Pharmacological screening using an FXN-EGFP cellular genomic reporter assay for the therapy of Friedreich ataxia

Copyright @ 2013 Li et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Friedreich ataxia (FRDA) is an autosomal recessive disorder characterized by neurodegeneration and cardiomyopathy. The presence of a GAA trinucleotide repeat expansion in the first intron of the FXN gene results in the inhibition of gene expression and an insufficiency of the mitochondrial protein frataxin. There is a correlation between expansion length, the amount of residual frataxin and the severity of disease. As the coding sequence is unaltered, pharmacological up-regulation of FXN expression may restore frataxin to therapeutic levels. To facilitate screening of compounds that modulate FXN expression in a physiologically relevant manner, we established a cellular genomic reporter assay consisting of a stable human cell line containing an FXN-EGFP fusion construct, in which the EGFP gene is fused in-frame with the entire normal human FXN gene present on a BAC clone. The cell line was used to establish a fluorometric cellular assay for use in high throughput screening (HTS) procedures. A small chemical library containing FDA-approved compounds and natural extracts was screened and analyzed. Compound hits identified by HTS were further evaluated by flow cytometry in the cellular genomic reporter assay. The effects on FXN mRNA and frataxin protein levels were measured in lymphoblast and fibroblast cell lines derived from individuals with FRDA and in a humanized GAA repeat expansion mouse model of FRDA. Compounds that were established to increase FXN gene expression and frataxin levels included several anti-cancer agents, the iron-chelator deferiprone and the phytoalexin resveratrol.Muscular Dystrophy Association (USA), the National Health and Medical Research Council (Australia), the Friedreich’s Ataxia Research Alliance (USA), the Brockhoff Foundation (Australia), the Friedreich Ataxia Research Association (Australasia), Seek A Miracle (USA) and the Victorian Government’s Operational Infrastructure Support Program

Rituximab for Children with Immune Thrombocytopenia: A Systematic Review

BACKGROUND: Rituximab has been widely used off-label as a second line treatment for children with immune thrombocytopenia (ITP). However, its role in the management of pediatric ITP requires clarification. To understand and interpret the available evidence, we conducted a systematic review to assess the efficacy and safety of rituximab for children with ITP. METHODOLOGY/PRINCIPAL FINDINGS: We searched MEDLINE, EMBASE, Cochrane Library, CBM, CNKI, abstract databases of American Society of Hematology, American Society of Clinical Oncology and Pediatric Academic Society. Clinical studies published in full text or abstract only in any language that met predefined inclusion criteria were eligible. Efficacy analysis was restricted to studies enrolling 5 or more patients. Safety was evaluated from all studies that reported data of toxicity. 14 studies (323 patients) were included for efficacy assessment in children with primary ITP. The pooled complete response (platelet count ≥ 100 × 10(9)/L) and response (platelet count ≥ 30 × 10(9)/L) rate after rituximab treatment were 39% (95% CI, 30% to 49%) and 68% (95%CI, 58% to 77%), respectively, with median response duration of 12.8 month. 4 studies (29 patients) were included for efficacy assessment in children with secondary ITP. 11 (64.7%) of 17 patients associated with Evans syndrome achieved response. All 6 patients with systemic lupus erythematosus associated ITP and all 6 patients with autoimmune lymphoproliferative syndrome associated ITP achieved response. 91 patients experienced 108 adverse events associated with rituximab, among that, 91 (84.3%) were mild to moderate, and no death was reported. CONCLUSIONS/SIGNIFICANCE: Randomized controlled studies on effect of rituximab for children with ITP are urgently needed, although a series of uncontrolled studies found that rituximab resulted in a good platelet count response both in children with primary and children secondary ITP. Most adverse events associated with rituximab were mild to moderate, and no death was reported

Enhanced upper genital tract pathologies by blocking Tim-3 and PD-L1 signaling pathways in mice intravaginally infected with Chlamydia muridarum

<p>Abstract</p> <p>Background</p> <p>Although Tim-3 & PD-L1 signaling pathways play important roles in negatively regulating immune responses, their roles in chlamydial infection have not been evaluated.</p> <p>Methods</p> <p>Neutralization antibodies targeting Tim-3 and PD-L1 were used to treat mice. Following an intravaginal infection with <it>C. muridarum </it>organisms, mice with or without the dual antibody treatment were compared for live chlamydial organism shedding from the lower genital tract and inflammatory pathology in the upper genital tract.</p> <p>Results</p> <p>Mice treated with anti-Tim-3 and anti-PD-L1 antibodies displayed a time course of live organism shedding similar to that of mice treated with equivalent amounts of isotype-matched IgG molecules. The combined antibody blocking failed to alter either the lower genital tract cytokine or systemic humoral and cellular adaptive responses to <it>C. muridarum </it>infection. However, the antibody blocking significantly enhanced <it>C. muridarum</it>-induced pathologies in the upper genital tract, including more significant hydrosalpinx and inflammatory infiltration in uterine horn and oviduct tissues.</p> <p>Conclusions</p> <p>The Tim-3 and PD-L1-mediated signaling can significantly reduce pathologies in the upper genital tract without suppressing immunity against chlamydial infection, suggesting that Tim-3 and PD-L1-mediated negative regulation may be manipulated to attenuate tubal pathologies in women persistently infected with <it>C. trachomatis </it>organisms.</p

