Online motor control in children with developmental coordination disorder: chronometric analysis of double-step reaching performance.
Although there are a number of plausible accounts to explain movement clumsiness in children [or developmental coordination disorder (DCD)], the cause(s) of the disorder remain(s) an issue of debate. One aspect of motor control that is particularly important to the fluid expression of skill is rapid online control (ROC). Data on DCD have been conflicting. While some recent work using double-step reaching suggests no difficulty in online control, others suggest deficits (e.g. based on sequential pointing). To help resolve this debate, we suggest two things: use of recent neuro-computational models as a framework for investigating motor control in DCD, and more rigorous investigation of double-step reaching. Our working assumption here is that ROC is only viable through the seamless integration of predictive (or forward) models of movement and feedback-based mechanisms. The aim of this chronometric study was to explore ROC in children with DCD using a double-step reaching paradigm. We predicted slower online adjustments in DCD based on the argument that these children manifest a core difficulty in predictive control. Participants were a group of 17 children with DCD and 27 typically developing children aged between 7 and 12 years. Visual targets were presented on a 17-inch LCD touch screen, inclined to an angle of 15° from horizontal. The children were instructed to press each target as it appeared as quickly and accurately as possible. For 80% of the trials, the central target location remained unchanged for the duration of the movement (non-jump trials), while for the remaining 20% of trials, the target jumped at movement onset to one of the two peripheral locations (jump trials). Reaction time (RT), movement time (MT) and reaching errors were recorded. For both groups, RT did not vary according to trial condition, while children with DCD were slower to initiate movement. Further, the MT of children with DCD was prolonged to a far greater extent on jump trials relative to controls, with a large effect size. As well, children with DCD committed significantly more errors, notably a reduced ability to inhibit central responses on jump trials. Our findings help reconcile some disparate findings in the literature using similar tasks. The pattern of performance in children with DCD suggests impairment in the ability to make rapid online adjustments that are based on a predictive (or internal) model of the action. These results pave the way for future kinematic investigation.


Autopsy in suspected COVID-19 cases.
The severe acute respiratory syndrome (SARS)-coronavirus-2 (CoV-2) outbreak in Wuhan, China has now spread to many countries across the world including the UK with an increasing death toll. This will inevitably lead to an increase in the number of suspected coronavirus disease 2019 (COVID-19)-related deaths at autopsy. The Royal College of Pathologists has responded to this concern with the release of a briefing on autopsy practice relating to COVID-19. The following article is a summary and interpretation of these guidelines. It includes a description of hazard group 3 organisms, the category to which SARS-CoV-2 has been assigned, a brief description of what is currently known about the pathological and autopsy findings in COVID-19, a summary of the recommendations for conducting autopsies in suspected COVID-19 cases and the techniques for making the diagnosis at autopsy. It concludes by considering the clinicopathological correlation and notification of such cases.


Chemsex behaviours among men who have sex with men: A systematic review of the literature.
'Chemsex' is the use of drugs before or during planned sexual events to facilitate, enhance, prolong and sustain the experience. Drugs associated with chemsex are methamphetamine, GHB/GBL, mephedrone, cocaine and ketamine. This review syntheses published research on the antecedents, behaviours and consequences associated with chemsex behaviours among men who have sex with men (MSM). Papers from high income countries which were published between January 2000 and September 2018 reporting the use of chemsex drugs before or during sex were identified through Medline, Web of Science, CINAHL and Central. Results were synthesised using a narrative approach and conceptualised using a behavioural analysis framework. The search identified 2492 publications, of which 38 were included in the final synthesis. There were wide variations in chemsex prevalence estimates due to the heterogeneous sampling in the studies. Chemsex participants have expectations that the drugs will positively affect their sexual encounters and HIV positive MSM are more likely to engage in the behaviour than HIV negative MSM. There were wide ranging prevalence estimates on injecting drugs for sexual purposes and the sharing of injecting equipment with some evidence of unsafe injecting practices. Participants were more likely to engage in condomless anal intercourse than men who do not engage in chemsex. This may increase the risk of transmission for HIV and other sexually transmitted infections. A minority of MSM appear to engage in chemsex behaviours but they are at risk of this negatively impacting on their health and well-being. Further research is required to examine high risk chemsex behaviours, impact of chemsex on psycho-social well-being and if chemsex influences uptake of PrEP, PEP and sexual health screening.


An excess of widespread pain among South Asians: are low levels of vitamin D implicated?
Anecdotal reports from rheumatologists in the United Kingdom suggest that patients from South Asian backgrounds are more likely to report widespread body pain. To confirm the presence of an excess of widespread pain in South Asians, and to evaluate the relationship of their symptoms with levels of 25-OH vitamin D. Two population studies involving over 3135 subjects were carried out in the North West and Midlands areas of England. The first study confirmed an excess of widespread pain among South Asians (OR = 1.6, 95% CI 1.3 to 2.1). The second smaller study conducted only among young women also showed a similar excess of widespread pain among South Asians (OR = 1.8, 95% CI 0.7 to 4.7) and found that low levels of 25-OH vitamin D (<10 ng/ml) were more common among those with widespread pain (OR = 3.5, 95% CI 0.4 to 31.0). Owing to the small numbers, the relationship between 25-OH vitamin D and widespread pain must be considered preliminary and requires further investigation. However, it may be one potentially treatable cause of widespread pain.


Towards a landscape scale management of pesticides: ERA using changes in modelled occupancy and abundance to assess long-term population impacts of pesticides.
Pesticides are regulated in Europe and this process includes an environmental risk assessment (ERA) for non-target arthropods (NTA). Traditionally a non-spatial or field trial assessment is used. In this study we exemplify the introduction of a spatial context to the ERA as well as suggest a way in which the results of complex models, necessary for proper inclusion of spatial aspects in the ERA, can be presented and evaluated easily using abundance and occupancy ratios (AOR). We used an agent-based simulation system and an existing model for a widespread carabid beetle (Bembidion lampros), to evaluate the impact of a fictitious highly-toxic pesticide on population density and the distribution of beetles in time and space. Landscape structure and field margin management were evaluated by comparing scenario-based ERAs for the beetle. Source-sink dynamics led to an off-crop impact even when no pesticide was present off-crop. In addition, the impacts increased with multi-year application of the pesticide whereas current ERA considers only maximally one year. These results further indicated a complex interaction between landscape structure and pesticide effect in time, both in-crop and off-crop, indicating the need for NTA ERA to be conducted at landscape- and multi-season temporal-scales. Use of AOR indices to compare ERA outputs facilitated easy comparison of scenarios, allowing simultaneous evaluation of impacts and planning of mitigation measures. The landscape and population ERA approach also demonstrates that there is a potential to change from regulation of a pesticide in isolation, towards the consideration of pesticide management at landscape scales and provision of biodiversity benefits via inclusion and testing of mitigation measures in authorisation procedures.


TAK1 (MAP3K7) signaling regulates hematopoietic stem cells through TNF-dependent and -independent mechanisms.
A cytokine/stress signaling kinase Tak1 (Map3k7) deficiency is known to impair hematopoietic progenitor cells. However, the role of TAK1 signaling in the stem cell function of the hematopoietic system is not yet well defined. Here we characterized hematopoietic stem cells (HSCs) harboring deletion of Tak1 and its activators, Tak1 binding proteins 1 and 2 (Tab1 and Tab2) using a competitive transplantation assay in a mouse model. Tak1 single or Tab1/Tab2 double deletions completely eliminated the reconstitution activity of HSCs, whereas Tab1 or Tab2 single deletion did not cause any abnormality. Tak1 single or Tab1/Tab2 double deficient lineage-negative, Sca-1(+), c-Kit(+) (LSK) cells did not proliferate and underwent cell death. We found that Tnfr1 deficiency restored the reconstitution activity of Tak1 deficient bone marrow cells for 6-18 weeks. However, the reconstitution activity of Tak1- and Tnfr1-double deficient bone marrow cells declined over the long term, and the number of phenotypically identified long-term hematopoietic stem cells were diminished. Our results indicate that TAB1- or TAB2-dependent activation of TAK1 is required for maintenance of the hematopoietic system through two mechanisms: one is prevention of TNF-dependent cell death and the other is TNF-independent maintenance of long-term HSC.


Age-gender differences in the postural sway during squat and stand-up movement.
Incidence of falling among elderly female has been reported to be much higher than that of elderly male. Although the gender differences in the elderly were reported for the static postural sway, there has been no investigation of the gender difference for the dynamic postural sway. This study investigates how age and gender affect the postural sway during dynamic squat and stand-up movement. 124 subjects (62 subjects for each of young and elderly) performed consecutive squat and stand-up movement, 2 times in one session, and 2 sessions per subject. Center of pressure (COP) was measured using force platform during the test. Outcome measures included peak-to-peak sways of the COP (COP sway) in the sagittal plane (anteroposterior) and frontal plane (mediolateral) and also those normalized by body height. Two-way ANOVA and post-hoc comparisons were performed for the outcome measures with the independent factors of age and gender. All outcome measures, excluding mediolateral COP sway, showed significant interaction of age and gender (p<0.05). Post-hoc test revealed that only female showed increase in COP sway with age. When normalized by height, increase in COP sways (both directions) with age significant only in women resulted in greater sways in elderly female than elderly male. This may be related to the greater fall rate of elderly female than that of elderly men while performing dynamic activities.


Vibrational Activation of Methane Chemisorption: The Role of Symmetry.
Quantum state-resolved reactivity measurements probe the role of vibrational symmetry on the vibrational activation of the dissociative chemisorption of CH4 on Pt(111). IR-IR double resonance excitation in a molecular beam is used to prepare CH4 in all three different vibrational symmetry components A1, E, and F2 of the 2ν3 antisymmetric stretch overtone vibration. Methyl dissociation products chemisorbed on the cold Pt(111) surface are detected via reflection absorption infrared spectroscopy (RAIRS). We observe similar reactivity for CH4 prepared in the A1 and F2 sublevels but up to a factor of 2 lower reactivity for excitation of the E sublevel. It is suggested that differences in the localization of the C-H stretch amplitudes for the three states at the transition state leads to the observed difference in reactivity rather than state-specific vibrational energy transfer to electronic excitation of the metal.


