Engineering Extracellular Vesicles by Three-dimensional Dynamic Culture of Human Mesenchymal Stem Cells
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE185874 | GEO | 2021/10/15
REPOSITORIES: GEO
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Project description:Extracellular vesicles (EVs) contain proteins, enzymes and metabolites that contribute to the therapeutic potential of human mesenchymal stem cells (hMSCs). However, scale-up production of hMSC EVs has become a major challenge. In current study, hMSCs were grown as 3D aggregates under wave motion to promote EV secretion (3D EVs). mRNA sequencing reveals global transcriptome alterations (e.g., upregulated Wnt, TNF, and Hippo signaling and downregulated cellular senescence) for 3D aggregates. Compared to 2D EVs, the quantity of 3D EVs was enhanced significantly with smaller size, higher miR-21 and miR-22 expression, and the altered protein cargo revealed by proteomics. 3D hMSC aggregates promote activation of the endosomal sorting complexes required for transport (ESCRT) pathway and ESCRT-independent pathway. In addition, 3D EVs rejuvenated stem cells expressing cellular senescence and modulated immune response determined by T lymphocyte and macrophage phenotype assays. In summary, this study provides a promising strategy for high-quality EV production from hMSCs with enhanced therapeutic potentials.
2022-08-12 | PXD028975 | Pride
Project description:Under physiological conditions, extracellular vesicles (EVs) are present simultaneously in the extracellular compartment together with cytokines. Thus, we hypothesized that EVs in combination with cytokines induce different responses of monocyte cells compared to EVs or cytokines alone. Human monocyte U937 cells were incubated with EV-containing or EV-free CCRF human T-cell supernatant, with or without the addition of TNF. U937 cells cultured in EV-free supernatant, supernatant containing CCRF t-cell derived EVs, TNF or both. Each treatment option was measured in 3 replicates.
2013-06-19 | E-GEOD-47897 | biostudies-arrayexpress
Project description:Osteolineage cells represent one of the critical bone marrow niche components that support maintenance of hematopoietic stem and progenitor cells (HSPCs). Recent studies demonstrate that extracellular vesicles (EVs) regulate stem cell development via horizontal transfer of bioactive cargo, including microRNAs (miRNAs). Here, we characterize the miRNA profile of EVs secreted by human osteoblasts and study their biological effect of on human umbilical cord blood-derived CD34+ HSPCs by sequencing, gene expression and biochemical analyses. Using next-generation sequencing we show that osteoblast-derived EVs contain highly abundant miRNAs specifically enriched in EVs, including critical regulators of hematopoietic proliferation (e.g., miR-29a). EV treatment of CD34+ HSPCs alters the expression of candidate miRNA targets, such as HBP1, BCL2 and PTEN. Furthermore, EVs enhance proliferation of CD34+ cells and their immature subsets in growth factor-driven ex vivo expansion cultures. Importantly, EV-expanded cells retain their differentiation capacity in vitro and show successful engraftment in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice in vivo. These discoveries reveal a novel osteoblast-derived EV-mediated mechanism for regulation of HSPC proliferation and warrant consideration of EV-miRNAs for the development of expansion strategies to treat hematological disorders.
