Identifying and quantifying heterogeneity in high content analysis: Application of heterogeneity indices to drug discovery

One of the greatest challenges in biomedical research, drug discovery and diagnostics is understanding how seemingly identical cells can respond differently to perturbagens including drugs for disease treatment. Although heterogeneity has become an accepted characteristic of a population of cells, in drug discovery it is not routinely evaluated or reported. The standard practice for cell-based, high content assays has been to assume a normal distribution and to report a well-to-well average value with a standard deviation. To address this important issue we sought to define a method that could be readily implemented to identify, quantify and characterize heterogeneity in cellular and small organism assays to guide decisions during drug discovery and experimental cell/tissue profiling. Our study revealed that heterogeneity can be effectively identified and quantified with three indices that indicate diversity, non-normality and percent outliers. The indices were evaluated using the induction and inhibition of STAT3 activation in five cell lines where the systems response including sample preparation and instrument performance were well characterized and controlled. These heterogeneity indices provide a standardized method that can easily be integrated into small and large scale screening or profiling projects to guide interpretation of the biology, as well as the development of therapeutics and diagnostics. Understanding the heterogeneity in the response to perturbagens will become a critical factor in designing strategies for the development of therapeutics including targeted polypharmacology. © 2014 Gough et al

Evolution of Th2 responses : Characterization of IL-4/13 in sea bass (Dicentrarchus labrax L.) and studies of expression and biological activity

Acknowledgements This research was funded by the European Commission under the 7th Framework Programme for Research and Technological Development (FP7) of the European Union (Grant Agreement 311993 TARGETFISH). T.W. received funding from the MASTS pooling initiative (The Marine Alliance for Science and Technology for Scotland). MASTS is funded by the Scottish Funding Council (grant reference number HR09011) and contributing institutions.Peer reviewedPublisher PD

Framing Male Circumcision to Promote its Adoption in Different Settings

The effectiveness of male circumcision in preventing transmission of HIV from females to males has been established. Those who are now advocating its widespread use face many challenges in convincing policy-makers and the public of circumcision’s value. We suggest that frames are a useful lens for communicating public health messages that may help promote adoption of circumcision. Frames relate to how individuals and societies perceive and understand the world. Existing frames are often hard to shift, and should be borne in mind by advocates and program implementers as they attempt to promote male circumcision by invoking new frames. Frames differ across and within societies, and advocates must find ways of delivering resonant messages that take into account prior perceptions and use the most appropriate means of communicating the benefits and value of male circumcision to different audiences

A novel 3-hydroxypropionic acid-inducible promoter regulated by the LysR-type transcriptional activator protein MmsR of Pseudomonas denitrificans

MmsR (33.3 kDa) is a putative LysR-type transcriptional activator of Pseudomonas denitrificans. With the help of 3-hydroxypropionic acid (3-HP), an important platform chemical, MmsR positively regulates the expression of mmsA, which encodes methylmalonylsemialdehyde dehydrogenase, the enzyme involved in valine degradation. In the present study, the cellular function of MmsR and its binding to the regulatory DNA sequence of mmsA expression were investigated both in vivo and in vitro. Transcription of the mmsA was enhanced >140-fold in the presence of 3-HP. In the MmsR-responsive promoter region, two operators showing dyad symmetry, designated O-1 and O-2 and centered at the -79 and -28 positions, respectively, were present upstream of the mmsA transcription start site. An electrophoretic mobility shift assay indicated that MmsR binds to both operator sites for transcription activation, probably in cooperative manner. When either O-1 or O-2 or both regions were mutated, the inducibility by the MmsR-3-HP complex was significantly reduced or completely removed, indicating that both sites are required for transcription activation. A 3-HP sensor was developed by connecting the activation of MmsR to a green fluorescent readout. A more than 50-fold induction by 25 mM 3-HP was observed

The Hubble Constant

I review the current state of determinations of the Hubble constant, which gives the length scale of the Universe by relating the expansion velocity of objects to their distance. There are two broad categories of measurements. The first uses individual astrophysical objects which have some property that allows their intrinsic luminosity or size to be determined, or allows the determination of their distance by geometric means. The second category comprises the use of all-sky cosmic microwave background, or correlations between large samples of galaxies, to determine information about the geometry of the Universe and hence the Hubble constant, typically in a combination with other cosmological parameters. Many, but not all, object-based measurements give $H_0$ values of around 72-74km/s/Mpc , with typical errors of 2-3km/s/Mpc. This is in mild discrepancy with CMB-based measurements, in particular those from the Planck satellite, which give values of 67-68km/s/Mpc and typical errors of 1-2km/s/Mpc. The size of the remaining systematics indicate that accuracy rather than precision is the remaining problem in a good determination of the Hubble constant. Whether a discrepancy exists, and whether new physics is needed to resolve it, depends on details of the systematics of the object-based methods, and also on the assumptions about other cosmological parameters and which datasets are combined in the case of the all-sky methods.Comment: Extensively revised and updated since the 2007 version: accepted by Living Reviews in Relativity as a major (2014) update of LRR 10, 4, 200

