Objective. To investigate the in vivo effect of recombinant human interleukin-1 receptor antagonist (rHuIL-1Ra) on the development of lesions and the expression of metalloproteases in the canine experimental osteoarthritis (OA) model.Methods. The right anterior cruciate ligament was sectioned percutaneously in 3 groups of dogs. The control group (n = 5) received an intraarticular injection of sterile physiologic saline (1 ml) twice weekly for 4 weeks starting on the day of surgery. The remaining 2 groups received intraarticular injections of either 2 mg (n = 6) or 4 mg (n = 5) rHuIL-1Ra in 1 ml of physiologic saline according to the same schedule as the first group. All dogs were killed 4 weeks after surgery. The macroscopic appearance of femoral condyle osteophytes and the size and severity of cartilage lesions on femoral condyles and tibial plateaus were evaluated, as were the histologic features of cartilage and synovial membrane. Levels of collagenase-1 and stromelysin-1 messenger RNA expression in cartilage and synovium were determined by Northern blotting.Results. Recombinant human IGlRa exerted a dose-dependent protective effect on the development of osteophytes and cartilage lesions in vivo. Treatment with rHuIL-1Ra reduced the incidence (saline-treated Conclusion. This study demonstrated that intraarticular injections of rHuIL-1Ra can protect against the development of experimentally induced OA lesions. This effect could result, at least in part, from a reduction of collagenase-1 expression. However, other catabolic processes involved in the degradation of OA cartilage may also be affected.Osteoarthritis (OA) is the most common rheumatic disease and is characterized by progressive depletion of articular cartilage matrix macromolecules. Accumulating evidence suggests that an important component of the matrix loss process is related to activity of proteolytic enzymes which degrade the principal matrix

There is increasing evidence suggesting that chondrocyte death may contribute to the progression of osteoarthritis (OA). This study focused on the characterization of signaling cascade during NO-induced cell death in human OA chondrocytes. The NO generator, sodium nitroprusside (SNP), promoted chondrocyte death in association with DNA fragmentation, caspase-3 activation, and down-regulation of Bcl-2. Both caspase-3 inhibitor Z-Asp(OCH3)-Glu(OCH3)-Val-Asp(OCH3)-CH2F and caspase-9 inhibitor Z-Leu-Glu(OCH3)-His-Asp(OCH3)-CH2F prevented the chondrocyte death. Blocking the mitogen-activated protein kinase pathway by the mitogen-activated protein kinase kinase 1/2 inhibitor PD98059 or p38 kinase inhibitor SB202190 also inhibited the SNP-mediated cell death, suggesting possible requirements of both extracellular signal-related protein kinase 1/2 and p38 kinase for the NO-induced cell death. Furthermore, the selective inhibition of cyclooxygenase (COX)-2 by NS-398 or the inhibition of COX-1/COX-2 by indomethacin blocked the SNP-induced cell death. The chondrocyte death induced by SNP was associated with an overexpression of COX-2 protein (as determined by Western blotting) and an increase in PGE2 release. PD98059 and SB202190, but neither Z-DEVD FMK nor Z-LEHD FMK completely inhibited the SNP-mediated PGE2 production. Analysis of interactions between PGE2 and the cell death showed that PGE2 enhanced the SNP-mediated cell death, whereas PGE2 alone did not induce the chondrocyte death. These data indicate that NO-induced chondrocyte death signaling includes PGE2 production via COX-2 induction and suggest that both extracellular signal-related protein kinase 1/2 and p38 kinase pathways are upstream signaling of the PGE2 production. The results also demonstrate that exogenous PGE2 may sensitize human OA chondrocytes to the cell death induced by NO.

