Genome-wide interrogation of extracellular vesicle biology using barcoded miRNAs.
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ABSTRACT:
SUBMITTER: Lu A
PROVIDER: S-EPMC6312402 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Cardiovascular disease (CVD) is the leading cause of mortality worldwide claiming almost 17. 9 million deaths annually. A primary cause is atherosclerosis within the coronary arteries, which restricts blood flow to the heart muscle resulting in myocardial infarction (MI) and cardiac cell death. Despite substantial progress in the management of coronary heart disease (CHD), there is still a significant number of patients developing chronic heart failure post-MI. Recent research has been focused on promoting neovascularisation post-MI with the ultimate goal being to reduce the extent of injury and improve function in the failing myocardium. Cardiac cell transplantation studies in pre-clinical models have shown improvement in cardiac function; nonetheless, poor retention of the cells has indicated a paracrine mechanism for the observed improvement. Cell communication in a paracrine manner is controlled by various mechanisms, including extracellular vesicles (EVs). EVs have emerged as novel regulators of intercellular communication, by transferring molecules able to influence molecular pathways in the recipient cell. Several studies have demonstrated the ability of EVs to stimulate angiogenesis by transferring microRNA (miRNA, miR) molecules to endothelial cells (ECs). In this review, we describe the process of neovascularisation and current developments in modulating neovascularisation in the heart using miRNAs and EV-bound miRNAs. Furthermore, we critically evaluate methods used in cell culture, EV isolation and administration.
| S-EPMC7546892 | biostudies-literature
Project description:BackgroundBreast milk has abundant extracellular vesicles (EVs) containing various biological molecules (cargo), including miRNAs. EVs are not degraded in the gastrointestinal system and circulation; thus, breast milk EVs (bEVs) are expected to interact with other organs in breastfed infants and modify the gene expression of recipient cells using miRNAs. Maternal pre-pregnancy BMI is a critical factor influencing the composition of breast milk. Thus, in mothers with obesity, miRNAs in bEVs can be altered, which might be associated with adverse health outcomes in infants. In this study, we examined 798 miRNAs to determine which miRNAs are altered in the bEVs of mothers with obesity and their potential impact on breastfed infants.MethodsWe recruited healthy nursing mothers who were either of normal weight (BMI < 25) or with obesity (BMI ≥ 30) based on their pre-pregnancy BMI, and delivered a singleton baby in the prior 6 months. EVs were isolated from breast milk with ultracentrifugation. bEV characteristics were examined by flow cytometry and fluorescence imaging of EV markers. A total of 798 miRNAs were screened using a NanoString human miRNA panel to find differentially expressed miRNAs in bEVs of mothers with obesity compared to mothers of normal weight.ResultsWe included 65 nursing mothers: 47 of normal weight and 18 with obesity based on pre-pregnancy BMI. After bEV isolation, we confirmed the expression of various EV markers. Out of 37 EV markers, CD326 (EpCaM) was the most highly expressed in bEVs. The most abundant miRNAs in bEVs include miR-30b-5p, miR-4454, miR-494-3p, and let-7 miRNAs. Target genes of the top 10 miRNAs were associated with cancer, prolactin pathway, EGFR, ErbB, and FoxO signaling pathway. In bEVs of mothers with obesity, 19 miRNAs were differentially expressed (adjusted p < 0.05 cut-off), which include miR-575, miR-630, miR-642a-3p, and miR-652-5p. These miRNAs and their target genes were associated with neurological diseases and psychological disorders.ConclusionIn this study, we characterized bEVs and demonstrated altered miRNAs in bEVs of mothers with obesity and identified the pathways of their potential target genes. Our findings will provide insight for future studies investigating the role of bEVs in breastfed infants.
