Glycosylated proteins preserved over millennia: N-glycan analysis of Tyrolean Iceman, Scythian Princess and Warrior.

An improved understanding of glycosylation will provide new insights into many biological processes. In the analysis of oligosaccharides from biological samples, a strict regime is typically followed to ensure sample integrity. However, the fate of glycans that have been exposed to environmental conditions over millennia has not yet been investigated. This is also true for understanding the evolution of the glycosylation machinery in humans as well as in any other biological systems. In this study, we examined the glycosylation of tissue samples derived from four mummies which have been naturally preserved: - the 5,300 year old "Iceman called Oetzi", found in the Tyrolean Alps; the 2,400 year old "Scythian warrior" and "Scythian Princess", found in the Altai Mountains; and a 4 year old apartment mummy, found in Vienna/Austria. The number of N-glycans that were identified varied both with the age and the preservation status of the mummies. More glycan structures were discovered in the contemporary sample, as expected, however it is significant that glycan still exists in the ancient tissue samples. This discovery clearly shows that glycans persist for thousands of years, and these samples provide a vital insight into ancient glycosylation, offering us a window into the distant past

Analysis of factors influencing the ultrasonic fetal weight estimation

Objective: The aim of our study was the evaluation of sonographic fetal weight estimation taking into consideration 9 of the most important factors of influence on the precision of the estimation. Methods: We analyzed 820 singleton pregnancies from 22 to 42 weeks of gestational age. We evaluated 9 different factors that potentially influence the precision of sonographic weight estimation ( time interval between estimation and delivery, experts vs. less experienced investigator, fetal gender, gestational age, fetal weight, maternal BMI, amniotic fluid index, presentation of the fetus, location of the placenta). Finally, we compared the results of the fetal weight estimation of the fetuses with poor scanning conditions to those presenting good scanning conditions. Results: Of the 9 evaluated factors that may influence accuracy of fetal weight estimation, only a short interval between sonographic weight estimation and delivery (0-7 vs. 8-14 days) had a statistically significant impact. Conclusion: Of all known factors of influence, only a time interval of more than 7 days between estimation and delivery had a negative impact on the estimation

The need to promote behaviour change at the cultural level: one factor explaining the limited impact of the MEMA kwa Vijana adolescent sexual health intervention in rural Tanzania. A process evaluation

Background - Few of the many behavioral sexual health interventions in Africa have been rigorously evaluated. Where biological outcomes have been measured, improvements have rarely been found. One of the most rigorous trials was of the multi-component MEMA kwa Vijana adolescent sexual health programme, which showed improvements in knowledge and reported attitudes and behaviour, but none in biological outcomes. This paper attempts to explain these outcomes by reviewing the process evaluation findings, particularly in terms of contextual factors. Methods - A large-scale, primarily qualitative process evaluation based mainly on participant observation identified the principal contextual barriers and facilitators of behavioural change. Results - The contextual barriers involved four interrelated socio-structural factors: culture (i.e. shared practices and systems of belief), economic circumstances, social status, and gender. At an individual level they appeared to operate through the constructs of the theories underlying MEMA kwa Vijana - Social Cognitive Theory and the Theory of Reasoned Action – but the intervention was unable to substantially modify these individual-level constructs, apart from knowledge. Conclusion - The process evaluation suggests that one important reason for this failure is that the intervention did not operate sufficiently at a structural level, particularly in regard to culture. Recently most structural interventions have focused on gender or/and economics. Complementing these with a cultural approach could address the belief systems that justify and perpetuate gender and economic inequalities, as well as other barriers to behaviour change

18^{18}F-Fluoride and 18^{18}F-Fluorodeoxyglucose Positron Emission Tomography After Transient Ischemic Attack or Minor Ischemic Stroke: Case-Control Study.

