Validation of the SCID-hu Thy/Liv mouse model with four classes of licensed antiretrovirals.

BackgroundThe SCID-hu Thy/Liv mouse model of HIV-1 infection is a useful platform for the preclinical evaluation of antiviral efficacy in vivo. We performed this study to validate the model with representatives of all four classes of licensed antiretrovirals.Methodology/principal findingsEndpoint analyses for quantification of Thy/Liv implant viral load included ELISA for cell-associated p24, branched DNA assay for HIV-1 RNA, and detection of infected thymocytes by intracellular staining for Gag-p24. Antiviral protection from HIV-1-mediated thymocyte depletion was assessed by multicolor flow cytometric analysis of thymocyte subpopulations based on surface expression of CD3, CD4, and CD8. These mice can be productively infected with molecular clones of HIV-1 (e.g., the X4 clone NL4-3) as well as with primary R5 and R5X4 isolates. To determine whether results in this model are concordant with those found in humans, we performed direct comparisons of two drugs in the same class, each of which has known potency and dosing levels in humans. Here we show that second-generation antiretrovirals were, as expected, more potent than their first-generation predecessors: emtricitabine was more potent than lamivudine, efavirenz was more potent than nevirapine, and atazanavir was more potent than indinavir. After interspecies pharmacodynamic scaling, the dose ranges found to inhibit viral replication in the SCID-hu Thy/Liv mouse were similar to those used in humans. Moreover, HIV-1 replication in these mice was genetically stable; treatment of the mice with lamivudine did not result in the M184V substitution in reverse transcriptase, and the multidrug-resistant NY index case HIV-1 retained its drug-resistance substitutions.ConclusionGiven the fidelity of such comparisons, we conclude that this highly reproducible mouse model is likely to predict clinical antiviral efficacy in humans

Symbolism in bank marketing and architecture: the headquarters of National Provincial Bank of England

This article undertakes an analysis of the symbolism present in the architecture and design in nineteenth century British banking. It focuses upon the headquarters of National Provincial Bank of England, which was built in the 1860s. It explores the symbols and messages that those at the bank wished to communicate to those that viewed the building. The analysis finds that those at the bank impressed its national identity, achieved through its extensive branch network, as its key message which differentiated it from its rivals. Other symbols emphasized that it had adapted to the local market and was equal in terms of competency and richness in comparison to its competitors. We argue that these messages became part of the organization’s identity and its brand, as well as the culture of the City of London more broadly. The article provides a new explanation for symbolic meanings represented by bank architecture. It integrates the existing discussion of bank architecture in historical research with the theoretical frameworks and literature being developed in organizational identity and branding

Ventilatory frequency as a measure of the response of tammar wallabies (Macropus eugenii) to the odour of potential predators

This study uses changes in ventilatory frequency to quantify the physiological response of an Australian terrestrial herbivore, the tammar wallaby (Macropus eugenii), to olfactory cues suggesting the presence of potential predators. Ventilatory frequency proved to be a quantifiable measure to assess the response of this macropod marsupial to olfactory cues. Ventilatory frequency increased from mean resting levels of 45 ± 5.1 breaths min–1 to 137 ± 11.2 breaths min–1 during the first minute of exposure to all odours. These physiological responses diminished over time, with ventilatory frequency in the first minute after introduction of the scents greater than that during the subsequent four, suggesting that the initial reaction was due to disturbance and was investigative in nature. However, the ratio of ventilatory frequency in the remaining 4 min after introduction of the odours compared with before was greater for fox (3.58 ± 0.918) and cat (2.44 ± 0.272) odours than for snake (2.27 ± 0.370), distilled water (1.81 ± 0.463) and quoll (1.71 ± 0.245) odours, suggesting that fox and cat odour provoked a greater response. However, the wallabies’ response to the odour of these introduced predators and to horse odour (2.40 ± 0.492) did not differ. Our study indicates that a long period of co-history with particular predators is not a prerequisite for detection of potentially threatening species. We do not find any support for the hypothesis that an inability to interpret olfactory cues to detect and respond to potential predation by introduced predators is responsible for the decline of these macropod marsupials

