Involvement of testosterone in the induction of hepatic microsomal cytochrome P-450 2B1/2 (P-450 2B1/2) by 1-benzylimidazole in male and female rats: sex-differentiated induction of P-450 2B1/2 species

Collagen-induced arthritis (CIA) is an animal model for the human autoimmune disease rheumatoid arthritis (RA). CIA can be induced in several species including primates by immunization with heterologous type-II collagen (CII). Polyclonal antibodies are formed upon immunization with CII that exhibit a broad range of epitope specificities (some that cross-react with hose CII); however, only antibodies directed against certain specific epitopes on CII are arthritogenic. Recently, the importance of cognate interactions between T-cells and B-cells to the induction of CIA was demonstrated by administration of monoclonal antibodies against a T-cell surface protein, gp39. Blocking the interaction of T-cell gp39, with its receptor/ligand on the surface of B-cells (CD40), completely blocked induction of CIA in mice. A concomitant reduction in the level of anti-CII IgG produced in anti-gp39-treated animals was observed, demonstrating the crucial importance of T-cell:B-cell interactions via gp39:CD-40 binding to the primary immune response to CII in vivo and therefore to the induction of CIA. Other features of CIA are important in elucidating the condition and this article will deal with some important issues.     

Nitrosoalkanes as Fe(II) ligands in the 455-nm-absorbing cytochrome P-450 complexes formed from nitroalkanes in reducing conditions

Primary and secondary aliphatic nitroalkanes RR'CHNO2, react with microsomal cytochrome p-450 in the presence of dithionite leading to new complexes with a Soret peak at 455 nm. The formation of these complexes is inhibited completely by CO and partially by metyrapone. However, once formed, their exogenous ligand is not displaced by excess CO. By deoxycholate treatment they are transformed into 423-nm-absorbing cytochrome P-420 complexes, which are spectrally similar to the corresponding RR'CHNO2-derived myoglobin complexes. The 455-nm-absorbing complexes are equally produced from RR'CHNO2 reduction, microsomal NADPH-dependent oxidation of the corresponding hydroxylamine RR'CHNHOH, or interaction of the nitrosodimer (RR'CHNO)2 with reduced cytochrome P-450. All the reported results are consistent with the involvement of a new class of ligands of cytochrome P-450-Fe(II), which are the unstable aliphatic nitrosomonomers RR'CHNO, whose nitroso group is isoelectronic with dioxygen and whose stabilisation results from their strong binding to heme-Fe(II), thus explaining the observed inhibition of the hydroxylating function of cytochrome P-450.     

Iron(II) oxidation by SO2/O2 for use in uranium leaching

Iron(III) can be used as an oxidant in the leaching of uranium ore in an acid medium. The oxidation of iron(II) to iron(III) using an SO₂/O₂ gas mixture was investigated in order to provide an iron(III) stream for uranium extraction. The effects of pH, temperature and SO₂/O₂ volumetric ratios were considered. Oxidation of iron(II) by SO₂/O₂ was controlled by diffusion of SO₂ or O₂ at pH 2 and 40 °C. However as the pH decreased below pH 1, the reaction was controlled by a slow chemical step and the reaction rate decreased. Increasing the temperature increased the oxidation rate at pH 0.8, and at 70 °C the rate again became dependent on SO₂ or O₂ diffusion. The oxygen efficiency for a fixed reactor set-up was dependent on the SO₂/O₂ ratio and total flow rate of the gas. In leach tests, the uranium extraction achieved with iron(III) solution prepared by SO₂/O₂ oxidation was the same as that for a standard uranium leach with conventional oxidant.     

