Female sex pheromone of Cameraria ohridella Desch. and Dim. (Lepidoptera: Gracillariidae): structure confirmation,synthesis and biological activity of (8E,10Z)-8,10-tetradecadienal and some analogues

Mass spectrometric investigations confirmed the structure of the female produced sex pheromone of the horse-chestnut leafminer Cameraria ohridella Desch. and Dim. to be (8E,IOZ)-8,10-tetradecadienal. Pure samples, prepared in a straightforward synthesis, were highly attractive in field tests and proved to be suitable for monitoring of flight activities and population dynamics. In mixtures with the synthetic pheromone, analogues like 9-tridecynal and 7-dodecynyl formate were shown to reduce trap catches. In electroantennographic experiments, pheromone analogues were less active than the pheromone. 9-Tridecynal was the most EAG active analogue tested, followed by 7-dodecyn-1-yl formate and 7-undecyn-1-yl formate.     

Mapping of a hapten-binding site: molecular modeling and site-directed mutagenesis study of an anti-atrazine antibody

A three-dimensional model of the variable domain of the atrazine-specific Fab fragment K411B was constructed by molecular modeling using known structures of highly homologous immunoglobulins as templates. Molecular dynamic simulations and cross-reactivity data were used to predict residues responsible for the binding of the hapten 4-chloro-6-(isopropylamino)-1,3,5-triazine-2-(6-aminohexanecarboxylic acid) (iPr/Cl/C6) instead of atrazine. Specific binding pockets could be defined for the chlorine, the isopropylamino group and the C6-spacer of the hapten. The influence of various amino acids on hapten binding was investigated by site-directed mutagenesis, and the effect of these mutations was analyzed by capture ELISA using the hapten iPr/Cl/C6 and 4-amino-6-chloro-1,3,5-triazine-2-(6-aminohexanecarboxylic acid) (H/Cl/C6). GlyH100a seems to be important in determining the conformation of the heavy-chain complementarity determining region H3; replacing it with any other residue prevented the binding of the hapten. Altering residues responsible for the binding of the chlorine atom (TrpH33, GluH50 and TyrL96) decreased the affinity significantly. Hapten-spacer recognition can be attributed to the interaction with PheL32; replacing PheL32 by leucine reduced the affinity towards iPr/Cl/C6. A triple mutant Fab fragment (GlnL89Glu, ValH37Ile and GluL3Val) showed an affinity 5-fold greater towards iPr/Cl/C6 compared to the wild-type K411B, as a result of better recognition of the isopropylamino group of iPr/Cl/C6.     

Combining theoretical and experimental approaches to understand the circadian clock

This review is intended as a summary of our work carried out as part of the German Research Association (DFG) Center Program on Circadian Rhythms. Over the last six years, our approach to understanding circadian systems combined theoretical and experimental tools, and Gonyaulax and Neurospora have proven ideal for these efforts. Both of these model organisms demonstrate that even simple circadian systems can have multiple light input pathways and more than one rhythm generator. They have both been used to elaborate basic circadian features in conjunction with formal models. The models introduce the “zeitnehmer,” i.e., a clock-regulated input pathway, to the conceptual framework of circadian systems, and proposes networks of individual feedbacks as the basis for circadian rhythmicity.     

Diversification of cardiac insulin signaling involves the p85 alpha/beta subunits of phosphatidylinositol 3-kinase

