Killing of Escherichia coli by Crohn's Disease Monocyte-derived Macrophages and Its Enhancement by Hydroxychloroquine and Vitamin D

BACKGROUND: Crohn's disease (CD) is associated with defective innate immunity, including impaired neutrophil chemotaxis, and mucosal invasion by bacteria, particularly adherent and invasive Escherichia coli that replicate inside macrophage phagolysosomes. We compared CD and healthy control (HC) macrophages for their abilities to kill E. coli and generate neutrophil chemoattractants and also assessed the effects of hydroxychloroquine (HCQ) and vitamin D on killing of phagocytosed E. coli. METHODS: Peripheral blood monocyte-derived macrophages from CD and HC were compared for bacterial killing and generation of neutrophil chemoattractants in response to CD-derived E. coli. Escherichia coli replication was also assessed in the presence and absence of HCQ, alone and with antibiotics, and vitamin D. RESULTS: Monocyte-derived macrophages from patients with CD were similar to HC in allowing replication of phagocytosed CD-derived E. coli: HM605 {CD: N = 10, mean fold replication in 3 hr = 1.08 (95% confidence interval [CI], 0.39–1.78); HC: N = 9, 1.50 (95% CI, 1.02–1.97); P = 0.15} and also in generation of neutrophil chemoattractants in response to E. coli (mean fold chemotaxis relative to control: CD = 2.55 [95% CI, 2.31–2.80]; HC = 2.65 [95% CI, 2.46–2.85], P = 0.42). HCQ and 1,25 OH(2)-vitamin D(3) both caused dose-dependent inhibition of intramacrophage E. coli replication 3-hour postinfection; HCQ: 73.9% inhibition (P < 0.001) at 1 μg/mL, accompanied by raised intraphagosomal pH, and 1,25 OH(2)-vitamin D(3): 80.7% inhibition (P < 0.05) at 80 nM. HCQ had synergistic effects with doxycycline and ciprofloxacin. CONCLUSIONS: CD and HC macrophages perform similarly in allowing replication of phagocytosed E. coli and generating neutrophil chemoattractants. Replication of phagocytosed E. coli was substantially decreased by HCQ and vitamin D. These warrant further therapeutic trials in CD in combination with relevant antibiotics

One-step zero-background IgG reformatting of phage-displayed antibody fragments enabling rapid and high-throughput lead identification

 A superconducting pair in high-temperature superconductivity structure was supposed to have an electrical quadrupole moment. It was shown, that under certain conditions, at the part of acoustic phonons, conduction electrons in high-temperature superconductivity structure can form superconducting pairs, when the interaction energy of the quadrupole moment of a superconducting pair with the structure electric field exceeds the energy of Coulomb repulsion between electrons. The electrical field parameters in a CuO2 plane of high-temperature superconductivity structure have been obtained by the method of effective point charges. The distance between electrons in superconducting pair in a CuO2 plane have been estimated in ~ 75 ÷ 130 Å. Within the framework of the suggested model it was shown that high-temperature superconductivity IR-spectra, which were observed at 500 cm-1, can be caused by the quadrupole resonance of superconducting pairs. Выдвинуто предположение, что электрический квадрупольный момент куперовской пары в структуре высокотемпературных сверхпроводников отличен от нуля. Показано, что при определенных условиях при участии акустических фононов электроны проводимости в структуре высокотемпературных сверхпроводников могут образовывать куперовские пары, когда энергия взаимодействия квадрупольного момента куперовской пары превышает энергию кулоновского отталкивания между электронами. Параметры электрического поля в плоскости CuO2 в высокотемпературных сверхпроводниках рассчитаны методом эффективных точечных зарядов. Расстояние между электронами в куперовской паре в плоскости CuO2 равно ~ 75 ÷ 130 Å. В соответствии с предлагаемой моделью инфракрасные спектры, наблюдаемые в районе 500 см-1 для высокотемпературных сверхпроводников, могут быть обусловлены квадрупольным резонансом куперовских пар. Висунуто припущення, що електричний квадрупольний момент куперівської пари в структурі високо-температурних надпровідників відмінний від нуля. Показано, що за певних умов за участю акустичних фононів електрони провідності в структурі високотемпературних надпровідників можуть утворювати куперівські пари, коли енергія взаємодії квадрупольного моменту куперівської пари перевищує енергію кулонівського відштовхування між електронами. Параметри електричного поля в площині CuО2 у високотемпературних надпровідниках розраховані методом ефективних точкових зарядів. Відстань між електронами в куперівській парі в площині CuО2 становить 75–130 Å. Відповідно до запропонованої моделі показано, що інфрачервоні спектри, які спостерігаються в районі 500 см-1 для високотемпера-турних надпровідників, можуть бути обумовлені квадрупольним резонансом куперівських пар

Primary structure of ovine pituitary basic fibroblast growth factor

AbstractThe complete amino acid sequence of basic FGF (146 residues) from ovine pituitary glands has been established. This has been achieved by the sequence analysis of subnanomole amounts of the intact molecule and of peptides derived by enzymatic digestions with clostripain, chymotrypsin, pepsin and Staphylococcus aureus V8 protease. Microbore HPLC, employing 1–2 mm i.d. columns, was used to purify, concentrate and buffer-exchange the FGF peptides. A novel application of ion-pairing chromatography was employed to isolate peptides which were not retained on conventional reversed-phase systems. There is only one positional difference between the ovine and bovine basic FGFs, but there are 3 positional differences between ovine and human basic FGFS

An optimized hepatitis C virus E2 glycoprotein core adopts a functional homodimer that efficiently blocks virus entry

The hepatitis C virus (HCV) envelope glycoprotein E2 is the major target of broadly neutralizing antibodies in vivo and is the focus of efforts in the rational design of a universal B cell vaccine against HCV. The E2 glycoprotein exhibits a high degree of amino acid variability which localizes to three discrete regions: hypervariable region 1 (HVR1), hypervariable region 2 (HVR2), and the intergenotypic variable region (igVR). All three variable regions contribute to immune evasion and/or isolate-specific structural variations, both important considerations for vaccine design. A high-resolution structural definition of the intact HCV envelope glycoprotein complex containing E1 and E2 remains to be elucidated, while crystallographic structures of a recombinant E2 ectodomain failed to resolve HVR1, HVR2, and a major neutralization determinant adjacent to HVR1. To obtain further information on E2, we characterized the role of all three variable regions in E2 ectodomain folding and function in the context of a recombinant ectodomain fragment (rE2). We report that removal of the variable regions accelerates binding to the major host cell receptor CD81 and that simultaneous deletion of HVR2 and the igVR is required to maintain wild-type CD81-binding characteristics. The removal of the variable regions also rescued the ability of rE2 to form a functional homodimer. We propose that the rE2 core provides novel insights into the role of the variable motifs in the higher-order assembly of the E2 ectodomain and may have implications for E1E2 structure on the virion surface. IMPORTANCE Hepatitis C virus (HCV) infection affects ∼2% of the population globally, and no vaccine is available. HCV is a highly variable virus, and understanding the presentation of key antigenic sites at the virion surface is important for the design of a universal vaccine. This study investigates the role of three surface-exposed variable regions in E2 glycoprotein folding and function in the context of a recombinant soluble ectodomain. Our data demonstrate the variable motifs modulate binding of the E2 ectodomain to the major host cell receptor CD81 and have an impact on the formation of an E2 homodimer with high-affinity binding to CD81