Activation of the innate immune receptor Dectin-1 upon formation of a 'phagocytic synapse'.

Innate immune cells must be able to distinguish between direct binding to microbes and detection of components shed from the surface of microbes located at a distance. Dectin-1 (also known as CLEC7A) is a pattern-recognition receptor expressed by myeloid phagocytes (macrophages, dendritic cells and neutrophils) that detects β-glucans in fungal cell walls and triggers direct cellular antimicrobial activity, including phagocytosis and production of reactive oxygen species (ROS). In contrast to inflammatory responses stimulated upon detection of soluble ligands by other pattern-recognition receptors, such as Toll-like receptors (TLRs), these responses are only useful when a cell comes into direct contact with a microbe and must not be spuriously activated by soluble stimuli. In this study we show that, despite its ability to bind both soluble and particulate β-glucan polymers, Dectin-1 signalling is only activated by particulate β-glucans, which cluster the receptor in synapse-like structures from which regulatory tyrosine phosphatases CD45 and CD148 (also known as PTPRC and PTPRJ, respectively) are excluded (Supplementary Fig. 1). The 'phagocytic synapse' now provides a model mechanism by which innate immune receptors can distinguish direct microbial contact from detection of microbes at a distance, thereby initiating direct cellular antimicrobial responses only when they are required

Persistence of anticancer activity in berry extracts after simulated gastrointestinal digestion and colonic fermentation

Fruit and vegetable consumption is associated at the population level with a protective effect against colorectal cancer. Phenolic compounds, especially abundant in berries, are of interest due to their putative anticancer activity. After consumption, however, phenolic compounds are subject to digestive conditions within the gastrointestinal tract that alter their structures and potentially their function. However, the majority of phenolic compounds are not efficiently absorbed in the small intestine and a substantial portion pass into the colon. We characterized berry extracts (raspberries, strawberries, blackcurrants) produced by in vitro-simulated upper intestinal tract digestion and subsequent fecal fermentation. These extracts and selected individual colonic metabolites were then evaluated for their putative anticancer activities using in vitro models of colorectal cancer, representing the key stages of initiation, promotion and invasion. Over a physiologically-relevant dose range (0–50 µg/ml gallic acid equivalents), the digested and fermented extracts demonstrated significant anti-genotoxic, anti-mutagenic and anti-invasive activity on colonocytes. This work indicates that phenolic compounds from berries undergo considerable structural modifications during their passage through the gastrointestinal tract but their breakdown products and metabolites retain biological activity and can modulate cellular processes associated with colon cancer

Prediction of adverse maternal outcomes in pre-eclampsia: development and validation of the fullPIERS model.

BACKGROUND: Pre-eclampsia is a leading cause of maternal deaths. These deaths mainly result from eclampsia, uncontrolled hypertension, or systemic inflammation. We developed and validated the fullPIERS model with the aim of identifying the risk of fatal or life-threatening complications in women with pre-eclampsia within 48 h of hospital admission for the disorder. METHODS: We developed and internally validated the fullPIERS model in a prospective, multicentre study in women who were admitted to tertiary obstetric centres with pre-eclampsia or who developed pre-eclampsia after admission. The outcome of interest was maternal mortality or other serious complications of pre-eclampsia. Routinely reported and informative variables were included in a stepwise backward elimination regression model to predict the adverse maternal outcome. We assessed performance using the area under the curve (AUC) of the receiver operating characteristic (ROC). Standard bootstrapping techniques were used to assess potential overfitting. FINDINGS: 261 of 2023 women with pre-eclampsia had adverse outcomes at any time after hospital admission (106 [5%] within 48 h of admission). Predictors of adverse maternal outcome included gestational age, chest pain or dyspnoea, oxygen saturation, platelet count, and creatinine and aspartate transaminase concentrations. The fullPIERS model predicted adverse maternal outcomes within 48 h of study eligibility (AUC ROC 0·88, 95% CI 0·84-0·92). There was no significant overfitting. fullPIERS performed well (AUC ROC >0·7) up to 7 days after eligibility. INTERPRETATION: The fullPIERS model identifies women at increased risk of adverse outcomes up to 7 days before complications arise and can thereby modify direct patient care (eg, timing of delivery, place of care), improve the design of clinical trials, and inform biomedical investigations related to pre-eclampsia. FUNDING: Canadian Institutes of Health Research; UNDP/UNFPA/WHO/World Bank Special Programme of Research, Development, and Research Training in Human Reproduction; Preeclampsia Foundation; International Federation of Obstetricians and Gynecologists; Michael Smith Foundation for Health Research; and Child and Family Research Institute

