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Mesenchymal stem or stromal cells are currently under clinical investigation for multiple diseases. While their mechanism of action is still not fully elucidated, vesicles secreted by MSCs are believed to recapitulate their therapeutic potentials to some extent. Microvesicles (MVs), also called as microparticles or ectosome, are among secreted vesicles that could transfer cytoplasmic cargo, including RNA and proteins, from emitting (source) cells to recipient cells. Given the importance of MVs, we here attempted to establish a method to isolate and characterize MVs secreted from unmodified human bone marrow derived MSCs (referred to as native MSCs, and their microvesicles as Native-MVs) and IFNγ stimulated MSCs (referred to as IFNγ-MSCs, and their microvesicles as IFNγ-MVs). We first describe an ultracentrifugation technique to isolate MVs from the conditioned cell culture media of MSCs. Next, we describe characterization and quality control steps to analyze the protein and RNA content of MVs. Finally, we examined the potential of MVs to exert immunomodulatory effects through induction of regulatory T cells (Tregs). Secretory vesicles from MSCs are promising alternatives for cell therapy with applications in drug delivery, regenerative medicine, and immunotherapy.
Pubmed ID: 31669353
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A software package and server used to identify and characterize proteins from primary sequence databases using mass spectrometry data. Mascot integrates peptide mass fingerprinting, sequence querying, and MS/MS ion searching in order to search for proteins in databases like SwissProt, NCBInr, EMBL EST divisions, contaminants, and cRAP. If a license is purchased, users may: search data sets that exceed the 1200 spectrum limit of the free version; set up automated, high throughput work; add and edit proteins and quantification methods; and search a preferred collection of sequence databases. The software package works with instruments from AB Sciex, Agilent, Bruker, Jeol, Shimadzu, Thermo Scientific, and Waters.
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