Mitochondrial function after associating liver partition and portal vein ligation for staged hepatectomy in an experimental model.
Associating liver partition and portal vein ligation for staged hepatectomy (ALPPS) is a two-stage strategy to induce rapid regeneration of the remnant liver. The technique has been associated with high mortality and morbidity rates. This study aimed to evaluate mitochondrial function, biogenesis and morphology during ALPPS-induced liver regeneration. Male Wistar rats (n = 100) underwent portal vein ligation (PVL) or ALPPS. The animals were killed at 0 h (without operation), and 24, 48, 72 or 168 h after intervention. Regeneration rate and proliferation index were assessed. Mitochondrial oxygen consumption and adenosine 5'-triphosphate (ATP) production were measured. Mitochondrial biogenesis was evaluated by protein level measurements of peroxisome proliferator-activated receptor γ co-activator (PGC) 1-α, nuclear respiratory factor (NRF) 1 and 2, and mitochondrial transcription factor α. Mitochondrial morphology was evaluated by electron microscopy. Regeneration rate and Ki-67 index were significantly raised in the ALPPS group compared with the PVL group (regeneration rate at 168 h: mean(s.d.) 291·2(21·4) versus 245·1(13·8) per cent, P < 0·001; Ki-67 index at 24 h: 86·9(4·6) versus 66·2(4·9) per cent, P < 0·001). In the ALPPS group, mitochondrial function was impaired 48 h after the intervention compared with that in the PVL group (induced ATP production); (complex I: 361·9(72·3) versus 629·7(165·8) nmol per min per mg, P = 0·038; complex II: 517·5(48·8) versus 794·8(170·4) nmol per min per mg, P = 0·044). Markers of mitochondrial biogenesis were significantly lower 48 and 72 h after ALPPS compared with PVL (PGC1-α at 48 h: 0·61-fold decrease, P = 0·045; NRF1 at 48 h: 0·48-fold decrease, P = 0·028). Mitochondrial size decreased significantly after ALPPS (0·26(0·05) versus 0·40(0·07) μm2 ; P = 0·034). Impaired mitochondrial function and biogenesis, along with the rapid energy-demanding cell proliferation, may cause hepatocyte dysfunction after ALPPS. Surgical relevance Associating liver partition and portal vein ligation for staged hepatectomy (ALPPS) is a well known surgical strategy that combines liver partition and portal vein ligation. This method induces immense regeneration in the future liver remnant. The rapid volume increase is of benefit for resectability, but the mortality and morbidity rates of ALPPS are strikingly high. Moreover, lagging functional recovery of the remnant liver has been reported recently. In this translational study, ALPPS caused an overwhelming inflammatory response that interfered with the peroxisome proliferator-activated receptor γ co-activator 1-α-coordinated, stress-induced, mitochondrial biogenesis pathway. This resulted in the accumulation of immature and malfunctioning mitochondria in hepatocytes during the early phase of liver regeneration (bioenergetic destabilization). These findings might explain some of the high morbidity if confirmed in patients.


Age-Related Changes in Bimanual Instrument Playing with Rhythmic Cueing.
Deficits in bimanual coordination of older adults have been demonstrated to significantly limit their functioning in daily life. As a bimanual sensorimotor task, instrument playing has great potential for motor and cognitive training in advanced age. While the process of matching a person's repetitive movements to auditory rhythmic cueing during instrument playing was documented to involve motor and attentional control, investigation into whether the level of cognitive functioning influences the ability to rhythmically coordinate movement to an external beat in older populations is relatively limited. Therefore, the current study aimed to examine how timing accuracy during bimanual instrument playing with rhythmic cueing differed depending on the degree of participants' cognitive aging. Twenty one young adults, 20 healthy older adults, and 17 older adults with mild dementia participated in this study. Each participant tapped an electronic drum in time to the rhythmic cueing provided using both hands simultaneously and in alternation. During bimanual instrument playing with rhythmic cueing, mean and variability of synchronization errors were measured and compared across the groups and the tempo of cueing during each type of tapping task. Correlations of such timing parameters with cognitive measures were also analyzed. The results showed that the group factor resulted in significant differences in the synchronization errors-related parameters. During bimanual tapping tasks, cognitive decline resulted in differences in synchronization errors between younger adults and older adults with mild dimentia. Also, in terms of variability of synchronization errors, younger adults showed significant differences in maintaining timing performance from older adults with and without mild dementia, which may be attributed to decreased processing time for bimanual coordination due to aging. Significant correlations were observed between variability of synchronization errors and performance of cognitive tasks involving executive control and cognitive flexibility when asked for bimanual coordination in response to external timing cues at adjusted tempi. Also, significant correlations with cognitive measures were more prevalent in variability of synchronization errors during alternative tapping compared to simultaneous tapping. The current study supports that bimanual tapping may be predictive of cognitive processing of older adults. Also, tempo and type of movement required for instrument playing both involve cognitive and motor loads at different levels, and such variables could be important factors for determining the complexity of the task and the involved task requirements for interventions using instrument playing.


Reaction Mechanisms on [3 + 2] Cycloaddition of Azides with Metal Carbyne Complexes: Significant Effects of Aromaticity, Substituent, and Metal Center.
Density functional theory calculations were used to investigate the reaction mechanisms on [3 + 2] cycloaddition reactions of azides with metal carbyne complexes. Our results reveal that the formation of a 1,4-metallatriazole regioisomer is a kinetically favorable process in comparison with the formation of 1,5-metallatriazole. Aromaticity plays an important role in stabilizing the products in these reactions. Further analyses show that the electron-donating ligand on metal centers or the electron-withdrawing group on the azide could accelerate the [3 + 2] cycloaddition reaction. All of these findings could be useful for experimental chemists to develop "click reactions" in organometallic chemistry.


Learning about Biomolecular Solvation from Water in Protein Crystals.
Water occupies typically 50% of a protein crystal and thus significantly contributes to the diffraction signal in crystallography experiments. Separating its contribution from that of the protein is, however, challenging because most water molecules are not localized and are thus difficult to assign to specific density peaks. The intricateness of the protein-water interface compounds this difficulty. This information has, therefore, not often been used to study biomolecular solvation. Here, we develop a methodology to surmount in part this difficulty. More specifically, we compare the solvent structure obtained from diffraction data for which experimental phasing is available to that obtained from constrained molecular dynamics (MD) simulations. The resulting spatial density maps show that commonly used MD water models are only partially successful at reproducing the structural features of biomolecular solvation. The radial distribution of water is captured with only slightly higher accuracy than its angular distribution, and only a fraction of the water molecules assigned with high reliability to the crystal structure is recovered. These differences are likely due to shortcomings of both the water models and the protein force fields. Despite these limitations, we manage to infer protonation states of some of the side chains utilizing MD-derived densities.


Potential Cost-Effectiveness of RSV Vaccination of Infants and Pregnant Women in Turkey: An Illustration Based on Bursa Data.
Worldwide, respiratory syncytial virus (RSV) is considered to be the most important viral cause of respiratory morbidity and mortality among infants and young children. Although no active vaccine is available on the market yet, there are several active vaccine development programs in various stages. To assess whether one of these vaccines might be a future asset for national immunization programs, modeling the costs and benefits of various vaccination strategies is needed. To evaluate the potential cost-effectiveness of RSV vaccination of infants and/or pregnant women in Turkey. A multi-cohort static Markov model with cycles of one month was used to compare the cost-effectiveness of vaccinated cohorts versus non-vaccinated cohorts. The 2014 Turkish birth cohort was divided by twelve to construct twelve monthly birth cohorts of equal size (111,459 new-borns). Model input was based on clinical data from a multicenter prospective study from Bursa, Turkey, combined with figures from the (inter)national literature and publicly available data from the Turkish Statistical Institute (TÜÏK). Incremental cost-effectiveness ratios (ICERs) were expressed in Turkish Lira (TL) per quality-adjusted life year (QALY) gained. Vaccinating infants at 2 and 4 months of age would prevent 145,802 GP visits, 8,201 hospitalizations and 48 deaths during the first year of life, corresponding to a total gain of 1650 QALYs. The discounted ICER was estimated at 51,969 TL (26,220 US $ in 2013) per QALY gained. Vaccinating both pregnant women and infants would prevent more cases, but was less attractive from a pure economic point of view with a discounted ICER of 61,653 TL (31,106 US $ in 2013) per QALY. Vaccinating only during pregnancy would result in fewer cases prevented than infant vaccination and a less favorable ICER. RSV vaccination of infants and/or pregnant women has the potential to be cost-effective in Turkey. Although using relatively conservative assumptions, all evaluated strategies remained slightly below the threshold of 3 times the GDP per capita.


Sarcomere number adaptation after retinaculum transection in adult mice.
Skeletal muscle has been shown to adjust serial sarcomere number in response to chronic static length changes. However, the adaptive responses to alterations in the dynamic environment are less well defined. The adaptations of the adult mouse tibialis anterior (TA) muscle to altered length and excursion were investigated by surgical transection of the flexor retinaculum. TA moment arm and muscle excursion increased by 38 +/- 7% (mean +/- S.E.M.) and fully extended (plantarflexed) muscle length was decreased by 8% after flexor retinaculum transection. In spite of the significant shortening of the muscle in full plantar- and dorsiflexion, serial sarcomere number decreased by 10 +/- 1% after 2 weeks of recovery. Gait analysis of these transected animals revealed a 14 +/- 3% decrease in dorsiflexion angular velocity after transection. The decrease in angular velocity was less than the increase in moment arm and, as a result, muscle velocity was calculated to increase by 20 +/- 4%. These data suggested that the muscle adapted in response to the underlying change in length, irrespective of the altered excursion or velocity.