Ventricular cardiomyocytes and cardiac tissue of lean and genetically obese (fa/fa) Zucker rats were used 1) to study the role of the p85 regulatory subunit isoforms p85 alpha and p85 beta for insulin signaling through the phosphatidylinositol (PI) 3-kinase pathway, and 2) to elucidate the implications of these mechanisms for cardiac insulin resistance. Western blot analysis of cardiomyocyte lysates revealed expression of p85 alpha and p85 beta but no detectable amounts of the splice variants of p85 alpha. Essentially no p85 alpha subunit of PI 3-kinase was found to be associated with insulin receptor substrate (IRS)-1 or IRS-2 in basal and insulin-stimulated (5 min) cardiomyocytes. Instead, insulin produced a twofold increase in p85 beta associated with IRS-1, leading to a three- to fourfold increase in p85 beta-associated PI 3-kinase activity. This response was significantly reduced in obese animals. Comparable results were obtained in the intact heart after in vivo stimulation. In GLUT-4-containing vesicles, an increased abundance (3.7 +/- 0.7-fold over basal) of p85 alpha was observed after insulin stimulation of lean animals, with no significant effect in the obese group. No p85 beta could be detected in GLUT-4-containing vesicles. Recruitment of the p110 catalytic subunit of PI 3-kinase and a twofold increase in enzyme activity in GLUT-4-containing vesicles by insulin was observed only in lean rats. We conclude that, in the heart, p85 alpha recruits PI 3-kinase activity to GLUT-4 vesicles, whereas p85 beta represents the main regulator of IRS-1- and IRS-2-mediated PI 3-kinase activation. Furthermore, multiple defects of PI 3-kinase activation, involving both the p85 alpha and the p85 beta adaptor subunits, may contribute to cardiac insulin resistance.     

Flow cytometric determination of E-selectin, vascular cell adhesion molecule-1, and intercellular cell adhesion molecule-1 in formaldehyde-fixed endothelial cell monolayers

BACKGROUND: Endothelial cell adhesion molecules are involved in initiation and progression of vascular diseases. The purpose of this study was to determine conditions of fixation and dissociation of human umbilical vein endothelial cell (HUVEC) monolayers that permit a reliable flow cytometric determination of intracellular and surface content of E-selectin, vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1). METHODS: TNFalpha-treated HUVEC monolayers were fixed with 0.5% formaldehyde at the end of the experimental incubation. Subsequently, either the monolayer was trypsinized and thereafter the cells were subjected to indirect fluorescence labeling or the monolayer was first labeled and then dissociated by trypsinization. Cell integrity was assessed by vimentin staining. Total adhesion molecule content was detected in saponin-permeabilized cells. RESULTS: HUVEC integrity was maintained when the fixation time of the monolayer did not exceed 5 min and trypsin/EDTA was used for dissociation. Surface adhesion molecules were partially hydrolyzed by trypsin when trypsinization preceded labeling but antibody binding protected adhesion molecules from degradation. VCAM-1 and E-selectin exhibited substantial trypsin-sensitive surface fractions but surface ICAM-1 was mainly trypsin resistant. Permeabilization with 0.06% saponin allowed the detection of considerable intracellular pools of the investigated adhesion molecules. CONCLUSIONS: The described method permits the reliable determination of surface and intracellular fractions of adhesion molecules in formaldehyde-fixed HUVEC monolayers and may be used for studies on the regulation of adhesion molecule expression.     

On the Subjective Acceptance during Cardiovascular Magnetic Resonance Imaging at 7.0 Tesla

Purpose

This study examines the subjective acceptance during UHF-CMR in a cohort of healthy volunteers who underwent a cardiac MR examination at 7.0T.

Methods

Within a period of two-and-a-half years (January 2012 to June 2014) a total of 165 healthy volunteers (41 female, 124 male) without any known history of cardiac disease underwent UHF-CMR. For the assessment of the subjective acceptance a questionnaire was used to examine the participants experience prior, during and after the UHF-CMR examination. For this purpose, subjects were asked to respond to the questionnaire in an exit interview held immediately after the completion of the UHF-CMR examination under supervision of a study nurse to ensure accurate understanding of the questions. All questions were answered with “yes” or “no” including space for additional comments.