On the general theory of the origins of retroviruses

<p>Abstract</p> <p>Background</p> <p>The order retroviridae comprises viruses based on ribonucleic acids (RNA). Some, such as HIV and HTLV, are human pathogens. Newly emerged human retroviruses have zoonotic origins. As far as has been established, both repeated infections (themselves possibly responsible for the evolution of viral mutations <b>(Vm) </b>and host adaptability <b>(Ha)</b>); along with interplay between <it>inhibitors </it>and <it>promoters </it>of cell tropism, are needed to effect retroviral cross-species transmissions. However, the exact <it>modus operadi </it>of intertwine between these factors at molecular level remains to be established. Knowledge of such intertwine could lead to a better understanding of retrovirology and possibly other infectious processes. This study was conducted to derive the mathematical equation of a general theory of the origins of retroviruses.</p> <p>Methods and results</p> <p>On the basis of an arbitrarily non-Euclidian geometrical "thought experiment" involving the cross-species transmission of simian foamy virus (sfv) from a non-primate species <it>Xy </it>to <it>Homo sapiens </it>(<it>Hs</it>), initially excluding all social factors, the following was derived. At the port of exit from <it>Xy </it>(where the species barrier, SB, is defined by the <it>Index of Origin</it>, IO), sfv shedding is (1) enhanced by two transmitting tensors <b>(Tt)</b>, (i) virus-specific immunity (VSI) and (ii) evolutionary defenses such as APOBEC, RNA interference pathways, and (when present) expedited therapeutics (denoted e²D); and (2) opposed by the five accepting scalars <b>(At)</b>: (a) genomic integration hot spots, gIHS, (b) nuclear envelope transit <b>(</b>NMt) vectors, (c) virus-specific cellular biochemistry, VSCB, (d) virus-specific cellular receptor repertoire, VSCR, and (e) pH-mediated cell membrane transit, (↓_pHCMat). Assuming <b>As </b>and <b>Tt </b>to be independent variables, <b>IO = Tt/As</b>. The same forces acting in an opposing manner determine SB at the port of sfv entry (defined here by the <it>Index of Entry</it>, <b>IE = As/Tt</b>). Overall, If sfv encounters no unforeseen effects on transit between X<it>y </it>and <it>Hs</it>, then the square root of the combined index of sfv transmissibility (√<b>|RTI|) </b>is proportional to the product IO* IE (or ~Vm* Ha* ∑Tt*∑As*<b>Ω</b>), where <b>Ω </b>is the retrovirological constant and ∑ is a function of the ratio Tt/As or As/Tt for sfv transmission from <it>Xy </it>to <it>Hs</it>.</p> <p>Conclusions</p> <p>I present a mathematical formalism encapsulating the general theory of the origins of retroviruses. It summarizes the choreography for the intertwined interplay of factors influencing the probability of retroviral cross-species transmission: <b>Vm, Ha, Tt, As, </b>and <b>Ω</b>.</p

Novel HIV-1 Knockdown Targets Identified by an Enriched Kinases/Phosphatases shRNA Library Using a Long-Term Iterative Screen in Jurkat T-Cells

HIV-1 is a complex retrovirus that uses host machinery to promote its replication. Understanding cellular proteins involved in the multistep process of HIV-1 infection may result in the discovery of more adapted and effective therapeutic targets. Kinases and phosphatases are a druggable class of proteins critically involved in regulation of signal pathways of eukaryotic cells. Here, we focused on the discovery of kinases and phosphatases that are essential for HIV-1 replication but dispensable for cell viability. We performed an iterative screen in Jurkat T-cells with a short-hairpin-RNA (shRNA) library highly enriched for human kinases and phosphatases. We identified 14 new proteins essential for HIV-1 replication that do not affect cell viability. These proteins are described to be involved in MAPK, JNK and ERK pathways, vesicular traffic and DNA repair. Moreover, we show that the proteins under study are important in an early step of HIV-1 infection before viral integration, whereas some of them affect viral transcription/translation. This study brings new insights for the complex interplay of HIV-1/host cell and opens new possibilities for antiviral strategies

Structural Analysis of the UBA Domain of X-linked Inhibitor of Apoptosis Protein Reveals Different Surfaces for Ubiquitin-Binding and Self-Association

