A preclinical model of TB meningitis to determine drug penetration and activity at the sites of disease.

Tuberculosis meningitis (TBM) is essentially treated with the first-line regimen used against pulmonary tuberculosis, with a prolonged continuation phase. However, clinical outcomes are poor in comparison, for reasons that are only partially understood, highlighting the need for improved preclinical tools to measure drug distribution and activity at the site of disease. A predictive animal model of TBM would also be of great value to prioritize promising drug regimens to be tested in clinical trials, given the healthy state of the development pipeline for the first time in decades. Here, we report the optimization of a rabbit model of TBM disease induced via inoculation of Mycobacterium tuberculosis into the cisterna magna, recapitulating features typical of clinical TBM: neurological deterioration within months post-infection, acid-fast bacilli in necrotic lesions in the brain and spinal cord, and elevated lactate levels in cerebrospinal fluid (CSF). None of the infected rabbits recovered or controlled the disease. We used young adult rabbits, the size of which allows for spatial drug quantitation in critical compartments of the central nervous system that cannot be collected in clinical studies. To illustrate the translational value of the model, we report the penetration of linezolid from plasma into the CSF, meninges, anatomically distinct brain areas, cervical spine, and lumbar spine. Across animals, we measured the bacterial burden concomitant with neurological deterioration, offering a useful readout for drug efficacy studies. The model thus forms the basis for building a preclinical platform to identify improved regimens and inform clinical trial design

Dynamic phenotypic heterogeneity and the evolution of multiple RNA subtypes in Hepatocellular Carcinoma: the PLANET study

Intra-tumor heterogeneity (ITH) is a key challenge in cancer treatment, but previous studies have focused mainly on the genomic alterations without exploring phenotypic (transcriptomic and immune) heterogeneity. Using one of the largest prospective surgical cohorts for Hepatocellular Carcinoma (HCC) with multi-region sampling, we sequenced whole genomes and paired transcriptomes from 67 HCC patients (331 samples). We found that while genomic ITH was rather constant across TNM stages, phenotypic ITH had a very different trajectory and quickly diversified in stage II patients. Most strikingly, 30% patients were found to contain more than one transcriptomic subtype within a single tumor. Such phenotypic ITH was found to be much more informative in predicting patient survival than genomic ITH and explains the poor efficacy of single-target systemic therapies in HCC. Taken together, we not only revealed an unprecedentedly dynamic landscape of phenotypic heterogeneity in HCC, but also highlighted the importance of studying phenotypic evolution across cancer types

Effect of troglitazone on tumor growth and pulmonary metastasis development of the mouse osteosarcoma cell line LM8

<p>Abstract</p> <p>Background</p> <p>Osteosarcoma often develops micrometastases in the lung prior to diagnosis, causing a fatal outcome. Therefore, the prevention of pulmonary metastases is critical for the improvement of the prognosis of patients with osteosarcoma. The purpose of this study was to investigate whether troglitazone (TGZ) is considered as possible therapeutics in the treatment of growth and metastasis of osteosarcoma.</p> <p>Methods</p> <p>LM8 cells were treated for 3 days with various concentrations of TGZ. The effect of TGZ on cell proliferation was determined by DNA measurement in the cultures and 5-bromo-2'-deoxyuridine incorporation study. The assay of cell invasion and motility was performed using either the Matrigel-coated cell culture inserts or the uncoated cell culture inserts in the invasion chambers. The effect of TGZ on Akt signaling was assessed by Western blot analysis of Akt and p-Akt. The effects of oral administration of either TGZ (TGZ group) or ethanol (control group) on the growth of primary tumor and the development of pulmonary metastasis were examined in nude mice implanted with LM8 cells on their backs. The expression and activity of matrix metalloproteinase 2 (MMP-2) within the tumor were determined by immunohistochemistry and zymography. The microvessel density (MVD) within the tumor was determined by immunohistochemistry for CD34.</p> <p>Results</p> <p>TGZ dose-dependently inhibits cell proliferation. TGZ-treated cells were less invasive and less motile than untreated cells. The activity of MMP-2 secreted by TGZ-treated cells was lower than that secreted by untreated cells. TGZ decreased the level of p-Akt. The primary tumor mass was smaller in the TGZ group than in the control group. The TGZ group had less metastatic tumors in the lung compared with the control group. The expression and activity of MMP-2 within the tumor of the TGZ group were lower than those of the control group. The MVD within the tumor of the TGZ group was lower than that of the control group.</p> <p>Conclusions</p> <p>Inhibition of Akt signaling by TGZ may decrease the secretion of MMP-2, resulting in the decrease of invasiveness and motility in LM8 cells. Treatment of tumor-bearing mice with TGZ decreases the expression and activity of MMP-2 within the tumor, and inhibits primary tumor growth and pulmonary metastasis development. TGZ may offer a new approach in chemotherapy for osteosarcoma.</p

