Functional interaction of a novel 15.5kD [U4/U6.U5] tri-snRNP protein with the 5' stem-loop of U4 snRNA.
Activation of the spliceosome for splicing catalysis requires the dissociation of U4 snRNA from the U4/U6 snRNA duplex prior to the first step of splicing. We characterize an evolutionarily conserved 15.5 kDa protein of the HeLa [U4/U6.U5] tri-snRNP that binds directly to the 5' stem-loop of U4 snRNA. This protein shares a novel RNA recognition motif with several RNP-associated proteins, which is essential, but not sufficient for RNA binding. The 15.5kD protein binding site on the U4 snRNA consists of an internal purine-rich loop flanked by the stem of the 5' stem-loop and a stem comprising two base pairs. Addition of an RNA oligonucleotide comprising the 5' stem-loop of U4 snRNA (U4SL) to an in vitro splicing reaction blocked the first step of pre-mRNA splicing. Interestingly, spliceosomal C complex formation was inhibited while B complexes accumulated. This indicates that the 15.5kD protein, and/or additional U4 snRNP proteins associated with it, play an important role in the late stage of spliceosome assembly, prior to step I of splicing catalysis. Our finding that the 15.5kD protein also efficiently binds to the 5' stem-loop of U4atac snRNA indicates that it may be shared by the [U4atac/U6atac.U5] tri-snRNP of the minor U12-type spliceosome.


Predictive value of S100-B and copeptin for outcomes following seizure: the BISTRO International Cohort Study.
To evaluate the performance of S100-B protein and copeptin, in addition to clinical variables, in predicting outcomes of patients attending the emergency department (ED) following a seizure. We prospectively included adult patients presented with an acute seizure, in four EDs in France and the United Kingdom. Participants were followed up for 28 days. The primary endpoint was a composite of seizure recurrence, all-cause mortality, hospitalization or rehospitalisation, or return visit in the ED within seven days. Among the 389 participants included in the analysis, 156 (40%) experienced the primary endpoint within seven days and 195 (54%) at 28 days. Mean levels of both S100-B (0.11 μg/l [95% CI 0.07-0.20] vs 0.09 μg/l [0.07-0.14]) and copeptin (23 pmol/l [9-104] vs 17 pmol/l [8-43]) were higher in participants meeting the primary endpoint. However, both biomarkers were poorly predictive of the primary outcome with a respective area under the receiving operator characteristic curve of 0.57 [0.51-0.64] and 0.59 [0.54-0.64]. Multivariable logistic regression analysis identified higher age (odds ratio [OR] 1.3 per decade [1.1-1.5]), provoked seizure (OR 4.93 [2.5-9.8]), complex partial seizure (OR 4.09 [1.8-9.1]) and first seizure (OR 1.83 [1.1-3.0]) as independent predictors of the primary outcome. A second regression analysis including the biomarkers showed no additional predictive benefit (S100-B OR 3.89 [0.80-18.9] copeptin OR 1 [1.00-1.00]). The plasma biomarkers S100-B and copeptin did not improve prediction of poor outcome following seizure. Higher age, a first seizure, a provoked seizure and a partial complex seizure are independently associated with adverse outcomes.


Perfectly conducting diffraction grating formalisms extended to good conductors via the surface impedance boundary condition.
The surface impedance boundary condition is used to include the effect of high conductivity of meals in the integral theory of perfectly conducting gratings. As an intuitive approach, the diffraction formalism proposed by Petit for the treatment of infinitely conducting gratings in P polarization is extended to highly conducting materials by introducing the concept of equivalent surface current density. Then, integral equations for both polarizations are deduced in a mathematically rigorous way. The new method is used to calculate the efficiencies of sinusoidal gratings at infrared and visible light, and the numerical results are compared with those obtained using Maxwell boundary conditions and also with the perfect conductivity model.


Using high frequency Raman spectra for colonic neoplasia detection.
Raman systems have tremendous potential as adjunct devices for endoscopes to improve the identification of early colon cancers. However, the traditional low frequency (LF) measurement range has several obstacles that make it challenging to develop a routine clinical tool. An alternative is to use high frequency (HF) range. To test this idea Raman spectra were obtained in both the LF and HF ranges from the same colon lesions. Multivariate analyses predicted the pathology with high sensitivity and specificity for both the LF and HF data sets. This suggests that Raman systems that measure HF spectra, and are simpler to adopt into the clinic, could be used in vivo to improve the identification of neoplastic lesions.


Decision from Models: Generalizing Probability Information to Novel Tasks.
We investigate a new type of decision under risk where-to succeed-participants must generalize their experience in one set of tasks to a novel set of tasks. We asked participants to trade distance for reward in a virtual minefield where each successive step incurred the same fixed probability of failure (referred to as hazard). With constant hazard, the probability of success (the survival function) decreases exponentially with path length. On each trial, participants chose between a shorter path with smaller reward and a longer (more dangerous) path with larger reward. They received feedback in 160 training trials: encountering a mine along their chosen path resulted in zero reward and successful completion of the path led to the reward associated with the path chosen. They then completed 600 no-feedback test trials with novel combinations of path length and rewards. To maximize expected gain, participants had to learn the correct exponential model in training and generalize it to the test conditions. We compared how participants discounted reward with increasing path length to the predictions of nine choice models including the correct exponential model. The choices of a majority of the participants were best accounted for by a model of the correct exponential form although with marked overestimation of the hazard rate. The decision-from-models paradigm differs from experience-based decision paradigms such as decision-from-sampling in the importance assigned to generalizing experience-based information to novel tasks. The task itself is representative of everyday tasks involving repeated decisions in stochastically invariant environments.


Acute psychotic episode in a patient with thyrotoxicosis factitia.
Psychotic syndromes in the context of hyperthyroidism are seldom mentioned in medical textbooks and only a few cases have been published. Typically, such cases present as an affective psychosis. Schizophrenia-like psychosis is a rare occurrence in hyperthyroidism and the link between these two conditions is still poorly understood.We report the case of a female patient with a known history of chronic lymphocytic thyroiditis. The patient presented to our emergency department with an acute schizophrenia-like psychosis. Elevated levels of T4 and free T4 were found. These resulted from the patient's voluntary intake of excess levothyroxine as an attempt to lose weight (thyrotoxicosis factitia). Normalisation of thyroid hormone levels and antipsychotic treatment led to prompt remission of the psychosis. Even though the patient stopped the antipsychotics, she remained free of symptoms during the follow-up. Similar cases are briefly reviewed and some of the data from basic research is also considered.


Modeling the Steady-State Effects of Mean Arterial Pressure on the Kidneys.
Goal: We describe the relationship between mean arterial pressure (MAP) and glomerular filtration rate (GFR) since therapies affecting MAP can have large effects on kidney function. Methods: We developed a closed-loop, steady-state mechanistic model of the human kidney with a reduced parameter set estimated from measurements. Results: The model was first validated against literature models. Further, GFR was validated against intensive care patient data (root mean squared error (RMSE) 13.5 mL/min) and against hypertensive patients receiving sodium nitroprusside (SNP) (RMSE less than 5 mL/min). A sensitivity analysis of the model reinforced the fact that vascular resistance is inversely related to GFR and showed that changes to either vascular resistance or renal autoregulation cause a significant change in sodium concentration in the descending limb of Henle. Conclusions: This model can be used to determine the impact of MAP on GFR and overall kidney health. The modeling framework lends itself to personalization of the model to a specific human.


Drug-resistant AML cells and primary AML specimens are killed by 111In-anti-CD33 monoclonal antibodies modified with nuclear localizing peptide sequences.
Multidrug resistance (MDR) is a major challenge to the successful treatment of acute myeloid leukemia (AML). Our purpose was to determine whether (111)In-HuM195 anti-CD33 antibodies modified with peptides harboring nuclear localizing sequences (NLS) could kill drug-resistant AML cell lines and primary AML patient specimens expressing MDR transporters through the emission of Auger electrons. HuM195, M195, and irrelevant mouse IgG (mIgG) were conjugated to 10+/-3 NLS peptides and then labeled with (111)In by diethylenetriaminepentaacetic acid substitution to a specific activity of 1-8 MBq/microg. The binding affinity of HuM195 and M195 was determined for HL-60 and mitoxantrone-resistant HL-60-MX-1 cells. Nuclear localization of (111)In-NLS-HuM195, (111)In-NLS-M195, (111)In-HuM195, and (111)In-M195 was measured by subcellular fractionation. The antiproliferative effects of (111)In-NLS-HuM195, (111)In-NLS-M195, (111)In-HuM195, and (111)In-M195 (2.5-250 kBq/well) on HL-60 and HL-60-MX-1 were studied using the WST-1 assay. Clonogenic survival of HL-60 and HL-60-MX-1 leukemic cells and 10 primary AML specimens with MDR phenotype (assessed by flow cytometry) was determined after exposure for 3 h at 37 degrees C to 2.5-250 mBq/cell of (111)In-NLS-HuM195, (111)In-HuM195, or (111)In-NLS-mIgG. Clonogenic survival versus the amount of radioactivity incubated with the cells (mBq/cell) was plotted, and the mean lethal amount of radioactivity and the lower asymptote of the curve (plateau) were determined. The (111)In-labeled anti-CD33 monoclonal antibodies HuM195 and M195 modified with NLS were efficiently routed to the nucleus of HL-60 cells and their mitoxantrone-resistant clone after CD33-mediated internalization. The following are the principal findings of our study: (111)In-NLS-HuM195 was more effective at killing HL-60 and HL-60-MX-1 cells than was (111)In-HuM195, a strong correlation between the specific activity of the (111)In-labeled radioimmunoconjugates and their cytotoxicity toward AML cells existed, and leukemic cells from patients were killed by (111)In-NLS-M195 or (111)In-M195, but the cytotoxic response among specimens was heterogeneous. NLS conjugation enhanced the nuclear uptake and cytotoxicity of (111)In-HuM195 and (111)In-M195 toward drug-resistant AML cell lines as well as patient specimens expressing a diversity of MDR phenotypes, including Pgp-170, BCRP1, or MRP1 transporters. Targeted Auger electron radioimmunotherapy using (111)In-labeled anti-CD33 monoclonal antibodies modified with NLS may be able to overcome MDR and provide a means of treating chemotherapy-resistant myeloid leukemias in patients.