Speciation of the Fe(II)–Fe(III)–H2SO4–H2O system at 25 and 50 °C

This work presents theoretical and experimental results on the speciation of the Fe(II)–Fe(III)–H₂SO₄–H₂O system in concentrated solutions (up to 2.2 m H₂SO₄ and 1.3 m Fe). The aim was to study the chemical equilibria of iron at 25 and 50 °C in synthetic aqueous sulphuric acid solutions that contain dissolved ferric and ferrous ion species. Raman spectroscopy, volumetric titration and conductivity measurements have been carried out in order to study the presence of specific ions and to characterize the ionic equilibrium. A thermochemical equilibrium model incorporating an extended Debye–Hückel relationship was used to calculate the activities of ionic species in solution. Model calculations were compared with experimental results. Model simulations indicate that anions, cations and neutral complexes coexisted in the studied system, where the dominant species were HSO₄⁻, H⁺, Fe²⁺ and FeH(SO₄)₂⁰. This indicated that these solutions showed a high buffer capacity due to the existence of bisulphate ions (HSO₄⁻), which presented the highest concentration. A decrease in the concentration of H⁺ and Fe³⁺ took place with increasing temperature due to the formation of complex species. Standard equilibrium constants for the formation of FeH(SO₄)₂⁰ were obtained in this work: log K_f⁰ = 8.1 ± 0.3 at 25 °C and 10.0 ± 0.3 at 50 °C.     

The mechanism of a P-450 enzyme-aromatase; a molecular modelling perspective for the removal of the C(19) methyl and aromatisation of the steroid A ring

1 Here we compared the effects of various inhibitors of the activity of protein tyrosine kinase on (i) the expression of the activity of the inducible isoform of nitric oxide (NO) synthase (iNOS) caused by endotoxin (lipopolysaccharide, LPS) in cultured macrophages, (ii) the induction of iNOS and cyclooxygenase 2 (COX-2) protein and activity in rats with endotoxaemia, and (iii) the circulatory failure and organ dysfunction caused by LPS in the anesthetized rat. 2 Activation of murine cultured macrophages with LPS (1 microgram ml-1) resulted, within 24 h, in a significant increase in nitrite (an indicator of the formation of NO) in the cell supernatant. This increase in nitrate was attenuated by the tyrphostins AG126, AG556, AG490 or AG1641 or by genistein in a dose-dependent fashion (IC50: approximately 15 microM). In contrast, tyrphostin A1 (an analogue of tyrphostin AG126) or daidzein (an analogue of genistein) had no effect on the rise in nitrite caused by LPS. 3 Administration of LPS (E. coli, 10 mg kg-1, i.v.) caused hypotension and a reduction of the pressor responses elicited by noradrenaline (NA, 1 microgram kg-1, i.v.). Pretreatment of rats with the tyrphostins AG126, AG490, AG556, AG1641 or A1 attenuated the circulatory failure caused by LPS. Although genistein attenuated the vascular hyporeactivity to NA, it did not affect the hypotension caused by LPS. Daidzein did not affect the circulatory failure caused by LPS. 4 Endotoxaemia for 360 min resulted in rises in the serum levels of (i) urea and creatinine (indicators of renal failure), (ii) alanine aminotransferase (ALT), aspartate aminotransferase (AST), bilirubin and gamma-glutamyl transferase (gamma GT) (indicators of liver injury/dysfunction), lipase (an indicator of pancreatic injury) as well as lactate (an indicator of tissue hypoxia). None of the tyrosine kinase inhibitors tested had a significant effect on the rise i the serum levels of urea, but the tyrphostins AG126, AG556 or A1 significantly attenuated the rises in the serum level of creatinine caused by LPS. In addition, all tyrphostins and genistein attenuated the liver injury/failure, the pancreatic injury, the hypoglycaemia and the lactic acidosis caused by LPS. In contrast, daidzein did not reduce the organ injury/dysfunction or the lactic acidosis caused by LPS. 5 Injection of LPS resulted (within 90 min) in a substantial increase in the serum level of tumor necrosis factor alpha (TNF alpha), which was attenuated by pretreatment of LPS-rats with any of the tyrphostins used. Genistein, but not daidzein, also reduced the rise in the serum levels of TNF alpha caused by LPS. Endotoxaemia for 6 h also resulted in a substantial increase in the expression of iNOS and COX-2 protein and activity in the lung, which was attenuated by pretreatment of LPS-rats with the tyrphostins AG126, AG556 or genistein, but not by daidzein. 6 Thus, tyrphostins (AG126, AG556, AG1641 or A1) and genistein, but not daidzein (inactive analogue of genistein), prevent the (i) circulatory failure, (ii) the multiple organ dysfunction (liver and pancreatic dysfunction/injury lactacidosis, hypoglycaemia), as well as (iii) the induction of iNOS and COX-2 protein and activity in rats with endotoxic shock.     

