Various species of the intestinal microbiota have been associated with the development of colorectal cancer
, but it has not been demonstrated that bacteria have a direct role in the occurrence of ...oncogenic mutations. Escherichia coli can carry the pathogenicity island pks, which encodes a set of enzymes that synthesize colibactin
. This compound is believed to alkylate DNA on adenine residues
and induces double-strand breaks in cultured cells
. Here we expose human intestinal organoids to genotoxic pks
E. coli by repeated luminal injection over five months. Whole-genome sequencing of clonal organoids before and after this exposure revealed a distinct mutational signature that was absent from organoids injected with isogenic pks-mutant bacteria. The same mutational signature was detected in a subset of 5,876 human cancer genomes from two independent cohorts, predominantly in colorectal cancer. Our study describes a distinct mutational signature in colorectal cancer and implies that the underlying mutational process results directly from past exposure to bacteria carrying the colibactin-producing pks pathogenicity island.
Escherichia coli sequence type 131 (ST131) is a worldwide pandemic clone, causing predominantly community-onset antimicrobial-resistant infection. Its pandemic spread was identified in 2008 by ...utilizing multilocus sequence typing (MLST) of CTX-M-15 extended-spectrum β-lactamase-producing E. coli from three continents. Subsequent research has confirmed the worldwide prevalence of ST131 harbouring a broad range of virulence and resistance genes on a transferable plasmid. A high prevalence of the clone (∼30%-60%) has been identified amongst fluoroquinolone-resistant E. coli. In addition, it potentially harbours a variety of β-lactamase genes; most often, these include CTX-M family β-lactamases, and, less frequently, TEM, SHV and CMY β-lactamases. Our knowledge of ST131's geographical distribution is incomplete. A broad distribution has been demonstrated amongst antimicrobial-resistant E. coli from human infection in Europe (particularly the UK), North America, Canada, Japan and Korea. High rates are suggested from limited data in Asia, the Middle East and Africa. The clone has also been detected in companion animals, non-companion animals and foods. The clinical spectrum of disease described is similar to that for other E. coli, with urinary tract infection predominant. This can range from cystitis to life-threatening sepsis. Infection occurs in humans of all ages. Therapy must be tailored to the antimicrobial resistance phenotype of the infecting isolate and the site of infection. Phenotypic detection of the ST131 clone is not possible and DNA-based techniques, including MLST and PCR, are described.
Aims/hypothesis
Several glucose-sensing pathways have been implicated in glucose-triggered secretion of glucagon-like peptide-1 (GLP-1) from intestinal L cells. One involves glucose metabolism and ...closure of ATP-sensitive K
+
channels, and another exploits the electrogenic nature of Na
+
-coupled glucose transporters (SGLTs). This study aimed to elucidate the role of these distinct mechanisms in glucose-stimulated GLP-1 secretion.
Methods
Glucose uptake into L cells (either GLUTag cells or cells in primary cultures, using a new transgenic mouse model combining proglucagon promoter-driven
Cre
recombinase with a ROSA26tdRFP reporter) was monitored with the FLII
12
Pglu-700μδ6 glucose sensor. Effects of pharmacological and genetic interference with SGLT1 or facilitative glucose transport (GLUT) on intracellular glucose accumulation and metabolism (measured by NAD(P)H autofluorescence), cytosolic Ca
2+
(monitored with Fura2) and GLP-1 secretion (assayed by ELISA) were assessed.
Results
L cell glucose uptake was dominated by GLUT-mediated transport, being abolished by phloretin but not phloridzin. NAD(P)H autofluorescence was glucose dependent and enhanced by a glucokinase activator. In GLUTag cells, but not primary L cells, phloretin partially impaired glucose-dependent secretion, and suppressed an amplifying effect of glucose under depolarising high K
+
conditions. The key importance of SGLT1 in GLUTag and primary cells was evident from the impairment of secretion by phloridzin or
Sglt1
knockdown and failure of glucose to trigger cytosolic Ca
2+
elevation in primary L cells from
Sglt1
knockout mice.
Conclusions/interpretation
SGLT1 acts as the luminal glucose sensor in L cells, but intracellular glucose concentrations are largely determined by GLUT activity. Although L cell glucose metabolism depends partially on glucokinase activity, this plays only a minor role in glucose-stimulated GLP-1 secretion.