Functional SNPs of the breast cancer resistance protein-therapeutic effects and inhibitor development.
Breast cancer resistance protein (BCRP) is a half-molecule ATP-binding cassette transporter that pumps out various anticancer agents such as 7-ethyl-10-hydroxycamptothecin, topotecan and mitoxantrone. We have previously identified three polymorphisms within the BCRP gene, G34A (substituting Met for Val-12), C376T (substituting a stop codon for Gln-126) and C421A (substituting Lys for Gln-141). C421A BCRP-transfected murine fibroblast PA317 cells showed markedly decreased protein expression and low-level drug resistance when compared with wild-type BCRP-transfected cells. In contrast, G34A BCRP-transfected PA317 cells showed a similar protein expression and drug resistance profile to wild-type. The C376T polymorphism would be expected to have a considerable impact as active BCRP protein will not be expressed from a T376 allele. Hence, people with C376T and/or C421A polymorphisms may express low levels of BCRP, resulting in hypersensitivity of normal cells to BCRP-substrate anticancer agents. Estrogens, estrone and 17beta-estradiol, were previously found to restore drug sensitivity levels in BCRP-transduced cells by increasing the cellular accumulation of anticancer agents. BCRP transports sulfated estrogens but not free estrogens and in a series of screening experiments for synthesized and natural estrogenic compounds, several tamoxifen derivatives and phytoestrogens/flavonoids were identified that effectively circumvent BCRP-mediated drug resistance. The kinase inhibitors gefitinib and imatinib mesylate also interact with BCRP. Gefitinib, an inhibitor of epidermal growth factor receptor-tyrosine kinase, inhibits its transporter function and reverses BCRP-mediated drug resistance both in vitro and in vivo. BCRP-transfected human epidermoid carcinoma A431 cells and BCRP-transfected human non-small cell lung cancer PC-9 cells show gefitinib resistance. Imatinib, an inhibitor of BCR-ABL tyrosine kinase, also inhibits BCRP-mediated drug transport. Hence, both functional SNPs and inhibitors of BCRP reduce its transporter function and thus modulate substrate pharmacokinetics and pharmacodynamics.


New denoising scheme for magnetic resonance spectroscopy signals.
A new scheme for denoising magnetic resonance spectroscopy (MRS) signals is presented. This scheme is based on projecting noisy MRS signals in different domains, consecutively, and performing noise filtering operations in these domains. The domains are chosen such that the noise portion, which is inseparable from the desired signal in one domain, is separable in the other. A set of stable, linear, time-frequency (SLTF) transforms with different resolutions was selected for these projections as an example. Scheme evaluation was performed using extensive MRS signals with various noise levels. Compared with one domain denoising, it was observed that the proposed scheme gives superior results that compensate for the excess computational requirements. The proposed scheme supersedes also the wavelet packet denoising schemes.


Capturing sexual violence experiences among battered women using the revised sexual experiences survey and the revised conflict tactics scales.
The assessment of intimate partner sexual violence (IPSV) has garnered increased attention in recent years. However, uncertainty about which measure best captures experiences of IPSV remains. The present study focused on the direct comparison of two widely used measures of IPSV: the revised Sexual Experiences Survey (SES) and the revised Conflict Tactics Scales (CTS2). A secondary aim of the study was to extend the scope of IPSV acts by evaluating the presence of pornographic acts and experiences of forced sexual relations with other individuals. The current sample consisted of 138 battered women using the services of shelters. Results indicated that 79.7 % of women reported at least one incident of IPSV on either the CTS2 or the SES. The concordance rate between both measures was 76.8 %, with the highest concordance being for severe sexual violence. The Sexual Violence scale of the CTS2, which is more concise than the SES, identified 16.7 % more cases of IPSV. In addition, 26.1 % of women reported at least one incident involving pornography and 9.4 % had been forced to engage in sexual activities with other individuals. Women who reported experiences associated with pornography were 12-20 times more likely to be victims of severe sexual violence on the two measures. Such findings confirm the high prevalence of sexual violence among this population and indicate how rates can vary depending on the measures used. This study also underscores the relevance of investigating diverse types of violent acts to better understand how IPSV manifests itself.


From cellulose to kerogen: molecular simulation of a geological process.
The process by which organic matter decomposes deep underground to form petroleum and its underlying kerogen matrix has so far remained a no man's land to theoreticians, largely because of the geological (Myears) timescale associated with the process. Using reactive molecular dynamics and an accelerated simulation framework, the replica exchange molecular dynamics method, we simulate the full transformation of cellulose into kerogen and its associated fluid phase under prevailing geological conditions. We observe in sequence the fragmentation of the cellulose crystal and production of water, the development of an unsaturated aliphatic macromolecular phase and its aromatization. The composition of the solid residue along the maturation pathway strictly follows what is observed for natural type III kerogen and for artificially matured samples under confined conditions. After expulsion of the fluid phase, the obtained microporous kerogen possesses the structure, texture, density, porosity and stiffness observed for mature type III kerogen and a microporous carbon obtained by saccharose pyrolysis at low temperature. As expected for this variety of precursor, the main resulting hydrocarbon is methane. The present work thus demonstrates that molecular simulations can now be used to assess, almost quantitatively, such complex chemical processes as petrogenesis in fossil reservoirs and, more generally, the possible conversion of any natural product into bio-sourced materials and/or fuel.


Spectral characteristics of terahertz radiation from plasmonic photomixers.
We present a comprehensive analysis of spectral characteristics of terahertz radiation from plasmonic photomixers. We fabricate plasmonic photomixer prototypes with plasmonic contact electrode gratings on a low temperature grown GaAs substrate and characterize the spectral properties of the generated terahertz radiation by use of a heterodyne detection scheme. Our analysis shows that linewidth, stability, and frequency tuning range of the generated terahertz radiation are directly determined by linewidth, stability, and wavelength tuning range of optical pump beam and not affected by device geometry, substrate properties, optical pump power level and other operational settings. Our study indicates the crucial role of optical sources in realizing high performance terahertz spectroscopy and wireless communication systems based on plasmonic photomixers.


Recombination and pseudorecombination driving the evolution of the begomoviruses Tomato severe rugose virus (ToSRV) and Tomato rugose mosaic virus (ToRMV): two recombinant DNA-A components sharing the same DNA-B.
Begomoviruses are dicot-infecting, whitefly-transmitted viruses with a genome comprised of one or two molecules of circular, single-stranded DNA. In Brazil, tomato-infecting begomoviruses have emerged as serious pathogens since the introduction of a new biotype of the insect vector in the mid-1990's. Tomato rugose mosaic virus (ToRMV) and Tomato severe rugose virus (ToSRV) are often found in tomato fields. The complete sequence of the DNA-B components of ToSRV and ToRMV show an identity of 98.2%. Additionally, the high nucleotide identity (96.2%) between their common regions indicates that these two viruses may share the same DNA-B. Tomato seedlings were biolistically inoculated with ToSRV (DNA-A and DNA-B) and ToRMV (DNA-A and DNA-B) infectious clones in every possible combination of single or mixed infection. Symptom expression was evaluated for up to 35 days post-inoculation (dpi). DNA was extracted at 28 dpi and the presence of each viral genomic component was examined by rolling circle amplification (RCA) followed by digestion, as well as by quantitative, real-time PCR. Sequence comparisons, recombination and phylogenetic analyzes were performed using EMBOSS needle, RDP program and maximum likelihood inference, respectively. Symptoms in tomato plants inoculated with the different combinations of ToRMV and ToSRV DNA-A and DNA-B components consisted of a typical mosaic in all combinations. Pseudorecombinants were formed in all possible combinations. When two DNA-A or two DNA-B components were inoculated simultaneously, the ToRMV components were detected preferentially in relation to the ToSRV components. The combination of minor changes in both the Rep protein and the CR may be involved in the preferential replication of ToRMV components. Recombination and phylogenetic analyzes support the exchange of genetic material between ToRMV and ToSRV. ToRMV and ToSRV form viable pseudorecombinants in their natural host (Solanum lycopersicum) and share the same DNA-B. ToRMV DNA components are preferentially replicated over ToSRV components. These results indicate that the emergence of ToRMV involved both recombination and pseudorecombination, further highlighting the importance of these mechanisms in the emergence and adaptation of begomoviruses.


Optimal time for early laparoscopic cholecystectomy for acute cholecystitis.
There is growing evidence in support of performing early laparoscopic cholecystectomy (LC) for acute cholecystitis. However, the definition of early LC varies from 0 through 10 days depending on the research protocol. The optimum time to perform early LC is still unclear. To determine whether outcomes after early LC for acute cholecystitis vary depending on time from presentation to surgery and to determine the optimum time to perform LC for acute cholecystitis. We performed a retrospective review of prospectively collected data from the Nationwide Inpatient Sample (NIS) for 2005 through 2009. The population-based sample included 95,523 adults (18 years and older) who underwent LC within 10 days of presentation for acute cholecystitis. Patients were categorized and analyzed in 2 ways based on length of time from presentation to surgery. First, patients were categorized into 3 groups: 0 through 1 day, 2 through 5 days, and 6 through 10 days. Second, we compared outcomes for each incremental preoperative day (days 0-5). Outcomes of interest were mortality, length of stay, complications, and cost. Propensity score matching and generalized linear modeling were used. The hypothesis being tested was formulated after data collection was complete. A total of 95,523 patients were selected. After matching the 3 groups based on propensity scores, patients who underwent surgery during days 2 through 5 and days 6 through 10 had increasingly worse outcomes when compared with those undergoing surgery on days 0 through 1. The odds of mortality were 1.26 (95% CI, 1.00-1.58) and 1.93 (95% CI, 1.38-2.68), and the odds of postoperative infections were 0.88 (95% CI, 0.69-1.12) and 1.53 (95% CI, 1.05-2.23) for days 2 through 5 and days 6 through 10, respectively. Adjusted mean hospital cost increased from $8974 (days 0-1) to $17,745 (days 6-10). Analysis by each incremental day revealed the optimal time of surgery to be within the first 48 hours of presentation. Laparoscopic cholecystectomy performed within 2 days of presentation of acute cholecystitis yielded the best outcomes and lowest costs. Although causality could not be established, delaying LC was associated with more complications, higher mortality, and higher costs.


Effectiveness of primary monocanalicularnasal intubation with Monoka tubes and nasal endoscopic findings for congenital nasolacrimal duct obstruction with enlarged lacrimal sac and chronic dacryocystitis.
We report successful outcomes of monocanalicularnasal intubation with Monoka tubes and endoscopic findings for Hasner valve area. This method is presented as a primary treatment modality in congenital nasolacrimal duct obstruction (CNLDO) with an enlarged lacrimal sac and chronic dacryocystitis. In a prospective noncomparative consecutive case series, 23 eyes of 18 patients with CNLDO with an enlarged lacrimal sac and chronic dacryocystitis underwent endoscopic probing and monocanalicularnasal intubation under general anesthesia. Demographic information, including age, sex, duration of preoperative symptoms, operative endoscopic findings, timing of silicone tube removal, follow-up periods, complications, and outcomes, was analyzed. The study included 23 eyes of 18 patients. Mean (SD) age was 19.7 (12.2) months (range, 9-48 mo). Under nasal endoscopic view, the probe tip protruded through a balloon-like nasal mucosa around the Hasner valve. In all cases, tubes were removed in an office setting under topical anesthesia. Mean follow-up period was 24.6 (5.4) weeks. Complications, such as punctal slitting and corneal abrasion, were not observed. All patients showed successful results until the final follow-up. In cases of CNLDO with an enlarged lacrimal sac and chronic dacryocystitis, monocanalicular intubation is recommended rather than simple probing. Monoka tube intubation has the advantages of technical ease of insertion and tube removal. Moreover, the tubing does not threaten the unprobed part of the lacrimal drainage system. Endoscopic findings revealed that the probe tip protruded through a balloon-like nasal mucosa with pus around the Hasner valve.


