Quantitative Chemically-Specific Coherent Diffractive Imaging of Buried Interfaces using a Tabletop EUV Nanoscope

Characterizing buried layers and interfaces is critical for a host of applications in nanoscience and nano-manufacturing. Here we demonstrate non-invasive, non-destructive imaging of buried interfaces using a tabletop, extreme ultraviolet (EUV), coherent diffractive imaging (CDI) nanoscope. Copper nanostructures inlaid in SiO2 are coated with 100 nm of aluminum, which is opaque to visible light and thick enough that neither optical microscopy nor atomic force microscopy can image the buried interfaces. Short wavelength (29 nm) high harmonic light can penetrate the aluminum layer, yielding high-contrast images of the buried structures. Moreover, differences in the absolute reflectivity of the interfaces before and after coating reveal the formation of interstitial diffusion and oxidation layers at the Al-Cu and Al-SiO2 boundaries. Finally, we show that EUV CDI provides a unique capability for quantitative, chemically-specific imaging of buried structures, and the material evolution that occurs at these buried interfaces, compared with all other approaches.Comment: 12 pages, 8 figure

Xenoestrogenic activity in blood of European and Inuit populations.

Human exposure to persistent organic pollutants (POPs) is ubiquitous and found in all individuals. Studies have documented endocrine disrupting effects and impact on reproduction. The aim of the present study was to compare the level of xenoestrogenic activity in serum of groups with varying POP exposure, and to evaluate correlations to the POP biomarkers, 2,2',4,4',5,5'-hexachlorobiphenyl (CB-153) and 1,1-dichloro-2,2-bis (p-chlorophenyl)-ethylene (p,p'-DDE). No strong consistent association between xenoestrogenic net activity and the two POP markers was found. The results showed that the selected POP markers alone can not predict the integrated xenoestrogenic serum activity. Correlations to the POP markers were found at the extreme edge; the Inuit's and Warsaw study groups eliciting high frequency of samples with ER antagonistic and agonistic activity, respectively. We suggest that the variation in xenoestrogenic serum activity reflects differences in POP exposure mixture, genetic factors and/or life style factors

Investigating the biological properties of carbohydrate derived fulvic acid (CHD-FA) as a potential novel therapy for the management of oral biofilm infections.

Background: A number of oral diseases, including periodontitis, derive from microbial biofilms and are associated with increased antimicrobial resistance. Despite the widespread use of mouthwashes being used as adjunctive measures to control these biofilms, their prolonged use is not recommended due to various side effects. Therefore, alternative broad-spectrum antimicrobials that minimise these effects are highly sought after. Carbohydrate derived fulvic acid (CHD-FA) is an organic acid which has previously demonstrated to be microbiocidal against Candida albicans biofilms, therefore, the aims of this study were to evaluate the antibacterial activity of CHD-FA against orally derived biofilms and to investigate adjunctive biological effects.<p></p> Methods: Minimum inhibitory concentrations were evaluated for CHD-FA and chlorhexidine (CHX) against a range of oral bacteria using standardised microdilution testing for planktonic and sessile. Scanning electron microscopy was also employed to visualise changes in oral biofilms after antimicrobial treatment. Cytotoxicity of these compounds was assessed against oral epithelial cells, and the effect of CHD-FA on host inflammatory markers was assessed by measuring mRNA and protein expression.<p></p> Results: CHD-FA was highly active against all of the oral bacteria tested, including Porphyromonas gingivalis, with a sessile minimum inhibitory concentration of 0.5%. This concentration was shown to kill multi-species biofilms by approximately 90%, levels comparable to that of chlorhexidine (CHX). In a mammalian cell culture model, pretreatment of epithelial cells with buffered CHD-FA was shown to significantly down-regulate key inflammatory mediators, including interleukin-8 (IL-8), after stimulation with a multi-species biofilm.<p></p> Conclusions: Overall, CHD-FA was shown to possess broad-spectrum antibacterial activity, with a supplementary function of being able to down-regulate inflammation. These properties offer an attractive spectrum of function from a naturally derived compound, which could be used as an alternative topical treatment strategy for oral biofilm diseases. Further studies in vitro and in vivo are required to determine the precise mechanism by which CHD-FA modulates the host immune response.<p></p&gt

Triosephosphate Isomerase Gene Characterization and Potential Zoonotic Transmission of Giardia duodenalis

