Activation of proteinase-activated receptor 2 in human osteoarthritic cartilage upregulates catabolic and proinflammatory pathways capable of inducing cartilage degradation: a basic science study
1 Osteoarthritis Research Unit, University of Montreal Hospital Centre, Notre-Dame Hospital, 1560 Sherbrooke Street East, Montreal, Quebec, H2L 4M1, Canada
2 Pavillon des Charmilles, 1487, boul. des Laurentides, Vimont, Quebec, H7M 2Y3, Canada
Arthritis Research & Therapy 2007, 9:R121 doi:10.1186/ar2329Published: 21 November 2007
Proteinase-activated receptors (PARs) belong to a family of G protein-coupled receptors. PARs are activated by a serine-dependent cleavage generating a tethered activating ligand. PAR-2 was shown to be involved in inflammatory pathways. We investigated the in situ levels and modulation of PAR-2 in human normal and osteoarthritis (OA) cartilage/chondrocytes. Furthermore, we evaluated the role of PAR-2 on the synthesis of the major catabolic factors in OA cartilage, including metalloproteinase (MMP)-1 and MMP-13 and the inflammatory mediator cyclooxygenase 2 (COX-2), as well as the PAR-2-activated signalling pathways in OA chondrocytes. PAR-2 expression was determined using real-time reverse transcription-polymerase chain reaction and protein levels by immunohistochemistry in normal and OA cartilage. Protein modulation was investigated in OA cartilage explants treated with a specific PAR-2-activating peptide (PAR-2-AP), SLIGKV-NH2 (1 to 400 μM), interleukin 1 beta (IL-1β) (100 pg/mL), tumor necrosis factor-alpha (TNF-α) (5 ng/mL), transforming growth factor-beta-1 (TGF-β1) (10 ng/mL), or the signalling pathway inhibitors of p38 (SB202190), MEK1/2 (mitogen-activated protein kinase kinase) (PD98059), and nuclear factor-kappa B (NF-κB) (SN50), and PAR-2 levels were determined by immunohistochemistry. Signalling pathways were analyzed on OA chondrocytes by Western blot using specific phospho-antibodies against extracellular signal-regulated kinase 1/2 (Erk1/2), p38, JNK (c-jun N-terminal kinase), and NF-κB in the presence or absence of the PAR-2-AP and/or IL-1β. PAR-2-induced MMP and COX-2 levels in cartilage were determined by immunohistochemistry. PAR-2 is produced by human chondrocytes and is significantly upregulated in OA compared with normal chondrocytes (p < 0.04 and p < 0.03, respectively). The receptor levels were significantly upregulated by IL-1β (p < 0.006) and TNF-α (p < 0.002) as well as by the PAR-2-AP at 10, 100, and 400 μM (p < 0.02) and were downregulated by the inhibition of p38. After 48 hours of incubation, PAR-2 activation significantly induced MMP-1 and COX-2 starting at 10 μM (both p < 0.005) and MMP-13 at 100 μM (p < 0.02) as well as the phosphorylation of Erk1/2 and p38 within 5 minutes of incubation (p < 0.03). Though not statistically significant, IL-1β produced an additional effect on the activation of Erk1/2 and p38. This study documents, for the first time, functional consequences of PAR-2 activation in human OA cartilage, identifies p38 as the major signalling pathway regulating its synthesis, and demonstrates that specific PAR-2 activation induces Erk1/2 and p38 in OA chondrocytes. These results suggest PAR-2 as a potential new therapeutic target for the treatment of OA.