Objective To determine the effects of peroxisome proliferator–activated receptor γ (PPARγ) agonists on interleukin‐1 (IL‐1) induction of nitric oxide (NO) and matrix metalloproteinase 13 (MMP‐13) in human chondrocytes. Methods PPARγ expression and synthesis in human chondrocytes were determined by reverse transcriptase–polymerase chain reaction (RT‐PCR) and immunohistochemistry, respectively. Chondrocytes were cultured with IL‐1β, tumor necrosis factor α (TNFα), and IL‐17 in the presence or absence of PPARγ agonists, and NO and MMP‐13 synthesis and expression levels were measured. Transient transfection experiments were performed with the 7‐kb inducible NO synthase (iNOS) and 1.6‐kb MMP‐13 human promoters, as well as with the PPARγ expression vector and the activator protein 1 (AP‐1) and nuclear factor κB (NF‐κB) reporter constructs. Results RT‐PCR and immunohistochemical analysis revealed that human chondrocytes expressed and produced PPARγ. Treatment of chondrocytes with PPARγ ligands BRL 49653 and 15‐deoxy‐Δ12,14‐prostaglandin J2 (15d‐PGJ2), but not with PPARα ligand Wy 14643, decreased IL‐1β–induced NO and MMP‐13 production in a dose‐dependent manner. In addition, both iNOS and MMP‐13 messenger RNA were inhibited in the presence of 15d‐PGJ2. The inhibitory effect of PPARγ activation was not restricted to IL‐1β, since TNFα‐ and IL‐17–induced NO and MMP‐13 production were also inhibited by 15d‐PGJ2. In transient transfection experiments, we showed that a constitutively active form of mitogen‐activated protein kinase kinase kinase 1 (ΔMEKK‐1) induced the MMP‐13 and iNOS human promoter activity. This process was reduced by 15d‐PGJ2 and further inhibited by cotransfection with a PPARγ expression vector. Similarly, in a PPARγ‐dependent manner, 15d‐PGJ2 inhibited ΔMEKK‐1–induced AP‐1– and NF‐κB–luciferase reporter plasmid activation. Conclusion The findings of this study demonstrate that PPARγ agonists inhibit IL‐1β induction of both NO and MMP‐13 in human chondrocytes. The inhibition occurs at least at the transcriptional level through a PPARγ‐dependent pathway, probably by interfering with the activation of AP‐1 and NF‐κB.

Objective. To examine the effect of intraarticular injections of methylprednisolone acetate (MA) on osteoarthritic lesions and chondrocyte stromelysin synthesis in experimental osteoarthritis (OA).Methods. In 15 mongrel dogs, the anterior cruciate ligament of the right knee was sectioned by a stab wound. Eight dogs received intraarticular injections of MA (20 mg) at the time of surgery and 4 weeks later; 7 had no treatment. The dogs were killed 8 weeks after surgery. Five normal dogs were used as controls. Macroscopic evaluation of the lesions, including measurements of osteophytes and areas of surface lesions on the condyles and plateaus, was conducted, along with histologic evaluation of the severity of lesions. Immunohistochemical analysis was carried out using a rabbit polyclonal antibody against stromelysin, followed by evaluation of matrix and chondrocyte staining using morphometric analysis.Results. Treatment with MA significantly reduced the incidence (P < 0.0004) and size (P < 0.0001) of osteophytes. The histologic grading of cartilage lesions was also significantly reduced both on condyles (P < 0.01) and on plateaus (P < 0.002). Immunohisto- Address reprint requests to Jean-Pieme Pelletier, MD, Unitt des maladies rhumatismales, HGpital Notre-Dame, 1560 est, rue Sherbrooke, Montrtal, Qutbec, Canada H2L 4K8.Submitted for publication October 6, 1992; accepted in revised form August 10, 1993. chemical studies revealed, for OA cartilage, a marked increase (P < 0.002) in the percentage of chondrocytes positive for stromelysin and in the intensity of staining throughout all the layers of the cartilage, as well as specific matrix staining (P < 0.005). Treatment with MA reduced staining at both the chondrocyte (P < 0.002) and the matrix (P < 0.01) levels toward normal.