Combining Membrane Potential Imaging with l-Glutamate or GABA Photorelease

Combining membrane potential imaging using voltage sensitive dyes with photolysis of l-glutamate or GABA allows the monitoring of electrical activity elicited by the neurotransmitter at different sub-cellular sites. Here we describe a simple system and some basic experimental protocols to achieve these measurements. We show how to apply the neurotransmitter and how to vary the dimension of the area of photolysis. We assess the localisation of photolysis and of the recorded membrane potential changes by depolarising the dendrites of cerebellar Purkinje neurons with l-glutamate photorelease using different experimental protocols. We further show in the apical dendrites of CA1 hippocampal pyramidal neurons how l-glutamate photorelease can be used to calibrate fluorescence changes from voltage sensitive dyes in terms of membrane potential changes (in mV) and how GABA photorelease can be used to investigate the phenomenon of shunting inhibition. We also show how GABA photorelease can be used to measure chloride-mediated changes of membrane potential under physiological conditions originating from different regions of a neuron, providing important information on the local intracellular chloride concentrations. The method and the proof of principle reported here open the gateway to a variety of important applications where the advantages of this approach are necessary

Slow GABAA mediated synaptic transmission in rat visual cortex

<p>Abstract</p> <p>Background</p> <p>Previous reports of inhibition in the neocortex suggest that inhibition is mediated predominantly through GABA_Areceptors exhibiting fast kinetics. Within the hippocampus, it has been shown that GABA_Aresponses can take the form of either fast or slow response kinetics. Our findings indicate, for the first time, that the neocortex displays synaptic responses with slow GABA_Areceptor mediated inhibitory postsynaptic currents (IPSCs). These IPSCs are kinetically and pharmacologically similar to responses found in the hippocampus, although the anatomical specificity of evoked responses is unique from hippocampus. Spontaneous slow GABA_AIPSCs were recorded from both pyramidal and inhibitory neurons in rat visual cortex.</p> <p>Results</p> <p>GABA_Aslow IPSCs were significantly different from fast responses with respect to rise times and decay time constants, but not amplitudes. Spontaneously occurring GABA_Aslow IPSCs were nearly 100 times less frequent than fast sIPSCs and both were completely abolished by the chloride channel blocker, picrotoxin. The GABA_Asubunit-specific antagonist, furosemide, depressed spontaneous and evoked GABA_Afast IPSCs, but not slow GABA_A-mediated IPSCs. Anatomical specificity was evident using minimal stimulation: IPSCs with slow kinetics were evoked predominantly through stimulation of layer 1/2 apical dendritic zones of layer 4 pyramidal neurons and across their basal dendrites, while GABA_Afast IPSCs were evoked through stimulation throughout the dendritic arborization. Many evoked IPSCs were also composed of a combination of fast and slow IPSC components.</p> <p>Conclusion</p> <p>GABA_Aslow IPSCs displayed durations that were approximately 4 fold longer than typical GABA_Afast IPSCs, but shorter than GABA_B-mediated inhibition. The anatomical and pharmacological specificity of evoked slow IPSCs suggests a unique origin of synaptic input. Incorporating GABA_Aslow IPSCs into computational models of cortical function will help improve our understanding of cortical information processing.</p

An Extensive Circuitry for Cell Wall Regulation in Candida albicans

Protein kinases play key roles in signaling and response to changes in the external environment. The ability of Candida albicans to quickly sense and respond to changes in its environment is key to its survival in the human host. Our guiding hypothesis was that creating and screening a set of protein kinase mutant strains would reveal signaling pathways that mediate stress response in C. albicans. A library of protein kinase mutant strains was created and screened for sensitivity to a variety of stresses. For the majority of stresses tested, stress response was largely conserved between C. albicans, Saccharomyces cerevisiae, and Schizosaccharomyces pombe. However, we identified eight protein kinases whose roles in cell wall regulation (CWR) were not expected from functions of their orthologs in the model fungi Saccharomyces cerevisiae and Schizosaccharomyces pombe. Analysis of the conserved roles of these protein kinases indicates that establishment of cell polarity is critical for CWR. In addition, we found that septins, crucial to budding, are both important for surviving and are mislocalized by cell wall stress. Our study shows an expanded role for protein kinase signaling in C. albicans cell wall integrity. Our studies suggest that in some cases, this expansion represents a greater importance for certain pathways in cell wall biogenesis. In other cases, it appears that signaling pathways have been rewired for a cell wall integrity response

