Deciphering interplay between Salmonella invasion effectors

Bacterial pathogens have evolved a specialized type III secretion system (T3SS) to translocate virulence effector proteins directly into eukaryotic target cells. Salmonellae deploy effectors that trigger localized actin reorganization to force their own entry into non-phagocytic host cells. Six effectors (SipC, SipA, SopE/2, SopB, SptP) can individually manipulate actin dynamics at the plasma membrane, which acts as a ‘signaling hub’ during Salmonella invasion. The extent of crosstalk between these spatially coincident effectors remains unknown. Here we describe trans and cis binary entry effector interplay (BENEFIT) screens that systematically examine functional associations between effectors following their delivery into the host cell. The results reveal extensive ordered synergistic and antagonistic relationships and their relative potency, and illuminate an unexpectedly sophisticated signaling network evolved through longstanding pathogen–host interaction

PAin SoluTions In the Emergency Setting (PASTIES)--patient controlled analgesia versus routine care in emergency department patients with non-traumatic abdominal pain: randomised trial

© Smith et al 2015. OBJECTIVE: To determine whether patient controlled analgesia (PCA) is better than routine care in providing effective analgesia for patients presenting to emergency departments with moderate to severe non-traumatic abdominal pain.DESIGN: Pragmatic, multicentre, parallel group, randomised controlled trialSETTING: Five English hospitals.PARTICIPANTS: 200 adults (66% (n=130) female), aged 18 to 75 years, who presented to the emergency department requiring intravenous opioid analgesia for the treatment of moderate to severe non-traumatic abdominal pain and were expected to be admitted to hospital for at least 12 hours.INTERVENTIONS: Patient controlled analgesia or nurse titrated analgesia (treatment as usual).MAIN OUTCOME MEASURES: The primary outcome was total pain experienced over the 12 hour study period, derived by standardised area under the curve (scaled from 0 to 100) of each participant's hourly pain scores, captured using a visual analogue scale. Pre-specified secondary outcomes included total morphine use, percentage of study period in moderate or severe pain, percentage of study period asleep, length of hospital stay, and satisfaction with pain management.RESULTS: 196 participants were included in the primary analyses (99 allocated to PCA and 97 to treatment as usual). Mean total pain experienced was 35.3 (SD 25.8) in the PCA group compared with 47.3 (24.7) in the treatment as usual group. The adjusted between group difference was 6.3 (95% confidence interval 0.7 to 11.9). Participants in the PCA group received significantly more morphine (mean 36.1 (SD 22.4) v 23.6 (13.1) mg; mean difference 12.3 (95% confidence interval 7.2 to 17.4) mg), spent less of the study period in moderate or severe pain (32.6% v 46.9%; mean difference 14.5% (5.6% to 23.5%)), and were more likely to be perfectly or very satisfied with the management of their pain (83% (73/88) v 66% (57/87); adjusted odds ratio 2.56 (1.25 to 5.23)) in comparison with participants in the treatment as usual group.CONCLUSIONS: Significant reductions in pain can be achieved by PCA compared with treatment as usual in patients presenting to the emergency department with non-traumatic abdominal pain. Trial registration European Clinical Trials Database EudraCT2011-000194-31; Current Controlled Trials ISRCTN25343280

The relationship of primary health care use with persistence of insomnia: a prospective cohort study