BACKGROUND: Combined positron emission tomography (PET) and computed tomography (CT) can assess both anatomy and biology of carotid atherosclerosis. We sought to assess whether $^{18}$F-fluoride or $^{18}$F-fluorodeoxyglucose can identify culprit and high-risk carotid plaque. METHODS AND RESULTS: We performed $^{18}$F-fluoride and $^{18}$F-fluorodeoxyglucose PET/CT in 26 patients after recent transient ischemic attack or minor ischemic stroke: 18 patients with culprit carotid stenosis awaiting carotid endarterectomy and 8 controls without culprit carotid atheroma. We compared standardized uptake values in the clinically adjudicated culprit to the contralateral asymptomatic artery, and assessed the relationship between radiotracer uptake and plaque phenotype or predicted cardiovascular risk (ASSIGN score [Assessing Cardiovascular Risk Using SIGN Guidelines to Assign Preventive Treatment]). We also performed micro PET/CT and histological analysis of excised plaque. On histological and micro PET/CT analysis, $^{18}$F-fluoride selectively highlighted microcalcification. Carotid $^{18}$F-fluoride uptake was increased in clinically adjudicated culprit plaques compared with asymptomatic contralateral plaques (log$_{10}$ standardized uptake value$_{mean}$ 0.29±0.10 versus 0.23±0.11, P=0.001) and compared with control patients (log$_{10}$ standardized uptake value$_{mean}$ 0.29±0.10 versus 0.12±0.11, P=0.001). $^{18}$F-Fluoride uptake correlated with high-risk plaque features (remodeling index [r=0.53, P=0.003], plaque burden [r=0.51, P=0.004]), and predicted cardiovascular risk [r=0.65, P=0.002]). Carotid $^{18}$F-fluorodeoxyglucose uptake appeared to be increased in 7 of 16 culprit plaques, but no overall differences in uptake were observed in culprit versus contralateral plaques or control patients. However, $^{18}$F-fluorodeoxyglucose did correlate with predicted cardiovascular risk (r=0.53, P=0.019), but not with plaque phenotype. CONCLUSIONS: $^{18}$F-Fluoride PET/CT highlights culprit and phenotypically high-risk carotid plaque. This has the potential to improve risk stratification and selection of patients who may benefit from intervention.Dr Vesey and the study were funded by program grants from the British Heart Foundation (PG12/8/29371) and Chest Heart and Stroke Scotland (R13/A147). Dr Jenkins, Vesey, Dweck, and Newby are supported by the British Heart Foundation (FS/14/78/31020, CH/09/002) and the Wellcome Trust (WT103782AIA). Dr Dweck is the recipient of the Sir Jules Thorn Biomedical Research Award 2015. The Wellcome Trust Clinical Research Facility and the Clinical Research Imaging Centre are supported by National Health Service (NHS) Research Scotland (NRS) through NHS Lothian. Dr Beek is supported by the Scottish Imaging Network—a Platform of Scientific Excellence (SINAPSE). Dr Rudd is part-supported by the National Institute for Health Research Cambridge Biomedical Research Centre, the British Heart Foundation, and the Wellcome Trust

Introducing systems approaches

Systems Approaches to Managing Change brings together five systems approaches to managing complex issues, each having a proven track record of over 25 years. The five approaches are: System Dynamics (SD) developed originally in the late 1950s by Jay Forrester Viable Systems Model (VSM) developed originally in the late 1960s by Stafford Beer Strategic Options Development and Analysis (SODA: with cognitive mapping) developed originally in the 1970s by Colin Eden Soft Systems Methodology (SSM) developed originally in the 1970s by Peter Checkland Critical Systems Heuristics (CSH) developed originally in the late 1970s by Werner Ulrich

Selective Inhibitors of Protozoan Protein N-myristoyltransferases as Starting Points for Tropical Disease Medicinal Chemistry Programs

Inhibition of N-myristoyltransferase has been validated pre-clinically as a target for the treatment of fungal and trypanosome infections, using species-specific inhibitors. In order to identify inhibitors of protozoan NMTs, we chose to screen a diverse subset of the Pfizer corporate collection against Plasmodium falciparum and Leishmania donovani NMTs. Primary screening hits against either enzyme were tested for selectivity over both human NMT isoforms (Hs1 and Hs2) and for broad-spectrum anti-protozoan activity against the NMT from Trypanosoma brucei. Analysis of the screening results has shown that structure-activity relationships (SAR) for Leishmania NMT are divergent from all other NMTs tested, a finding not predicted by sequence similarity calculations, resulting in the identification of four novel series of Leishmania-selective NMT inhibitors. We found a strong overlap between the SARs for Plasmodium NMT and both human NMTs, suggesting that achieving an appropriate selectivity profile will be more challenging. However, we did discover two novel series with selectivity for Plasmodium NMT over the other NMT orthologues in this study, and an additional two structurally distinct series with selectivity over Leishmania NMT. We believe that release of results from this study into the public domain will accelerate the discovery of NMT inhibitors to treat malaria and leishmaniasis. Our screening initiative is another example of how a tripartite partnership involving pharmaceutical industries, academic institutions and governmental/non-governmental organisations such as Medical Research Council and Wellcome Trust can stimulate research for neglected diseases

