Monitoring phagocytic uptake of amyloid beta into glial cell lysosomes in real time

Phagocytosis by glial cells is essential to regulate brain function during health and disease. Therapies for Alzheimer's disease (AD) have primarily focused on targeting antibodies to amyloid β (Aβ) or inhibitng enzymes that make it, and while removal of Aβ by phagocytosis is protective early in AD it remains poorly understood. Impaired phagocytic function of glial cells during later stages of AD likely contributes to worsened disease outcome, but the underlying mechanisms of how this occurs remain unknown. We have developed a human Aβ_{1-42} analogue (Aβ^{pH}) that exhibits green fluorescence upon internalization into the acidic organelles of cells but is non-fluorescent at physiological pH. This allowed us to image, for the first time, glial uptake of Aβ^{pH} in real time in live animals. We find that microglia phagocytose more AβpH than astrocytes in culture, in brain slices and in vivo. Aβ^{pH} can be used to investigate the phagocytic mechanisms responsible for removing Aβ from the extracellular space, and thus could become a useful tool to study Aβ clearance at different stages of AD

Titin mutations in iPS cells define sarcomere insufficiency as a cause of dilated cardiomyopathy

Human mutations that truncate the massive sarcomere protein titin [TTN-truncating variants (TTNtvs)] are the most common genetic cause for dilated cardiomyopathy (DCM), a major cause of heart failure and premature death. Here we show that cardiac microtissues engineered from human induced pluripotent stem (iPS) cells are a powerful system for evaluating the pathogenicity of titin gene variants. We found that certain missense mutations, like TTNtvs, diminish contractile performance and are pathogenic. By combining functional analyses with RNA sequencing, we explain why truncations in the A-band domain of TTN cause DCM, whereas truncations in the I band are better tolerated. Finally, we demonstrate that mutant titin protein in iPS cell-derived cardiomyocytes results in sarcomere insufficiency, impaired responses to mechanical and {beta}-adrenergic stress, and attenuated growth factor and cell signaling activation. Our findings indicate that titin mutations cause DCM by disrupting critical linkages between sarcomerogenesis and adaptive remodeling

Carrier concentration dependence of optical Kerr nonlinearity in indium tin oxide films

Optical Kerr nonlinearity (n2) in n-type indium tin oxide (ITO) films coated on glass substrates has been measured using Z-scans with 200-fs laser pulses at wavelengths ranging from 720 to 780 nm. The magnitudes of the measured nonlinearity in the ITO films were found to be dependent on the carrier concentration with a maximum n2-value of 4.1 x 10-5 cm2/GW at 720-nm wavelength and an electron density of Nd = 5.8 x 1020 cm-3. The Kerr nonlinearity was also observed to be varied with the laser wavelength. By employing a femtosecond time-resolved optical Kerr effect (OKE) technique, the relaxation time of OKE in the ITO films is determined to be ~1 ps. These findings suggest that the Kerr nonlinearity in ITO can be tailored by controlling the carrier concentration, which should be highly desirable in optoelectronic devices for ultrafast all-optical switching.Comment: 15 pages, 1 table, 4 figure

Interaction of the tetracyclines with double-stranded RNAs of random base sequence: new perspectives on the target and mechanism of action

The 16S rRNA binding mechanism proposed for the antibacterial action of the tetracyclines does not explain their mechanism of action against non-bacterial pathogens. In addition, several contradictory base pairs have been proposed as their binding sites on the 16S rRNA. This study investigated the binding of minocycline and doxycycline to short double-stranded RNAs (dsRNAs) of random base sequences. These tetracyclines caused a dose-dependent decrease in the fluorescence intensities of 6-carboxyfluorescein (FAM)-labelled dsRNA and ethidium bromide (EtBr)-stained dsRNA, indicating that both drugs bind to dsRNA of random base sequence in a manner that is competitive with the binding of EtBr and other nucleic acid ligands often used as stains. This effect was observable in the presence of Mg2+. The binding of the tetracyclines to dsRNA changed features of the fluorescence emission spectra of the drugs and the CD spectra of the RNA, and inhibited RNase III cleavage of the dsRNA. These results indicate that the double-stranded structures of RNAs may have a more important role in their interaction with the tetracyclines than the specific base pairs, which had hitherto been the subject of much investigation. Given the diverse functions of cellular RNAs, the binding of the tetracyclines to their double-stranded helixes may alter the normal processing and functioning of the various biological processes they regulate. This could help to explain the wide range of action of the tetracyclines against various pathogens and disease condition