Getting there is half the battle: recent advances in delivering therapeutics.
Delivery of therapeutic molecules at the right time, place and at the correct dosage is critical to improve the effectiveness of therapeutic regimens. Barriers in the body, normally needed to maintain function, often impede delivery of therapeutic payloads to their target areas. Designing innovative solutions to circumvent these environmental factors, and ensure the timely delivery of therapeutic doses, is an essential element in improving human health. Here we highlight recent studies that focus on bypassing different barriers crucial for improving therapeutic delivery, by temporarily modifying the in vivo microenvironment, re-designing therapeutic carrier vehicles to improve control characteristics and on-demand delivery, and developing convenience-based strategies to improve patient compliance and access to therapeutics.


Klebsiella quasipneumoniae Provides a Window into Carbapenemase Gene Transfer, Plasmid Rearrangements, and Patient Interactions with the Hospital Environment.
Several emerging pathogens have arisen as a result of selection pressures exerted by modern health care. Klebsiella quasipneumoniae was recently defined as a new species, yet its prevalence, niche, and propensity to acquire antimicrobial resistance genes are not fully described. We have been tracking inter- and intraspecies transmission of the Klebsiella pneumoniae carbapenemase (KPC) gene, blaKPC, between bacteria isolated from a single institution. We applied a combination of Illumina and PacBio whole-genome sequencing to identify and compare K. quasipneumoniae from patients and the hospital environment over 10- and 5-year periods, respectively. There were 32 blaKPC-positive K. quasipneumoniae isolates, all of which were identified as K. pneumoniae in the clinical microbiology laboratory, from 8 patients and 11 sink drains, with evidence for seven separate blaKPC plasmid acquisitions. Analysis of a single subclade of K. quasipneumoniae subsp. quasipneumoniae (n = 23 isolates) from three patients and six rooms demonstrated seeding of a sink by a patient, subsequent persistence of the strain in the hospital environment, and then possible transmission to another patient. Longitudinal analysis of this strain demonstrated the acquisition of two unique blaKPC plasmids and then subsequent within-strain genetic rearrangement through transposition and homologous recombination. Our analysis highlights the apparent molecular propensity of K. quasipneumoniae to persist in the environment as well as acquire carbapenemase plasmids from other species and enabled an assessment of the genetic rearrangements which may facilitate horizontal transmission of carbapenemases.


Survey of Phortica drosophilid flies within and outside of a recently identified transmission area of the eye worm Thelazia callipaeda in Switzerland.
Phortica drosophilid flies are the intermediate hosts and vectors of the eye worm Thelazia callipaeda. This nematode originates from Asia and was first detected in southern Europe in 1989. The aim of the study was to assess the presence and the population dynamics of Phortica flies in a recently discovered new endemic area (Ticino, Southern Switzerland, south of the Alps) of T. callipaeda (site 1), at its border (site 2), at higher altitudes (beyond 1100 meters above sea level) within (site 3) or outside (site 4) the endemic area, and in a site north of the Alps (site 5). Flies were captured using two types of fruit-baited traps, the bait being changed once per week, and by netting around the eyes of a dog and human. A total of 1695 Phortica flies were collected. One of the fruit-baited traps, which can easily be assembled with cheap components, was found to be efficient for catching Phortica spp. At site 1, 644 such flies were collected with this trap during 34 weekly catches from April to October. The number of flies caught was highest at site 2 (n=903) and it was significantly lower (n=36) at site 5 north of the Alps. Virtually no Phortica at all were caught at higher altitudes (sites 3 and 4). Females were all in all predominant in the traps, accounting for 72.6% of Phortica flies (1150/1584), although males became dominant late in the season (male/female ratio 1.26 in October). In contrast, 80.2% of Phortica flies collected around the eyes of dog and human baits by netting (n=111) were males. No female at all was captured by netting until September. PCR for T. callipaeda was negative with all Phortica flies. Morphological examination of the 523 male flies based on features of the eye margin and the number of particular genital sensilla identified 89.1% P. semivirgo, 5.7% P. variegata but also 5.2% intermediate forms. Genetic analyses of partial mitochondrial cox1 and rDNA internal transcribed spacer 1 sequences revealed that these three morphotypes were genetically not distinguishable. This study confirms the presence of Phortica spp. north to the Alps and therefore the potential risk of T. callipaeda infection outside the currently known endemic region, depending on local abundance and longevity of the drosophilid vectors.


Arteriolosclerosis that affects multiple brain regions is linked to hippocampal sclerosis of ageing.
Hippocampal sclerosis of ageing is a prevalent brain disease that afflicts older persons and has been linked with cerebrovascular pathology. Arteriolosclerosis is a subtype of cerebrovascular pathology characterized by concentrically thickened arterioles. Here we report data from multiple large autopsy series (University of Kentucky Alzheimer's Disease Centre, Nun Study, and National Alzheimer's Coordinating Centre) showing a specific association between hippocampal sclerosis of ageing pathology and arteriolosclerosis. The present analyses incorporate 226 cases of autopsy-proven hippocampal sclerosis of ageing and 1792 controls. Case-control comparisons were performed including digital pathological assessments for detailed analyses of blood vessel morphology. We found no evidence of associations between hippocampal sclerosis of ageing pathology and lacunar infarcts, large infarcts, Circle of Willis atherosclerosis, or cerebral amyloid angiopathy. Individuals with hippocampal sclerosis of ageing pathology did not show increased rates of clinically documented hypertension, diabetes, or other cardiac risk factors. The correlation between arteriolosclerosis and hippocampal sclerosis of ageing pathology was strong in multiple brain regions outside of the hippocampus. For example, the presence of arteriolosclerosis in the frontal cortex (Brodmann area 9) was strongly associated with hippocampal sclerosis of ageing pathology (P < 0.001). This enables informative evaluation of anatomical regions outside of the hippocampus. To assess the morphology of brain microvasculature far more rigorously than what is possible using semi-quantitative pathological scoring, we applied digital pathological (Aperio ScanScope) methods on a subsample of frontal cortex sections from hippocampal sclerosis of ageing (n = 15) and control (n = 42) cases. Following technical studies to optimize immunostaining methods for small blood vessel visualization, our analyses focused on sections immunostained for smooth muscle actin (a marker of arterioles) and CD34 (an endothelial marker), with separate analyses on grey and white matter. A total of 43 834 smooth muscle actin-positive vascular profiles and 603 798 CD34-positive vascular profiles were evaluated. In frontal cortex of cases with hippocampal sclerosis of ageing, smooth muscle actin-immunoreactive arterioles had thicker walls (P < 0.05), larger perimeters (P < 0.03), and larger vessel areas (P < 0.03) than controls. Unlike the arterioles, CD34-immunoreactive capillaries had dimensions that were unchanged in cases with hippocampal sclerosis of ageing versus controls. Arteriolosclerosis appears specific to hippocampal sclerosis of ageing brains, because brains with Alzheimer's disease pathology did not show the same morphological alterations. We conclude that there may be a pathogenetic change in aged human brain arterioles that impacts multiple brain areas and contributes to hippocampal sclerosis of ageing.


Inhibition of entry of HIV-1 in neural cell lines by antibodies against galactosyl ceramide.
Although the CD4 molecule is the principal cellular receptor for the human immunodeficiency virus (HIV), several CD4-negative cell lines are susceptible to infection with one or more HIV strains. These findings indicate that there are alternate modes of viral entry, perhaps involving one or more receptor molecules. Antibodies against galactosyl ceramide (galactocerebroside, or GalC) inhibited viral internalization and infection in two CD4-negative cell lines derived from the nervous system: U373-MG and SK-N-MC. Furthermore, recombinant HIV surface glycoprotein gp120 bound to GalC but not to other glycolipids. These results suggest a role for GalC or a highly related molecule in HIV entry into neural cells.


Headache and symptoms of temporomandibular disorder: an epidemiological study.
A population-based cross-sectional study was conducted to estimate the prevalence of migraine, episodic tension-type headaches (ETTH), and chronic daily headaches (CDH), as well as the presence of symptoms of temporomandibular disorders (TMD) in the adult population. The potential comorbidity of headache syndromes and TMD has been established mostly based on clinic-based studies. A representative sample of 1230 inhabitants (51.5% women) was interviewed by a validated phone survey. TMD symptoms were assessed through 5 questions, as recommended by the American Academy of Orofacial Pain, in an attempt to classify possible TMD. Primary headaches were diagnosed based on the International Classification of Headache Disorders. When at least 1 TMD symptom was reported, any headache happened in 56.5% vs 31.9% (P < .0001) in those with no symptoms. For 2 symptoms, figures were 65.1% vs 36.3% (P < .0001); for 3 or more symptoms, the difference was even more pronounced: 72.8% vs 37.9%. (P < .0001). Taking individuals without headache as the reference, the prevalence of at least 1 TMD symptom was increased in ETTH (prevalence ratio = 1.48, 95% confidence interval = 1.20-1.79), migraine (2.10, 1.80-2.47) and CDH (2.41, 1.84-3.17). At least 2 TMD symptoms also happened more frequently in migraine (4.4, 3.0-6.3), CDH (3.4; 1.5-7.6), and ETTH (2.1; 1.3-3.2), relative to individuals with no headaches. Finally, 3 or more TMD symptoms were also more common in migraine (6.2; 3.8-10.2) than in no headaches. Differences were significant for ETTH (2.7 1.5-4.8), and were numerically but not significant for CDH (2.3; 0.66-8.04). Temporomandibular disorder symptoms are more common in migraine, ETTH, and CDH relative to individuals without headache. Magnitude of association is higher for migraine. Future studies should clarify the nature of the relationship.


