Good genes and the maternal effects of polyandry on offspring reproductive success in the bulb mite.
Genetic benefits are potentially the most robust explanation of the controversial issue of evolutionary maintenance of polyandry, but the unambiguous demonstration of such benefits has been hindered by the possibility of their confusion with maternal effects. Previous research has shown that polyandrous bulb mite females produce daughters with higher fecundity than monandrous females. Here, we investigate whether this effect arises because polyandrous females invest more in their offspring, or because their offspring inherit 'good genes' from their fathers. Females were mated with either one or four (different) males. However, by sterilizing three of the four males with ionizing radiation, we eliminated any chance of sexual selection (in the polyandrous treatment) so that any differences in the female mating regimes must have been owing to maternal effects. Polyandry had no significant effect on daughter fecundity, thus indicating that any previously documented effects must have been genetic. This was further supported by a significant association between fathers' offensive sperm-competitive ability and the fecundity of their daughters. The association with fathers' sperm defensive ability was not significant, and neither was the association between fathers' sperm competitiveness and sons' reproductive success. However, sons of polyandrous females had lower reproductive success than sons of monandrous females. This shows that the maternal effects of polyandry should be taken into account whenever its costs and benefits are being considered.


Efficient encapsulation of theranostic nanoparticles in cell-derived exosomes: leveraging the exosomal biogenesis pathway to obtain hollow gold nanoparticle-hybrids.
Exosomes can be considered natural targeted delivery systems able to carry exogenous payloads, drugs or theranostic nanoparticles (NPs). This work aims to combine the therapeutic capabilities of hollow gold nanoparticles (HGNs) with the unique tumor targeting properties provided by exosomes. Here, we tested different methods to encapsulate HGNs (capable of absorbing light in the NIR region for selective thermal ablation) into murine melanoma cells derived exosomes (B16-F10-exos), including electroporation, passive loading by diffusion, thermal shock, sonication and saponin-assisted loading. These methods gave less than satisfactory results: although internalization of relatively large NPs into B16-F10-exos was achieved by almost all the physicochemical methods tested, only about 15% of the exosomes were loaded with NPs and several of those processes had a negative effect regarding the morphology and integrity of the loaded exosomes. In a different approach, B16-F10 cells were pre-incubated with PEGylated HGNs (PEG-HGNs) in an attempt to incorporate the NPs into the exosomal biogenesis pathway. The results were highly successful: exosomes recovered from the supernatant of the cell culture showed up to 50% of HGNs internalization. The obtained hybrid HGN-exosome vectors were characterized with a battery of techniques to make sure that internalization of HGNs did not affect exosome characteristics compared with other strategies. PEG-HGNs were released through the endosomal-exosome biogenesis pathway confirming that the isolated vesicles were exosomes.


Association of Zinc Finger Antiviral Protein Binding to Viral Genomic RNA with Attenuation of Replication of Echovirus 7.
Previous studies have implicated both zinc finger antiviral protein (ZAP) and oligoadenylate synthetase 3 (OAS3)/RNase L in the attenuation of RNA viruses with elevated CpG and UpA dinucleotides. Mechanisms and interrelationships between these two pathways were investigated using an echovirus 7 (E7) replicon with compositionally modified sequences inserted into the 3' untranslated region. ZAP and OAS3 immunoprecipitation (IP) assays provided complementary data on dinucleotide composition effects on binding. Elevated frequencies of alternative pyrimidine/purine (CpA and UpG) and reversed (GpC and ApU) dinucleotides showed no attenuating effect on replication or specific binding to ZAP by IP. However, the bases 3' and 5' of CpG motifs influenced replication and ZAP binding; UCGU enhanced CpG-mediated attenuation and ZAP binding, while A residues shielded CpGs from ZAP recognition. Attenuating effects of elevated frequencies of UpA on replication occurred independently of CpG dinucleotides and bound noncompetitively with CpG-enriched RNA, consistent with a separate recognition site from CpG. Remarkably, immunoprecipitation with OAS3 antibody reproduced the specific binding to CpG- and UpA-enriched RNA sequences. However, OAS3 and ZAP were coimmunoprecipitated in both ZAP and OAS3 IP and colocalized with E7 and stress granules (SGs) by confocal microscopy analysis of infected cells. ZAP's association with larger cellular complexes may mediate the recruitment of OAS3/RNase L, KHNYN, and other RNA degradation pathways.IMPORTANCE We recently discovered that the OAS3/RNase L antiviral pathway is essential for restriction of CpG- and UpA-enriched viruses, in addition to the requirement for zinc finger antiviral protein (ZAP). The current study provides evidence for the specific dinucleotide and wider recognition contexts associated with virus recognition and attenuation. It further documents the association of ZAP and OAS3 and association with stress granules and a wider protein interactome that may mediate antiviral effects in different cellular compartments. The study provides a striking reconceptualization of the pathways associated with this aspect of antiviral defense.


Ultrasound imaging velocimetry: effect of beam sweeping on velocity estimation.
As an emerging flow-mapping tool that can penetrate deep into optically opaque media such as human tissue, ultrasound imaging velocimetry has promise in various clinical applications. Previous studies have shown that errors occur in velocity estimation, but the causes have not been well characterised. In this study, the error in velocity estimation resulting from ultrasound beam sweeping in image acquisition is quantitatively investigated. The effects on velocity estimation of the speed and direction of beam sweeping relative to those of the flow are studied through simulation and experiment. The results indicate that a relative error in velocity estimation of up to 20% can be expected. Correction methods to reduce the errors under steady flow conditions are proposed and evaluated. Errors in flow estimation under unsteady flow are discussed.


Laboratory Value Effectiveness in Predicting Early Postoperative Periprosthetic Joint Infection After Total Hip Arthroplasty.
Diagnosing early periprosthetic joint infection (PJI) after primary total hip arthroplasty (THA) remains challenging. We sought to validate optimal laboratory value cutoffs for detecting early PJIs in a series of primary THAs from one institution. We retrospectively identified 22,795 primary THAs performed between 2000 and 2019. Within 12 weeks, 43 hips (43 patients) underwent arthrocentesis. Patients were divided into 2 groups: evaluation ≤6 weeks or 6-12 weeks following THA. The 2011 Musculoskeletal Infection Society major criteria for PJI diagnosed PJI in 15 patients. Mann-Whitney U-tests were used to compare median laboratory values and receiver operating characteristic curve analysis was used to evaluate optimal cutoff values. Both within 6 weeks and between 6 and 12 weeks postoperatively, median C-reactive protein (CRP), erythrocyte sedimentation rate, synovial white blood cell (WBC) count, neutrophil percentage, and absolute neutrophil count (ANC) values were significantly higher in infected THAs. Optimal cutoffs within 6 weeks were: CRP ≥100 mg/L, synovial WBCs ≥4390 cells/μL, neutrophil percentage ≥74%, and ANC ≥3249 cells/μL. Between 6 and 12 weeks, optimal cutoffs were: CRP ≥33 mg/L, synovial WBCs ≥26,995 cells/μL, neutrophil percentage ≥93%, and ANC ≥25,645 cells/μL. Early PJI following THA should be suspected within 6 weeks with CRP ≥100 mg/L or synovial WBCs ≥4390 cells/μL. Between 6 and 12 weeks postoperatively, cutoffs of CRP ≥33 mg/L, synovial fluid WBC ≥26,995 cells/μL, and neutrophil percentage ≥93% diagnosed PJI with high accuracy. Level IV Diagnostic.


Palaeococcus ferrophilus gen. nov., sp. nov., a barophilic, hyperthermophilic archaeon from a deep-sea hydrothermal vent chimney.
A novel barophilic, hyperthermophilic archaeon was isolated from a deep-sea hydrothermal vent chimney at the Myojin Knoll in the Ogasawara-Bonin Arc, Japan. The cells were found to be irregular cocci and motile with multiple polar flagella. Growth was observed between 60 and 88 degrees C (opt. 83 degrees C; 30 min doubling time), pH 4.0 and 8.0 (opt. pH 6.0), 20 and 73 g sea salts l-1 (opt. 47 g l-1) and 0.1 and 60 MPa (opt. 30 MPa). The isolate was a strictly anaerobic chemoorganotroph capable of utilizing proteinaceous substrates such as yeast extract, peptone, tryptone and casein in the presence of elemental sulfur or ferrous iron. The G + C content of the genomic DNA was 53.5 mol%. Phylogenetic analysis based on 16S rDNA sequences indicated that the isolate was a member of an ancient lineage of the Thermococcales that diverged prior to the formation of the two genera Thermococcus and Pyrococcus. On the basis of the physiological and molecular properties of the new isolate, the name Palaeococcus ferrophilus gen. nov., sp. nov. is proposed. The type strain is strain DMJT (= JCM 10417) [corrected].


Evidence suggesting that galanin (GAL), melanin-concentrating hormone (MCH), neurotensin (NT), proopiomelanocortin (POMC) and neuropeptide Y (NPY) are targets of leptin signaling in the hypothalamus.
Leptin (OB protein) reduces food intake by acting at the hypothalamic level. The purpose of the present study was to identify potential targets of leptin signaling in the hypothalamus in ad-lib fed rats. Central administration of leptin (5 microg) for 3 days decreased food intake and body weight gain in association with a decrease in hypothalamic galanin (GAL), melanin-concentrating hormone (MCH), proopiomelanocortin (POMC) and neuropeptide Y (NPY) gene expression and with an increase in neurotensin (NT) gene expression. In pair-fed rats, NPY gene expression was increased and there was no change in either MCH, GAL, POMC or NT gene expression. This study identifies GAL, MCH, POMC and NT as non-NPY targets of leptin signaling and suggests that leptin's action on food intake and body weight is most likely mediated by inhibiting excitatory (e.g. NPY, MCH, GAL, POMC) and stimulating inhibitory (e.g., NT) signals in the feeding circuitry.


1,3-Dimethyl-3-silapiperidine: synthesis, molecular structure, and conformational analysis by gas-phase electron diffraction, low temperature NMR, IR and Raman spectroscopy, and quantum chemical calculations.
The first Si-H-containing azasilaheterocycle, 1,3-dimethyl-3-silapiperidine 1, was synthesized, and its molecular structure and conformational properties were studied by gas-phase electron diffraction (GED), low temperature NMR, IR and Raman spectroscopy and quantum chemical calculations. The compound exists as a mixture of two conformers possessing the chair conformation with the equatorial NMe group and differing by axial or equatorial position of the SiMe group. In the gas phase, the SiMe(ax) conformer predominates (GED: ax/eq = 65(7):35(7)%, ΔG = 0.36(18) kcal/mol; IR: ax/eq = 62(5):38(5)%, ΔG = 0.16(7) kcal/mol). In solution, at 143 K the SiMe(eq) conformer predominates in the frozen equilibrium (NMR: ax/eq = 31.5(1.5):68.5(1.5)%, ΔG = -0.22(2) kcal/mol). Thermodynamic parameters of the ring inversion are determined (ΔG(‡) = 8.9-9.0 kcal/mol, ΔH(‡) = 9.6 kcal/mol, ΔS(‡) = 2.1 eu). High-level quantum chemical calculations (MP2, G2, CCSD(T)) nicely reproduce the experimental geometry and the predominance of the axial conformer in the gas phase.


