Developing influenza and respiratory syncytial virus activity thresholds for syndromic surveillance in England.

Influenza and respiratory syncytial virus (RSV) are common causes of respiratory tract infections and place a burden on health services each winter. Systems to describe the timing and intensity of such activity will improve the public health response and deployment of interventions to these pressures. Here we develop early warning and activity intensity thresholds for monitoring influenza and RSV using two novel data sources: general practitioner out-of-hours consultations (GP OOH) and telehealth calls (NHS 111). Moving Epidemic Method (MEM) thresholds were developed for winter 2017-2018. The NHS 111 cold/flu threshold was breached several weeks in advance of other systems. The NHS 111 RSV epidemic threshold was breached in week 41, in advance of RSV laboratory reporting. Combining the use of MEM thresholds with daily monitoring of NHS 111 and GP OOH syndromic surveillance systems provides the potential to alert to threshold breaches in real-time. An advantage of using thresholds across different health systems is the ability to capture a range of healthcare-seeking behaviour, which may reflect differences in disease severity. This study also provides a quantifiable measure of seasonal RSV activity, which contributes to our understanding of RSV activity in advance of the potential introduction of new RSV vaccines.

Impact of Centralizing Gastric Cancer Surgery on Treatment, Morbidity, and Mortality.

INTRODUCTION: Centralization of gastric cancer surgery is thought to improve outcome and has been imposed in the Netherlands since 2012. This study analyzes the effect of centralization in terms of treatment outcome and survival in the Eastern part of the Netherlands. METHODS: All gastric cancer patients without distant metastases who underwent a gastrectomy in six hospitals in the Eastern part of the Netherlands between 2008 and 2011 (pre-centralization) and 2013-2016 (post-centralization) were selected from the Netherlands Cancer Registry. Patient and tumor characteristics and treatment outcomes (duration of surgery, blood loss, resection margin, lymphadenectomy, chemotherapy, postoperative complications and hospital stay, and overall and disease-free survival) were analyzed and compared between pre- and post-centralization. RESULTS: One hundred forty-four patients were included pre-centralization and 106 patients post-centralization. Patient and tumor characteristics were almost similar in the two periods. After centralization, more patients were treated with perioperative chemotherapy (25 vs. 42% p < 0.01). The proportion of patients treated with an adequate lymphadenectomy (21 vs. 93% p < 0.01) and laparoscopic surgery (6 vs. 40% p < 0.01) increased significantly (p < 0.01). The amount of cardiac complications (16 vs. 7.5% p < 0.05) decreased; however, complications needing a re-intervention were comparable (42 vs. 40% p = 0.79). Median hospital stay decreased from 10 to 8 days (p < 0.01). A 30-day mortality did not differ significantly (4.2 vs. 1.9%). A 1-year overall (78 vs. 80% p = 0.17) and disease-free survival (73 vs. 74% p = 0.66) remained stable. DISCUSSION: Centralizing gastric cancer treatment in the Eastern part of the Netherlands resulted in improved lymph node harvesting and a successful introduction of laparoscopic gastrectomies. Centralization has not translated into improved mortality, and other variables may also have led to these improved outcomes. Further research using a nationwide population-based study will be needed to confirm these data.

The burden of seasonal respiratory infections on a national telehealth service in England.

Seasonal respiratory illnesses present a major burden on primary care services. We assessed the burden of respiratory illness on a national telehealth system in England and investigated the potential for providing early warning of respiratory infection. We compared weekly laboratory reports for respiratory pathogens with telehealth calls (NHS 111) between week 40 in 2013 and week 29 in 2015. Multiple linear regression was used to identify which pathogens had a significant association with respiratory calls. Children aged <5 and 5-14 years, and adults over 65 years were modelled separately as were time lags of up to 4 weeks between calls and laboratory specimen dates. Associations with respiratory pathogens explained over 83% of the variation in cold/flu, cough and difficulty breathing calls. Based on the first two seasons available, the greatest burden was associated with respiratory syncytial virus (RSV) and influenza, with associations found in all age bands. The most sensitive signal for influenza was calls for 'cold/flu', whilst for RSV it was calls for cough. The best-fitting models showed calls increasing a week before laboratory specimen dates. Daily surveillance of these calls can provide early warning of seasonal rises in influenza and RSV, contributing to the national respiratory surveillance programme.

