diamide and Inflammation

A series of novel non-peptide diamide compounds was synthesized and evaluated as antibradykinin agents by utilizing guinea-pig ileum smooth muscle. Among the final compounds, (Z)-4-(4-(bis(4-fluorophenyl)methyl)piperazin-1-yl)-4-oxo-N-(4-phenylbutan-2-yl)but-2-enamide showed most favorable bradykinin inhibitory activity and demonstrated analgesic efficacies in the rat models of inflammatory and neuropathic pain.

Topics: Analgesics; Animals; Anti-Inflammatory Agents; Bradykinin; Diamide; Disease Models, Animal; Guinea Pigs; Ileum; Inflammation; Molecular Structure; Muscle, Smooth; Pain; Rats; Stereoisomerism; Structure-Activity Relationship

Resolving inflammation is a vital step in preventing the persistence of inflammatory disorders. Neutrophils play a major role in tissue damage associated with an inflammatory response. Their death by apoptosis is central to the final resolution of this response. Thiol depletion with diethylmaleate (DEM) or diamide represent important triggers for neutrophil apoptosis. The mechanism by which this process occurs remains unknown. The apoptotic cascade is associated with a number of cellular changes, including caspase activation and mitochondrial permeability. The aims of this study were to determine the role of mitochondrial permeability and the caspase cascade in thiol depletion-induced neutrophil apoptosis. Total cellular glutathione was reduced by DEM and diamide. This reduction was associated with neutrophil apoptosis and an increase in caspase 3 activity. The effects of DEM were blocked by the caspase 3 inhibitor, Z-DEVD-FMK. Mitochondrial permeability that occurred was also increased during this induction of apoptosis. Bongkrekic acid, a mitochondrial membrane stabilizer, inhibited DEM-induced apoptosis. The inhibitors' effects of LPS or GM-CSF on spontaneous neutrophil apoptosis was reversed by DEM, which was mediated by an increase in caspase 3 activity and independent of mitochondrial disruption. Caspase activation is an important step in glutathione depletion-induced apoptosis in resting and inflammatory neutrophils. Regulation of caspase activity may represent a possible target to trigger apoptosis and resolve inflammatory disorders.

Topics: Apoptosis; Caspase 3; Caspase Inhibitors; Caspases; Cysteine Proteinase Inhibitors; Diamide; Glutathione; Humans; In Vitro Techniques; Inflammation; Maleates; Mitochondria; Neutrophils; Oligopeptides; Permeability

The present study underlines the importance of reactive oxygen species in cytokine-mediated degradation of sphingomyelin (SM) to ceramide. Treatment of rat primary astrocytes with tumor necrosis factor-alpha (TNF-alpha) or interleukin-1beta led to marked alteration in cellular redox (decrease in intracellular GSH) and rapid degradation of SM to ceramide. Interestingly, pretreatment of astrocytes with N-acetylcysteine (NAC), an antioxidant and efficient thiol source for glutathione, prevented cytokine-induced decrease in GSH and degradation of sphingomyelin to ceramide, whereas treatment of astrocytes with diamide, a thiol-depleting agent, alone caused degradation of SM to ceramide. Moreover, potent activation of SM hydrolysis and ceramide generation were observed by direct addition of an oxidant like hydrogen peroxide or a prooxidant like aminotriazole. Similar to NAC, pyrrolidinedithiocarbamate, another antioxidant, was also found to be a potent inhibitor of cytokine-induced degradation of SM to ceramide indicating that cytokine-induced hydrolysis of sphingomyelin is redox-sensitive. Besides astrocytes, NAC also blocked cytokine-mediated ceramide production in rat primary oligodendrocytes, microglia, and C6 glial cells. Inhibition of TNF-alpha- and diamide-mediated depletion of GSH, elevation of ceramide level, and DNA fragmentation (apoptosis) in primary oligodendrocytes by NAC, and observed depletion of GSH, elevation of ceramide level, and apoptosis in banked human brains from patients with neuroinflammatory diseases (e.g. X-adrenoleukodystrophy and multiple sclerosis) suggest that the intracellular level of GSH may play a critical role in the regulation of cytokine-induced generation of ceramide leading to apoptosis of brain cells in these diseases.

Topics: Acetylcysteine; Adrenoleukodystrophy; Amitrole; Animals; Antioxidants; Apoptosis; Brain; Cells, Cultured; Ceramides; Cytokines; Diamide; DNA Fragmentation; Glutathione; Glutathione Disulfide; Hydrogen Peroxide; Inflammation; Interleukin-1; Multiple Sclerosis; Oxidation-Reduction; Pyrrolidines; Rats; Reactive Oxygen Species; Sphingomyelins; Thiocarbamates; Tumor Necrosis Factor-alpha