Search

Activated NF-kappaB plays an important role in the expression of matrix metalloproteinase (MMP)-1 and MMP-13 in rheumatoid arthritis and osteoarthritis. The objective of this study was to determine the effects of the NF-kappaB inhibitor pyrrolidine dithiocarbamate (PDTC) on the expression of MMPs in IL-1beta-stimulated fibroblast-like synoviocytes (FLSs) of rheumatoid arthritis patients. FLSs were treated with IL-1beta (10 ng/ml) for 24 h in the presence or absence of PDTC. The level of MMP-1 and MMP-13 increased in response to PDTC in time- and dose-dependent manners in IL-1beta-stimulated FLSs; the expressions of IL-6 and vascular endothelial growth factor (VEGF) decreased in a PDTC concentration-dependent manner. However, PDTC-mediated repression of IL-6 and VEGF expression was not observed in TNF-alpha-stimulated rheumatoid arthritis FLSs. In contrast, other NF-kappaB inhibitors, such as fenofibrate, N-acetylcysteine and MG132, decreased MMP expression in IL-1beta-stimulated FLSs. The stimulatory effect of PDTC on MMP expression was not mimicked by specific inhibitors of the mitogen-activated protein kinase (MAPK) signaling pathway. Treatments with 100 muM PDTC did not inhibit the phosphorylation of p-ERK1/2, p-P38, and p-JNK, or the transnuclear migration of NF-kappaB through degradation of IkappaB-alpha in IL-1beta-stimulated FLSs. These results suggest that the increase of MMP expression may occur in a stimuli-specific manner or by an NF-kappaB independent mechanism. Therefore, therapeutic NF-kappaB inhibitors should be thoroughly studied before their clinical use in treating rheumatoid arthritis, as undesirable genes may be upregulated through unknown mechanisms, possibly resulting in worse symptoms.

Pyrrolidine dithiocarbamate (PDTC) can form a complex with metal ions and then act as a proteasome inhibitor, which leads to tumor cell apoptosis, and could therefore be developed as an anticancer agent. In our efforts to find factors that induce PDTC-mediated apoptosis of tumor cells, the effect of serum concentration on the apoptotic activity of PDTC was investigated. PDTC could not induce MCF-7 breast tumor cell death in serum-free media but significantly induced cell death in a dose-dependent manner at concentrations of ≥25 μM in media containing 10% fetal bovine serum. PDTC-mediated cell death was also dependent on serum concentration. PDTC-mediated cell death occurred through apoptosis. Similar to that in normal FBS, PDTC-mediated apoptotic cell death was also induced in media containing dialyzed FBS, indicating that PDTC-mediated apoptosis does not require metal ions or salts, but rather proteins in fetal bovine serum. In addition, differential apoptotic effects of PDTC were not observed with inhibitors of NF-κB activation such as N-acetylcysteine (NAC), Fenofibrate and carbobenzoxyl-l-leucyl-l-leucyl-l-leucinal (MG132) or with the metal-binding agent, 5-chloro-7-iodo-8-hydroxyquinoline (Clioquinol). These results indicate that serum is required for PDTC-mediated apoptosis and that zinc-binding compounds such as PDTC, N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) and Clioquinol may each have their own mechanisms by which they induce tumor cell death, even though they are all classified as zinc-binding compounds.

Inflammation in the joint of rheumatoid arthritis is a complex immune reaction facilitated by various factors, such as cytokines, cells and hypoxia. Thus, we evaluated their relative capacity to produce proinflammatory mediators in response to IL-1beta, TNF-alpha or IL-17 under hypoxia or normoxia in fibroblast-like synoviocytes (FLSs) and macrophages. The level of IL-6 expression was strongly increased in both FLSs and THP-1 macrophages in response to IL-1beta and TNF-alpha, but the level by TNF-alpha was less than that by IL-1beta. In contrast, the expression of IL-8 in both cell types was strongly stimulated by both IL-1beta and TNF-alpha. In FLSs, PGE(2) production increased only in response to IL-1beta; and no effect was observed in THP-1 cells and TNF-alpha-stimulated FLSs. In addition, the production by IL-17 was extremely low when compared with those induced by IL-1beta or TNF-alpha in FLSs and THP-1 cells. Hypoxia (2% O(2)) decreased IL-1beta-stimulated production of PGE(2), even though it increased the expression of mRNA and protein of COX-2. These results suggest that IL-1beta and TNF-alpha differentially regulate gene expression in FLSs and macrophages under hypoxia or normoxia.

