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Cartilage Matrix Degradation (cartilage + matrix_degradation)
Selected AbstractsDiscoidin domain receptor 2 mediates the collagen II-dependent release of interleukin-6 in primary human chondrocytes,THE JOURNAL OF PATHOLOGY, Issue 2 2009Andreas R Klatt Abstract We deciphered constituent parts of a signal transduction cascade that is initiated by collagen II and results in the release of various pro-inflammatory cytokines, including interleukin-6 (IL-6), in primary human chondrocytes. This cascade represents a feed-forward mechanism whereby cartilage matrix degradation is exacerbated by the mutually inducing effect of released collagen II fragments and pro-inflammatory cytokines. We previously proposed discoidin domain receptor 2 as a central mediator in this event. Since this cascade plays a prominent role in the pathogenesis of osteoarthritis, our study further investigates the hypothesis that discoidin domain receptor 2 is a candidate receptor for collagen II, and that transcription factor NF,B, lipid kinase PI3K, and the MAP kinases are constituent parts of this very signal transduction cascade. To accomplish this, we selectively knocked down the molecules of interest in primary human chondrocytes, induced the specified cascade by incubating primary human chondrocytes with collagen II, and observed the outcome, specifically the changes in interleukin-6 release. Knockdown was performed by siRNA-mediated gene silencing in the case of discoidin domain receptor 2 (DDR2) or by using specific inhibitors for the remainder of the molecules. Results indicated that discoidin domain receptor 2 mediates the collagen II-dependent release of interleukin-6 in primary human chondrocytes and that MAP kinases p38, JNK and ERK, as well as transcription factor NF,B, are integral components of intracellular collagen II signalling. Given the detrimental role of these molecules in osteoarthritis, our findings provide new targets for more specific therapeutics, which may have fewer side effects than those currently applied. Copyright © 2009 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. [source] Matriptase is a novel initiator of cartilage matrix degradation in osteoarthritisARTHRITIS & RHEUMATISM, Issue 7 2010Jennifer M. Milner Objective Increasing evidence implicates serine proteinases in pathologic tissue turnover. The aim of this study was to assess the role of the transmembrane serine proteinase matriptase in cartilage destruction in osteoarthritis (OA). Methods Serine proteinase gene expression in femoral head cartilage obtained from either patients with hip OA or patients with fracture to the neck of the femur (NOF) was assessed using a low-density array. The effect of matriptase on collagen breakdown was determined in cartilage degradation models, while the effect on matrix metalloproteinase (MMP) expression was analyzed by real-time polymerase chain reaction. ProMMP processing was determined using sodium dodecyl sulfate,polyacrylamide gel electrophoresis/N-terminal sequencing, while its ability to activate proteinase-activated receptor 2 (PAR-2) was determined using a synovial perfusion assay in mice. Results Matriptase gene expression was significantly elevated in OA cartilage compared with NOF cartilage, and matriptase was immunolocalized to OA chondrocytes. We showed that matriptase activated proMMP-1 and processed proMMP-3 to its fully active form. Exogenous matriptase significantly enhanced cytokine-stimulated cartilage collagenolysis, while matriptase alone caused significant collagenolysis from OA cartilage, which was metalloproteinase-dependent. Matriptase also induced MMP-1, MMP-3, and MMP-13 gene expression. Synovial perfusion data confirmed that matriptase activates PAR-2, and we demonstrated that matriptase-dependent enhancement of collagenolysis from OA cartilage is blocked by PAR-2 inhibition. Conclusion Elevated matriptase expression in OA and the ability of matriptase to activate selective proMMPs as well as induce collagenase expression make this serine proteinase a key initiator and inducer of cartilage destruction in OA. We propose that the indirect effects of matriptase are mediated by