Cells and exosomes after cell-free culture
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ABSTRACT:
PROVIDER: PRJNA259429 | ENA |
REPOSITORIES: ENA
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Project description:The goal of this study is to report that breast cancer associated exosomes contain microRNAs (miRNAs) associated with the RISC Loading Complex (RLC) and display cell-independent capacity to process precursor microRNAs (pre-miRNAs) into mature miRNAs. Pre-miRNAs, along with Dicer, AGO2, and TRBP, are present in exosomes of cancer cells. CD43 mediates the accumulation of Dicer specifically in cancer exosomes. Cancer exosomes mediate an efficient and rapid silencing of mRNAs to reprogram the target cell transcriptome. Exosomes derived from cells and sera of patients with breast cancer instigate non-tumorigenic epithelial cells to form tumors in a Dicer-dependent manner. These findings offer opportunities for the development of exosomes based biomarkers and therapies.
2014-08-27 | GSE60715 | GEO
Project description:After isolation, islets were cultured in a serum-exosomes-free culture media for one week. Collected culture media were centrifuged first at 300g for 20 min to pellets cells and then at 10000g for 20 min to discard dead cells and cell debris. Exosomes were then isolated from the supernatant by ultracentrifugation at 110000g for 70 min. Exosomes were collected in a minimal volume of PBS,and added three times the volume of Trizol LS to extract exosomes RNA.
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2015-05-19 | E-GEOD-68979 | biostudies-arrayexpress
Project description:To investigate the difference of miRNA expression in exosomes derived from A549 cells and its DDP-resistant cell strain A549/DDP, we have employed miRNA microarray expression to discover the difference expression of miRNAs in exosomes derived from A549 and A549/DDP. We conducted RT-qPCR to examine the expression levels of top differential expressed miRNAs, namely, miR-197-5p, miR-4443, miR-642a-3p, miR-27b-3p and miR-100-5p, confirming low variability between two methods. The A549/DDP was established from A549 in our laboratory, by exposing A549 to gradually increasing DDP concentrations, until the final concentration at 1μg/ml. To avoid the influence of drug to the A549/DDP cells, they were cultured in a drug-free medium for at least two weeks before gene expression analysis. After being incubated for 48-72h, the culture medium of cells was harvested. Exosomes were isolated by ultracentrifugation. And miRNA expression of exosomes derived from A549 and A549/DDP was then analzyed.
2016-08-16 | E-GEOD-85604 | biostudies-arrayexpress
Project description:Aim: EBV encode at least 44 miRNAs involved in immune regulation and disease progression. Exosomes can be used as carriers of EBV-miRNA-BART intercellular transmission and affect the biological behavior of cells. We characterized exosomes and established a co-culture experiment of exosomes to explore the mechanism of miR-BART1-3p transmission through the exosome pathway and its influence on tumor cell proliferation and invasion. Materials and methods: Exosomes of EBV-positive and EBV-negative gastric cancer cells were characterized by transmission electron microscopy, Nanosight and western blotting, and miRNA expression profiles in exosomes were sequenced with high throughput. Exosomes with high or low expression of miR-BART1-3p were co-cultured with AGS cells to study the effects on proliferation, invasion and migration of gastric cancer cells. The target genes of EBV-miR-BART1-3p were screened and predicted by PITA, miRanda, RNAhybrid, virBase and Diana-Tarbase v.8 databases, and the expression of the target genes after co-culture was detected by qPCR. Results：The exosomes secreted by EBV positive and negative gastric cancer cells range in diameter from 30 nm to 150nm and express the exosomal signature proteins CD63 and CD81. High throughput sequencing showed that exosomes expressed some human miRNAs, among which has-miR-23b-3p, has-miR-320a-3p and has-miR-4521 were highly expressed in AGS-exo. has-miR-21-5p, has-miR-148a-3p and has-miR-7-5p were highly expressed in SNU-719-exo. All EBV miRNAs were expressed in SNU-719 cells and their exosomes, among which EBV- miR-Bart1-5p, EBV- miR-bart22 and EBV- miR-bart16 were the highest in SNU-719 cells. EBV-miR-BART1-5p, EBV-miR-BART10-3p and EBV-miR-BART16 were the highest in SNU-719-exo. After miR-BART1-3p silencing in gastric cancer cells, the proliferation, healing, migration and invasion of tumor cells were significantly improved. Laser confocal microscopy showed that exosomes could carry miRNA into recipient cells. After co-culture with miR-BART1-3p silenced exosomes, the proliferation, healing, migration and invasion of gastric cancer cells were significantly improved. The target gene of miR-BART1-3p was FMA168A, MACC1, CPEB3, ANKRD28 and USP37 after screening by targeted database. CPEB3 was not expressed in all exosome co-cultured cells, while ANKRD28, USP37, MACC1 and FAM168A were all expressed to varying degrees. USP37 and MACC1 are down-regulated after up-regulation of miR-BART1-3p, which may be the key target genes for miR-BART1-3p to regulate the proliferation of gastric cancer cells through exosomes. Conclusions: miR-BART1-3p can affect the growth of tumor cells through exosome pathway. The proliferation, healing, migration and invasion of gastric cancer cells were significantly improved after co-culture with exosomes of miR-BART1-3p silenced expression. USP37 and MACC1 may be potential target genes of miR-BART1-3p in regulating cell proliferation.