The scattering of dark matter particles off nuclei in direct detection experiments can be described in terms of a multidimensional effective field theory (EFT). A new systematic analysis technique is ...developed using the EFT approach and Bayesian inference methods to exploit, when possible, the energy-dependent information of the detected events, experimental efficiencies, and backgrounds. Highly dimensional likelihoods are calculated over the mass of the weakly interacting massive particle (WIMP) and multiple EFT coupling coefficients, which can then be used to set limits on these parameters and choose models (EFT operators) that best fit the direct detection data. Expanding the parameter space beyond the standard spin-independent isoscalar cross section and WIMP mass reduces tensions between previously published experiments. Combining these experiments to form a single joint likelihood leads to stronger limits than when each experiment is considered on its own. Simulations using two nonstandard operators (O3 and O8) are used to test the proposed analysis technique in up to five dimensions and demonstrate the importance of using multiple likelihood projections when determining constraints on WIMP mass and EFT coupling coefficients. In particular, this shows that an explicit momentum dependence in dark matter scattering can be identified.
Aims/hypothesis Glucose-dependent insulinotropic polypeptide (GIP) is an incretin hormone with anti-apoptotic effects on the pancreatic beta cell. The aim of this study was to generate transgenic ...mice with fluorescently labelled GIP-secreting K cells and to use these to investigate pathways by which K cells detect nutrients. Methods Transgenic mice were generated in which the GIP promoter drives the expression of the yellow fluorescent protein Venus. Fluorescent cells were purified by flow cytometry and analysed by quantitative RT-PCR. GIP secretion was assayed in primary cultures of small intestine. Results Expression of Venus in transgenic mice was restricted to K cells, as assessed by immunofluorescence and measurements of the Gip mRNA and GIP protein contents of purified cells. K cells expressed high levels of mRNA for Kir6.2 (also known as Kcnj11), Sur1 (also known as Abcc8), Sglt1 (also known as Slc5a1), and of the G-protein-coupled lipid receptors Gpr40 (also known as Ffar1), Gpr119 and Gpr120. In primary cultures, GIP release was stimulated by glucose, glutamine and linoleic acid, and potentiated by forskolin plus 3-isobutyl-1-methylxanthine (IBMX), but was unaffected by the artificial sweetener sucralose. Secretion was half-maximal at 0.6 mmol/l glucose and partially mimicked by α-methylglucopyranoside, suggesting the involvement of SGLT1. Tolbutamide triggered secretion under basal conditions, whereas diazoxide suppressed responses in forskolin/IBMX. Conclusions/interpretation These transgenic mice and primary culture techniques provide novel opportunities to interrogate the mechanisms of GIP secretion. Glucose-triggered GIP secretion was SGLT1-dependent and modulated by KATP channel activity but not determined by sweet taste receptors. Synergistic stimulation by elevated cAMP and glucose suggests that targeting appropriate G-protein-coupled receptors may provide opportunities to modulate GIP release in vivo.
Self-assembled enantiomers of an asymmetric di-iron metallohelix differ in their antiproliferative activities against HCT116 colon cancer cells such that the compound with Λ-helicity at the metals ...becomes more potent than the Δ compound with increasing exposure time. From concentration- and temperature-dependent
Fe isotopic labelling studies of cellular accumulation we postulate that while the more potent Λ enantiomer undergoes carrier-mediated efflux, for Δ the process is principally equilibrative. Cell fractionation studies demonstrate that both enantiomers localise in a similar fashion; compound is observed mostly within the cytoskeleton and/or genomic DNA, with significant amounts also found in the nucleus and membrane, but with negligible concentration in the cytosol. Cell cycle analyses using flow cytometry reveal that the Δ enantiomer induces mild arrest in the G
phase, while Λ causes a very large dose-dependent increase in the G
/M population at a concentration significantly below the relevant IC
. Correspondingly, G
-M checkpoint failure as a result of Λ-metallohelix binding to DNA is shown to be feasible by linear dichroism studies, which indicate, in contrast to the Δ compound, a quite specific mode of binding, probably in the major groove. Further, spindle assembly checkpoint (SAC) failure, which could also be responsible for the observed G
/M arrest, is established as a feasible mechanism for the Λ helix
drug combination (synergy) studies and the discovery of tubulin and actin inhibition. Here, while the Λ compound stabilizes F-actin and induces a distinct change in tubulin architecture of HCT116 cells, Δ promotes depolymerization and more subtle changes in microtubule and actin networks.
Intention-to-treat analyses do not address adherence. Per protocol analyses treat nonadherence as a protocol deviation and assess if the intervention is effective if followed.
To determine the rate ...of early preterm birth (EPTB, <34 weeks gestation) and preterm birth (PTB, <37 weeks gestation) in participants who adhered to a randomly assigned docosahexaenoic acid (DHA) dose of 1000 mg/day.