National trends in the treatment of penile prosthesis infections by explantation alone vs. immediate salvage and reimplantation.
A penile prosthesis infection (PPI) is either treated with explantation of the prosthesis with a possible delayed reimplantation or a salvage procedure with an immediate reimplantation of the prosthesis. We used a large, all-payer national database to investigate the use of the salvage procedure in the setting of PPI. The study used years 2000-2009 of the Nationwide Inpatient Sample to identify PPIs treated with immediate salvage or explantation alone. Admissions were then stratified by various parameters to compare differences in the salvage rates. Salvage Rate of Penile Prosthesis infection. A total of 1,557 patients were treated with an explantation only (82.7%) or salvage (17.3%) for PPI, a proportion that remained stable over the study period. The patients treated with salvage were younger (60.4 vs. 65.1 years), more likely to be discharged home (87.3% vs. 61.9%), and were less likely to have a severe presentation (7.2% vs. 31.6%) than those who were explanted only (P < 0.001). These factors were confirmed on multivariate regression analysis. The regression also revealed that treatment at rural hospitals had lower odds of salvage than treatment at urban teaching hospitals. Race, comorbid diabetes, and insurance status did not independently affect the salvage rate. There was no significant difference in total hospital charges between groups. Salvage rates have remained low over the past decade. Our study elucidated several factors decreasing the chances of salvage after PPI including age, severity of presentation, and hospital setting.


Testicular embryonal carcinoma presenting as chest wall subcutaneous mass.
Embryonal carcinoma affects young males in the prime of life with a majority of these tumors already having metastasis at the time of diagnosis. Subcutaneous metastasis from embryonal carcinoma are very rare and often associated with wide spread disease and poor prognosis. We report a case of chest wall subcutaneous metastasis of embryonal carcinoma in a 27 year-old man that was the first presentation of the disease and was diagnosed on fine needle aspiration cytology (FNAC). Subsequent search led to the discovery of the primary in the testis. The cytomorphological features of embryonal carcinoma are quite distinctive and FNAC plays a vital role in early diagnosis. The criteria for diagnosis includes presence of cellular smears exhibiting disperse cells as well as cell aggregates forming microglandular patterns. Cells have large nuclei with prominent nucleoli and pale indistinct often vacuolated cytoplasm. Early diagnosis and treatment with platinum based chemotherapy in conjunction with radiotherapy and surgery have high cure rate.


Illness coping strategies and hypochondriacal traits among medical inpatients.
The Illness Coping Strategies scale (ICS) is an 18-Likert-item scale developed to examine illness appraisal and coping by medical patients more comprehensively than instruments which screen for hypochondriacal traits in this population. This study has examined the association of hypochondriacal traits with illness coping strategies addressed by the ICS among 101 randomly selected inpatients drawn from a general medicine unit of a teaching general hospital. Despite the exclusion of patients with substance abuse or organic mental disorder, or referral for psychiatric consultations, hypochondriacal traits were prevalent in this inpatient sample. Five factor-derived subscales of the ICS (disease vigilance, limit activity, overresponsiveness, self-treatment and obsessive worry) were extracted, and internal consistency and test-retest reliabilities were determined. Multiple regression analysis on a composite measure of hypochondriasis revealed that the five ICS factor scales accounted for 26 percent of the variance. Patients with a high hypochondriacal index had associated high scores on symptom vigilance, limit activity and obsessive worry subscales, suggesting that these illness coping strategies may represent an important aspect of hypochondriacal illness presentation. The self-treatment subscale appeared to act as a suppressor variable, contributing to the prediction of the hypochondriacal index by extracting self treatment aspects from the other illness coping factors. Illness coping responses found to be associated with hypochondriacal traits in this study were rather passive strategies which promoted increased vigilance and concern about symptoms while providing few opportunities to reduce uncertainty regarding health status.


Sex-based differences in antiretroviral therapy initiation, switching and treatment interruptions: global overview from the International Epidemiologic Databases to Evaluate AIDS (IeDEA).
In 2015, the World Health Organization recommended that all HIV-infected individuals consider ART initiation as soon as possible after diagnosis. Sex differences in choice of initial ART regimen, indications for switching, time to switching and choice of second-line regimens have not been well described. The aims of this study were to describe first-line ART and CD4 count at ART initiation by sex, calendar year and region, and to analyse time to change or interruption in first-line ART, according to sex in each region. Participating cohorts included: Southern, East and West Africa (IeDEA-Africa), North America (NA-ACCORD), Caribbean, Central/South America (CCASAnet) and Asia-Pacific including Australia (IeDEA Asia-Pacific). The primary outcomes analysed for each region and according to sex were choice of initial ART, time to switching and time to discontinuation of the first-line regimen. The combined cohort data set comprised of 715,252 participants across seven regions from low- to high-income settings. The median CD4 count at treatment initiation was lower in men compared with women in nearly all regions and time periods. Women from North America and Southern Africa were more likely to switch ART compared to men (p < 0.001) with approximately 90% of women reporting a major change after 10 years in North America. Overall, after 8 years on ART, >50% of HIV- positive men and women from Southern Africa, East Africa, South and Central America remained on their original regimen. Men were more likely to have a treatment interruption compared with women in low- and middle-income countries from the Asia/Pacific region (p < 0.001) as were men from Southern Africa (p < 0.001). Greater than 75% of men and women did not report a treatment interruption after 10 years on ART from all regions except North America and Southern Africa. There are regional variations in the ART regimen commenced at baseline and rates of major change and treatment interruption according to sex. Some of this is likely to reflect changes in local and international antiretroviral guideline recommendations but other sex-specific factors such as pregnancy may contribute to these differences.


Integrin Mechano-chemical Signaling Generates Plasma Membrane Nanodomains that Promote Cell Spreading.
Glycosylphosphatidylinositol-anchored proteins (GPI-APs) are a major class of lipid-anchored plasma membrane proteins. GPI-APs form nanoclusters generated by cortical acto-myosin activity. While our understanding of the physical principles governing this process is emerging, the molecular machinery and functional relevance of GPI-AP nanoclustering are unknown. Here, we first show that a membrane receptor signaling pathway directs nanocluster formation. Arg-Gly-Asp motif-containing ligands bound to the β1-integrin receptor activate src and focal adhesion kinases, resulting in RhoA signaling. This cascade triggers actin-nucleation via specific formins, which, along with myosin activity, drive the nanoclustering of membrane proteins with actin-binding domains. Concurrently, talin-mediated activation of the mechano-transducer vinculin is required for the coupling of the acto-myosin machinery to inner-leaflet lipids, thereby generating GPI-AP nanoclusters. Second, we show that these nanoclusters are functional; disruption of their formation either in GPI-anchor remodeling mutants or in vinculin mutants impairs cell spreading and migration, hallmarks of integrin function.


Post mortem DNA degradation of human tissue experimentally mummified in salt.
Mummified human tissues are of great interest in forensics and biomolecular archaeology. The aim of this study was to analyse post mortem DNA alterations in soft tissues in order to improve our knowledge of the patterns of DNA degradation that occur during salt mummification. In this study, the lower limb of a female human donor was amputated within 24 h post mortem and mummified using a process designed to simulate the salt dehydration phase of natural or artificial mummification. Skin and skeletal muscle were sampled at multiple time points over a period of 322 days and subjected to genetic analysis. Patterns of genomic fragmentation, miscoding lesions, and overall DNA degradation in both nuclear and mitochondrial DNA was assessed by different methods: gel electrophoresis, multiplex comparative autosomal STR length amplification, cloning and sequence analysis, and PCR amplification of different fragment sizes using a damage sensitive recombinant polymerase. The study outcome reveals a very good level of DNA preservation in salt mummified tissues over the course of the experiment, with an overall slower rate of DNA fragmentation in skin compared to muscle.


Co-occupancy of two Pumilio molecules on a single hunchback NRE.
Pumilio controls a number of processes in eukaryotes, including the translational repression of hunchback (hb) mRNA in early Drosophila embryos. The Pumilio Puf domain binds to a pair of 32 nucleotide (nt) Nanos response elements (NRE1 and NRE2) within the 3' untranslated region of hb mRNA. Despite the elucidation of structures of human Pumilio Puf domain in complex with hb RNA elements, the nature of hb mRNA recognition remains unclear. In particular, the site that mediates regulation in vivo is significantly larger than the 8-10-nt RNA elements bound to single Puf molecules in crystal structures. Here we present biophysical and biochemical data that partially resolve the paradox. We show that each NRE is composed of two binding sites (Box A and Box B) and that two Puf domains can co-occupy a single NRE. The Puf domains have a higher affinity for the 3' Box B site than the 5' Box A site; binding to the intact NRE appears to be cooperative (at least in some experiments). We suggest that the 2 Pumilio:1 NRE complex is the functional regulatory unit in vivo.


Apnea after immunization of preterm infants.
To determine the frequency of adverse reactions, particularly the occurrence of apnea, among preterm infants after immunization with diphtheria and tetanus toxoids and whole cell pertussis vaccine adsorbed (DTP) and Haemophilus influenzae type b conjugate (HibC) vaccine in the neonatal intensive care unit. After the occurrence of apnea in two preterm infants following immunization with DTP and HibC, a prospective surveillance of 97 preterm infants younger than 37 weeks of gestation who were immunized with DTP (94 also received HibC at the same time) in the neonatal intensive care unit was performed to assess the frequency of adverse reactions and in particular, the occurrence of apnea. For each infant, data were recorded for a 3-day period before and after receipt of the immunization. The majority of preterm infants tolerated immunizations with DTP and HibC without ill effects. However, 12 (12%) infants experienced a recurrence of apnea, and 11 (11%) had at least a 50% increase in the number of apneic and bradycardic episodes in the 72 hours after immunization. This occurred primarily among smaller preterm infants who were immunized at a lower weight (p = 0.01), had experienced more severe apnea of prematurity (p = 0.01), and had chronic lung disease (p = 0.03). The temporal association observed between immunization of preterm infants and a transient increase or recurrence of apnea after vaccination merits further study. Cardiorespiratory monitoring of these infants after immunization may be advisable.


Genetic hypothesis of idiopathic schizophrenia: its exorphin connection.
This brief overview proposes a testable oligogenic model of the inheritance of susceptibility to idiopathic schizophrenia: "abnormal" genes at each of a few complementary loci. The model is based on my assumptions as to the likely genetic abnormalities at possibly four or five interacting loci that would permit exorphins, the opioid peptides from some food proteins, especially glutens and possibly caseins, to go from gut to brain and cause symptoms of schizophrenia. Exorphins may reach the brain cerebrospinal fluid (CSF) in harmful amounts because of their genetically increased, receptor-mediated transcellular passage across the gut epithelial barrier plus decreased catabolism by genetically defective enzymes. A schizophrenia-specific, genetically enhanced affinity for exorphins by opioid receptors influencing dopaminergic and other neurons would permit sustained dysfunction at low CSF exorphin concentrations. Tests of each postulated genetic abnormality are suggested. This model is supported by a variety of evidence, including a significant effect of gluten or its absence on relapsed schizophrenic patients, the high correlation of changes in first admission rates for schizophrenia with changes in grain consumption rates, and the rarity of cases of schizophrenia where grains and milk are rare.