An investigation of the thermodynamics of solvent extraction of metals II. Calculation of the activity coefficient of non-electrolytes in the UO2(NO3)2(C7H15)2SO system

The thermodynamic method used in Part I for evaluating activity coefficients of mixed non-electrolytes in the ternary organic phase of a metal extraction system has been extended successfully to a quaternary organic phase. Experimental and calculated results indicate that both the Scatchard-Hildebrand model and the Guggenheim quasi-lattice model are applicable for calculation of the activity coefficients in the organic phase of the (C₂H₁₅)₂SOC₆H₄(CH₃)₂UO₂(NO₃)₂ · 2 (C₇H₁₅)₂SOH₂O system.     

Phase equilibrium data and liquidus for the system “MnO”-CaO-(Al2O3 + SiO2) at Al2O3/SiO2 = 0.41

Phase-equilibrium data and the liquidus for the system “MnO”-CaO-(Al₂O₃-SiO₂) at a manganese-rich alloy saturation have been determined in the temperature range from 1423 to 1723 K. The results are presented in the form of a pseudoternary section “MnO”-CaO-(Al₂O₃ + SiO₂) with an Al₂O₃/SiO₂ weight ratio of 0.41. The following primary phases are present in the range of conditions investigated: 3Al₂O₃·2SiO₂; SiO₂; MnO·Al₂O₃·2SiO₂; (Mn,Ca)O·SiO₂; 2(Mn,Ca)O·SiO₂; MnO·Al₂O₃; (Mn,Ca)O; α-2CaO·SiO₂; α′-2CaO·SiO₂; 2CaO·Al₂O₃·SiO₂; CaO·SiO₂, and CaO·Al₂O₃·2SiO₂. The presence of alumina in this system is shown to have a significant effect on the liquidus compared to the system “MnO”-CaO-SiO₂, leading to the stabilization of the anorthite and gehlenite phases.     

Empirical model for the extraction system YCl3ErCl3HClH2ODI-(2-ethylhexyl)phosphoric acid—n-heptane

Equilibrium data were obtained for the extraction of the binary rare earth mixture yttrium chloride-erbium chloride from 2 kmol/m³ HClH₂O solutions by 1 kmol/m³ di(2-ethylhexyl)phosphoric acid in n-heptane. Total rare earth concentration in the aqueous phase was 0.5 kmol/m³. It was found that yttrium concentrations in the organic phase exhibit negative deviations whereas erbium concentrations exhibit positive deviations from ideality. Empirical correlations for predicting these deviations were developed. The separation factor was calculated.     

Experimental study of phase equilibria in the PbO-ZnO-“Fe2O3”-(CaO + SiO2) system in air for the lead and zinc blast furnace sinters (CaO/SiO2 weight ratio of 0.933 and PbO/(CaO + SiO2) ratios of 2.0 and 3.2)

Experimental studies on phase equilibria and liquidus in the multicomponent system PbO-ZnO-CaO-SiO₂-FeO-Fe₂O₃ in air have been conducted over the temperature range between 1323 K (1050 °C) and 1623 K (1350 °C) to characterize the phase relations of the complex slag systems encountered in lead and zinc blast furnace sinters. The liquidus in two pseudoternary sections ZnO-“Fe₂O₃”-(PbO + CaO + SiO₂) with the CaO/SiO₂ weight ratio of 0.933 and PbO/(CaO + SiO₂) weight ratios of 2.0 and 3.2 have been constructed.     

The N-terminal part of the enzyme component (C2I) of the binary Clostridium botulinum C2 toxin interacts with the binding component C2II and functions as a carrier system for a Rho ADP-ribosylating C3-like fusion toxin

PURPOSE: The major obstacle to successful discordant kidney xenotransplantation is hyperacute rejection (HAR). Complement plays a key role in the induction of HRA, defined by endothelial cell activation, loss of vascular integrity, hemorrhage and thrombosis. The activation of complement is tightly controlled by a number of species-specific regulatory proteins which inhibit, at different points, the cascade of events leading to the formation of the membrane attack complex (MAC). We have tested the hypothesis that kidneys derived from transgenic mice expressing two human complement inhibitors, Decay Accelerating Factor (hDAF) and Membrane Cofactor Protein (MCP), could be protected from human complement-mediated damage. MATERIALS AND METHODS: Control and transgenic mice were perfused with human plasma by cannulation of the right jugular vein, at a perfusion rate of 10 microL./min. for two hours. Complement C3 deposition was detected on kidney sections by immunohistochemistry using specific FITC antibody. Complement-induced tissue damage was evaluated by histopathological examination. RESULTS: Heavy deposition of complement C3 was observed on kidneys derived from perfused control mice. This was associated with a characteristic HAR pathology of severe interstitial hemorrhage, inflammatory reaction, loss of glomerula and tubuli structure. Kidneys derived from mice transgenic for hDAF or hMCP were partially protected from both complement C3 deposition and tissue damage. The expression of both hDAF and hMCP in double transgenic mice significantly increases the protection from human complement-mediated damage. CONCLUSION: A novel model of in vivo perfusion with human plasma has been adopted to recreate the initial event of HAR. Our data show that this murine model could be very valuable to determine the effect of transgenic human molecules in protecting vascularized organs from human complement attack.     