To address the source of infection in humans and public health importance of Giardia duodenalis parasites from animals, nucleotide sequences of the triosephosphate isomerase (TPI) gene were generated for 37 human isolates, 15 dog isolates, 8 muskrat isolates, 7 isolates each from cattle and beavers, and 1 isolate each from a rat and a rabbit. Distinct genotypes were found in humans, cattle, beavers, dogs, muskrats, and rats. TPI and small subunit ribosomal RNA (SSU rRNA) gene sequences of G. microti from muskrats were also generated and analyzed. Phylogenetic analysis on the TPI sequences confirmed the formation of distinct groups. Nevertheless, a major group (assemblage B) contained most of the human and muskrat isolates, all beaver isolates, and the rabbit isolate. These data confirm that G. duodenalis from certain animals can potentially infect humans and should be useful in the detection, differentiation, and taxonomy of Giardia spp

ALKBH3 partner ASCC3 mediates P-body formation and selective clearance of MMS-induced 1-methyladenosine and 3-methylcytosine from mRNA

BACKGROUND: Reversible enzymatic methylation of mammalian mRNA is widespread and serves crucial regulatory functions, but little is known to what degree chemical alkylators mediate overlapping modifications and whether cells distinguish aberrant from canonical methylations. METHODS: Here we use quantitative mass spectrometry to determine the fate of chemically induced methylbases in the mRNA of human cells. Concomitant alteration in the mRNA binding proteome was analyzed by SILAC mass spectrometry. RESULTS: MMS induced prominent direct mRNA methylations that were chemically identical to endogenous methylbases. Transient loss of 40S ribosomal proteins from isolated mRNA suggests that aberrant methylbases mediate arrested translational initiation and potentially also no-go decay of the affected mRNA. Four proteins (ASCC3, YTHDC2, TRIM25 and GEMIN5) displayed increased mRNA binding after MMS treatment. ASCC3 is a binding partner of the DNA/RNA demethylase ALKBH3 and was recently shown to promote disassembly of collided ribosomes as part of the ribosome quality control (RQC) trigger complex. We find that ASCC3-deficient cells display delayed removal of MMS-induced 1-methyladenosine (m CONCLUSIONS: Our findings conform to a model in which ASCC3-mediated disassembly of collided ribosomes allows demethylation of aberrant

Development of an in vitro periodontal biofilm model for assessing antimicrobial and host modulatory effects of bioactive molecules

Background: Inflammation within the oral cavity occurs due to dysregulation between microbial biofilms and the host response. Understanding how different oral hygiene products influence inflammatory properties is important for the development of new products. Therefore, creation of a robust host-pathogen biofilm platform capable of evaluating novel oral healthcare compounds is an attractive option. We therefore devised a multi-species biofilm co-culture model to evaluate the naturally derived polyphenol resveratrol (RSV) and gold standard chlorhexidine (CHX) with respect to anti-biofilm and anti-inflammatory properties.<p></p> Methods: An in vitro multi-species biofilm containing <i>S. mitis, F. nucleatum, P. Gingivalis</i> and <i>A. Actinomycetemcomitans</i> was created to represent a disease-associated biofilm and the oral epithelial cell in OKF6-TERT2. Cytotoxicity studies were performed using RSV and CHX. Multi-species biofilms were either treated with either molecule, or alternatively epithelial cells were treated with these prior to biofilm co-culture. Biofilm composition was evaluated and inflammatory responses quantified at a transcriptional and protein level.<p></p> Results: CHX was toxic to epithelial cells and multi-species biofilms at concentrations ranging from 0.01-0.2%. RSV did not effect multi-species biofilm composition, but was toxic to epithelial cells at concentrations greater than 0.01%. In co-culture, CHX-treated biofilms resulted in down regulation of the inflammatory chemokine IL-8 at both mRNA and protein level. RSV-treated epithelial cells in co-culture were down-regulated in the release of IL-8 protein, but not mRNA.<p></p> Conclusions: CHX possesses potent bactericidal properties, which may impact downstream inflammatory mediators. RSV does not appear to have bactericidal properties against multi-species biofilms, however it did appear to supress epithelial cells from releasing inflammatory mediators. This study demonstrates the potential to understand the mechanisms by which different oral hygiene products may influence gingival inflammation, thereby validating the use of a biofilm co-culture model.<p></p&gt

