The effects of PDTC on interleukin-1β-induced nitric oxide production in chondrocytes

Abstract

In order to find new drugs to inhibit nitric oxide (NO) production, the effects of pyrrolidine dithiocarbamate (PDTC), a nuclear factor-kappa B (NF-κB) inhibitor, on recombinant human interleukin-1β (rhIL-1β)-induced NO production in chondrocytes were investigated. Rat chondrocytes were isolated and cultured, divided into control, P0, P1, P2, P3 and P4 groups. The chondrocytes in the P0, P1, P2, P3 and P4 groups were treated with different concentrations of PDTC (0, 3, 10, 30, and 50 μmol/L respectively) for 1 h and then incubated with 5 U/mL rhIL-1β for 24 h. NO assay kit and RT-PCR were used to detect the NO content and the iNOS mRNA expression in the chondrocytes. The expression level of iNOS mRNA in control, P0, P1, P2, P3 and P4 groups was 0.02±0.01, 1.24±0.13, 1.21±0.14, 0.61±0.11, 0.40±0.09, 0.21±0.06, and the relative content of NO was 15.8±2.7, 100±14.8, 92.6±9.3, 68.3±14.2, 27.5±9.8, 19.8±3.6, respectively. In the P0, P1, P2, P3 and P4 groups, the expression of iNOS mRNA and NO production were significantly increased as compared with those in the control group. As compared with the P0 group, the expression of iNOS mRNA and NO content in control group were lower. In the P2, P3 and P4 groups, PDTC could significantly inhibit the expression of iNOS and NO production induced by rhIL-1β in a concentration-dependent manner. It is suggested that PDTC can inhibit NO production and iNOS mRNA expression induced by IL-1β, which may provide an alternative method for the treatment of osteoarthritis.

Keywords

pyrrolidine dithiocarbamate / chondrocyte / nitric oxide / nuclear factor-kappa B

References

[1] CastroR.R., CunhaF.Q., SilvaF.S.Jr, et al. . A quantitative approach to measure joint pain in experimental osteoarthritis—evidence of a role for nitric oxide. Osteoarthritis Cartilage, 2006, 14(8): 769-776

[2] MinB.H., KimH.J., LimH., et al. . Effects of ageing and arthritic disease on nitric oxide production by human articular chondrocytes. Exp Mol Med, 2001, 33(4): 299-302

[3] NotoyaK., JovanovicD.V., ReboulP., et al. . The induction of cell death in human osteoarthritis chondrocytes by nitric oxide is related to the production of prostaglandin E2 via the induction of cyclooxygenase-2. J Immunol, 2000, 165(6): 3402-3410

[4] PelletierJ.P., JovanovicD., FernandesJ.C., et al. . Reduced progression of experimental osteoarthritis in vivo by selective inhibition of inducible nitric oxide synthase. Arthritis Rheum, 1998, 41(7): 1275-1286

[5] XieQ.W., KashiwabaraY., NathanC.. Role of transcription factor NF-kappa B/Rel in induction of nitric oxide synthase. J Biol Chem, 1994, 269(7): 4705-4708

[6] BlackwellT.S., ChristmanJ.W.. The role of nuclear factor-kappa B in cytokine gene regulation. Am J Respir Cell Mol Biol, 1997, 17(1): 3-9

[7] SchreckR., MeierB., MannelD.N., et al. . Dithiocarbamates as potent inhibitors of nuclear factor kapaB activation in intact cells. J Exp Med, 1992, 175(5): 1181-1194

[8] NurmiA., VartiainenN., PihlajaR., et al. . Pyrrolidine dithiocarbamate inhibits translocation of nuclear factor kappa-B in neurons and protects against brain ischaemia with a wide therapeutic time window. J Neurochem, 2004, 91(3): 755-765

[9] NedelecE., AbidA., CipollettaC., et al. . Stimulation of cyclooxygenase-2-activity by nitric oxide-derived species in rat chondrocyte: lack of contribution to loss of cartilage anabolism. Biochem Pharmacol, 2001, 61(8): 965-978

[10] ShikhmanA.R., KuhnK., AlaaeddineN., et al. . N-Acetylglucosamine prevents IL-1beta-mediated activation of human chondrocytes. J Immunol, 2001, 166(8): 5155-5160

[11] BadgerA.M., CookM.N., SwiftB.A., et al. . Inhibition of interleukin-1-induced proteoglycan degradation and nitric oxide production in bovine articular cartilage/chondrocyte cultures by the natural product, hymenialdisine. J Pharmacol Exp Ther, 1999, 290(2): 587-593

[12] SaklatvalaJ.. Tumor necrosis factor alpha stimulates resorption and inhibits synthesis of proteoglycan in cartilage. Nature, 1986, 322(6079): 547-549

[13] GoldringM.B.. The role of cytokine as inflammatory mediators in osteoarthritis: lessons from animal modes. Connect Tissue Res, 1999, 40(1): 1-11

[14] Di MarcoS., MazrouiR., DallaireP., et al. . NF-kappa B-mediated MyoD decay during muscle wasting requires nitric oxide synthase mRNA stabilization, HuR protein, and nitric oxide release. Mol Cell Biol, 2005, 25(15): 6533-6545

[15] YamamotoY., GaynorR.B.. IkB kinases: key regulators of the NF-kappaB pathway. Trends Biochem Sci, 2004, 29(2): 72-79

[16] BouwmeesterT., BauchA., RuffnerH., et al. . A physical and functional map of the human TNF-alpha/NF-kappa B signal transduction pathway. Nat Cell Biol, 2004, 6(2): 97-105

[17] TakP.P., FiresteinG.S.. NF-kB: a key role in inflammatory disease. J Clin Invest, 2001, 107(1): 7-11

[18] ClancyR.M., AminA.R., AbramsonS.B.. The role of nitric oxide in inflammation and immunity. Arthritis Rheum, 1998, 41(7): 1141-1151

[19] StuderR., JaffursD., Stefanovic-RacicM., et al. . Nitric oxide in osteoarthritis. Osteoarthritis Cartilage, 1999, 7(4): 377-379

[20] PelletierJ.P., Lascau-ComanV., JovanovicD., et al. . Selective inhibition of inducible nitric oxide synthase in experimental osteoarthritis is associated with reduction in tissue levels of catabolic factors. J Rheumatol, 1999, 26(9): 2002-2014

[21] PelltierJ.P., JovanovicD.V., Lascau-ComanV., et al. . Selective inhibitor of inducible nitric oxide synthase reduces progression of experimental osteoarthritis in vivo: possible link with the reduction in chondrocyte apoptosis and caspase 3 level. Arthritis Rheum, 2000, 43(6): 1290-1299

[22] CuzzoreaS., ChatterjeeP.K., MazzonE., et al. . Pyrrolidine dithiocarbamate attenuates the developmentof acute and chronic inflammation. Br J Pharmacol, 2002, 135(2): 496-510