Open-label, clinical phase I studies of tasquinimod in patients with castration-resistant prostate cancer

Background:Tasquinimod is a quinoline-3-carboxamide derivative with anti-angiogenic activity. Two open-label phase I clinical trials in patients were conducted to evaluate the safety and tolerability of tasquinimod, with additional pharmacokinetic and efficacy assessments.Methods:Patients with castration-resistant prostate cancer with no previous chemotherapy were enrolled in this study. The patients received tasquinimod up to 1 year either at fixed doses of 0.5 or 1.0 mg per day or at an initial dose of 0.25 mg per day that escalated to 1.0 mg per day.Results:A total of 32 patients were enrolled; 21 patients were maintained for >/=4 months. The maximum tolerated dose was determined to be 0.5 mg per day; but when using stepwise intra-patient dose escalation, a dose of 1.0 mg per day was well tolerated. The dose-limiting toxicity was sinus tachycardia and asymptomatic elevation in amylase. Common treatment-emergent adverse events included transient laboratory abnormalities, anaemia, nausea, fatigue, myalgia and pain. A serum prostate-specific antigen (PSA) decline of >/=50% was noted in two patients. The median time to PSA progression (>25%) was 19 weeks. Only 3 out of 15 patients (median time on study: 34 weeks) developed new bone lesions.Conclusion:Long-term continuous oral administration of tasquinimod seems to be safe, and the overall efficacy results indicate that tasquinimod might delay disease progression.British Journal of Cancer advance online publication, 15 September 2009; doi:10.1038/sj.bjc.6605322 www.bjcancer.com

Expression of SPIG1 Reveals Development of a Retinal Ganglion Cell Subtype Projecting to the Medial Terminal Nucleus in the Mouse

Visual information is transmitted to the brain by roughly a dozen distinct types of retinal ganglion cells (RGCs) defined by a characteristic morphology, physiology, and central projections. However, our understanding about how these parallel pathways develop is still in its infancy, because few molecular markers corresponding to individual RGC types are available. Previously, we reported a secretory protein, SPIG1 (clone name; D/Bsp120I #1), preferentially expressed in the dorsal region in the developing chick retina. Here, we generated knock-in mice to visualize SPIG1-expressing cells with green fluorescent protein. We found that the mouse retina is subdivided into two distinct domains for SPIG1 expression and SPIG1 effectively marks a unique subtype of the retinal ganglion cells during the neonatal period. SPIG1-positive RGCs in the dorsotemporal domain project to the dorsal lateral geniculate nucleus (dLGN), superior colliculus, and accessory optic system (AOS). In contrast, in the remaining region, here named the pan-ventronasal domain, SPIG1-positive cells form a regular mosaic and project exclusively to the medial terminal nucleus (MTN) of the AOS that mediates the optokinetic nystagmus as early as P1. Their dendrites costratify with ON cholinergic amacrine strata in the inner plexiform layer as early as P3. These findings suggest that these SPIG1-positive cells are the ON direction selective ganglion cells (DSGCs). Moreover, the MTN-projecting cells in the pan-ventronasal domain are apparently composed of two distinct but interdependent regular mosaics depending on the presence or absence of SPIG1, indicating that they comprise two functionally distinct subtypes of the ON DSGCs. The formation of the regular mosaic appears to be commenced at the end of the prenatal stage and completed through the peak period of the cell death at P6. SPIG1 will thus serve as a useful molecular marker for future studies on the development and function of ON DSGCs

Limits of Calcium Clearance by Plasma Membrane Calcium ATPase in Olfactory Cilia

BACKGROUND: In any fine sensory organelle, a small influx of Ca(2+) can quickly elevate cytoplasmic Ca(2+). Mechanisms must exist to clear the ciliary Ca(2+) before it reaches toxic levels. One such organelle has been well studied: the vertebrate olfactory cilium. Recent studies have suggested that clearance from the olfactory cilium is mediated in part by plasma membrane Ca(2+)-ATPase (PMCA). PRINCIPAL FINDINGS: In the present study, electrophysiological assays were devised to monitor cytoplasmic free Ca(2+) in single frog olfactory cilia. Ca(2+) was allowed to enter isolated cilia, either through the detached end or through membrane channels. Intraciliary Ca(2+) was monitored via the activity of ciliary Ca(2+)-gated Cl(-) channels, which are sensitive to free Ca(2+) from about 2 to 10 microM. No significant effect of MgATP on intraciliary free Ca(2+) could be found. Carboxyeosin, which has been used to inhibit PMCA, was found to substantially increase a ciliary transduction current activated by cyclic AMP. This increase was ATP-independent. CONCLUSIONS: Alternative explanations are suggested for two previous experiments taken to support a role for PMCA in ciliary Ca(2+) clearance. It is concluded that PMCA in the cilium plays a very limited role in clearing the micromolar levels of intraciliary Ca(2+) produced during the odor response