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Project description:Mapping the genomic locations of chromatin-associated proteins, such as transcription factors and histone modifications, is key to understanding the mechanisms of transcriptional regulation. ChIPmentation offers a simple and robust way of investigating the genomic binding sites of a protein using relatively low-input material. Here, we present a detailed protocol for the key steps that lead to a successful ChIPmentation experiment, as well as a quick analysis pipeline to examine the data. For complete details on the use and execution of this protocol, please refer to Schmidl et al. (2015). For example data produced by this protocol, please refer to Henriksson et al. (2019) and Zhang et al. (2019).
| S-EPMC7757419 | biostudies-literature
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| S-BSST164 | biostudies-other
Project description:During the chronic stage of Schistosoma infection, the female lays fertile eggs, triggering a strong anti-parasitic Type 2 helper T cell (Th2) immune response. It is unclear how this Th2 response gradually declines even though the worms live for years and continue to produce eggs. Here, we show that S. mansoni downregulates Th2 differentiation in an antigen presenting cell-independent manner, by modulating the Th2-specific transcriptional program. Adult schistosomes secrete miRNA-harboring extracellular vesicles that are internalized by Th cells in-vitro. Schistosomal-miRNAs are found also in T helper cells isolated from Peyer's patches and mesenteric lymph nodes of infected mice. In T helper cells, the schistosomal miR-10 targets MAP3K7 and consequently down-modulates NF-kB activity, a critical transcription factor for Th2 differentiation and function. Our results explain, at least partially, how schistosomes tune down the Th2 response, and provide further insight into the reciprocal geographical distribution between high prevalence of parasitic infections and immune disorders such as allergy. Furthermore, this worm-host crosstalk mechanism can be harnessed to develop diagnostic and therapeutic approaches for human schistosomiasis and Th2-associated diseases.
| S-SCDT-EMBOR-2019-47882V1 | biostudies-other
Project description:To gain insight into the microRNA expression profile of small extracellular vesicle derived from different cell types and to verify their mechanism, we utilized the miRNA sequencing technology to analyze the miRNA profiles of different human cell derived small extracellular vesicle
2024-05-01 | GSE203445 | GEO
Project description:During the chronic stage of Schistosoma infection, the female lays fertile eggs, triggering a strong anti-parasitic type 2 helper T-cell (Th2) immune response. It is unclear how this Th2 response gradually declines even though the worms live for years and continue to produce eggs. Here, we show that Schistosoma mansoni downregulates Th2 differentiation in an antigen-presenting cell-independent manner, by modulating the Th2-specific transcriptional program. Adult schistosomes secrete miRNA-harboring extracellular vesicles that are internalized by Th cells in vitro. Schistosomal miRNAs are found also in T helper cells isolated from Peyer's patches and mesenteric lymph nodes of infected mice. In T helper cells, the schistosomal miR-10 targets MAP3K7 and consequently downmodulates NF-?B activity, a critical transcription factor for Th2 differentiation and function. Our results explain, at least partially, how schistosomes tune down the Th2 response, and provide further insight into the reciprocal geographic distribution between high prevalence of parasitic infections and immune disorders such as allergy. Furthermore, this worm-host crosstalk mechanism can be harnessed to develop diagnostic and therapeutic approaches for human schistosomiasis and Th2-associated diseases.
| S-EPMC6944914 | biostudies-literature
Project description:Cells can communicate with neighboring or distant cells using extracellular vesicles (EVs), mainly attributed to their containing miRNAs. Given that diets can change host circulatory miRNA profiling, and EVs are the major miRNA carriers in serum, we hypothesized that different diets could change bovine circulating EV-miRNA expression. We partly replaced alfalfa hay with whole cotton seed and soybean hull in the feed formula of the tested cows. Blood EVs were isolated using a polyethylene glycol precipitation kit. Particle size analysis revealed exosomes were dominant in bovine serum EVs. Small RNAs were enriched in bovine serum EVs, including miRNAs, snRNAs, tiRNAs, Cis-regulatory elements, piRNAs, etc. In total, 359 types of Bos taurus miRNAs were identified by Solexa sequencing. Each cow in the control group contained about 244 types of serum EV-miRNAs, compared to 246 types in the tested group. There were 15 immune-related miRNAs in the top 20 serum EV-miRNAs, accounting for about 80% of the total. Seven differently expressed known miRNAs were detected in responding to different diets. An analysis of the Kyoto Encyclopedia of Genes and Genomes (KEGG) showed differently expressed miRNAs were related to hormone signal pathways and protein metabolism. Bovine serum EVs are abundant with miRNAs, most of which are immune-related. Different diets eventually change the miRNA profiling of bovine serum EVs.