2023-03-13 | GSE227017 | GEO
Project description:Human liver sinusoidal endothelial cells (LSECs) were grown in culture medium supplemented with 10% FBS (depleted of endogenous exosomes by overnight centrifugation at 100,000 g) and were treated with or without 1,000 U/ml IFN- alpha (PBL Interferon Source, Piscataway, NJ, USA) for 48 h. The exosomes from the cell culture supernatants were isolated by differential centrifugation as follow, at 300g for 10 min, 2,000g for 10 min, 10,000g for 30 min, and 100,000g for 70 min, washed once with PBS, and purified by centrifugation at 100,000g for 70 min. Total RNA of purified exosomes were extracted and Agilent Human 4x44K Gene Expression Array.
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2022-06-09 | PXD033124 | Pride
Project description:The goal of this study is to report that breast cancer associated exosomes contain microRNAs (miRNAs) associated with the RISC Loading Complex (RLC) and display cell-independent capacity to process precursor microRNAs (pre-miRNAs) into mature miRNAs. Pre-miRNAs, along with Dicer, AGO2, and TRBP, are present in exosomes of cancer cells. CD43 mediates the accumulation of Dicer specifically in cancer exosomes. Cancer exosomes mediate an efficient and rapid silencing of mRNAs to reprogram the target cell transcriptome. Exosomes derived from cells and sera of patients with breast cancer instigate non-tumorigenic epithelial cells to form tumors in a Dicer-dependent manner. These findings offer opportunities for the development of exosomes based biomarkers and therapies. Exosomes from cancer cells and non-tumorigenic cells were isolated using established ultracentrifugation methods. The global miRNA content of cancer exosomes and normosomes was investigated. Profiling of cells themselves was also used as a control. Exosomes with Dicer down regulation (MCF10AshDicer and MDA-MB-231shDicer exosomes), as well as MDA-MB-231 exosomes that contain a Dicer antibody inside were used to study the function Dicer protein in the microRNA biogenesis in exosomes.
2014-08-27 | E-GEOD-60714 | biostudies-arrayexpress
Project description:The therapeutic functions of stem cells were mainly depending on paracrine effects rather than their own proliferation and differentiation on site. In advantage of easy to collected from conditioned medium, have contents of cytokines, growth factors, signaling lipids, mRNAs, and regulatory miRNAs, exosomes from stem cells were considered to be a promising material for stem cell-based cell-free therapies. In traditional Chinese medicine, antler extracts have been used to alleviates aging and aging related diseases. We have demonstrated that exosomes from antler stem cells (ASCs), the source of the annual full regeneration of deer antler, could alleviate cellular senescence and aging related diseases. With exosomes treatment, the senescent phenotype of primary MSCs and hMSCs at late passage were significantly mitigated. When injected into articular cavity, exosomes from ASCs markedly promote joint function recovery and bone and cartilage regeneration. By LC-MS/MS assay, we found that exosomes from ASCs contain various proteins which in the regulation of biological processes including cell proliferation and migration, immunoregulation, wound healing and angiogenesis. With the advantages of abundant sources, low risk of tumorigenicity and immunogenicity, exosomes from ASCs may consider as a promising bio-material for cell-free therapies in the future.
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