Identification of secreted proteins regulated by cAMP in glioblastoma cells using glycopeptide capture and label-free quantification.
Exposure of glioblastoma U87MG cells to a cAMP analog leads to a decrease in proliferation, invasion, and angiogenic potential. Here, we apply a label-free MS-based approach to identify formerly N-linked glycopeptides that change in abundance upon cAMP treatment. Over 150 unique glycopeptides in three biological repetitions were quantified, leading to the identification of 14 upregulated proteins and 21 downregulated proteins due to cAMP treatment. Of these, eight have been validated, either through comparison with microarray data or by Western blot. We estimate our ability to identify differentially expressed peptides at greater than 85% in a single biological repetition, while the analysis of multiple biological repetitions lowers the false positive rate to approximately 2%. Many of the proteins identified in this study are involved in cell signaling and some, such as Tenascin C, Cathepsin L, Neuroblastoma suppressor of tumorigenicity, and AXL/UFO tyrosine-protein kinase receptor, have been previously shown to be involved in glioblastoma progression. We also identify several semitryptic peptides that increase in abundance upon cAMP treatment, suggesting that cAMP regulates protease activity in these cells. Overall, these results demonstrate the benefits of using a highly specific enrichment method for quantitative proteomic experiments.


Patterns of hepatitis C virus RNA levels during acute infection: the InC3 study.
Understanding the patterns of HCV RNA levels during acute hepatitis C virus (HCV) infection provides insights into immunopathogenesis and is important for vaccine design. This study evaluated patterns of HCV RNA levels and associated factors among individuals with acute infection. Data were from an international collaboration of nine prospective cohorts of acute HCV (InC3 Study). Participants with well-characterized acute HCV infection (detected within three months post-infection and interval between the peak and subsequent HCV RNA levels ≤ 120 days) were categorised by a priori-defined patterns of HCV RNA levels: i) spontaneous clearance, ii) partial viral control with persistence (≥ 1 log IU/mL decline in HCV RNA levels following peak) and iii) viral plateau with persistence (increase or <1 log IU/mL decline in HCV RNA levels following peak). Factors associated with HCV RNA patterns were assessed using multinomial logistic regression. Among 643 individuals with acute HCV, 162 with well-characterized acute HCV were identified: spontaneous clearance (32%), partial viral control with persistence (27%), and viral plateau with persistence (41%). HCV RNA levels reached a high viraemic phase within two months following infection, with higher levels in the spontaneous clearance and partial viral control groups, compared to the viral plateau group (median: 6.0, 6.2, 5.3 log IU/mL, respectively; P = 0.018). In the two groups with persistence, Interferon lambda 3 (IFNL3) CC genotype was independently associated with partial viral control compared to viral plateau (adjusted odds ratio [AOR]: 2.75; 95%CI: 1.08, 7.02). In the two groups with viral control, female sex was independently associated with spontaneous clearance compared to partial viral control (AOR: 2.86; 95%CI: 1.04, 7.83). Among individuals with acute HCV, a spectrum of HCV RNA patterns is evident. IFNL3 CC genotype is associated with initial viral control, while female sex is associated with ultimate spontaneous clearance.


Effects of a combined treatment with mTOR inhibitor RAD001 and tamoxifen in vitro on growth and apoptosis of human cancer cells.
Interactions between estrogen receptor signaling and the PI3K/Akt pathway are present in estrogen-dependent cancer cells. Therapeutical inhibition of each of these pathways has been proven to exert antitumoral effects. Inhibition of mammalian target of rapamycin (mTOR), a downstream target of Akt, is able to restore tamoxifen response in tamoxifen-resistant breast cancer cells. Given that Akt and mTOR phosphorylation also is frequently detected in ovarian and endometrial cancer, we intended to find out to what extent mTOR inhibitor RAD001 (everolimus) and tamoxifen add to each other's effects on growth and apoptosis of cancer cell lines derived from these tissues when given concomitantly. OVCAR-3 and SK-OV-3 ovarian cancer cells, HEC-1A endometrial adenocarcinoma cells and MCF-7 breast cancer cells were treated with different concentrations of mTOR inhibitor RAD001 alone or in combination with 4-OH tamoxifen. Relative numbers of viable cells were assessed by means of the resazurin-based Cell Titer Blue assay, cellular apoptosis was examined by measurement of activated caspases 3 and 7 by means of the luminometric Caspase-Glo assay. Treatment with RAD001 resulted in growth inhibition of all employed cancer cell lines in a dose-dependent manner, and SK-OV-3 ovarian cancer cells proved to be most sensitive to this drug. Moreover, we report the observation of additive, but not synergistical growth inhibitory effects of a combination treatment with RAD001 and 4-OH TAM on SK-OV-3 and OVCAR-3 ovarian cancer cells and MCF-7 breast cancer cells in vitro, whereas no such effect was observed in HEC-1A endometrial adenocarcinoma cells. Combination treatment with both drugs was demonstrated to be superior to single treatment with lower concentrations (0.1 and 1 nM) of RAD001 or standard concentrations of 4-OH TAM. Furthermore, RAD001 increased the apoptotic effect triggered by high 4-OH TAM concentrations in SK-OV-3 ovarian cancer cells. Combination treatment with RAD001 and 4-OH TAM in vitro exerts an additive antitumoral effect on ovarian cancer cells and MCF-7 breast cancer cells. The significance of these data in the clinical situation has to be evaluated in further studies.


Smoothened mutants reveal redundant roles for Shh and Ihh signaling including regulation of L/R asymmetry by the mouse node.
Genetic analyses in Drosophila have demonstrated that the multipass membrane protein Smoothened (Smo) is essential for all Hedgehog signaling. We show that Smo acts epistatic to Ptc1 to mediate Shh and Ihh signaling in the early mouse embryo. Smo and Shh/Ihh compound mutants have identical phenotypes: embryos fail to turn, arresting at somite stages with a small, linear heart tube, an open gut and cyclopia. The absence of visible left/right (L/R) asymmetry led us to examine the pathways controlling L/R situs. We present evidence consistent with a model in which Hedgehog signaling within the node is required for activation of Gdf1, and induction of left-side determinants. Further, we demonstrate an absolute requirement for Hedgehog signaling in sclerotomal development and a role in cardiac morphogenesis.[Dedicated to Rosa Beddington, a pioneer in mammalian embryology].


Human adipose tissue-derived mesenchymal stromal cells promote B-cell motility and chemoattraction.
Mesenchymal stromal cells hold special interest for cell-based therapy because of their tissue-regenerative and immunosuppressive abilities. B-cell involvement in chronic inflammatory and autoimmune pathologies makes them a desirable target for cell-based therapy. Mesenchymal stromal cells are able to regulate B-cell function; although the mechanisms are little known, they imply cell-to-cell contact. We studied the ability of human adipose tissue-derived mesenchymal stromal cells (ASCs) to attract B cells. We show that ASCs promote B-cell migration through the secretion of chemotactic factors. Inflammatory/innate signals do not modify ASC capacity to mediate B-cell motility and chemotaxis. Analysis of a panel of B cell-related chemokines showed that none of them appeared to be responsible for B-cell motility. Other ASC-secreted factors able to promote cell motility and chemotaxis, such as the cytokine interleukin-8 and prostaglandin E2, did not appear to be implicated. We propose that ASC promotion of B-cell migration by undefined secreted factors is crucial for ASC regulation of B-cell responses.


A comparison of thermodilution and pulsed Doppler cardiac output measurement in critically ill children.
To evaluate the pulsed Doppler cardiac output method as a noninvasive means for determining cardiac output in critically ill children, we performed paired pulsed Doppler and thermodilution cardiac output determinations in 17 critically ill children. Commercially available equipment, specifically designed for this purpose, was employed. Forty paired thermodilution and pulsed Doppler determinations were made. There was a significant correlation between the two measurements (pulsed Doppler = 0.84 thermodilution + 0.39; r = 0.79, p less than 0.01). The ranges of thermodilution measurements (1.02 to 6.26 L/min; median 2.77 L/min) and pulsed Doppler measurements (1.13 to 6.35 L/min; median 2.57 L/min) were not different (p = 0.25). However, differences between individual paired thermodilution and pulsed Doppler measurements were large (-3.13 to 2.03 L/min; median 0.12 L/min), and the percentage difference between individual paired thermodilution and pulsed Doppler measurements ranged from 0.41% to 102.5% (median 12.7%). A discrepancy of 15% or more between thermodilution and pulsed Doppler was encountered in 18 (45%) of 40 of paired measurements (95% confidence interval: 29% to 61%), and one fourth of the paired measurements differed by more than 25%. We conclude that, as employed in this study, pulsed Doppler cardiac output determination is not sufficiently representative of the thermodilution output to be employed for hemodynamic monitoring in critically ill children.


S-allylcysteine induces cytotoxic effects in two human lung cancer cell lines via induction of oxidative damage, downregulation of Nrf2 and NF-κB, and apoptosis.
In this study, we investigated the putative cytotoxic effect elicited by the garlic-derived compound S-allylcysteine (SAC) in two human cancer cell lines (HCC827 and NCI-H1975) in order to develop an experimental approach to the therapeutic potential of this molecule for lung cancer. Cells were incubated for 24, 48 and 72 h in the presence of SAC (10 or 20 mM), which resulted in a concentration- and time-dependent decrease in cell viability and culture confluence in both cell lines. These effects were contrasted with - and validated through - those observed in an immortalized but nontumorigenic epithelial cell line from human bronchial epithelium (BEAS-2B, negative control) and an adenocarcinoma human alveolar basal epithelial cell line (A549, positive control). SAC (20 mM at 72 h) also increased the oxidative damage to lipids, augmented apoptosis, and decreased the expression of the nuclear factor erythroid 2-related factor 2 (Nrf2) and the nuclear factor kappa B (NF-κB) proteins in HCC827 and NCI-H1975 cells. Our results establish the efficacy of SAC in reducing malignant growth and proliferation of lung tumor cells. This effect is mediated by the induction of oxidative damage associated with the downregulation of Nrf2 and NF-κB and their corresponding signaling pathways.


Clinical and pathological disappearance of peritoneal dissemination in a patient with advanced gastric cancer receiving chemotherapy with S-1 and low-dose cisplatin.
A 54-year-old woman with severe abdominal distention suffered from massive ascites. Cytological examination revealed adenocarcinoma cells, leading to a diagnosis of peritonitis carcinomatosa. Gastrointestinal fiberscopy (GIF) resulted in a histological diagnosis of type 4 advanced gastric cancer with signet-ring cell carcinoma. The clinical diagnosis was confirmed to be cT3(SE)cN1cM0cH0cP1, cStage IV gastric cancer, type 4, according to the Japanese classification of gastric carcinoma. The patient was treated with S-1 and low-dose cisplatin (CDDP) in order to alleviate the critical state of the disease. After the third cycle of the regimen, the clinical response of P1 was classified as a partial response (PR) according to the World Health Organization (WHO) criteria. The patient's appetite loss and abdominal discomfort were markedly alleviated. The patient experienced grade 2 leukocytopenia throughout the regimen. Surgery was performed. Ascites and peritoneal disseminated lesions were not observed, and cytological examination of the peritoneal washes was negative. Total gastrectomy with D1 lymph node dissection was performed, and the surgical diagnosis was sT3(SE)sN0sM0sH0sP0; sStage II. Microscopically, viable cancer cells were found to be scattered throughout the subserosal-serosal layers in the resected stomach. All of the samples from lesions that were potentially cancers involving peritoneal dissemination were diagnosed as fibrous scar tissues without any viable cancer cells. The patient is alive without recurrence at 10 months after surgery and 14 months after the initial chemotherapy. Thus, systemic chemotherapy with S-1/low-dose CDDP achieved desirable control of peritoneal disseminated cells, as assessed microscopically, suggesting that the regimen may be an effective strategy for the treatment of advanced gastric cancer with peritonitis carcinomatosa.