Results

Transient muscular contraction was documented in 12.7% of the questionnaires. Muscular contraction was reported to occur only during periods of scanning with the magnetic field gradients being rapidly switched. Dizziness during the study was reported by 12.7% of the subjects. Taste of metal was reported by 10.1% of the study population. Light flashes were reported by 3.6% of the entire cohort. 13% of the subjects reported side effects/observations which were not explicitly listed in the questionnaire but covered by the question about other side effects. No severe side effects as vomiting or syncope after scanning occurred. No increase in heart rate was observed during the UHF-CMR exam versus the baseline clinical examination.

Conclusions

This study adds to the literature by detailing the subjective acceptance of cardiovascular magnetic resonance imaging examinations at a magnetic field strength of 7.0T. Cardiac MR examinations at 7.0T are well tolerated by healthy subjects. Broader observational and multi-center studies including patient cohorts with cardiac diseases are required to gain further insights into the subjective acceptance of UHF-CMR examinations.     

Mass HIV Treatment and Sex Disparities in Life Expectancy: Demographic Surveillance in Rural South Africa

Background

Women have better patient outcomes in HIV care and treatment than men in sub-Saharan Africa. We assessed—at the population level—whether and to what extent mass HIV treatment is associated with changes in sex disparities in adult life expectancy, a summary metric of survival capturing mortality across the full cascade of HIV care. We also determined sex-specific trends in HIV mortality and the distribution of HIV-related deaths in men and women prior to and at each stage of the clinical cascade.

Methods and Findings

Data were collected on all deaths occurring from 2001 to 2011 in a large population-based surveillance cohort (52,964 women and 45,688 men, ages 15 y and older) in rural KwaZulu-Natal, South Africa. Cause of death was ascertained by verbal autopsy (93% response rate). Demographic data were linked at the individual level to clinical records from the public sector HIV treatment and care program that serves the region. Annual rates of HIV-related mortality were assessed for men and women separately, and female-to-male rate ratios were estimated in exponential hazard models. Sex-specific trends in adult life expectancy and HIV-cause-deleted adult life expectancy were calculated. The proportions of HIV deaths that accrued to men and women at different stages in the HIV cascade of care were estimated annually.Following the beginning of HIV treatment scale-up in 2004, HIV mortality declined among both men and women. Female adult life expectancy increased from 51.3 y (95% CI 49.7, 52.8) in 2003 to 64.5 y (95% CI 62.7, 66.4) in 2011, a gain of 13.2 y. Male adult life expectancy increased from 46.9 y (95% CI 45.6, 48.2) in 2003 to 55.9 y (95% CI 54.3, 57.5) in 2011, a gain of 9.0 y. The gap between female and male adult life expectancy doubled, from 4.4 y in 2003 to 8.6 y in 2011, a difference of 4.3 y (95% CI 0.9, 7.6). For women, HIV mortality declined from 1.60 deaths per 100 person-years (95% CI 1.46, 1.75) in 2003 to 0.56 per 100 person-years (95% CI 0.48, 0.65) in 2011. For men, HIV-related mortality declined from 1.71 per 100 person-years (95% CI 1.55, 1.88) to 0.76 per 100 person-years (95% CI 0.67, 0.87) in the same period. The female-to-male rate ratio for HIV mortality declined from 0.93 (95% CI 0.82–1.07) in 2003 to 0.73 (95% CI 0.60–0.89) in 2011, a statistically significant decline (p = 0.046). In 2011, 57% and 41% of HIV-related deaths occurred among men and women, respectively, who had never sought care for HIV in spite of the widespread availability of free HIV treatment. The results presented here come from a poor rural setting in southern Africa with high HIV prevalence and high HIV treatment coverage; broader generalizability is unknown. Additionally, factors other than HIV treatment scale-up may have influenced population mortality trends.

Conclusions

Mass HIV treatment has been accompanied by faster declines in HIV mortality among women than men and a growing female–male disparity in adult life expectancy at the population level. In 2011, over half of male HIV deaths occurred in men who had never sought clinical HIV care. Interventions to increase HIV testing and linkage to care among men are urgently needed.