A vector-valued ground motion intensity measure incorporating normalized spectral area

A vector-valued intensity measure is presented, which incorporates a relative measure represented by the normalized spectral area. The proposed intensity measure is intended to have high correlation with specific relative engineering demand parameters, which collectively can provide information regarding the damage state and collapse potential of the structure. Extensive dynamic analyses are carried out on a single-degree-of-freedom system with a modified Clough–Johnston hysteresis model, using a dataset of 40 ground motions, in order to investigate the proposed intensity measure characteristics. Response is expressed using the displacement ductility, and the normalized hysteretic energy, both of which are relative engineering demand parameters. Through regression analysis the correlation between the proposed intensity measure and the engineering demand parameters is evaluated. Its domain of applicability is investigated through parametric analysis, by varying the period and the strain-hardening stiffness. Desirable characteristics such as efficiency, sufficiency, and statistical independence are examined. The proposed intensity measure is contrasted to another one, with respect to its correlation to the engineering demand parameters. An approximate procedure for estimating the optimum normalized spectral area is also presented. It is demonstrated that the proposed intensity measure can be used in intensity-based assessments, and, with proper selection of ground motions, in scenario-based assessments

SUPPORT Tools for evidence-informed health Policymaking (STP) 5: Using research evidence to frame options to address a problem

This article is part of a series written for people responsible for making decisions about health policies and programmes and for those who support these decision makers

Effect of a micronutrient-rich snack taken preconceptionally and throughout pregnancy on ultrasound measures of fetal growth: The Mumbai Maternal Nutrition Project (MMNP)

Improving micronutrient intakes of under‐nourished mothers in low‐ and middle‐income countries increases birth weight, but there is little data on the nature and timing during gestation of any effects on fetal growth. Ultrasound measures of fetal size were used to determine whether and when a food‐based supplement affected fetal growth. Non‐pregnant women living in Mumbai slums, India (N = 6,513), were randomly assigned to receive either a daily micronutrient‐rich snack containing green leafy vegetables, fruit, and milk (treatment) or a snack made from lower‐micronutrient vegetables (control) in addition to their usual diet from before pregnancy until delivery. From 2,291 pregnancies, the analysis sample comprised 1,677 fetuses (1,335 fetuses of women supplemented for ≥3 months before conception). First‐trimester (median: 10 weeks, interquartile range: 9–12 weeks) fetal crown‐rump length was measured. Fetal head circumference, biparietal diameter, femur length, and abdominal circumference were measured during the second (19, 19–20 weeks) and third trimesters (29, 28–30 weeks). The intervention had no effect on fetal size or growth at any stage of pregnancy. In the second trimester, there were interactions between parity and allocation group for biparietal diameter (p = .02) and femur length (p = .04) with both being smaller among fetuses of primiparous women and larger among those of multiparous women, in the treatment group compared with the controls. Overall, a micronutrient‐rich supplement did not increase standard ultrasound measures of fetal size and growth at any stage of pregnancy. Additional ultrasound measures of fetal soft tissues (fat and muscle) may be informative

A Differential Drug Screen for Compounds That Select Against Antibiotic Resistance

Antibiotics increase the frequency of resistant bacteria by providing them a competitive advantage over sensitive strains. Here, we develop a versatile assay for differential chemical inhibition of competing microbial strains, and use it to identify compounds that preferentially inhibit tetracycline-resistant relative to sensitive bacteria, thus “inverting” selection for resistance. Our assay distinguishes compounds selecting directly against specific resistance mechanisms and compounds whose selection against resistance is based on their physiological interaction with tetracycline and is more general with respect to resistance mechanism. A pilot screen indicates that both types of selection-inverting compounds are secreted by soil microbes, suggesting that nature has evolved a repertoire of chemicals that counteracts antibiotic resistance. Finally, we show that our assay can more generally permit simple, direct screening for drugs based on their differential activity against different strains or targets

Modulation of enhancer looping and differential gene targeting by Epstein-Barr virus transcription factors directs cellular reprogramming