<p>Abstract</p> <p>Background</p> <p>Prevalence of insomnia symptoms in the general population is high. Insomnia is linked with high health care use and within primary care there are a number of treatment options available. The objective of this study was to determine the association of persistence and remission of insomnia with primary health care using a longitudinal study.</p> <p>Methods</p> <p>A postal survey of registered adult (over 18 years) populations of five UK general practices, repeated after 1 year, linked to primary care records. Baseline survey responders were assessed for persistence of insomnia symptoms at 12 months. The association of primary care consultation or prescription for any mood disorder (defined as anxiety, depression, stress, neurosis, or insomnia) in the 12 months between baseline and follow-up surveys with persistence of insomnia was determined.</p> <p>Results</p> <p>474 participants reporting insomnia symptoms at baseline were followed up at 12 months. 131(28%) consulted for mood problem(s) or received a relevant prescription. Of these 100 (76%) still had insomnia symptoms at one year, compared with 227 (66%) of those with no contact with primary care for this condition (OR 1.37; 95% CI 0.83, 2.27). Prescription of hypnotics showed some evidence of association with persistence of insomnia at follow-up (OR 3.18; 95% CI 0.93, 10.92).</p> <p>Conclusion</p> <p>Insomniacs continue to have problems regardless of whether or not they have consulted their primary care clinician or received a prescription for medication over the year. Hypnotics may be associated with persistence of insomnia. Further research is needed to determine more effective methods of identifying and managing insomnia in primary care. There may however be a group who have unmet need such as depression who would benefit from seeking primary health care.</p

Genes and Gene Ontologies Common to Airflow Obstruction and Emphysema in the Lungs of Patients with COPD

Chronic obstructive pulmonary disease (COPD) is a major public health problem with increasing prevalence worldwide. The primary aim of this study was to identify genes and gene ontologies associated with COPD severity. Gene expression profiling was performed on total RNA extracted from lung tissue of 18 former smokers with COPD. Class comparison analysis on mild (n = 9, FEV1 80–110% predicted) and moderate (n = 9, FEV1 50–60% predicted) COPD patients identified 46 differentially expressed genes (p<0.01), of which 14 genes were technically confirmed by quantitative real-time-PCR. Biological replication in an independent test set of 58 lung samples confirmed the altered expression of ten genes with increasing COPD severity, with eight of these genes (NNMT, THBS1, HLA-DPB1, IGHD, ETS2, ELF1, PTGDS and CYRBD1) being differentially expressed by greater than 1.8 fold between mild and moderate COPD, identifying these as candidate determinants of COPD severity. These genes belonged to ontologies potentially implicated in COPD including angiogenesis, cell migration, proliferation and apoptosis. Our secondary aim was to identify gene ontologies common to airway obstruction, indicated by impaired FEV1 and KCO. Using gene ontology enrichment analysis we have identified relevant biological and molecular processes including regulation of cell-matrix adhesion, leukocyte activation, cell and substrate adhesion, cell adhesion, angiogenesis, cell activation that are enriched among genes involved in airflow obstruction. Exploring the functional significance of these genes and their gene ontologies will provide clues to molecular changes involved in severity of COPD, which could be developed as targets for therapy or biomarkers for early diagnosis

Identification of stromally expressed molecules in the prostate by tag-profiling of cancer-associated fibroblasts, normal fibroblasts and fetal prostate.

The stromal microenvironment has key roles in prostate development and cancer, and cancer-associated fibroblasts (CAFs) stimulate tumourigenesis via several mechanisms including the expression of pro-tumourigenic factors. Mesenchyme (embryonic stroma) controls prostate organogenesis, and in some circumstances can re-differentiate prostate tumours. We have applied next-generation Tag profiling to fetal human prostate, normal human prostate fibroblasts (NPFs) and CAFs to identify molecules expressed in prostatic stroma. Comparison of gene expression profiles of a patient-matched pair of NPFs vs CAFs identified 671 transcripts that were enriched in CAFs and 356 transcripts whose levels were decreased, relative to NPFs. Gene ontology analysis revealed that CAF-enriched transcripts were associated with prostate morphogenesis and CAF-depleted transcripts were associated with cell cycle. We selected mRNAs to follow-up by comparison of our data sets with published prostate cancer fibroblast microarray profiles as well as by focusing on transcripts encoding secreted and peripheral membrane proteins, as well as mesenchymal transcripts identified in a previous study from our group. We confirmed differential transcript expression between CAFs and NPFs using QrtPCR, and defined protein localization using immunohistochemistry in fetal prostate, adult prostate and prostate cancer. We demonstrated that ASPN, CAV1, CFH, CTSK, DCN, FBLN1, FHL1, FN, NKTR, OGN, PARVA, S100A6, SPARC, STC1 and ZEB1 proteins showed specific and varied expression patterns in fetal human prostate and in prostate cancer. Colocalization studies suggested that some stromally expressed molecules were also expressed in subsets of tumour epithelia, indicating that they may be novel markers of EMT. Additionally, two molecules (ASPN and STC1) marked overlapping and distinct subregions of stroma associated with tumour epithelia and may represent new CAF markers

