Synthesis of isoindole and benzoisoindole derivatives of teicoplanin pseudoaglycon with remarkable antibacterial and antiviral activities

The primary amino function of teicoplanin pseudoaglycon has been transformed into arylthioisoindole or benzoisoindole and glycosylthioisoindole derivatives, in a reaction with o-phthalaldehyde or naphtalene-2,3-dicarbaldehyde and various thiols. All of the obtained semisynthetic antibiotics exhibited potent antibacterial activities against Gram-positive bacteria in the ng per ml concentration range. A few of them showed antiviral activity, in particular against influenza virus.     

TGFβ Activated Kinase 1 (TAK1) at the Crossroad of B Cell Receptor and Toll-Like Receptor 9 Signaling Pathways in Human B Cells

B cell development and activation are regulated by combined signals mediated by the B cell receptor (BCR), receptors for the B-cell activating factor of the tumor necrosis factor family (BAFF-R) and the innate receptor, Toll-like receptor 9 (TLR9). However, the underlying mechanisms by which these signals cooperate in human B cells remain unclear. Our aim was to elucidate the key signaling molecules at the crossroads of BCR, BAFF-R and TLR9 mediated pathways and to follow the functional consequences of costimulation.Therefore we stimulated purified human B cells by combinations of anti-Ig, B-cell activating factor of the tumor necrosis factor family (BAFF) and the TLR9 agonist, CpG oligodeoxynucleotide. Phosphorylation status of various signaling molecules, B cell proliferation, cytokine secretion, plasma blast generation and the frequency of IgG producing cells were investigated. We have found that BCR induced signals cooperate with BAFF-R- and TLR9-mediated signals at different levels of cell activation. BCR and BAFF- as well as TLR9 and BAFF-mediated signals cooperate at NFκB activation, while BCR and TLR9 synergistically costimulate mitogen activated protein kinases (MAPKs), ERK, JNK and p38. We show here for the first time that the MAP3K7 (TGF beta activated kinase, TAK1) is responsible for the synergistic costimulation of B cells by BCR and TLR9, resulting in an enhanced cell proliferation, plasma blast generation, cytokine and antibody production. Specific inhibitor of TAK1 as well as knocking down TAK1 by siRNA abrogates the synergistic signals. We conclude that TAK1 is a key regulator of receptor crosstalk between BCR and TLR9, thus plays a critical role in B cell development and activation.     

Fluorescent ligands for adenosine receptors

Interest is increasing in developing fluorescent ligands for characterization of adenosine receptors (ARs), which hold a promise of usefulness in the drug discovery process. The size of a strategically labeled AR ligand can be greatly increased after the attachment of a fluorophore. The choice of dye moiety (e.g. Alexa Fluor 488), attachment point and linker length can alter the selectivity and potency of the parent molecule. Fluorescent derivatives of adenosine agonists and antagonists (e.g. XAC and other heterocyclic antagonist scaffolds) have been synthesized and characterized pharmacologically. Some are useful AR probes for flow cytometry, fluorescence correlation spectroscopy, fluorescence microscopy, fluorescence polarization, fluorescence resonance energy transfer, and scanning confocal microscopy. Thus, the approach of fluorescent labeled GPCR ligands, including those for ARs, is a growing dynamic research field.     

Limited Promiscuity of HLA-DRB1 Presented Peptides Derived of Blood Coagulation Factor VIII

The formation of inhibitory antibodies directed against coagulation factor VIII (FVIII) is a severe complication in the treatment of hemophilia A patients. The induction of anti-FVIII antibodies is a CD4⁺ T cell-dependent process. Activation of FVIII-specific CD4⁺ T cells is dependent on the presentation of FVIII-derived peptides on MHC class II by antigen-presenting cells. Previously, we have shown that FVIII-pulsed human monocyte-derived dendritic cells can present peptides from several FVIII domains. In this study we show that FVIII peptides are presented on immature as well as mature dendritic cells. In immature dendritic cells half of the FVIII-loaded MHC class II molecules are retained within the cell, whereas in LPS-matured dendritic cells the majority of MHC class II/peptide complexes is present on the plasma membrane. Time-course studies revealed that presentation of FVIII-derived peptides was optimal between 12 and 24 hours after maturation but persisted for at least 96 hours. We also show that macrophages are able to internalize FVIII as efficiently as dendritic cells, however FVIII was presented on MHC class II with a lower efficiency and with different epitopes compared to dendritic cells. In total, 48 FVIII core-peptides were identified using a DCs derived of 8 different donors. Five HLA-promiscuous FVIII peptide regions were found – these were presented by at least 4 out of 8 donors. The remaining 42 peptide core regions in FVIII were presented by DCs derived from a single (30 peptides) or two to three donors (12 peptides). Overall, our findings show that a broad repertoire of FVIII peptides can be presented on HLA-DR.     

FAD oxidizes the ERO1-PDI electron transfer chain: the role of membrane integrity

The molecular steps of the electron transfer in the endoplasmic reticulum from the secreted proteins during their oxidation are relatively unknown. We present here that flavine adenine dinucleotide (FAD) is a powerful oxidizer of the oxidoreductase system, Ero1 and PDI, besides the proteins of rat liver microsomes and HepG2 hepatoma cells. Inhibition of FAD transport hindered the action of FAD. Microsomal membrane integrity was mandatory for all FAD-related oxidation steps downstream of Ero1. The PDI inhibitor bacitracin could inhibit FAD-mediated oxidation of microsomal proteins and PDI, but did not hinder the FAD-driven oxidation of Ero1. Our data demonstrated that Ero1 can utilize FAD as an electron acceptor and that FAD-driven protein oxidation goes through the Ero1-PDI pathway and requires the integrity of the endoplasmic reticulum membrane. Our findings prompt further studies to elucidate the membrane-dependent steps of PDI oxidation and the role of FAD in redox folding.     