Epstein-Barr virus (EBV) epigenetically reprogrammes B-lymphocytes to drive immortalization and facilitate viral persistence. Host-cell transcription is perturbed principally through the actions of EBV EBNA 2, 3A, 3B and 3C, with cellular genes deregulated by specific combinations of these EBNAs through unknown mechanisms. Comparing human genome binding by these viral transcription factors, we discovered that 25% of binding sites were shared by EBNA 2 and the EBNA 3s and were located predominantly in enhancers. Moreover, 80% of potential EBNA 3A, 3B or 3C target genes were also targeted by EBNA 2, implicating extensive interplay between EBNA 2 and 3 proteins in cellular reprogramming. Investigating shared enhancer sites neighbouring two new targets (WEE1 and CTBP2) we discovered that EBNA 3 proteins repress transcription by modulating enhancer-promoter loop formation to establish repressive chromatin hubs or prevent assembly of active hubs. Re-ChIP analysis revealed that EBNA 2 and 3 proteins do not bind simultaneously at shared sites but compete for binding thereby modulating enhancer-promoter interactions. At an EBNA 3-only intergenic enhancer site between ADAM28 and ADAMDEC1 EBNA 3C was also able to independently direct epigenetic repression of both genes through enhancer-promoter looping. Significantly, studying shared or unique EBNA 3 binding sites at WEE1, CTBP2, ITGAL (LFA-1 alpha chain), BCL2L11 (Bim) and the ADAMs, we also discovered that different sets of EBNA 3 proteins bind regulatory elements in a gene and cell-type specific manner. Binding profiles correlated with the effects of individual EBNA 3 proteins on the expression of these genes, providing a molecular basis for the targeting of different sets of cellular genes by the EBNA 3s. Our results therefore highlight the influence of the genomic and cellular context in determining the specificity of gene deregulation by EBV and provide a paradigm for host-cell reprogramming through modulation of enhancer-promoter interactions by viral transcription factors

Anti-malarial activity of Holarrhena antidysenterica and Viola canescens, plants traditionally used against malaria in the Garhwal region of north-west Himalaya

<p>Abstract</p> <p>Background</p> <p>The increasing number of multidrug-resistant <it>Plasmodium </it>strains warrants exploration of new anti-malarials. Medicinal plant research has become more important, particularly after the development of Chinese anti-malarial drug artemisnin from <it>Artemisia annua</it>. The present study shows evaluation of anti-malarial effects of two plants commonly used against malaria in the Garhwal region of north-west Himalaya, in order to discover the herbal-based medicine.</p> <p>Methods</p> <p><it>In vitro </it>anti-plasmodial sensitivity of plant extracts was assessed using schizont maturation and parasite lactate dehydrogenase (pLDH) assay. Cytotoxic activities of the examined extracts were determined on L-6 cells of rat skeletal muscle myoblast. The 4-day test for anti-malarial activity against a chloroquine sensitive <it>Plasmodium berghei </it>NK65 strain in Swiss albino mice was used for monitoring <it>in vivo </it>activity of plant extracts.</p> <p>Results</p> <p>Chloroform extract of <it>H. antidysenterica </it>(HA-2) and petroleum ether extract of <it>V. canescens </it>(VC-1) plants significantly reduced parasitaemia in <it>P. berghei </it>infected mice. The extract HA-2 showed <it>in vitro </it>anti-plasmodial activity with its IC₅₀value 5.5 μg/ml using pLDH assay and ED₅₀value 18.29 mg/kg in <it>P. berghei </it>infected Swiss albino mice. Similarly petroleum ether extract of <it>V. canescens </it>(VC-1) showed <it>in vitro </it>anti-plasmodial activity with its IC₅₀value 2.76 μg/ml using pLDH assay and ED₅₀15.8 mg/kg in <it>P. berghei </it>infected mice. The extracts coded as HA-2 at 30 mg/kg and VC-1 at 20 mg/kg exhibited parasite inhibition in mice: 73.2% and 63.0% respectively. Of these two plant extracts, petroleum ether extract of <it>V. canescens </it>was found slightly cytotoxic.</p> <p>Conclusion</p> <p>The present investigation reflects the use of these traditional medicinal plants against malaria and these plants may work as potential source in the development of variety of herbal formulations for the treatment of malaria.</p