Bridging developmental boundaries: lifelong dietary patterns modulate life histories in a parthenogenetic insect.
Determining the effects of lifelong intake patterns on performance is challenging for many species, primarily because of methodological constraints. Here, we used a parthenogenetic insect (Carausius morosus) to determine the effects of limited and unlimited food availability across multiple life-history stages. Using a parthenogen allowed us to quantify intake by juvenile and adult females and to evaluate the morphological, physiological, and life-history responses to intake, all without the confounding influences of pair-housing, mating, and male behavior. In our study, growth rate prior to reproductive maturity was positively correlated with both adult and reproductive lifespans but negatively correlated with total lifespan. Food limitation had opposing effects on lifespan depending on when it was imposed, as it protracted development in juveniles but hastened death in adults. Food limitation also constrained reproduction regardless of when food was limited, although decreased fecundity was especially pronounced in individuals that were food-limited as late juveniles and adults. Additional carry-over effects of juvenile food limitation included smaller adult size and decreased body condition at the adult molt, but these effects were largely mitigated in insects that were switched to ad libitum feeding as late juveniles. Our data provide little support for the existence of a trade-off between longevity and fecundity, perhaps because these functions were fueled by different nutrient pools. However, insects that experienced a switch to the limited diet at reproductive maturity seem to have fueled egg production by drawing down body stores, thus providing some evidence for a life-history trade-off. Our results provide important insights into the effects of food limitation and indicate that performance is modulated by intake both within and across life-history stages.


Methods used for suicide by farmers in England and Wales. The contribution of availability and its relevance to prevention.
Farmers in the UK have an elevated risk of suicide. It has been suggested that this may be related to their ease of access to dangerous means for suicidal behaviour. The extent to which farmers use these means and changes in their use may be relevant to suicide prevention. Data on 719 deaths in farmers of both genders in England and Wales between 1981 and 1993 in which a verdict of suicide or undetermined cause (open verdict) was recorded were analysed. Of 702 deaths in male farmers, firearms were involved in 40.0%, hanging in 29.6%, carbon monoxide in 16.4%, poisoning in 8.0% (over half of which involved agricultural or horticultural poisons) and other methods in 6.1%. There was a considerable excess of deaths due to firearms compared with the distribution of methods of suicide and open verdict deaths in males in the general population. Hanging was also somewhat more frequent. During the study period there was a reduction in firearm death rates, particularly after 1989 when there was national legislation on firearm ownership, registration and storage. There were also fewer farming suicides after this date. By the end of the study period hanging was more frequent than deaths involving firearms. Farmers who commit suicide tend to use methods to which they have easy access. Restriction of the ready availability of such methods, particularly in farmers known to be depressed or otherwise at risk, might prevent some suicides.


Atomic resolution structure of a protein prepared by non-enzymatic His-tag removal. Crystallographic and NMR study of GmSPI-2 inhibitor.
Purification of suitable quantity of homogenous protein is very often the bottleneck in protein structural studies. Overexpression of a desired gene and attachment of enzymatically cleavable affinity tags to the protein of interest made a breakthrough in this field. Here we describe the structure of Galleria mellonella silk proteinase inhibitor 2 (GmSPI-2) determined both by X-ray diffraction and NMR spectroscopy methods. GmSPI-2 was purified using a new method consisting in non-enzymatic His-tag removal based on a highly specific peptide bond cleavage reaction assisted by Ni(II) ions. The X-ray crystal structure of GmSPI-2 was refined against diffraction data extending to 0.98 Å resolution measured at 100 K using synchrotron radiation. Anisotropic refinement with the removal of stereochemical restraints for the well-ordered parts of the structure converged with R factor of 10.57% and Rfree of 12.91%. The 3D structure of GmSPI-2 protein in solution was solved on the basis of 503 distance constraints, 10 hydrogen bonds and 26 torsion angle restraints. It exhibits good geometry and side-chain packing parameters. The models of the protein structure obtained by X-ray diffraction and NMR spectroscopy are very similar to each other and reveal the same β2αβ fold characteristic for Kazal-family serine proteinase inhibitors.


New viral vectors for infectious diseases and cancer.
Since the discovery in 1796 by Edward Jenner of vaccinia virus as a way to prevent and finally eradicate smallpox, the concept of using a virus to fight another virus has evolved into the current approaches of viral vectored genetic vaccines. In recent years, key improvements to the vaccinia virus leading to a safer version (Modified Vaccinia Ankara, MVA) and the discovery that some viruses can be used as carriers of heterologous genes encoding for pathological antigens of other infectious agents (the concept of 'viral vectors') has spurred a new wave of clinical research potentially providing for a solution for the long sought after vaccines against major diseases such as HIV, TB, RSV and Malaria, or emerging infectious diseases including those caused by filoviruses and coronaviruses. The unique ability of some of these viral vectors to stimulate the cellular arm of the immune response and, most importantly, T lymphocytes with cell killing activity, has also reawakened the interest toward developing therapeutic vaccines against chronic infectious diseases and cancer. To this end, existing vectors such as those based on Adenoviruses have been improved in immunogenicity and efficacy. Along the same line, new vectors that exploit viruses such as Vesicular Stomatitis Virus (VSV), Measles Virus (MV), Lymphocytic choriomeningitis virus (LCMV), cytomegalovirus (CMV), and Herpes Simplex Virus (HSV), have emerged. Furthermore, technological progress toward modifying their genome to render some of these vectors incompetent for replication has increased confidence toward their use in infant and elderly populations. Lastly, their production process being the same for every product has made viral vectored vaccines the technology of choice for rapid development of vaccines against emerging diseases and for 'personalised' cancer vaccines where there is an absolute need to reduce time to the patient from months to weeks or days. Here we review the recent developments in viral vector technologies, focusing on novel vectors based on primate derived Adenoviruses and Poxviruses, Rhabdoviruses, Paramixoviruses, Arenaviruses and Herpesviruses. We describe the rationale for, immunologic mechanisms involved in, and design of viral vectored gene vaccines under development and discuss the potential utility of these novel genetic vaccine approaches in eliciting protection against infectious diseases and cancer.


A gene fusion at a homeobox locus: alterations in leaf shape and implications for morphological evolution.
Compound leaves are seen in many angiosperm genera and are thought to be either fundamentally different from simple leaves or elaborations of simple leaves. The knotted1-like homeobox (knox) genes are known to regulate plant development. When overexpressed in homologous or heterologous species, this family of genes can cause changes in leaf morphology, including excessive leaf compounding in tomato. We describe here an instance of a spontaneously arisen fusion between a gene encoding a metabolic enzyme and a homeodomain protein. We show that the fusion results in overexpression of the homeodomain protein and a change in morphology that approximates the changes caused by overexpression of the same gene under the control of the cauliflower mosaic virus 35S promoter in transgenic plants. Exon-shuffling events can account for the modularity of proteins. If the shuffled exons are associated with altered promoters, changes in gene expression patterns can result. Our results show that gene fusions of this nature can cause changes in expression patterns that lead to altered morphology. We suggest that such phenomena may have played a role in the evolution of form.


Extraction of phase derivative data from interferometer images using a continuous wavelet transform to determine two-dimensional refractive index profiles.
Two-dimensional refractive index profiles of ion exchanged channel waveguides in glass have been obtained from the analysis of interferometer data. To obtain depth data, a shallow bevel is produced in the glass by polishing. The refractive index profile information that is contained within the derivative of the phase data is extracted directly using a continuous wavelet transform algorithm. The algorithm used to characterize and smooth the wavelet ridge is discussed in detail.


Loss of the desmosomal protein perp enhances the phenotypic effects of pemphigus vulgaris autoantibodies.
Pemphigus vulgaris (PV) is an autoimmune bullous disease in which autoantibodies against proteins of the desmosomal adhesion complex perturb desmosomal function, leading to intercellular adhesion defects in the oral mucosa and skin. Previous studies have demonstrated a central role for downregulation of the desmosomal cadherin desmoglein 3 (DSG3) in the pathogenesis of PV. However, the effects of non-cadherin desmosomal proteins in modulating the cellular manifestations of PV remain poorly understood. Here, we characterize the expression and functional importance of Perp, a newly discovered tetraspan desmosomal protein, in PV. Our data demonstrate that PV autoantibodies disrupt Perp expression at the membrane and trigger its internalization along with DSG3 into the endosomal pathway, where it is ultimately targeted to the lysosome for degradation. We further show that Perp deficiency exacerbates the pathogenic effects of PV autoantibodies on keratinocytes by enhancing both the depletion of desmosomal DSG3 and intercellular adhesion defects. Together, our findings highlight the importance of non-cadherin desmosomal proteins in modulating PV phenotypes and provide new insight into Perp's role in the desmosome.


Numerical modelling of tooth enamel subsurface lesion formation induced by dental plaque.
Using a one-dimensional mathematical model that couples tooth demineralisation and remineralisation with metabolic processes occurring in the dental plaque, two mechanisms for subsurface lesion formation were evaluated. It was found that a subsurface lesion can develop only as the result of alternating periods of demineralisation (acid attack during sugar consumption) and remineralisation (resting period) in tooth enamel with uniform mineral composition. It was also shown that a minimum plaque thickness that can induce an enamel lesion exists. The subsurface lesion formation can also be explained by assuming the existence of a fluoride-containing layer at the tooth surface that decreases enamel solubility. A nearly constant thickness of the surface layer was obtained with both proposed mechanisms. Sensitivity analysis showed that surface layer formation is strongly dependent on the length of remineralisation and demineralisation cycles. The restoration period is very important and the numerical simulations support the observation that often consumption of sugars is a key factor in caries formation. The calculated profiles of mineral content in enamel are similar to those observed experimentally. Most probably, both studied mechanisms interact in vivo in the process of caries development, but the simplest explanation for subsurface lesion formation remains the alternation between demineralisation and remineralisation cycles without any pre-imposed gradients.


