m-40403 and Inflammation

In this review we describe the potential role(s) of superoxide in inflammatory disorders.

Topics: Anti-Inflammatory Agents; Free Radical Scavengers; Humans; Inflammation; Manganese; Molecular Structure; Organometallic Compounds; Superoxide Dismutase; Superoxides

Gram-negative bacterial infection predisposes to the development of shock and acute lung injury with multiple organ dysfunction in the critically ill. Although overexpression of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1beta, IL-6, IL-8, and other mediators is causally implicated in the pathogenesis of shock and lung injury, the underlying mechanisms following cellular exposure to gram-negative endotoxin remain unclear. De novo generation of reactive oxygen species (ROS) by monocytes/macrophages in particular has been proposed as a pivotal regulatory mechanism by which enhanced transactivation of redox-sensitive genes culminates in augmented cytokine expression within the lower respiratory tract. Here we sought to characterize the mechanism of action of a synthetic, nonpeptide, low-molecular-weight, Mn-containing superoxide dismutase mimetic (SODm), M40403, in modulating E. coli lipopolysaccharide serotype 0111:B4 (LPS)-induced cytokine production by cultured rat alveolar macrophages. Intracellular superoxide (O2) ion generation was measured using hydroethidine (HE) dye, and the dose-dependent effects of M40403 on TNF-alpha and IL-6 biosynthesis by ELISAs. Upstream redox-sensitive signaling events involving the pleiotropic transcription factor NF-kappaB were determined in nuclear extracts by electrophoretic mobility shift assays (EMSAs) and p65 subunit Western blot. The levels of the cytosolic inhibitory protein IkappaB-alpha were also assessed by Western analysis. We found that M40403 potently suppressed the production of superoxide, TNF-alpha, and IL-6 in LPS-stimulated alveolar macrophages, suggesting a key role for superoxide in endotoxin-induced cytokine production in the distal air spaces. In addition, M40403 decreased E. coli LPS-induced activation of NF-kappaB, and this effect was associated with modest suppression of cytoplasmic IkappaB-alpha degradation. Together, these results suggest that removal of superoxide by M40403 inhibits endotoxin-induced production of TNF-alpha and IL-6 in alveolar macrophages by a mechanism involving suppression of redox-sensitive NF-kappaB transactivation or signaling.

Topics: Animals; Blotting, Western; Cell Nucleus; Coloring Agents; Cytokines; Cytoplasm; Endotoxins; Enzyme Activation; Enzyme-Linked Immunosorbent Assay; Escherichia coli; I-kappa B Proteins; Immunoblotting; Inflammation; Interleukin-1; Interleukin-6; Interleukin-8; Lipopolysaccharides; Macrophages; Macrophages, Alveolar; Manganese; Models, Biological; NF-kappa B; NF-KappaB Inhibitor alpha; Organometallic Compounds; Oxidation-Reduction; Oxygen; Phenanthridines; Rats; Reactive Oxygen Species; Signal Transduction; Superoxide Dismutase; Superoxides; Time Factors; Transcription Factor RelA; Tumor Necrosis Factor-alpha

The nuclear factor-kappaB (NF-kappaB) is a transcription factor that plays a pivotal role in the induction of genes involved in physiological processes, as well as in the response to inflammation. In this study, we used a selective nonpeptidyl superoxide dismutase mimetic, M40403, to investigate the role of superoxide anion in NF-kappaB activation during acute inflammation in mice. Injection of carrageenan into the pleural cavity of mice induced an acute inflammatory response characterized by fluid accumulation in the pleural cavity that contained a large number of neutrophils, as well as an increased production of tumor necrosis factor-alpha and interleukin-1beta. All parameters of inflammation were attenuated by M40403 (10 mg/kg i. p., 30 min prior to carrageenan administration). These inflammatory events were associated with the activation of NF-kappaB in the lung. In particular, the appearance of inhibitory protein kappaB-alpha (IkappaB-alpha) in homogenates of lung tissues was investigated by immunoblot analysis at 4 h after carrageenan administration. IkappaB-alpha levels were substantially reduced in the lung tissue from carrageenan-treated mice in comparison with sham-treated mice. Furthermore, to detect NF-kappaB/DNA binding activity, whole extracts from lung tissue of each mouse were analyzed by electrophoretic mobility-shift assay. The DNA binding activity significantly increased in whole extracts obtained from lung tissues of vehicle-treated mice 4 h after carrageenan administration. Treatment of mice with M40403 caused a significant inhibition of carrageenan-induced IkappaB-alpha degradation and NF-kappaB/DNA binding activity. These data confirm that M40403 exerts a potent antiinflammatory activity and clearly demonstrate that the reduction of the inflammatory process is associated with modification of the activation of signal transduction pathways.

Topics: Animals; Carrageenan; I-kappa B Proteins; Inflammation; Interleukin-1; Lung; Male; Manganese; Mice; NF-kappa B; NF-KappaB Inhibitor alpha; Organometallic Compounds; Pleurisy; Random Allocation; Signal Transduction; Superoxide Dismutase; Superoxides; Tumor Necrosis Factor-alpha

Many human diseases are associated with the overproduction of oxygen free radicals that inflict cell damage. A manganese(II) complex with a bis(cyclohexylpyridine)-substituted macrocyclic ligand (M40403) was designed to be a functional mimic of the superoxide dismutase (SOD) enzymes that normally remove these radicals. M40403 had high catalytic SOD activity and was chemically and biologically stable in vivo. Injection of M40403 into rat models of inflammation and ischemia-reperfusion injury protected the animals against tissue damage. Such mimics may result in better clinical therapies for diseases mediated by superoxide radicals.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Cytoprotection; Dinoprostone; Dose-Response Relationship, Drug; Drug Design; Drug Stability; Inflammation; Interleukin-1; L-Lactate Dehydrogenase; Male; Manganese; Molecular Mimicry; Neutrophils; Organometallic Compounds; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Splanchnic Circulation; Superoxide Dismutase; Superoxides; Time Factors; Tumor Necrosis Factor-alpha

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Computer Simulation; Drug Design; Inflammation; Intestines; Manganese; Molecular Mimicry; Organometallic Compounds; Rats; Reperfusion Injury; Superoxide Dismutase; Superoxides