BACKGROUND: Inhibitor of apoptosis proteins (IAPs) belong to a pivotal antiapoptotic protein family that plays a crucial role in tumorigenesis, cancer progression, chemoresistance and poor patient-survival. X-linked inhibitor of apoptosis protein (XIAP) is a prominent member of IAPs attracting intense research because it has been demonstrated to be a physiological inhibitor of caspases and apoptosis. Recently, an evolutionarily conserved ubiquitin-associated (UBA) domain was identified in XIAP and a number of RING domain-bearing IAPs. This has placed the IAPs in the group of ubiquitin binding proteins. Here, we explore the three-dimensional structure of the XIAP UBA domain (XIAP-UBA) and how it interacts with mono-ubiquitin and diubiquitin conjugates. PRINCIPAL FINDINGS: The solution structure of the XIAP-UBA domain was determined by NMR spectroscopy. XIAP-UBA adopts a typical UBA domain fold of three tightly packed alpha-helices but with an additional N-terminal 3(10) helix. The XIAP-UBA binds mono-ubiquitin as well as Lys48-linked and linear-linked diubiquitins at low-micromolar affinities. NMR analysis of the XIAP-UBA-ubiquitin interaction reveals that it involves the classical hydrophobic patches surrounding Ile44 of ubiquitin and the conserved MGF/LV motif surfaces on XIAP-UBA. Furthermore, dimerization of XIAP-UBA was observed. Mapping of the self-association surface of XIAP-UBA reveals that the dimerization interface is formed by residues in the N-terminal 3(10) helix, helix alpha1 and helix alpha2, separate from the ubiquitin-binding surface. CONCLUSION: Our results provide the first structural information of XIAP-UBA and map its interaction with mono-ubiquitin, Lys48-linked and linear-linked diubiquitins. The notion that XIAP-UBA uses different surfaces for ubiquitin-binding and self-association provides a plausible model to explain the reported selectivity of XIAP in binding polyubiquitin chains with different linkages.published_or_final_versio

Deciphering the Code for Retroviral Integration Target Site Selection

Upon cell invasion, retroviruses generate a DNA copy of their RNA genome and integrate retroviral cDNA within host chromosomal DNA. Integration occurs throughout the host cell genome, but target site selection is not random. Each subgroup of retrovirus is distinguished from the others by attraction to particular features on chromosomes. Despite extensive efforts to identify host factors that interact with retrovirion components or chromosome features predictive of integration, little is known about how integration sites are selected. We attempted to identify markers predictive of retroviral integration by exploiting Precision-Recall methods for extracting information from highly skewed datasets to derive robust and discriminating measures of association. ChIPSeq datasets for more than 60 factors were compared with 14 retroviral integration datasets. When compared with MLV, PERV or XMRV integration sites, strong association was observed with STAT1, acetylation of H3 and H4 at several positions, and methylation of H2AZ, H3K4, and K9. By combining peaks from ChIPSeq datasets, a supermarker was identified that localized within 2 kB of 75% of MLV proviruses and detected differences in integration preferences among different cell types. The supermarker predicted the likelihood of integration within specific chromosomal regions in a cell-type specific manner, yielding probabilities for integration into proto-oncogene LMO2 identical to experimentally determined values. The supermarker thus identifies chromosomal features highly favored for retroviral integration, provides clues to the mechanism by which retrovirus integration sites are selected, and offers a tool for predicting cell-type specific proto-oncogene activation by retroviruses

Effects of fluoxetine on functional outcomes after acute stroke (FOCUS): a pragmatic, double-blind, randomised, controlled trial

Background Results of small trials indicate that fluoxetine might improve functional outcomes after stroke. The FOCUS trial aimed to provide a precise estimate of these effects. Methods FOCUS was a pragmatic, multicentre, parallel group, double-blind, randomised, placebo-controlled trial done at 103 hospitals in the UK. Patients were eligible if they were aged 18 years or older, had a clinical stroke diagnosis, were enrolled and randomly assigned between 2 days and 15 days after onset, and had focal neurological deficits. Patients were randomly allocated fluoxetine 20 mg or matching placebo orally once daily for 6 months via a web-based system by use of a minimisation algorithm. The primary outcome was functional status, measured with the modified Rankin Scale (mRS), at 6 months. Patients, carers, health-care staff, and the trial team were masked to treatment allocation. Functional status was assessed at 6 months and 12 months after randomisation. Patients were analysed according to their treatment allocation. This trial is registered with the ISRCTN registry, number ISRCTN83290762. Findings Between Sept 10, 2012, and March 31, 2017, 3127 patients were recruited. 1564 patients were allocated fluoxetine and 1563 allocated placebo. mRS data at 6 months were available for 1553 (99·3%) patients in each treatment group. The distribution across mRS categories at 6 months was similar in the fluoxetine and placebo groups (common odds ratio adjusted for minimisation variables 0·951 [95% CI 0·839–1·079]; p=0·439). Patients allocated fluoxetine were less likely than those allocated placebo to develop new depression by 6 months (210 [13·43%] patients vs 269 [17·21%]; difference 3·78% [95% CI 1·26–6·30]; p=0·0033), but they had more bone fractures (45 [2·88%] vs 23 [1·47%]; difference 1·41% [95% CI 0·38–2·43]; p=0·0070). There were no significant differences in any other event at 6 or 12 months. Interpretation Fluoxetine 20 mg given daily for 6 months after acute stroke does not seem to improve functional outcomes. Although the treatment reduced the occurrence of depression, it increased the frequency of bone fractures. These results do not support the routine use of fluoxetine either for the prevention of post-stroke depression or to promote recovery of function. Funding UK Stroke Association and NIHR Health Technology Assessment Programme