Serum magnesium and calcium levels in infertile women during a cycle of reproductive assistance.
Magnesium (Mg) and calcium (Ca) are essential cations for women's preconception health. It is well known that, in blood, the concentration of ionized form of these two cations is temporally altered during menstrual cycle, suggesting a correlation between sex steroid hormones and serum calcium and magnesium levels. Evidence from literature suggests that in assisted reproductive technology increasing estrogens during ovarian hyperstimulation may also modulate serum magnesium and calcium levels. Therefore, we first examined total serum magnesium and calcium levels during follicular phase in a large population of infertile patients who underwent intrauterine insemination (IUI). The results were compared to a group of fertile women. Successively, we studied the total serum magnesium and calcium concentrations in infertile patients before and after ovarian hyperstimulation for in vitro fertilization (IVF). Results highlight that total serum concentration of magnesium and calcium does not seem altered in infertile women. During stimulation with gonadotropins, the values of the two cations do not change significantly in ovarian-stimulated women. However, we found a downward trend in the total magnesium and calcium levels in relation to the rising estrogens.


How multi-organ microdevices can help foster drug development.
Multi-organ microdevices can mimic tissue-tissue interactions that occur as a result of metabolite travel from one tissue to other tissues in vitro. These systems are capable of simulating human metabolism, including the conversion of a pro-drug to its effective metabolite as well as its subsequent therapeutic actions and toxic side effects. Since tissue-tissue interactions in the human body can play a significant role in determining the success of new pharmaceuticals, the development and use of multi-organ microdevices present an opportunity to improve the drug development process. The devices have the potential to predict potential toxic side effects with higher accuracy before a drug enters the expensive phase of clinical trials as well as to estimate efficacy and dose response. Multi-organ microdevices also have the potential to aid in the development of new therapeutic strategies by providing a platform for testing in the context of human metabolism (as opposed to animal models). Further, when operated with human biopsy samples, the devices could be a gateway for the development of individualized medicine. Here we review studies in which multi-organ microdevices have been developed and used in a ways that demonstrate how the devices' capabilities can present unique opportunities for the study of drug action. We will also discuss challenges that are inherent in the development of multi-organ microdevices. Among these are how to design the devices, and how to create devices that mimic the human metabolism with high authenticity. Since single organ devices are testing platforms for tissues that can later be combined with other tissues within multi-organ devices, we will also mention single organ devices where appropriate in the discussion.


Calcium binding activity of the epidermal growth factor-like domains of the apicomplexan microneme protein EtMIC4.
Microneme proteins are secreted from apicomplexan parasites during invasion of host cells and they play crucial roles in parasite-host cell adhesion. EtMIC4 is a 240 kDa transmembrane protein from Eimeria tenella that contains 31 tandemly arranged epidermal growth factor (EGF), like repeats within its extracellular domain. The majority of these repeats have calcium binding (cb) consensus sequences. Little is known about cbEGFs in apicomplexan parasites but their presence in microneme proteins suggests that they may contribute to parasite-host interactions. To investigate the potential role of cbEGFs we have expressed and correctly refolded a cbEGF triplet from EtMIC4 (cbEGF7-9) and demonstrated that this triplet binds calcium. Circular dichroism spectroscopic analysis of cbEGF7-9 demonstrates that the molecule undergoes a gradual change in conformation with increasing levels of calcium. In the presence of calcium, the triplet becomes resistant to proteolytic degradation by a variety of proteases, a characteristic feature of cbEGF repeats from higher eukaryotic proteins, such as fibrillin, suggesting that calcium binding induces the formation of a rigid conformation. Moreover, mass spectrometric mapping of the cleavage sites that are protected by calcium shows that these sites are located both close to and distant from the calcium binding sites, indicating that protection is not due to steric hindrance by calcium ions, but rather due to the overall conformation adopted by the triplet in the presence of calcium. Thus, the tandemly-arranged cbEGF repeats within EtMIC4 provide a mechanism whereby, in the calcium-rich extracellular environment, the molecule could adopt a protease-resistant, rigid structure that could favour its interaction with host cell ligands.


Health status, psychological symptoms, mood, and cognition in L-thyroxine-treated hypothyroid subjects.
Many hypothyroid subjects receiving L-thyroxine (L-T4) complain of psychological symptoms or cognitive dysfunction. However, there is limited validated information on these self-reports. Cross-sectional comparison of 20 euthyroid and 34 treated hypothyroid subjects, aged 20-45 years, with normal thyroid-stimulating hormone (TSH) levels. Subjects underwent the following validated measures: Short Form 36 (SF-36); Symptom Checklist 90-Revised (SCL-90-R); Profile of Mood States (POMS); and tests of declarative memory (Paragraph Recall, Complex Figure), working memory (N-Back, Subject Ordered Pointing, Digit Span Backwards), and motor learning (Pursuit Rotor). L-T4-treated subjects had higher mean TSH and free T4 levels, but free triiodothyronine (T3) levels were comparable to controls. L-T4-treated subjects had decrements on SF-36 and SCL-90-R summary scales and subscales. These subjects performed slightly worse on N-Back and Pursuit Rotor tests. Neither TSH nor thyroid hormone levels were associated with performance on psychological or cognitive measures. This group of L-T4-treated subjects had decrements in health status, psychological function, working memory, and motor learning compared to euthyroid controls. Higher mean TSH levels suggest this may be related to suboptimal treatment, although there were no correlations between TSH levels and outcomes. These findings are limited by potential selection bias, and randomized studies targeting different TSH levels and memory subdomains would clarify these issues.


Regulation of the trafficking and the function of the metalloprotease ADAM10 by tetraspanins.
By interacting directly with partner proteins and with one another, tetraspanins organize a network of interactions referred to as the tetraspanin web. ADAM10 (A Disintegrin And Metalloprotease 10), an essential membrane-anchored metalloprotease that cleaves off the ectodomain of a large variety of cell surface proteins including cytokines, adhesion molecules, the precursor of the β-amyloid peptide APP or Notch, has emerged as a major component of the tetraspanin web. Recent studies have shown that ADAM10 associates directly with all members (Tspan5, Tspan10, Tspan14, Tspan15, Tspan17 and Tspan33) of a subgroup of tetraspanins having eight cysteines in the large extracellular domain and referred to as TspanC8. All TspanC8 regulate ADAM10 exit from the endoplasmic reticulum, but differentially regulate its subsequent trafficking and its function, and have notably a different impact on Notch signaling. TspanC8 orthologs in invertebrates also regulate ADAM10 trafficking and Notch signaling. It may be possible to target TspanC8 tetraspanins to modulate in a tissue- or substrate-restricted manner ADAM10 function in pathologies such as cardiovascular diseases, cancer or Alzheimer's disease.


Freezing and desiccation tolerance in entomopathogenic nematodes: diversity and correlation of traits.
The ability of entomopathogenic nematodes to tolerate environmental stress such as desiccating or freezing conditions, can contribute significantly to biocontrol efficacy. Thus, in selecting which nematode to use in a particular biocontrol program, it is important to be able to predict which strain or species to use in target areas where environmental stress is expected. Our objectives were to (i) compare inter- and intraspecific variation in freeze and desiccation tolerance among a broad array of entomopathogenic nematodes, and (ii) determine if freeze and desiccation tolerance are correlated. In laboratory studies we compared nematodes at two levels of relative humidity (RH) (97% and 85%) and exposure periods (24 and 48 h), and nematodes were exposed to freezing temperatures (-2°C) for 6 or 24 h. To assess interspecific variation, we compared ten species including seven that are of current or recent commercial interest: Heterorhabditis bacteriophora (VS), H. floridensis, H. georgiana, (Kesha), H. indica (HOM1), H. megidis (UK211), Steinernema carpocapsae (All), S. feltiae (SN), S. glaseri (VS), S. rarum (17C&E), and S. riobrave (355). To assess intraspecific variation we compared five strains of H. bacteriophora (Baine, Fl1-1, Hb, Oswego, and VS) and four strains of S. carpocapsae (All, Cxrd, DD136, and Sal), and S. riobrave (355, 38b, 7-12, and TP). S. carpocapsae exhibited the highest level of desiccation tolerance among species followed by S. feltiae and S. rarum; the heterorhabditid species exhibited the least desiccation tolerance and S. riobrave and S. glaseri were intermediate. No intraspecific variation was observed in desiccation tolerance; S. carpocapsae strains showed higher tolerance than all H. bacteriophora or S. riobrave strains yet there was no difference detected within species. In interspecies comparisons, poor freeze tolerance was observed in H. indica, and S. glaseri, S. rarum, and S. riobrave whereas H. georgiana and S. feltiae exhibited the highest freeze tolerance, particularly in the 24-h exposure period. Unlike desiccation tolerance, substantial intraspecies variation in freeze tolerance was observed among H. bacteriophora and S. riobrave strains, yet within species variation was not detected among S. carpocapsae strains. Correlation analysis did not detect a relationship between freezing and desiccation tolerance.


Uneven mapping of magnocellular and parvocellular projections from the lateral geniculate nucleus to the striate cortex in the macaque monkey.
Central vision is substantially over represented in the lateral geniculate nucleus (dLGN) and striate cortex. The over representation could be accompanied by a selective expansion of central vision in parvocellular dLGN, in which case the ratio of parvocellular to magnocellular inputs to striate cortex should change with retinal eccentricity. To test this, sample ratios were determined from counts of neurons in dLGN labelled retrogradely with WGA-HRP from striate cortex at the cortical representations of various eccentricities. Parvocellular to magnocellular ratios decreased from a mean of 35:1 at the fovea to 5:1 at 15 degrees eccentricity. Furthermore, they exceeded the ratio of P beta to P alpha ganglion cells in central retina, but not in peripheral retina, showing that the uneven P to M ratio in the LGN does not merely mirror the distribution of ganglion cells in the retina. This provides direct evidence for selective over representation of central vision in parvocellular dLGN.