Hand2 elevates cardiomyocyte production during zebrafish heart development and regeneration.
Embryonic heart formation requires the production of an appropriate number of cardiomyocytes; likewise, cardiac regeneration following injury relies upon the recovery of lost cardiomyocytes. The basic helix-loop-helix (bHLH) transcription factor Hand2 has been implicated in promoting cardiomyocyte formation. It is unclear, however, whether Hand2 plays an instructive or permissive role during this process. Here, we find that overexpression of hand2 in the early zebrafish embryo is able to enhance cardiomyocyte production, resulting in an enlarged heart with a striking increase in the size of the outflow tract. Our evidence indicates that these increases are dependent on the interactions of Hand2 in multimeric complexes and are independent of direct DNA binding by Hand2. Proliferation assays reveal that hand2 can impact cardiomyocyte production by promoting division of late-differentiating cardiac progenitors within the second heart field. Additionally, our data suggest that hand2 can influence cardiomyocyte production by altering the patterning of the anterior lateral plate mesoderm, potentially favoring formation of the first heart field at the expense of hematopoietic and vascular lineages. The potency of hand2 during embryonic cardiogenesis suggested that hand2 could also impact cardiac regeneration in adult zebrafish; indeed, we find that overexpression of hand2 can augment the regenerative proliferation of cardiomyocytes in response to injury. Together, our studies demonstrate that hand2 can drive cardiomyocyte production in multiple contexts and through multiple mechanisms. These results contribute to our understanding of the potential origins of congenital heart disease and inform future strategies in regenerative medicine.


Cirrhosis as a Comorbidity in Conditions Subject to the Hospital Readmissions Reduction Program.
Although the Hospital Readmissions Reduction Program (HRRP) has decreased readmissions in targeted conditions, outcomes in high-risk subgroups are unknown. This study analyzed the impact of cirrhosis as a comorbidity on readmissions in conditions subjected to the HRRP. Using a longitudinal analysis of the New York, Florida, and Washington State inpatient databases from 2009 to 2013, adult Medicare beneficiaries with a diagnosis-related group of targeted conditions by the HRRP-pneumonia, congestive heart failure (CHF), and myocardial infarction (MI)-were included. Exclusion criteria included inability to assess for readmission, previous liver transplant, or having a readmission not subject to penalty under the HRRP. A sensitivity analysis used the International Classification of Diseases, 9th Revision, Clinical Modification codes to identify pneumonia, CHF, and MI hospitalizations. The primary outcome was 30-day readmission, with secondary outcomes including 90-day readmission, trends, and cirrhosis-specific risk factors for readmission. Of the 797,432 patients included, 8,964 (1.1%) had cirrhosis. Patients with cirrhosis had significantly higher 30-day readmissions overall (29.3% vs 23.8%, P < 0.001) and specifically for pneumonia and CHF, but not for MI. Thirty-day readmission rates significantly decreased in patients without cirrhosis (annual percent change -1.8%, P < 0.001), but not in patients with cirrhosis (P = 0.39). Similar findings were present for 90-day readmissions. A sensitivity analysis confirmed these findings. On multivariable analysis, cirrhosis was associated with significantly higher 30-day readmissions (odds ratio 1.13, P < 0.001). When cirrhosis is comorbid in patients with conditions subjected to the HRRP, readmissions are higher and have not improved. Focused efforts are needed to improve outcomes in cirrhosis and other high-risk comorbidities within the HRRP cohort.


Development and application of small-subunit rRNA probes for assessment of selected Thiobacillus species and members of the genus Acidiphilium.
Culture-dependent studies have implicated sulfur-oxidizing bacteria as the causative agents of acid mine drainage and concrete corrosion in sewers. Thiobacillus species are considered the major representatives of the acid-producing bacteria in these environments. Small-subunit rRNA genes from all of the Thiobacillus and Acidiphilium species catalogued by the Ribosomal Database Project were identified and used to design oligonucleotide DNA probes. Two oligonucleotide probes were synthesized to complement variable regions of 16S rRNA in the following acidophilic bacteria: Thiobacillus ferrooxidans and T. thiooxidans (probe Thio820) and members of the genus Acidiphilium (probe Acdp821). Using (32)P radiolabels, probe specificity was characterized by hybridization dissociation temperature (T(d)) with membrane-immobilized RNA extracted from a suite of 21 strains representing three groups of bacteria. Fluorochrome-conjugated probes were evaluated for use with fluorescent in situ hybridization (FISH) at the experimentally determined T(d)s. FISH was used to identify and enumerate bacteria in laboratory reactors and environmental samples. Probing of laboratory reactors inoculated with a mixed culture of acidophilic bacteria validated the ability of the oligonucleotide probes to track specific cell numbers with time. Additionally, probing of sediments from an active acid mine drainage site in Colorado demonstrated the ability to identify numbers of active bacteria in natural environments that contain high concentrations of metals, associated precipitates, and other mineral debris.


Arterolane-piperaquine-mefloquine versus arterolane-piperaquine and artemether-lumefantrine in the treatment of uncomplicated Plasmodium falciparum malaria in Kenyan children: a single-centre, open-label, randomised, non-inferiority trial.
Triple antimalarial combination therapies combine potent and rapidly cleared artemisinins or related synthetic ozonides, such as arterolane, with two, more slowly eliminated partner drugs to reduce the risk of resistance. We aimed to assess the safety, tolerability, and efficacy of arterolane-piperaquine-mefloquine versus arterolane-piperaquine and artemether-lumefantrine for the treatment of uncomplicated falciparum malaria in Kenyan children. In this single-centre, open-label, randomised, non-inferiority trial done in Kilifi County Hospital, Kilifi, coastal Kenya, children with uncomplicated Plasmodium falciparum malaria were recruited. Eligible patients were aged 2-12 years and had an asexual parasitaemia of 5000-250 000 parasites per μL. The exclusion criteria included the presence of an acute illness other than malaria, the inability to tolerate oral medications, treatment with an artemisinin derivative in the previous 7 days, a known hypersensitivity or contraindication to any of the study drugs, and a QT interval corrected for heart rate (QTc interval) longer than 450 ms. Patients were randomly assigned (1:1:1), by use of blocks of six, nine, and 12, and opaque, sealed, and sequentially numbered envelopes, to receive either arterolane-piperaquine, arterolane-piperaquine-mefloquine, or artemether-lumefantrine. Laboratory staff, but not the patients, the patients' parents or caregivers, clinical or medical officers, nurses, or trial statistician, were masked to the intervention groups. For 3 days, oral artemether-lumefantrine was administered twice daily (target dose 5-24 mg/kg of bodyweight of artemether and 29-144 mg/kg of bodyweight of lumefantrine), and oral arterolane-piperaquine (arterolane dose 4 mg/kg of bodyweight; piperaquine dose 20 mg/kg of bodyweight) and oral arterolane-piperaquine-mefloquine (mefloquine dose 8 mg/kg of bodyweight) were administered once daily. All patients received 0·25 mg/kg of bodyweight of oral primaquine at hour 24. All patients were admitted to Kilifi County Hospital for at least 3 consecutive days and followed up at day 7 and, thereafter, weekly for up to 42 days. The primary endpoint was 42-day PCR-corrected efficacy, defined as the absence of treatment failure in the first 42 days post-treatment, of arterolane-piperaquine-mefloquine versus artemether-lumefantrine, and, along with safety, was analysed in the intention-to-treat population, which comprised all patients who received at least one dose of a study drug. The 42-day PCR-corrected efficacy of arterolane-piperaquine-mefloquine versus arterolane-piperaquine was an important secondary endpoint and was also analysed in the intention-to-treat population. The non-inferiority margin for the risk difference between treatments was -7%. The study is registered in ClinicalTrials.gov, NCT03452475, and is completed. Between March 7, 2018, and May 2, 2019, 533 children with P falciparum were screened, of whom 217 were randomly assigned to receive either arterolane-piperaquine (n=73), arterolane-piperaquine-mefloquine (n=72), or artemether-lumefantrine (n=72) and comprised the intention-to-treat population. The 42-day PCR-corrected efficacy after treatment with arterolane-piperaquine-mefloquine (100%, 95% CI 95-100; 72/72) was non-inferior to that after treatment with artemether-lumefantrine (96%, 95% CI 88-99; 69/72; risk difference 4%, 95% CI 0-9; p=0·25). The 42-day PCR-corrected efficacy of arterolane-piperaquine-mefloquine was non-inferior to that of arterolane-piperaquine (100%, 95% CI 95-100; 73/73; risk difference 0%). Vomiting rates in the first hour post-drug administration were significantly higher in patients treated with arterolane-piperaquine (5%, 95% CI 2-9; ten of 203 drug administrations; p=0·0013) or arterolane-piperaquine-mefloquine (5%, 3-9; 11 of 209 drug administrations; p=0·0006) than in patients treated with artemether-lumefantrine (1%, 0-2; three of 415 drug administrations). Upper respiratory tract complaints (n=26 for artemether-lumefantrine; n=19 for arterolane-piperaquine-mefloquine; n=23 for arterolane-piperaquine), headache (n=13; n=4; n=5), and abdominal pain (n=7; n=5; n=5) were the most frequently reported adverse events. There were no deaths. This study shows that arterolane-piperaquine-mefloquine is an efficacious and safe treatment for uncomplicated falciparum malaria in children and could potentially be used to prevent or delay the emergence of antimalarial resistance. UK Department for International Development, The Wellcome Trust, The Bill & Melinda Gates Foundation, Sun Pharmaceutical Industries.