Carbohydrate metabolism in the isolated perfused rat kidney.
1. Anaerobic formation of lactate from glucose by isolated perfused rat kidney (411mumol/h per g dry wt.) was three times as fast as in aerobic conditions (138mumol/h per g). 2. In aerobic or in anaerobic conditions, the ratio of lactate production to glucose utilization was about 2. 3. Starvation or acidosis caused a decline of about 30% in the rate of aerobic glycolysis. 4. The rate of formation of glucose from lactate by perfused kidney from a well-fed rat, in the presence of 5mm-acetoacetate (83mumol/h per g dry wt.), was of the same order as the rate of aerobic glycolysis. 5. During perfusion with physiological concentrations of glucose (5mm) and lactate (2mm) there were negligible changes in the concentration of either substrate. 6. Comparison of kidneys perfused with lactate, from well-fed or starved rats, showed no major differences in contents of intermediates of gluconeogenesis. 7. The tissue concentrations of hexose monophosphates and C(3) phosphorylated glycolytic intermediates (except triose phosphate) were decreased in anaerobic conditions. 8. Aerobic metabolism of fructose by perfused kidney was rapid: the rate of glucose formation was 726mumol/h per g dry wt. and of lactate formation 168mumol/h per g (dry wt.). Glycerol and d-glyceraldehyde were also released into the medium. 9. Aerobically, fructose generated high concentrations of glycolytic intermediates. 10. Anaerobic production of lactate from fructose (74mumol/h per g dry wt.) was slower than the aerobic rate. 11. In both anaerobic and aerobic conditions the ratio [lactate]/[pyruvate] in kidney or medium was lower during perfusion with fructose than with glucose. 12. These results are discussed in terms of the regulation of renal carbohydrate metabolism.


Chromatin immunoprecipitation from fixed clinical tissues reveals tumor-specific enhancer profiles.
Extensive cross-linking introduced during routine tissue fixation of clinical pathology specimens severely hampers chromatin immunoprecipitation followed by next-generation sequencing (ChIP-seq) analysis from archived tissue samples. This limits the ability to study the epigenomes of valuable, clinically annotated tissue resources. Here we describe fixed-tissue chromatin immunoprecipitation sequencing (FiT-seq), a method that enables reliable extraction of soluble chromatin from formalin-fixed paraffin-embedded (FFPE) tissue samples for accurate detection of histone marks. We demonstrate that FiT-seq data from FFPE specimens are concordant with ChIP-seq data from fresh-frozen samples of the same tumors. By using multiple histone marks, we generate chromatin-state maps and identify cis-regulatory elements in clinical samples from various tumor types that can readily allow us to distinguish between cancers by the tissue of origin. Tumor-specific enhancers and superenhancers that are elucidated by FiT-seq analysis correlate with known oncogenic drivers in different tissues and can assist in the understanding of how chromatin states affect gene regulation.


Hydrocarbon-degrading bacteria enriched by the Deepwater Horizon oil spill identified by cultivation and DNA-SIP.
The massive influx of crude oil into the Gulf of Mexico during the Deepwater Horizon (DWH) disaster triggered dramatic microbial community shifts in surface oil slick and deep plume waters. Previous work had shown several taxa, notably DWH Oceanospirillales, Cycloclasticus and Colwellia, were found to be enriched in these waters based on their dominance in conventional clone and pyrosequencing libraries and were thought to have had a significant role in the degradation of the oil. However, this type of community analysis data failed to provide direct evidence on the functional properties, such as hydrocarbon degradation of organisms. Using DNA-based stable-isotope probing with uniformly (13)C-labelled hydrocarbons, we identified several aliphatic (Alcanivorax, Marinobacter)- and polycyclic aromatic hydrocarbon (Alteromonas, Cycloclasticus, Colwellia)-degrading bacteria. We also isolated several strains (Alcanivorax, Alteromonas, Cycloclasticus, Halomonas, Marinobacter and Pseudoalteromonas) with demonstrable hydrocarbon-degrading qualities from surface slick and plume water samples collected during the active phase of the spill. Some of these organisms accounted for the majority of sequence reads representing their respective taxa in a pyrosequencing data set constructed from the same and additional water column samples. Hitherto, Alcanivorax was not identified in any of the previous water column studies analysing the microbial response to the spill and we discuss its failure to respond to the oil. Collectively, our data provide unequivocal evidence on the hydrocarbon-degrading qualities for some of the dominant taxa enriched in surface and plume waters during the DWH oil spill, and a more complete understanding of their role in the fate of the oil.


Food Service Pork Chops from Three U.S. Regions Harbor Similar Levels of Antimicrobial Resistance Regardless of Antibiotic Use Claims.
Pork products from animals "raised without antibiotics" (RWA) are assumed to harbor lower levels of antimicrobial resistance (AMR) than conventional (CONV) pork products with no claims regarding use of antimicrobial agents during production. A total of 372 pork chop samples from CONV (n = 190) and RWA (n = 182) production systems were collected over 13 months from three food service suppliers. The following bacteria were cultured: Escherichia coli, tetracycline-resistant (TETr) E. coli, third-generation cephalosporin-resistant (3GCr) E. coli, Salmonella enterica, TETr Salmonella, 3GCr Salmonella, nalidixic acid-resistant Salmonella, Enterococcus spp., TETr Enterococcus, erythromycin-resistant Enterococcus, Staphylococcus aureus, and methicillin-resistant S. aureus. Production system did not significantly impact the detection of cultured bacteria (P > 0.05). Metagenomic DNA was isolated from each sample, and equal amounts of metagenomic DNA were pooled by supplier, month, and production system for 75 pooled samples (38 CONV, 37 RWA). Quantitative PCR was used to assess the abundances of the following 10 AMR genes: aac(6')-Ie-aph(2″)-Ia, aadA1, blaCMY-2, blaCTX-M, blaKPC-2, erm(B), mecA, tet(A), tet(B), and tet(M). For all 10 AMR genes, abundances did not differ significantly (P > 0.05) between production systems. These results suggest that use of antimicrobial agents during swine production minimally impacts the AMR of bacteria in pork chops.


A Novel Nanocomposite of Activated Serpentine Mineral Decorated with Magnetic Nanoparticles for Rapid and Effective Adsorption of Hazardous Cationic Dyes: Kinetics and Equilibrium Studies.
A widely distributed mineral, serpentine, obtained from Wadi Ghadir (Eastern Desert in Egypt) was studied as a potential naturally and abundantly available source for the synthesis of an efficient adsorbent for aquatic remediation applications. A novel nanocomposite was synthesized after the exfoliation of the layered structure of serpentine by hydrogen peroxide treatment (serpentine (SP)), followed by decoration with magnetic Fe3O4 nanoparticles (MNP). The goal behind the utilization of the latter phase was to increase the environmental remediation capability and to incorporate magnetic properties at the final adsorbent, toward a better separation after the use. The fabricated composite (MNP/SP) was characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and transmission electron microscopy (TEM). The composite's potential adsorption application toward the removal of two cationic dyes, methylene blue (MB) and malachite green (MG), was investigated. The observed adsorption kinetics was fast, and the highest uptake was observed at pH = 8, with the capacities to reach 162 and 176 mg g-1 for MB and MG, respectively, values significantly higher than various other materials tested against these two cationic dyes. Compared to hydrogen peroxide-treated serpentine, the removal efficiency of the composite was higher by 157 and 127% for MB and MG, respectively. The MB and MG were adsorbed because of the favorable electrostatic interactions between MNP/SP active sites and the cationic dyes. The close value capacities suggest that the difference in chemistry of the two dyes does not affect the interactions, with the later occurring via the dyes' amine functionalities. With increasing ionic strength, the adsorption of the studied basic dyes was slightly decreased, suggesting only partial antagonistic ion effect. The sorbent can be easily regenerated and reused without significant deterioration of its adsorption efficiency, which makes MNP/SP a promising adsorbent for the removal of hazardous pollutants from aquatic environments.


Saponins from Sea Cucumber and Their Biological Activities.
Sea cucumbers, belonging to the phylum Echinodermata, have been valued for centuries as a nutritious and functional food with various bioactivities. Sea cucumbers can produce highly active substances, notably saponins, the main secondary metabolites, which are the basis of their chemical defense. The saponins are mostly triterpene glycosides with triterpenes or steroid in aglycone, which possess multiple biological properties including antitumor, hypolipidemic activity, improvement of nonalcoholic fatty liver, inhibition of fat accumulation, antihyperuricemia, promotion of bone marrow hematopoiesis, antihypertension, etc. Sea cucumber saponins have received attention due to their rich sources, low toxicity, high efficiency, and few side effects. This review summarizes current research on the structure and activities of sea cucumber saponins based on the physiological and pharmacological activities from source, experimental models, efficacy, and mechanisms, which may provide a valuable reference for the development of sea cucumber saponins.


Effect of oral N-acetyl cysteine on eradication of Helicobacter pylori in patients with dyspepsia.
Using mucolytic agents that decrease viscosity of the gastric mucous and therefore, increase the permeability of antibiotics through gastric membrane has been offered as an additive treatment to achieve a higher rate of eradication of Helicobacter pylori (H. Pylori) infection. The aim of this study was to determine the efficacy of oral N-acetyl cysteine (NAC) on eradication of H. pylori infections in patients suffering from dyspepsia. In this randomized double-blinded clinical trial, 60 H. pylori positive patients who were suffering from dyspepsia were included. They were divided into two groups. Both groups received three-drug regimen including pantoprazole 40 mg BD, ciprofloxacin 500 mg BD and bismuth subcitrate 120 mg two tablets BD. Experimental group (30 cases) received 600 mg of NAC besides three-drug regimen. Control group received placebo. The results of therapy were tested by 14C-UBT and were compared with each other two months after the first visit. H. pylori infection was eradicated in 21 (70%) and 17 (60.7%) patients in experimental and control groups, respectively (P=0.526). Regarding clinical and endoscopic variables, no significant difference was observed between the two groups except for erosive gastritis (0.041) and erosive esophagitis (0.031). Our findings offer that NAC has an additive effect on H. pylori triple therapy with pantoprazole, ciprofloxacin and bismuth subcitrate. Although NAC does not have any known activity against H. pylori, it can reduce the thickness of the mucus layer and increase the permeability of antibiotics at the site of infection. To evaluate this effect, more studies with larger sample size should be performed.


