APOBEC3G genetic variants and their influence on the progression to AIDS

The cytosine deaminase APOBEC3G, in the absence of the human immunodeficiency virus type 1 (HIV-1) accessory gene HIV-1 viral infectivity factor (vif), inhibits viral replication by introducing G-->A hypermutation in the newly synthesized HIV-1 DNA negative strand. We tested the hypothesis that genetic variants of APOBEC3G may modify HIV-1 transmission and disease progression. Single nucleotide polymorphisms were identified in the promoter region (three), introns (two), and exons (two). Genotypes were determined for 3,073 study participants enrolled in six HIV-AIDS prospective cohorts. One codon-changing variant, H186R in exon 4, was polymorphic in African Americans (AA) (f = 37%) and rare in European Americans (f < 3%) or Europeans (f = 5%). For AA, the variant allele 186R was strongly associated with decline in CD4 T cells (CD4 slope on square root scale: -1.86, P = 0.009), The 186R allele was also associated with accelerated progression to AIDS-defining conditions in AA. The in vitro antiviral activity of the 186R enzyme was not inferior to that of the common H186 variant. These studies suggest that there may be a modifying role of variants of APOBEC3G on HIV-1 disease progression that warrants further investigation.     

Identification of Regulators of Polyploidization Presents Therapeutic Targets for Treatment of AMKL

The mechanism by which cells decide to skip mitosis to become polyploid is largely undefined. Here we used a high-content image-based screen to identify small-molecule probes that induce polyploidization of megakaryocytic leukemia cells and serve as perturbagens to help understand this process. Our?study implicates five networks of kinases that?regulate the switch to polyploidy. Moreover, we find that dimethylfasudil (diMF, H-1152P) selectively increased polyploidization, mature cell-surface marker expression, and apoptosis of malignant megakaryocytes. An integrated target identification approach employing proteomic and shRNA screening revealed that a major target of diMF is Aurora kinase A (AURKA). We further find that MLN8237 (Alisertib), a selective inhibitor of AURKA, induced polyploidization and expression of mature megakaryocyte markers in acute megakaryocytic leukemia (AMKL) blasts and displayed potent anti-AMKL activity in?vivo. Our findings provide a rationale to support clinical trials of MLN8237 and other inducers of polyploidization and differentiation in AMKL.     

L-carnosine as an enhancer to computerized cognitive behaviour therapy in Japanese workers,an unjustified claim

This letter comments on the conclusion drawn by Shirotsuki et al. (2017) in their article entitled “The effect for Japanese workers of a self-help computerized cognitive behaviour therapy program with a supplement soft drink”, recently published in BioPsychoSocial Medicine. The authors concluded that their drink, containing L-carnosine, enhances the effects of a computerized cognitive-behavioural therapy (CCBT) on the psychological well-being of healthy Japanese workers. Yet, we argue that their conclusion is unfounded given their results and the methodological shortcomings of their study. Briefly, while the authors reported improvement on the tension-anxiety subscale of the Profile of Mood States (POMS) in the CCBT only group, they also observed a lack of improvement on this subscale in the CCBT+L-carnosine group suggesting that the drink washes out this beneficial effect of CCBT. Methodological issues include the uncontrolled levels of L-carnosine metabolized by participants jeopardize the study’s internal validity. Also, the clinical meaningfulness of the findings seems dubious as post-treatment scores remained within the range of the general Japanese population. Consequently, we argue that Shirotsuki et al.’s study should be re-conducted before drawing any valid conclusion.     

Evolution of Toxoplasma-PCR methods and practices: a French national survey and proposal for technical guidelines

The molecular diagnosis of toxoplasmosis lacks standardisation due to the use of numerous methods with variable performance. This diversity of methods also impairs robust performance comparisons between laboratories. The harmonisation of practices by diffusion of technical guidelines is a useful way to improve these performances. The knowledge of methods and practices used for this molecular diagnosis is an essential step to provide guidelines for Toxoplasma-PCR. In the present study, we aimed (i) to describe the methods and practices of Toxoplasma-PCR used by clinical microbiology laboratories in France and (ii) to propose technical guidelines to improve molecular diagnosis of toxoplasmosis. To do so, a yearly self-administered questionnaire-based survey was undertaken in proficient French laboratories from 2008 to 2015, and guidelines were proposed based on the results of those as well as previously published work. This period saw the progressive abandonment of conventional PCR methods, of Toxoplasma-PCR targeting the B1 gene and of the use of two concomitant molecular methods for this diagnosis. The diversity of practices persisted during the study, in spite of the increasing use of commercial kits such as PCR kits, DNA extraction controls and PCR inhibition controls. We also observed a tendency towards the automation of DNA extraction. The evolution of practices did not always go together with an improvement in those, as reported notably by the declining use of Uracil-DNA Glycosylase to avoid carry-over contamination. We here propose technical recommendations which correspond to items explored during the survey, with respect to DNA extraction, Toxoplasma-PCR and good PCR practices.     