Nanoscale, automated, high throughput synthesis and screening for the accelerated discovery of protein modifiers.
Hit finding in early drug discovery is often based on high throughput screening (HTS) of existing and historical compound libraries, which can limit chemical diversity, is time-consuming, very costly, and environmentally not sustainable. On-the-fly compound synthesis and in situ screening in a highly miniaturized and automated format has the potential to greatly reduce the medicinal chemistry environmental footprint. Here, we used acoustic dispensing technology to synthesize a library in a 1536 well format based on the Groebcke-Blackburn-Bienaymé reaction (GBB-3CR) on a nanomole scale. The unpurified library was screened by differential scanning fluorimetry (DSF) and cross-validated using microscale thermophoresis (MST) against the oncogenic protein-protein interaction menin-MLL. Several GBB reaction products were found as μM menin binder, and the structural basis of the interactions with menin was elucidated by co-crystal structure analysis. Miniaturization and automation of the organic synthesis and screening process can lead to an acceleration in the early drug discovery process, which is an alternative to classical HTS and a step towards the paradigm of continuous manufacturing.


Quantitative evaluation of spermatozoa ultrastructure after acupuncture treatment for idiopathic male infertility.
To evaluate the ultramorphologic sperm features of idiopathic infertile men after acupuncture therapy. Prospective controlled study. Christian-Lauritzen-Institut, Ulm, IVF center Munich, Germany, and Department of General Biology, University of Siena, Siena, Italy. Forty men with idiopathic oligospermia, asthenospermia, or teratozoospermia. Twenty eight of the patients received acupuncture twice a week over a period of 5 weeks. The samples from the treatment group were randomized with semen samples from the 12 men in the untreated control group. Quantitative analysis by transmission electron microscopy (TEM) was used to evaluate the samples, using the mathematical formula based on submicroscopic characteristics. Statistical evaluation of the TEM data showed a statistically significant increase after acupuncture in the percentage and number of sperm without ultrastructural defects in the total ejaculates. A statistically significant improvement was detected in acrosome position and shape, nuclear shape, axonemal pattern and shape, and accessory fibers of sperm organelles. However, specific sperm pathologies in the form of apoptosis, immaturity, and necrosis showed no statistically significant changes between the control and treatment groups before and after treatment. The treatment of idiopathic male infertility could benefit from employing acupuncture. A general improvement of sperm quality, specifically in the ultrastructural integrity of spermatozoa, was seen after acupuncture, although we did not identify specific sperm pathologies that could be particularly sensitive to this therapy.


Hepatic actinomycosis with immunoglobulin G4-related liver disease: A case report.
Immunoglobulin (Ig)G4-related pseudotumors of the liver are very rare diseases that are difficult to distinguish from malignant tumors. They can be usually improved by steroid therapy. Actinomycosis is a chronic, suppurative, granulomatous infection, for which immune suppression is a predisposing factor. It can also mimic malignant tumors. A 67-year-old man presented with mild abdominal discomfort and a 5-kg weight loss for 3 months. Initially, he visited another hospital and was treated with antibiotics under the assumption of a liver abscess. Symptom was not resolved. He diagnosed as having an IgG4-related pseudotumor of the right lobe of the liver after liver biopsy. Despite 2 months of steroid therapy, the liver mass was aggravated and invaded the right lung, as observed on follow-up computed tomography scan. We performed en bloc resection of the tumor under the assumption that it was a malignant tumor. The pathology of the tumor was revealed as actinomycotic colonies and IgG4-positive plasma cells of the liver. He recovered well and was discharged with ursodeoxycholic acid tablet for 14 days. After 3 months, he underwent postoperative follow-up CT and there was no remarkable finding in remnant left hepatic lobe. Hepatic actinomycosis and IgG4-related pseudotumors of the liver are both difficult to diagnose. As in our patient, combined diseases are more difficult to diagnose and to determine the optimal treatment. Since immunosuppression therapy of autoimmune diseases can cause and aggravate infection, management must be approached carefully. We can learn that various possibilities must be considered before diagnosing and treating a hepatic mass.


Photomorphogenesis.
As photoautotrophs, plants are exquisitely sensitive to their light environment. Light affects many developmental and physiological responses throughout plants' life histories. The focus of this chapter is on light effects during the crucial period of time between seed germination and the development of the first true leaves. During this time, the seedling must determine the appropriate mode of action to best achieve photosynthetic and eventual reproductive success. Light exposure triggers several major developmental and physiological events. These include: growth inhibition and differentiation of the embryonic stem (hypocotyl); maturation of the embryonic leaves (cotyledons); and establishment and activation of the stem cell population in the shoot and root apical meristems. Recent studies have linked a number of photoreceptors, transcription factors, and phytohormones to each of these events.


Probing battery chemistry with liquid cell electron energy loss spectroscopy.
We demonstrate the ability to apply electron energy loss spectroscopy (EELS) to follow the chemistry and oxidation states of LiMn2O4 and Li4Ti5O12 battery electrodes within a battery solvent. This is significant as the use and importance of in situ electrochemical cells coupled with a scanning/transmission electron microscope (S/TEM) has expanded and been applied to follow changes in battery chemistry during electrochemical cycling. We discuss experimental parameters that influence measurement sensitivity and provide a framework to apply this important analytical method to future in situ electrochemical studies.


Identification of Variants in RET and IHH Pathway Members in a Large Family With History of Hirschsprung Disease.
Hirschsprung disease (HSCR) is an inherited congenital disorder characterized by absence of enteric ganglia in the distal part of the gut. Variants in ret proto-oncogene (RET) have been associated with up to 50% of familial and 35% of sporadic cases. We searched for variants that affect disease risk in a large, multigenerational family with history of HSCR in a linkage region previously associated with the disease (4q31.3-q32.3) and exome wide. We performed exome sequencing analyses of a family in the Netherlands with 5 members diagnosed with HSCR and 2 members diagnosed with functional constipation. We initially focused on variants in genes located in 4q31.3-q32.3; however, we also performed an exome-wide analysis in which known HSCR or HSCR-associated gene variants predicted to be deleterious were prioritized for further analysis. Candidate genes were expressed in HEK293, COS-7, and Neuro-2a cells and analyzed by luciferase and immunoblot assays. Morpholinos were designed to target exons of candidate genes and injected into 1-cell stage zebrafish embryos. Embryos were allowed to develop and stained for enteric neurons. Within the linkage region, we identified 1 putative splice variant in the lipopolysaccharide responsive beige-like anchor protein gene (LRBA). Functional assays could not confirm its predicted effect on messenger RNA splicing or on expression of the mab-21 like 2 gene (MAB21L2), which is embedded in LRBA. Zebrafish that developed following injection of the lrba morpholino had a shortened body axis and subtle gut morphological defects, but no significant reduction in number of enteric neurons compared with controls. Outside the linkage region, members of 1 branch of the family carried a previously unidentified RET variant or an in-frame deletion in the glial cell line derived neurotrophic factor gene (GDNF), which encodes a ligand of RET. This deletion was located 6 base pairs before the last codon. We also found variants in the Indian hedgehog gene (IHH) and its mediator, the transcription factor GLI family zinc finger 3 (GLI3). When expressed in cells, the RET-P399L variant disrupted protein glycosylation and had altered phosphorylation following activation by GDNF. The deletion in GDNF prevented secretion of its gene product, reducing RET activation, and the IHH-Q51K variant reduced expression of the transcription factor GLI1. Injection of morpholinos that target ihh reduced the number of enteric neurons to 13% ± 1.4% of control zebrafish. In a study of a large family with history of HSCR, we identified variants in LRBA, RET, the gene encoding the RET ligand (GDNF), IHH, and a gene encoding a mediator of IHH signaling (GLI3). These variants altered functions of the gene products when expressed in cells and knockout of ihh reduced the number of enteric neurons in the zebrafish gut.


The impact of implementing a 24/7 open trauma bed protocol in the surgical intensive care unit on throughput and outcomes.
Increased emergency department (ED) length of stay (LOS) has been associated with increased mortality in trauma patients. In 2010, we implemented a 24/7 open trauma bed protocol in our designated trauma intensive care units (TICUs) to facilitate rapid admission from the ED. This required maintenance of a daily bump list and timely transferring of patients out of the TICU. We hypothesized that ED LOS and mortality would decrease after implementation. The following data from patients admitted directly from the ED to any ICU were retrospectively compared before (2009) and after (2011) the implementation of a trauma bed protocol at a Level I trauma center: age, sex, Glasgow Coma Scale (GCS) score, shock on admission (systolic blood pressure < 90 mm Hg), mechanism, injury severity scores (Injury Severity Score [ISS] and Abbreviated Injury Scale [AIS] score), ED LOS, ICU readmission rates, and mortality. Of the patients, 267 (17%) of 1,611 before and 262 (21%) of 1,266 (p < 0.01) after the protocol were admitted directly to the ICU, despite similar characteristics. ED LOS decreased from 4.2 ± 4.0 hours to 3.1 ± 2.1 hours (p < 0.01) in all patients as well as patients with an ISS of greater than 24 (3.1 ± 2.5 vs. 2.2 ± 1.6, p < 0.05) and a head AIS score of greater than 2 (4.2 ± 4.9 vs. 3.1 ± 2.0, p = 0.01). Mortality was unchanged for all patients (9% vs. 8%, p = 0.58) but trends toward improved mortality were found after protocol implementation inpatients with an ISS of greater than 24 (30% vs. 13%, p = 0.07) and in patients with a head AIS score of greater than 2 (12% vs. 6%, p = 0.08). A greater proportion of total patients were admitted to a designated TICU after implementation (83% vs. 93%, p < 0.01). ICU readmissions were unchanged (0.3% vs. 1.5%, p = 0.21). The implementation of a 24/7 open trauma bed protocol in the surgery ICU was associated with a decreased ED LOS and increased admissions to designated TICUs in all patients. Improved throughput was achieved without increases in ICU readmissions. Therapeutic study, level IV.


The mannitol-specific enzyme II (mtlA) gene and the mtlR gene of the PTS of Streptococcus mutans.
The phosphoenolpyruvate-dependent phosphotransferase system (PTS) is widely found among Gram-positive bacteria. It is the major source of carbohydrate transport in the dental pathogen Streptococcus mutans. The transported carbohydrates are fermented to produce large amounts of lactic acid which initiates dental caries. The authors have isolated the S. mutans gene for the mannitol-specific Enzyme II (EII) component of the PTS, mtlA, and the adjacent mtlR gene, which is located in the same operon. The mtlR gene is located between mtlA and the genes mtlF and mtlD. The nucleotide sequence of the mtlA and mtlR loci has been determined. The deduced mtlA gene product of S. mutans consists of 589 amino acids with a molecular mass of 62.0 kDa. It exhibits similarity with the mtlA gene products from other organisms. However, the similarity between these proteins is generally restricted to the 470 amino-terminal residues of the S. mutans protein. This region would correspond to the EIICB domains of the PTS. The authors have previously shown that the S. mutans mtlF gene product exhibits 76.6% similarity to the carboxyl-terminal 143 amino acids of the Escherichia coli mtlA product and that the mtlF gene encodes the EIIA domain of the PTS. Thus, the genes that encode the EIICB and the EIIA domains are separated by approximately 2250 bp. In many organisms, all of the EII domains may be fused together to form one molecule. The fact that these domains are separated by this distance in S. mutans supports the hypothesis that various functional domains of the PTS have been rearranged during evolution. The sequence of the 119 carboxyl-terminal amino acids of the S. mutans mtlA gene product also displays homology to the carboxyl-terminal end of the EIIB domain of various mannitol PTSs. Thus, this domain may have been duplicated in S. mutans during evolution of the operon. The mtlR gene is located in the same operon structure as mtlA but these loci are separated by an intragenic space. The precise 5' end of the mtlR locus cannot be determined either by in vitro transcription-translation assays or based upon nucleotide sequence analysis because of the apparent lack of a ribosome-binding site preceding the gene. The deduced mtlR gene product, which consists of approximately 650 amino acids with a molecular mass of 75.3 kDa, exhibits limited similarity to several potential transcriptional regulators. However, the exact function of this locus is currently unknown.


