P183 Expression Patterns of Notch Receptors, Their Ligands and HES5 in Human Articular Cartilage (original) (raw)

2006, Osteoarthritis and Cartilage

via four G-protein-coupled receptors, EP1-EP4. We investigated role of PGE2 on mitochondrial function and matrix degradation in chondrocytes. Methods: Cartilage explants were prepared from OA patients undergoing joint replacement surgery. In selected experiments chondrocytes were isolated using collagenase digestion and cultured in alginate beads. PGE2 receptor (EP1-4) expression was analyzed by FACS. Mitochondrial function was assessed by the fluorescent probe JC-1 and ATP generation. Results: Immunostaining for enzymes COX-1, COX-2, cPGES and mPGES confirmed the presence of PG biosynthetic pathway(s) in OA cartilage. All four PGE2 EP receptors were expressed in both normal and OA cartilage and confirmed in OA chondrocytes by FACS. However, the EP4 receptor was upregulated (2-3-fold) in OA compared to normal. Addition of PGE2 (0.1-10uM) exerted the following deleterious effects on OA chondrocytes: 1) decreased mitochondrial membrane activity (fluorescent probe JC-1) and ATP generation 2) inhibition of proteoglycan synthesis (35 S incorporation) by chondrocyte cultures to levels of 75% of control (p<0.01), 3) increased type II collagen degradation as assessed by C12C ELISA. 4) inhibited expression of aggrecan and type II collagen mRNA and 5) induced production of pro-MMP-13, IL-6 and IL-8, but inhibited MMP-1 secretion. The potency of PGE2 effects were comparable to that of IL-1b, and the inhibitory effects of a combination of IL-1 and PGE2 (10uM) were additive.To examine the effects of endogenous PGE2 produced in response to IL-1, we performed experiments in the presence or absence of the EP4 antagonist, A23858 or COX-2 selective inhibitor celecoxib. The addition of IL-1 reduced chondrocyte proteoglycan synthesis to 45% of control values; pretreatment with 10uM A23858 or 2uM celecoxib, reversed this inhibition to 67% and 65% respectively of control values (p<0.01). Both A23858 and celecoxib reversed other catabolic effects of IL-1 (1-4 above), including mitochondrial activity, ATP generation and inhibition of PGE2 dependent production of MMP-13. Conclusions: PGE2, the predominant eicosanoid produced by OA chondrocytes, exerts catabolic effects within cartilage. These deleterious effects are mediated, in part via the EP4 receptor, which should be considered as a potential target for disease modifying agents in OA.