Ultrastructural changes in blood vessels of perinephritic hypertensive dogs.
Mesenteric arteries and saphenous veins of control and perinephritic hypertensive dogs were studied with light and electron microscopy. 2 weeks after surgery there are ultrastructural changes, both in arteries and veins which are thought to be of hydroelectrolytic origin. These alterations are more marked 7 weeks after surgery and disappear thereafter. There are signs of increased protein synthesis activity in the smooth muscle of the mesenteric arteries, but not of the saphenous veins obtained 7 and 14 weeks after surgery, leading to a progressive hypertrophy of the muscular cells. The connective tissue augmented proportionally. Degeneration of adrenergic nerves in the arteries, accompanied by a parallel decrease in their noradrenaline content, constituted an unexpected finding clearly deserving further study.


Using calcium chloride injection to improve tenderness of beef from mature cows.
The objective of this investigation was to determine the effect of calcium chloride (CaCl2) injection on Warner-Bratzler shear force (WBS), sensory panel ratings, and collagen traits of mature cow beef. Within 30 min of exsanguination, subprimals (top round, TR; top sirloin, TS; strip loin, SL) from alternate sides of the carcass were injected with a .3 M CaCl2 solution (10% of the subprimal weight) and aged for 1, 7, or 14 d. The corresponding cold-boned cuts of the other side served as a control. Injecting CaCl2 eliminated the requirement for extended postmortem storage, as indicated by d 1 WBS. During the 14-d aging period, WBS of noninjected cuts decreased by 2.59 kg, whereas WBS of CaCl2-injected samples decreased by only .35 kg. Compared with control cuts, CaCl2 injection improved (P less than .05) d-14 WBS of steaks from SL, TS, and TR by 41.1, 40.1, and 15.3%, respectively. Additionally, CaCl2-injected subprimals exhibited higher (P less than .05) sensory panel tenderness ratings, lower (P less than .05) amounts of detectable connective tissue, and shorter (P less than .05) sarcomere lengths. No differences (P greater than .05) were observed in any quantitative collagen traits between CaCl2-injected and control cuts. These results indicate that CaCl2 injection improved ultimate tenderness and sensory ratings of meat from mature cow cuts.


Pests, diseases, and aridity have shaped the genome of Corymbia citriodora.
Corymbia citriodora is a member of the predominantly Southern Hemisphere Myrtaceae family, which includes the eucalypts (Eucalyptus, Corymbia and Angophora; ~800 species). Corymbia is grown for timber, pulp and paper, and essential oils in Australia, South Africa, Asia, and Brazil, maintaining a high-growth rate under marginal conditions due to drought, poor-quality soil, and biotic stresses. To dissect the genetic basis of these desirable traits, we sequenced and assembled the 408 Mb genome of Corymbia citriodora, anchored into eleven chromosomes. Comparative analysis with Eucalyptus grandis reveals high synteny, although the two diverged approximately 60 million years ago and have different genome sizes (408 vs 641 Mb), with few large intra-chromosomal rearrangements. C. citriodora shares an ancient whole-genome duplication event with E. grandis but has undergone tandem gene family expansions related to terpene biosynthesis, innate pathogen resistance, and leaf wax formation, enabling their successful adaptation to biotic/abiotic stresses and arid conditions of the Australian continent.


Olfactory marker protein expression is an indicator of olfactory receptor-associated events in non-olfactory tissues.
Olfactory receptor (OR)-associated events are mediated by well-conserved components in the olfactory epithelium, including olfactory G-protein (Golf), adenylate cyclase III (ACIII), and olfactory marker protein (OMP). The expression of ORs has recently been observed in non-olfactory tissues where they are involved in monitoring extracellular chemical cues. The large number of OR genes and their sequence similarities illustrate the need to find an effective and simple way to detect non-olfactory OR-associated events. In addition, expression profiles and physiological functions of ORs in non-olfactory tissues are largely unknown. To overcome limitations associated with using OR as a target protein, this study used OMP with Golf and ACIII as targets to screen for potential OR-mediated sensing systems in non-olfactory tissues. Here, we show using western blotting, real-time PCR, and single as well as double immunoassays that ORs and OR-associated proteins are co-expressed in diverse tissues. The results of immunohistochemical analyses showed OMP (+) cells in mouse heart and in the following cells using the corresponding marker proteins c-kit, keratin 14, calcitonin, and GFAP in mouse tissues: interstitial cells of Cajal of the bladder, medullary thymic epithelial cells of the thymus, parafollicular cells of the thyroid, and Leydig cells of the testis. The expression of ORs in OMP (+) tissues was analyzed using a refined microarray analysis and validated with RT-PCR and real-time PCR. Three ORs (olfr544, olfr558, and olfr1386) were expressed in the OMP (+) cells of the bladder and thyroid as shown using a co-immunostaining method. Together, these results suggest that OMP is involved in the OR-mediated signal transduction cascade with olfactory canonical signaling components between the nervous and endocrine systems. The results further demonstrate that OMP immunohistochemical analysis is a useful tool for identifying expression of ORs, suggesting OMP expression is an indicator of potential OR-mediated chemoreception in non-olfactory systems.


Homocysteine is associated with severity of microvasculopathy in sickle cell disease patients.
The pathophysiology of sickle cell disease (SCD) includes vasculopathy as well as anaemia. Elevated plasma homocysteine is a risk factor for vascular disease and may be associated with increased risk of vascular complications in SCD patients. In the present study, microvascular characteristics were assessed in the bulbar conjunctiva of 18 paediatric and 18 adult SCD patients, using the non-invasive technique of computer-assisted intravital microscopy. A vasculopathy severity index (SI) was computed to quantify the degree of microvasculopathy in each patient. Plasma homocysteine and several of its determinants [serum folate and vitamin B12, plasma pyridoxal-5'-phosphate (vitamin B6 status) and creatinine (kidney function)] were measured. Age was strongly correlated with microvasculopathy in the SCD patients, with the SI increasing about 0·1 unit per one-year increase in age (P < 0·001). After adjusting for age, gender, B-vitamin status and creatinine, homocysteine concentration was directly correlated with severity index (P < 0·05). Age and homocysteine concentration were independent predictors of microvasculopathy in SCD patients. It remains to be determined whether lowering homocysteine concentrations using appropriate B-vitamin supplements (folate and vitamins B12 and B6) - particularly if started early in life - could ameliorate microvasculopathy and its associated complications in SCD patients.


Comparison of intervention methods for reducing human exposure to Mycobacterium bovis through milk in pastoralist households of Tanzania.
Bovine tuberculosis (bTB), caused by Mycobacterium bovis, is a disease of zoonotic concern, especially in countries with no control programs in livestock and where routine pasteurization of milk is not practiced. In Tanzania, bTB is widespread in livestock and has been diagnosed in humans; however, herd bTB testing is primarily carried out for bTB-free certification in commercial dairy herds at the expense of the dairy cattle owner. For rural livestock holders, such an expense is prohibitive, and consequently there is no control of bTB in most areas. Although effective long-term solutions to control bTB in livestock are desirable, there is a need to assess the effect of preventive measures on reducing human exposure to bTB in such settings. We utilized locally relevant cattle herd characteristics and management data from the Health for Animals and Livelihood Improvement (HALI) project in south-central Tanzania to build a Reed-Frost model that compared the efficacy of alternative methods aimed at reducing the exposure of humans to infectious milk from a typical pastoralist cattle herd. During a 10-year simulation period, the model showed that boiling milk 80% of the time is necessary to obtain a reduction in liters of infectious milk approximately equivalent to what would be obtained with a standard 2-year testing and removal regimen, and that boiling milk was more effective than animal test and removal early in the time period. In addition, even with testing and removing infected cattle, a residual risk of exposure to infectious milk remained due to imperfect sensitivity of the skin test and a continuous risk of introduction of infectious animals from other herds. The model was sensitive to changes in initial bTB prevalence but not to changes in herd size. In conclusion, continuous complimentary treatment of milk may be an effective strategy to reduce human exposure to M. bovis-infected milk in settings where bTB is endemic and a comprehensive bTB control program is yet to be implemented.


Validation of a simplified food frequency questionnaire as used in the Nutrition and Health Survey in Taiwan (NAHSIT) for the elderly.
A 28-item simplified food frequency questionnaire (SFFQ) combined with 9 open questions about basic commodities was designed for the Elderly Nutrient and Health Survey in Taiwan (NAHSIT) to collect participants' usual intake of the previous month. We have examined the validity this SFFQ as assessed by multiple 24-hour dietary recall (n=81) and biomarkers (n=1473). All questionnaires were completed by face-to-face interview and fasting blood taken. Thirty seven males and 44 females were randomly selected from NAHSIT participants. Of these, 31 and 50 subjects completed 2 or 3 24-hour dietary recalls within one month, respectively. Mean daily intake frequencies for each food group were calculated from the SFFQ and 24-hr recalls, respectively. Spearman rank correlation coefficients between frequencies of food intake from the FFQ and dietary recalls ranged from 0.132 to 0.678 for men; 0.052 to 0.759 for women. For food weight, the trends were similar to frequency findings. For biomarkers, the most correlated were dairy intake frequency judged by vitamin B-2 and calcium intakes and by erythrocyte glutathione reductase (EGRAC) for B-2 functionality, where the correlation coefficients were, respectively, 0.533, 0.518 and -0.205 for men; 0.494, 0.475 and -0.174 for women; fish and fruit followed in overall validity. The SFFQ measured the food patterns of elderly NAHSIT elders with validity high for dairy and good for fish, and fruit intakes in both genders.


Simvastatin induces proliferation inhibition and apoptosis in C6 glioma cells via c-jun N-terminal kinase.
The lipid-lowering drugs, statins, induce apoptosis in a variety of tumor cells. Here we investigated the apoptotic effect of the lipophilic statin, simvastatin, in C6 glioma cells and the underlying effects on intracellular signal transduction. Simvastatin inhibited cell proliferation totally after 20h of treatment as shown by the decrease in proliferating cell nuclear antigen expression in the nucleus. Subsequently, simvastatin caused apoptotic cell death by shrinkage of cytoplasm and condensation of chromatin, and DNA fragmentation. The features of apoptosis were visible only after 48 h of treatment, possibly reflecting a requirement for cell commitment to growth arrest. In immunocytochemical and immunoblotting experiments we have shown that simvastatin markedly increased the phosphorylation of ATF-2 and c-jun in the nucleus of the C6 glioma cells at early time points which was preserved even 24 h after treatment. In contrast, activities of protein kinases Erk1/2 and AKT in the cell survival pathway remained unchanged throughout the treatment. Selective inhibitor of JNK, but not p38 kinase, reduced simvastatin-induced cell death and ATF-2 and c-jun phosphorylation suggesting that JNK-dependent activation of ATF-2 and c-jun may play an important role in simvastatin-induced proliferation inhibition and apoptosis in C6 glioma cells. These observations suggest that statins may have clinical significance in the prevention of glial tumors beyond their cholesterol-lowering effect and JNK may be a rational target for sensitizing glioma cells to chemotherapeutic agents.