Activation of sickle red blood cell adhesion via integrin-associated protein/CD47-induced signal transduction.
Peripheral human red blood cells (RBCs) are not generally known to become activated and adhesive in response to cell signaling. We show, however, that soluble thrombospondin via integrin-associated protein (IAP; CD47) increases the adhesiveness of sickle RBCs (SS RBCs) by activating signal transduction in the SS RBC. This stimulated adhesion requires occupancy of IAP and shear stress and is mediated by the activation of large G proteins and tyrosine kinases. Reticulocyte-enriched RBCs derived from sickle-cell disease (SCD) patients are most responsive to IAP-induced activation. These studies therefore establish peripheral SS RBCs as signaling cells that respond to a novel synergy between IAP-induced signal transduction and shear stress, suggesting new therapeutic targets in SCD.


Factors associated with mental health outcomes across healthcare settings in Oman during COVID-19: frontline versus non-frontline healthcare workers.
This study aims to assess and compare demographic and psychological factors and sleep status of frontline healthcare workers (HCWs) in relation to non-frontline HCWs. This cross-sectional study was conducted from 8 April 2020 to 17 April 2020 using an online survey across varied healthcare settings in Oman accruing 1139 HCWs.The primary and secondary outcomes were mental health status and sociodemographic data, respectively. Mental health status was assessed using the Depression, Anxiety, and Stress Scale (DASS-21), and insomnia was evaluated by the Insomnia Severity Index. Samples were categorised into the frontline and non-frontline groups. χ2 and t-tests were used to compare groups by demographic data. The Mantel-Haenszel OR was used to compare groups by mental health outcomes adjusted by all sociodemographic factors. This study included 1139 HCWs working in Oman. While working during the pandemic period, a total of 368 (32.3%), 388 (34.1%), 271 (23.8%) and 211 (18.5%) respondents were reported to have depression, anxiety, stress and insomnia, respectively. HCWs in the frontline group were 1.5 times more likely to report anxiety (OR=1.557, p=0.004), stress (OR=1.506, p=0.016) and insomnia (OR=1.586, p=0.013) as compared with those in the non-frontline group. No significant differences in depression status were found between the frontline and non-frontline groups (p=0.201). To our knowledge, this is the first study to explore the differential impacts of the COVID-19 pandemic on different grades of HCWs. This study suggests that frontline HCWs are disproportionally affected compared to non-frontline HCWs, with managing sleep-wake cycles and anxiety symptoms being highly endorsed among frontline HCWs. As psychosocial interventions are likely to be constrained owing to the pandemic, mental healthcare must first be directed to frontline HCWs.


Host restriction, pathogenesis and chronic carriage of typhoidal Salmonella.
While conjugate vaccines against typhoid fever have recently been recommended by the World Health Organization for deployment, the lack of a vaccine against paratyphoid, multidrug resistance and chronic carriage all present challenges for the elimination of enteric fever. In the past decade, the development of in vitro and human challenge models has resulted in major advances in our understanding of enteric fever pathogenesis. In this review, we summarise these advances, outlining mechanisms of host restriction, intestinal invasion, interactions with innate immunity and chronic carriage, and discuss how this knowledge may progress future vaccines and antimicrobials.


Hypothalamic-pituitary-adrenal axis suppression in asthmatic school children.
Hypothalamic-pituitary-adrenal axis suppression (HPAS) when treating children with corticosteroids is thought to be rare. Our objective was to determine the prevalence of and predictive factors for various degrees of HPAS. Clinical features of HPAS, doses, adherence, asthma score, and lung functions were recorded in 143 asthmatic children. The overnight metyrapone test was performed if morning cortisol was >83 nmol/L. Spearman correlations coefficients (r) were calculated between 3 postmetyrapone outcomes and each continuous variable. A multiple linear regression model of √postmetyrapone adrenocorticotropic hormone (ACTH) and a logistic regression model for HPAS were developed. Hypocortisolemia was seen in 6.1% (1.8-10.5), hypothalamic-pituitary suppression (HPS) in 22.2% (14.5-29.9), adrenal suppression in 32.3% (23.7-40.9), HPAS in 16.3% (9.3-23.3), and any hypothalamic-pituitary-adrenal axis dysfunction in 65.1% (56.5-72.9). Log daily nasal steroid (NS) dose/m(2) was associated with HPAS in the logistic regression model (odds ratio = 3.7 [95% confidence interval: 1.1-13.6]). Daily inhaled corticosteroids (ICSs) + NS dose/m(2) predicted HPAS in the univariate logistic regression model (P = .038). Forced expiratory volume in 1 second/forced vital capacity <80% was associated with HPAS (odds ratio = 4.1 [95% confidence interval: 1.0-14.8]). Daily ICS + NS/m(2) dose was correlated with the postmetyrapone ACTH (r = -0.29, P < .001). BMI (P = .048) and percent adherence to ICS (P < .001) and NS (P = .002) were predictive of √postmetyrapone ACTH (R(2) = .176). Two-thirds of children on corticosteroids may have hypothalamic-pituitary-adrenal axis dysfunction. In one-third, central function had recovered but adrenal suppression persisted. Predictive factors for HPAS are NS use, BMI, and adherence to ICS and NS.


Phospho-Akt overexpression is prognostic and can be used to tailor the synergistic interaction of Akt inhibitors with gemcitabine in pancreatic cancer.
There is increasing evidence of a constitutive activation of Akt in pancreatic ductal adenocarcinoma (PDAC), associated with poor prognosis and chemoresistance. Therefore, we evaluated the expression of phospho-Akt in PDAC tissues and cells, and investigated molecular mechanisms influencing the therapeutic potential of Akt inhibition in combination with gemcitabine. Phospho-Akt expression was evaluated by immunohistochemistry in tissue microarrays (TMAs) with specimens tissue from radically-resected patients (n = 100). Data were analyzed by Fisher and log-rank test. In vitro studies were performed in 14 PDAC cells, including seven primary cultures, characterized for their Akt1 mRNA and phospho-Akt/Akt levels by quantitative-RT-PCR and immunocytochemistry. Growth inhibitory effects of Akt inhibitors and gemcitabine were evaluated by SRB assay, whereas modulation of Akt and phospho-Akt was investigated by Western blotting and ELISA. Cell cycle perturbation, apoptosis-induction, and anti-migratory behaviors were studied by flow cytometry, AnnexinV, membrane potential, and migration assay, while pharmacological interaction with gemcitabine was determined with combination index (CI) method. Immunohistochemistry of TMAs revealed a correlation between phospho-Akt expression and worse outcome, particularly in patients with the highest phospho-Akt levels, who had significantly shorter overall and progression-free-survival. Similar expression levels were detected in LPC028 primary cells, while LPC006 were characterized by low phospho-Akt. Remarkably, Akt inhibitors reduced cancer cell growth in monolayers and spheroids and synergistically enhanced the antiproliferative activity of gemcitabine in LPC028, while this combination was antagonistic in LPC006 cells. The synergistic effect was paralleled by a reduced expression of ribonucleotide reductase, potentially facilitating gemcitabine cytotoxicity. Inhibition of Akt decreased cell migration and invasion, which was additionally reduced by the combination with gemcitabine. This combination significantly increased apoptosis, associated with induction of caspase-3/6/8/9, PARP and BAD, and inhibition of Bcl-2 and NF-kB in LPC028, but not in LPC006 cells. However, targeting the key glucose transporter Glut1 resulted in similar apoptosis induction in LPC006 cells. These data support the analysis of phospho-Akt expression as both a prognostic and a predictive biomarker, for the rational development of new combination therapies targeting the Akt pathway in PDAC. Finally, inhibition of Glut1 might overcome resistance to these therapies and warrants further studies.


What shadows reveal about object structure.
In a scene observed from a fixed viewpoint, the set of shadow boundaries in an image changes as a point light source (nearby or at infinity) assumes different locations. We show that for any finite set of point light sources illuminating an object viewed under either orthographic or perspective projection, there is an equivalence class of object shapes having the same set of shadows. Members of this equivalence class differ by a four-parameter family of projective transformations, and the shadows of a transformed object are identical when the same transformation is applied to the light source locations. Under orthographic projection, this family is the generalized bas-relief (GBR) transformation, and we show that the GBR transformation is the only family of transformations of an object's shape for which the complete set of imaged shadows is identical. Finally, we show that given multiple images under differing and unknown light source directions, it is possible to reconstruct both an object's surface and the light source locations up to this family of transformations from the shadows alone.


GAF domains: cyclic nucleotides come full circle.
Cyclic nucleotides are ubiquitous signaling molecules that are present in life forms ranging from bacteria to yeast to humans. In higher eukaryotes, conserved molecular machinery processes signals through both cyclic adenosine monophosphate (AMP) and cyclic guanosine monophosphate (GMP). Hurley describes how one widespread cyclic nucleotide recognition module, the GAF (cyclic GMP, adenylyl cyclase, FhlA) domain, formerly thought to bind only cyclic GMP, has now been found to bind cyclic AMP as well. Recent structural analysis of a cyclic GMP-binding GAF domain shows how conserved elements among the cyclic nucleotide-binding subgroup of GAF domains recognize the common chemical moieties in the two compounds.