Characterization of TNSALP expression, localization, and activity in ovine utero-placental tissues.
Tissue-nonspecific alkaline phosphatase (TNSALP; encoded by ALPL gene) has a critical role in the regulation of phosphate homeostasis postnatally. However, the utero-placental expression of TNSALP and the role in phosphate transport in pregnancy is poorly understood. Estrous cycles of ewes were synchronized, and ewes were euthanized and hysterectomized on Day 1, 9, or 14 of the estrous cycle or bred to fertile rams and euthanized and hysterectomized on Day 9, 12, 17, 30, 50, 70, 90, 110, or 125 of pregnancy. The expression of ALPL mRNA, immunolocalization of TNSALP protein, and quantification and localization of TNSALP enzymatic activity was performed on ovine endometria and placentomes. Day of the estrous cycle did not alter ALPL mRNA expression or enzymatic activity of TNSALP. TNSALP protein localized to uterine epithelial and stromal cells, blood vessels, myometrium, caruncular and cotyledonary stroma. TNSALP activity was localized to uterine epithelia, blood vessels, caruncular stroma (from Day 70 of gestation), and the apical surface of chorionic epithelia (from Day 50 of gestation). TNSALP protein and activity localized to the apical surface of uterine epithelia during the estrous cycle and in early pregnancy. Endometrial TNSALP enzymatic activity was downregulated on Day 17 and 30 of gestation (P < 0.05). Expression of ALPL mRNA decreased in late gestation in endometria and placentomes (P < 0.05). TNSALP activity peaked in placentomes on Days 70 and 90 of gestation. Collectively, these results suggest a potential role of TNSALP in the regulation of phosphate transport and homeostasis at the maternal-conceptus interface in ruminants.


Molecular basis of rifampin resistance in Mycobacterium leprae.
Rifampin is currently the most potent drug used in leprosy control programs. We show that the rifampin resistance which emerged in nine patients with lepromatous leprosy, who had received rifampin monotherapy, stemmed from mutations in the rpoB gene, which encodes the beta subunit of RNA polymerase of Mycobacterium leprae. In eight cases missense mutations were found to affect a serine residue, Ser-425, while in the remaining mutant a small insertion was found close to this site. These findings will be of use for the development of a rapid screening procedure, involving the polymerase chain reaction, for monitoring the emergence of rifampin-resistant M. leprae strains.


Cooking and future risk of all-cause and cardiopulmonary mortality.
Cooking is practiced worldwide and is associated with multiple social, economic and environmental factors; thus, understanding cooking-related health effects would have broad public health implications. Here, we show that after an average 9.9 years of follow-up for 510,106 Chinese adults, always cooking with clean fuels was associated with lower risks of all-cause (0.90 [95% confidence interval 0.87-0.93]; P = 1.39 × 10-9), cardiovascular (0.83 [0.78-0.87]; P = 6.83 × 10-11) and respiratory (0.88 [0.79-0.99]; P = 0.026) mortality compared with non-cooking, of which 50.1% (14.5-85.6%) to 66.0% (38.5-85.8%) could be attributed to increased household physical activity. The mortality risks decreased with extended duration of cooking with clean fuels in dose-response manners, with the lowest hazard ratios of 0.74 (0.68-0.80; P = 1.20 × 10-13) for all-cause and 0.62 (0.55-0.71; P = 3.15 × 10-12) for cardiovascular mortality among never-smokers reported over 25 years of cooking. Our findings suggest lower future mortality risks may be gained only when cooking with clean fuels.


Biological Characterization and Clinical Relevance of Circulating Tumor Cells: Opening the Pandora's Box of Multiple Myeloma.
Bone marrow (BM) aspirates are the gold standard for patient prognostication and genetic characterization in multiple myeloma (MM). However, they represent an important limitation for periodic disease monitoring because they entail an aggressive procedure. Moreover, recent findings show that a single BM aspirate is unable to reflect the complex MM heterogeneity. Recent advances in flow cytometry, microfluidics, and "omics" technologies have opened Pandora's box of MM: The detection and isolation of circulating tumor cells (CTCs) offer a promising and minimally invasive alternative for tumor assessment and metastasis study. CTCs are detectable in premalignant and active MM states, and their enumeration has strong prognostic value, to the extent that it is challenging current stratification systems. In addition, CTCs reflect with high precision both intra- and extra-medullary disease at the phenotypic, genomic, and transcriptomic levels. Despite this high resemblance between tumor clones in distinct locations, some subtle (not random) differences might shed some light on the metastatic process. Thus, it has been suggested that a hypoxic and pro-inflammatory microenvironment could induce an arrest in proliferation forcing tumor cells to recirculate. Herein, we summarize data on the characterization of MM CTCs as well as their clinical and research potential.


Resistance and inactivation kinetics of bacterial strains isolated from the non-chlorinated and chlorinated effluents of a WWTP.
The microbiological quality of water from a wastewater treatment plant that uses sodium hypochlorite as a disinfectant was assessed. Mesophilic aerobic bacteria were not removed efficiently. This fact allowed for the isolation of several bacterial strains from the effluents. Molecular identification indicated that the strains were related to Aeromonas hydrophila, Escherichia coli (three strains), Enterobacter cloacae, Kluyvera cryocrescens (three strains), Kluyvera intermedia, Citrobacter freundii (two strains), Bacillus sp. and Enterobacter sp. The first five strains, which were isolated from the non-chlorinated effluent, were used to test resistance to chlorine disinfection using three sets of variables: disinfectant concentration (8, 20 and 30 mg·L(-1)), contact time (0, 15 and 30 min) and water temperature (20, 25 and 30 °C). The results demonstrated that the strains have independent responses to experimental conditions and that the most efficient treatment was an 8 mg·L(-1) dose of disinfectant at a temperature of 20 °C for 30 min. The other eight strains, which were isolated from the chlorinated effluent, were used to analyze inactivation kinetics using the disinfectant at a dose of 15 mg·L(-1) with various retention times (0, 10, 20, 30, 60 and 90 min). The results indicated that during the inactivation process, there was no relationship between removal percentage and retention time and that the strains have no common response to the treatments.


Changes in the Chemical Barrier Composition of Tears in Alzheimer's Disease Reveal Potential Tear Diagnostic Biomarkers.
Alzheimer's disease (AD) is one of the most common neurodegenerative diseases, with increasing prevalence affecting millions of people worldwide. Currently, only autopsy is able to confirm the diagnosis with a 100% certainty, therefore, biomarkers from body fluids obtained by non-invasive means provide an attractive alternative for the diagnosis of Alzheimer`s disease. Global changes of the protein profile were examined by quantitative proteomics; firstly, electrophoresis and LC-MS/MS were used, thereafter, SRM-based targeted proteomics method was developed and applied to examine quantitative changes of tear proteins. Alterations in the tear flow rate, total tear protein concentration and composition of the chemical barrier specific to AD were demonstrated, and the combination of lipocalin-1, dermcidin, lysozyme-C and lacritin was shown to be a potential biomarker, with an 81% sensitivity and 77% specificity.


A pilot study assessing the bioavailability and pharmacokinetics of diazepam after intranasal and intravenous administration in healthy volunteers.
Diazepam rectal gel (Diastat(®)) is the only FDA-approved product indicated for acute repetitive seizures. Despite its proven efficacy, most older children and adults object to this route of administration. As a result, many patients do not realize the benefit of a therapy that can improve outcomes and decrease healthcare costs. Intranasal administration of benzodiazepines offers a potential alternative. The primary objective of this study was to compare the bioavailability and pharmacokinetics of two novel intranasal (IN) diazepam (DZP) formulations versus intravenous (IV) administration in healthy volunteers. Twenty-four healthy volunteers were randomized into an open-label, three-way crossover study. 10mg doses of two investigational intranasal DZP formulations (solution, suspension) and a 5mg IV dose of commercially available DZP injectable, USP were given. A two-week washout period separated treatments. Plasma samples for DZP analysis were collected pre-dose and at regular intervals up to 240 h post-dose. DZP concentration-time data were analyzed using a non-compartmental pharmacokinetics approach. Exposure following administration of DZP IN solution (absolute bioavailability - 97%) was greater than the IN suspension (absolute bioavailability - 67%). Mean Cmaxvalues for the suspension and solution formulations were 221 ng/mL and 272 ng/mL, respectively. Median time to maximum concentration (Tmax) was 1h and 1.5h for suspension and solution formulation, respectively. Both investigational intranasal formulations were well tolerated. The results of this pilot study indicate that development of an intranasal diazepam formulation with high bioavailability, reasonable variability, and good tolerability is feasible.


The utility of a health risk assessment in providing care for a rural free clinic population.
Free clinics are an important part of our country's health safety net, serving a working poor uninsured population. With limited resources and heavily dependent upon volunteer health care providers, these clinics have historically focused on stopgap, band-aid solutions to the population's health problems. Embracing a new paradigm, free clinics are now prioritizing resources for disease prevention and health promotion. We initiated a Healthy Friday Clinic project in a rural, southwest Virginia free clinic. The clinic operated every Friday and was open to all people eligible for care in the free clinic. Each participant completed a 43 question Health Risk Appraisal which was used to calculate current risk age (age as determined by current lifestyle choices), optimal risk age (age with optimal lifestyle choices) and potential risk years gained (current risk age - optimal risk age) as well as a ranked listing of modifiable risk factors. The total sum of potential risk years gained in the free clinic population of 186 subjects was 371.4. Frequency distributions on potential risk years gained by each of the eleven modifiable risk factors revealed the following, in order of impact: quitting smoking could result in a total of 173.5 risk years gained; reducing alcohol consumption, 64.2 years gained; reducing blood pressure, 50.8 years gained; increasing seatbelt use, 38.2 years gained; weight reduction, 24.7 years gained; having regular mammograms, 6.8 years gained; reducing cholesterol levels, 5.8 years gained; reducing frequency of speeding while driving, 3.5 years gained; having regular pap tests, 2.3 years gained; improving HDL levels, 0.9 years gained; and reducing use of smokeless tobacco, 0.8 years gained. Each person received an individualized letter explaining his evaluation along with resources for making changes. Health risk assessments play a role in changing health beliefs and behaviors by providing subjects with individualized feedback on how their lifestyle choices impact their health and well-being. Summed data from health risk appraisals can also be a useful tool in determining the allocation of limited health resources. Whether health risk assessments impact health outcomes directly needs to be studied.


Influenza vaccination might reduce the risk of ischemic stroke in patients with atrial fibrillation: A population-based cohort study.
Atrial fibrillation (AF) is associated with the risk of ischemic stroke, regardless of the administration of appropriate antithrombotic prophylaxis. This study investigated whether influenza vaccination is associated with the risk of ischemic stroke, to determine a solution to reduce this risk in patients with AF. We used data from the Taiwan National Health Insurance Research Database. The study cohort comprised all patients diagnosed as having AF (n = 14 454) before January 1, 2005; these patients were followed until December 31, 2012. The index date was January 1, 2005. A propensity score was derived using a logistic regression model to estimate the effect of vaccination by accounting for covariates that predict receiving the intervention (vaccine). A Cox proportional hazard model was used to calculate the hazard ratios (HRs) of ischemic stroke in vaccinated and unvaccinated patients with AF. We included 6570 patients (2547 [38.77%] with and 4023 [61.23%] without influenza vaccination). The adjusted HRs (aHRs) of ischemic stroke were lower in the vaccinated patients than in the unvaccinated patients (influenza season, noninfluenza season, and all seasons: aHRs = 0.59, 0.50, and 0.55; P < 0.001, P < 0.001, and P < 0.001, respectively). Influenza vaccination might exert a dose-response effect against ischemic stroke in patients with AF who have risk factors for ischemic stroke by reducing the incidence of ischemic stroke, particularly in those aged 65-74 and ≥75 y.


