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Novel Mouse Model for Tissue-specific Extracellular Vesicle Isolation and Characterization

ABSTRACT: Novel Mouse Model for Tissue-specific Extracellular Vesicle Isolation and Characterization

PROVIDER: PRJNA1078063 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR28009834_1.fastq.gz	Fastqsanger.gz
	SRR28009834_2.fastq.gz	Fastqsanger.gz
	SRR28009835_1.fastq.gz	Fastqsanger.gz
	SRR28009835_2.fastq.gz	Fastqsanger.gz
	SRR28009836_1.fastq.gz	Fastqsanger.gz

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Novel Mouse Model for Tissue-specific Extracellular Vesicle Isolation and Characterization

Project description:Novel Mouse Model for Tissue-specific Extracellular Vesicle Isolation and Characterization

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Novel Mouse Model for Tissue-specific Extracellular Vesicle Isolation and Characterization

Project description:Extracellular vesicles (EVs) are key mediators of intercellular communication, with important roles in numerous physiological and pathological processes, including profound effects on bone metabolism. These small membrane-bound vesicles are produced and released in the extracellular environment by virtually all cell types, including cells in the osteogenic lineage such as bone marrow-derived mesenchymal stem cells (MSCs), osteoblasts, osteoclasts and osteocytes. EVs serve as potent carriers of bioactive molecules, such as nucleic acids, proteins, lipids and metabolites, where they can influence recipient cells through fusion with target cell membranes to deliver these functional biomolecules. However, once released, the source cell of the EV is difficult to ascertain with any certainty. To overcome this obstacle, we developed a conditional (e.g. Cre-mediated) mouse model that expresses an EV tag, containing a fusion of CD81 and multiple C-terminal tags, termed the “Snorkel-tag”. By crossing with a Cre of interest, representing a specific cell-type or tissue, the specific EV subpopulations that are released can be isolated using antibody affinity columns. We crossed the CAGS-Snorkel mouse with Prx1- and Ocn-Cre, representing cell-types in the early vs late stages of osteoblast differentiation, isolated EVs from bone marrow plasma, and treated mouse bone marrow stromal cells (mBMSCs) with Prx1-EVs, Ocn-EVs or All-EVs (isolated using a Pan EV Isolation Kit [Miltenyi Biotec]) for 3 days and performed bulk RNA-sequencing. We found unique transcriptional and pathway signatures elicited by the different EV subpopulations in the mBMSCs, suggesting that EVs from diverse sources have distinct biological activities.

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Novel Mouse Model for Tissue-specific Extracellular Vesicle Isolation and Characterization

Project description:Extracellular vesicles (EVs) are key mediators of intercellular communication, with important roles in numerous physiological and pathological processes, including profound effects on bone metabolism. These small membrane-bound vesicles are produced and released in the extracellular environment by virtually all cell types, including cells in the osteogenic lineage such as bone marrow-derived mesenchymal stem cells (MSCs), osteoblasts, osteoclasts and osteocytes. EVs serve as potent carriers of bioactive molecules, such as nucleic acids, proteins, lipids and metabolites, where they can influence recipient cells through fusion with target cell membranes to deliver these functional biomolecules. However, once released, the source cell of the EV is difficult to ascertain with any certainty. To overcome this obstacle, we developed a conditional (e.g. Cre-mediated) mouse model that expresses an EV tag, containing a fusion of CD81 and multiple C-terminal tags, termed the “Snorkel-tag”. By crossing with a Cre of interest, representing a specific cell-type or tissue, the specific EV subpopulation that is released can be isolated using antibody affinity columns. We crossed the CAGS-Snorkel mouse with Prx1-Cre and Ocn-Cre, representing cell-types in the early vs late stages of osteoblast differentiation. Isolation of Prx1-EVs and Ocn-EVs was performed from the mouse bone marrow plasma. We performed miRNA-sequencing to determine the specific miRNA cargo in these different EV subpopulations and found miRNAs involved in bone metabolism and function to be expressed, some of which are enriched in the Ocn-EVs. In summary, the CAGS-Snorkel mouse model will be useful in the characterization of EVs from diverse cell- and tissue-types in the mouse.

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