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Time-Course Mass Spectrometry Data of Adipose Mesenchymal Stem Cells Acquiring Chondrogenic Phenotype

ABSTRACT: Project description: Adipose mesenchymal stem cells (ADSCs) are fast gaining popularity in regenerative medicine for their pluripotency and relative use of collection. Currently, we are addressing the use of ADSCs in otolaryngology, for the treatment of vocal fold (VF) paralysis. VF can result from accidentally sectioning the recurrent laryngeal nerve (RLN) during thyroidectomy. This prevents closure of the upper larynx and promotes food aspiration into the lungs. Volume augmenting materials are sometimes injected in order to bring the paralyzed VF in close proximity to the counter lateral VF. The procedure is called injection medialization laryngoplasty (IML); we have recently developed a rabbit IML model for testing biological reagents in order to improve outcomes. As part of these studies ADSCs were differentiated into chondrocytes and their fate was studied in vitro and in situ. We used mass spectrometry proteomics because microarrays based gene expression platforms do not report actual protein levels. The data show protein levels in controls and chondrogenically differentiating rabbit ADSCs at 0, 14 and 28 days, analyzed using nano-flow liquid chromatography electrospray ionization tandem mass spectrometry (nLC-MS/MS).

INSTRUMENT(S): Thermo Scientific Q-Exactive Mass Spectrometer (Thermo Fisher Scientific, Bremen, Germany) coupled to a Thermo Ultimate 3000 RSLC nano HPLC system

ORGANISM(S): Oryctolagus Cuniculus (ncbitaxon:9986)

SUBMITTER: Serban San-Marina

PROVIDER: MSV000082505 | MassIVE | Tue Jun 26 13:06:00 BST 2018

SECONDARY ACCESSION(S): PXD010236

REPOSITORIES: MassIVE

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Project description:Osteoarthritis (OA) is an aging-associated disease of diarthrodial joints that involves changes in the bone, cartilage and soft tissue. Chondrocytes are the only cell type in cartilage and play a pivotal role in tissue homeostasis and cartilage remodeling in OA. Age- or OA-dependent metabolic changes affecting extracelluar matrix composition as well as variations in the response to cytokines and growth factors have been investigated in human chondrocytes. However, the molecular mechanisms causing age-dependent variation of the chondrocyte phenotype are poorly characterized. The proinflammatory mediator nitric oxide (NO) is produced when chondrocytes are exposed to cytokines such as IL-1. NO affects the energy metabolism of cells through interfering with mitochondrial respiration and glycolysis and has been implicated in extracellular matrix synthesis and degradation and chondrocyte death. NO may contribute to the aging process by compromising the energy balance ! in chondrocytes through the generation of an oxidizing environment and this may in turn affect matrix gene expression. In human chondrocytes NO effects on the expression of extacelluar matrix genes or genes related to matrix structure have not been addressed comprehensively. In chondrocytes NO has profound effects on extracellular matrix: Interleukin-1 induced the production of nitric oxide in human articular chondrocytes in alginate culture and simultaneously inhibited the biosynthesis of proteoglycans. NG-monomethyl-L-arginine (L-NMMA) inhibited nitric oxide formation and the suppression of proteoglycan synthesis. The nitric oxide donor, S-nitrosyl-acetyl-D,L- penicillamine (SNAP) also inhibited proteoglycan biosynthesis suggesting that interleukin-1 suppresses matrix synthesis through mechanisms involving the production of NO. Slices of rabbit articular cartilage synthesized large quantities of nitric oxide (NO) following exposure to IL-1 and strongly suppressed the incorporation of 35SO4 into the glycosaminoglycans (GAGs) of cartilage proteoglycans. Simultaneous treatment of cartilage fragments with IL-1 and L-NMMA inhibited NO synthesis in response to IL-1 and restored proteoglycan synthesis. SNAP reversibly mimicked the effect! of IL-1 on proteoglycan synthesis. These data suggest that endogenously synthesized NO is the mediator which reduces cartilage proteoglycan synthesis in response to cytokines such as IL-1. Aggrecan and type II collagen synthesis are also affected by NO. Our hypothesis entails that in human chondrocytes endogenous NO modulates the expression of matrix genes as well as enzymes involved in extracellular matrix remodelling. Experimental approach: In this experiment cultured chondrocytes from 3 donors was left unstimulated (control) or stimulated with IL-1, L-NMMA or IL-1 + L-NMMA for a period of 8 days in order to achieve sufficiently high levels of intracellular NO and stable expression of a subset of genes. Total RNA was isolated using the Trizol procedure. Gene expression will be probed using the Glycov2 chip. In this study cultured chondrocytes from three donors will either be left unstimulated (control) or stimulated with IL-1, L-NMMA or IL-1 L-NMMA for a period of 8 days in order to achieve sufficiently high levels of intracellular NO and stable expression of a subset of genes. 12 Samples were hybridized and analyzed using the GLYCOv2 array, three from each class.

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Time-Course Mass Spectrometry Data of Adipose Mesenchymal Stem Cells Acquiring Chondrogenic Phenotype

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