Intercellular transfer of mitochondria via tunneling nanotubes protects against cobalt nanoparticle-induced neurotoxicity and mitochondrial damage.
Broad applications of cobalt nanoparticles (CoNPs) have raised increased concerns regarding their potential toxicity. However, the underlining mechanisms of their toxicity have yet to be characterized. Here, we demonstrated that CoNPs reduced cell viability and induced membrane leakage. CoNPs induced oxidative stress, as indicated by the generation of reactive oxygen species (ROS) secondary to the increased expression of hypoxia-induced factor 1 alpha. Moreover, CoNPs led to mitochondrial damage, including generation of mitochondrial ROS, reduction in ATP content, morphological damage and autophagy. Interestingly, exogenous mitochondria were observed between neurons and astrocytes upon CoNPs exposure. Concomitantly, tunneling nanotubes (TNTs)-like structures were observed between neurons and astrocytes upon CoNPs exposure. These structures were further verified to be TNTs as they were found to be F-actin rich and lacking tubulin. We then demonstrated that TNTs were utilized for mitochondrial transfer between neurons and astrocytes, suggesting a novel crosstalk phenomenon between these cells. Moreover, we found that the inhibition of TNTs (using actin-depolymerizing drug latrunculin B) intensified apoptosis triggered by CoNPs. Therefore, we demonstrate, for the first time, that the inhibition of intercellular mitochondrial transfer via TNTs aggravates CoNPs-induced cellular and mitochondrial toxicity in neuronal cells, implying a novel intercellular protection mechanism in response to nanoparticle exposure.


Partnerships for Global Child Health.
Child mortality remains a global health challenge and has resulted in demand for expanding the global child health (GCH) workforce over the last 3 decades. Institutional partnerships are the cornerstone of sustainable education, research, clinical service, and advocacy for GCH. When successful, partnerships can become self-sustaining and support development of much-needed training programs in resource-constrained settings. Conversely, poorly conceptualized, constructed, or maintained partnerships may inadvertently contribute to the deterioration of health systems. In this comprehensive, literature-based, expert consensus review we present a definition of partnerships for GCH, review their genesis, evolution, and scope, describe participating organizations, and highlight benefits and challenges associated with GCH partnerships. Additionally, we suggest a framework for applying sound ethical and public health principles for GCH that includes 7 guiding principles and 4 core practices along with a structure for evaluating GCH partnerships. Finally, we highlight current knowledge gaps to stimulate further work in these areas. With awareness of the potential benefits and challenges of GCH partnerships, as well as shared dedication to guiding principles and core practices, GCH partnerships hold vast potential to positively impact child health.


Astrocyte glutamine synthetase: pivotal in health and disease.
The multifunctional properties of astrocytes signify their importance in brain physiology and neurological function. In addition to defining the brain architecture, astrocytes are primary elements of brain ion, pH and neurotransmitter homoeostasis. GS (glutamine synthetase), which catalyses the ATP-dependent condensation of ammonia and glutamate to form glutamine, is an enzyme particularly found in astrocytes. GS plays a pivotal role in glutamate and glutamine homoeostasis, orchestrating astrocyte glutamate uptake/release and the glutamate-glutamine cycle. Furthermore, astrocytes bear the brunt of clearing ammonia in the brain, preventing neurotoxicity. The present review depicts the central function of astrocytes, concentrating on the importance of GS in glutamate/glutamine metabolism and ammonia detoxification in health and disease.


Novel molecular imaging platform for monitoring oncological kinases.
Recent advances in oncology have lead to identification of a plethora of alterations in signaling pathways that are critical to oncogenesis and propagation of malignancy. Among the biomarkers identified, dysregulated kinases and associated changes in signaling cascade received the lion's share of scientific attention and have been under extensive investigations with goal of targeting them for anti-cancer therapy. Discovery of new drugs is immensely facilitated by molecular imaging technology which enables non-invasive, real time, dynamic imaging and quantification of kinase activity. Here, we review recent development of novel kinase reporters based on conformation dependent complementation of firefly luciferase to monitor kinase activity. Such reporter system provides unique insights into the pharmacokinetics and pharmacodynamics of drugs that modulate kinase signaling and have a huge potential in drug discovery, validation, and drug-target interactions.


Dual influence of the yeast Cat1p (Snf1p) protein kinase on carbon source-dependent transcriptional activation of gluconeogenic genes by the regulatory gene CAT8.
The CSRE (carbon source-responsive element) is a sequence motif responsible for the transcriptional activation of gluconeogenic structural genes in Saccharomyces cerevisiae. We have isolated a regulatory gene, DIL1 (derepression of isocitrate lyase, = CAT8), which is specifically required for derepression of CSRE-dependent genes. Expression of CAT8 is carbon source regulated and requires a functional Cat1p (Snf1p) protein kinase. The derepression defect of CAT8 in a cat1 mutant could be suppressed by a mutant Mig1p repressor protein. Derepression of CAT8 also requires a functional HAP2 gene, suggesting a regulatory connection between respiratory and gluconeogenic genes. Carbon source-dependent protein-CSRE complexes detected in a gel retardation analysis with wild-type extracts were absent in cat8 mutant extracts. However, similar experiments with an epitope-tagged CAT8 gene product in the presence of tag-specific antibodies gave evidence against a direct binding of Cat8p to the CSRE. A constitutively expressed GAL4-CAT8 fusion gene revealed a carbon source-dependent transcriptional activation of a UAS(GAL)-containing reporter gene. Activation mediated by Cat8p was no longer detectable in a cat1 mutant. Thus, biosynthetic control of CAT8 as well as transcriptional activation by Cat8p requires a functional Cat1p protein kinase. A model proposing CAT8 as a specific activator of a transcription factor(s) binding to the CSRE is discussed.


Streamlining radioembolization in UNOS T1/T2 hepatocellular carcinoma by eliminating lung shunt estimation.
Pre-treatment Tc-99m macroaggregated albumin (MAA) scans are routinely performed prior to transarterial radioembolization (TARE) to estimate lung shunt fraction (LSF) and lung dose. In this study, we investigate LSF observed in early hepatocellular carcinoma (HCC) and provide the scientific rationale for eliminating this step from routine practice. Patients with HCC who underwent Y90 from 2004 to 2018 were reviewed. Inclusion criteria were early stage HCC (UNOS T1/T2/Milan criteria: solitary ≤5 cm, 3 nodules ≤3 cm). LSF was determined using MAA in all patients. Associations between LSF and baseline characteristics were investigated. A "no-MAA" paradigm was then proposed based on a homogenous group that expressed very low LSF. Of 1,175 patients with HCC treated with TARE, 448 patients met inclusion criteria. Mean age was 65.6 years and 303 (68%) were males. A total of 352 (79%) had solitary lesions and 406 (91%) unilobar disease. Two-hundred and forty-three (54%), 178 (40%) and 27 (6%) patients were Child-Pugh class A, B and C, respectively. Median LSF was 3.9% (IQR 2.4-6%). Median administered activity was 0.9 GBq (IQR 0.6-1.4), for a median segmental volume of 170 cm3 (range: 60-530). Median lung dose was 1.9 Gy (IQR: 1.0-3.3). The presence of a transjugular intrahepatic portosystemic shunt (TIPS; n = 38) was associated with LSF >10% (odds ratio 12.2; 95% CI 5.2-28.6; p <0.001). Median LSF was 3.8% (IQR: 2.4-5.7%) and 6% (IQR: 3.8-15.3%) in no-TIPS vs. TIPS patients (p <0.001). LSF is clinically negligible in patients with UNOS T1/T2 HCC without TIPS. When segmental injections are planned, this step can be eliminated, thereby reducing time-to-treatment, number of procedures, and improving convenience for patients traveling from faraway. Transarterial radioembolization is a microembolic transarterial treatment for hepatocellular carcinoma. In our study, we found that early stage patients, where segmental injections are planned, exhibited low lung shunting, effectively eliminating the risk of radiation pneumonitis. We propose that the lung shunt study be eliminated in this subgroup, thus leading to fewer procedures, a cost reduction and improved convenience for patients.


Spectroscopic Photoacoustic Molecular Imaging of Breast Cancer using a B7-H3-targeted ICG Contrast Agent.
Purpose: Breast cancer imaging methods lack diagnostic accuracy, in particular for patients with dense breast tissue, and improved techniques are critically needed. The purpose of this study was to evaluate antibody-indocyanine green (ICG) conjugates, which undergo dynamic absorption spectrum shifts after cellular endocytosis and degradation, and spectroscopic photoacoustic (sPA) imaging to differentiate normal breast tissue from breast cancer by imaging B7-H3, a novel breast cancer associated molecular target. Methods: Quantitative immunohistochemical staining of endothelial and epithelial B7-H3 expression was assessed in 279 human breast tissue samples, including normal (n=53), benign lesions (11 subtypes, n=129), and breast cancers (4 subtypes, n=97). After absorption spectra of intracellular and degraded B7-H3-ICG and Isotype control-ICG (Iso-ICG) were characterized, sPA imaging in a transgenic murine breast cancer model (FVB/N-Tg(MMTVPyMT)634Mul) was performed and compared to imaging of control conditions [B7-H3-ICG in tumor negative animals (n=60), Iso-ICG (n=30), blocking B7-H3+B7-H3-ICG (n=20), and free ICG (n=20)] and validated with ex vivo histological analysis. Results: Immunostaining showed differential B7-H3 expression on both the endothelium and tumor epithelium in human breast cancer with an area under the ROC curve of 0.93 to differentiate breast cancer vs non-cancer. Combined in vitro/in vivo imaging showed that sPA allowed specific B7-H3-ICG detection down to the 13 nM concentration and differentiation from Iso-ICG. sPA molecular imaging of B7-H3-ICG showed a 3.01-fold (P<0.01) increase in molecular B7-H3-ICG signal in tumors compared to control conditions. Conclusions: B7-H3 is a promising target for both vascular and epithelial sPA imaging of breast cancer. Leveraging antibody-ICG contrast agents and their dynamic optical absorption spectra allows for highly specific sPA imaging of breast cancer.


A new mutation in mgrb mediating polymyxin resistance in Klebsiella variicola.
Polymyxin resistance is a public health concern - present in humans, animals and the environment - caused by chromosomal-encoding or plasmid-encoding mechanisms. Chromosomal alterations in MgrB are frequently detected in Klebsiella spp., but not yet reported and characterised in Klebsiella variicola (K. variicola). This study performed microbiological and genomic characterisation of three polymyxin-resistant K. variicola isolates (M14, M15 and M50) recovered from the microbiota of migratory birds in Brazil. The isolates were submitted to SpeI-PFGE, broth microdilution and whole genome sequencing using Illumina MiSeq for analysis of genetic relatedness, sequence typing and detection of antimicrobial-resistance genes. K. variicola isolates belonged to two clones, and susceptibility tests showed resistance only for polymyxins. Sequences of chromosomal two-component systems (PmrAB, PhoPQ, RstAB, CrrAB) and MgrB were evaluated by blastN and blastP against a polymyxin-susceptible K. variicola (A58243), and mutations with biological effect were checked by the PROVEAN tool. K. variicola isolates belonged to two clones, and susceptibility tests showed resistance for polymyxins. In M14 and M15, phoQ deleterious mutations (D90N, I122S and G385S) were identified, while an mgrB variant containing a single deletion (C deletion on position 93) leading to the production of a non-functional protein was detected in M50. mgrB complementation studies showed restoration of polymyxin susceptibility (64 to ≤ 0.25 mg/L) as a wild-type mgrB was inserted into the mgrB-deficient M50. This study confirmed the role of a non-functional mgrB variant in conferring polymyxin resistance, stressing the role of this regulator in K. variicola and drawing attention to novel polymyxin resistance mechanisms emerging in wildlife.