Chemoprevention of rat liver toxicity and carcinogenesis by Spirulina.
Spirulina platensis (SP) is a filamentous cyanobacterium microalgae with potent dietary phyto-antioxidant, anti-inflammatory and anti-cancerous properties. The present study aimed to investigate the chemopreventive effect of SP against rat liver toxicity and carcinogenesis induced by dibutyl nitrosamine (DBN) precursors, and further characterized its underlying mechanisms of action in HepG2 cell line. Investigation by light and electron microscopy showed that DBN treatment induced severe liver injury and histopathological abnormalities, which were prevented by SP supplementation. The incidence of liver tumors was significantly reduced from 80 to 20% by SP. Immunohistochemical results indicated that both PCNA and p53 were highly expressed in the liver of DBN-treated rats, but were significantly reduced by SP supplementation. Molecular analysis indicated that SP treatment inhibited cell proliferation, which was accompanied by increased p21 and decreased Rb expression levels at 48hrs post-treatment. In addition, SP increased Bax and decreased Bcl-2 expression, indicating induction of apoptosis by 48hrs. This is the first report of the in vivo chemopreventive effect of SP against DBN-induced rat liver cytotoxicity and carcinogenesis, suggesting its potential use in chemoprevention of cancer.


A study of time management: the correlation between video game usage and academic performance markers.
This study analyzes the correlation between video game usage and academic performance. Scholastic Aptitude Test (SAT) and grade-point average (GPA) scores were used to gauge academic performance. The amount of time a student spends playing video games has a negative correlation with students' GPA and SAT scores. As video game usage increases, GPA and SAT scores decrease. A chi-squared analysis found a p value for video game usage and GPA was greater than a 95% confidence level (0.005 < p < 0.01). This finding suggests that dependence exists. SAT score and video game usage also returned a p value that was significant (0.01 < p < 0.05). Chi-squared results were not significant when comparing time spent studying and an individual's SAT score. This research suggests that video games may have a detrimental effect on an individual's GPA and possibly on SAT scores. Although these results show statistical dependence, proving cause and effect remains difficult, since SAT scores represent a single test on a given day. The effects of video games maybe be cumulative; however, drawing a conclusion is difficult because SAT scores represent a measure of general knowledge. GPA versus video games is more reliable because both involve a continuous measurement of engaged activity and performance. The connection remains difficult because of the complex nature of student life and academic performance. Also, video game usage may simply be a function of specific personality types and characteristics.


Efficient presentation of soluble antigen by cultured human dendritic cells is maintained by granulocyte/macrophage colony-stimulating factor plus interleukin 4 and downregulated by tumor necrosis factor alpha.
Using granulocyte/macrophage colony-stimulating factor (GM-CSF) and interleukin 4 we have established dendritic cell (DC) lines from blood mononuclear cells that maintain the antigen capturing and processing capacity characteristic of immature dendritic cells in vivo. These cells have typical dendritic morphology, express high levels of major histocompatibility complex (MHC) class I and class II molecules, CD1, Fc gamma RII, CD40, B7, CD44, and ICAM-1, and lack CD14. Cultured DCs are highly stimulatory in mixed leukocyte reaction (MLR) and are also capable of triggering cord blood naive T cells. Most strikingly, these DCs are as efficient as antigen-specific B cells in presenting tetanus toxoid (TT) to specific T cell clones. Their efficiency of antigen presentation can be further enhanced by specific antibodies via FcR-mediated antigen uptake. Incubation of these cultured DCs with tumor necrosis factor alpha (TNF-alpha) or soluble CD40 ligand (CD40L) for 24 h results in an increased surface expression of MHC class I and class II molecules, B7, and ICAM-1 and in the appearance of the CD44 exon 9 splice variant (CD44-v9); by contrast, Fc gamma RII is markedly and sometimes completely downregulated. The functional consequences of the short contact with TNF-alpha are in increased T cell stimulatory capacity in MLR, but a 10-fold decrease in presentation of soluble TT and a 100-fold decrease in presentation of TT-immunoglobulin G complexes.


Detection of plasma EGFR mutations for personalized treatment of lung cancer patients without pathologic diagnosis.
Next-generation sequencing (NGS) and digital polymerase chain reaction (PCR) based platforms have been used to detect EGFR mutations in plasma circulating tumor DNA (ctDNA) with high accuracy. Generally, molecular testing is performed after histopathological analysis. However, many patients with suspected advanced nonsmall cell lung cancer are unable to undergo biopsy thus forgoing potential treatment with highly effective tyrosine kinase inhibitors (TKIs) in patients with sensitizing EGFR mutations. We examined the utility of ctDNA testing to detect EGFR mutations in patients' plasma, where tissue biopsy is not feasible. We conducted a single-center, prospective study of 30 Chinese patients with suspected advanced lung cancer, who were unable to undergo a biopsy for initial diagnosis due to comorbidities or poor performance status. Patients with plasma EGFR sensitizing mutations were treated with first-generation EGFR TKIs. Twenty of 30 patients enrolled had sensitizing EGFR mutations in ctDNA and were started on EGFR TKIs. After a median follow-up of 12 months, median progression-free survival (PFS) was 10 months and median overall survival (OS) was not reached. The median OS for the 10 untreated patients was 3 months. In our study, patients with plasma EGFR mutations treated with TKIs showed disease control rate (DCR) and PFS similar to historical controls that were treated based on tissue testing. This is the first prospective study showing that ctDNA genotyping provides a feasible diagnostic approach for frail lung cancer patients who are unable to undergo biopsy, which subsequently leads to EGFR-targeted therapy, and improved outcomes in this subgroup of patients.


Prevalence of multiple antibiotic resistance in 443 Campylobacter spp. isolated from humans and animals.
In view of recent findings that a multidrug efflux pump CmeABC exists in Campylobacter jejuni, 391 C. jejuni and 52 Campylobacter coli of human and animal origin were examined for a multidrug resistance phenotype. The MICs of ampicillin, chloramphenicol, ciprofloxacin, erythromycin, kanamycin, tetracycline, cetrimide, triclosan, acridine orange, paraquat and ethidium bromide were determined. Resistance to organic solvents and the effect of salicylate (known inducer of the marRAB operon in Escherichia coli and Salmonella) were also examined. Two C. coli and 13 C. jejuni isolates, mainly from pigs or poultry, were resistant to three or more antibiotics and 12 of these strains had reduced susceptibility to acridine orange and/or ethidium bromide. Strains (n = 20) that were less susceptible to acridine orange, ethidium bromide and triclosan were significantly more resistant (P < 0.05) to ampicillin, chloramphenicol, ciprofloxacin, erythromycin, nalidixic acid and tetracycline, with two- to four-fold increases in MIC values compared with strains (n = 20) most susceptible to acridine orange, ethidium bromide and triclosan. Growth of strains with 1 mM salicylate caused a small (up to two-fold) but statistically significant (P < or = 0.005) increase in the MICs of chloramphenicol, ciprofloxacin, erythromycin and tetracycline. These data indicate that multiple antibiotic resistant (MAR)-like Campylobacter strains occur and it may be postulated that these may overexpress cmeABC or another efflux system.


2-Aminopurine-modified DNA homopolymers for robust and sensitive detection of mercury and silver.
Heavy metal detection is a key topic in analytical chemistry. DNA-based metal recognition has advanced significantly producing many specific metal ligands, such as thymine for Hg2+ and cytosine for Ag+. For practical applications, however, robust sensors that can work in a diverse range of salt concentrations need to be developed, while most current sensing strategies cannot meet this requirement. In this work, 2-aminopurine (2AP) is used as a fluorescence label embedded in the middle of four 10-mer DNA homopolymers. 2AP can be quenched up to 98% in these DNA without an external quencher. The interaction between 2AP and all common metal ions is studied systematically for both free 2AP base and 2AP embedded DNA homopolymers. With such low background, Hg2+ induces up to 14-fold signal enhancement for the poly-T DNA, and Ag+ enhances up to 10-fold for the poly-C DNA. A detection limit of 3nM is achieved for both metals. With these four probes, silver and mercury can be readily discriminated from the rest. A comparison with other signaling methods was made using fluorescence resonance energy transfer, graphene oxide, and SYBR Green I staining, respectively, confirming the robustness of the 2AP label. Detection of Hg2+ in Lake Huron water was also achieved with a similar sensitivity. This work has provided a comprehensive fundamental understanding of using 2AP as a label for metal detection, and has achieved the highest fluorescence enhancement for non-protein targets.


Limb salvage in octogenarians and nonagenarians.
Although advanced age has often been a relative contraindication to attempts at limb salvage, we have not regarded it as an important deterrent to arterial reconstruction. Our 6-year experience with 168 consecutive patients over 80 years of age who underwent arterial reconstruction or percutaneous transluminal angioplasty represented 18% of all patients treated with limb-threatening ischemia during this period. The average age was 84 years, with 14 patients over 90 years of age. Sixty-eight patients were men (41%) and 100 were women (59%). Indications for treatment in 189 limbs were restricted to limb salvage. One hundred eighty-two operative procedures were performed consisting of 84 femoropopliteal, 72 femorotibial, 12 axillofemoral, 11 femorofemoral, two axillopopliteal and one iliofemoral bypass. Percutaneous transluminal angioplasty was performed in 12 iliac and 14 femoral or popliteal arteries as an alternative (seven) or adjunct (19) to vascular reconstruction. The 30-day procedural mortality rate was 6%. The cumulative life table survival rate of all patients who underwent an attempt at limb salvage was 78% at 1 year, 65% at 2 years, and 54% at 3 years. Cumulative life table limb salvage rates were 84% at 1 year, 74% at 2 years, and 71% at 3 years. Overall graft patency for 182 arterial reconstructive operations was 80% at 1 year and 62% at 3 years. Of patients in whom limb salvage was attempted, 65% lived more than 1 year and 51% more than 2 years with a functional limb. Of patients who died within 5 years of treatment, 76% did so with their previously threatened limb intact. These data support an aggressive approach to arterial reconstruction in elderly patients and indicate that advanced age alone should not be considered a contraindication to attempts at limb salvage.


From eye to face: The impact of face outline, feature number, and feature saliency on the early neural response to faces.
The LIFTED model of early face perception postulates that the face-sensitive N170 event-related potential may reflect underlying neural inhibition mechanisms which serve to regulate holistic and featural processing. It remains unclear, however, what specific factors impact these neural inhibition processes. Here, N170 peak responses were recorded whilst adults maintained fixation on a single eye using a gaze-contingent paradigm, and the presence/absence of a face outline, as well as the number and type of parafoveal features within the outline, were manipulated. N170 amplitudes and latencies were reduced when a single eye was fixated within a face outline compared to fixation on the same eye in isolation, demonstrating that the simple presence of a face outline is sufficient to elicit a shift towards a more face-like neural response. A monotonic decrease in the N170 amplitude and latency was observed with increasing numbers of parafoveal features, and the type of feature(s) present in parafovea further modulated this early face response. These results support the idea of neural inhibition exerted by parafoveal features onto the foveated feature as a function of the number, and possibly the nature, of parafoveal features. Specifically, the results suggest the use of a feature saliency framework (eyes > mouth > nose) at the neural level, such that the parafoveal eye may play a role in down-regulating the response to the other eye (in fovea) more so than the nose or the mouth. These results confirm the importance of parafoveal features and the face outline in the neural inhibition mechanism, and provide further support for a feature saliency mechanism guiding early face perception.