PAR-2, and a more detailed understanding of these mechanisms may highlight important new therapeutic targets for OA treatment. [source] Involvement of protein kinase C, in interleukin-1, induction of ADAMTS-4 and type 2 nitric oxide synthase via NF-,B signaling in primary human osteoarthritic chondrocytesARTHRITIS & RHEUMATISM, Issue 12 2007Priya S. Chockalingam Objective Protein kinase C, (PKC,), an atypical PKC, has been found to be transcriptionally up-regulated in human osteoarthritic (OA) articular cartilage. This study was undertaken to examine the role of PKC, in interleukin-1, (IL-1,),induced NF-,B signaling in human OA chondrocytes, and ultimately to better understand its function in the regulation of downstream mediators of cartilage matrix degradation. Methods Pharmacologic inhibitors or genetic knockdown techniques were used to investigate the role of PKC,. Western blot analysis was used to evaluate phosphorylation of PKC, and NF-,B. Quantitative polymerase chain reaction (PCR) and activity assays were used to evaluate ADAMTS-4 expression and aggrecanase activity, respectively. Quantitative PCR, biochemical identification, and Western blot analysis were used to evaluate type 2 nitric oxide synthase (NOS2) and NO production. Results Phosphorylation of PKC, and NF-,B was induced by IL-1, treatment in a time-dependent manner, and was specifically inhibited by inhibitors of atypical PKCs. Inhibition of PKC, suppressed IL-1,,induced up-regulation of ADAMTS-4 messenger RNA (mRNA) and aggrecanase activity. Inhibitors of atypical PKCs also inhibited IL-1,,induced NO production and NOS2 mRNA expression, demonstrating a novel link between PKC, and NO production. Furthermore, small interfering RNA, or short hairpin RNA,mediated knockdown of PKC, mRNA resulted in significant repression of both ADAMTS-4 and NOS2 mRNA expression. Conclusion Our results show that PKC, is involved in the regulation of IL-1,,induced NF-,B signaling in human OA chondrocytes, which in turn regulates downstream expression of ADAMTS-4 and NOS2. Therefore, inhibition of PKC, could potentially regulate the production of matrix-degrading enzymes as well as NO production and have a profound effect on disease progression in OA. [source] Neutrophil gelatinase,associated lipocalin is expressed in osteoarthritis and forms a complex with matrix metalloproteinase 9ARTHRITIS & RHEUMATISM, Issue 10 2007Kalpana Gupta Objective Expression of matrix metalloproteinase 9 (MMP-9) is up-regulated in osteoarthritis (OA) and usually presents as multiple bands when synovial fluid (SF) from OA patients is analyzed by zymography. Among these bands is an ,125,130,kd band for high molecular weight (HMW) gelatinase, which has not been characterized. This study was undertaken to characterize the HMW MMP activity in OA SF. Methods MMP activity in OA SF was determined by gelatin zymography. Recombinant MMPs were used to identify MMP activity on the zymogram. Western immunoblotting, immunoprecipitation, and immunodepletion analyses were performed using antibodies specific for human MMP-9 and human neutrophil gelatinase,associated lipocalin (NGAL). Human cartilage matrix degradation was determined by dimethylmethylene blue assay. Results Zymographic analysis showed that the HMW gelatinase in OA SF comigrated with a purified NGAL,MMP-9 complex. Results of Western immunoblotting showed that the HMW gelatinase was also recognized by antibodies specific for human NGAL or human MMP-9. These same antibodies also immunoprecipitated the HMW gelatinase activity from OA SF. The NGAL,MMP-9 complex was reconstituted in vitro in gelatinase buffer. In the presence of NGAL, MMP-9 activity was stabilized; in the absence of NGAL, rapid loss of MMP-9 activity occurred. MMP-9,mediated release of cartilage matrix proteoglycans was significantly higher in the presence of NGAL (P < 0.05). Conclusion Our findings demonstrate that the HMW gelatinase activity in OA SF represents a complex of NGAL and MMP-9. The ability of NGAL to protect MMP-9 activity is relevant to cartilage matrix degradation in OA and may represent an important mechanism by which NGAL may contribute to the loss of cartilage matrix proteins in OA. [source] | ||||||||||