Fluoride mouthrinses for preventing dental caries in children and adolescents.
Fluoride mouthrinses have been used extensively as a caries-preventive intervention in school-based programmes and individually at home. To determine the effectiveness and safety of fluoride mouthrinses in the prevention of dental caries in children and to examine factors potentially modifying their effect. We searched the Cochrane Oral Health Group's Trials Register (May 2000), the Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library Issue 2, 2000), MEDLINE (1966 to January 2000), plus several other databases. We handsearched journals, reference lists of articles and contacted selected authors and manufacturers. Randomised or quasi-randomised controlled trials with blind outcome assessment, comparing fluoride mouthrinse with placebo or no treatment in children up to 16 years during at least 1 year. The main outcome was caries increment measured by the change in decayed, missing and filled tooth surfaces (D(M)FS). Inclusion decisions, quality assessment and data extraction were duplicated in a random sample of one third of studies, and consensus achieved by discussion or a third party. Authors were contacted for missing data. The primary measure of effect was the prevented fraction (PF) that is the difference in mean caries increments between the treatment and control groups expressed as a percentage of the mean increment in the control group. Random effects meta-analyses were performed where data could be pooled. Potential sources of heterogeneity were examined in random effects metaregression analyses. Thirty-six studies were included. For the 34 that contributed data for meta-analysis (involving 14,600 children) the D(M)FS pooled PF was 26% (95% confidence interval (CI), 23% to 30%; p < 0.0001). Heterogeneity was not substantial, but confirmed statistically (p = 0.008). No significant association between estimates of D(M)FS prevented fractions and baseline caries severity, background exposure to fluorides, rinsing frequency and fluoride concentration was found in metaregression analyses. A funnel plot of the 34 studies indicated no relationship between prevented fraction and study precision. There is little information concerning possible adverse effects or acceptability of treatment in the included trials. This review suggests that the supervised regular use of fluoride mouthrinse at two main strengths and rinsing frequencies is associated with a clear reduction in caries increment in children. In populations with caries increment of 0.25 D(M)FS per year, 16 children will need to use a fluoride mouthrinse (rather than a non-fluoride rinse) to avoid one D(M)FS; in populations with a caries increment of 2.14 D(M)FS per year, 2 children will need to rinse to avoid one D(M)FS. There is a need for complete reporting of side effects and acceptability data in fluoride mouthrinse trials.


A Combination of Metagenomic and Cultivation Approaches Reveals Hypermutator Phenotypes within Vibrio cholerae-Infected Patients.
Vibrio cholerae can cause a range of symptoms, from severe diarrhea to asymptomatic infection. Previous studies using whole-genome sequencing (WGS) of multiple bacterial isolates per patient showed that V. cholerae can evolve modest genetic diversity during symptomatic infection. To further explore the extent of V. cholerae within-host diversity, we applied culture-based WGS and metagenomics to a cohort of both symptomatic and asymptomatic cholera patients from Bangladesh. While metagenomics allowed us to detect more mutations in symptomatic patients, WGS of cultured isolates was necessary to detect V. cholerae diversity in asymptomatic carriers, likely due to their low V. cholerae load. Using both metagenomics and isolate WGS, we report three lines of evidence that V. cholerae hypermutators evolve within patients. First, we identified nonsynonymous mutations in V. cholerae DNA repair genes in 5 out of 11 patient metagenomes sequenced with sufficient coverage of the V. cholerae genome and in 1 of 3 patients with isolate genomes sequenced. Second, these mutations in DNA repair genes tended to be accompanied by an excess of intrahost single nucleotide variants (iSNVs). Third, these iSNVs were enriched in transversion mutations, a known hallmark of hypermutator phenotypes. While hypermutators appeared to generate mostly selectively neutral mutations, nonmutators showed signs of convergent mutation across multiple patients, suggesting V. cholerae adaptation within hosts. Our results highlight the power and limitations of metagenomics combined with isolate sequencing to characterize within-patient diversity in acute V. cholerae infections, while providing evidence for hypermutator phenotypes within cholera patients. IMPORTANCE Pathogen evolution within patients can impact phenotypes such as drug resistance and virulence, potentially affecting clinical outcomes. V. cholerae infection can result in life-threatening diarrheal disease or asymptomatic infection. Here, we describe whole-genome sequencing of V. cholerae isolates and culture-free metagenomic sequencing from stool of symptomatic cholera patients and asymptomatic carriers. Despite the typically short duration of cholera, we found evidence for adaptive mutations in the V. cholerae genome that occur independently and repeatedly within multiple symptomatic patients. We also identified V. cholerae hypermutator phenotypes within several patients, which appear to generate mainly neutral or deleterious mutations. Our work sets the stage for future studies of the role of hypermutators and within-patient evolution in explaining the variation from asymptomatic carriage to symptomatic cholera.


Clinical comparison of hydroxyapatite-coated titanium dental implants placed in fresh extraction sockets and healed sites.
The placement of hydroxyapatite-coated dental implants in fresh extraction sockets was compared to placement in adjacent healed sites in 14 patients. Systematic sequential documentation was obtained regarding periodontal health, radiographic bone levels, and implant stability at the time of implant placement, at uncovering, and from 8 to 24 months (mean 16 months) after loading and restoration delivery. There were no significant differences in any clinical parameter between those implants placed in fresh extraction sockets and those placed in healed areas. Periodontal health, maintenance of crestal bone levels, and implant stability were excellent for implants placed in both types of recipient sites. The results of this study suggest that hydroxyapatite-coated dental implants can be successfully placed in fresh extraction sockets utilizing otherwise standard implant placement techniques, and that they appear to clinically perform equally well in fresh sockets and healed sites.


Trends in Intravenous Antibiotic Duration for Urinary Tract Infections in Young Infants.
To assess trends in the duration of intravenous (IV) antibiotics for urinary tract infections (UTIs) in infants ≤60 days old between 2005 and 2015 and determine if the duration of IV antibiotic treatment is associated with readmission. Retrospective analysis of infants ≤60 days old diagnosed with a UTI who were admitted to a children's hospital and received IV antibiotics. Infants were excluded if they had a previous surgery or comorbidities, bacteremia, or admission to the ICU. Data were analyzed from the Pediatric Health Information System database from 2005 through 2015. The primary outcome was readmission within 30 days for a UTI. The proportion of infants ≤60 days old receiving 4 or more days of IV antibiotics (long IV treatment) decreased from 50% in 2005 to 19% in 2015. The proportion of infants ≤60 days old receiving long IV treatment at 46 children's hospitals varied between 3% and 59% and did not correlate with readmission (correlation coefficient 0.13; P = .37). In multivariable analysis, readmission for a UTI was associated with younger age and female sex but not duration of IV antibiotic therapy (adjusted odds ratio for long IV treatment: 0.93 [95% confidence interval 0.52-1.67]). The proportion of infants ≤60 days old receiving long IV treatment decreased substantially from 2005 to 2015 without an increase in hospital readmissions. These findings support the safety of short-course IV antibiotic therapy for appropriately selected neonates.


Outpatient antibiotic use in the four administrations of the UK: cross-sectional and longitudinal analysis.
We compared community antibiotic use in the four administrations of the UK with that in other European countries. A cross-sectional analysis of 24 European countries and UK administrations in 2005 with longitudinal analysis of data from Belgium and UK from 1997 was performed. Antibiotic use was measured as defined daily doses per 1000 inhabitants per day (DID) or as prescriptions or packages per 1000 inhabitants per day (PID) with aggregate data from patients who received antibiotics in the primary care or outpatient setting. In 2005, there were marked differences in antibiotic prescriptions between the four UK administrations, for example, in descending order of DIDs, Northern Ireland and England ranked 8th and 24th, respectively, out of the 28 countries. DIDs for Northern Ireland were 37% greater than that for England. Longitudinal analysis showed that differences were present before devolution in 1999. Increase in the age of exemption from prescription charges in Wales in 2002 was not associated with significant change in use in comparison with the other UK countries. There were discrepancies between changes in DID and changes in PID, particularly in Belgium. This suggests that some changes in DID were due to changes in dosing or duration of the treatment rather than in the number of people treated and shows the importance of using both measures. The European Surveillance of Antimicrobial Consumption project has for the first time made data about antibiotic use in the four UK administrations publicly available. This reveals important practice variations that should stimulate research to explain differences and assess their consequences.


Testosterone-induced neuroendocrine changes in the medial preoptic area precede song activation and plasticity in song control nuclei of female canaries.
Testosterone plays a key role in the control of seasonal changes in singing behavior and its underlying neural circuitry. After administration of exogenous testosterone, song quality and song control nuclei volumes change over the course of weeks, but song rate increases within days. The medial preoptic nucleus (POM) controls sexual motivation and testosterone action in POM increases sexually motivated singing. In this study, we investigated the time course of testosterone action in the song control nuclei and POM, at the gross anatomical and cellular level. Photosensitive female canaries were injected with BrdU to label newborn neurons. One day later they were transferred to a long-day photoperiod and implanted with testosterone-filled or empty implants. Brains and blood were collected 1, 2, 9 or 21 days later. Testosterone increased POM volume within 1 day, whereas the volume of song control nuclei increased significantly only on day 21 even if a trend was already observed for HVC on day 9. The density of newborn neurons in HVC, labeled by Bromodeoxyuridine (BrdU) and doublecortin, was increased by testosterone on days 9 and 21 although a trend was already detectable on day 2. In POM, testosterone increased the number and size of aromatase-immunoreactive neurons already after 1 day. This rapid action of testosterone in POM supports its proposed role in controlling singing motivation. Although testosterone increased the number of newborn neurons in HVC rapidly (9, possibly 2 days), it is unlikely that these new neurons affect singing behavior before they mature and integrate into functional circuits.