Adolescent deviant peer clustering as an amplifying mechanism underlying the progression from early substance use to late adolescent dependence.
Early substance use co-occurs with youths' self-organization into deviant peer groups in which substance use is central to social interaction. We hypothesized that the social dynamics of deviant peer groups amplify the risk of progressing from early use to later dependence, and that this influence occurs over and above escalations in use that typically accompany early substance use and membership in deviant groups. Our study used a longitudinal, multimethod dataset consisting of 998 adolescents and their families. Participants were recruited from middle schools in a large metropolitan area in the Pacific Northwest. The sample was 47.3% female and ethnically diverse (42.3% European American, 29.1% African American, and 28.6% other, including biracial). We examined deviant peer clustering as a mediator between early substance use and later dependence, controlling for proximal levels of use, SES, early antisocial behavior, and parental monitoring. Tobacco, alcohol, and marijuana use were assessed at ages 12, 13, and 16-17. Past-year nicotine, alcohol, and marijuana dependence (DSM-IV) was assessed at age 19. Youth and parent reports and observational data were used to assess deviant peer clustering at age 16-17, and youth reported on antisocial behavior and parental monitoring at ages 12 and 13. Early substance use predicted increased likelihood of dependence on tobacco, alcohol, and marijuana by late adolescence. Deviant peer affiliation mediated these links, even when accounting for proximal levels of substance use. Early substance use not only promotes escalations in use across adolescence but also provides entry into a deviant social context that contributes to increased risk of dependence. Our results emphasize the importance of identifying and intervening in early substance use before it becomes an organizing factor in friendship selection and interaction. Deviant peer clusters are clearly an important avenue for intervention when seeking to interrupt the progression to substance dependence.


Dendritic tree extraction from noisy maximum intensity projection images in C. elegans.
Maximum Intensity Projections (MIP) of neuronal dendritic trees obtained from confocal microscopy are frequently used to study the relationship between tree morphology and mechanosensory function in the model organism C. elegans. Extracting dendritic trees from noisy images remains however a strenuous process that has traditionally relied on manual approaches. Here, we focus on automated and reliable 2D segmentations of dendritic trees following a statistical learning framework. Our dendritic tree extraction (DTE) method uses small amounts of labelled training data on MIPs to learn noise models of texture-based features from the responses of tree structures and image background. Our strategy lies in evaluating statistical models of noise that account for both the variability generated from the imaging process and from the aggregation of information in the MIP images. These noisy models are then used within a probabilistic, or Bayesian framework to provide a coarse 2D dendritic tree segmentation. Finally, some post-processing is applied to refine the segmentations and provide skeletonized trees using a morphological thinning process. Following a Leave-One-Out Cross Validation (LOOCV) method for an MIP databse with available "ground truth" images, we demonstrate that our approach provides significant improvements in tree-structure segmentations over traditional intensity-based methods. Improvements for MIPs under various imaging conditions are both qualitative and quantitative, as measured from Receiver Operator Characteristic (ROC) curves and the yield and error rates in the final segmentations. In a final step, we demonstrate our DTE approach on previously unseen MIP samples including the extraction of skeletonized structures, and compare our method to a state-of-the art dendritic tree tracing software. Overall, our DTE method allows for robust dendritic tree segmentations in noisy MIPs, outperforming traditional intensity-based methods. Such approach provides a useable segmentation framework, ultimately delivering a speed-up for dendritic tree identification on the user end and a reliable first step towards further morphological characterizations of tree arborization.


Psychological distress in women with polycystic ovary syndrome from imam khomeini hospital, tehran.
Polycystic ovary syndrome (PCOS) is a complex, multifaceted, heterogeneous disorder, affecting 4%-18% of reproductive-aged women and it is associated with reproductive, metabolic and psychological dysfunctions. PCOS affects quality of life and can worsen anxiety and depression either due to the features of PCOS or due to the diagnosis of a chronic disease. In this descriptive-analytical study, 81 patients with PCOS were recruited from Vali-e-Asr Reproductive Health Research Center. A questionnaire with items related to pieces of information about stress was used for data collection. Stress symptoms were assessed using the Understanding Yourself questionnaire. Statistical analyses were performed using SPSS Ver. 13.0 (SPSS Inc., Chicago, ILL, USA). The data are presented as mean±SD or as frequency with percentages. A p-value less than 0.05 was considered as statistically significant. The descriptive results showed that 8 (9.9%) participants did not have any signs of stress, 32 (39.5%) had neurotic stress, 29 (35.8%) had high and 12 (14.8%) had extremely high levels of stress. The odds of high levels of anxiety in women with hirsutism was 3.1 (95% CI, 1.00-9.59). The odds of high levels of obsession in overweight patients was 3.2 (95% CI, 1.12-9.234). The odds of high levels of worries in patients with touchy personality was 3.4 (95% CI, 1.10 - 11.19) obsession score. The present study showed that clinical signs of PCOS were most closely associated with psychological distress which has important implications in the diagnosis and treatment of disorders.


DNA translocation through low-noise glass nanopores.
The effect of electron irradiation-induced shrinking on glass nanocapillaries with diameters ranging from 75 to 14 nm was analyzed by measuring the conductance characteristics with and without DNA translocation. We have investigated nanocapillary shrinking with a scanning electron microscope from several perspectives to understand the geometry of the shrunken nanocapillary. On the basis of this observation, the conductance was modeled with respect to the nanocapillary diameter, which allowed reproducing the experimental results. We then translocated DNA through the shrunken nanocapillaries and measured higher conductance drops for smaller diameters, reaching 1.7 nS for the 14 nm diameter nanocapillary. A model taking into account the conical shape of the shrunken nanocapillaries also supported this dependence. Next, we calculated the noise in the form of the standard deviation of the ionic conductance (between 0.04 and 0.15 nS) to calculate a signal-to-noise ratio (SNR) and compared it with nanopores embedded in 20 nm thick silicon nitride membranes. This shows that although nanocapillaries have smaller signal amplitudes due to their conical shape, they benefit from a lower noise. The glass nanocapillaries have a good SNR of about 25 compared with the SNR of 15 for smaller sized nanopores in silicon nitride membranes. The ability to use a modified model of nanopores to mimic the block conductance by DNA translocation provides a theoretical framework to support experimental results from translocating polymers such as DNA.


Utility of international normative 20 m shuttle run values for identifying youth at increased cardiometabolic risk.
The purpose of this study was to examine the ability of international normative centiles for the 20 m shuttle run test (20mSRT) to identify youth at increased cardiometabolic risk. This was a cross-sectional study involving 961 children aged 10-17 years (53% girls) from the United Kingdom. Receiver operating characteristic (ROC) curves determined the discriminatory ability of cardiorespiratory fitness percentiles for predicting increased cardiometabolic risk. ROC analysis demonstrated a significant but poor discriminatory accuracy of cardiorespiratory fitness in identifying low/high cardiometabolic risk in girls (AUC = 0.58, 95% CI: 0.54-0.63; p = 0.04), and in boys (AUC = 0.59, 95% CI: 0.54-0.63; p = 0.03). The cardiorespiratory fitness cut-off associated with high cardiometabolic risk was the 55th percentile (sensitivity = 33.3%; specificity = 84.5%) in girls and the 60th percentile (sensitivity = 42.9%; specificity = 73.6%) in boys. These 20mSRT percentile thresholds can be used to identify children and adolescents who may benefit from lifestyle intervention. Nonetheless, further work involving different populations and cardiometabolic risk scores comprising of different variables are needed to confirm our initial findings.


Enzymatic Treatment of Spent Green Tea Leaves and Their Use in High-Fibre Cookie Production.
By-products of food industry have been studied as sources of high fibre and antioxidant ingredients for healthy food products, because of their economic and environmental benefits. However, the soluble dietary fibre content of these materials is usually lower than the recommended value that is claimed to bring positive health effects. Enzymatic treatment could be an efficient method for modifying insoluble and soluble dietary fibre contents of these materials. The purpose of this study is to investigate the effects of enzymatic treatment conditions on soluble, insoluble and total dietary fibre mass fractions in spent green tea leaves, and evaluate the quality of dough and cookies when different mass fractions of untreated and treated leaves were added to the recipe. The mass fractions of soluble, insoluble and total dietary fibre in spent tea leaf powder was evaluated after the leaves were treated with cellulase amount of 0-25 U/g for 0 to 2 h. Wheat flour was replaced by untreated and treated spent tea leaf powder at 0, 10, 20, 30 and 40% in cookie formulation. Textural properties of dough, proximate composition, physical properties and overall acceptability of cookies were analysed. The appropriate conditions for enzymatic treatment were enzyme loading of 20 U/g and biocatalytic time of 1.5 h, under which the mass fraction of soluble dietary fibre in spent tea leaves increased by 144.5% compared to that of the control sample. The addition of spent tea leaves led to the increase in dough hardness. Increase in the spent tea leaf amount also enhanced fibre mass fraction, antioxidant activity and hardness of cookies but reduced their overall acceptability. Moreover, the enzymatic treatment of spent tea leaves improved the soluble to total dietary fibre ratio of the cookies, which influenced their textural properties and health benefits. The cookies with added 20% untreated or treated spent tea leaves were overall accepted by the panel. For the first time, spent tea leaves have been treated with enzymes to improve their soluble to total dietary fibre ratio. The treated spent tea leaves are a new promising high-fibre antioxidant ingredient for cookie preparation.


Protein subunit interfaces: A statistical analysis of hot spots in Sm proteins.
The distinguishing property of Sm protein associations is very high stability. In order to understand this property, we analyzed the interfaces and compared the properties of Sm protein interfaces with those of a test set, the Binding Interface Database (BID). The comparison revealed that the main differences between the interfaces of Sm proteins and those of the BID set are the content of charged residues, the coordination numbers of the residues, knowledge-based pair potentials, and the conservation scores of hot spots. In Sm proteins, the interfaces have more hydrophobic and fewer charged residues than the surfaces, which is also the case for the BID test set and other proteins. However, in the interfaces, the content of charged residues in Sm proteins (26%) is substantially larger than that in the BID set (22%). Hot spots are residues that make up a small fraction of the interfaces, but they contribute most of the binding energy. These residues are critical to protein-protein interactions. Analyses of knowledge-based pair potentials of hot spot and non-hot spot residues in Sm proteins show that they are significantly different; their mean values are 31.5 and 11.3, respectively. In the BID set, this difference is smaller; in this case, the mean values for hot spot and non-hot spot residues are 20.7 and 12.4, respectively. Hence, the pair potentials of hot spots differ significantly for the Sm and BID data sets. In the interfaces of Sm proteins, the amino acids are tightly packed, and the coordination numbers are larger in Sm proteins than in the BID set for both hot spots and non-hot spots. At the same time, the coordination numbers are higher for hot spots; the average coordination number of the hot spot residues in Sm proteins is 7.7, while it is 6.1 for the non-hot spot residues. The difference in the calculated average conservation score for hot spots and non-hot spots in Sm proteins is significantly larger than it is in the BID set. In Sm proteins, the average conservation score for the hot spots is 7.4. Hot spots are surrounded by residues that are moderately conserved (5.9). The average conservation score for the other interface residues is 5.6. The conservation scores in the BID set do not show a significant distinction between hot and non-hot spots: the mean values for hot and non-hot spot residues are 5.5 and 5.2, respectively. These data show that structurally conserved residues and hot spots are significantly correlated in Sm proteins.


