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Quantitative proteomic characterization of lung-MSC and bone marrow-MSC using DIA-mass spectrometry.

ABSTRACT: Mesenchymal stromal cells (MSC) are ideal candidates for cell therapies, due to their immune-regulatory and regenerative properties. We have previously reported that lung-derived MSC are tissue-resident cells with lung-specific properties compared to bone marrow-derived MSC. Assessing relevant molecular differences between lung-MSC and bone marrow-MSC is important, given that such differences may impact their behavior and potential therapeutic use. Here, we present an in-depth mass spectrometry (MS) based strategy to investigate the proteomes of lung-MSC and bone marrow-MSC. The MS-strategy relies on label free quantitative data-independent acquisition (DIA) analysis and targeted data analysis using a MSC specific spectral library. We identified several significantly differentially expressed proteins between lung-MSC and bone marrow-MSC within the cell layer (352 proteins) and in the conditioned medium (49 proteins). Bioinformatics analysis revealed differences in regulation of cell proliferation, which was functionally confirmed by decreasing proliferation rate through Cytochrome P450 stimulation. Our study reveals important differences within proteome and matrisome profiles between lung- and bone marrow-derived MSC that may influence their behavior and affect the clinical outcome when used for cell-therapy.

SUBMITTER: Rolandsson Enes S

PROVIDER: S-EPMC5570998 | biostudies-literature | 2017 Aug

REPOSITORIES: biostudies-literature

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Quantitative proteomic characterization of lung-MSC and bone marrow-MSC using DIA-mass spectrometry.

Rolandsson Enes Sara S Åhrman Emma E Palani Anitha A Hallgren Oskar O Bjermer Leif L Malmström Anders A Scheding Stefan S Malmström Johan J Westergren-Thorsson Gunilla G

Scientific reports 20170824 1

Mesenchymal stromal cells (MSC) are ideal candidates for cell therapies, due to their immune-regulatory and regenerative properties. We have previously reported that lung-derived MSC are tissue-resident cells with lung-specific properties compared to bone marrow-derived MSC. Assessing relevant molecular differences between lung-MSC and bone marrow-MSC is important, given that such differences may impact their behavior and potential therapeutic use. Here, we present an in-depth mass spectrometry ...[more]

PMID: 28839187

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Project description:BACKGROUND:Astrocytes are the most abundant cells in the central nervous system and are responsible for a wide range of functions critical to normal neuronal development, synapse formation, blood-brain barrier regulation, and brain homeostasis. They are also actively involved in initiating and perpetuating neuroinflammatory responses. However, information about their proteomic phenotypes under inflammation is currently limited. METHOD:Data-independent acquisition mass spectrometry was applied to extensively characterize the profile of more than 4000 proteins in immortalized human fetal astrocytes under distinct inflammatory conditions induced by TNF, IL-1?, and LPS, while multiplex immunoassay-based screening was used to quantify a wide range of cytokines released under these inflammatory conditions. Then, immunocytochemistry and western blotting were used to verify the activation of canonical and non-canonical NF-?B upon exposure to the different stimuli. Finally, an in vitro model of the blood-brain barrier consisting of a co-culture of primary human brain microvascular endothelial cells and primary human astrocytes was used to verify the inflammatory response of astrocytes upon LPS exposure in a more complex in vitro system. RESULTS:We reported on a set of 186 proteins whose levels were significantly modulated by TNF, IL-1?, and LPS. These three stimuli induced proteome perturbations, which led to an increased abundance of key inflammatory proteins involved in antigen presentation and non-canonical NF-?B pathways. TNF and IL-1?, but not LPS, also activated the canonical NF-?B pathway, which in turn led to an extensive inflammatory response and dysregulation of cytoskeletal and adhesion proteins. In addition, TNF and LPS, but not IL-1?, increased the abundance of several interferon-stimulated gene products. Finally, TNF and IL-1? similarly upregulated the secretion of several cytokines and chemokines, whereas LPS only induced a moderate increase in IL-8, IFN-?, and IL-1? secretion. Upregulation of proteins associated with type I IFN and non-canonical NF-?B signaling was also observed in primary astrocytes co-cultured with primary brain microvascular endothelial cells exposed to LPS. CONCLUSIONS:The present study provides comprehensive information about the proteomic phenotypes of human astrocytes upon exposure to inflammatory stimuli both in monoculture and in co-culture with human brain microvascular endothelial cells.

Dataset Information

Quantitative proteomic characterization of lung-MSC and bone marrow-MSC using DIA-mass spectrometry.

Publications

Quantitative proteomic characterization of lung-MSC and bone marrow-MSC using DIA-mass spectrometry.

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