Allocation of Internal Medicine Resident Time in a Swiss Hospital: A Time and Motion Study of Day and Evening Shifts.
Little current evidence documents how internal medicine residents spend their time at work, particularly with regard to the proportions of time spent in direct patient care versus using computers. To describe how residents allocate their time during day and evening hospital shifts. Time and motion study. Internal medicine residency at a university hospital in Switzerland, May to July 2015. 36 internal medicine residents with an average of 29 months of postgraduate training. Trained observers recorded the residents' activities using a tablet-based application. Twenty-two activities were categorized as directly related to patients, indirectly related to patients, communication, academic, nonmedical tasks, and transition. In addition, the presence of a patient or colleague and use of a computer or telephone during each activity was recorded. Residents were observed for a total of 696.7 hours. Day shifts lasted 11.6 hours (1.6 hours more than scheduled). During these shifts, activities indirectly related to patients accounted for 52.4% of the time, and activities directly related to patients accounted for 28.0%. Residents spent an average of 1.7 hours with patients, 5.2 hours using computers, and 13 minutes doing both. Time spent using a computer was scattered throughout the day, with the heaviest use after 6:00 p.m. The study involved a small sample from 1 institution. At this Swiss teaching hospital, internal medicine residents spent more time at work than scheduled. Activities indirectly related to patients predominated, and about half the workday was spent using a computer. Information Technology Department and Department of Internal Medicine of Lausanne University Hospital.


Increases in the circulating testosterone of maturing male guinea pigs appear neither necessary nor sufficient for heightened maternally directed sexual and social/courtship behavior.
Periadolescent male guinea pigs housed continuously with their mother since birth exhibit little maternally directed sexual behavior. However, if rehoused apart from the mother for 24 h, they show elevations in circulating testosterone concentrations and display frequent sexual responses and increased social/courtship behavior upon reunion with her. We investigated the role of testosterone in the disinhibition of maternally directed sexual and social/courtship behavior. Subcutaneous implants of testosterone (Experiment 1) did not trigger maternally directed sexual behavior or an increase in social/courtship behavior among males housed continuously with their mothers. Further, neither blocking androgen receptors (Experiment 2) nor preventing the surge in testosterone (Experiment 3) prevented males housed without the mother from exhibiting increased maternally directed sexual and social/courtship behavior upon reunion. These findings indicate that the increase in testosterone that males exhibit when rehoused apart from the mother is neither sufficient nor necessary for the disinhibition of maternally directed sexual and social/courtship behavior observed when mother and son are reunited.


Severe population collapses and species extinctions in multihost epidemic dynamics.
Most infectious diseases, including more than half of known human pathogens, are not restricted to just one host, yet much of the mathematical modeling of infections has been limited to a single species. We investigate consequences of a single epidemic propagating in multiple species and compare and contrast it with the endemic steady state of the disease. We use the two-species susceptible-infected-recovered model to calculate the severity of postepidemic collapses in populations of two-host species as a function of their initial population sizes, the times individuals remain infectious, and the matrix of infection rates. We derive the criteria for a very large, extinction-level, population collapse in one or both of the species. The main conclusion of our study is that a single epidemic could drive a species with high mortality rate to local or even global extinction provided that it is coinfected with an abundant species. Such collapse-driven extinctions depend on factors different than those in the endemic steady state of the disease.


Nsp1 protein of SARS-CoV-2 disrupts the mRNA export machinery to inhibit host gene expression.
The ongoing unprecedented severe acute respiratory syndrome caused by the SARS-CoV-2 outbreak worldwide has highlighted the need for understanding viral-host interactions involved in mechanisms of virulence. Here, we show that the virulence factor Nsp1 protein of SARS-CoV-2 interacts with the host messenger RNA (mRNA) export receptor heterodimer NXF1-NXT1, which is responsible for nuclear export of cellular mRNAs. Nsp1 prevents proper binding of NXF1 to mRNA export adaptors and NXF1 docking at the nuclear pore complex. As a result, a significant number of cellular mRNAs are retained in the nucleus during infection. Increased levels of NXF1 rescues the Nsp1-mediated mRNA export block and inhibits SARS-CoV-2 infection. Thus, antagonizing the Nsp1 inhibitory function on mRNA export may represent a strategy to restoring proper antiviral host gene expression in infected cells.


Enzymes Catalyzing the TCA- and Urea Cycle Influence the Matrix Composition of Biofilms Formed by Methicillin-Resistant Staphylococcus aureus USA300.
In methicillin-sensitive Staphylococcus aureus (MSSA), the tricarboxylic acid (TCA) cycle is known to negatively regulate production of the major biofilm-matrix exopolysaccharide, PIA/PNAG. However, methicillin-resistant S. aureus (MRSA) produce a primarily proteinaceous biofilm matrix, and contribution of the TCA-cycle therein remains unclear. Utilizing USA300-JE2 Tn-mutants (NARSA) in genes encoding TCA- and urea cycle enzymes for transduction into a prolific biofilm-forming USA300 strain (UAS391-Erys), we studied the contribution of the TCA- and urea cycle and of proteins, eDNA and PIA/PNAG, to the matrix. Genes targeted in the urea cycle encoded argininosuccinate lyase and arginase (argH::Tn and rocF::Tn), and in the TCA-cycle encoded succinyl-CoA synthetase, succinate dehydrogenase, aconitase, isocitrate dehydrogenase, fumarate hydratase class II, and citrate synthase II (sucC::Tn, sdhA/B::Tn, acnA::Tn, icd::Tn, fumC::Tn and gltA::Tn). Biofilm formation was significantly decreased under no flow and flow conditions by argH::Tn, fumC::Tn, and sdhA/B::Tn (range OD492 0.374-0.667; integrated densities 2.065-4.875) compared to UAS391-EryS (OD492 0.814; integrated density 10.676) (p ≤ 0.008). Cellular and matrix stains, enzymatic treatment (Proteinase K, DNase I), and reverse-transcriptase PCR-based gene-expression analysis of fibronectin-binding proteins (fnbA/B) and the staphylococcal accessory regulator (sarA) on pre-formed UAS391-Erys and Tn-mutant biofilms showed: (i) < 1% PIA/PNAG in the proteinaceous/eDNA matrix; (ii) increased proteins under no flow and flow in the matrix of Tn mutant biofilms (on average 50 and 51 (±11)%) compared to UAS391-Erys (on average 22 and 25 (±4)%) (p < 0.001); and (iii) down- and up-regulation of fnbA/B and sarA, respectively, in Tn-mutants compared to UAS391-EryS (0.62-, 0.57-, and 2.23-fold on average). In conclusion, we show that the biofilm matrix of MRSA-USA300 and the corresponding Tn mutants is PIA/PNAG-independent and are mainly composed of proteins and eDNA. The primary impact of TCA-cycle inactivation was on the protein component of the biofilm matrix of MRSA-USA300.


Lactose in diet influences the degradation of mixed linked β(1-3;1-4)-D-glucan in the small intestine of pigs.
The objective was to study the cause of variation in digestibility of mixed linked β(1-3;1-4)-D-glucan (β-glucan) in the small intestine of growing pigs. The β-glucan is an important cell wall [dietary fiber (DF)] component of the endosperm of barley (Hordeum vulgare) and oats (Avena sativa). The digestibility of β-glucan in the small intestine from both cereals is among the highest of all DF components, but in 1 study with oat-based diets it was lower (P < 0.001) than in other studies. In this study, whey protein containing lactose was used as protein supplement. Lactose is slowly digestible in the small intestine. To investigate if lactose might cause the lower digestibility of β-glucan in the study with whey protein, the lactose in diets was analyzed together with lactose and organic acids (lactic acids and short-chain fatty acids) in digesta samples from the small intestine (the small intestine was by length divided in 3 equal segments: SI(1), SI(2), and SI(3)) and ileal digesta. Diets containing lactose were based on oat goats, oat flour, and oat bran (12 to 38 g lactose/kg DM) whereas the reference diets were based on rolled oats, rolled oats and oat bran, wheat (Triticum aestivum) flour with added oat bran, and wheat flour with added β-glucan (0 to 1 g lactose/kg DM). Lactose was identified in digesta up to SI(2) but disappeared in digesta from SI(3) and ileum. Digestibility of β-glucan did not differ among diets up to SI(3) (18% average) whereas digestibility in ileum was 64% in diets without lactose and 27% in diets containing lactose (P < 0.001). The β-glucan was virtually completely digested in the cecum (96% average) in all diets. The concentration of organic acids did not differ between diets either in SI(3), ileum, or cecum. In conclusion, slowly digestible lactose was the most likely cause of the reduced digestibility of β-glucan in oat diets containing lactose.


Surgical decision making in premenopausal BRCA carriers considering risk-reducing early salpingectomy or salpingo-oophorectomy: a qualitative study.
Acceptance of the role of the fallopian tube in 'ovarian' carcinogenesis and the detrimental sequelae of surgical menopause in premenopausal women following risk-reducing salpingo-oophorectomy (RRSO) has resulted in risk-reducing early-salpingectomy with delayed oophorectomy (RRESDO) being proposed as an attractive alternative risk-reducing strategy in women who decline/delay oophorectomy. We present the results of a qualitative study evaluating the decision-making process among BRCA carriers considering prophylactic surgeries (RRSO/RRESDO) as part of the multicentre PROTECTOR trial (ISRCTN:25173360). In-depth semistructured 1:1 interviews conducted using a predeveloped topic-guide (development informed by literature review and expert consultation) until informational saturation reached. Wording and sequencing of questions were left open with probes used to elicit additional information. All interviews were audio-recorded, transcribed verbatim, transcripts analysed using an inductive theoretical framework and data managed using NVIVO-v12. Informational saturation was reached following 24 interviews. Seven interconnected themes integral to surgical decision making were identified: fertility/menopause/cancer risk reduction/surgical choices/surgical complications/sequence of ovarian-and-breast prophylactic surgeries/support/satisfaction. Women for whom maximising ovarian cancer risk reduction was relatively more important than early menopause/quality-of-life preferred RRSO, whereas those more concerned about detrimental impact of menopause chose RRESDO. Women managed in specialist familial cancer clinic settings compared with non-specialist settings felt they received better quality care, improved hormone replacement therapy access and were more satisfied. Multiple contextual factors (medical, physical, psychological, social) influence timing of risk-reducing surgeries. RRESDO offers women delaying/declining premenopausal oophorectomy, particularly those concerned about menopausal effects, a degree of ovarian cancer risk reduction while avoiding early menopause. Care of high-risk women should be centralised to centres with specialist familial gynaecological cancer risk management services to provide a better-quality, streamlined, holistic multidisciplinary approach.


