Structure of Cu2(indomethacin)4 and the reaction with superoxide in aprotic systems

The copper complex of indomethacin (1-(p-chlorobenzoyl)-5-methoxy-2-methyl-indole acetate), a common anti-inflammatory drug, was prepared and characterized. Crystal structure determination revealed the dimeric form of the 1:2 complex, namely Cu₂(indomethacin)₄ · L₂, in the unit cell. Suprisingly, the copper-copper distance (263 pm) was very close to metallic copper (256 pm). The two coordination sites in the copper-copper axis can be readily replaced by superoxide. An intriguing similarity to Cu₂(acetate)₄ was seen.Due to the lipophilic nature of the indomethacin ligand, this copper complex reacted with superoxide in aprotic solvents. The superoxide dismutating activity was successfully demonstrated in Me₂SO/water and acetonitrile/water mixtures using the nitro-blue tetrazolium assay and pulse radiolysis. The second-order rate constant of 6 · 10⁹ M^?1 · s^?1 in strictly aqueous systems dropped only slightly to 1.1 · 10⁹ M^?1 · s^?1 when aprotic solvents were used. This is the fastest rate constant ever observed for a copper-dependent dismutation of superoxide. The KO₂-induced lipid peroxidation in both erythrocytes and liver microsomes was suppressed by 70% in the presence of 1 · 10^?10 mol · ml^?1 of Cu₂(indomethacin)₄. The inhibitory action dropped to 25% when Cu₂Zn₂superoxide dismutase was employed. The formation of copper · indomethacin in rat serum after administration of indomethacin was shown in vitro and in vivo.     

The Administration of an α-MSH Analogue Reduces the Serum Release of IL-1α and TNFα Induced by the Injection of a Sublethal Dose of Lipopolysaccharides in the BALB/c Mouse

The injection of α-MSH or of one of its analogues ([Nle₄-D.Phe₇] α-MSH_4–10) reduced, in vivo, the release of two cytokines (IL-1α and TNFα) involved in inflammation. The inflammatory state was induced in BALB/c mice by intraperitoneal injection of a sublethal dose of lipopolysaccharides (LPS). The assay of these cytokines by ELISA showed a reduction of 20% with α-MSH and between 30 and 60% with the α-MSH analogue. The α-MSH or the analogue was administered in one of two ways: intravenously or subcutaneously. The most efficient method seemed to be the subcutaneous one because it improved the activity 10,000 times more than the intravenous method. Moreover, the analogue induced a regression of mortality in the animals treated by the intravenous method. Our results show that α-MSH and one of its analogues inhibit IL-1α and TNFα, and can be used as anti-inflammatory molecules.     

Dicoumarol derivatives: Green synthesis and molecular modelling studies of their anti-LOX activity

Dicoumarol derivatives were synthesized in the InCl₃ catalyzed pseudo three-component reactions of 4-hydroxycoumarin with aromatic aldehydes in excellent yields. The reactions were performed in water under microwave irradiation. All synthesized compounds were characterized using NMR, IR, and UV–Vis spectroscopy, as well as with TD-DFT. Obtained dicoumarols were subjected to evaluation of their in vitro lipid peroxidation and soybean lipoxygenase inhibition activities. It was shown that five of ten examined compounds (3e, 3h, 3b, 3d, 3f) possess significant potential of antilipid peroxidation (84–97%), and that compounds 3b, 3e, 3h provided the highest soybean lipoxygenase (LOX-Ib) inhibition (IC₅₀ = 52.5 µM) and 3i somewhat lower activity (IC₅₀ = 55.5 µM). The bioactive conformations of the best LOX-Ib inhibitors were obtained by means of molecular docking and molecular dynamics. It was shown that, within the bioactive conformations interior to LOX-Ib active site, the most active compounds form the pyramidal structure made of two 4-hydroxycoumarin cores and a central phenyl substituent. This form serves as a spatial barrier which prevents LOX-Ib Fe²⁺/Fe³⁺ ion activity to generate the coordinative bond with the C13 hydroxyl group of the α-linoleate. It is worth pointing out that the most active compounds 3b, 3e, 3h and 3i can be candidates for further examination of their in vitro and in vivo anti-inflammatory activity and that molecular modeling study results provide possibility to screen bioactive conformations and elucidate the mechanism of dicoumarols anti-LOX activity.     

Isopimarane-type diterpenoids from Callicarpa macrophylla vahl

Three new isopimarane-type diterpenoids, named callicapene M1 (1), callicapene M2 (2), and callicapene M3 (3), together with four known isopimarane-type diterpenoids (4, 5, 6, 7), were isolated from the Callicarpa macrophylla Vahl. Their structures were elucidated by spectroscopic techniques (IR, UV, MS, 1D, 2D NMR). The isolated compounds 6 and 7 exhibited potent inhibitory activity with inhibition rates of 40.23–46.78% on NO production in LPS-activated RAW 264.7 macrophage cells by using MTT assays.     

Anti-inflammatory action of gentamycin through inhibitory effect on neutrophil NADPH oxidase activity

The effects of gentamycin on the NADPH oxidase (EC 1.6.99.6) from human neutrophils in both whole-cell and fully soluble (cell-free) systems were investigated. Gentamycin was found to inhibit, concentration-dependently, the superoxide generation of neutrophils exposed to phorbol myristate acetate in a whole-cell system and the activation of superoxide-generating NADPH oxidase by sodium dodecyl sulfate in a cell-free system. The concentrations of the drug required for 50% inhibition of the oxidase (IC₅₀) were 150 μM in the whole-cell system and 10 μM in the cell-free system. In addition, in the cell-free system, the drug did not change the K_m value for NADPH of the oxidase. However, gentamycin did not the superoxide generation of NADPH oxidase after its activation in the cell-free system, suggesting that the drug do not have superoxide-scavenger action. These results suggest that gentamycin, an aminoglycoside antibiotic, may exhibit an anti-inflammatory action due to inhibition of neutrophil NADPH oxidase activation.     