Childhood gender nonconformity and harassment as predictors of suicidality among gay, lesbian, bisexual, and heterosexual Austrians.
The role of childhood gender role nonconformity (CGNC) and childhood harassment (CH) in explaining suicidality (suicide ideation, aborted suicide attempts, and suicide attempts) was examined in a sample of 142 lesbian, gay, and bisexual (LGB) adults and 148 heterosexual adults in Austria. Current and previous suicidality, CGNC, and CH were significantly greater in LGB participants compared to heterosexual participants. After controlling for CGNC, the effect of sexual orientation on CH diminished. CGNC correlated significantly with current suicidality in the LGB but not in the heterosexual group, and only non-significant correlations were found for CGNC with previous suicidality. Controlling for CH and CGNC diminished the effect of sexual orientation on current suicidality. Bayesian multivariate analysis indicated that current suicidality, but not previous suicidality, depended directly on CGNC. CH and CGNC are likely implicated in the elevated levels of current suicidality among adult LGB participants. As for previous suicidality, the negative impact of CGNC on suicidality might be overshadowed by stress issues affecting sexual minorities around coming out. The association of CGNC with current suicidality suggests an enduring effect of CGNC on the mental health and suicide risk of LGB individuals.


Consumerism: forcing medical practices toward patient-centered care.
Consumerism has been apart of many industries over the years; now consumerism may change the way many medical practices deliver healthcare. With the advent of consumer-driven healthcare, employers are shifting the decision-making power to their employees. Benefits strategies like health savings accounts and high-deductible insurance plans now allow the patients to control how and where they spend their money on medical care. Practices that seek to attract the more affluent and informed consumers are beginning to institute patient-centered systems designs that invite patients to actively participate in their healthcare. This article will outline the changes in the healthcare delivery system facing medical practices, the importance of patient-centered care, and six strategies to implement to change toward more patient-centered care.


Altering telomere structure allows telomerase to act in yeast lacking ATM kinases.
Telomerase is a ribonucleoprotein that copies a short RNA template into telomeric DNA, maintaining eukaryotic chromosome ends and preventing replicative senescence. Telomeres differentiate chromosome ends from DNA double-stranded breaks. Nevertheless, the DNA damage-responsive ATM kinases Tel1p and Mec1p are required for normal telomere maintenance in Saccharomyces cerevisiae. We tested whether the ATM kinases are required for telomerase enzyme activity or whether it is their action on the telomere that allows telomeric DNA synthesis. Cells lacking Tel1p and Mec1p had wild-type levels of telomerase activity in vitro. Furthermore, altering telomere structure in three different ways showed that telomerase can function in ATM kinase-deleted cells: tel1 mec1 cells senesced more slowly than tel1 mec1 cells that also lacked TLC1, which encodes telomerase RNA, suggesting that tel1 mec1 cells have residual telomerase function; deleting the telomere-associated proteins Rif1p and Rif2p in tel1 mec1 cells prevented senescence; we isolated a point mutation in the telomerase RNA template domain (tlc1-476A) that altered telomeric DNA sequences, causing uncontrolled telomeric DNA elongation and increasing single strandedness. In tel1 mec1 cells, tlc1-476A telomerase was also capable of uncontrolled synthesis, but only after telomeres had shortened for >30 generations. Our results show that, without Tel1p and Mec1p, telomerase is still active and can act in vivo when the telomere structure is disrupted by various means. Hence, a primary function of the ATM-family kinases in telomere maintenance is to act on the substrate of telomerase, the telomere, rather than to activate the enzymatic activity of telomerase.


Light-sheet microscopy-based 3D single-cell tracking reveals a correlation between cell cycle and the start of endoderm cell internalization in early zebrafish development.
Controlling the initiation of cell migration plays a fundamental role in shaping the tissue during embryonic development. During gastrulation in zebrafish, some mesendoderm cells migrate inward to form the endoderm as the innermost germ layer along the yolk syncytial layer. However, how the initiation of inward migration is regulated is poorly understood. In this study, we performed light-sheet microscopy-based 3D single-cell tracking consisting of (a) whole-embryo time-lapse imaging with light-sheet microscopy and (b) three-dimensional single cell tracking in the zebrafish gastrula in which cells are marked with histone H2A-mCherry (nuclei) and the sox17:EGFP transgene (expressed in endoderm cells). We analyzed the correlation between the timing of cell internalization and cell division. Most cells that differentiated into endoderm cells began to internalize during the first half of the cell cycle, where the length of a cell cycle was defined by the period between two successive cell divisions. By contrast, the timing of other internalized cells was not correlated with a certain phase of the cell cycle. These results suggest the possibility that cell differentiation is associated with the relationship between cell cycle progression and the start of internalization. Moreover, the 3D single-cell tracking approach is useful for further investigating how cell migration is integrated with cell proliferation to shape tissues in zebrafish embryos.


R5-SHIV induces multiple defects in T cell function during early infection of rhesus macaques including accumulation of T reg cells in lymph nodes.
HIV-1 is a pathogen that T cell responses fail to control. HIV-1gp120 is the surface viral envelope glycoprotein that interacts with CD4 T cells and mediates entry. HIV-1gp120 has been implicated in immune dysregulatory functions that may limit anti-HIV antigen-specific T cell responses. We hypothesized that in the context of early SHIV infection, immune dysregulation of antigen-specific T-effector cell and regulatory functions would be detectable and that these would be associated or correlated with measurable concentrations of HIV-1gp120 in lymphoid tissues. Rhesus macaques were intravaginally inoculated with a Clade C CCR5-tropic simian-human immunodeficiency virus, SHIV-1157ipd3N4. HIV-1gp120 levels, antigen-specificity, levels of apoptosis/anergy and frequency and function of Tregs were examined in lymph node and blood derived T cells at 5 and 12 weeks post inoculation. We observed reduced responses to Gag in CD4 and gp120 in CD8 lymph node-derived T cells compared to the peripheral blood at 5 weeks post-inoculation. Reduced antigen-specific responses were associated with higher levels of PD-1 on lymph node-derived CD4 T cells as compared to peripheral blood and uninfected lymph node-derived CD4 T cells. Lymph nodes contained increased numbers of Tregs as compared to peripheral blood, which positively correlated with gp120 levels; T regulatory cell depletion restored CD8 T cell responses to Gag but not to gp120. HIV gp120 was also able to induce T regulatory cell chemotaxis in a dose-dependent, CCR5-mediated manner. These studies contribute to our broader understanding of the ways in which HIV-1 dysregulates T cell function and localization during early infection.


MUC4 is a novel prognostic factor of extrahepatic bile duct carcinoma.
Many of the patients with extrahepatic bile duct carcinoma (EHBDC) show a poor outcome. We have reported that MUC4 is a novel prognostic factor of pancreatic adenocarcinoma and intrahepatic cholangiocarcinoma. The aim of this study is to evaluate the prognostic significance of MUC4 expression in EHBDC. We examined the expression profile of MUC4 in EHBDC tissues from 70 patients using immunohistochemistry. MUC4 is a membrane mucin like MUC1. In addition, MUC4 is an intramembrane ligand for receptor tyrosine kinase ErbB2 and is related with regulation of p27. We compared the MUC4 expression with MUC1, ErbB2, or p27 expression in EHBDC. MUC4 was expressed in 36 of the 70 patients with EHBDC. There was no significant correlation between the MUC4 expression and MUC1, ErbB2, or p27 expression. The survival of 19 patients with high MUC4 expression (>or=20% of carcinoma cells stained) was significantly worse than that of the 51 patients with low MUC4 expression (under 20% of carcinoma cells stained; P = 0.0072). The univariate analysis showed that high MUC4 expression (P = 0.0072), high MUC1 expression (P = 0.0092), histologic grading (P = 0.0029), surgical margin involvement (P = 0.0137), and nodal metastasis (P = 0.0036) were statistically significant risk factors. The backward stepwise multivariate analysis showed that high MUC4 expression (P = 0.0195) and surgical margin involvement (P = 0.0358) were statistically significant independent risk factors. MUC4 expression in EHBDC is a new independent factor for poor prognosis and predicts the outcome of patients with EHBDC.


Additive prognostic value of coronary flow reserve in patients with chest pain syndrome and normal or near-normal coronary arteries.
In patients with angiographically normal coronary arteries and chest pain, pharmacologic stress echocardiography can identify a subgroup of patients with a less benign prognosis. Coronary flow reserve (CFR) in the left anterior descending artery (LAD) can currently be combined with wall motion analysis during vasodilator stress echocardiography. The aim of this study was to assess the prognostic value of CFR response in patients with normal coronary arteries and normal wall motion during stress. We selected 394 patients (171 men, 61 +/- 11 years of age) who underwent dipyridamole stress echocardiography (0.84 mg/kg over 6 minutes) with 2-dimensional echocardiography and CFR evaluation of the LAD by Doppler. All had angiographically nonsignificant (<50% quantitatively assessed) stenosis in any major vessel, normal left ventricular function (wall motion score index 1), and test negativity for conventional wall motion criteria. Images were independently read by a core laboratory for wall motion and a core laboratory for CFR. Mean CFR was 2.5 +/- 0.6 and 87 patients (22%) had an abnormal CFR <2. During a median follow-up of 51 months, 31 events occurred, namely 4 deaths and 27 nonfatal myocardial infarctions (3 ST-elevated myocardial infarctions and 24 non-ST-elevated myocardial infarctions). Kaplan-Meier survival estimates for hard events showed a better outcome for those patients with a normal CFR compared with those with an abnormal CFR (96% vs 55%, p = 0.001, at 48 months of follow-up). In conclusion, in patients with angiographically normal or near-normal coronary arteries and preserved at-rest regional and global left ventricular function at baseline and during stress, CFR adds incremental value to the prognostic stratification achieved with clinical and angiographic data.