Eosinophil recovery in hospitalized COVID-19 patients is associated with lower rates of ICU admission and in-hospital mortality: An observational cohort analysis.
Admission eosinopenia (<100 cells/μL) is associated with poor clinical outcomes in hospitalized COVID-19 patients. However, the effects of eosinophil recovery (defined as reaching ≥50 eosinophils/μL) during hospitalization on COVID-19 outcomes have been inconsistent. The study included 1,831 patients admitted to UCLA hospitals between February 2020 and February 2021 with PCR-confirmed COVID-19. Using competing risk regression and modeling eosinophil recovery as a time-dependent covariate, we evaluated the longitudinal relationship between eosinophil recovery and in-hospital outcomes including ICU admission, need for mechanical ventilation, and in-hospital mortality. All analyses were adjusted for covariates including age, BMI, tobacco smoke exposure, comorbidities known to be risk factors for COVID-19 mortality, and treatments including dexamethasone and remdesivir. Eosinophil recovery was evaluated in patients with <50 eosinophils/μL on admission (n = 1282). These patients cumulatively amassed 11,633 hospital patient-days; 3,985 of those days qualified as eosinophil recovery events, which were represented by 781 patients achieving at least one instance of eosinophil recovery during hospitalization. Despite no significant difference in the rate of mechanical ventilation, eosinophil recoverers had significantly lower rates of in-hospital mortality (aHR: 0.44 [0.29, 0.65], P = 0.001) and ICU admission (aHR: 0.25 [0.11, 0.61], P = 0.002). Trending eosinophil counts during hospitalization is simple and can be performed in resource-limited healthcare settings to track the inflammatory status of a patient. Lack of eosinophil recovery events can identify those at risk for future progression to severe COVID.


A molecular basis underpinning the T cell receptor heterogeneity of mucosal-associated invariant T cells.
Mucosal-associated invariant T (MAIT) cells express an invariant T cell receptor (TCR) α-chain (TRAV1-2 joined to TRAJ33, TRAJ20, or TRAJ12 in humans), which pairs with an array of TCR β-chains. MAIT TCRs can bind folate- and riboflavin-based metabolites restricted by the major histocompatibility complex (MHC)-related class I-like molecule, MR1. However, the impact of MAIT TCR and MR1-ligand heterogeneity on MAIT cell biology is unclear. We show how a previously uncharacterized MR1 ligand, acetyl-6-formylpterin (Ac-6-FP), markedly stabilized MR1, potently up-regulated MR1 cell surface expression, and inhibited MAIT cell activation. These enhanced properties of Ac-6-FP were attributable to structural alterations in MR1 that subsequently affected MAIT TCR recognition via conformational changes within the complementarity-determining region (CDR) 3β loop. Analysis of seven TRBV6-1(+) MAIT TCRs demonstrated how CDR3β hypervariability impacted on MAIT TCR recognition by altering TCR flexibility and contacts with MR1 and the Ag itself. Ternary structures of TRBV6-1, TRBV6-4, and TRBV20(+) MAIT TCRs in complex with MR1 bound to a potent riboflavin-based antigen (Ag) showed how variations in TRBV gene usage exclusively impacted on MR1 contacts within a consensus MAIT TCR-MR1 footprint. Moreover, differential TRAJ gene usage was readily accommodated within a conserved MAIT TCR-MR1-Ag docking mode. Collectively, MAIT TCR heterogeneity can fine-tune MR1 recognition in an Ag-dependent manner, thereby modulating MAIT cell recognition.


The gut of the finch: uniqueness of the gut microbiome of the Galápagos vampire finch.
Darwin's finches are a clade of 19 species of passerine birds native to the Galápagos Islands, whose biogeography, specialized beak morphologies, and dietary choices-ranging from seeds to blood-make them a classic example of adaptive radiation. While these iconic birds have been intensely studied, the composition of their gut microbiome and the factors influencing it, including host species, diet, and biogeography, has not yet been explored. We characterized the microbial community associated with 12 species of Darwin's finches using high-throughput 16S rRNA sequencing of fecal samples from 114 individuals across nine islands, including the unusual blood-feeding vampire finch (Geospiza septentrionalis) from Darwin and Wolf Islands. The phylum-level core gut microbiome for Darwin's finches included the Firmicutes, Gammaproteobacteria, and Actinobacteria, with members of the Bacteroidetes at conspicuously low abundance. The gut microbiome was surprisingly well conserved across the diversity of finch species, with one exception-the vampire finch-which harbored bacteria that were either absent or extremely rare in other finches, including Fusobacterium, Cetobacterium, Ureaplasma, Mucispirillum, Campylobacter, and various members of the Clostridia-bacteria known from the guts of carnivorous birds and reptiles. Complementary stable isotope analysis of feathers revealed exceptionally high δ15N isotope values in the vampire finch, resembling top marine predators. The Galápagos archipelago is also known for extreme wet and dry seasons, and we observed a significant seasonal shift in the gut microbial community of five additional finch species sampled during both seasons. This study demonstrates the overall conservatism of the finch gut microbiome over short (< 1 Ma) divergence timescales, except in the most extreme case of dietary specialization, and elevates the evolutionary importance of seasonal shifts in driving not only species adaptation, but also gut microbiome composition.


Endogenous Viral Elements Are Widespread in Arthropod Genomes and Commonly Give Rise to PIWI-Interacting RNAs.
Arthropod genomes contain sequences derived from integrations of DNA and nonretroviral RNA viruses. These sequences, known as endogenous viral elements (EVEs), have been acquired over the course of evolution and have been proposed to serve as a record of past viral infections. Recent evidence indicates that EVEs can function as templates for the biogenesis of PIWI-interacting RNAs (piRNAs) in some mosquito species and cell lines, raising the possibility that EVEs may serve as a source of immunological memory in these organisms. However, whether piRNAs are derived from EVEs or serve an antiviral function in other arthropod species is unknown. Here, we used publicly available genome assemblies and small RNA sequencing data sets to characterize the repertoire and function of EVEs across 48 arthropod genomes. We found that EVEs are widespread in arthropod genomes and primarily correspond to unclassified single-stranded RNA (ssRNA) viruses and viruses belonging to the Rhabdoviridae and Parvoviridae families. Additionally, EVEs were enriched in piRNA clusters in a majority of species, and we found that production of primary piRNAs from EVEs is common, particularly for EVEs located within piRNA clusters. While the abundance of EVEs within arthropod genomes and the frequency with which EVEs give rise to primary piRNAs generally support the hypothesis that EVEs contribute to an antiviral response via the piRNA pathway, limited nucleotide identity between currently described viruses and EVEs identified here likely limits the extent to which this process plays a role during infection with known viruses in the arthropod species analyzed.IMPORTANCE Our results greatly expand the knowledge of EVE abundance, diversity, and function in an exceptionally wide range of arthropod species. We found that while previous findings in mosquitoes regarding the potential of EVEs to serve as sources of immunological memory via the piRNA pathway may be generalized to other arthropod species, speculation regarding the antiviral function of EVE-derived piRNAs should take into context the fact that EVEs are, in the vast majority of cases, not similar enough to currently described viruses at the nucleotide level to serve as sources of antiviral piRNAs against them.


Injury-associated reacquiring of intestinal stem cell function.
Epithelial layer of the intestine relies upon stem cells for maintaining homeostasis and regeneration. Two types of stem cells are currently defined in intestinal crypts: the cycling crypt base columnar cells and quiescent cells. Though several candidate markers and regulators of rapidly cycling and quiescent stem cells have been identified so far, the exact nature of quiescent cells is still questionable since investigations mainly focused on candidate markers rather than the label-retaining population itself. Recent results, however, have strengthened the argument for functional plasticity. Using a lineage tracing strategy label-retaining cells (LRCs) of the intestinal epithelium were marked, then followed by a pulse-chase analysis it was found that during homeostasis, LRCs were Lgr5-positive and were destined to become Paneth and neuroendocrine cells. Nevertheless, it was demonstrated that LRCs are capable of clonogenic growth by recall to the self-renewing pool of stem cells in case of epithelial injury. These new findings highlight on the hierarchical and spatial organization of intestinal epithelial homeostasis and the important plasticity of progenitors during tissue regeneration, moreover, provide a motivation for studying their role in disorders like colorectal cancer.


Adult health outcomes and their implications for experiences of childhood nutritional stress in Jamaica.
With insights from the developmental origins of health and disease paradigm (DOHaD), this study explores the impact of childhood nutritional stress on adult health outcomes in Jamaica. Jamaica experienced a lengthy period of political and economic instability beginning in the postcolonial period of the early 1960s. This study tests whether decreased government spending on public resources and limited access to imported food products during the early postcolonial period will be reflected in increased adiposity and body mass index among Jamaican adults. Ethnographic and anthropometric data were collected from individuals born between 1958 and 1988. Variability in health outcomes was assessed using Z-score values for body mass index and summed skinfold thickness measures. Age was employed as both a continuous and categorical independent variable. In partial correlation models controlling for economic status, body mass index values and summed skinfold thickness increased with age. Birth cohort and gender effects were also apparent. Women born between 1959 and 1968 had higher body mass index Z-score values than younger women. Both men and women born between 1959 and 1968 had significantly higher skinfold thickness measures than younger individuals. Individuals born between 1959 and 1968 were children during the immediate postcolonial era in Jamaica. Experiences of nutritional stress during critical developmental periods may have contributed to the observed age-related increases in adipose tissue and body mass index values. This study informs our understanding of the ways that fluctuations in the sociopolitical environment during development can mediate and contribute to poor adult health outcomes.


Sildenafil decreases rat tracheal hyperresponsiveness to carbachol and changes canonical transient receptor potential gene expression after antigen challenge.
Inhibition of type-5 phosphodiesterase by sildenafil decreases capacitative Ca2+ entry mediated by transient receptor potential proteins (TRPs) in the pulmonary artery. These families of channels, especially the canonical TRP (TRPC) subfamily, may be involved in the development of bronchial hyperresponsiveness, a hallmark of asthma. In the present study, we evaluated i) the effects of sildenafil on tracheal rings of rats subjected to antigen challenge, ii) whether the extent of TRPC gene expression may be modified by antigen challenge, and iii) whether inhibition of type-5 phosphodiesterase (PDE5) may alter TRPC gene expression after antigen challenge. Sildenafil (0.1 µM to 0.6 mM) fully relaxed carbachol-induced contractions in isolated tracheal rings prepared from naive male Wistar rats (250-300 g) by activating the NO-cGMP-K+ channel pathway. Rats sensitized to antigen by intraperitoneal injections of ovalbumin were subjected to antigen challenge by ovalbumin inhalation, and their tracheal rings were used to study the effects of sildenafil, which more effectively inhibited contractions induced by either carbachol (10 µM) or extracellular Ca2+ restoration after thapsigargin (1 µM) treatment. Antigen challenge increased the expression of the TRPC1 and TRPC4 genes but not the expression of the TRPC5 and TRPC6 genes. Applied before the antigen challenge, sildenafil increased the gene expression, which was evaluated by RT-PCR, of TRPC1 and TRPC6, decreased TRPC5 expression, and was inert against TRPC4. Thus, we conclude that PDE5 inhibition is involved in the development of an airway hyperresponsive phenotype in rats after antigen challenge by altering TRPC gene expression.