Characterization of phosphotyrosine binding motifs in the cytoplasmic domain of platelet/endothelial cell adhesion molecule-1 (PECAM-1) that are required for the cellular association and activation of the protein-tyrosine phosphatase, SHP-2

Recent studies have shown that the Src homology-2 (SH2) domain-containing protein-tyrosine phosphatase, SHP-2, associates with the cytoplasmic domain of PECAM-1 as it becomes tyrosine-phosphorylated during platelet aggregation: a process that can be mimicked in part by small synthetic phosphopeptides corresponding to the cytoplasmic domain of PECAM-1 encompassing tyrosine residues Tyr-663 or Tyr-686. To further examine the molecular requirements for PECAM-1/SHP-2 interactions, we generated human embryonic kidney (HEK)-293 cell lines that stably expressed mutant forms of PECAM-1 harboring tyrosine to phenylalanine (Tyr --> Phe) mutations in the cytoplasmic domain. Y663F and Y686F forms of PECAM-1 were tyrosine-phosphorylated to a somewhat lesser extent than wild-type PECAM-1, and a doubly substituted Y663,686F form of PECAM-1 failed to become tyrosine-phosphorylated, suggesting that the PECAM-1 cytoplasmic domain tyrosine residues 596, 636 and 701 do not serve as substrates for cellular kinases. Interestingly, SHP-2 binding was lost when either Tyr-663 or Tyr-686 were changed to phenylalanine, indicating that both residues are required for SHP-2/PECAM-1 association. Although PECAM-1 phosphopeptides NSDVQpY663TEVQV and DTETVpY686SEVRK stimulated the catalytic activity of the phosphatase to a similar extent, surface plasmon resonance studies revealed that the Tyr-663-containing peptide had approximately 10-fold higher affinity for SHP-2 than did the Tyr-686 peptide. Finally, peptido-precipitation analysis showed that the NH2-terminal SH2 domain of SHP-2 reacted preferentially with the Tyr-663 PECAM-1 phosphopeptide, while the Tyr-686 phosphopeptide associated only with the COOH-terminal SH2 domain of the phosphatase. Together, these data provide a molecular model for PECAM-1/SHP-2 interactions that may shed light on the downstream events that follow PECAM-1-mediated interactions of vascular cells.     

Experimental study of phase equilibria in the PbO-ZnO-“Fe2O3”-CaO-SiO2 system in air for high lead smelting slags (CaO/SiO2=0.35 and PbO/(CaO + SiO2)=5.0 by weight)

Experimental studies on phase equilibria in the multicomponent system PbO-ZnO-CaO-SiO₂-FeO-Fe₂O₃ in air have been conducted to characterize the phase relations of a complex slag system used in commercial lead oxidation smelting. The liquidus in the pseudo-ternary section ZnO-“Fe₂O₃”-(PbO + CaO + SiO₂) with the CaO/SiO₂ weight ratio of 0.35 and the PbO/(CaO + SiO₂) weight ratio of 5.0 has been constructed using results of over 100 high-temperature equilibration and quenching experiments followed by electron probe X-ray microanalysis. The liquidus in this pseudoternary section contains primary phase fields of spinel (zinc ferrite) Zn _x Fe_3−x O_4+y , zincite Zn _u Fe_1−u O, melilite Pb _v Ca_2−v Zn _w Fe_1−w Si₂O₇, hematite Fe₂O₃, magneto-plumbite PbFe₁₀O₁₆, and dicalcium silicate Ca_2−t Pb _t SiO₄. The laboratory results are compared with the slags obtained from an industrial reactor.