Potential Antiviral Options against SARS-CoV-2 Infection

As of June 2020, the number of people infected with severe acute respiratory coronavirus 2 (SARS-CoV-2) continues to skyrocket, with more than 6.7 million cases worldwide. Both the World Health Organization (WHO) and United Nations (UN) has highlighted the need for better control of SARS-CoV-2 infections. However, developing novel virus-specific vaccines, monoclonal antibodies and antiviral drugs against SARS-CoV-2 can be time-consuming and costly. Convalescent sera and safe-in-man broad-spectrum antivirals (BSAAs) are readily available treatment options. Here, we developed a neutralization assay using SARS-CoV-2 strain and Vero-E6 cells. We identified the most potent sera from recovered patients for the treatment of SARS-CoV-2-infected patients. We also screened 136 safe-in-man broad-spectrum antivirals against the SARS-CoV-2 infection in Vero-E6 cells and identified nelfinavir, salinomycin, amodiaquine, obatoclax, emetine and homoharringtonine. We found that a combination of orally available virus-directed nelfinavir and host-directed amodiaquine exhibited the highest synergy. Finally, we developed a website to disseminate the knowledge on available and emerging treatments of COVID-19

Unique Arrangement of α- and β-Cells in Human Islets of Langerhans

OBJECTIVE: It is generally admitted that the endocrine cell organization in human islets is different from that of rodent islets. However, a clear description of human islet architecture has not yet been reported. The aim of this work was to describe our observations on the arrangement of human islet cells. RESEARCH DESIGN AND METHODS: Human pancreas specimens and isolated islets were processed for histology. Sections were analyzed by fluorescence microscopy after immunostaining for islet hormones and endothelial cells. RESULTS: In small human islets (40-60 mum in diameter), beta-cells had a core position, alpha-cells had a mantle position, and vessels laid at their periphery. In bigger islets, alpha-cells had a similar mantle position but were found also along vessels that penetrate and branch inside the islets. As a consequence of this organization, the ratio of beta-cells to alpha-cells was constantly higher in the core than in the mantle part of the islets, and decreased with increasing islet diameter. This core-mantle segregation of islet cells was also observed in type 2 diabetic donors but not in cultured isolated islets. Three-dimensional analysis revealed that islet cells were in fact organized into trilaminar epithelial plates, folded with different degrees of complexity and bordered by vessels on both sides. In epithelial plates, most beta-cells were located in a central position but frequently showed cytoplasmic extensions between outlying non-beta-cells. CONCLUSIONS: Human islets have a unique architecture allowing all endocrine cells to be adjacent to blood vessels and favoring heterologous contacts between beta- and alpha-cells, while permitting homologous contacts between beta-cells

Social Pedagogy: An Approach Without Fixed Recipes

A historical and theoretical reconstruction of the specificity and peculiarity of the discipline of social pedagogy, as it has developed in Denmark. Social pedagogy takes its departure from the idea that the individual person and the community are complementary but at the same time opposed to each other, so the task of social pedagogy is rebalancing the dynamics between the two. Social pedagogy is also characterised as a discipline with three dimensions: a practical dimension, a theoretical dimension and a professional dimension. The professional’s task is neither to apply theory in practice nor to uphold the usual practice; it is to mediate between theory and practice. The specificity of the discipline gives rise to particular challenges and dilemmas that theorists make understandable and transparent and practitioners have to deal with. A big challenge for social pedagogy is the quest for evidence-based methods that overrides the specificity of the social pedagogical approach. Balancing different forms of knowledge implies that programmes and methods are used as inspiration that can be contained in a social pedagogical approach

Cystathionine beta-synthase mutants exhibit changes in protein unfolding: conformational analysis of misfolded variants in crude cell extracts

Protein misfolding has been proposed to be a common pathogenic mechanism in many inborn errors of metabolism including cystathionine β-synthase (CBS) deficiency. In this work, we describe the structural properties of nine CBS mutants that represent a common molecular pathology in the CBS gene. Using thermolysin in two proteolytic techniques, we examined conformation of these mutants directly in crude cell extracts after expression in E. coli. Proteolysis with thermolysin under native conditions appeared to be a useful technique even for very unstable mutant proteins, whereas pulse proteolysis in a urea gradient had limited values for the study of the majority of CBS mutants due to their instability. Mutants in the active core had either slightly increased unfolding (p.A114V, p.E302K and p.G307S) or extensive unfolding with decreased stability (p.H65R, p.T191M, p.I278T and p.R369C). The extent of the unfolding inversely correlated with the previously determined degree of tetrameric assembly and with the catalytic activity. In contrast, mutants bearing aminoacid substitutions in the C-terminal regulatory domain (p.R439Q and p.D444N) had increased global stability with decreased flexibility. This study shows that proteolytic techniques can reveal conformational abnormalities even for CBS mutants that have activity and/or a degree of assembly similar to the wild-type enzyme. We present here a methodological strategy that may be used in cell lysates to evaluate properties of proteins that tend to misfold and aggregate and that may be important for conformational studies of disease-causing mutations in the field of inborn errors of metabolism