Investigations into a putative role for the novel BRASSIKIN pseudokinases in compatible pollen-stigma interactions in Arabidopsis thaliana.

BACKGROUND: In the Brassicaceae, the early stages of compatible pollen-stigma interactions are tightly controlled with early checkpoints regulating pollen adhesion, hydration and germination, and pollen tube entry into the stigmatic surface. However, the early signalling events in the stigma which trigger these compatible interactions remain unknown. RESULTS: A set of stigma-expressed pseudokinase genes, termed BRASSIKINs (BKNs), were identified and found to be present in only core Brassicaceae genomes. In Arabidopsis thaliana Col-0, BKN1 displayed stigma-specific expression while the BKN2 gene was expressed in other tissues as well. CRISPR deletion mutations were generated for the two tandemly linked BKNs, and very mild hydration defects were observed for wild-type Col-0 pollen when placed on the bkn1/2 mutant stigmas. In further analyses, the predominant transcript for the stigma-specific BKN1 was found to have a premature stop codon in the Col-0 ecotype, but a survey of the 1001 Arabidopsis genomes uncovered three ecotypes that encoded a full-length BKN1 protein. Furthermore, phylogenetic analyses identified intact BKN1 orthologues in the closely related outcrossing Arabidopsis species, A. lyrata and A. halleri. Finally, the BKN pseudokinases were found to be plasma-membrane localized through the dual lipid modification of myristoylation and palmitoylation, and this localization would be consistent with a role in signaling complexes. CONCLUSION: In this study, we have characterized the novel Brassicaceae-specific family of BKN pseudokinase genes, and examined the function of BKN1 and BKN2 in the context of pollen-stigma interactions in A. thaliana Col-0. Additionally, premature stop codons were identified in the predicted stigma specific BKN1 gene in a number of the 1001 A. thaliana ecotype genomes, and this was in contrast to the out-crossing Arabidopsis species which carried intact copies of BKN1. Thus, understanding the function of BKN1 in other Brassicaceae species will be a key direction for future studies

A breakthrough on Amanita phalloides poisoning: an effective antidotal effect by polymyxin B