HPLC study on Fenton-reaction initiated oxidation of salicylic acid. Biological relevance of the reaction in intestinal biotransformation of salicylic acid

Abstract

Fenton-reaction initiated in vitro oxidation and in vivo oxidative biotransformation of salicylic acid was investigated by HPLC-UV-Vis method. By means of the developed high performance liquid chromatography (HPLC) method salicylic acid, catechol, and all the possible monohydroxylated derivatives of salicylic acid can be separated. Fenton oxidations were performed in acidic medium (pH 3.0) with two reagent molar ratios: (1) salicylic acid: iron: hydrogen peroxide 1:3:1 and (2) 1:0.3:1. The incubation samples were analysed at different time points of the reactions. The biological effect of elevated reactive oxygen species concentration on the intestinal metabolism of salicylic acid was investigated by an experimental diabetic rat model. HPLC-MS analysis of the in vitro samples revealed presence of 2,3- and 2,5-dihydroxybenzoic acids. The results give evidence for nonenzyme catalysed intestinal hydroxylation of xenobiotics.     

Maternal compensation for hatching asynchrony in the collared flycatcher Ficedula albicollis

Several hypotheses have been proposed to explain the adaptiveness of hatching asynchrony for the parents, but delayed hatching is generally detrimental for the late hatched young. These offspring often experience competitive disadvantage and delayed development. If hatching asynchrony has a reason other than producing competitive differences among offspring, it would be advantageous, not only for the offspring but even for the parents, to compensate for its detrimental effects. In some species, increasing investment into later laid eggs has been reported and discussed as a compensation mechanism, but its effect on nestling growth and fledging size has not been examined in details. In this study we investigated nestling growth and size at fledging in terms of body mass and length of primaries in relation to the accurate laying and hatching order in collared flycatcher Ficedula albicollis broods. We found that females laid larger eggs at the end of the laying sequence, and this helped to decrease the disadvantages for the last offspring. The last offspring had lower body mass growth rate and fledged with shorter feathers, but in both cases the larger the last egg was, the smaller the lag of the offspring was. We conclude, that even if females were not able to fully compensate for the detrimental effects of hatching asynchrony, larger eggs may improve the survival prospects of late hatched nestlings.     

Comparison of Predictive Immunohistochemical Marker Expression of Primary Breast Cancer and Paired Distant Metastasis using Surgical Material: A Practice-Based Study

Parallel studies of primary breast carcinomas and corresponding distant metastases samples reveal considerable differences. Our aim was to highlight this issue from another perspective and provide further data based on 98 patient samples: 69 primary breast carcinoma and 85 distant metastases from bone, central nervous system (CNS) and lung (56 paired). Two independent series of immunohistochemical reactions with different antibodies for estrogen receptor (ER), progesterone receptor (PgR) and human epidermal growth factor receptor 2 (Her2), along with HER2 fluroscence in situ hybridization (FISH) were performed on tissue microarrays to classify breast carcinoma and distant metastases samples into Luminal A, Luminal B-proliferating, Luminal B-HER2+, HER2+ and triple negative (TNBC) surrogate breast cancer groups. Correlation and agreement between the two assessments of ER and PgR were fair-to-moderate, and almost perfect for HER2 and Ki67. There was 40% discordance concerning immunophenotype between breast carcinomas and distant metastases. Most common metastatic site of ER+ breast carcinoma was the skeletal system (59.2%), whereas that of TNBCs was the CNS (58.8%) and lungs (23.5%). Distant metastases in bones were mostly luminal (54.3%), in the CNS, Luminal B (53.2%), and in the lung, TNBC (37.5%). The change of drugable properties of primary breast cancers in the respective bone and CNS metastases suggests that characterization of the metastasis is necessary for appropriate treatment planning.     

Temperature Dependence of Soil Respiration Modulated by Thresholds in Soil Water Availability Across European Shrubland Ecosystems

Soil respiration (SR) is a major component of the global carbon cycle and plays a fundamental role in ecosystem feedback to climate change. Empirical modelling is an essential tool for predicting ecosystem responses to environmental change, and also provides important data for calibrating and corroborating process-based models. In this study, we evaluated the performance of three empirical temperature–SR response functions (exponential, Lloyd–Taylor and Gaussian) at seven shrublands located within three climatic regions (Atlantic, Mediterranean and Continental) across Europe. We investigated the performance of SR models by including the interaction between soil moisture and soil temperature. We found that the best fit for the temperature functions depended on the site-specific climatic conditions. Including soil moisture, we identified thresholds in the three different response functions that improved the model fit in all cases. The direct soil moisture effect on SR, however, was weak at the annual time scale. We conclude that the exponential soil temperature function may only be a good predictor for SR in a narrow temperature range, and that extrapolating predictions for future climate based on this function should be treated with caution as modelled outputs may underestimate SR. The addition of soil moisture thresholds improved the model fit at all sites, but had a far greater ecological significance in the wet Atlantic shrubland where a fundamental change in the soil CO₂ efflux would likely have an impact on the whole carbon budget.