| S-EPMC6940744 | biostudies-literature
Project description:BackgroundGenome-wide DNA methylation profiling has recently been developed into a tool that allows tumor classification in central nervous system tumors. Extracellular vesicles (EVs) are released by tumor cells and contain high molecular weight DNA, rendering EVs a potential biomarker source to identify tumor subgroups, stratify patients and monitor therapy by liquid biopsy. We investigated whether the DNA in glioblastoma cell-derived EVs reflects genome-wide tumor methylation and mutational profiles and allows noninvasive tumor subtype classification.MethodsDNA was isolated from EVs secreted by glioblastoma cells as well as from matching cultured cells and tumors. EV-DNA was localized and quantified by direct stochastic optical reconstruction microscopy. Methylation and copy number profiling was performed using 850k arrays. Mutations were identified by targeted gene panel sequencing. Proteins were differentially quantified by mass spectrometric proteomics.ResultsGenome-wide methylation profiling of glioblastoma-derived EVs correctly identified the methylation class of the parental cells and original tumors, including the MGMT promoter methylation status. Tumor-specific mutations and copy number variations (CNV) were detected in EV-DNA with high accuracy. Different EV isolation techniques did not affect the methylation profiling and CNV results. DNA was present inside EVs and on the EV surface. Proteome analysis did not allow specific tumor identification or classification but identified tumor-associated proteins that could potentially be useful for enriching tumor-derived circulating EVs from biofluids.ConclusionsThis study provides proof of principle that EV-DNA reflects the genome-wide methylation, CNV, and mutational status of glioblastoma cells and enables their molecular classification.
| S-EPMC8673443 | biostudies-literature
Project description:BackgroundGestational diabetes mellitus (GDM) is a serious public health issue affecting 9-15% of all pregnancies worldwide. Recently, it has been suggested that extracellular vesicles (EVs) play a role throughout gestation, including mediating a placental response to hyperglycaemia. Here, we investigated the EV-associated miRNA profile across gestation in GDM, assessed their utility in developing accurate, multivariate classification models, and determined the signaling pathways in skeletal muscle proteome associated with the changes in the EV miRNA profile.MethodsDiscovery: A retrospective, case-control study design was used to identify EV-associated miRNAs that vary across pregnancy and clinical status (i.e. GDM or Normal Glucose Tolerance, NGT). EVs were isolated from maternal plasma obtained at early, mid and late gestation (n = 29) and small RNA sequencing was performed. Validation: A longitudinal study design was used to quantify expression of selected miRNAs. EV miRNAs were quantified by real-time PCR (cases = 8, control = 14, samples at three times during pregnancy) and their individual and combined classification efficiencies were evaluated. Quantitative, data-independent acquisition mass spectrometry was use to establish the protein profile in skeletal muscle biopsies from normal and GDM.ResultsA total of 2822 miRNAs were analyzed using a small RNA library, and a total of 563 miRNAs that significantly changed (p < 0.05) across gestation and 101 miRNAs were significantly changed between NGT and GDM. Analysis of the miRNA changes in NGT and GDM separately identified a total of 256 (NGT-group), and 302 (GDM-group) miRNAs that change across gestation. A multivariate classification model was developed, based on the quantitative expression of EV-associated miRNAs, and the accuracy to correctly assign samples was > 90%. We identified a set of proteins in skeletal muscle biopsies from women with GDM associated with JAK-STAT signaling which could be targeted by the miRNA-92a-3p within circulating EVs. Interestingly, overexpression of miRNA-92a-3p in primary skeletal muscle cells increase insulin-stimulated glucose uptake.ConclusionsDuring early pregnancy, differently-expressed, EV-associated miRNAs may be of clinical utility in identifying presymptomatic women who will subsequently develop GDM later in gestation. We suggest that miRNA-92a-3p within EVs might be a protected mechanism to increase skeletal muscle insulin sensitivity in GDM.
| S-EPMC8377872 | biostudies-literature