Dataset Information

MicroRNA and mRNA cargo of extracellular vesicles from porcine adipose tissue-derived mesenchymal stem cells.

ABSTRACT: Mesenchymal stromal/stem cells (MSCs) are clinically useful for cell-based therapy, but concerns regarding their ability to replicate limit their human application. MSCs release extracellular vesicles (EVs) that mediate at least in part the paracrine effects of the parental cells. To understand the molecular basis of their biological properties, we characterized the RNA cargo of EVs from porcine adipose-tissue derived MSCs. Comprehensive characterization of mRNA and miRNA gene expression using high-throughput RNA sequencing (RNA-seq) revealed that EVs are selectively enriched for distinct classes of RNAs. For example, EVs preferentially express mRNA for transcription factors (e.g. MDFIC, POU3F1, NRIP1) and genes involved in angiogenesis (e.g. HGF, HES1, TCF4) and adipogenesis (e.g. CEBPA, KLF7). EVs also express Golgi apparatus genes (ARRB1, GOLGA4) and genes involved in TGF-? signaling. In contrast, mitochondrial, calcium signaling, and cytoskeleton genes are selectively excluded from EVs, possibly because these genes remain sequestered in organelles or intracellular compartments. RNA-seq generated reads for at least 386 annotated miRNAs, but only miR148a, miR532-5p, miR378, and let-7f were enriched in EVs compared to MSCs. Gene ontology analysis indicates that these miRNAs target transcription factors and genes that participate in several cellular pathways, including angiogenesis, cellular transport, apoptosis, and proteolysis. Our data suggest that EVs transport gene regulatory information to modulate angiogenesis, adipogenesis, and other cell pathways in recipient cells. These observations may contribute to development of regenerative strategies using EVs to overcome potential complications of cell-based therapy.

SUBMITTER: Eirin A

PROVIDER: S-EPMC4174680 | biostudies-literature | 2014 Nov

REPOSITORIES: biostudies-literature

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MicroRNA and mRNA cargo of extracellular vesicles from porcine adipose tissue-derived mesenchymal stem cells.

Eirin Alfonso A Riester Scott M SM Zhu Xiang-Yang XY Tang Hui H Evans Jared M JM O'Brien Daniel D van Wijnen Andre J AJ Lerman Lilach O LO

Gene 20140823 1

Mesenchymal stromal/stem cells (MSCs) are clinically useful for cell-based therapy, but concerns regarding their ability to replicate limit their human application. MSCs release extracellular vesicles (EVs) that mediate at least in part the paracrine effects of the parental cells. To understand the molecular basis of their biological properties, we characterized the RNA cargo of EVs from porcine adipose-tissue derived MSCs. Comprehensive characterization of mRNA and miRNA gene expression using h ...[more]

PMID: 25158130

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Project description:Obesity is a chronic disease that interferes with normal repair processes, including adipose mesenchymal stem/stromal cells (ASCs) function. ASCs produce extracellular vesicles (EVs) that activate a repair program in recipient cells partly via their micro-RNA (miRNA) cargo. We hypothesized that obesity alters the miRNA expression profile of human ASC-derived EVs, limiting their capacity to repair injured cells. Human ASCs were harvested from obese and age- and gender-matched non-obese (lean) subjects during bariatric or cosmetic surgeries, respectively (n = 5 each), and their EVs isolated. Following high-throughput sequencing analysis, differentially expressed miRNAs were identified and their gene targets classified based on cellular component, molecular function, and biological process. The capacity of human lean- and obese-EVs to modulate inflammation, apoptosis, as well as mitogen-activated protein kinase (MAPK) and Wnt signaling in injured human proximal tubular epithelial (HK2) cells was evaluated in vitro. The number of EVs released from lean- and obese-ASCs was similar, but obese-EVs were smaller compared to lean-EVs. Differential expression analysis revealed 8 miRNAs upregulated (fold change > 1.4, p < 0.05) and 75 downregulated (fold change < 0.7, p < 0.05) in obese-EVs vs. lean-EVs. miRNAs upregulated in obese-EVs participate in regulation of NFk-B and MAPK signaling, cytoskeleton organization, and apoptosis, whereas those downregulated in obese-EVs are implicated in cell cycle, angiogenesis, and Wnt and MAPK signaling. Treatment of injured HK2 cells with obese-EVs failed to decrease inflammation, and they decreased apoptosis and MAPK signaling significantly less effectively than their lean counterparts. Obesity alters the size and miRNA cargo of human ASC-derived EVs, as well as their ability to modulate important injury pathways in recipient cells. These observations may guide development of novel strategies to improve healing and repair in obese individuals.

Project description:Background: Extracellular vesicles (EVs) isolated from mesenchymal stem/stromal cells (MSCs) contribute to recovery of damaged tissue in animals models of human disease. We have previously shown that EVs isolated from porcine MSCs transport mRNA and miRNA capable of modulating several cellular pathways in recipient cells, yet their proteome remained to be profiled. Using a quantitative proteomic strategy, we sought to study the protein cargo of porcine MSC-derived EVs to identify candidate molecules for mediating their therapeutic effect. Methods: Autologous MSCs were collected from abdominal fat of 3 female domestic pigs, and MSC-derived EVs were subsequently isolated, cultured, and characterized by the expression of typical MSC and EV markers. LC-MS/MS proteomic analysis was performed and proteins classified using the Panther Classification System. Functional pathway analysis was performed with DAVID 6.7. Three candidate proteins were selected for validation and their expression in EVs and MSCs confirmed by Western blot. Results: Proteomics analysis identified 5,469 protein groups in MSCs and 4,937 in EVs. Average protein intensity was higher in MSCs compared to EVs (p<0.0001). Differential expression analysis revealed 128 proteins upregulated in EVs vs. MSCs (log2 fold change>10, p<0.05), whereas 563 proteins were excluded from EVs (log2 fold change<-10, p<0.05). Biological functional analysis of proteins enriched in EVs indicated a broad distribution, with the most frequently represented categories being proteins involved in angiogenesis, blood coagulation, apoptosis, extracellular matrix remodeling, and regulation of inflammatory responses. Proteins excluded from EVs were mostly nuclear proteins and proteins involved in nucleotide binding and RNA splicing. Conclusions: The present study provides novel proteomic characterization of the biological signatures of porcine adipose MSC-derived EVs. The selective cargo that EVs shuttle may define the spectrum of their roles in mediating MSC intercellular communication.

Dataset Information

MicroRNA and mRNA cargo of extracellular vesicles from porcine adipose tissue-derived mesenchymal stem cells.

Publications

MicroRNA and mRNA cargo of extracellular vesicles from porcine adipose tissue-derived mesenchymal stem cells.

Similar Datasets

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