Multivariable risk prediction can greatly enhance the statistical power of clinical trial subgroup analysis

BACKGROUND: When subgroup analyses of a positive clinical trial are unrevealing, such findings are commonly used to argue that the treatment's benefits apply to the entire study population; however, such analyses are often limited by poor statistical power. Multivariable risk-stratified analysis has been proposed as an important advance in investigating heterogeneity in treatment benefits, yet no one has conducted a systematic statistical examination of circumstances influencing the relative merits of this approach vs. conventional subgroup analysis. METHODS: Using simulated clinical trials in which the probability of outcomes in individual patients was stochastically determined by the presence of risk factors and the effects of treatment, we examined the relative merits of a conventional vs. a "risk-stratified" subgroup analysis under a variety of circumstances in which there is a small amount of uniformly distributed treatment-related harm. The statistical power to detect treatment-effect heterogeneity was calculated for risk-stratified and conventional subgroup analysis while varying: 1) the number, prevalence and odds ratios of individual risk factors for risk in the absence of treatment, 2) the predictiveness of the multivariable risk model (including the accuracy of its weights), 3) the degree of treatment-related harm, and 5) the average untreated risk of the study population. RESULTS: Conventional subgroup analysis (in which single patient attributes are evaluated "one-at-a-time") had at best moderate statistical power (30% to 45%) to detect variation in a treatment's net relative risk reduction resulting from treatment-related harm, even under optimal circumstances (overall statistical power of the study was good and treatment-effect heterogeneity was evaluated across a major risk factor [OR = 3]). In some instances a multi-variable risk-stratified approach also had low to moderate statistical power (especially when the multivariable risk prediction tool had low discrimination). However, a multivariable risk-stratified approach can have excellent statistical power to detect heterogeneity in net treatment benefit under a wide variety of circumstances, instances under which conventional subgroup analysis has poor statistical power. CONCLUSION: These results suggest that under many likely scenarios, a multivariable risk-stratified approach will have substantially greater statistical power than conventional subgroup analysis for detecting heterogeneity in treatment benefits and safety related to previously unidentified treatment-related harm. Subgroup analyses must always be well-justified and interpreted with care, and conventional subgroup analyses can be useful under some circumstances; however, clinical trial reporting should include a multivariable risk-stratified analysis when an adequate externally-developed risk prediction tool is available

Crystal Structure of PrgI-SipD: Insight into a Secretion Competent State of the Type Three Secretion System Needle Tip and its Interaction with Host Ligands

Many infectious Gram-negative bacteria, including Salmonella typhimurium, require a Type Three Secretion System (T3SS) to translocate virulence factors into host cells. The T3SS consists of a membrane protein complex and an extracellular needle together that form a continuous channel. Regulated secretion of virulence factors requires the presence of SipD at the T3SS needle tip in S. typhimurium. Here we report three-dimensional structures of individual SipD, SipD in fusion with the needle subunit PrgI, and of SipD:PrgI in complex with the bile salt, deoxycholate. Assembly of the complex involves major conformational changes in both SipD and PrgI. This rearrangement is mediated via a π bulge in the central SipD helix and is stabilized by conserved amino acids that may allow for specificity in the assembly and composition of the tip proteins. Five copies each of the needle subunit PrgI and SipD form the T3SS needle tip complex. Using surface plasmon resonance spectroscopy and crystal structure analysis we found that the T3SS needle tip complex binds deoxycholate with micromolar affinity via a cleft formed at the SipD:PrgI interface. In the structure-based three-dimensional model of the T3SS needle tip, the bound deoxycholate faces the host membrane. Recently, binding of SipD with bile salts present in the gut was shown to impede bacterial infection. Binding of bile salts to the SipD:PrgI interface in this particular arrangement may thus inhibit the T3SS function. The structures presented in this study provide insight into the open state of the T3SS needle tip. Our findings present the atomic details of the T3SS arrangement occurring at the pathogen-host interface