Human thrombopoietin knockin mice efficiently support human hematopoiesis in vivo.
Hematopoietic stem cells (HSCs) both self-renew and give rise to all blood cells for the lifetime of an individual. Xenogeneic mouse models are broadly used to study human hematopoietic stem and progenitor cell biology in vivo. However, maintenance, differentiation, and function of human hematopoietic cells are suboptimal in these hosts. Thrombopoietin (TPO) has been demonstrated as a crucial cytokine supporting maintenance and self-renewal of HSCs. We generated RAG2(-/-)γ(c)(-/-) mice in which we replaced the gene encoding mouse TPO by its human homolog. Homozygous humanization of TPO led to increased levels of human engraftment in the bone marrow of the hosts, and multilineage differentiation of hematopoietic cells was improved, with an increased ratio of myelomonocytic verus lymphoid lineages. Moreover, maintenance of human stem and progenitor cells was improved, as demonstrated by serial transplantation. Therefore, RAG2(-/-)γ(c)(-/-) TPO-humanized mice represent a useful model to study human hematopoiesis in vivo.


Early effects of transcatheter aortic valve implantation and aortic valve replacement on myocardial function and aortic valve hemodynamics: insights from cardiovascular magnetic resonance imaging.
There remains a paucity of mechanistic data on the effect of transcatheter aortic valve implantation (TAVI) on early left and right ventricular function and quantitative aortic valve regurgitation. We sought to assess and compare the early effects on myocardial function and aortic valve hemodynamics of TAVI and aortic valve replacement (AVR) using serial cardiovascular magnetic resonance (CMR) imaging and echocardiography. A prospective comparison study of 47 patients with severe aortic stenosis undergoing either TAVI (n = 26) or high-risk AVR (n = 21). CMR (for left ventricle/right ventricle function, left ventricular mass, left atrial volume, and aortic regurgitation) was carried out before the procedure and early postprocedure (<14 days). Groups were similar with respect to Society of Thoracic Surgeons score (TAVI, 7.7 vs AVR, 5.9; P = .11). Preoperative left ventricular (TAVI, 69% ± 13% vs AVR, 73% ± 10%; P = .10) and right ventricular (TAVI, 61% ± 11% vs AVR, 59% ± 8%; P = .5) ejection fractions were similar. Postoperative left ventricular ejection fraction was preserved in both groups. In contrast, decline in right ventricular ejection fraction was more significant in the TAVI group (61%-54% vs 59%-58%; P = .01). Postprocedure aortic regurgitant fraction was significantly greater in the TAVI group (16% vs 4%; P = .001), as was left atrial size (110 vs 84 mL; P = .02). Further analysis revealed a significant relationship between the increased aortic regurgitant fraction and greater left atrial size (P = .006), and a trend toward association between the decline in right ventricle dysfunction and increased postprocedure aortic regurgitation (P = .08). There was no significant difference in early left ventricular systolic function between techniques. Whereas right ventricle systolic function was preserved in the AVR group, it was significantly impaired early after TAVI, possibly reflecting a clinically important pathophysiologic consequence of paravalvular aortic regurgitation.


Physical interaction between retinoic acid receptor and Sp1: mechanism for induction of urokinase by retinoic acid.
Induction of urokinase plasminogen activator (uPA) by retinoic acid (RA) is the initial event preceding certain subsequent biological changes in vascular endothelial cells. We investigated the molecular mechanism by which RA stimulates the expression of uPA, which lacks a canonical RA receptor (RAR)-responsive element, in bovine and human aortic endothelial cells. Upon stimulation with RA, mRNA levels of RARalpha and beta transiently increased in parallel with the induction of uPA, and this increase was inhibited by cycloheximide. Results of transient transfection of RAR/RXR cDNAs and experiments using specific agonists and antagonists suggested that uPA induction is dependent upon RAR (initially, RARalpha) with the help of RXRalpha. Deletion analysis of the uPA promoter suggested that RAR/RXR acts on GC box region within the uPA promoter. This was further supported by inhibition of Sp1 binding to this region. Coimmunoprecipitation studies, glutathione S-transferase pull-down experiment, and mammalian two-hybrid assays suggested a physical interaction between RAR/RXR and Sp1. Furthermore, gel shift studies showed that the binding of Sp1 to the uPA GC box is significantly potentiated in the presence of RARs/RXRs. Finally, Sp1 and RAR/RXR synergistically enhanced the transactivation activity of the uPA promoter. These results suggest that (1) RA induces RARs mainly via RARalpha and that (2) RAR/RXR physically and functionally interact with Sp1, resulting in a potentiation of uPA transcription.


Longitudinal associations between activity and cognition vary by age, activity type, and cognitive domain.
The demonstration of correlated change is critical to understanding the relationship between activity engagement and cognitive functioning in older adulthood. Changes in activity have been shown to be related to changes in cognition, but little attention has been devoted to how this relationship may vary between specific activity types, cognitive domains, and age groups. Participants initially aged 65-98 years (M = 77.46 years) from the Australian Longitudinal Study of Ageing (n = 1,321) completed measurements of activity (i.e., cognitive, group social, one-on-one social, and physical) and cognition (i.e., perceptual speed, and immediate and delayed episodic memory) at baseline, 2, 8, 11, and 15 years later. Bivariate latent growth curve models covarying for education, sex, and baseline age and medical conditions revealed multiple positive-level relations between activity and cognitive performance, but activity level was not related to later cognitive change. Change in perceptual speed over 15 years was positively associated with change in cognitive activity, and change in immediate episodic memory was positively associated with change in one-on-one social activity. Old-old adults showed a stronger change-change covariance for mentally stimulating activity in relation to perceptual speed than did young-old adults. The differentiation by activity type, cognitive domain, and age contributes to the growing evidence that there is variation in the way cognitive ability at different ages is related to activity.


Antioxidant diet supplementation influences blood iron status in endurance athletes.
The aim of this work was to check the effects of antioxidant supplementation (vitamins E and C, and beta-carotene) on the basal iron status of athletes prior to and following their training and competition season (3 months). Eighteen amateur trained male athletes were randomly distributed in 2 groups: placebo (lactose) and antioxidant supplemented (vitamin E, 500 mg/d; vitamin C, 1 g/d; and beta -carotene, 30 mg/d). The study was double blind. Hematological parameters, dietary intake, physical activity intensity, antioxidant status (GSH/GSSG ratio), and basal iron status (serum iron, transferrin, ferritin, and iron saturation index) were determined before and after the intervention trials. Exercise decreased antioxidant defenses in the placebo group but not in the antioxidant-supplemented group. No changes were found in the number of erythrocytes, hematocrit, or hemoglobin concentration, or in values of serum iron parameters, after taking the antioxidant cocktail for 3 months, in spite of the exercise completed. The placebo group showed a high oxidative stress index, and decreases in serum iron (24%) and iron saturation index (28%), which can neither be attributed to aspects of the athletes' usual diet, nor to hemoconcentration. Antioxidant supplementation prevents the decrease of serum iron and the iron saturation index, and a link between iron metabolism and oxidative stress may also be suggested.


A Mechatronic System for Studying Energy Optimization During Walking.
A general principle of human movement is that our nervous system is able to learn optimal coordination strategies. However, how our nervous system performs this optimization is not well understood. Here we design, build, and test a mechatronic system to probe the algorithms underlying the optimization of energetic cost in walking. The system applies controlled fore-aft forces to a hip-belt worn by a user, decreasing their energetic cost by pulling forward, or increasing it by pulling backward. The system controls the forces, and thus energetic cost as a function of how the user is moving. In testing, we found that the system can quickly, accurately, and precisely apply target forces within a walking step. We next controlled the forces as a function of the user's step frequency and found that we could predictably reshape their energetic cost landscape. Finally, we tested whether users adapted their walking in response to the new cost landscapes created by our system, and found that users shifted their step frequency toward the new energetic minima. Our system design appears to be effective for reshaping energetic cost landscapes in human walking to study how the nervous system optimizes movement.


Effect of novel restoration techniques on the fracture resistance of teeth treated endodontically: An in vitro study.
The aim was to evaluate the effects of fiber-reinforced composite restorations and a bulk-fill resin composite on the fracture strength of mandibular premolars treated endodontically. Standard mesio-occluso-distal (MOD) cavities were prepared in 48 mandibular premolars. Following root canal treatment, teeth were assigned to four groups: Group 1, nano-hybrid resin composite; Group 2, polyethylene woven fiber plus nano-hybrid resin composite; Group 3, short fiber-reinforced resin composite plus nano-hybrid resin composite; and Group 4, bulk-fill resin composite plus nano-hybrid resin composite. Then, the teeth were subjected to the fracture toughness test. The data were analyzed statistically using one-way ANOVA, followed by Tukey's post-hoc test. The fiber-reinforced groups had better results than the nano-hybrid and bulk-fill composites (p<0.05), while the bulk-fill and nano-hybrid composite restorations gave similar results (p>0.05). Fiber-reinforcement improved the fracture strength of teeth with large MOD cavities treated endodontically. Bulk-fill composites can be used reliably as well as nano-hybrid composites.


Involvement of the multilineage CD38 molecule in a unique pathway of cell activation and proliferation.
We report clear evidence that the interaction of the CD38 molecule with the specific mAb A10 on normal human cells and lines modulates the expression of surface activation markers relevant to T, NK, and plasma cell biology and functions. Moreover A10 mAb binding is followed by proliferation effects on all the target cells analyzed, and the phenomenon is accessory cell and IL-2 dependent. The effects of A10 mAb synergizing both CD2 and CD3 activation pathways indicate that CD38 signal transduction mechanism(s) are apparently different from the aforementioned. Nevertheless the decreased A10-driven proliferation after CD3-Ti modulation suggests a possible functional interdependence between these activation pathways. Taken together, the results indicate that the CD38 molecule might play a physiologic role in T, NK, and plasma cell regulation.


