Plasmodium falciparum: Antimalarial activity in culture of sinefungin and other methylation inhibitors

A culture line of Plasmodium falciparum (FCR-3/Gambia) was exposed in vitro for a 2-day period to several analogs of adenosylhomocysteine. Minimal concentrations giving complete inhibition of growth were 0.2 mM for 3-deazaadenosine, 0.2 mM for 5′-deoxy-5′-(isobutylthio)-3-deazaadenosine, and 0.3 μM for sinefungin. The effects of the first two of these compounds were potentiated by homocysteine-thiolactone, suggesting that they were inhibiting methylation reaction(s) indirectly via adenosylhomocysteine hydrolase (EC 3.3.1.1).     

Catecholamine metabolism and in vitro induction of premature cuticle melanization in wild type and pigmentation mutants of Drosophila melanogaster

The major pathway leading to adult cuticle melanization in Drosophila melanogaster has been investigated by a combination of biochemical and genetic approaches. By comparing catecholamine pools in newly emerged flies and in frass (excreta) collected 1 to 4 days after eclosion from wild type with those obtained from several pigmentation mutants, the major flow of catecholamines through the pathway to an unidentified final catabolite was determined. We also demonstrate that incubation with dopamine in vitro induces premature melanization in wild type unpigmented pharate adults several hours before the developmentally programmed onset of melanization, supporting the hypothesis that the availability of catecholamines may be the limiting factor determining the onset of melanization and that the major enzymatic activities that act downstream of dopa decarboxylase in the pathway are deposited into the cuticle before pigmentation begins. In vitro melanization studies with various pigmentation mutants that are associated with critical enzymatic steps in Drosophila catecholamine metabolism are consistent with their proposed function and suggest a central role of N-β-alanyldopamine in adult cuticle pigmentation. © 1996 Wiley-Liss, Inc.     

Currently used in clinical practice beam rate changes have no significant effect on the reduction of clonogenic capacity of PNT1A cells in vitro

BackgroundDue to the lack of selectivity of ionizing radiation between normal and cancer cells, it is important to improve the existing radiation patterns. Lowering the risk of cancer recurrence and comfort during treatment are priorities in radiotherapy.Materials and methodsIn the experiment we used dose verification to determine the irradiation time calculated by a treatment planning system for 6XFFF and 10XFFF beams. Cells cultured under standard conditions were irradiated with a dose of 2 Gy at different beam rates 400 MU/min, 600 MU/min, 800 MU/min, 1000 MU/min, 1400 MU/min, 1600 MU/min and 2400 MU/min using 6XFFF, 10XFFF and 6XFF beams.ResultsThe experiment was aimed at comparing the biological response of normal prostate cells after clinically applied radiation patterns. No statistically significant differences in the cellular response were observed. The wide range of beam rates as well as the beam profiles did not significantly affect cell proliferation.ConclusionsHigh beam rates, without significantly affecting the clonogenic capacity of cells, have an impact on the quality of patient’s treatment. With the increasing beam rate the irradiation time is shortened, which has an important impact on patients’ health. This experiment can have a practical significance.     

Establishing reference intervals for von Willebrand factor multimers

Backgroundvon Willebrand factor (VWF) multimers (VWF:MM) methodologies are technically difficult, laborious, time consuming, non-standardized and results vary between laboratories. A new semi automated VWF:MM assay is available for routine use (Sebia). Due to lack of reference values for VWF:MM fractions, results interpretation can be challenging in some cases. The aim of this study was to determine reference intervals for low molecular weight (LMWM), intermediate molecular weight (IMWM) and high molecular weight (HMWM) multimers.MethodsBy the international cooperation initiated between 4 countries (Estonia, Latvia, France, and USA) 131 samples of relatively healthy individuals were analyzed for VWF:MM (in total 51 males and 80 non-pregnant females aged 17-69 years). Reference intervals were calculated according to CLSI C28-A3 standard.ResultsThe proposed reference intervals for VWF:MM were calculated for LMWM 10.4-22.5%, IMWM 22.6-37.6%, HMWM 45.6-66.6%. Age related differences were seen in IMWM and HMWM (p<0.001 and 0.038). There was no gender related difference observed. Geographically LMWM results of France were different from the other regions (p<0.05).ConclusionsQuantification of VWF:MM fractions, in addition to qualitative assessment of VWF:MM patterns, has the potential to aid in differential diagnosis of von Willebrand disease (VWD) subtypes. The reference values calculated in this study can be used in future research to establish clinical decision limits.     

