Blockade of recombinant human IL-6 by tocilizumab suppresses matrix metalloproteinase-9 production in the C28/I2 immortalized human chondrocyte cell line

Abstract

Two immortalized human juvenile chondrocyte cell lines, T/C28a2 and C28/I2, were employed to determine the extent to which recombinant human (rh) IL-6 or rh-TNF-α increased the production of matrix metalloproteinase-9 (MMP-9). The effect of rhIL-6 on neutrophil gelatinase-associated lipocalin (NGAL) was also assessed. Although C28/I2 chondrocytes incubated with rhIL-6 (50 ng/ml) increased MMP-9 production which could not be mimicked by the T/C28a2 chondrocyte line, the effect of rhTNF-α on MMP-9 was more robust than with rhIL-6. The combinations of rhIL-6 and soluble IL-6 receptor-α (sIL-6Rα) or rhIL-6 and tocilizumab (TCZ), a fully-humanized recombinant monoclonal antibody that neutralizes the interaction between IL-6 and IL-6R significantly reduced MMP-9 production by C28/I2 chondrocytes. However, TCZ had no effect on rhTNF-α-induced MMP-9 production. By contrast, rhIL-6 did not increase the production of NGAL by C28/I2 chondrocytes although the number of NGAL-positive cells was significantly reduced by sIL-6R compared to its control group, but not by the combination of rhIL-6 plus TCZ compared to rhIL-6. In summary, these results showed that rhIL-6 stimulated the production of MMP-9, but not NGAL, in the C28/I2 chondrocyte line. TCZ or sIL-6Rα suppressed rhIL-6-induced MMP-9 production.

Anti-inflammatory and anti-osteoarthritis effect ofMollugo pentaphyllaextract

“…The production of IL-6 in diseased joint tissues is generally mediated by chondrocytes, osteoblasts and macrophages in response to IL-1b and TNF-a (Guerne et al 1990). These inflammatory cytokines stimulate a major cascade of events involving the degradation of cartilage matrix and the destruction of articular cartilage, such as the synthesis of MMPs and ECM proteins, which are absent in normal cartilage, and the release of other inflammatory mediators, including COX-2 (Largo et al 2003;Akhtar et al 2011;Meszaros et al 2015;Zeng et al 2015). We evaluated the therapeutic potential of MPE for pain relief and chondroprotective effects in MIA-induced OA in rats by measuring the weight-bearing distribution, inflammatory cytokines and mediator levels in serum, inflammation-related gene expression in knee joints, and histopathological parameters.…”

Matrix Metalloproteinase-9 Production by Immortalized Human Chondrocyte Lines

“…PANC-1 was incubated with phorbol myristate acetate. PANC-1 was employed as the positive control for MMP-9 production [ 14 ]. The pro-inflammatory cytokines, rhIL-6 and rhTNF-α were obtained from various commercial vendors as was sIL-6R as previously described [ 14 ].…”

“…PANC-1 was employed as the positive control for MMP-9 production [ 14 ]. The pro-inflammatory cytokines, rhIL-6 and rhTNF-α were obtained from various commercial vendors as was sIL-6R as previously described [ 14 ]. U0126, a small molecule inhibitor of MEK1/2, an upstream protein kinase required for the phosphorylation of ERK1/2 was purchased from Cell Signaling Technology.…”

U0126, an Inhibitor of MEK1/2, Increases Tumor Necrosis Factor-?-Induced Apoptosis, but not Interleukin-6 Induced Apoptosis in C-28/I2 Human Chondrocytes

“…Thus, one limitation on the interpretation of the DAPI/TUNEL analysis may reflect, in part, the concentration of these pro-inflammatory cytokines early on in RA pathogenesis, but may not be an accurate indicator of the level of these cytokines after medical therapy of RA is initiated by either by TNF-α blockade or neutralization of the IL-6/IL-6R interaction. The concentration of tocilizumab [200 ng/ml] employed in this study to determine whether tocilizumab altered the pro-caspase-3 activation and PARP degradation after treating C-28/I2 chondrocytes with rhIL-6 (Table 1) was based on the results of our prior study [31] which showed that this concentration of tocilizumab [i.e., 200 ng/ml] inhibited rhIL-6-inducd MMP-9 production.…”