Automated Machine Learning: A Case Study of Genomic "Image-Based" Prediction in Maize Hybrids.
Machine learning methods such as multilayer perceptrons (MLP) and Convolutional Neural Networks (CNN) have emerged as promising methods for genomic prediction (GP). In this context, we assess the performance of MLP and CNN on regression and classification tasks in a case study with maize hybrids. The genomic information was provided to the MLP as a relationship matrix and to the CNN as "genomic images." In the regression task, the machine learning models were compared along with GBLUP. Under the classification task, MLP and CNN were compared. In this case, the traits (plant height and grain yield) were discretized in such a way to create balanced (moderate selection intensity) and unbalanced (extreme selection intensity) datasets for further evaluations. An automatic hyperparameter search for MLP and CNN was performed, and the best models were reported. For both task types, several metrics were calculated under a validation scheme to assess the effect of the prediction method and other variables. Overall, MLP and CNN presented competitive results to GBLUP. Also, we bring new insights on automated machine learning for genomic prediction and its implications to plant breeding.


miRNA regulation of liver growth after 50% partial hepatectomy and small size grafts in rats.
The molecular mechanisms underlying the growth of small size grafts and the remaining livers are poorly understood. MicroRNAs (miRNAs) negatively modulate expression of genes that are involved in cellular function and metabolism. The aim of this study is to identify critical miRNA species that modulate the growth of small grafts and the remaining livers after partial hepatectomy (PH). Small size graft liver transplantation was performed in rats. Liver tissue was harvested after transplant or PH for the determination of miRNA expression profile, and the data were confirmed by quantitative reverse-transcriptase polymerase chain reaction. The genes involved in cell cycle and proliferation were analyzed by quantitative reverse-transcriptase polymerase chain reaction and immunohistochemical staining. Compared with control liver, miR_122a, Let_7b, and miR_26a were reduced by more than 90% in 45% volume grafts. In the remaining livers after 50% PH, 30 miRNAs were down-regulated by more than 50%, and among them, miR_22a, miR_26a, miR_30b, Let_7f, and Let_7g were markedly decreased. A negative correlation existed between down-regulated miRNAs and highly up-regulated genes involved in cell cycle and proliferation in the remaining livers. Moreover, overexpression of miR_26a markedly down-regulated cyclin E2 protein levels and significantly decreased proliferation of HepG2 cells. Down-regulated miRNAs play a pivotal role in promoting the growth of small size grafts and the remaining livers. The negative correlation between down-regulated miRNAs and up-regulated genes suggests that these specific miRNAs participate in the modulation of a growth response in both living donors and small size graft recipients.


Are there causal relationships between the development of the inflammatory diseases amyotrophic lateral sclerosis and asthma?
Amyotrophic lateral sclerosis (ALS) and asthma are inflammatory diseases. ALS is a fatal progressive, neurodegenerative disease with inflammation around the upper and lower motor neurons leading to their degeneration, muscle atrophy, paralysis, and death. Asthma is a chronic inflammatory disease with reversible airway obstruction and nonspecific airway hyper-reactivity. The local release of sensory neuropeptides from capsaicin-sensitive primary afferents causes motor neuron pathophysiology and airway inflammation and hyper-reactivity. While there is no cure for ALS, asthma is managed according to its symptoms and severity, to decrease the symptoms, improve pulmonary function, and reduce morbidity. To determine whether understanding asthma may provide insights into how to clinically deal with ALS, the authors examined the etiologies of ALS and asthma, and the factors that exacerbate the symptoms. Although no direct correlations were found, the similar multifactorial triggers, and the critical roles of neuronal inflammation, suggest that one or more exists.


PERTURBATION AND SCALED COOK'S DISTANCE.
Cook's (Cook, 1977) distance is one of the most important diagnostic tools for detecting influential individual or subsets of observations in linear regression for cross-sectional data. However, for many complex data structures (e.g., longitudinal data), no rigorous approach has been developed to address a fundamental issue: deleting subsets with different numbers of observations introduces different degrees of perturbation to the current model fitted to the data and the magnitude of Cook's distance is associated with the degree of the perturbation. The aim of this paper is to address this issue in general parametric models with complex data structures. We propose a new quantity for measuring the degree of the perturbation introduced by deleting a subset. We use stochastic ordering to quantify the stochastic relationship between the degree of the perturbation and the magnitude of Cook's distance. We develop several scaled Cook's distances to resolve the comparison of Cook's distance for different subset deletions. Theoretical and numerical examples are examined to highlight the broad spectrum of applications of these scaled Cook's distances in a formal influence analysis.


Determining the influence of rainfall patterns and carbendazim on the surface activity of the earthworm Lumbricus terrestris.
Carbendazim is highly toxic to earthworms and is used as a standard control substance when running field-based trials of pesticides, but results using carbendazim are highly variable. In the present study, impacts of timing of rainfall events following carbendazim application on earthworms were investigated. Lumbricus terrestris were maintained in soil columns to which carbendazim and then deionized water (a rainfall substitute) were applied. Carbendazim was applied at 4 kg/ha, the rate recommended in pesticide field trials. Three rainfall regimes were investigated: initial and delayed heavy rainfall 24 h and 6 d after carbendazim application, and frequent rainfall every 48 h. Earthworm mortality and movement of carbendazim through the soil was assessed 14 d after carbendazim application. No detectable movement of carbendazim occurred through the soil in any of the treatments or controls. Mortality in the initial heavy and frequent rainfall was significantly higher (approximately 55%) than in the delayed rainfall treatment (approximately 25%). This was due to reduced bioavailability of carbendazim in the latter treatment due to a prolonged period of sorption of carbendazim to soil particles before rainfall events. The impact of carbendazim application on earthworm surface activity was assessed using video cameras. Carbendazim applications significantly reduced surface activity due to avoidance behavior of the earthworms. Surface activity reductions were least in the delayed rainfall treatment due to the reduced bioavailability of the carbendazim. The nature of rainfall events' impacts on the response of earthworms to carbendazim applications, and details of rainfall events preceding and following applications during field trials should be made at a higher level of resolution than is currently practiced according to standard International Organization for Standardization protocols.


Retrovirus in salivary glands from patients with Sjögren's syndrome.
To investigate the possibility of an immune response to retroviral antigens or of detecting retrovirus in Sjögren's syndrome. Retroviruses were sought in labial salivary glands and peripheral blood mononuclear cells from patients with Sjögren's syndrome by immunoblotting assay, immunohistochemical assay, polymerase chain reaction (PCR), reverse transcriptase (RT) activity assay, and transmission electron microscopy. Sera from five of 15 patients with Sjögren's syndrome (33%) reacted against p24 group specific antigen (gag) of human immunodeficiency virus (HIV). Labial salivary gland biopsy specimens from seven of the 15 patients with Sjögren's syndrome (47%) contained an epithelial cytoplasmic protein reactive with a monoclonal antibody to p24 of HIV. PCR was performed to detect HIV and human T lymphotropic virus type I (HTLV-I) genes from salivary gland tissues and peripheral blood mononuclear cells from patients with Sjögren's syndrome. Mn2+ dependent, Mg2+ independent RT activity was detected in the salivary gland tissues in three of 10 patients. A-type-like retroviral particles were observed in epithelial cells of salivary glands by transmission electron microscopy. Target genes for HIV and HTLV-I were not found in any of the salivary gland tissues or peripheral blood mononuclear cells from Sjögren's syndrome patients. The data suggest the presence of an unknown retrovirus similar to HIV in the salivary gland which might be involved in the pathogenesis of a subpopulation in Sjögren's syndrome.


Neural correlates of metacontrast masking across different contrast polarities.
Metacontrast masking is a powerful illusion to investigate the dynamics of perceptual processing and to control conscious visual perception. However, the neural mechanisms underlying this fundamental investigative tool are still debated. In the present study, we examined metacontrast masking across different contrast polarities by employing a contour discrimination task combined with EEG (Electroencephalography). When the target and mask had the same contrast polarity, a typical U-shaped metacontrast function was observed. A change in mask polarity (i.e., opposite mask polarity) shifted this masking function to a monotonic increasing function such that the target visibility was strongly suppressed at stimulus onset asynchronies less than 50 ms. This transition in metacontrast function has been typically interpreted as an increase in intrachannel inhibition of the sustained activities functionally linked to object visibility and identity. Our EEG analyses revealed an early (160-300 ms) and a late (300-550 ms) spatiotemporal cluster associated with this effect of polarity. The early cluster was mainly over occipital and parieto-occipital scalp sites. On the other hand, the later modulations of the evoked activities were centered over parietal and centro-parietal sites. Since both of these clusters were beyond 160 ms, the EEG results point to late recurrent inhibitory mechanisms. Although the findings here do not directly preclude other proposed mechanisms for metacontrast, they highlight the involvement of recurrent intrachannel inhibition in metacontrast masking.


WE-G-213CD-03: A Dual Complementary Verification Method for Dynamic Tumor Tracking on Vero SBRT.
to use complementary cine EPID and gimbals log file analysis for in-vivo tracking accuracy monitoring. A clinical prototype of dynamic tracking (DT) was installed on the Vero SBRT system. This prototype version allowed tumor tracking by gimballed linac rotations using an internal-external correspondence model. The DT prototype software allowed the detailed logging of all applied gimbals rotations during tracking. The integration of an EPID on the vero system allowed the acquisition of cine EPID images during DT. We quantified the tracking error on cine EPID (E-EPID) by subtracting the target center (fiducial marker detection) and the field centroid. Dynamic gimbals log file information was combined with orthogonal x-ray verification images to calculate the in-vivo tracking error (E-kVLog). The correlation between E-kVLog and E-EPID was calculated for validation of the gimbals log file. Further, we investigated the sensitivity of the log file tracking error by introducing predefined systematic tracking errors. As an application we calculate gimbals log file tracking error for dynamic hidden target tests to investigate gravity effects and decoupled gimbals rotation from gantry rotation. Finally, calculating complementary cine EPID and log file tracking errors evaluated the clinical accuracy of dynamic tracking. A strong correlation was found between log file and cine EPID tracking error distribution during concurrent measurements (R=0.98). We found sensitivity in the gimbals log files to detect a systematic tracking error up to 0.5 mm. Dynamic hidden target tests showed no gravity influence on tracking performance and high degree of decoupled gimbals and gantry rotation during dynamic arc dynamic tracking. A submillimetric agreement between clinical complementary tracking error measurements was found. Redundancy of the internal gimbals log file with x-ray verification images with complementary independent cine EPID images was implemented to monitor the accuracy of gimballed tumor tracking on Vero SBRT. Research was financially supported by the Flemish government (FWO), Hercules Foundation and BrainLAB AG.


