The complex effects of the slow-releasing hydrogen sulfide donor GYY4137 in a model of acute joint inflammation and in human cartilage cells.
Bottom Line: We have now assessed the effect of GYY4137 (a slow-releasing H2 S donor) on lipopolysaccharide (LPS)-evoked release of inflammatory mediators from human synoviocytes (HFLS) and articular chondrocytes (HAC) in vitro.GYY4137 (0.1-0.5 mM) decreased LPS-induced production of nitrite (NO2 (-) ), PGE2 , TNF-α and IL-6 from HFLS and HAC, reduced the levels and catalytic activity of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and reduced LPS-induced NF-κB activation in vitro.In the CFA-treated mouse, GYY4137 (50 mg/kg, i.p.) injected 1 hr prior to CFA increased knee joint swelling while an anti-inflammatory effect, as demonstrated by reduced synovial fluid myeloperoxidase (MPO) and N-acetyl-β-D-glucosaminidase (NAG) activity and decreased TNF-α, IL-1β, IL-6 and IL-8 concentration, was apparent when GYY4137 was injected 6 hrs after CFA.
Affiliation: Pharmaceutical Science Research Division, King's College London, London, England.Show MeSH
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Mentions: To determine whether GYY4137 affected the levels of COX-2 or iNOS enzymes, intracellular levels of these proteins were determined in LPS-treated cells by ELISA. Treatment of HFLS or HAC with GYY4137 for 1 hr prior to LPS significantly reduced LPS-induced COX-2 (Fig. 3A) and iNOS (Fig. 3B) protein levels in addition to reducing PGE2 and NO2− levels. Although treatment of HFLS with a cocktail of cytokines (TNF-α, IL-1β and IFN-γ) induced an increase in the levels of TACE protein (Fig. 3C), the level of this enzyme was unaffected by GYY4137. To examine further the possibility that the attenuation of LPS-stimulated increases in PGE2, •NO and TNF-α (Figs 1 and 2) were due to an inhibitory effect of GYY4137 on COX-2, iNOS and TACE activity, respectively, we incubated human recombinant COX-2, iNOS and TACE with GYY4137 and determined residual catalytic activity. GYY4137 and Na2S significantly inhibited COX-2, iNOS and TACE activity (Fig. 3D–F), respectively, suggesting that H2S could directly inhibit enzyme activity and consequently cellular synthesis/secretion of PGE2, NO2− and TNF-α which is independent of any effect of this drug on COX-2 or iNOS protein levels.
Affiliation: Pharmaceutical Science Research Division, King's College London, London, England.