Developing and validating a new national remote health advice syndromic surveillance system in England.

Background: Public Health England (PHE) coordinates a suite of real-time national syndromic surveillance systems monitoring general practice, emergency department and remote health advice data. We describe the development and informal evaluation of a new syndromic surveillance system using NHS 111 remote health advice data. Methods: NHS 111 syndromic indicators were monitored daily at national and local level. Statistical models were applied to daily data to identify significant exceedances; statistical baselines were developed for each syndrome and area using a multi-level hierarchical mixed effects model. Results: Between November 2013 and October 2014, there were on average 19 095 NHS 111 calls each weekday and 43 084 each weekend day in the PHE dataset. There was a predominance of females using the service (57%); highest percentage of calls received was in the age group 1-4 years (14%). This system was used to monitor respiratory and gastrointestinal infections over the winter of 2013-14, the potential public health impact of severe flooding across parts of southern England and poor air quality episodes across England in April 2014. Conclusions: This new system complements and supplements the existing PHE syndromic surveillance systems and is now integrated into the routine daily processes that form this national syndromic surveillance service.

Measuring residual activity of topical antimicrobials: is the residual activity of chlorhexidine an artefact of laboratory methods?

Residual activity of chlorhexidine gluconate (CHG) was evaluated by pretreating hands with CHG and then touching Staphylococcus aureus dried on to stainless steel discs. By this method, no reduction in bacteria was observed up to 15 min, suggesting that residual CHG does not offer protection against contamination with transient micro-organisms in clinical practice.

A survey of UK dental health professionals using a medicines information service: what questions do they ask and do they get useful answers?

Dentists prescribe a limited range of medicines but it is important that they consider the effects of all medicines their patients are taking when providing dental care. In the UK, a national medicines information (UKMi) service funded by the National Health Service is available to advise health professionals on prescribing and to support evidence-based practice. This paper presents the results of a survey of 151 dental health professionals who contacted the UKMi service for advice. Enquiries most commonly involved antibiotics (32%), but dental health professionals also asked for advice on legal issues relating to medicines (10%), and on managing patients receiving bisphosphonates (9%), local anaesthetics (6%) and antiplatelet drugs (5%). One hundred and forty-six (97%) enquirers used the advice provided: for managing current patients, planning the care of future patients, for continuing professional development and teaching others. Two thirds of enquirers used the information provided to check if current or proposed management was appropriate, one half to change therapy and over one quarter to identify, manage or avoid adverse effects or drug interactions.

Per-arnt-sim (PAS) domain-containing protein kinase is downregulated in human islets in type 2 diabetes and regulates glucagon secretion.