Introduction The objective of this study was to determine the anti-inflammatory, nociceptive, and antiarthritic effects of piperine, the active phenolic component in black pepper extract. Methods The in vitro anti-inflammatory activity of piperine was tested on interleukin 1β (IL1β)-stimulated fibroblast-like synoviocytes derived form patients with rheumatoid arthritis. The levels of IL6, matrix metalloproteinase (MMPs), cyclo-oxygenase 2 (COX-2), and prostaglandin E2 (PGE2) were investigated by ELISA and RT-PCR analysis. The analgesic and antiarthritic activities of piperine were investigated on rat models of carrageenan-induced acute paw pain and arthritis. The former were evaluated with a paw pressure test, and the latter by measuring the squeaking score, paw volume, and weight distribution ratio. Piperine was administrated orally to rats at 20 and 100 mg/kg/day for 8 days. Results Piperine inhibited the expression of IL6 and MMP13 and reduced the production of PGE2 in a dose dependant manner at concentrations of 10 to 100 μg/ml. In particular, the production of PGE2 was significantly inhibited even at 10 μg/ml of piperine. Piperine inhibited the migration of activator protein 1 (AP-1), but not nuclear factor (NF)κB, into the nucleus in IL1β-treated synoviocytes. In rats, piperine significantly reduced nociceptive and arthritic symptoms at days 8 and 4, respectively. Histological staining showed that piperine significantly reduced the inflammatory area in the ankle joints. Conclusions These results suggest that piperine has anti-inflammatory, antinociceptive, and antiarthritic effects in an arthritis animal model. Thus, piperine should be further studied with regard to use either as a pharmaceutical or as a dietary supplement for the treatment of arthritis.

It has been suggested that taurine chloramine (TauCl) plays an important role in the downregulation of proinflammatory mediators. However, little is known about its effect on the expression of matrix metalloproteinases (MMPs). In this study, we investigated the effects of TauCl on synovial expression of MMPs. The effects of TauCl on MMP expression in IL-1beta stimulated fibroblast-like synoviocytes (FLSs) were studied using the following techniques. Real-time PCR and semi-quantitative PCR were employed to analyze the mRNA expression of MMPs. ELISA was used to determine protein levels of MMPs. Western blot analyses were performed to analyze the mitogen-activated protein kinase and inhibitor of nuclear factor-kappaB (IkappaB) kinase signalling pathways. Finally, electrophoretic mobility shift assay and immunohistochemistry were used to assess localization of transcription factors. IL-1beta increased the transcriptional and translational levels of MMP-1 and MMP-13 in rheumatoid arthritis FLSs, whereas the levels of MMP-2 and MMP-9 were unaffected. TauCl at a concentration of 400 to 600 micromol/l greatly inhibited the transcriptional and translational expression of MMP-13, but the expression of MMP-1 was significantly inhibited at 800 micromol/l. At a concentration of 600 micromol/l, TauCl did not significantly inhibit phosphorylation of mitogen-activated protein kinase or IkappaB degradation in IL-1beta stimulated rheumatoid arthritis FLSs. The degradation of IkappaB was significantly inhibited at a TauCl concentration of 800 micromol/l. The inhibitory effect of TauCl on IkappaB degradation was confirmed by electrophoretic mobility shift assay and immunochemical staining for localization of nuclear factor-kappaB. TauCl differentially inhibits the expression of MMP-1 and MMP-13, and inhibits expression of MMP-1 primarily through the inhibition of IkappaB degradation, whereas it inhibits expression of MMP-13 through signalling pathways other than the IkappaB pathway.