Congenital anomalies: Impact of prenatal diagnosis on mode of delivery.
An important aspect of prenatal diagnosis is the avoidance of emergency caesarean delivery (CD) where the abnormality is considered lethal and the infant will not survive. A consecutive cohort of 211,163 women delivered of infants weighing 500 grams or more in three tertiary referral centers from 01/95 to 12/04, was analyzed for perinatal death attributed to congenital malformations. In the group that died in the neonatal period, the emergency CD rate was significantly lower where anomaly was detected versus undetected (17.5% versus 31%). Further, in contrast to undiagnosed anomalies, the indication for emergency CD was more often maternal in the diagnosed group (42% versus 19%, p=0.019). When a diagnosis of lethal congenital anomaly has been made in the prenatal period, the reduction in the emergency CD rate by almost half in this study supports a pivotal role for prenatal diagnosis in optimizing maternal care.


Decreased anionic sites in Bruch's membrane of spontaneous and drug-induced diabetes.
The basal laminae of capillaries, glomeruli, and nerves are thickened in diabetes. Previous studies have shown that diabetic tissues produce increased levels of basal lamina collagen and reduced levels of proteoglycans. Since heparan sulfate and chondroitin sulfate proteoglycans are thought to act as anionic barriers regulating passage of proteins across Bruch's membrane of the eye, the cationic electron microscope tracers, polyethyleneimine (PEI) and ruthenium red, were used to study the distribution of anionic sites in Bruch's membrane of spontaneously diabetic Bio-Breeding/Worcester (BB-W) rats and streptozotocin-diabetic rats. The distribution of the two tracers is similar. In Bruch's membrane of control rats, electron dense particles are present at regular intervals along both sides of the basal laminae of the retinal pigment epithelium (RPE) and of the choriocapillary endothelium (CE), and along collagen fibers in the zone between the two basal laminae. Within 3-6 months after the onset of hyperglycemia in both diabetic rat models, quantitative analysis shows a significant reduction in binding sites along both RPE and CE basal laminae, while thickness of both basal laminae is significantly increased. Because reductions in anionic binding sites along basal laminae in the renal glomerulus have been found to accompany changes in glomerular filtration, these changes suggest that filtration through Bruch's membrane is altered in diabetes.


Transmission of the nocturnal periodic strain of Wuchereria bancrofti by Culex quinquefasciatus: establishing the potential for urban filariasis in Thailand.
Control programmes have reduced the prevalence of Bancroftian filariasis in Thailand to low levels. Recently, there has been an influx of more than one million Myanmar immigrants into urban centres of Thailand. The prevalence of patent Wuchereria bancrofti infection in these immigrants (2-5%) has prompted concern in the public health community that the potential now exists for a re-emergence of Bancroftian filariasis in Thailand. It is possible that an urban cycle of transmission could become established. The Myanmar immigrants are infected with the nocturnal periodic (urban) type W. bancrofti for which Culex quinquefasciatus serves as the main vector. The Thai strains of Cx. quinquefasciatus have never been reported to transmit Bancroftian filariasis. Our results of feeding experiments demonstrated that the Thai Cx. quinquefasciatus are permissive for the development of Myanmar W. bancrofti to infective third-stage larvae thus establishing the potential for establishing an urban cycle of transmission in Thailand. We also adapted the SspI repeat PCR assay for the identification of infective mosquitoes that was capable of detecting a single infective stage larvae in a pool of 100 mosquitoes.


Retrospective Study on the Effects of Long-Term Use of Methylprednisolone Acetate on the Blood Work of 25 Cats.
Twenty-five cats at a private animal sanctuary received multiple nonimmunosuppressive doses of parenteral methylprednisolone acetate for at least 3 yr. Complete blood count, chemistry, and T4 results from these cats were examined to look for statistically significant changes. Results found significant changes in triglycerides, amylase, and monocytes. However, these changes remained within the reference interval. All other values showed no significant changes. These results suggest that after 3 yr of chronic parenteral administration of nonimmunosuppressive doses of methylprednisolone acetate, the complete blood count, chemistry, and T4 values in 25 cats were not significantly affected and did not result in abnormal laboratory values.


Preventing Gender-Based Violence Among Adolescents and Young Adults: Lessons From 25 Years of Program Development and Evaluation.
Effective prevention of intimate partner violence (IPV) among adolescents and young adults is a key strategy for reducing rates of gender-based violence (GBV). Numerous initiatives have been developed and evaluated over the past 25 years. There is emerging evidence about effective strategies for universal prevention of dating violence in high school settings and effective bystander interventions on university and college campuses. In addition, there have been some effective practices identified for specific groups of youth who are vulnerable to victimization (either based on past experiences of exposure to domestic violence or previous dating victimization). At the same time, though our evidence about school and college-based interventions has grown, there are significant gaps in our knowledge of effective prevention among marginalized groups. For example, there is a lack of evidence-based strategies for preventing IPV among Indigenous youth; lesbian, gay, bisexual, transgender, questioning+ [LGBTQ+] youth; and young women with disabilities, even though these groups are at elevated risk for experiencing violence. Our review of the current state of evidence for effective GBV prevention among adolescents and young adults suggests significant gaps. Our analysis of these gaps highlights the need to think more broadly about what constitutes evidence. We identify some strategies and a call to action for moving the field forward and provide examples from our work with vulnerable youth in a variety of settings.


Drug use patterns at major rock concert events.
To describe alcohol and drug use patterns in patients presenting to first aid stations at major rock concerts. We retrospectively reviewed all charts generated at the first aid stations of five major rock concerts featuring the rock groups Pink Floyd, the Grateful Dead, and the Rolling Stones. The first aid stations, located at a sports stadium, were staffed by paramedics, emergency medicine nurses, and physicians. We recorded the following data: patient demographics, history of drug or ethanol use, time spent by patient in first aid station, treatment rendered, diagnosis, and patient disposition. A total of 253, 286 spectators attended the five concert events. The rate of use of the first aid station was 1.2 per 1,000 patrons. The average age of the patrons was 26.3 +/- 7.9 years (range, 3 to 56 years). The most common diagnoses were minor trauma 130 (42%) and ethanol or illicit drug intoxication 98 (32%). Of the patients treated, 147 (48%) admitted to using illicit drugs or ethanol while attending the concerts. The median time spent in the first aid station was 15 +/- 22.5 minutes (range, 5 to 150 minutes). One hundred patients (32.5%) were treated and released, 98 (32%) were transported to emergency departments, and 110 (35.5%) signed out against medical advice. Minor trauma and the use of illicit drugs and ethanol were common in spectators presenting to first aid stations at these concert events. Physicians and paramedical personnel working at rock concerts should be aware of the current drug use patterns and should be trained in treating such drug use.


Plant neighbor identity influences plant biochemistry and physiology related to defense.
Chemical and biological processes dictate an individual organism's ability to recognize and respond to other organisms. A small but growing body of evidence suggests that plants may be capable of recognizing and responding to neighboring plants in a species specific fashion. Here we tested whether or not individuals of the invasive exotic weed, Centaurea maculosa, would modulate their defensive strategy in response to different plant neighbors. In the greenhouse, C. maculosa individuals were paired with either conspecific (C. maculosa) or heterospecific (Festuca idahoensis) plant neighbors and elicited with the plant defense signaling molecule methyl jasmonate to mimic insect herbivory. We found that elicited C. maculosa plants grown with conspecific neighbors exhibited increased levels of total phenolics, whereas those grown with heterospecific neighbors allocated more resources towards growth. To further investigate these results in the field, we conducted a metabolomics analysis to explore chemical differences between individuals of C. maculosa growing in naturally occurring conspecific and heterospecific field stands. Similar to the greenhouse results, C. maculosa individuals accumulated higher levels of defense-related secondary metabolites and lower levels of primary metabolites when growing in conspecific versus heterospecific field stands. Leaf herbivory was similar in both stand types; however, a separate field study positively correlated specialist herbivore load with higher densities of C. maculosa conspecifics. Our results suggest that an individual C. maculosa plant can change its defensive strategy based on the identity of its plant neighbors. This is likely to have important consequences for individual and community success.


Expression, crystallization, and three-dimensional structure of the catalytic domain of human plasma kallikrein.
Plasma kallikrein is a serine protease that has many important functions, including modulation of blood pressure, complement activation, and mediation and maintenance of inflammatory responses. Although plasma kallikrein has been purified for 40 years, its structure has not been elucidated. In this report, we described two systems (Pichia pastoris and baculovirus/Sf9 cells) for expression of the protease domain of plasma kallikrein, along with the purification and high resolution crystal structures of the two recombinant forms. In the Pichia pastoris system, the protease domain was expressed as a heterogeneously glycosylated zymogen that was activated by limited trypsin digestion and treated with endoglycosidase H deglycosidase to reduce heterogeneity from the glycosylation. The resulting protein was chromatographically resolved into four components, one of which was crystallized. In the baculovirus/Sf9 system, homogeneous, crystallizable, and nonglycosylated protein was expressed after mutagenizing three asparagines (the glycosylation sites) to glutamates. When assayed against the peptide substrates, pefachrome-PK and oxidized insulin B chain, both forms of the protease domain were found to have catalytic activity similar to that of the full-length protein. Crystallization and x-ray crystal structure determination of both forms have yielded the first three-dimensional views of the catalytic domain of plasma kallikrein. The structures, determined at 1.85 A for the endoglycosidase H-deglycosylated protease domain produced from P. pastoris and at 1.40 A for the mutagenically deglycosylated form produced from Sf9 cells, show that the protease domain adopts a typical chymotrypsin-like serine protease conformation. The structural information provides insights into the biochemical and enzymatic properties of plasma kallikrein and paves the way for structure-based design of protease inhibitors that are selective either for or against plasma kallikrein.


Differential modulation in human primary and secondary somatosensory cortices during the preparatory period of self-initiated finger movement.
To elucidate the mechanisms underlying sensorimotor integration, we investigated modulation in the primary (SI) and secondary (SII) somatosensory cortices during the preparatory period of a self-initiated finger extension. Electrical stimulation of the right median nerve was applied continuously, while the subjects performed a self-initiated finger extension and were instructed not to pay attention to the stimulation. The preparatory period was divided into five sub-periods from the onset of the electromyogram to 3000 ms before movement and the magnetoencephalogram signals following stimulation in each sub-period were averaged. Multiple source analysis indicated that the equivalent current dipoles (ECDs) were located in SI and bilateral SII. Although the ECD moment for N 20 m (the upward deflection peaking at around 20 ms) was not significantly changed, that for P 30 m (the downward deflection peaking at around 30 m) was significantly smaller in the 0- to -500-ms sub-period than the -2000- to -3000-ms sub-period. As for SII, the ECD moment for the SII ipsilateral to movement showed no significant change, while that for the contralateral SII was significantly larger in the 0- to -500-ms sub-period than the -1500- to -2000-ms or -2000- to -3000-ms sub-period. The opposite effects of movement on SI and SII cortices indicated that these cortical areas play a different role in the function of the sensorimotor integration and are affected differently by the centrifugal process.