AIMS/HYPOTHESIS: We assessed whether per-arnt-sim (PAS) domain-containing protein kinase (PASK) is involved in the regulation of glucagon secretion. METHODS: mRNA levels were measured in islets by quantitative PCR and in pancreatic beta cells obtained by laser capture microdissection. Glucose tolerance, plasma hormone levels and islet hormone secretion were analysed in C57BL/6 Pask homozygote knockout mice (Pask-/-) and control littermates. Alpha-TC1-9 cells, human islets or cultured E13.5 rat pancreatic epithelia were transduced with anti-Pask or control small interfering RNAs, or with adenoviruses encoding enhanced green fluorescent protein or PASK. RESULTS: PASK expression was significantly lower in islets from human type 2 diabetic than control participants. PASK mRNA was present in alpha and beta cells from mouse islets. In Pask-/- mice, fasted blood glucose and plasma glucagon levels were 25 ± 5% and 50 ± 8% (mean ± SE) higher, respectively, than in control mice. At inhibitory glucose concentrations (10 mmol/l), islets from Pask-/- mice secreted 2.04 ± 0.2-fold (p < 0.01) more glucagon and 2.63 ± 0.3-fold (p < 0.01) less insulin than wild-type islets. Glucose failed to inhibit glucagon secretion from PASK-depleted alpha-TC1-9 cells, whereas PASK overexpression inhibited glucagon secretion from these cells and human islets. Extracellular insulin (20 nmol/l) inhibited glucagon secretion from control and PASK-deficient alpha-TC1-9 cells. PASK-depleted alpha-TC1-9 cells and pancreatic embryonic explants displayed increased expression of the preproglucagon (Gcg) and AMP-activated protein kinase (AMPK)-alpha2 (Prkaa2) genes, implying a possible role for AMPK-alpha2 downstream of PASK in the control of glucagon gene expression and release. CONCLUSIONS/INTERPRETATION: PASK is involved in the regulation of glucagon secretion by glucose and may be a useful target for the treatment of type 2 diabetes.

Tolerance to 3,4-methylenedioxymethamphetamine in rats exposed to single high-dose binges.

3,4-Methylenedioxymethamphetamine (MDMA or ecstasy) stimulates the transporter-mediated release of monoamines, including 5-HT. High-dose exposure to MDMA causes persistent 5-HT deficits (e.g. depletion of brain 5-HT) in animals, yet the functional and clinical relevance of such deficits are poorly defined. Here we examine functional consequences of MDMA-induced 5-HT depletions in rats. Male rats received binges of three i.p. injections of MDMA or saline, one injection every 2 h; MDMA was given at a threshold pharmacological dose (1.5 mg/kgx3, low dose) or at a fivefold higher amount (7.5 mg/kgx3, high dose). One week later, jugular catheters and intracerebral guide cannulae were implanted. Two weeks after binges, rats received acute i.v. challenge injections of 1 and 3 mg/kg MDMA. Neuroendocrine effects evoked by i.v. MDMA (prolactin and corticosterone secretion) were assessed via serial blood sampling, while neurochemical effects (5-HT and dopamine release) were assessed via microdialysis in brain. MDMA binges elevated core temperatures only in the high-dose group, with these same rats exhibiting approximately 50% loss of forebrain 5-HT 2 weeks later. Prior exposure to MDMA did not alter baseline plasma hormones or dialysate monoamines, and effects of i.v. MDMA were similar in saline and low-dose groups. By contrast, rats pretreated with high-dose MDMA displayed significant reductions in evoked hormone secretion and 5-HT release when challenged with i.v. MDMA. As tolerance developed only in rats exposed to high-dose binges, hyperthermia and 5-HT depletion are implicated in this phenomenon. Our results suggest that MDMA tolerance in humans may reflect 5-HT deficits which could contribute to further dose escalation.

Regulation by Per-Arnt-Sim (PAS) kinase of pancreatic duodenal homeobox-1 nuclear import in pancreatic beta-cells.

The transcription factor PDX-1 (pancreatic duodenal homeobox-1) is required for normal pancreatic development and for the function of insulin-producing islet beta-cells in mammals. We have shown previously that glucose regulates insulin gene expression in part through the activation and translocation of PDX-1 from the nuclear periphery to the nucleoplasm. We have also found that PASK [PAS (Per-Arnt-Sim) kinase], a member of the nutrient-regulated family of protein kinases, is activated in response to glucose challenge in beta-cells and is involved in the regulation of expression of PDX-1. Purified PASK efficiently phosphorylated recombinant PDX-1 in vitro on a single site (Thr-152). To determine the impact of phosphorylation at this site, we generated wild-type and mutant (T152A, T152D and T152E) forms of PDX-1 and examined the distribution of each of these in clonal MIN6 beta-cells by immunocytochemical analysis. Unexpectedly, only the T152D mutation significantly affected subcellular distribution, increasing the ratio of nuclear/cytosolic labelling at low and high glucose concentrations, suggesting that phosphorylation at Thr-152 inhibits nuclear uptake in response to glucose. Based on these results, experiments to examine the contribution of Thr-152 to the overall phosphorylation of PDX-1 in intact cells will be undertaken.

