An efficient method for generation of bi-allelic null mutant mouse embryonic stem cells and its application for investigating epigenetic modifiers.

Mouse embryonic stem (ES) cells are a popular model system to study biological processes, though uncovering recessive phenotypes requires inactivating both alleles. Building upon resources from the International Knockout Mouse Consortium (IKMC), we developed a targeting vector for second allele inactivation in conditional-ready IKMC 'knockout-first' ES cell lines. We applied our technology to several epigenetic regulators, recovering bi-allelic targeted clones with a high efficiency of 60% and used Flp recombinase to restore expression in two null cell lines to demonstrate how our system confirms causality through mutant phenotype reversion. We designed our strategy to select against re-targeting the 'knockout-first' allele and identify essential genes in ES cells, including the histone methyltransferase Setdb1. For confirmation, we exploited the flexibility of our system, enabling tamoxifen inducible conditional gene ablation while controlling for genetic background and tamoxifen effects. Setdb1 ablated ES cells exhibit severe growth inhibition, which is not rescued by exogenous Nanog expression or culturing in naive pluripotency '2i' media, suggesting that the self-renewal defect is mediated through pluripotency network independent pathways. Our strategy to generate null mutant mouse ES cells is applicable to thousands of genes and repurposes existing IKMC Intermediate Vectors

Chronic Leukemias

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/66325/1/j.1365-4362.1982.tb03146.x.pd

Distinguishing Asthma Phenotypes Using Machine Learning Approaches.

Asthma is not a single disease, but an umbrella term for a number of distinct diseases, each of which are caused by a distinct underlying pathophysiological mechanism. These discrete disease entities are often labelled as asthma endotypes. The discovery of different asthma subtypes has moved from subjective approaches in which putative phenotypes are assigned by experts to data-driven ones which incorporate machine learning. This review focuses on the methodological developments of one such machine learning technique-latent class analysis-and how it has contributed to distinguishing asthma and wheezing subtypes in childhood. It also gives a clinical perspective, presenting the findings of studies from the past 5 years that used this approach. The identification of true asthma endotypes may be a crucial step towards understanding their distinct pathophysiological mechanisms, which could ultimately lead to more precise prevention strategies, identification of novel therapeutic targets and the development of effective personalized therapies

A Threshold Value for the Time Delay to TB Diagnosis

The original publication is available at http:/www.plosone.orgIncludes bibliographyBackgound. In many communities where TB occurs at high incidence, the major force driving the epidemic is transmission. It is plausible that the typical long delay from the onset of infectious disease to diagnosis and commencement of treatment is almost certainly the major factor contributing to the high rate of transmission. Methodology/Principal Findings. This study is confined to communities which are epidemiologically relatively isolated and which have low HIV incidence. The consequences of delays to diagnosis are analyzed and the existence of a threshold delay value is demonstrated. It is shown that unless a sufficient number of cases are detected before this threshold, the epidemic will escalate. The method used for the analysis avoids the standard computer integration of systems of differential equations since the intention is to present a line of reasoning that reveals the essential dynamics of an epidemic in an intuitively clear way that is nevertheless quantitatively realistic. Conclusions/Significance. The analysis presented here shows that typical delays to diagnosis present a major obstacle to the control of a TB epidemic. Control can be achieved by optimizing the rapid identification of TB cases together with measures to increase the threshold value. A calculated and aggressive program is therefore necessary in order to bring about a reduction in the prevalence of TB in a community by decreasing the time to diagnosis in all its ramifications. Intervention strategies to increase the threshold value relative to the time to diagnosis and which thereby decrease disease incidence are discussed. © 2007 Uys et al.Publishers' Versio

Tnni3k Modifies Disease Progression in Murine Models of Cardiomyopathy

The Calsequestrin (Csq) transgenic mouse model of cardiomyopathy exhibits wide variation in phenotypic progression dependent on genetic background. Seven heart failure modifier (Hrtfm) loci modify disease progression and outcome. Here we report Tnni3k (cardiac Troponin I-interacting kinase) as the gene underlying Hrtfm2. Strains with the more susceptible phenotype exhibit high transcript levels while less susceptible strains show dramatically reduced transcript levels. This decrease is caused by an intronic SNP in low-transcript strains that activates a cryptic splice site leading to a frameshifted transcript, followed by nonsense-mediated decay of message and an absence of detectable protein. A transgenic animal overexpressing human TNNI3K alone exhibits no cardiac phenotype. However, TNNI3K/Csq double transgenics display severely impaired systolic function and reduced survival, indicating that TNNI3K expression modifies disease progression. TNNI3K expression also accelerates disease progression in a pressure-overload model of heart failure. These combined data demonstrate that Tnni3k plays a critical role in the modulation of different forms of heart disease, and this protein may provide a novel target for therapeutic intervention