Complete Characterization of Polyacyltrehaloses from Mycobacterium tuberculosis H37Rv Biofilm Cultures by Multiple-Stage Linear Ion-Trap Mass Spectrometry Reveals a New Tetraacyltrehalose Family.
Polyacylated trehaloses in Mycobacterium tuberculosis play important roles in pathogenesis and structural roles in the cell envelope, promoting the intracellular survival of the bacterium, and are potential targets for drug development. Herein, we describe a linear ion-trap multiple-stage mass spectrometric approach (LIT MSn) with high-resolution mass spectrometry to the structural characterization of a glycolipid family that includes a 2,3-diacyltrehalose, 2,3,6-triacyltrehalose, 2,3,6,2',4'-petaacyltrehalose, and a novel 2,3,6,2'-tetraacyltrehalose (TetraAT) subfamily isolated from biofilm cultures of M. tuberculosis H37Rv. The LIT MSn spectra (n = 2, 3, or 4) provide structural information to unveil the location of the palmitoyl/stearoyl and one to four multiple methyl-branched fatty acyl substituents attached to the trehalose backbone, leading to the identification of hundreds of glycolipid species with many isomeric structures. We identified a new TetraAT subfamily whose structure has not been previously defined. We also developed a strategy for defining the structures of the multiple methyl-branched fatty acid substituents, leading to the identification of mycosanoic acid, mycolipenic acid, mycolipodienoic acid, mycolipanolic acid, and a new cyclopropyl-containing acid. The observation of the new TetraAT family, and the realization of the structural similarity between the various subfamilies, may have significant implications in the biosynthetic pathways of this glycolipid family.


Gluten- and casein-free diets for autistic spectrum disorder.
It has been suggested that peptides from gluten and casein may have a role in the origins of autism and that the physiology and psychology of autism might be explained by excessive opioid activity linked to these peptides. Research has reported abnormal levels of peptides in the urine and cerebrospinal fluid of persons with autism. If this is the case, diets free of gluten and /or casein should reduce the symptoms associated with autism. To determine the efficacy of gluten- and/or casein- free diets as an intervention to improve behaviour, cognitive and social functioning in individuals with autism. Electronic searching of abstracts from the Cochrane Library (Issue 3, 2003), PsycINFO (1971- May 2003), EMBASE (1974- May 2003), CINAHL (1982- May 2003), MEDLINE (1986- May 2003), ERIC (1965-2003), LILACS (to 2003) and the specialist register of the Cochrane Complementary Medicine Field (January 2004). Review bibliographies were also examined to identify potential trials. All randomised controlled trials involving programmes which eliminated gluten, casein or both gluten and casein from the diets of individuals diagnosed with autistic spectrum disorder. Abstracts of studies identified in searches of electronic databases were read and assessed to determine whether they might meet the inclusion criteria. The authors independently selected the relevant studies from the reports identified in this way. As only one trial fitted the inclusion criteria, no meta-analysis is currently possible and data are presented in narrative form. The one trial included reported results on four outcomes. Unsurprisingly in such a small-scale study, the results for three of these outcomes (cognitive skills, linguistic ability and motor ability) had wide confidence intervals that spanned the line of nil effect. However, the fourth outcome, reduction in autistic traits, reported a significant beneficial treatment effect for the combined gluten- and casein- free diet. This is an important area of investigation and large scale, good quality randomised controlled trials are needed.


Endovascular treatment for chronic pulmonary hypertension: a focus on angioplasty for chronic thromboembolic pulmonary hypertension.
Percutaneous transluminal pulmonary angioplasty (PTPA) was introduced for the treatment of chronic thromboembolic pulmonary hypertension (CTEPH) in the late 20(th) century, and first attempts in collective patients were made in 2001 with beneficial effects but a moderate amount of complications. It was refined around 2010, and has been recently established as an effective and safe treatment. The indication was originally inoperable CTEPH with peripheral lesions, but has now widened to symptomatic or hypoxic patients. The lesion is typically a meshwork-like structure of organized thrombi and is sometimes not seen as a stenosis angiographically, necessitating other means of investigation such as measurement of distal pressure. The technique to treat lesions is the same as for coronary angioplasty except in several ways. Expert commentary: The effects of PTPA are comparable to those of surgical endarterectomy, and the complications of reperfusion pulmonary edema and vascular injury are now controlled by several strategies and based on experience.


Cell-cycle reprogramming for PI3K inhibition overrides a relapse-specific C481S BTK mutation revealed by longitudinal functional genomics in mantle cell lymphoma.
Despite the unprecedented clinical activity of the Bruton tyrosine kinase (BTK) inhibitor ibrutinib in mantle cell lymphoma (MCL), acquired resistance is common. By longitudinal integrative whole-exome and whole-transcriptome sequencing and targeted sequencing, we identified the first relapse-specific C481S mutation at the ibrutinib binding site of BTK in MCL cells at progression following a durable response. This mutation enhanced BTK and AKT activation and tissue-specific proliferation of resistant MCL cells driven by CDK4 activation. It was absent, however, in patients with primary resistance or progression following transient response to ibrutinib, suggesting alternative mechanisms of resistance. Through synergistic induction of PIK3IP1 and inhibition of PI3K-AKT activation, prolonged early G1 arrest induced by PD 0332991 (palbociclib) inhibition of CDK4 sensitized resistant lymphoma cells to ibrutinib killing when BTK was unmutated, and to PI3K inhibitors independent of C481S mutation. These data identify a genomic basis for acquired ibrutinib resistance in MCL and suggest a strategy to override both primary and acquired ibrutinib resistance. We have discovered the first relapse-specific BTK mutation in patients with MCL with acquired resistance, but not primary resistance, to ibrutinib, and demonstrated a rationale for targeting the proliferative resistant MCL cells by inhibiting CDK4 and the cell cycle in combination with ibrutinib in the presence of BTK(WT) or a PI3K inhibitor independent of BTK mutation. As drug resistance remains a major challenge and CDK4 and PI3K are dysregulated at a high frequency in human cancers, targeting CDK4 in genome-based combination therapy represents a novel approach to lymphoma and cancer therapy. Cancer Discov; 4(9); 1022-35. ©2014 AACR. This article is highlighted in the In This Issue feature, p. 973.


Pressure ulcer prevalence, use of preventive measures, and mortality risk in an acute care population: a quality improvement project.
The primary aim of this quality improvement project was to determine pressure prevalence, risk of mortality, and use of preventive measures in a group of hospitalized patients. Two hundred fifty-eight patients recruited from Skaraborg Hospital in Sweden were assessed. A 1-day point prevalence study was carried out using a protocol advocated by the European PU Advisory Panel. Patients' age, gender, severity of PU (grades I-IV), anatomical location of PU, and use of preventive measures were recorded. The Swedish language version of the Modified Norton Scale was used for PU risk assessment. Data were collected by nurses trained according to the Web-based training: PU classification, "ePuclas2." After 21 months, a retrospective audit of the electronic records for patients identified with pressure ulcers was completed. The point prevalence of pressure ulcers was 23%. The total number of ulcers was 85, most were grade 1 (n = 39). The most common locations were the sacrum (n = 15) and the heel (n = 10). Three percent of patients (n = 9) had been assessed during their current hospital stay using a risk assessment tool. There was a statistically significant relationship between pressure ulcer occurrence and a low total score on the Modified Norton Scale. The patients' ages correlated significantly to the presence of a pressure ulcer. Patients with a pressure ulcer had a 3.6-fold increased risk of dying within 21 months, as compared with those without a pressure ulcer. Based on results from this quality improvement project, we recommend routine pressure ulcer risk assessment for all patients managed in a hospital setting such as ours. We further recommend that particular attention should be given to older and frail patients who are at higher risk for pressure ulcer occurrence and mortality.


Aortic Disease Presentation and Outcome Associated With ACTA2 Mutations.
ACTA2 mutations are the major cause of familial thoracic aortic aneurysms and dissections. We sought to characterize these aortic diseases in a large case series of individuals with ACTA2 mutations. Aortic disease, management, and outcome associated with the first aortic event (aortic dissection or aneurysm repair) were abstracted from the medical records of 277 individuals with 41 various ACTA2 mutations. Aortic events occurred in 48% of these individuals, with the vast majority presenting with thoracic aortic dissections (88%) associated with 25% mortality. Type A dissections were more common than type B dissections (54% versus 21%), but the median age of onset of type B dissections was significantly younger than type A dissections (27 years versus 36 years). Only 12% of aortic events were repair of ascending aortic aneurysms, which variably involved the aortic root, ascending aorta, and aortic arch. Overall, cumulative risk of an aortic event at age 85 years was 0.76 (95% confidence interval, 0.64-0.86). After adjustment for intrafamilial correlation, sex and race, mutations disrupting p.R179 and p.R258 were associated with significantly increased risk for aortic events, whereas p.R185Q and p.R118Q mutations showed significantly lower risk of aortic events compared with other mutations. ACTA2 mutations are associated with high risk of presentation with an acute aortic dissection. The lifetime risk for an aortic event is only 76%, suggesting that additional environmental or genetic factors play a role in expression of aortic disease in individuals with ACTA2 mutations.