Synergistic induction of matrix metalloproteinase 1 by interleukin-1alpha and oncostatin M in human chondrocytes involves signal transducer and activator of transcription and activator protein 1 transcription factors via a novel mechanism.

OBJECTIVE: To investigate the mechanism of interleukin-1alpha (IL-1alpha) and oncostatin M (OSM) synergistic regulation of matrix metalloproteinase 1 (MMP-1) in human chondrocytes. METHODS: Using an immortalized human chondrocyte cell line (T/C28a4), we investigated regulation of the MMP-1 gene. Northern blotting and flow cytometric analysis were used to assess changes in receptor, MMP-1, and c-fos expression. Transient transfections using MMP-1 promoter/luciferase constructs, electrophoretic mobility shift assay, and site-directed mutagenesis were used to investigate MMP-1 promoter activation. RESULTS: We found no alteration in the expression of receptors used by these cytokines after stimulation with IL-1alpha/OSM. Using MMP-1 promoter/luciferase reporter constructs, we found that the proximal (-517/+63) region of the MMP-1 promoter was sufficient to support a synergistic activation. A role for activated signal transducers and activators of transcription (STAT-3) was demonstrated, although no binding of STAT-3 to the MMP-1 promoter was found. However, constitutive binding of activator protein 1 (AP-1) was detected, and changes in c-fos expression could modulate promoter activity. CONCLUSION: Since no changes in receptor expression were observed, receptor modulation cannot account for the IL-1alpha/OSM synergy observed. Instead, the interplay of various intracellular signaling pathways is a more likely explanation. STAT activation is required, but STAT proteins do not interact directly with the MMP-1 promoter. We propose that activated STATs stimulate c-fos expression, and changes in expression of the AP-1 components regulate MMP-1 expression. We highlight a new mechanism for MMP-1 regulation in human chondrocytes that could provide potential new therapeutic targets.

Regulation of clock and NPAS2 DNA binding by the redox state of NAD cofactors.

Clock:BMAL1 and NPAS2:BMAL1 are heterodimeric transcription factors that control gene expression as a function of the light-dark cycle. Although built to fluctuate at or near a 24-hour cycle, the clock can be entrained by light, activity, or food. Here we show that the DNA-binding activity of the Clock:BMAL1 and NPAS2:BMAL1 heterodimers is regulated by the redox state of nicotinamide adenine dinucleotide (NAD) cofactors in a purified system. The reduced forms of the redox cofactors, NAD(H) and NADP(H), strongly enhance DNA binding of the Clock:BMAL1 and NPAS2:BMAL1 heterodimers, whereas the oxidized forms inhibit. These observations raise the possibility that food, neuronal activity, or both may entrain the circadian clock by direct modulation of cellular redox state.

PAS kinase: an evolutionarily conserved PAS domain-regulated serine/threonine kinase.

PAS domains regulate the function of many intracellular signaling pathways in response to both extrinsic and intrinsic stimuli. PAS domain-regulated histidine kinases are common in prokaryotes and control a wide range of fundamental physiological processes. Similarly regulated kinases are rare in eukaryotes and are to date completely absent in mammals. PAS kinase (PASK) is an evolutionarily conserved gene product present in yeast, flies, and mammals. The amino acid sequence of PASK specifies two PAS domains followed by a canonical serine/threonine kinase domain, indicating that it might represent the first mammalian PAS-regulated protein kinase. We present evidence that the activity of PASK is regulated by two mechanisms. Autophosphorylation at two threonine residues located within the activation loop significantly increases catalytic activity. We further demonstrate that the N-terminal PAS domain is a cis regulator of PASK catalytic activity. When the PAS domain-containing region is removed, enzyme activity is significantly increased, and supplementation of the purified PAS-A domain in trans selectively inhibits PASK catalytic activity. These studies define a eukaryotic signaling pathway suitable for studies of PAS domains in a purified in vitro setting.