Low loss coatings for the VIRGO large mirrors

présentée par L. PinardThe goal of the VIRGO program is to build a giant Michelson type interferometer (3 kilometer long arms) to detect gravitational waves. Large optical components (350 mm in diameter), having extremely low loss at 1064 nm, are needed. Today, the Ion beam Sputtering is the only deposition technique able to produce optical components with such performances. Consequently, a large ion beam sputtering deposition system was built to coat large optics up to 700 mm in diameter. The performances of this coater are described in term of layer uniformity on large scale and optical losses (absorption and scattering characterization). The VIRGO interferometer needs six main mirrors. The first set was ready in June 2002 and its installation is in progress on the VIRGO site (Italy). The optical performances of this first set are discussed. The requirements at 1064 nm are all satisfied. Indeed, the absorption level is close to 1 ppm (part per million), the scattering is lower than 5 ppm and the R.M.S. wavefront of these optics is lower than 8 nm on 150 mm in diameter. Finally, some solutions are proposed to further improve these performances, especially the absorption level (lower than 0.1 ppm) and the mechanical quality factor Q of the mirrors (thermal noise reduction)

New Insights into the Organization, Recombination, Expression and Functional Mechanism of Low Molecular Weight Glutenin Subunit Genes in Bread Wheat

The bread-making quality of wheat is strongly influenced by multiple low molecular weight glutenin subunit (LMW-GS) proteins expressed in the seeds. However, the organization, recombination and expression of LMW-GS genes and their functional mechanism in bread-making are not well understood. Here we report a systematic molecular analysis of LMW-GS genes located at the orthologous Glu-3 loci (Glu-A3, B3 and D3) of bread wheat using complementary approaches (genome wide characterization of gene members, expression profiling, proteomic analysis). Fourteen unique LMW-GS genes were identified for Xiaoyan 54 (with superior bread-making quality). Molecular mapping and recombination analyses revealed that the three Glu-3 loci of Xiaoyan 54 harbored dissimilar numbers of LMW-GS genes and covered different genetic distances. The number of expressed LMW-GS in the seeds was higher in Xiaoyan 54 than in Jing 411 (with relatively poor bread-making quality). This correlated with the finding of higher numbers of active LMW-GS genes at the A3 and D3 loci in Xiaoyan 54. Association analysis using recombinant inbred lines suggested that positive interactions, conferred by genetic combinations of the Glu-3 locus alleles with more numerous active LMW-GS genes, were generally important for the recombinant progenies to attain high Zeleny sedimentation value (ZSV), an important indicator of bread-making quality. A higher number of active LMW-GS genes tended to lead to a more elevated ZSV, although this tendency was influenced by genetic background. This work provides substantial new insights into the genomic organization and expression of LMW-GS genes, and molecular genetic evidence suggesting that these genes contribute quantitatively to bread-making quality in hexaploid wheat. Our analysis also indicates that selection for high numbers of active LMW-GS genes can be used for improvement of bread-making quality in wheat breeding

Plasma mid-regional pro-atrial natriuretic peptide and N-terminal pro-brain natriuretic peptide improve discrimination of lone atrial fibrillation

10.1016/j.ijcard.2015.03.415International Journal of Cardiology18810-1

Association between childhood trauma and risk for obesity: a putative neurocognitive developmental pathway.

BACKGROUND: Childhood trauma increases the risk for adult obesity through multiple complex pathways, and the neural substrates are yet to be determined. METHODS: Participants from three population-based neuroimaging cohorts, including the IMAGEN cohort, the UK Biobank (UKB), and the Human Connectome Project (HCP), were recruited. Voxel-based morphometry analysis of both childhood trauma and body mass index (BMI) was performed in the longitudinal IMAGEN cohort; validation of the findings was performed in the UKB. White-matter connectivity analysis was conducted to study the structural connectivity between the identified brain region and subdivisions of the hypothalamus in the HCP. RESULTS: In IMAGEN, a smaller frontopolar cortex (FPC) was associated with both childhood abuse (CA) (β = - .568, 95%CI - .942 to - .194; p = .003) and higher BMI (β = - .086, 95%CI - .128 to - .043; p < .001) in male participants, and these findings were validated in UKB. Across seven data collection sites, a stronger negative CA-FPC association was correlated with a higher positive CA-BMI association (β = - 1.033, 95%CI - 1.762 to - .305; p = .015). Using 7-T diffusion tensor imaging data (n = 156), we found that FPC was the third most connected cortical area with the hypothalamus, especially the lateral hypothalamus. A smaller FPC at age 14 contributed to higher BMI at age 19 in those male participants with a history of CA, and the CA-FPC interaction enabled a model at age 14 to account for some future weight gain during a 5-year follow-up (variance explained 5.8%). CONCLUSIONS: The findings highlight that a malfunctioning, top-down cognitive or behavioral control system, independent of genetic predisposition, putatively contributes to excessive weight gain in a particularly vulnerable population, and may inform treatment approaches