Mice with a disruption of the imprinted Grb10 gene exhibit altered body composition, glucose homeostasis, and insulin signaling during postnatal life

The Grb10 adapter protein is capable of interacting with a variety of receptor tyrosine kinases, including, notably, the insulin receptor. Biochemical and cell culture experiments have indicated that Grb10 might act as an inhibitor of insulin signaling. We have used mice with a disruption of the Grb10 gene (Grb10Delta2-4 mice) to assess whether Grb10 might influence insulin signaling and glucose homeostasis in vivo. Adult Grb10Delta2-4 mice were found to have improved whole-body glucose tolerance and insulin sensitivity, as well as increased muscle mass and reduced adiposity. Tissue-specific changes in insulin receptor tyrosine phosphorylation were consistent with a model in which Grb10, like the closely related Grb14 adapter protein, prevents specific protein tyrosine phosphatases from accessing phosphorylated tyrosines within the kinase activation loop. Furthermore, insulin-induced IRS-1 tyrosine phosphorylation was enhanced in Grb10Delta2-4 mutant animals, supporting a role for Grb10 in attenuation of signal transmission from the insulin receptor to IRS-1. We have previously shown that Grb10 strongly influences growth of the fetus and placenta. Thus, Grb10 forms a link between fetal growth and glucose-regulated metabolism in postnatal life and is a candidate for involvement in the process of fetal programming of adult metabolic health.     

Evidence of inbreeding depression on human height

Stature is a classical and highly heritable complex trait, with 80%-90% of variation explained by genetic factors. In recent years, genome-wide association studies (GWAS) have successfully identified many common additive variants influencing human height; however, little attention has been given to the potential role of recessive genetic effects. Here, we investigated genome-wide recessive effects by an analysis of inbreeding depression on adult height in over 35,000 people from 21 different population samples. We found a highly significant inverse association between height and genome-wide homozygosity, equivalent to a height reduction of up to 3 cm in the offspring of first cousins compared with the offspring of unrelated individuals, an effect which remained after controlling for the effects of socio-economic status, an important confounder (χ(2) = 83.89, df = 1; p = 5.2 × 10(-20)). There was, however, a high degree of heterogeneity among populations: whereas the direction of the effect was consistent across most population samples, the effect size differed significantly among populations. It is likely that this reflects true biological heterogeneity: whether or not an effect can be observed will depend on both the variance in homozygosity in the population and the chance inheritance of individual recessive genotypes. These results predict that multiple, rare, recessive variants influence human height. Although this exploratory work focuses on height alone, the methodology developed is generally applicable to heritable quantitative traits (QT), paving the way for an investigation into inbreeding effects, and therefore genetic architecture, on a range of QT of biomedical importance.     

Zeolin. A new recombinant storage protein constructed using maize gamma-zein and bean phaseolin

The major seed storage proteins of maize (Zea mays) and bean (Phaseolus vulgaris), zein and phaseolin, accumulate in the endoplasmic reticulum (ER) and in storage vacuoles, respectively. We show here that a chimeric protein composed of phaseolin and 89 amino acids of gamma-zein, including the repeated and the Pro-rich domains, maintains the main characteristics of wild-type gamma-zein: It is insoluble unless its disulfide bonds are reduced and forms ER-located protein bodies. Unlike wild-type phaseolin, the protein, which we called zeolin, accumulates to very high amounts in leaves of transgenic tobacco (Nicotiana tabacum). A relevant proportion of the ER chaperone BiP is associated with zeolin protein bodies in an ATP-sensitive fashion. Pulse-chase labeling confirms the high affinity of BiP to insoluble zeolin but indicates that, unlike structurally defective proteins that also extensively interact with BiP, zeolin is highly stable. We conclude that the gamma-zein portion is sufficient to induce the formation of protein bodies also when fused to another protein. Because the storage proteins of cereals and legumes nutritionally complement each other, zeolin can be used as a starting point to produce nutritionally balanced and highly stable chimeric storage proteins.     

Novel 1,2-dihydroquinazolin-2-ones: Design,synthesis, and biological evaluation against Trypanosoma brucei

In 2014, a published report of the high-throughput screen of >42,000 kinase inhibitors from GlaxoSmithKline against T. brucei identified 797 potent and selective hits. From this rich data set, we selected NEU-0001101 (1) for hit-to-lead optimization. Through our preliminary compound synthesis and SAR studies, we have confirmed the previously reported activity of 1 in a T. brucei cell proliferation assay and have identified alternative groups to replace the pyridyl ring in 1. Pyrazole 24 achieves improvements in both potency and lipophilicity relative to 1, while also showing good in vitro metabolic stability. The SAR developed on 24 provides new directions for further optimization of this novel scaffold for anti-trypanosomal drug discovery.