Impact Study: MK-0646 (Dalotuzumab), Insulin Growth Factor 1 Receptor Antibody Combined with Pemetrexed and Cisplatin in Stage IV Metastatic Non-squamous Lung Cancer.
Insulin-like growth factor 1 receptor (IGF-1R) regulates cell growth, proliferation, and apoptosis. Adenocarcinoma and never-smokers have a higher expression of IGF-1R, which is associated with worse overall survival. Dalotuzumab-MK0646 (D) is a humanized monoclonal antibody that targets IGF-1R. Pemetrexed (P) has higher activity in non-squamous lung cancer (NSQL). We initiated a randomized phase II trial to test the combination of P and Cisplatin (C) ± D in NSQL. Eligibility criteria were untreated NSQL stage IV, ECOG 0 or 1, measurable disease, adequate renal, hepatic and hematologic function, and no other intercurrent illness. P at 500 mg/m(2) and C at 75 mg/m(2) IV were given every 3 weeks. D was given at 10 mg/kg IV weekly on days 1, 8, and 15 of every 3-week cycle in the experimental group. The patients had a radiographic assessment after every two cycles and were treated for a maximum of six cycles if there was a response or stable disease. The primary objective of the study was to compare the clinical response rates of PC vs. PC + D. From 1/2009 to 2/2011, the study accrued 26 subjects: 16 male and 10 female, with a median age of 59; 14 were treated with PC and 12 were treated with PC + D. We observed two partial responses (PR), seven stable disease (SD), three progressive disease (PD), and two not evaluable (NE) in the PC arm. In comparison, for the PC + D arm, there were three PR, four SD, four PD, and one NE. The hematologic toxicity was similar in both groups. There was higher incidence of hyperglycemia in the experimental group; four cases with grade 3 and one case with grade 4. PC + D had a similar response rate compared to PC, with a higher rate of hyperglycemia. Identification of responders using predictive markers would be key to continuing the study of D in NSQL. NCT00799240, clinicaltrials.gov.


Analysis of the colorimetric properties of goniochromatic colors using the MacAdam limits under different light sources.
Technological innovation in all areas has led to the appearance in recent years of new metallic and pearlescent materials, yet no exhaustive studies have been conducted to assess their colorimetric capabilities. The chromatic variability of these special-effect pigments may largely be due to the three-dimensional effect of their curved shapes and orientations when they are directionally or diffusely illuminated. Our study examines goniochromatic colors using the optimal colors (MacAdam limits) associated with normal colors (photometric scale of relative spectral reflectance from 0 to 1) under certain conventional illuminants and other light sources. From a database of 91 metallic and interference samples and using a multi-gonio-spectrophotometer, we analyzed samples with lightness values of more than 100 and others with lightness values of less than 100, but with higher chromaticities than optimal colors, which places them beyond the MacAdam limits. Our study thus demonstrates the existence of chromatic perceptions beyond the normal solid color associated with these materials and independent of the light source. The challenge for future research, therefore, is to replicate and render these color appearances in current and future color reproduction technologies for computer graphics.


Physician complicity in misrepresentation and omission of evidence of torture in postdetention medical examinations in Turkey.
Between June 1994 and October 1995, representatives of Physicians for Human Rights studied the problem of physician complicity in torture (ie, misrepresentation and omission of medical evidence in postdetention examinations of detainees) in Turkey. The research consisted of a survey of forensic documentation of torture, interviews with individual physicians who examine detainees, analyses of official medical reports of detainees, and interviews with survivors of torture. Results from the survey, interviews, and medical report analyses provide evidence that torture of political and criminal detainees continues to occur in Turkey and that Turkish physicians are coerced to ignore, misrepresent, and omit evidence of torture in their examinations of detainees to certify that there are no physical signs of torture.


Evaluating the utility of ICD-10 diagnostic criteria for postconcussion syndrome following mild traumatic brain injury.
The present study investigated the utility of the International Classification of Diseases and Related Health Problems, 10th edition (ICD-10) diagnostic criteria for postconcussion syndrome (PCS) symptoms by comparing symptom endorsement rates in a group of patients with mild traumatic brain injury (MTBI) to those of a noninjured control group at one month and three months post-injury. The 110 MTBI patients and 118 control participants were group-matched on age, gender, and education level. Seven of the nine self-reported ICD-10 PCS symptoms differentiated the groups at one month post-injury and two symptoms differentiated the groups at three months post-injury: symptom endorsement rates were higher in the MTBI group at both time periods. Fatiguing quickly and dizziness/vertigo best differentiated the groups at both time periods, while depression and anxiety/tension failed to differentiate the groups at either time period. Collectively, the ICD-10 PCS symptoms accurately classified the MTBI patients at one month post-injury, with the optimal positive test threshold of endorsement of five symptoms coinciding with a sensitivity and specificity of 73% and 61%, respectively. The ICD-10 PCS symptoms were unable to accurately classify the MTBI patients at three months post-injury.


In vitro toxicity and genotoxicity assessment of disinfection by-products, organic N-chloramines.
Disinfection by-products (DBPs) are of concern to both water industries and health authorities. Although several classes of DBPs have been studied, and there are regulated safe levels in disinfected water for some, a large portion of DBPs are not characterized, and need further investigation. Organic N-chloramines are a group of DBPs, which can be formed during common disinfection processes such as chlorination and chloramination, but little is known in terms of their toxicological significance if consumed in drinking water. Only a few in vitro studies using bacterial assays have reported some genotoxic potential of organic N-chloramines, largely in the context of inflammatory processes in the body rather than exposure through drinking water. In this study, we investigated 16 organic N-chloramines produced by chlorination of model amino acids and amines. It was found that within the drinking water-relevant micromolar concentration range, four compounds were both cytotoxic and genotoxic to mammalian cells. A small reduction of cellular GSH was also observed in the treatment with these four compounds, but not of a magnitude to account for the cytotoxicity and genotoxicity. The results presented in this study demonstrate that some organic N-chloramines, at low concentrations that might be present in disinfected water, can be harmful to mammalian cells.


Bismuth-Silicon and Bismuth-Polyurethane Composite Shields for Breast Protection in Chest Computed Tomography Examinations.
The article aims at constructing protective composite shields for breasts in chest computed tomography and investigating the effects of applying these new bismuth composites on dose and image quality. Polyurethane and silicon with 5% of bismuth were fabricated as a protective shield. At first, their efficiency in attenuating the X-ray beam was investigated by calculating the total attenuation coefficients at diagnostic energy range. Then, a physical chest phantom was scanned without and with these shields at tube voltage of 120 kVp, and image parameters together with dose values were studied. The results showed that these two shields have great effects on attenuating the X-ray beam, especially for lower energies (<40 kV), and in average, the attenuation coefficients of bismuth-polyurethane composite are higher in this energy range. The maximum relative differences between the average Hounsfield units (HUs) and noises of images without and with shield for both composites in 13 regions of interest were 4.5% and 15.7%, respectively. Moreover, primary investigation confirmed the ability of both shields (especially polyurethane-bismuth composite) in dose reduction. Comparing these two composites regarding the amount of dose reduction, the changes in HU and noise, and attenuation coefficients in diagnostic energy range, it seems that polyurethane composite is more useful for dose reduction, especially for higher tube voltages.


The Hypoglycemic and Synergistic Effect of Loganin, Morroniside, and Ursolic Acid Isolated from the Fruits of Cornus officinalis.
Hypoglycemic activity-guided separation of ethanol extracts from the fruits of Cornus officinalis Sieb. et Zucc (CO) led to the isolation of loganin, morroniside, and ursolic acid. The antidiabetic capacity of CO extracts and related compounds was further investigated in diabetes mellitus mice. The results suggested that both CO extracts and pure compounds could ameliorate diabetes-associated damages and complications. Oral administration of loganin and morroniside decreased fasting blood glucose levels in diabetes mellitus mice. Ursolic acid exhibited the highest reactive oxygen species scavenging activity and α-glucosidase inhibitory activity. Notably, we noticed an interesting synergistic effect between loganin and ursolic acid. Given these favorable hypoglycemic properties, C. officinalis, a food and medicinal plant in China, may be used as a valuable food supplement for the treatment of diabetes mellitus.


The human immunodeficiency virus type 1-specific protein vpu is required for efficient virus maturation and release.
A deletion mutation affecting vpu was introduced into an infectious molecular clone of human immunodeficiency virus type 1, and the resultant phenotype was examined after infection of human T lymphocytes. The absence of vpu resulted in an accumulation of cell-associated viral proteins and impaired the release of progeny virions. Both electron microscopic and biochemical analyses indicated that a large proportion of the mutant particles was attached to the surface of infected cells. Significant variation in the size and shape of these progeny virions was observed. In addition, intracytoplasmic particles, some of which formed aberrant budding structures, were visualized in T cells infected with the vpu mutant. Indirect immunofluorescence analyses of cultures inoculated with wild-type virus with use of a vpu-specific antiserum demonstrated that vpu is mainly localized to a perinuclear region in the cytoplasm of virus-producing cells.


Genome-wide analysis reveals PADI4 cooperates with Elk-1 to activate c-Fos expression in breast cancer cells.
Peptidylarginine deiminase IV (PADI4) catalyzes the conversion of positively charged arginine and methylarginine residues to neutrally charged citrulline, and this activity has been linked to the repression of a limited number of target genes. To broaden our knowledge of the regulatory potential of PADI4, we utilized chromatin immunoprecipitation coupled with promoter tiling array (ChIP-chip) analysis to more comprehensively investigate the range of PADI4 target genes across the genome in MCF-7 breast cancer cells. Results showed that PADI4 is enriched in gene promoter regions near transcription start sites (TSSs); and, surprisingly, this pattern of binding is primarily associated with actively transcribed genes. Computational analysis found potential binding sites for Elk-1, a member of the ETS oncogene family, to be highly enriched around PADI4 binding sites; and coimmunoprecipitation analysis then confirmed that Elk-1 physically associates with PADI4. To better understand how PADI4 may facilitate gene transactivation, we then show that PADI4 interacts with Elk-1 at the c-Fos promoter and that, following Epidermal Growth Factor (EGF) stimulation, PADI4 catalytic activity facilitates Elk-1 phosphorylation, histone H4 acetylation, and c-Fos transcriptional activation. These results define a novel role for PADI4 as a transcription factor co-activator.


Tunable composite nanoparticle for plasmonics.
We present a numerical study of the tunability properties of a plasmonic subwavelength particle deposited on a metallic slab. The particle is composed of a metallic part, supporting a localized plasmon mode, separated from the slab by a dielectric spacer. It is shown that the position of the resonance wavelength can be modified over a large spectral range by changing either the spacer thickness by a few tens of nanometers or its susceptibility within the range of usual dielectrics. A linear relation is observed between the resonance wavelength and the spacer permittivity.