Effects of galanin and the galanin receptor antagonist galantide on plasma catecholamine levels during a psychosocial stress stimulus in rats.
The aim of this study was to investigate the role played by the neuropeptide galanin (GAL) in the regulation of sympathoadrenal function. We evaluated the effects of rat GAL (rGAL) and of the putative GAL receptor antagonist galantide (GLT) on epinephrine (E) and norepinephrine (NE) plasma levels in conscious freely moving male rats, during a psychosocial stress condition. Four groups of male rats were challenged by a stress stimulus, obtained by exposing the animals to a resident conspecific fighter (intruder model), following an intravenous injection with (1) rGAL + saline (SAL), (2) GLT + SAL, (3) rGAL + GLT, or (4) SAL + SAL. Plasma levels of both E and NE were also measured in an additional group of male rats not exposed to any stressor stimulus. The results (mean+/-SEM) showed that rats exposed to the stressor stimulus (intruder rats) exhibited a significant increase above baseline in circulating levels of both E (peak values of 834.13+/-115.13 pmol/l vs. basal values of 309.31+/-32.93 pmol/l; p < 0.01) and NE (peak values of 5,299.03+/-450.62 pmol/l vs. basal values of 2,798.24+/-311.56 pmol/l; p < 0.01) in comparison to control, nonstressed rats. The comparison of the areas under the curve response (AUC) among treatments in the intruder rats revealed that rGAL + SAL injections resulted in a further increase in E levels when compared to SAL + SAL treatment (AUC values: 8.26+/-0.64 vs. 25.38+/-5.52 nmol/ 1/20 min in SAL + SAL vs. rGAL + SAL treatment, respectively; p < 0.02). No significant changes in stress-induced E plasma levels were found following GLT + SAL treatment in comparison to SAL + SAL injections. When the intruder rats were submitted to rGAL + GLT injections, the increments in E levels were found to be higher than those observed following SAL + SAL treatment (AUC values: 8.26+/-0.64 vs. 36.00+/-13.76 nmol/ 1/20 min in SAL + SAL vs. rGAL + GLT treatment, p < 0.03); however, the values were not significantly different from those observed in rGAL + SAL-injected rats. No significant changes in stress-induced NE levels were found in either treatment group when compared to SAL + SAL-injected intruder rats. The results of this study demonstrate that rGAL administration leads to an increase in the E response to the stress stimulus without any effect on NE response. Galantide does not affect either the physiological stress-induced elevation of plasma catecholamines or the effects of rGAL on E plasma levels in response to stress. Therefore, GLT does not appear to behave as a GAL receptor antagonist in the regulation of sympathoadrenal function in rats.


A multicenter evaluation of how many infants with permanent hearing loss pass a two-stage otoacoustic emissions/automated auditory brainstem response newborn hearing screening protocol.
Ninety percent of all newborns in the United States are now screened for hearing loss before they leave the hospital. Many hospitals use a 2-stage protocol for newborn hearing screening in which all infants are screened first with otoacoustic emissions (OAE). No additional testing is done with infants who pass the OAE, but infants who fail the OAE next are screened with automated auditory brainstem response (A-ABR). Infants who fail the A-ABR screening are referred for diagnostic testing to determine whether they have permanent hearing loss (PHL). Those who pass the A-ABR are considered at low risk for hearing loss and are not tested further. The objective of this multicenter study was to determine whether a substantial number of infants who fail the initial OAE and pass the A-ABR have PHL at approximately 9 months of age. Seven birthing centers with successful newborn hearing screening programs using a 2-stage OAE/A-ABR screening protocol participated. During the study period, 86634 infants were screened for hearing loss at these sites. Of those infants who failed the OAE but passed the A-ABR in at least 1 ear, 1524 were enrolled in the study. Data about prenatal, neonatal, and socioeconomic factors, plus hearing loss risk indicators, were collected for all enrolled infants. When the infants were an average of 9.7 months of age, diagnostic audiologic evaluations were done for 64% of the enrolled infants (1432 ears from 973 infants). Twenty-one infants (30 ears) who had failed the OAE but passed the A-ABR during the newborn hearing screening were identified with permanent bilateral or unilateral hearing loss. Twenty-three (77%) of the ears had mild hearing loss (average of 1 kHz, 2 kHz, and 4 kHz < or =40-decibel hearing level). Nine (43%) infants had bilateral as opposed to unilateral loss, and 18 (86%) infants had sensorineural as opposed to permanent conductive hearing loss. If all infants were screened for hearing loss using the 2-stage OAE/A-ABR newborn hearing screening protocol currently used in many hospitals, then approximately 23% of those with PHL at approximately 9 months of age would have passed the A-ABR. This happens in part because much of the A-ABR screening equipment in current use was designed to identify infants with moderate or greater hearing loss. Thus, program administrators should be certain that the screening program, equipment, and protocols are designed to identify the type of hearing loss targeted by their program. The results also show the need for continued surveillance of hearing status during childhood.


Tissue Drug Concentrations of Anti-tumor Necrosis Factor Agents Are Associated with the Long-term Outcome of Patients with Crohn's Disease.
Many reports indicate that a high-serum trough level of anti-tumor necrosis factor (TNF) agents is required for sustained remission in patients with Crohn's disease The pharmacokinetics of anti-TNF agents in inflamed intestinal tissue, however, is not well investigated. We investigated the association between the tissue concentration of anti-TNF agents and long-term disease outcome. This was a prospective single-center study that enrolled 25 patients with Crohn's disease who were administered infliximab or adalimumab. All participants underwent endoscopy 2 weeks after administration of the anti-TNF agents, and biopsy samples were obtained from both inflamed and noninflamed intestinal tissue. Tissue concentrations of anti-TNF agents were evaluated and the correlation with serum trough levels was compared. The relation between the tissue drug concentration and clinical course over 24 months was also investigated. Concentrations of anti-TNF agents were significantly higher in inflamed tissue than in noninflamed tissue. Patients with high-serum trough concentrations of anti-TNF agents had significantly higher drug levels in the noninflamed tissue than those with low-serum trough concentrations, but no difference in the levels was detected in the inflamed tissue. Patients with high-drug levels in the noninflamed tissue had a significantly higher sustained response rate than patients with low-drug levels. Concentrations of anti-TNF agents in the noninflamed tissue can reflect sustained remission and may be a useful biomarker for monitoring therapeutic intensity in patients with Crohn's disease treated with anti-TNF agents (see Video Abstract, Supplemental Digital Content 1, http://links.lww.com/IBD/B623).


Serum miR-181а and miR-25 Levels in Patients with Breast Cancer or Benign Breast Disease.
Circulating miR-181а and miR-25, which reflect regulation of the expression of carcinogenesis-related genes, were assayed in patients with invasive carcinoma of no specific type (ICNT) or benign breast diseases (BBDs) and in subjects without pathologies of the mammary gland (controls). miR-181а expression level proved to be higher compared to control in patients with fibroadenoma and adenosis with low, but not high, risk of malignant transformation, as well as in patients with luminal HER2-negative type B (Lum B HER2-), HER2-positive type (HER2+), and triple-negative breast cancer (TNBC) than in the controls and luminal-type (Lum A) breast cancer. MiR-25 expression level prevailed in patients with Lum B HER2- compared to control, Lum A, and TNBC patients compared to Lum A. Thus, miR-181а and miR-25 expression levels may be risk indicators of malignant transformation in some patients with BBD, whereas in patients with ICNT, these levels reflect pathological processes of different directions within the tumor.


Three-year clinical evaluation of different restorative resins in class I restorations.
The aim of the present study was to evaluate the three-year clinical performance of a nanofilled resin composite, a packable resin composite, and silorane-based resin restorations in Class I occlusal cavities. Twenty-eight patients with at least three similar-sized occlusal lesions in molar teeth participated in the study. A total of 84 Class I occlusal restorations were placed: 28 with nanofilled resin composite (Filtek Supreme), 28 with packable resin composite (P60), and 28 with silorane-based resin (Filtek Silorane). Filtek Supreme and P60 were used with their respective etch-and-rinse adhesive system, Adper Single Bond 2, and Filtek Silorane was used with its respective self-etch adhesive, Filtek Silorane Adhesive. All restorations were placed by the same operator. The restorations were evaluated at baseline, at six months, and annually for three years according to modified US Public Health Service criteria by two calibrated examiners who did not know which restorative resin had been used. The three restorative materials for each category were compared using the χ (2) test at a significance level of 0.05. Cochran's Q test was used to compare the changes across the five time points for each restorative material. McNemar's test followed by Bonferroni adjustment was used when significance differences were found. At the end of the three years, 60 restorations were evaluated in 20 patients, with a recall rate of 71.4%. The retention rate was 100% for all restorative resins. Eight restorations from the P60 group, ten from the Filtek Supreme group, and nine from the Filtek Silorane group were rated Bravo for marginal discoloration. For marginal adaptation, three P60, five Filtek Supreme, and 11 Filtek Silorane restorations were rated Bravo. No statistically significant differences in overall clinical performance were found between the restorative materials except for marginal adaptation. P60 showed the best marginal adaptation at the end of the three years. No differences were observed between the restorative resins for any of the evaluation criteria tested (p>0.05). None of the restorations showed postoperative sensitivity, secondary caries, or loss of anatomic form. All restorative resins performed equally well in clinical conditions during the three-year evaluation, and no significant differences were found among them, except for marginal adaptation, in which P60 showed superior results.


Carotenoid modulation of immune function and sexual attractiveness in zebra finches.
One hypothesis for why females in many animal species frequently prefer to mate with the most elaborately ornamented males predicts that availability of carotenoid pigments is a potentially limiting factor for both ornament expression and immune function. An implicit assumption of this hypothesis is that males that can afford to produce more elaborate carotenoid-dependent displays must be healthier individuals with superior immunocompetence. However, whether variation in circulating carotenoid levels causes variation in both immune function and sexual attractiveness has not been determined in any species. In this study, we show that manipulation of dietary carotenoid supply invokes parallel changes in cell-mediated immune function and sexual attractiveness in male zebra finches (Taeniopygia guttata).


Discovery of cis-N-(1-(4-(methylamino)cyclohexyl)indolin-6-yl)thiophene-2-carboximidamide: a 1,6-disubstituted indoline derivative as a highly selective inhibitor of human neuronal nitric oxide synthase (nNOS) without any cardiovascular liabilities.
A series of 1,6-disubstituted indoline derivatives were synthesized and evaluated as inhibitors of human nitric oxide synthase (NOS) designed to mitigate the cardiovascular liabilities associated with previously reported tetrahydroquinoline-based selective neuronal NOS inhibitors due to higher lipophilicity ( J. Med. Chem. 2011 , 54 , 5562 - 5575 ). This new series produced similar potency and selectivity among the NOS isoforms and was devoid of any cardiovascular liabilities associated with QT prolongation due to hERG activity or endothelial NOS mediated vasoconstriction effect. The SAR studies led to the identification of cis-45, which was shown to reverse thermal hyperalgesia in vivo in the spinal nerve ligation model of neuropathic pain with excellent safety profile (off-target activities at 80 CNS related receptors/ion channels/transporters). The results presented in this report make cis-45 as an ideal tool for evaluating the potential role of selective nNOS inhibitors in CNS related disorders where excess NO produced by nNOS is thought to play a crucial role.