Transfer on hydrophobic substrates and AFM imaging of membrane proteins reconstituted in planar lipid bilayers

The lipid-layer technique allows reconstituting transmembrane proteins at a high density in microns size planar membranes and suspended to a lipid monolayer at the air/water interface. In this paper, we transferred these membranes onto two hydrophobic substrates for further structural analysis of reconstituted proteins by Atomic Force Microscopy (AFM). We used a mica sheet covered by a lipid monolayer or a sheet of highly oriented pyrolytic graphite (HOPG) to trap the lipid monolayer at the interface and the suspended membranes. In both cases, we succeeded in the transfer of large membrane patches containing densely packed or 2D-crystallized proteins. As a proof of concept, we transferred and imaged the soluble Shiga toxin bound to its lipid ligand and the ATP-binding cassette (ABC) transporter BmrA reconstituted into a planar bilayer. AFM imaging with a lateral resolution in the nanometer range was achieved. Potential applications of this technique in structural biology and nanobiotechnology are discussed.     

Variations temporelles de l'ultrastructure de Scenedesmus quadricauda exposée au cuivre lors d'une expérience à long terme

An experiment was conducted with a continuous culture of Scenedesmus quadricauda for 50 days in a copper concentration of 250 μg l⁻¹. Ultrastructural examinations were carried out at different times to obtain the sequence of events. After 1.5 h of exposure, the ultrastructural effects were obvious: the chloroplasts were retracted and all cell structures appeared diffuse. The same observations were reported after 3, 6, 12 and 24 hours. Between the 3rd and 15th days of the experiment, the cells showed major accumulations of starch, lipid and electron-dense inclusions in the vacuoles, and the cell walls became thicker. After 22,32 and 50 days the cells appeared normal, but the cytoplasm was densely packed with ribosomes and the endoplasmic reticulum was evident in most cells. However, atomic absorption analysis for copper showed that there was no bio-accumulation after 50 days of exposure. These findings suggest the implication of a second tolerance mechanism to copper (possibly extracellular complexation or removal) that is activated after long exposure, in contrast to electron-dense inclusions, which appear after a relatively short time (2 days).     

QTL mapping for traits associated with stress neuroendocrine reactivity in rats

In the present study we searched for quantitative trait loci (QTLs) that affect neuroendocrine stress responses in a 20-min restraint stress paradigm using Brown–Norway (BN) and Wistar–Kyoto–Hyperactive (WKHA) rats. These strains differed in their hypothalamic–pituitary–adrenal axis (plasma ACTH and corticosterone levels, thymus, and adrenal weights) and in their renin–angiotensin–aldosterone system reactivity (plasma renin activity, aldosterone concentration). We performed a whole-genome scan on a F₂ progeny derived from a WKHA × BN intercross, which led to the identification of several QTLs linked to plasma renin activity (Sr6, Sr8, Sr11, and Sr12 on chromosomes RNO2, 3, 19, and 8, respectively), plasma aldosterone concentration (Sr7 and Sr9 on RNO2 and 5, respectively), and thymus weight (Sr10, Sr13, and Srl4 on RNO5, 10, and 16, respectively). The type 1b angiotensin II receptor gene (Agtrlb) maps within the confidence intervals of QTLs on RNO2 linked to plasma renin activity (Sr6, highly significant; LOD = 5.0) and to plasma aldosterone level (Sr7, suggestive; LOD = 2.0). In vitro studies of angiotensin II–induced release of aldosterone by adrenal glomerulosa cells revealed a lower receptor potency (log EC₅₀ = −8.16 ± 0.11 M) and efficiency (E_max = 453.3 ± 25.9 pg/3 × 10⁴ cells/24 h) in BN than in WKHA (log EC₅₀ = −10.66 ± 0.18 M; E_max = 573.1 ± 15.3 pg/3 × 10⁴ cells/24 h). Moreover, differences in Agtr1b mRNA abundance and sequence reinforce the putative role of the Agtr1b gene in the differential plasma renin stress reactivity between the two rat strains.     

Differential pattern of Cu/Zn superoxide dismutase isoforms in relation to tidal spatio-temporal changes in the blue mussel Mytilus edulis

Inducible antioxidant defences in marine organisms such as mussel bivalves are commonly used as biomarkers of pollutant-induced oxidative stress and their variations proposed as one of the biological effect measurements for assessment of contamination impact in aquatic environments. Among them, the copper/zinc superoxide dismutases (Cu/Zn-SODs) are metalloenzymes which play a key role in the protection of cells in case of oxidative stress. In order to observe possible variations of an antioxidant response in relation to tidal oscillations, the copper/zinc superoxide dismutase activity (Cu/Zn-SOD) was characterized in the digestive gland and gills of blue mussels sampled at high and low shore throughout the tidal cycle. Determination of SOD activity was performed on gels after isoelectro-focusing, allowing the revelation of three isoforms. In both tissues, high-shore mussels exhibited a higher level of total SOD activity than low-shore mussels. During emersion, a decrease of total SOD activity appeared in digestive gland for both groups. In high-shore mussels, the less acidic form contributed to 75% of the total activity, the second one to 20% and the more acidic one to 5% in both tissues before air exposure. During emersion, the relative contribution of the three isoforms to the total activity was markedly changed with a significant decrease in intensity of the first isoform and parallel increases in the two other ones. After re-immersion a progressive recovery of proportions of SOD isoforms was observed. In low-shore mussels, the relative contribution of the three isoforms to the total SOD activity showed similar changes. The observed variations could correspond to changes in the redox status of the mussels during tidal oscillations.