Fitness Consequences of Advanced Ancestral Age over Three Generations in Humans

A rapid rise in age at parenthood in contemporary societies has increased interest in reports of higher prevalence of de novo mutations and health problems in individuals with older fathers, but the fitness consequences of such age effects over several generations remain untested. Here, we use extensive pedigree data on seven pre-industrial Finnish populations to show how the ages of ancestors for up to three generations are associated with fitness traits. Individuals whose fathers, grandfathers and great-grandfathers fathered their lineage on average under age 30 were ~13% more likely to survive to adulthood than those whose ancestors fathered their lineage at over 40 years. In addition, females had a lower probability of marriage if their male ancestors were older. These findings are consistent with an increase of the number of accumulated de novo mutations with male age, suggesting that deleterious mutations acquired from recent ancestors may be a substantial burden to fitness in humans. However, possible non-mutational explanations for the observed associations are also discussed

Bleeding disorders in the tribe: result of consanguineous in breeding

<p>Abstract</p> <p>Objective</p> <p>To determine the frequency and clinical features of bleeding disorders in the tribe as a result of consanguineous marriages.</p> <p>Design</p> <p>Cross Sectional Study</p> <p>Introduction</p> <p>Countries in which consanguinity is a normal practice, these rare autosomal recessive disorders run in close families and tribes. Here we describe a family, living in village Ali Murad Chandio, District Badin, labeled as haemophilia.</p> <p>Patients & Methods</p> <p>Our team visited the village & developed the pedigree of the whole extended family, up to seven generations. Performa was filled by incorporating patients, family history of bleeding, signs & symptoms, and bleeding from any site. From them 144 individuals were screened with CBC, bleeding time, platelet aggregation studies & RiCoF. While for PT, APTT, VWF assay and Factor VIII assay, samples were kept frozen at -70 degrees C until tested.</p> <p>Results</p> <p>The family tree of the seven generations comprises of 533 individuals, 63 subjects died over a period of 20 years and 470 were alive. Out of all those 144 subjects were selected on the basis of the bleeding history. Among them 98(68.1%) were diagnosed to have a bleeding disorder; 44.9% patients were male and 55.1% patients were female. Median age of all the patients was 20.81, range (4 months- 80 yrs). The results of bleeding have shown that majority had gum bleeding, epistaxis and menorrhagia. Most common bleeding disorder was Von Willebrand disease and Platelet functional disorders.</p> <p>Conclusion</p> <p>Consanguineous marriages keep all the beneficial and adversely affecting recessive genes within the family; in homozygous states. These genes express themselves and result in life threatening diseases. Awareness, education & genetic counseling will be needed to prevent the spread of such common occurrence of these bleeding disorders in the community.</p

Coronary–aortic interaction during ventricular isovolumic contraction

In earlier work, we suggested that the start of the isovolumic contraction period could be detected in arterial pressure waveforms as the start of a temporary pre-systolic pressure perturbation (AICstart, start of the Arterially detected Isovolumic Contraction), and proposed the retrograde coronary blood volume flow in combination with a backwards traveling pressure wave as its most likely origin. In this study, we tested this hypothesis by means of a coronary artery occlusion protocol. In six Yorkshire × Landrace swine, we simultaneously occluded the left anterior descending (LAD) and left circumflex (LCx) artery for 5 s followed by a 20-s reperfusion period and repeated this sequence at least two more times. A similar procedure was used to occlude only the right coronary artery (RCA) and finally all three main coronary arteries simultaneously. None of the occlusion protocols caused a decrease in the arterial pressure perturbation in the aorta during occlusion (P > 0.20) nor an increase during reactive hyperemia (P > 0.22), despite a higher deceleration of coronary blood volume flow (P = 0.03) or increased coronary conductance (P = 0.04) during hyperemia. These results show that the pre-systolic aortic pressure perturbation does not originate from the coronary arteries