Chapter 1: Introducing systems approaches

The five approaches covered in Systems Approaches to Making Change – System Dynamics (SD) Viable Systems Model (VSM), Strategic Options Development and Analysis (SODA: with cognitive mapping), Soft Systems Methodology (SSM), and Critical Systems Heuristics (CSH) – are introduced. The rationale for their inclusion is described based on their (i) common historic emergence in dealing with complex situations of change and uncertainty, (ii) shared potential and actual constructivist use of the systems idea, and (iii) pedigree of adaptability and versatility of tools in working with other approaches to making change

Delineation of the Innate and Adaptive T-Cell Immune Outcome in the Human Host in Response to Campylobacter jejuni Infection

BACKGROUND: Campylobacter jejuni is the most prevalent cause of bacterial gastroenteritis worldwide. Despite the significant health burden this infection presents, molecular understanding of C. jejuni-mediated disease pathogenesis remains poorly defined. Here, we report the characterisation of the early, innate immune response to C. jejuni using an ex-vivo human gut model of infection. Secondly, impact of bacterial-driven dendritic cell activation on T-cell mediated immunity was also sought. METHODOLOGY: Healthy, control paediatric terminal ileum or colonic biopsy tissue was infected with C. jejuni for 8-12 hours. Bacterial colonisation was followed by confocal microscopy and mucosal innate immune responses measured by ELISA. Marked induction of IFNγ with modest increase in IL-22 and IL-17A was noted. Increased mucosal IL-12, IL-23, IL-1β and IL-6 were indicative of a cytokine milieu that may modulate subsequent T-cell mediated immunity. C. jejuni-driven human monocyte-derived dendritic cell activation was followed by analyses of T cell immune responses utilising flow cytometry and ELISA. Significant increase in Th-17, Th-1 and Th-17/Th-1 double-positive cells and corresponding cytokines was observed. The ability of IFNγ, IL-22 and IL-17 cytokines to exert host defence via modulation of C. jejuni adhesion and invasion to intestinal epithelia was measured by standard gentamicin protection assay. CONCLUSIONS: Both innate and adaptive T cell-immunity to C. jejuni infection led to the release of IFNγ, IL-22 and IL-17A; suggesting a critical role for this cytokine triad in establishing host anti-microbial immunity during the acute and effectors phase of infection. In addition, to their known anti-microbial functions; IL-17A and IL-17F reduced the number of intracellular C. jejuni in intestinal epithelia, highlighting a novel aspect of how IL-17 family members may contribute to protective immunity against C. jejuni

Colocalization of Protein Kinase A with Adenylyl Cyclase Enhances Protein Kinase A Activity during Induction of Long-Lasting Long-Term-Potentiation

The ability of neurons to differentially respond to specific temporal and spatial input patterns underlies information storage in neural circuits. One means of achieving spatial specificity is to restrict signaling molecules to particular subcellular compartments using anchoring molecules such as A-Kinase Anchoring Proteins (AKAPs). Disruption of protein kinase A (PKA) anchoring to AKAPs impairs a PKA-dependent form of long term potentiation (LTP) in the hippocampus. To investigate the role of localized PKA signaling in LTP, we developed a stochastic reaction-diffusion model of the signaling pathways leading to PKA activation in CA1 pyramidal neurons. Simulations investigated whether the role of anchoring is to locate kinases near molecules that activate them, or near their target molecules. The results show that anchoring PKA with adenylyl cyclase (which produces cAMP that activates PKA) produces significantly greater PKA activity, and phosphorylation of both inhibitor-1 and AMPA receptor GluR1 subunit on S845, than when PKA is anchored apart from adenylyl cyclase. The spatial microdomain of cAMP was smaller than that of PKA suggesting that anchoring PKA near its source of cAMP is critical because inactivation by phosphodiesterase limits diffusion of cAMP. The prediction that the role of anchoring is to colocalize PKA near adenylyl cyclase was confirmed by experimentally rescuing the deficit in LTP produced by disruption of PKA anchoring using phosphodiesterase inhibitors. Additional experiments confirm the model prediction that disruption of anchoring impairs S845 phosphorylation produced by forskolin-induced synaptic potentiation. Collectively, these results show that locating PKA near adenylyl cyclase is a critical function of anchoring