The Immunosuppressant FTY720 (Fingolimod) enhances Glycosaminoglycan depletion in articular cartilage

Abstract

BackgroundFTY720 (Fingolimod) is a novel immunosuppressive drug investigated in clinical trials for organ transplantation and multiple sclerosis. It acts as a functional sphingosine-1-phosphate (S1P) receptor antagonist, thereby inhibiting the egress of lymphocytes from secondary lymphoid organs. As S1P is able to prevent IL-1beta induced cartilage degradation, we examined the direct impact of FTY720 on cytokine induced cartilage destruction.MethodsBovine chondrocytes were treated with the bioactive phosphorylated form of FTY720 (FTY720-P) in combination with IL-1beta or TNF-alpha. Expression of MMP-1,-3.-13, iNOS and ADAMTS-4,-5 and COX-2 was evaluated using quantitative real-time PCR and western blot. Glycosaminoglycan depletion from cartilage explants was determined using a 1,9-dimethylene blue assay and safranin O staining.ResultsFTY720-P significantly reduced IL-1beta and TNF-alpha induced expression of iNOS. In contrast FTY720-P increased MMP-3 and ADAMTS-5 mRNA expression. Furthermore depletion of glycosaminoglycan from cartilage explants by IL-1beta and TNF-alpha was significantly enhanced by FTY720-P in an MMP-3 dependent manner.ConclusionsOur results suggest that FTY720 may enhance cartilage degradation in pro-inflammatory environment.

Experimentally induced cartilage degeneration treated by pulsed electromagnetic field stimulation; an in vitro study on bovine cartilage

“…The concentration of IL-1β found in the synovial fluid of patients affected by OA is between 0.068 and 0.33 pg/ml [ 35 ]. However, most of the in vitro literature studies, that aimed to re-create an inflammatory microenvironment, added 10 ng/ml of IL-1β in the medium of cartilage explants [ 36 – 38 ], while there are other authors that employed 50 or 100 ng/ml [ 39 – 41 ]. The so high concentration of IL-1β used in the present study was chosen to employed an intermediate cytokine dosage and to evaluate if PEMF stimulation is able to restore cartilage ECM biological and morphological properties also with a huge concentration of IL-1β than the most employed one (10 ng/ml).…”

Prenatal Infection

“…First biochemical evidence from our group indeed suggests that abnormal proteolytic cleavage of Reelin through the serine (tissue Plasminogen Activator, tPA) and matrix metalloproteinase (A Disintegrin And Metalloproteinase with thrombo-spondin motif, ADAMTS4/5) alters the ability of Reelin to form dimers and facilitates its oligomerization [143]. In line with the inflammatory cytokine-mediated induction of tPA [144] and ADAMTS4/5 [145,146] as well as the strong association of tPA with neuronal injuries [147], chronic inflammatory conditions are expected to profoundly impair Reelin-dependent signaling through alterations in its proteolytic processing. In agreement with the prediction that accumulation of Reelin and its fragment in axonal terminals is restricted to projection zones of Reelin-expressing neurons, the earliest aging-associated neuritic varicosities are indeed seen along the olfactorylimbic pathways in which Reelin is normally secreted from axonal terminals [66].…”

Sphingosine 1‐Phosphate Counteracts the Effects of Interleukin‐1β in Human Chondrocytes

“…Four independent experiments with chondrocytes derived from 4 different patients were performed. RNA isolation and complementary DNA (cDNA) synthesis were performed as described in an earlier study (). For amplification, a ready‐to‐use Master Mix containing SYBR Green (Invitrogen) was used.…”

“…Supernatants were analyzed for nitrate concentrations as an indicator of NO release. Griess reaction was performed as described previously (). The same supernatants were also analyzed for ADAMTS activity using a commercially available enzyme‐linked immunosorbent assay (ELISA) (ProteaDetect Sensitive Aggrecanase Activity Assay; ProteaImmun).…”