Novel class of benzimidazole-thiazole hybrids: The privileged scaffolds of potent anti-inflammatory activity with dual inhibition of cyclooxygenase and 15-lipoxygenase enzymes

The present study includes design and synthesis of new molecular hybrids of 2-methylthiobenzimidazole linked to various anti-inflammatory pharmacophores through 2-aminothiazole linker, to investigate the effect of such molecular variation on cyclooxygenase (COX) and 15-lipoxygenase (15-LOX) enzymes inhibition as well as in vivo anti-inflammatory activity. The chemical structures of new hybrids were confirmed using different spectroscopic tools and elemental analyses. Benzimidazole-thiazole hybrids linked to acetyl moiety 13, phenyl thiosemicarbazone 14, 1,3-thiazolines 15a-c and 4-thiazolidinone 16 exhibited significant COX-2 inhibition (IC₅₀ = 0.045–0.075 µM) with significant COX-2 selectivity indices (SI = 142–294). All hybrids revealed potent 15-LOX inhibitory activity (IC₅₀ = 1.67–6.56 µM). Benzimidazole-thiazole hybrid 15b was the most potent dual COX-2 (IC₅₀ = 0.045 µM, SI = 294) inhibitor approximate to celecoxib (COX-2; IC₅₀ = 0.045 µM, SI = 327), with double inhibitory activity versus 15-LOX enzyme (IC₅₀ = 1.67 µM) relative to quercetin (IC₅₀ = 3.34 µM). Three hybrids (14, 15b & 16) were selected for in vivo screening using carrageenan-induced paw edema method. Benzimidazole-thiazole hybrid linked to 4-thiazolidinone 16 showed the maximum edema inhibition at both 3 h and 4 h intervals as well (~119% and 102% relative to indomethacin, respectively). The gastric ulcerogenic effect of benzimidazole-thiazole hybrid 16 was estimated compared with indomethacin showing superior gastrointestinal safety profile. In bases of molecular modeling; all new active hybrids were subjected to docking simulation into active sites of COX-2 and 15-LOX enzymes to study the binding mode of these novel potent dual COX-2/15-LOX inhibitors.     

肿瘤坏死因子α刺激基因/诱导蛋白-6的抗炎作用研究进展

炎症损伤是众多临床疾病的病理学基础,常可引起严重并发症甚至导致死亡。然而传统临床治疗不仅方法有限,且效果不佳。近年研究报道,肿瘤坏死因子α刺激基因/诱导蛋白-6(tumor necrosis factor alpha stimulated gene/inducible protein 6, TSG-6)可通过与体内相应的配体结合参与炎症反应的多个过程,并发挥抗炎和促进细胞外基质重塑等重要作用。本文就TSG-6的生物学特性、作用机制及其在病理性瘢痕、神经炎症、动脉粥样硬化和关节炎等多种疾病中发挥的抗炎作用作一综述。     

Zone Precipitation Chromatography: Its Use in the Isolation of Different Collagen Types

Zone Precipitation Chromatography is useful tech-nique for the initial isolation of the different colla-gen types in their native configuration. Small quan-tities of collagen mixtures can be rapidly separated into different collagen types with relatively high degree of purity, based upon stained protein patterns on sodium dodecyl sulfate polyacrylamide gel electro-phoresis (SDS-PAGE) slab gels. Tn the commonly used bulk salt preparative method for isolating the different collagens, 50 mg of starting material was needed. Three days were required to complete the procedure. The stained protein patterns on SDS-PAGE slab gels showed about 25% contamination with the bulk purified Type III fraction and 20% contamination with the bulk purified type AB collagen. With Zone Precipitation Chromatography 5 mg of starting material was used and in less than 4 hours the mixture was separated with Types III and AB fractions showing less than 10% contamination from other collagen types. The technique is patterned after the Zone Precinitation method reported by Porath seventeen years ago and utilizes a step-wise sodium chloride gradient to precipitate and redissolve the collagens, eluting from the interbead spaces of a molecular sieve column.     

Acidic Nonsteroidal Anti-inflammatory Drugs Inhibit Rat Brain Fatty Acid Amide Hydrolase in a pH-dependent Manner

Previous studies have demonstrated that fatty acid amide hydrolase, the enzyme responsible for the metabolism of anandamide, is inhibited by the acidic non-steroidal anti-inflammatory drug (NSAID) ibuprofen with a potency that increases as the assay pH is reduced. Here we show that (R) -, (S) - and (R, S) -flurbiprofen, indomethacin and niflumic acid show similar pH-dependent shifts in potency to that seen with ibuprofen. Thus, (S) -flurbiprofen inhibited 2 μM [3 H]anandamide metabolism with IC 50 values of 13 and 50 μM at assay pH values of 6 and 8, respectively. In contrast, the neutral compound celecoxib was a weak fatty acid amide hydrolase inhibitor and showed no pH dependency (IC 50 values ~300 μM at both assay pH). The cyclooxygenase-2-selective inhibitors nimesulide and SC-58125 did not inhibit fatty acid amide hydrolase activity at either pH. The data are consistent with the conclusion that the non-ionised forms of the acidic NSAIDs are responsible for the inhibition of fatty acid amide hydrolase.