Collection of genomic DNA from adults in epidemiological studies by buccal cytobrush and mouthwash.

Blood samples are an excellent source of large amounts of genomic DNA. However, alternative sources are often needed in epidemiological studies because of difficulties in obtaining blood samples. This report evaluates the buccal cytobrush and alcohol-containing mouthwash protocols for collecting DNA by mail. Several DNA extraction techniques are also evaluated. The study was conducted in two phases. In phase 1, we compared cytobrush and mouthwash samples collected by mail in two different epidemiological studies: (a) cytobrush samples (n = 120) from a United States case-control study of breast cancer; and (b) mouthwash samples (n = 40) from a prospective cohort of male United States farmers. Findings from phase 1 were confirmed in phase 2, where we randomized cytobrush (n = 28) and mouthwash (n = 25) samples among participants in the breast cancer study to directly compare both collection methods. The median human DNA yield determined by hybridization with a human DNA probe from phenol-chloroform extracts was 1.0 and 1.6 microg/2 brushes for phases 1 and 2, respectively, and 27.5 and 16.6 microg/mouthwash sample for phases 1 and 2, respectively. Most (94-100%) mouthwash extracts contained high molecular weight DNA (>23 kb), in contrast to 55-61% of the brush extracts. PCR success rates for amplification of beta-globin gene fragments (268, 536, and 989 bp) were similar for cytobrush and mouthwash phenol-chloroform extracts (range, 94.4-100%). Also, we obtained high success rates in determining the number of CAG repeats in the androgen receptor gene, characterizing tetranucleotide microsatellites in six gene loci, and screening for mutations in the BRCA1/2 genes in a subset of phenol-chloroform DNA extracts. Relative to DNA extracted by phenol-chloroform from cytobrush samples, DNA extracted by NaOH had lower molecular weight, decreased PCR success rates for most assays performed, and unreliably high spectrophotometer readings for DNA yields. In conclusion, although DNA isolated from either mouthwash or cytobrush samples collected by mail from adults is adequate for a wide range of PCR-based assays, a single mouthwash sample provides substantially larger amounts and higher molecular weight DNA than two cytobrush samples.

Manganese superoxide dismutase signals matrix metalloproteinase expression via H2O2-dependent ERK1/2 activation.

Manganese-superoxide dismutase (Sod2) removes mitochondrially derived superoxide (O(2)) at near-diffusion limiting rates and is the only antioxidant enzyme whose expression is regulated by numerous stimuli. Here it is shown that Sod2 also serves as a source of the intracellular signaling molecule H(2)O(2). Sod2-dependent increases in the steady-state levels of H(2)O(2) led to ERK1/2 activation and subsequent downstream transcriptional increases in matrix metalloproteinase-1 (MMP-1) expression, which were reversed by expression of the H(2)O(2)-detoxifying enzyme, catalase. In addition, a single nucleotide polymorphism has recently been identified (1G/2G) at base pair--1607 that creates an Ets site adjacent to an AP-1 site at base pair --1602 and has been shown to dramatically enhance transcription of the MMP-1 promoter. Luciferase promoter constructs containing either the 1G or 2G variation were 25- or 1000-fold more active when transiently transfected into Sod2-overexpressing cell lines, respectively. The levels of MMP-2, -3, and -7 were also increased in the Sod2-overexpressing cell lines, suggesting that Sod2 may function as a "global" redox regulator of MMP expression. In addition, Sod2(-/+) mouse embryonic fibroblasts failed to respond to the cytokine-mediated induction of the murine functional analog of MMP-1, MMP-13. This study provides evidence that the modulation of Sod2 activity by a wide array of pathogenic and inflammatory stimuli may be utilized by the cell as a primary signaling mechanism leading to matrix metalloproteinase expression.