Buried and trapped penis: a case report.
While buried penis cases are characterised by congenital normal attachments to penis, trapped penis cases are characterised by insufficiency of penile skin occurring as a complication after surgical operations such as circumcision. Unless diagnosed, circumcision procedures should be avoided in congenital concealed penis cases. Here we present a case of congenital buried penis with deteriorated clinical findings after two circumcision procedures at 1.5 years of age. The surgical treatment applied in this case is discussed.


The impact of supported employment and working on clinical and social functioning: results of an international study of individual placement and support.
Concerns are frequently expressed that working might worsen the mental health of people with severe mental illness (SMI). Several studies of Individual Placement and Support (IPS), however, have found associations between working and better nonvocational outcomes. IPS has been found to double the return to work of people with SMI in 6 European countries. To explore separately associations between IPS, returning to work, and clinical and social outcomes. Patients (n = 312) in a randomized controlled trial of IPS in 6 European centers were followed up for 18 months. There were no differences in clinical and social functioning between IPS and control patients at 18 months. Those who worked had better global functioning, fewer symptoms, and less social disability at final follow-up; greater job tenure was associated with better functioning. Working was associated with concurrently better clinical and social functioning, but this contrast was stronger in the control group, suggesting that IPS was better than the control service at helping more unwell patients into work. Working was associated with having been in remission and out of hospital for the previous 6 months. It was also associated with a slight decrease in depression and with being in remission over the subsequent 6 months. Concerns among clinicians about possible detrimental effects of working and supported employment have been misplaced. Although some of the associations found may have been selection effects, there is sufficient evidence of work having beneficial effects on clinical and social functioning to merit further exploration.


Comparison of phenotypic value changes in pure lines of Bombyx mori (Lepidoptera: Bombycidae) during consecutive generations following initial selection on cocoon weight.
The experiments reported here were conducted to investigate the effect of selection on three quantitative traits, namely cocoon weight, cocoon shell weight, and cocoon shell percentage, during four generations by rearing six pure breeds of domesticated silkworm, Bombyx mori L. (Lepidoptera: Bombycidae) of Chinese and Japanese origin compared with random unselected groups as controls. All stages of rearing and data recording were performed over four rearing periods, with generations 1-3 during successive spring seasons and generation 4 during the autumn season in year 3. Each pure line contained two groups of selected and random (control) groups. Comparisons included the effect of selection methods, pure line, and generation on the phenotypic values. We found strong main effects of pure line, generation, sex, and group and support for nearly all interactions between these main effects for all three response traits. The results indicated that cocoon weight and cocoon shell weight in the selected group were higher than in the control or nonselected group. Both selected and nonselected groups had the lowest cocoon weight, cocoon shell weight, and cocoon shell percentage in the fourth generation when environmental conditions during the autumn season were less favorable than spring. The cocoon weight and cocoon shell weight averages were higher for nonselected groups in the second and third generations, and for the selected group in the first generation due to the direct effect of selection.


Antiepileptic prophylaxis following severe traumatic brain injury within a military cohort.
Traumatic brain injury increases the risk of both early and late seizures. Antiepileptic prophylaxis reduces early seizures, but their use beyond 1 week does not prevent the development of post-traumatic epilepsy. Furthermore, prolonged prophylaxis exposes patients to side effects of the drugs and has occupational implications. The American Academy of Neurology recommends that antiepileptic prophylaxis should be started for patients with severe traumatic brain injury and discontinued after 1 week. An audit is presented here that investigates the use of prophylaxis in a cohort of military patients admitted to the UK Defence Medical Rehabilitation Centre (DMRC). Data were collected and analysed retrospectively from electronic and paper records between February 2009 and August 2012. The timing and duration of antiepileptic drug use and the incidence of seizures were recorded. During the study period, 52 patients with severe traumatic brain injury were admitted to the rehabilitation centre: 25 patients (48%) were commenced on prophylaxis during the first week following injury while 27 (52%) did not receive prophylaxis. Only one patient (2%) received prophylaxis for the recommended period of 1 week, 22 patients (42%) received prophylaxis for longer than 1 week with a mean duration of 6.2 months. Two patients (4%) had post-traumatic epilepsy and started on treatment at DMRC. The use of antiepileptic prophylaxis varies widely and is generally inconsistent with evidence-based guidance. This exposes some patients to a higher risk of early seizures and others to unnecessary use of antiepileptics. Better implementation of prophylaxis is required.


A Distinct Class of Genome Rearrangements Driven by Heterologous Recombination.
Erroneous DNA repair by heterologous recombination (Ht-REC) is a potential threat to genome stability, but evidence supporting its prevalence is lacking. Here we demonstrate that recombination is possible between heterologous sequences and that it is a source of chromosomal alterations in mitotic and meiotic cells. Mechanistically, we find that the RTEL1 and HIM-6/BLM helicases and the BRCA1 homolog BRC-1 counteract Ht-REC in Caenorhabditis elegans, whereas mismatch repair does not. Instead, MSH-2/6 drives Ht-REC events in rtel-1 and brc-1 mutants and excessive crossovers in rtel-1 mutant meioses. Loss of vertebrate Rtel1 also causes a variety of unusually large and complex structural variations, including chromothripsis, breakage-fusion-bridge events, and tandem duplications with distant intra-chromosomal insertions, whose structure are consistent with a role for RTEL1 in preventing Ht-REC during break-induced replication. Our data establish Ht-REC as an unappreciated source of genome instability that underpins a novel class of complex genome rearrangements that likely arise during replication stress.


Self-talk influences vertical jump performance and kinematics in male rugby union players.
We examined the effects of instructional and motivational self-talk on centre of mass displacement and hip kinematics during the vertical jump. Twenty-four male rugby union players (age 21.1 years, s = 3.5; body mass 81.0 kg, s = 8.9; height 1.80 m, s = 0.06) performed three vertical jump tests, with a 2 min rest between jumps. Before each jump, participants engaged in one of three counterbalanced interventions (motivational self-talk, instructional self-talk or no-intervention). Motivational self-talk led to greater centre of mass displacement (0.602 m, s = 0.076; P = 0.012) than the no-intervention control (0.583 m, s = 0.085). Centre of mass displacement did not differ between instructional self-talk and the control condition or between motivational and instructional self-talk. Motivational (100.75 degrees , s = 16.05; P = 0.001) and instructional self-talk (106.14 degrees , s = 17.04; P = 0.001) led to greater hip displacement than the no-intervention control (94.11 degrees , s = 17.14). There was also a significant difference in hip displacement between motivational and instructional self-talk (P = 0.014), although there was no difference between instructional self-talk and the control condition. Motivational (451.69 degrees /s, s = 74.34; P = 0.008) and instructional self-talk (462.01 degrees /s, s = 74.37; P = 0.001) led to greater hip rotation velocity than the no-intervention control (434.37 degrees /s, s = 75.37), although there was no difference between the two self-talk interventions. These results indicate that self-talk may influence performance and technique during the vertical jump in male rugby players.


Multiple adaptive routes of Salmonella enterica Typhimurium to biocide and antibiotic exposure.
Biocides and antibiotics are used to eradicate or prevent the growth of microbial species on surfaces (occasionally on catheters), or infected sites, either in combination or sequentially, raising concerns about the development of co-resistance to both antimicrobial types. The effect of such compounds on Salmonella enterica, a major food-borne and zoonotic pathogen, has been analysed in different studies, but only few works evaluated its biological cost, and the overall effects at the genomic and transcriptomic levels associated with diverse phenotypes resulting from biocide exposure, which was the aim of this work. Exposure to triclosan, clorhexidine, benzalkonium, (but not to hypochlorite) resulted in mutants with different phenotypes to a wide range of antimicrobials even unrelated to the selective agent. Most biocide-resistant mutants showed increased susceptibility to compounds acting on the cell wall (β-lactams) or the cell membranes (poly-L-lysine, polymyxin B, colistin or toxic anions). Mutations (SNPs) were found in three intergenic regions and nine genes, which have a role in energy production, amino acids, carbohydrates or lipids metabolism, some of them involved in membrane transport and pathogenicity. Comparative transcriptomics of biocide-resistant mutants showed over-expression of genes encoding efflux pumps (sugE), ribosomal and transcription-related proteins, cold-shock response (cpeE) and enzymes of microaerobic metabolism including those of the phosphotransferase system. Mainly ribosomal, metabolic and pathogenicity-related genes had affected expression in both in vitro-selected biocide mutants and field Salmonella isolates with reduced biocide susceptibility. Multiple pathways can be involved in the adaptation of Salmonella to biocides, mainly related with global stress, or involving metabolic and membrane alterations, and eventually causing "collateral sensitivity" to other antimicrobials. These changes might impact the bacterial-environment interaction, imposing significant bacterial fitness costs which may reduce the chances of fixation and spread of biocide resistant mutants.


Fabry disease: identification of 50 novel alpha-galactosidase A mutations causing the classic phenotype and three-dimensional structural analysis of 29 missense mutations.
Fabry disease, an X-linked recessive inborn error of glycosphingolipid catabolism, results from the deficient activity of the lysosomal exoglycohydrolase, alpha-galactosidase A (EC 22.214.171.124; alpha-Gal A). The molecular lesions in the alpha-Gal A gene causing the classic phenotype of Fabry disease in 66 unrelated families were determined. In 49 families, 50 new mutations were identified, including: 29 missense mutations (N34K, T41I, D93V, R112S, L166G, G171D, M187T, S201Y, S201F, D234E, W236R, D264Y, M267R, V269M, G271S, G271V, S276G, Q283P, A285P, A285D, M290I, P293T, Q312H, Q321R, G328V, E338K, A348P, E358A, Q386P); nine nonsense mutations (C56X, E79X, K127X, Y151X, Y173X, L177X, W262X, Q306X, E338X); five splicing defects (IVS4-1G>A, IVS5-2A>G, IVS5+3A>G, IVS5+4A>G, IVS6-1G>C); four small deletions (18delA, 457delGAC, 567delG, 1096delACCAT); one small insertion (996insC); one 3.1 kilobase Alu-Alu deletion (which included exon 2); and one complex mutation (K374R, 1124delGAG). In 18 families, 17 previously reported mutations were identified, with R112C occurring in two families. In two classically affected families, affected males were identified with two mutations: one with two novel mutations, D264Y and V269M and the other with one novel (Q312H) and one previously reported (A143T) mutation. Transient expression of the individual mutations revealed that D264Y and Q312H were localised in the endoplasmic reticulum and had no detectable or markedly reduced activity, whereas V269M and A143T were localised in lysosomes and had approximately 10 per cent and approximately 35 per cent of expressed wild-type activity, respectively. Structural analyses based on the enzyme's three-dimensional structure predicted the effect of the 29 novel missense mutations on the mutant glycoprotein's structure. Of note, three novel mutations (approximately 10 per cent) were predicted not to significantly alter the glycoprotein's structure; however, they were disease causing. These studies further define the molecular heterogeneity of the alpha-Gal A mutations in classical Fabry disease, permit precise heterozygote detection and prenatal diagnosis, and provide insights into the structural alterations of the mutant enzymes that cause the classic phenotype.