Sub region-specific modulation of synchronous neuronal burst firing after a kainic acid insult in organotypic hippocampal cultures

<p>Abstract</p> <p>Background</p> <p>Excitotoxicity occurs in a number of pathogenic states including stroke and epilepsy. The adaptations of neuronal circuits in response to such insults may be expected to play an underlying role in pathogenesis. Synchronous neuronal firing can be induced in isolated hippocampal slices and involves all regions of this structure, thereby providing a measure of circuit activity. The effect of an excitotoxic insult (kainic acid, KA) on Mg²⁺-free-induced synchronized neuronal firing was tested in organotypic hippocampal culture by measuring extracellular field activity in CA1 and CA3.</p> <p>Results</p> <p>Within 24 hrs of the insult regional specific changes in neuronal firing patterns were evident as: (i) a dramatic <it>reduction </it>in the ability of CA3 to generate firing; and (ii) a contrasting <it>increase </it>in the frequency and duration of synchronized neuronal firing events in CA1. Two distinct processes underlie the increased propensity of CA1 to generate synchronized burst firing; a lack of ability of the CA3 region to 'pace' CA1 resulting in an increased frequency of synchronized events; and a change in the 'intrinsic' properties limited to the CA1 region, which is responsible for increased event duration. Neuronal quantification using NeuN immunoflurescent staining and stereological confocal microscopy revealed no significant cell loss in hippocampal sub regions, suggesting that changes in the properties of neurons within this region were responsible for the KA-mediated excitability changes.</p> <p>Conclusion</p> <p>These results provide novel insight into adaptation of hippocampal circuits following excitotoxic injury. KA-mediated disruption of the interplay between CA3 and CA1 clearly increases the propensity to synchronized firing in CA1.</p

Changes in preterm birth and stillbirth during COVID-19 lockdowns in 26 countries

Preterm birth (PTB) is the leading cause of infant mortality worldwide. Changes in PTB rates, ranging from −90% to +30%, were reported in many countries following early COVID-19 pandemic response measures (‘lockdowns’). It is unclear whether this variation reflects real differences in lockdown impacts, or perhaps differences in stillbirth rates and/or study designs. Here we present interrupted time series and meta-analyses using harmonized data from 52 million births in 26 countries, 18 of which had representative population-based data, with overall PTB rates ranging from 6% to 12% and stillbirth ranging from 2.5 to 10.5 per 1,000 births. We show small reductions in PTB in the first (odds ratio 0.96, 95% confidence interval 0.95–0.98, P value <0.0001), second (0.96, 0.92–0.99, 0.03) and third (0.97, 0.94–1.00, 0.09) months of lockdown, but not in the fourth month of lockdown (0.99, 0.96–1.01, 0.34), although there were some between-country differences after the first month. For high-income countries in this study, we did not observe an association between lockdown and stillbirths in the second (1.00, 0.88–1.14, 0.98), third (0.99, 0.88–1.12, 0.89) and fourth (1.01, 0.87–1.18, 0.86) months of lockdown, although we have imprecise estimates due to stillbirths being a relatively rare event. We did, however, find evidence of increased risk of stillbirth in the first month of lockdown in high-income countries (1.14, 1.02–1.29, 0.02) and, in Brazil, we found evidence for an association between lockdown and stillbirth in the second (1.09, 1.03–1.15, 0.002), third (1.10, 1.03–1.17, 0.003) and fourth (1.12, 1.05–1.19, <0.001) months of lockdown. With an estimated 14.8 million PTB annually worldwide, the modest reductions observed during early pandemic lockdowns translate into large numbers of PTB averted globally and warrant further research into causal pathways