Actual and undiagnosed HIV prevalence in a community sample of men who have sex with men in Auckland, New Zealand

<p>Abstract</p> <p>Background</p> <p>The prevalence of HIV infection and how this varies between subgroups is a fundamental indicator of epidemic control. While there has been a rise in the number of HIV diagnoses among men who have sex with men (MSM) in New Zealand over the last decade, the actual prevalence of HIV and the proportion undiagnosed is not known. We measured these outcomes in a community sample of MSM in Auckland, New Zealand.</p> <p>Methods</p> <p>The study was embedded in an established behavioural surveillance programme. MSM attending a gay community fair day, gay bars and sex-on-site venues during 1 week in February 2011 who agreed to complete a questionnaire were invited to provide an anonymous oral fluid specimen for analysis of HIV antibodies. From the 1304 eligible respondents (acceptance rate 48.5%), 1049 provided a matched specimen (provision rate 80.4%).</p> <p>Results</p> <p>HIV prevalence was 6.5% (95% CI: 5.1-8.1). After adjusting for age, ethnicity and recruitment site, HIV positivity was significantly elevated among respondents who were aged 30-44 or 45 and over, were resident outside New Zealand, had 6-20 or more than 20 recent sexual partners, had engaged in unprotected anal intercourse with a casual partner, had had sex with a man met online, or had injected drugs in the 6 months prior to survey. One fifth (20.9%) of HIV infected men were undiagnosed; 1.3% of the total sample. Although HIV prevalence did not differ by ethnicity, HIV infected non-European respondents were more likely to be undiagnosed. Most of the small number of undiagnosed respondents had tested for HIV previously, and the majority believed themselves to be either "definitely" or "probably" uninfected. There was evidence of continuing risk practices among some of those with known HIV infection.</p> <p>Conclusions</p> <p>This is the first estimate of actual and undiagnosed HIV infection among a community sample of gay men in New Zealand. While relatively low compared to other countries with mature epidemics, HIV prevalence was elevated in subgroups of MSM based on behaviour, and diagnosis rates varied by ethnicity. Prevention should focus on raising condom use and earlier diagnosis among those most at risk, and encouraging safe behaviour after diagnosis.</p

Effect of prostaglandin E2 injection on the structural properties of the rat patellar tendon

Background: Increased tendon production of the inflammatory mediator prostaglandin E2 (PGE2) has been suggested to be a potential etiologic agent in the development of tendinopathy. Repeated injection of PGE2 into tendon has been proposed as a potential animal model for studying treatments for tendinopathy. In contrast, nonsteroidal anti-inflammatory drugs (NSAIDs) which inhibit PGE2 production and are commonly prescribed in treating tendinopathy have been shown to impair the healing of tendon after acute injury in animal models. The contradictory literature suggests the need to better define the functional effects of PGE2 on tendon. Our objective was to characterize the effects of PGE2 injection on the biomechanical and biochemical properties of tendon and the activity of the animals. Our hypothesis was that weekly PGE2 injection to the rat patellar tendon would lead to inferior biomechanical properties. Methods: Forty rats were divided equally into four groups. Three groups were followed for 4 weeks with the following peritendinous injection procedures: No injection (control), 4 weekly injections of saline (saline), 4 weekly injections of 800 ng PGE2 (PGE2-4 wks). The fourth group received 4 weekly injections of 800 ng PGE2 initially and was followed for a total of 8 weeks. All animals were injected bilaterally. The main outcome measurements included: the structural and material properties of the patellar tendon under tensile loading to failure, tendo

Cost-Effectiveness of Pooled Nucleic Acid Amplification Testing for Acute HIV Infection after Third-Generation HIV Antibody Screening and Rapid Testing in the United States: A Comparison of Three Public Health Settings

Angela Hutchinson and colleagues conducted a cost-effectiveness analysis of pooled nucleic acid amplification testing following HIV testing and show that it is not cost-effective at recommended antibody testing intervals for high-risk persons except in very high-incidence settings

Search for CP violation in D+→ϕπ+ and D+s→K0Sπ+ decays

A search for CP violation in D + → ϕπ + decays is performed using data collected in 2011 by the LHCb experiment corresponding to an integrated luminosity of 1.0 fb−1 at a centre of mass energy of 7 TeV. The CP -violating asymmetry is measured to be (−0.04 ± 0.14 ± 0.14)% for candidates with K − K + mass within 20 MeV/c 2 of the ϕ meson mass. A search for a CP -violating asymmetry that varies across the ϕ mass region of the D + → K − K + π + Dalitz plot is also performed, and no evidence for CP violation is found. In addition, the CP asymmetry in the D+s→K0Sπ+ decay is measured to be (0.61 ± 0.83 ± 0.14)%