Alterations in the chondrocyte surfaceome in response to pro-inflammatory cytokines.
Abstract: Chondrocytes are exposed to an inflammatory micro-environment in the extracellular matrix (ECM) of articular cartilage in joint diseases such as osteoarthritis (OA) and rheumatoid arthritis (RA). In OA, degenerative changes and low-grade inflammation within the joint transform the behaviour and metabolism of chondrocytes, disturb the balance between ECM synthesis and degradation, and alter the osmolality and ionic composition of the micro-environment. We hypothesize that chondrocytes adjust their physiology to the inflammatory microenvironment by modulating the expression of cell surface proteins, collectively referred to as the 'surfaceome'. Therefore, the aim of this study was to characterize the surfaceome of primary equine chondrocytes isolated from healthy joints following exposure to the pro-inflammatory cytokines interleukin-1-beta (IL-1β) and tumour necrosis factor-alpha (TNF-α). We employed combined methodology that we recently developed for investigating the surfaceome in stem cells. Membrane proteins were isolated using an aminooxy-biotinylation technique and analysed by mass spectrometry using high throughput shotgun proteomics. Selected proteins were validated by western blotting. Results: Amongst the 431 unique cell surface proteins identified, a high percentage of low-abundance proteins, such as ion channels, receptors and transporter molecules were detected. Data are available via ProteomeXchange with identifier PXD014773. A high number of proteins exhibited different expression patterns following chondrocyte stimulation with pro-inflammatory cytokines. Low density lipoprotein related protein 1 (LPR-1), thrombospondin-1 (TSP-1), voltage dependent anion channel (VDAC) 1-2 and annexin A1 were considered to be of special interest and were analysed further by western blotting. Conclusions: Our results provide, for the first time, a repository for proteomic data on differentially expressed low-abundance membrane proteins on the surface of chondrocytes in response to pro-inflammatory stimuli.
Publication Date: 2020-06-26 PubMed ID: 32586320PubMed Central: PMC7318434DOI: 10.1186/s12860-020-00288-9Google Scholar: Lookup
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Summary
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This research investigates how inflammatory conditions affect the expression of cell surface proteins, or ‘surfaceome’, in chondrocytes (cartilage cells). The researchers specifically study the effect of two pro-inflammatory chemicals (interleukin-1-beta and tumour necrosis factor-alpha) on the surfaceome of chondrocytes from healthy horse joints, using cutting-edge protein analysis techniques. Results showed that inflammation changed the expression patterns of many proteins on the cell surface.
Research Goal and Hypothesis
- The focus of this research is on chondrocytes, cells that exist within a joint’s cartilage. They hypothesized these cells adapt their physiology in response to inflammation by modulating the expression of cell surface proteins known as the ‘surfaceome’.
- Joint diseases like osteoarthritis (OA) and rheumatoid arthritis (RA) offer an inflammatory environment that influences the behavior of chondrocytes, affecting the balance between synthesis and degradation of ECM (extracellular matrix – a network of proteins and sugars that provides structural and biochemical support to cells).
Methods
- In order to investigate their hypothesis, they studied the surfaceome of primary equine chondrocytes, cells isolated from healthy horse joints, after exposure to pro-inflammatory cytokines interleukin-1-beta (IL-1β) and tumour necrosis factor-alpha (TNF-α).
- The researchers used an aminooxy-biotinylation technique to isolate membrane proteins, and mass spectrometry with high throughput shotgun proteomics for analysis.
- Their method also included validation of selected proteins using western blotting, a technique in molecular biology used to detect proteins in a sample.
Results and Conclusion
- From the 431 unique cell surface proteins identified, there were numerous low-abundance proteins, such as ion channels, receptors, and transporter molecules.
- The expression patterns of a significant number of proteins changed when chondrocytes were exposed to pro-inflammatory cytokines. Proteins such as Low density lipoprotein related protein 1 (LPR-1), thrombospondin-1 (TSP-1), voltage dependent anion channel (VDAC) 1-2, and annexin A1 were scrutinized further with western blotting.
- The results offer the first inventory of proteomic data about differentially expressed low-abundance membrane proteins on chondrocyte surfaces responding to pro-inflammatory stimuli, which may be helpful in understanding joint diseases better.
Cite This Article
APA
Jeremiasse B, Matta C, Fellows CR, Boocock DJ, Smith JR, Liddell S, Lafeber F, van Spil WE, Mobasheri A.
(2020).
Alterations in the chondrocyte surfaceome in response to pro-inflammatory cytokines.
BMC Mol Cell Biol, 21(1), 47.
https://doi.org/10.1186/s12860-020-00288-9 Publication
Researcher Affiliations
- Department of Rheumatology & Clinical Immunology, University Medical Centre Utrecht, Utrecht, The Netherlands.
- Department of Anatomy, Histology and Embryology, Faculty of Medicine, University of Debrecen, Debrecen, Hungary. matta.csaba@med.unideb.hu.
- Department of Veterinary Pre-Clinical Sciences, School of Veterinary Science and Medicine, University of Surrey, Guildford, UK.
- John van Geest Cancer Research Centre, Nottingham Trent University, Nottingham, NG11 8NS, UK.
- Bruker UK Limited, Coventry, UK.
- Exonate Ltd., Medicity, Thane Road, Nottingham, UK.
- Department of Rheumatology & Clinical Immunology, University Medical Centre Utrecht, Utrecht, The Netherlands.
- Department of Rheumatology & Clinical Immunology, University Medical Centre Utrecht, Utrecht, The Netherlands.
- Department of Rheumatology & Clinical Immunology, University Medical Centre Utrecht, Utrecht, The Netherlands. ali.mobasheri@oulu.fi.
- Research Unit of Medical Imaging, Physics and Technology, Faculty of Medicine, University of Oulu, Oulu, Finland. ali.mobasheri@oulu.fi.
- Department of Regenerative Medicine, State Research Institute Centre for Innovative Medicine, Vilnius, Lithuania. ali.mobasheri@oulu.fi.
- Centre for Sport, Exercise and Osteoarthritis Research Versus Arthritis, Queen's Medical Centre, Nottingham, UK. ali.mobasheri@oulu.fi.
- Department of Orthopedics, UMC Utrecht, Utrecht, The Netherlands. ali.mobasheri@oulu.fi.
MeSH Terms
- Animals
- Biomarkers
- Cartilage, Articular / cytology
- Cells, Cultured
- Chondrocytes / drug effects
- Chondrocytes / metabolism
- Cytokines / metabolism
- Cytokines / pharmacology
- Horses
- Inflammation / metabolism
- Interleukin-1beta / metabolism
- Interleukin-1beta / pharmacology
- Mass Spectrometry
- Membrane Proteins / drug effects
- Membrane Proteins / metabolism
- Osteoarthritis / diagnosis
- Osteoarthritis / pathology
- Primary Cell Culture
- Proteomics
- Tumor Necrosis Factor-alpha / metabolism
- Tumor Necrosis Factor-alpha / pharmacology
Grant Funding
- HEALTH.2012.2.4.5-2 / Seventh Framework Programme
- 20194 / Arthritis Research UK
- DOTSUT-215 / European Social Fund
- 625746 / FP7 People: Marie-Curie Actions
Conflict of Interest Statement
The authors declare that they have no competing interests. This paper was written by the authors within the scope of their academic and research positions. None of the authors have any relationships that could be construed as biased or inappropriate. The funding bodies were not involved in the study design, data collection, analysis and interpretation. The decision to submit the paper for publication was not influenced by any the funding bodies.
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