Abstract
In this exploratory study, we used discovery proteomics to follow the release of proteins from bovine knee articular cartilage in response to mechanical injury and cytokine treatment. We also studied the effect of the glucocorticoid Dexamethasone (Dex) on these responses. Bovine cartilage explants were treated with either cytokines alone (10 ng/ml TNFα, 20 ng/ml IL-6, 100 ng/ml sIL-6R), a single compressive mechanical injury, cytokines and injury, or no treatment, and cultured in serum-free DMEM supplemented with 1% ITS for 22 days. All samples were incubated with or without addition of 100 nM Dex. Mass spectrometry and western blot analyses were performed on medium samples for the identification and quantification of released proteins. We identified 500 unique proteins present in all three biological replicates. Many proteins involved in the catabolic response of cartilage degradation had increased release after inflammatory stress. Dex rescued many of these catabolic effects. The release of some proteins involved in anabolic and chondroprotective processes was inconsistent, indicating differential effects on processes that may protect cartilage from injury. Dex restored only a small fraction of these to the control state, while others had their effects exacerbated by Dex exposure. We identified proteins that were released upon cytokine treatment which could be potential biomarkers of the inflammatory contribution to cartilage degradation. We also demonstrated the imperfect rescue of Dex on the effects of cartilage degradation, with many catabolic factors being reduced, while other anabolic or chondroprotective processes were not.
Highlights
While glucocorticoids have been used for over 50 years to treat osteoarthritis (OA) pain, the prescription of glucocorticoids remains controversial because of potentially harmful side effects to multiple joint tissues, especially cartilage
principle component analysis (PCA) clustering on log2-transformed abundance data (Fig. 2B, all eight treatment groups in Supplemental Fig. S2) revealed cytokine treatment as the major determinant of medium composition: one cluster is seen for control and injury (N, I) and another cluster for samples treated by cytokines with and without injury (C, injury þ cytokines (IC))
C–C motif chemokine 5, ephrin-A1, inhibin, and transforming growth factor beta-2 (TGFβ-2)) and extracellular matrix (ECM). These results suggest that Group III proteins are those actively released from the cartilage after the induction of post-traumatic OA (PTOA): the degradation products of cleaved ECM components and the proteases, protease inhibitors, and other signaling factors associated with catabolic responses
Summary
While glucocorticoids have been used for over 50 years to treat osteoarthritis (OA) pain, the prescription of glucocorticoids remains controversial because of potentially harmful side effects to multiple joint tissues, especially cartilage. Dexamethasone (Dex) has been demonstrated to rescue the loss of aggrecan and collagen constituents as well as chondrocyte viability in human and bovine cartilage explant models of inflammatory tissue injury and post-traumatic OA (PTOA) [1,2], suggesting the possibility of Dex as a disease-modifying drug. The anti-catabolic versus pro-catabolic effects of Dex on the cartilage extracellular matrisome, as well as the fate of intracellular proteins in the presence of tissue injury, remain unexplored. The goals of the present study are to utilize a discovery proteomics approach to quantify the loss of extracellular and intracellular proteins from cartilage explants subjected to inflammatory cytokine challenge and impact mechanical injury, and to determine whether Dex protects against or exacerbates the response.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
