancer cases have a survival rate exceeding 70%, metastatic, refractory, and recurrent forms are associated with significantly poorer prognoses. Initially believed to be mere vehicles for cellular waste disposal, exosomes are now recognized as extracellular vesicles facilitating intercellular communication. These vesicles influence cellular behaviors by transporting various biomolecules, such as proteins, DNA, RNA, and lipids, among cells. The role of exosomes in regulating the progression of bone cancer is increasingly evident, impacting critical processes like tumorigenesis, proliferation, metastasis, angiogenesis, immune evasion, and drug resistance. Current research underscores the substantial potential of exosomes in promoting the progression and development of bone cancer. This review delves into the complex process of exosome biogenesis, the variety of cell-derived exosome sources, and their applications in drug delivery and therapeutics. It also examines ongoing clinical trials focused on exosome cargo levels and discusses the challenges and future directions in exosome research. Unlike costly and invasive traditional diagnostic methods, exosomal biomarkers offer a non-invasive, cost-effective, and readily accessible routine screening through simple fluid collection that aims to inspire researchers to investigate the potential of exosomes for cancer theragnostic. Through comprehensive exploration of these areas, the review seeks to enhance understanding and foster innovative solutions to cancer biology in the near future.
PubMed: 38947401
DOI: 10.7150/jca.95709 -
Frontiers in Immunology 2024CD39 plays an important role in the immunoregulation and inhibition of effector cells. It is expressed on immune cells, including Tregs, and on extracellular vesicles...
INTRODUCTION
CD39 plays an important role in the immunoregulation and inhibition of effector cells. It is expressed on immune cells, including Tregs, and on extracellular vesicles (EVs) budding from the plasma membrane. Platelet transfusion may induce alloimmunization against HLA-I antigens, leading to refractoriness to platelet transfusion with severe consequences for patients. Tregs may play a key role in determining whether alloimmunization occurs in patients with hematologic disorders. We hypothesized that CD39 EVs might play an immunoregulatory role, particularly in the context of platelet transfusions in patients with hematologic disorders. Such alloimmunization leads to the production of alloantibodies and is sensitive to the regulatory action of CD39.
METHODS
We characterized CD39 EVs in platelet concentrates by flow cytometry. The absolute numbers and cellular origins of CD39 EVs were evaluated. We also performed functional tests to evaluate interactions with immune cells and their functions.
RESULTS
We found that CD39 EVs from platelet concentrates had an inhibitory phenotype that could be transferred to the immune cells with which they interacted: CD4 and CD8 T lymphocytes (TLs), dendritic cells, monocytes, and B lymphocytes (BLs). Moreover, the concentration of CD39 EVs in platelet concentrates varied and was very high in 10% of concentrates. The number of these EVs present was determinant for EV-cell interactions. Finally, functional interactions were observed with BLs, CD4 TLs and CD39 EVs for immunoglobulin production and lymphoproliferation, with potential implications for the immunological management of patients.
Topics: Humans; Extracellular Vesicles; Blood Platelets; Tetraspanin 29; Cell Communication; Platelet Transfusion; Female; B-Lymphocytes; Male; Apyrase; T-Lymphocytes, Regulatory; Antigens, CD
PubMed: 38947317
DOI: 10.3389/fimmu.2024.1397967 -
World Journal of Gastroenterology Jun 2024Hepatocellular carcinoma (HCC) is the most common and deadliest subtype of liver cancer worldwide and, therefore, poses an enormous threat to global health....
Hepatocellular carcinoma (HCC) is the most common and deadliest subtype of liver cancer worldwide and, therefore, poses an enormous threat to global health. Understanding the molecular mechanisms underlying the development and progression of HCC is central to improving our clinical approaches. PIWI-interacting RNAs (piRNAs) are a class of small non-coding RNAs that bind to PIWI family proteins to regulate gene expression at transcriptional and post-transcriptional levels. A growing body of work shows that the dysregulation of piRNAs plays a crucial role in the progression of various human cancers. In this editorial, we report on the current knowledge of HCC-associated piRNAs and their potential clinical utility. Based on the editorial by Papadopoulos and Trifylli, on the role and clinical evaluation of exosomal circular RNAs in HCC, we highlight this other emerging class of non-coding RNAs.
Topics: Humans; Liver Neoplasms; Carcinoma, Hepatocellular; RNA, Small Interfering; Gene Expression Regulation, Neoplastic; Exosomes; RNA, Circular; Disease Progression; Biomarkers, Tumor
PubMed: 38947286
DOI: 10.3748/wjg.v30.i22.2843 -
Biomicrofluidics May 2024Small extracellular vesicles (sEVs) are extracellular vesicles with diameters ranging from 30 to 150 nm, harboring proteins and nucleic acids that reflect their source...