Shortened Modified Look-Locker Inversion recovery (ShMOLLI) for clinical myocardial T1-mapping at 1.5 and 3 T within a 9 heartbeat breathhold.
T1 mapping allows direct in-vivo quantitation of microscopic changes in the myocardium, providing new diagnostic insights into cardiac disease. Existing methods require long breath holds that are demanding for many cardiac patients. In this work we propose and validate a novel, clinically applicable, pulse sequence for myocardial T1-mapping that is compatible with typical limits for end-expiration breath-holding in patients. The Shortened MOdified Look-Locker Inversion recovery (ShMOLLI) method uses sequential inversion recovery measurements within a single short breath-hold. Full recovery of the longitudinal magnetisation between sequential inversion pulses is not achieved, but conditional interpretation of samples for reconstruction of T1-maps is used to yield accurate measurements, and this algorithm is implemented directly on the scanner. We performed computer simulations for 100 ms<T1 < 2.7 s and heart rates 40-100 bpm followed by phantom validation at 1.5T and 3T. In-vivo myocardial T1-mapping using this method and the previous gold-standard (MOLLI) was performed in 10 healthy volunteers at 1.5T and 3T, 4 volunteers with contrast injection at 1.5T, and 4 patients with recent myocardial infarction (MI) at 3T. We found good agreement between the average ShMOLLI and MOLLI estimates for T1 < 1200 ms. In contrast to the original method, ShMOLLI showed no dependence on heart rates for long T1 values, with estimates characterized by a constant 4% underestimation for T1 = 800-2700 ms. In-vivo, ShMOLLI measurements required 9.0 ± 1.1 s (MOLLI = 17.6 ± 2.9 s). Average healthy myocardial T1 s by ShMOLLI at 1.5T were 966 ± 48 ms (mean ± SD) and 1166 ± 60 ms at 3T. In MI patients, the T1 in unaffected myocardium (1216 ± 42 ms) was similar to controls at 3T. Ischemically injured myocardium showed increased T1 = 1432 ± 33 ms (p < 0.001). The difference between MI and remote myocardium was estimated 15% larger by ShMOLLI than MOLLI (p < 0.04) which suffers from heart rate dependencies for long T1. The in-vivo variability within ShMOLLI T1-maps was only 14% (1.5T) or 18% (3T) higher than the MOLLI maps, but the MOLLI acquisitions were twice longer than ShMOLLI acquisitions. ShMOLLI is an efficient method that generates immediate, high-resolution myocardial T1-maps in a short breath-hold with high precision. This technique provides a valuable clinically applicable tool for myocardial tissue characterisation.


Eugenol enhances the chemotherapeutic potential of gemcitabine and induces anticarcinogenic and anti-inflammatory activity in human cervical cancer cells.
Administration of natural or synthetic agents to inhibit, delay, block, or reverse the initiation and promotional events associated with carcinogenesis opens a new avenue for cancer prevention and treatment to reduce cancer morbidity and mortality. Eugenol, a potential chemopreventive agent, is a component of clove and several other spices such as basil, cinnamon, and bay leaves. A number of reports have shown that eugenol possesses antiseptic, analgesic, antibacterial, and anticancer properties. The present study was undertaken to evaluate the chemopreventive potential of eugenol alone and in combination with a chemotherapeutic agent such as gemcitabine. Eugenol showed dose-dependent selective cytotoxicity toward HeLa cells in comparison to normal cells, pointing to its safe cytotoxicity profile. A combination of eugenol and gemcitabine induced growth inhibition and apoptosis at lower concentrations, compared with the individual drugs. The analysis of the data using a combination index showed combination index values of <1 indicating strong synergistic interaction. The combination thus may enhance the efficacy of gemcitabine at lower doses and minimize the toxicity on normal cells. In addition, the expression analysis of genes involved in apoptosis and inflammation revealed significant downregulation of Bcl-2, COX-2, and IL-1β on treatment with eugenol. Thus, the results suggest that eugenol exerts its anticancer activities via apoptosis induction and anti-inflammatory properties and also provide the first evidence demonstrating synergism between eugenol and gemcitabine, which may enhance the therapeutic index of prevention and/or treatment of cervical cancer.


[Follow-up of child abuse victims: challenges for the pediatrician].
To review practical questions about the initial assistance and follow-up of child abuse victims and their families by pediatricians. A literature review was carried out using the MEDLINE and LILACS databases, including the years 2000 to 2005. Some articles from past years and books were included due to their importance. Initial assistance is one of the most important actions by health professionals for the protection of abused children in different healthcare sectors (community, outpatient clinics, emergency rooms and infirmary), and it is fundamental for the reduction of immediate and long-term negative consequences of violence. The protection services cannot monitor all the families under their responsibility and most child abuse cases are not even reported to those institutions; therefore, regular follow-up by a pediatrician is advisable. It is important to provide the family with support and guidance until the child is safe. The main challenges are: to be involved without causing more violence; to consider all the family as the focus of attention, including the family members who have committed the assault, helping them to change inadequate behaviors; to develop specific abilities to carry out this work, which must be multiprofessional, interdisciplinary and intersectoral. Families face difficulties when their children are abused and when the situation gains notoriety, demanding interventions from many institutions. In this process, a pediatrician can guide and help them to guarantee the protection and healthy development of their children. To overcome challenges, health professionals have to be technically and emotionally prepared.


Does Mandible-First Sequencing Increase Maxillary Surgical Accuracy in Bimaxillary Procedures?
In bimaxillary procedures, it is important to know how the chosen sequence affects the surgical outcome. The purpose of this study was to explore whether the theoretical advantages of using the mandible-first procedure were supported by clinical data. The authors performed a retrospective investigation on a cohort compiled from 3 published retrospective studies. The sample was composed of patients treated at the Radboud University Nijmegen Medical Centre (Nijmegen, the Netherlands) from 2010 to 2014 and the Odense University Hospital (Odense, Denmark) from 2011 to 2015. The inclusion criterion was bimaxillary surgery without maxillary segmentation. The exclusion criterion was lack of a virtual surgical plan. The primary outcome variable was surgical accuracy, defined as the mean difference between the obtained outcome and the virtual surgical plan. The primary predictor variable was the comparison between mandible-first and maxilla-first sequencing. Secondary predictors were inferior maxillary repositioning and counterclockwise (CCW) rotation. The confounding variable was the virtually planned reposition. Results were analyzed by mixed-model regression encompassing all variables, followed by a detailed analysis of positive results using 2-sample t tests. Overall, 145 patients were included for analysis (98 women; mean age, 28 years). Operating on the mandible first notably influenced maxillary positioning and placed the maxilla 1.5 mm posterior and with 1.4° of CCW rotation compared with virtual surgical planning. The interaction of surgical sequence with maxillary rotation showed similar surgical accuracy between maxilla-first surgery with clockwise rotation and mandible-first surgery with CCW rotation. Inferior maxillary repositioning resulted in the maxilla being placed 1.7 mm (maxilla-first sequence) and 2.0 mm (mandible-first sequence) posterior to the planned position. Surgical accuracy was considerably influenced by sequencing in bimaxillary procedures. It remains important to know how the chosen sequence affects the surgical outcome so that the virtual surgical plan can be adjusted accordingly.


Apoptosis and transmethylation metabolites in HL-60 cells.
To investigate the role of transmethylation metabolites in initiation of apoptosis in human leukemia HL-60 cells exposed to 3-deazaadenosine (c3 Ado) as single agent and c3Ado plus homocysteine thiolactone (Hcy), S-adenosylmethionine (AdoMet), S-adenosylhomocysteine (AdoHcy) and 3-deazaadenosylhomocysteine (c3AdoHcy), were measured by HPLC in HL-60 cells exposed to 1-100 microM c3Ado as single agent and 1 to 100 microM c3Ado plus 1 mM Hcy. 3-deaza-(+/-)-aristeromycin (c3Ari), a more specific and potent S-adenosylhomocysteine hydrolase inhibitor compared with c3Ado, was used to inhibit synthesis of c3AdoHcy. AdoMet increased to maximum values 1.5- and 2.3-fold control in cultures treated with c3Ado as single agent and c3Ado plus Hcy, respectively. AdoHcy did not change in cultures treated with c3Ado as single agent, but increased to maximum values 2.0-fold control when Hcy was added. The synthesis of c3AdoHcy was favored by Hcy, and 35- to 70-fold higher levels of c3AdoHcy were found in cultures treated with c3Ado plus Hcy vs. c3Ado as single agent. The amounts of c3AdoHcy in cultures treated with c3Ado at concentrations initiating apoptosis did not exceed c3AdoHcy levels in cultures treated with c3Ado plus Hcy at concentrations not initiating apoptosis. Pretreatment of c3Ado plus Hcy cultures with c3Ari diminished c3AdoHcy and almost completely abrogated apoptosis. Exposure of cells to 100 microM c3Ari as single agent resulted in an increase in AdoHcy to 8.4-fold control but no changes in AdoMet and no initiation of apoptosis. Our findings indicate that c3Ado as single agent exerts its apoptosis-initiating effect through a nontransmethylase-related bio-chemical action. c3AdoHcy seems to be related to biochemical events initiating apoptotic cell death of HL-60 cells when c3Ado and Hcy are combined.


Organization of lin genes and IS6100 among different strains of hexachlorocyclohexane-degrading Sphingomonas paucimobilis: evidence for horizontal gene transfer.
The organization of lin genes and IS6100 was studied in three strains of Sphingomonas paucimobilis (B90A, Sp+, and UT26) which degraded hexachlorocyclohexane (HCH) isomers but which had been isolated at different geographical locations. DNA-DNA hybridization data revealed that most of the lin genes in these strains were associated with IS6100, an insertion sequence classified in the IS6 family and initially found in Mycobacterium fortuitum. Eleven, six, and five copies of IS6100 were detected in B90A, Sp+, and UT26, respectively. IS6100 elements in B90A were sequenced from five, one, and one regions of the genomes of B90A, Sp+, and UT26, respectively, and were found to be identical. DNA-DNA hybridization and DNA sequencing of cosmid clones also revealed that S. paucimobilis B90A contains three and two copies of linX and linA, respectively, compared to only one copy of these genes in strains Sp+ and UT26. Although the copy number and the sequence of the remaining genes of the HCH degradative pathway (linB, linC, linD, and linE) were nearly the same in all strains, there were striking differences in the organization of the linA genes as a result of replacement of portions of DNA sequences by IS6100, which gave them a strange mosaic configuration. Spontaneous deletion of linD and linE from B90A and of linA from Sp+ occurred and was associated either with deletion of a copy of IS6100 or changes in IS6100 profiles. The evidence gathered in this study, coupled with the observation that the G+C contents of the linA genes are lower than that of the remaining DNA sequence of S. paucimobilis, strongly suggests that all these strains acquired the linA gene through horizontal gene transfer mediated by IS6100. The association of IS6100 with the rest of the lin genes further suggests that IS6100 played a role in shaping the current lin gene organization.