Dimethyl fumarate in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial

Dimethyl fumarate (DMF) inhibits inflammasome-mediated inflammation and has been proposed as a treatment for patients hospitalised with COVID-19. This randomised, controlled, open-label platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]), is assessing multiple treatments in patients hospitalised for COVID-19 (NCT04381936, ISRCTN50189673). In this assessment of DMF performed at 27 UK hospitals, adults were randomly allocated (1:1) to either usual standard of care alone or usual standard of care plus DMF. The primary outcome was clinical status on day 5 measured on a seven-point ordinal scale. Secondary outcomes were time to sustained improvement in clinical status, time to discharge, day 5 peripheral blood oxygenation, day 5 C-reactive protein, and improvement in day 10 clinical status. Between 2 March 2021 and 18 November 2021, 713 patients were enroled in the DMF evaluation, of whom 356 were randomly allocated to receive usual care plus DMF, and 357 to usual care alone. 95% of patients received corticosteroids as part of routine care. There was no evidence of a beneficial effect of DMF on clinical status at day 5 (common odds ratio of unfavourable outcome 1.12; 95% CI 0.86-1.47; p = 0.40). There was no significant effect of DMF on any secondary outcome

Tocilizumab in patients admitted to hospital with COVID-19 (RECOVERY): a randomised, controlled, open-label, platform trial

Background: In this study, we aimed to evaluate the effects of tocilizumab in adult patients admitted to hospital with COVID-19 with both hypoxia and systemic inflammation. Methods: This randomised, controlled, open-label, platform trial (Randomised Evaluation of COVID-19 Therapy [RECOVERY]), is assessing several possible treatments in patients hospitalised with COVID-19 in the UK. Those trial participants with hypoxia (oxygen saturation &lt;92% on air or requiring oxygen therapy) and evidence of systemic inflammation (C-reactive protein ≥75 mg/L) were eligible for random assignment in a 1:1 ratio to usual standard of care alone versus usual standard of care plus tocilizumab at a dose of 400 mg–800 mg (depending on weight) given intravenously. A second dose could be given 12–24 h later if the patient's condition had not improved. The primary outcome was 28-day mortality, assessed in the intention-to-treat population. The trial is registered with ISRCTN (50189673) and ClinicalTrials.gov (NCT04381936). Findings: Between April 23, 2020, and Jan 24, 2021, 4116 adults of 21 550 patients enrolled into the RECOVERY trial were included in the assessment of tocilizumab, including 3385 (82%) patients receiving systemic corticosteroids. Overall, 621 (31%) of the 2022 patients allocated tocilizumab and 729 (35%) of the 2094 patients allocated to usual care died within 28 days (rate ratio 0·85; 95% CI 0·76–0·94; p=0·0028). Consistent results were seen in all prespecified subgroups of patients, including those receiving systemic corticosteroids. Patients allocated to tocilizumab were more likely to be discharged from hospital within 28 days (57% vs 50%; rate ratio 1·22; 1·12–1·33; p&lt;0·0001). Among those not receiving invasive mechanical ventilation at baseline, patients allocated tocilizumab were less likely to reach the composite endpoint of invasive mechanical ventilation or death (35% vs 42%; risk ratio 0·84; 95% CI 0·77–0·92; p&lt;0·0001). Interpretation: In hospitalised COVID-19 patients with hypoxia and systemic inflammation, tocilizumab improved survival and other clinical outcomes. These benefits were seen regardless of the amount of respiratory support and were additional to the benefits of systemic corticosteroids. Funding: UK Research and Innovation (Medical Research Council) and National Institute of Health Research