Eleven hundred women with a singleton pregnancy were enrolled before 20-weeks’ gestation, provided a capsule with 200 mg/day DHA and randomly assigned to two additional capsules containing a placebo or 800 mg of DHA. In the Bayesian Adaptive Design, new randomization schedules were determined at prespecified intervals. In each randomization, the group with the most EPTB was assigned fewer participants than the other group. Adherence was defined a priori as a postpartum red blood cell phospholipid DHA (RBC-PL-DHA) ≥5.5%.and post hoc as ≥8.0% RBC-PL-DHA, the latter after examination of postpartum RBC-PL-DHA. Bayesian mixture models were fitted for gestational age and dichotomized for EPTB and PTB as a function of baseline RBC-PL-DHA and dose-adherence. Bayesian hierarchical models were also fitted for EPTB by dose adherence and quartiles of baseline RBC-PL-DHA.
Adherence to the high dose using both RBC-PL-DHA cut points resulted in less EPTB compared to 200 mg Bayesian posterior probability (pp) = 0.93 and 0.92, respectively. For participants in the two lowest quartiles of baseline DHA status, adherence to the higher dose resulted in lower EPTB (≥5.5% RBC-PL-DHA, quartiles 1 and 2, pp = 0.95 and 0.96; ≥8% RBC-PL-DHA, quartiles 1 and 2, pp = 0.94 and 0.95). Using the Bayesian model, EPTB was reduced by 65%, from 3.45% to 1.2%, using both cut points. Adherence also reduced PTB before 35, 36 and 37 weeks using both cut points (pp ≥ 0.95). In general, performance of the nonadherent subgroup mirrored that of participants assigned to 200 mg.
Adherence to high dose DHA reduced EPTB and PTB. The largest effect of adherence on reducing EPTB was observed in women with low baseline DHA levels. ClinicalTrials.gov (NCT02626299).
We investigate the origin of archaeological wool textiles preserved by anoxic waterlogging from seven medieval archaeological deposits in north-western Europe (c. 700-1600 AD), using geospatial ...patterning in carbon (δ13C), nitrogen (δ15N) and non-exchangeable hydrogen (δ2H) composition of modern and ancient sheep proteins. δ13C, δ15N and δ2H values from archaeological wool keratin (n = 83) and bone collagen (n = 59) from four sites were interpreted with reference to the composition of modern sheep wool from the same regions. The isotopic composition of wool and bone collagen samples clustered strongly by settlement; inter-regional relationships were largely parallel in modern and ancient samples, though landscape change was also significant. Degradation in archaeological wool samples, examined by elemental and amino acid composition, was greater in samples from Iceland (Reykholt) than in samples from north-east England (York, Newcastle) or northern Germany (Hessens). A nominal assignment approach was used to classify textiles into local/non-local at each site, based on maximal estimates of isotopic variability in modern sheep wool. Light element stable isotope analysis provided new insights into the origins of wool textiles, and demonstrates that isotopic provenancing of keratin preserved in anoxic waterlogged contexts is feasible. We also demonstrate the utility of δ2H analysis to understand the location of origin of archaeological protein samples.
Autism is a common disorder of childhood, affecting 1 in 500 children. Yet, it often remains unrecognized and undiagnosed until or after late preschool age because appropriate tools for routine ...developmental screening and screening specifically for autism have not been available. Early identification of children with autism and intensive, early intervention during the toddler and preschool years improves outcome for most young children with autism. This practice parameter reviews the available empirical evidence and gives specific recommendations for the identification of children with autism. This approach requires a dual process: 1) routine developmental surveillance and screening specifically for autism to be performed on all children to first identify those at risk for any type of atypical development, and to identify those specifically at risk for autism; and 2) to diagnose and evaluate autism, to differentiate autism from other developmental disorders.
A
bstract
The MicroBooNE liquid argon time projection chamber located at Fermilab is a neutrino experiment dedicated to the study of short-baseline oscillations, the measurements of neutrino cross ...sections in liquid argon, and to the research and development of this novel detector technology. Accurate and precise measurements of calorimetry are essential to the event reconstruction and are achieved by leveraging the TPC to measure deposited energy per unit length along the particle trajectory, with mm resolution. We describe the non-uniform calorimetric reconstruction performance in the detector, showing dependence on the angle of the particle trajectory. Such non-uniform reconstruction directly affects the performance of the particle identification algorithms which infer particle type from calorimetric measurements. This work presents a new particle identification method which accounts for and effectively addresses such non-uniformity. The newly developed method shows improved performance compared to previous algorithms, illustrated by a 93.7% proton selection efficiency and a 10% muon mis-identification rate, with a fairly loose selection of tracks performed on beam data. The performance is further demonstrated by identifying exclusive final states in
ν
μ
CC
interactions. While developed using MicroBooNE data and simulation, this method is easily applicable to future LArTPC experiments, such as SBND, ICARUS, and DUNE.
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