Amanita phalloides is responsible for more than 90 % of mushroom-related fatalities, and no effective antidote is available. a-Amanitin, the main toxin of A. phalloides, inhibits RNA polymerase II (RNAP II), causing hepatic and kidney failure. In silico studies included docking and molecular dynamics simulation coupled to molecular mechanics with generalized Born and surface area method energy decomposition on RNAP II. They were performed with a clinical drug that shares chemical similarities to a-amanitin, polymyxin B. The results show that polymyxin B potentially binds to RNAP II in the same interface of a-amanitin, preventing the toxin from binding to RNAP II. In vivo, the inhibition of the mRNA transcripts elicited by a-amanitin was efficiently reverted by polymyxin B in the kidneys. Moreover, polymyxin B significantly decreased the hepatic and renal a-amanitin-induced injury as seen by the histology and hepatic aminotransferases plasma data. In the survival assay, all animals exposed to a-amanitin died within 5 days, whereas 50 % survived up to 30 days when polymyxin B was administered 4, 8, and 12 h post-a-amanitin. Moreover, a single dose of polymyxin B administered concomitantly with a-amanitin was able to guarantee 100 % survival. Polymyxin B protects RNAP II from inactivation leading to an effective prevention of organ damage and increasing survival in a-amanitin-treated animals. The present use of clinically relevant concentrations of an already human-use-approved drug prompts the use of polymyxin B as an antidote for A. phalloides poisoning in humans.Juliana Garcia, Vera Marisa Costa, Ricardo Dinis-Oliveira and Ricardo Silvestre thank FCT-Foundation for Science and Technology-for their PhD grant (SFRH/BD/74979/2010), Post-doc grants (SFRH/BPD/63746/2009 and SFRH/BPD/110001/2015) and Investigator grants (IF/01147/2013) and (IF/00021/2014), respectively. This work was supported by the Fundacao para a Ciencia e Tecnologia (FCT) - project PTDC/DTPFTO/4973/2014 - and the European Union (FEDER funds through COMPETE) and National Funds (FCT, Fundacao para a Ciencia e Tecnologia) through project Pest-C/EQB/LA0006/2013

A water-based training program that include perturbation exercises to improve stepping responses in older adults: study protocol for a randomized controlled cross-over trial

<p>Abstract</p> <p>Background</p> <p>Gait and balance impairments may increase the risk of falls, the leading cause of accidental death in the elderly population. Fall-related injuries constitute a serious public health problem associated with high costs for society as well as human suffering. A rapid step is the most important protective postural strategy, acting to recover equilibrium and prevent a fall from initiating. It can arise from large perturbations, but also frequently as a consequence of volitional movements. We propose to use a novel water-based training program which includes specific perturbation exercises that will target the stepping responses that could potentially have a profound effect in reducing risk of falling. We describe the water-based balance training program and a study protocol to evaluate its efficacy (Trial registration number #NCT00708136).</p> <p>Methods/Design</p> <p>The proposed water-based training program involves use of unpredictable, multi-directional perturbations in a group setting to evoke compensatory and volitional stepping responses. Perturbations are made by pushing slightly the subjects and by water turbulence, in 24 training sessions conducted over 12 weeks. Concurrent cognitive tasks during movement tasks are included. Principles of physical training and exercise including awareness, continuity, motivation, overload, periodicity, progression and specificity were used in the development of this novel program. Specific goals are to increase the speed of stepping responses and improve the postural control mechanism and physical functioning. A prospective, randomized, cross-over trial with concealed allocation, assessor blinding and intention-to-treat analysis will be performed to evaluate the efficacy of the water-based training program. A total of 36 community-dwelling adults (age 65–88) with no recent history of instability or falling will be assigned to either the perturbation-based training or a control group (no training). Voluntary step reaction times and postural stability using stabiliogram diffusion analysis will be tested before and after the 12 weeks of training.</p> <p>Discussion</p> <p>This study will determine whether a water-based balance training program that includes perturbation exercises, in a group setting, can improve speed of voluntary stepping responses and improve balance control. Results will help guide the development of more cost-effective interventions that can prevent the occurrence of falls in the elderly.</p

The evolutionary significance of polyploidy

Polyploidy, or the duplication of entire genomes, has been observed in prokaryotic and eukaryotic organisms, and in somatic and germ cells. The consequences of polyploidization are complex and variable, and they differ greatly between systems (clonal or non-clonal) and species, but the process has often been considered to be an evolutionary 'dead end'. Here, we review the accumulating evidence that correlates polyploidization with environmental change or stress, and that has led to an increased recognition of its short-term adaptive potential. In addition, we discuss how, once polyploidy has been established, the unique retention profile of duplicated genes following whole-genome duplication might explain key longer-term evolutionary transitions and a general increase in biological complexity

Search for neutral MSSM Higgs bosons decaying to a pair of tau leptons in pp collisions

Peer reviewe

Measurement of top quark–antiquark pair production in association with a W or Z boson in pp collisions at √s=8 TeV

Peer reviewe