2016-10-05 | E-MTAB-4560 | biostudies-arrayexpress
Project description:Phenotypic changes induced by extracellular vesicles (EVs) have been implicated in the recovery of acute kidney injury (AKI) induced by mesenchymal stromal cells (MSCs). miRNAs are potential candidates for cell reprogramming towards a pro-regenerative phenotype. The aim of the present study was to evaluate whether miRNA de-regulation inhibits the regenerative potential of MSCs and derived-EVs in a model of glycerol-induced AKI in SCID mice. For this purpose, we generated MSCs depleted of Drosha, a critical enzyme of miRNA maturation, to alter miRNA expression within MSCs and EVs. Drosha knock-down MSCs (MSC-Dsh) maintained the phenotype and differentiation capacity. They produced EVs that did not differ from those of wild type cells in quantity, surface molecule expression and internalization within renal tubular epithelial cells. However, EVs derived from MSC-Dsh (EV-Dsh) showed global down-regulation of miRNAs. Whereas, wild type MSCs and derived EVs were able to induce morphological and functional recovery in AKI, MSC-Dsh and EV-Dsh were ineffective. RNA sequencing analysis showed that genes deregulated in the kidney of AKI mice were restored by treatment with MSCs and EVs but not by MSC-Dsh and EV-Dsh. Gene Ontology analysis showed that down-regulated genes in AKI were associated with fatty acid metabolism. The up-regulated genes in AKI were involved in inflammation, ECM-receptor interaction and cell adhesion molecules. These alterations were reverted by treatment with wild type MSCs and EVs, but not by the Drosha counterparts. In conclusion, miRNA depletion in MSCs and EVs significantly reduced their intrinsic regenerative potential in AKI, suggesting a critical role of miRNAs. RNA-seq
2015-05-07 | E-GEOD-59958 | biostudies-arrayexpress
Project description:Beyond forming bone, osteoblasts play pivotal roles in various biological processes, including hematopoiesis and bone metastasis. Extracellular vesicles (EVs) have recently been implicated in intercellular communication via transfer of proteins and nucleic acids between cells. Here, we focused on the proteomic characterization of non-mineralizing (NMOBs) and mineralizing (MOBs) human osteoblast (SV-HFOs) EVs and investigated their effect on human prostate cancer (PC3) cells by microscopic, proteomic and gene expression analyses. Proteomic analysis showed that 97% of the proteins were shared among NMOB and MOB EVs, and 30% were novel osteoblast-specific EV proteins. Label-free quantification demonstrated mineralization stage-dependent five-fold enrichment of 59 and 451 EV proteins in NMOBs and MOBs, respectively. Interestingly, bioinformatic analyses of the osteoblast EV proteomes and EV-regulated prostate cancer gene expression profiles showed that they converged on pathways involved in cell survival and growth. This was verified by in vitro proliferation assays where osteoblast EV uptake led to two-fold increase in PC3 cell growth compared to cell-free culture medium-derived vesicle controls. Our findings elucidate the mineralization stage-specific protein content of osteoblast-secreted EVs, show a novel way by which osteoblasts communicate with prostate cancer, and open up innovative avenues for therapeutic intervention. PC3 cells were treated with extracellular vesicles from non-mineralizing and mineralizing SV-HFOs for three different incubation times (4hrs, 24hrs, 48hr)
2014-10-28 | E-GEOD-56172 | biostudies-arrayexpress
Project description:Extracellular vesicles (EV) are membrane-surrounded vesicles secreted by cells that carry biologically important molecules to the target cells. EVs form a heterogenous group and they represent a novel way of intercellular communication. To study the importance of colorectal cancer (CRC)-derived EVs on stromal fibroblasts, we applied CRC patient-derived 3D organoid cultures and commercially available human colon fibroblasts. We studied the gene expression changes in fibroblasts in the presence and absence pf CRC-derived EVs.
2019-02-25 | GSE114979 | GEO
Project description:Mesenchymal stem cell (MSC) transplantation is a promising therapy for regenerative medicine. However, MSCs cultured in two-dimensional (2D) culture conditions are significantly different from the cell shape in the body, down-regulation of stemness genes and the secretion of paracrine factors. Here, we evaluated the effect of 3D culture using Cellhesion VP, a water-insoluble material composing of chitin-based polysaccharide fibers, on the characteristics of human Wharton’s jelly-derived MSCs (hWJ-MSCs). Cellhesion VP significantly increased the cell proliferation after retrieval. Transcriptome analysis revealed that genes involved in cell stemness, migration ability, and extracellular vesicle (EV) production appeared to be enhanced by 3D culture. Subsequent biochemical analyses showed that the expression levels of stemness genes including OCT4, NANOG, and SSEA4 were up-regulated, and migration capacity was elevated in 3D cultured hWJ-MSCs. In addition, the EV production was significantly increased in 3D cells, which contained a distinct protein profile from 2D cells. Gene and drug connectivity analysis revealed that the 2D and 3D EVs had similar functions as immunomodulators; however, 3D EVs had completely distinct therapeutic profiles on various infectious and metabolic diseases by activating the disease-associated signaling pathways. Therefore, EVs from Cellhesion VP-primed hWJ-MSCs appear as a new treatment for immune and metabolic diseases.