Small extracellular vesicles (sEVs) are extracellular vesicles with diameters ranging from 30 to 150 nm, harboring proteins and nucleic acids that reflect their source cells and act as vital mediators of intercellular communication. The comprehensive analysis of sEVs is hindered by the complex composition of biofluids that contain various extracellular vesicles. Conventional separation methods, such as ultracentrifugation and immunoaffinity capture, face routine challenges in operation complexity, cost, and compromised recovery rates. Microfluidic technologies, particularly viscoelastic microfluidics, offer a promising alternative for sEV separation due to its field-free nature, fast and simple operation procedure, and minimal sample consumption. In this context, we here introduce an innovative viscoelastic approach designed to exploit the viscosity gradient-induced force with size-dependent characteristics, thereby enabling the efficient separation of nano-sized particles and sEVs from larger impurities. We first seek to illustrate the underlying mechanism of the viscosity gradient-induced force, followed by experimental validation with fluorescent nanoparticles demonstrating separation results consistent with qualitative analysis. We believe that this work is the first to report such viscosity gradient-induced phenomenon in the microfluidic context. The presented approach achieves ∼80% for both target purity and recovery rate. We further demonstrate effective sEV separation using our device to showcase its efficacy in the real biological context, highlighting its potential as a versatile, label-free platform for sEV analysis in both fundamental biological research and clinical applications.
PubMed: 38947280
DOI: 10.1063/5.0208417 -
Journal of Extracellular Biology Jun 2024Congenital diaphragmatic hernia (CDH) is a life-threatening condition with high morbidity and mortality rates. The survival rate of neonates with severe CDH is...
Congenital diaphragmatic hernia (CDH) is a life-threatening condition with high morbidity and mortality rates. The survival rate of neonates with severe CDH is reportedly only 10%-15%. However, prenatal prediction of severe cases is difficult, and the discovery of new predictive markers is an urgent issue. In this study, we focused on microRNAs (miRNAs) in amniotic fluid-derived small EVs (AF-sEVs). We identified four miRNAs (hsa-miR-127-3p, hsa-miR-363-3p, hsa-miR-493-5p, and hsa-miR-615-3p) with AUC > 0.8 to classify good prognosis group and poor prognosis group in human study. The AUC for hsa-miR-127-3p and hsa-miR-615-3p, for predicting the poor prognosis, were 0.93 and 0.91, respectively. In addition, in the in vivo study, the miRNA profiles of the lung tissues of CDH rats were different from those of control rats. Additionally, two elevated miRNAs (rno-miR-215-5p and rno-miR-148a-3p) in the lung tissues of CDH rats were increased in the AF-sEVs of CDH rats. Our results suggest that severe CDH neonates can be predicted prenatally with high accuracy using miRNAs contained in AF-sEVs. Furthermore, miRNA profile changes in AF-sEVs reflected the lung status in CDH. Our findings may contribute to the development of advanced perinatal care for patients with CDH.
PubMed: 38947173
DOI: 10.1002/jex2.160 -
Journal of Extracellular Biology Jun 2024Chemoresistance is a common problem in ovarian cancer (OvCa) treatment, where resistant cells, in response to chemotherapy, secrete small extracellular vesicles (sEVs),...
Chemoresistance is a common problem in ovarian cancer (OvCa) treatment, where resistant cells, in response to chemotherapy, secrete small extracellular vesicles (sEVs), known as chemo-sEVs, that transfer resistance to recipient cells. sEVs are formed as intraluminal vesicles (ILVs) within multivesicular endosomes (MVEs), whose trafficking is regulated by Ras-associated binding (RAB) GTPases that mediate sEVs secretion or lysosomal degradation. A decrease in lysosomal function can promote sEVs secretion, but the relationship between MVEs trafficking pathways and sEVs secretion in OvCa chemoresistance is unclear. Here, we show that A2780cis cisplatin (CCDP) resistant OvCa cells had an increased number of MVEs and ILVs structures, higher levels of Endosomal Sorting Complex Required for Transport (ESCRTs) machinery components, and RAB27A compared to A2780 CDDP-sensitive OvCa cells. CDDP promoted the secretion of chemo-sEVs in A2780cis cells, enriched in DNA damage response proteins. A2780cis cells exhibited poor lysosomal function with reduced levels of RAB7, essential in MVEs-Lysosomal trafficking. The silencing of RAB27A in A2780cis cells prevents the Chemo-EVs secretion, reduces its chemoresistance and restores lysosomal function and levels of RAB7, switching them into an A2780-like cellular phenotype. Enhancing lysosomal function with rapamycin reduced chemo-sEVs secretion. Our results suggest that adjusting the balance between secretory MVEs and lysosomal MVEs trafficking could be a promising strategy for overcoming CDDP chemoresistance in OvCa.
PubMed: 38947172
DOI: 10.1002/jex2.157