Transmural atrial insulating lesions using epicardially applied radiofrequency energy coupled with saline irrigation.
The Maze procedure has been demonstrated to eliminate atrial fibrillation (AF) refractory to medical management. The procedure, however, is technically difficult and requires prolonged cardiopulmonary bypass (CPB). We investigated the use of epicardially applied radiofrequency (RF) energy delivered using a saline-irrigated probe to functionally reproduce the linear lesions of the surgical Maze procedure. The goals of this study were to safely produce transmural atrial insulation lines without CPB and to investigate whether they could retain their electrical insulation properties and structural integrity over an acute time period. Baseline pacing thresholds were measured on the atria of six swine. Then, circular insulating lesions (n = 11) were created around each pacing site using a saline-irrigated RF probe and specifically designed radiofrequency generator. Pacing thresholds were repeated at the same atrial sites, and in adjacent areas, immediately following lesion creation and at 7 days. At sacrifice the areas of interest were removed for histopathology. Pacing thresholds increased from 0.9 +/- 0.09 V to 9.59 +/- 0.87 V immediately following lesion formation (p <.05) and remained supraphysiologic at 4.05 +/- 0.36 V after 7 days (p <.05). Thresholds from outside the lesions were unchanged from baseline. There was no evidence of perforation and histologic analysis demonstrated transmural necrosis with advancing granulation tissue. Our data suggest that epicardially applied RF energy coupled with saline irrigation can produce transmural electrical insulation lines for up to 7 days. However, the decreasing trend in thresholds toward baseline suggests further investigation is needed to verify insulation line continuity over a chronic time period. These lines were created safely and efficiently without CPB, and may be applicable to a minimally invasive Maze procedure.


Hyaluronan synthase mediates dye translocation across liposomal membranes.
Hyaluronan (HA) is made at the plasma membrane and secreted into the extracellular medium or matrix by phospolipid-dependent hyaluronan synthase (HAS), which is active as a monomer. Since the mechanism by which HA is translocated across membranes is still unresolved, we assessed the presence of an intraprotein pore within HAS by adding purified Streptococcus equisimilis HAS (SeHAS) to liposomes preloaded with the fluorophore Cascade Blue (CB). CB translocation (efflux) was not observed with mock-purified material from empty vector control E. coli membranes, but was induced by SeHAS, purified from membranes, in a time- and dose-dependent manner. CB efflux was eliminated or greatly reduced when purified SeHAS was first treated under conditions that inhibit enzyme activity: heating, oxidization or cysteine modification with N-ethylmaleimide. Reduced CB efflux also occurred with SeHAS K48E or K48F mutants, in which alteration of K48 within membrane domain 2 causes decreased activity and HA product size. The above results used liposomes containing bovine cardiolipin (BCL). An earlier study testing many synthetic lipids found that the best activating lipid for SeHAS is tetraoleoyl cardiolipin (TO-CL) and that, in contrast, tetramyristoyl cardiolipin (TM-CL) is an inactivating lipid (Weigel et al, J. Biol. Chem. 281, 36542, 2006). Consistent with the effects of these CL species on SeHAS activity, CB efflux was more than 2-fold greater in liposomes made with TO-CL compared to TM-CL. The results indicate the presence of an intraprotein pore in HAS and support a model in which HA is translocated to the exterior by HAS itself.


Major genetic susceptibility for venous thromboembolism in men: a study of Danish twins.
Although several genetic determinants (mutations or polymorphisms) have been associated with increased risk of venous thromboembolism, the overall influence of genetic factors on this disease is unknown. We linked the Danish Twin Registry, which includes twins born 1870-1953, with the Danish National Registry of Patients, comprising all hospitalizations in Denmark since 1977. We then determined the risk of venous thromboembolism as determined from discharge diagnosis. We identified 26,982 twins who were alive on 1 January 1977, and computed measures of familial and genetic association of venous thrombotic disorders. Individuals were classified according to zygosity and hospitalization with venous thromboembolism. Since 1977, 678 twins were hospitalized with an episode of venous thromboembolism. Of these, only 545 pairs (281 male pairs and 264 female pairs) were alive in 1977. For men, the concordance rates for mono- and dizygotic twin pairs, respectively, were 0.22 (95% confidence interval = 0.14 to 0.30) and 0.08 (0.04-0.12). The odds ratio (interpreted as the relative risk of venous thromboembolism for one twin, given venous thromboembolism in the partner twin) was 13.5 (7.3-24.8) among monozygotic twins and 3.8 (1.8-8.3) among dizygotic twins. The respective correlations for venous thromboembolism were 0.55 (0.38-0.70) and 0.26 (0.09-0.42). The proportion of the variance attributable to genetic effects on venous thromboembolism in males was 55% (39%-68%). The remaining variation could be attributed to men's nonfamilial environments. In contrast, for women there was no intra-twin pair similarity for venous thromboembolism. We found differences in genetic susceptibility to venous thromboembolism between the sexes, with genetic factors playing a substantially stronger role in males than in females.


Broad Hemagglutinin-Specific Memory B Cell Expansion by Seasonal Influenza Virus Infection Reflects Early-Life Imprinting and Adaptation to the Infecting Virus.
Memory B cells (MBCs) are key determinants of the B cell response to influenza virus infection and vaccination, but the effect of different forms of influenza antigen exposure on MBC populations has received little attention. We analyzed peripheral blood mononuclear cells and plasma collected following human H3N2 influenza infection to investigate the relationship between hemagglutinin-specific antibody production and changes in the size and character of hemagglutinin-reactive MBC populations. Infection produced increased concentrations of plasma IgG reactive to the H3 head of the infecting virus, to the conserved stalk, and to a broad chronological range of H3s consistent with original antigenic sin responses. H3-reactive IgG MBC expansion after infection included reactivity to head and stalk domains. Notably, expansion of H3 head-reactive MBC populations was particularly broad and reflected original antigenic sin patterns of IgG production. Findings also suggest that early-life H3N2 infection "imprints" for strong H3 stalk-specific MBC expansion. Despite the breadth of MBC expansion, the MBC response included an increase in affinity for the H3 head of the infecting virus. Overall, our findings indicate that H3-reactive MBC expansion following H3N2 infection is consistent with maintenance of response patterns established early in life, but nevertheless includes MBC adaptation to the infecting virus.IMPORTANCE Rapid and vigorous virus-specific antibody responses to influenza virus infection and vaccination result from activation of preexisting virus-specific memory B cells (MBCs). Understanding the effects of different forms of influenza virus exposure on MBC populations is therefore an important guide to the development of effective immunization strategies. We demonstrate that exposure to the influenza hemagglutinin via natural infection enhances broad protection through expansion of hemagglutinin-reactive MBC populations that recognize head and stalk regions of the molecule. Notably, we show that hemagglutinin-reactive MBC expansion reflects imprinting by early-life infection and that this might apply to stalk-reactive, as well as to head-reactive, MBCs. Our findings provide experimental support for the role of MBCs in maintaining imprinting effects and suggest a mechanism by which imprinting might confer heterosubtypic protection against avian influenza viruses. It will be important to compare our findings to the situation after influenza vaccination.


The Role of Endoplasmic Reticulum Stress and Unfolded Protein Response in Atherosclerosis.
Pathogenesis of atherosclerosis is a complex process involving several metabolic and signalling pathways. Accumulating evidence demonstrates that endoplasmic reticulum stress and associated apoptosis can be induced in the pathological conditions of atherosclerotic lesions and contribute to the disease progression. Notably, they may play a role in the development of vulnerable plaques that induce thrombosis and are therefore especially dangerous. Endoplasmic reticulum stress response is regulated by several signaling mechanisms that involve protein kinases and transcription factors. Some of these molecules can be regarded as potential therapeutic targets to improve treatment of atherosclerosis. In this review we will discuss the role of endoplasmic reticulum stress and apoptosis in atherosclerosis development in different cell types and summarize the current knowledge on potential therapeutic agents targeting molecules regulating these pathways and their possible use for anti-atherosclerotic therapy.


Mechanical properties of lower limb dermis following static and cyclic compression.
Lower extremity amputations and foot ulcers are complications associated with diabetes, and have been shown to affect diabetic African Americans (AA) three times as often as diabetic non-Hispanic Whites (NHW). Possible causes for the increased risk include ethnic differences in structure and function within the dermis of the lower extremity. Testing this hypothesis requires studying the mechanical properties of skin from different ethnic groups with and without the diagnosis of noninsulin-dependent diabetes. The purpose of this study was to develop a testing method to investigate changes in tensile mechanical properties resulting from static and cyclic compression of dermis harvested from patients undergoing lower extremity amputations. Full thickness dermal samples were obtained from 15 patients undergoing below-knee amputations. Sections of each sample were conditioned with a compressive static pressure (170 mmHg) or cyclic pressures (110-170 mmHg) for 4 hours to elicit collagen bundle remodeling. Skin samples were then tested in tension to obtain sub-plastic stress vs. strain mechanical behavior. Length of the stress-strain toe-region was examined to quantify the effect of collagen bundle remodeling. Toe-region mean lengths were 0.141±0.041, 0.146±0.034, and 0.164±0.064 strains for the control, cyclic, and statically compressed samples respectively (p>.05). These results suggest that the preconditioning regimes did not produce sufficient collagen remodeling to affect the tensile properties of full-thickness dermis. Future work will examine histology from each specimen to identify microstructural features associated with this trend.