2021-06-30 | GSE152507 | GEO
Project description:Exosomes and microvesicles (i.e., extracellular vesicles; EVs) have been identified within ovarian follicular fluid, and recent evidence suggests that EVs are able to elicit profound effects on ovarian cell function. While existence of miRNA within EVs has been reported, it remains unknown if EV size and concentration as well as their cargos (i.e., proteins and RNA) change during antral follicle growth. Extracellular vesicles isolated from follicular fluid of small, medium and large bovine follicles were similar in size, while concentration of EVs decreased progressively as follicle size increased. Electron microscopy indicated a highly purified population of the lipid bilayer enclosed vesicles that were enriched in exosome biomarkers including CD81 and Alix. Small RNA sequencing identified a large number of known and novel miRNAs that changed in the EVs of different size follicles. Ingenuity Pathway Analysis (IPA) indicated that miRNA abundant in small follicle EV preparations were associated with cell proliferation pathways, while those miRNA abundant in large follicle preparations were related to inflammatory response pathways. These studies are the first to demonstrate that EVs change in their levels and makeup during antral follicle development and point to the potential for a unique vesicle-mediated cell-to-cell communication network within the ovarian follicle. Examination of small RNA population in bovine follicular fluid extracellular vesicles isolated from antral follicles
2016-02-09 | E-GEOD-74879 | biostudies-arrayexpress
Project description:Patient-derived prostate fibroblast primary cultures PCF-54 and PCF-55 were established from two specimens of PC tissues. Urinary EVs were isolated from urine samples of 3 patients with PC and 2 healthy males and used for the treatment of prostate fibroblast primary cultures and normal foreskin fibroblasts. Normoxic and hypoxic EVs were isolated from cell culture medium of PC3 and LNCaP prostate cancer cell lines, cultivated in normoxic and hypoxic conditions respectively. The EV-treated fibroblasts were subjected to RNA sequencing analysis.
2023-04-01 | E-MTAB-11317 | biostudies-arrayexpress
Project description:Background: Acute coronary syndromes (ACS) are associated with aberrant gene expression and epigenetic mechanisms. In particular, de novo DNA methylation is typically linked to gene silencing, but its role in heart disease remains not fully understood. Extracellular vesicles (EVs) are active components in cellular communication for their ability to carry a plethora of signalling biomolecules, thus representing a promising new diagnostic/therapeutic approach in cardiovascular diseases (CVDs). Indeed, there is the need of novel biomarkers for ACS prediction and timely detection. Purpose: We hypothesized that specific epigenetic signals can be carried by EVs. In this regard, we isolated and characterized circulating EVs from ACS patients and evaluated their potential role to influence DNA methylation in target cells. Methods: Circulating EVs were recovered, by ultracentrifugation, from plasma samples of 19 ACS patients and 50 healthy subjects (HS). Nanoparticle tracking analysis (NTA) and western blot (WB) were used to confirm the EVs integrity and purity. Peripheral blood mononuclear cells (PBMCs) of volunteer donors were treated with both ACS and HS derived EVs and genomic DNA was extracted to perform epigenome wide analysis through Reduced Representation Bisulfite Sequencing. ShinyGO, PANTHER, and STRING tools were interrogated to perform GO and PPI network analyses. Flow Cytometry, qRT-PCR, and WB analysis were also performed to evaluate and validate both intra-vesicular and intra-cellular signals. Results: Plasma ACS-derived EVs showed a significant up-regulation of DNA methyltransferases (DNMTs) gene expression levels as compared to HS (P<0.001). Specifically, de novo methylation transcripts, as DNMT3A and DNMT3B were significantly increased in plasma ACS-EVs. DNA methylation analysis of PBMCs from volunteer donors treated with HS- and ACS-derived EVs showed that RNF166 and CCND3 genes resulted the most hyper- and hypo-methylated, respectively, after by ACS-EV treatment. In addition, PPI network analysis specifically evidenced the subnetwork with SRC, as a hub gene, connecting it to NOTCH1, FOXO3, CDC42, IKBKG, RXRA, DGKG, known as important genes already involved in the onset of CVDs. Surprising, other novel genes, BAIAP2, SYP, CHL1, and SHB, which were hypomethylated, were found significantly overexpressed in PBMCs (P<0.005), underlying the fundamental modulating properties of EV cargo in atherosclerosis. Conclusions: These findings support the significant role of ACS plasma-derived EVs, inducing de novo DNA methylation signals, and modulating specific signaling pathways in target cells.
2023-04-03 | E-MTAB-11641 | biostudies-arrayexpress