Inhibition of sphingosine 1-phosphate protects mice against chondrocyte catabolism and osteoarthritis

Summary Objective Cartilage loss observed in osteoarthritis (OA) is prevented when osteoclasts in the subchondral bone are inhibited in mice. Here, we investigated the role of the osteoclast secretome and of the lipid mediator sphingosine 1-phosphate (S1P) in chondrocyte metabolism and OA. Materials and methods We used SphK1LysMCre and wild type mice to assess the effect of murine osteoclast secretome in chondrocyte metabolism. Gene and protein expressions of matrix metalloproteinase (Mmp) were quantified in chondrocytes and explants by RT-qPCR and Western blots. SphK1LysMCre mice or wild type mice treated with S1P2 receptor inhibitor JTE013 or anti-S1P neutralizing antibody sphingomab are analyzed by OA score and immunohistochemistry. Results The osteoclast secretome increased the expression of Mmp3 and Mmp13 in murine chondrocytes and cartilage explants and activated the JNK signaling pathway, which led to matrix degradation. JTE013 reversed the osteoclast-mediated chondrocyte catabolism and protected mice against OA, suggesting that osteoclastic S1P contributes to cartilage damage in OA via S1P/S1P2 signaling. The activity of sphingosine kinase 1 (SphK1) increased with osteoclast differentiation, and its expression was enhanced in subchondral bone of mice with OA. The expression of Mmp3 and Mmp13 in chondrocytes was low upon stimulation with the secretome of Sphk1-lacking osteoclasts. Cartilage damage was significantly reduced in SphK1LysMCre mice, but not the synovial inflammation. Finally, intra-articular administration of sphingomab inhibited the cartilage damage and synovial inflammation. Conclusions Lack of S1P in myeloid cells and local S1P neutralization alleviates from osteoarthritis in mice. These data identify S1P as a therapeutic target in OA.The authors thank Alexandre Garcia for measurements of S1P. The work was supported by the Sybil SP7 European project and the “Fondation de l’Avenir”. JT and SV received grants from the Deutsche Forschungsgemeinschaft within the collaborative research center SFB1149.Peer reviewe

Upregulation of MMP-13 and TIMP-1 expression in response to mechanical strain in MC3T3-E1 osteoblastic cells

<p>Abstract</p> <p>Background</p> <p>Mechanical strain plays a significant role in the regulation of bone matrix turnover, which is mediated in part by matrix metalloproteinase (MMP)-13 and tissue inhibitors of matrix metalloproteinase (TIMP)-1. However, little is known about the correlation between mechanical strain and osteoblastic cell activities, including extracellular matrix (ECM) metabolism. Herein, we determined the effect of different magnitudes of cyclic tensile strain (0%, 6%, 12%, and 18%) on MMP-13 and TIMP-1 mRNA and protein expression in MC3T3-E1 osteoblasts. Furthermore, we employed specific inhibitors to examine the role of distinct signal transduction pathways known to mediate cellular responses to mechanical strain.</p> <p>Results</p> <p>We identified a magnitude-dependent increase in MMP-13 and TIMP-1 mRNA and protein levels in response to mechanical strains corresponding to 6%, 12%, and 18% elongation. The strain-induced increases in MMP-13 and TIMP-1 mRNA expression were inhibited by PD098059 and cycloheximide, respectively.</p> <p>Conclusions</p> <p>Our results suggest a mechanism for the regulation of bone matrix metabolism mediated by the differential expression of MMP-13 and TIMP-1 in response to increasing magnitudes of mechanical strain.</p

Glucocorticoids in T cell apoptosis and function

Glucocorticoids (GCs) are a class of steroid hormones which regulate a variety of essential biological functions. The profound anti-inflammatory and immunosuppressive activity of synthetic GCs, combined with their power to induce lymphocyte apoptosis place them among the most commonly prescribed drugs worldwide. Endogenous GCs also exert a wide range of immunomodulatory activities, including the control of T cell homeostasis. Most, if not all of these effects are mediated through the glucocorticoid receptor, a member of the nuclear receptor superfamily. However, the signaling pathways and their cell type specificity remain poorly defined. In this review, we summarize our present knowledge on GC action, the mechanisms employed to induce apoptosis and the currently discussed models of how they may participate in thymocyte development. Although our knowledge in this field has substantially increased during recent years, we are still far from a comprehensive picture of the role that GCs play in T lymphocytes

Non-contact universal sample presentation for room temperature macromolecular crystallography using acoustic levitation

Macromolecular Crystallography is a powerful and valuable technique to assess protein structures. Samples are commonly cryogenically cooled to minimise radiation damage effects from the X-ray beam, but low temperatures hinder normal protein functions and this procedure can introduce structural artefacts. Previous experiments utilising acoustic levitation for beamline science have focused on Langevin horns which deliver significant power to the confined droplet and are complex to set up accurately. In this work, the low power, portable TinyLev acoustic levitation system is used in combination with an approach to dispense and contain droplets, free of physical sample support to aid protein crystallography experiments. This method facilitates efficient X-ray data acquisition in ambient conditions compatible with dynamic studies. Levitated samples remain free of interference from fixed sample mounts, receive negligible heating, do not suffer significant evaporation and since the system occupies a small volume, can be readily installed at other light sources