Involvement of site-specific FAK phosphorylation in sphingosine-1 phosphate- and thrombin-induced focal adhesion remodeling: role of Src and GIT.
Sphingosine-1 phosphate (S1P) and thrombin are agents with profound but divergent effects on vascular endothelial cell (EC) barrier properties. We have previously reported that S1P-induced focal adhesion (FA) remodeling involves interactions between focal adhesion kinase (FAK), paxillin, and G-protein-coupled receptor kinase-interacting proteins GIT1 and GIT2 and suggested a critical involvement of focal adhesions in the EC barrier regulation. In this study, we examined redistribution of FA proteins (FAK, paxillin, GIT1, and GIT2) and site-specific FAK tyrosine phosphorylation in human pulmonary artery endothelial cells stimulated with thrombin. In contrast to S1P, which we have shown to induce peripheral translocation of FA proteins associated with cortical actin ring formation, thrombin caused the redistribution of FA proteins to the ends of the newly formed massive stress fibers. S1P and thrombin induced distinct patterns of FAK site-specific phosphorylation with the FAK Y576 phosphorylation site targeted by SIP challenge and phosphorylation of three FAK sites (Y397, Y576, and Y925) in response to thrombin stimulation. Pharmacological inhibition of Src with Src-specific inhibitor PP2 abolished S1P-induced translocation of FA proteins, cortical actin ring formation, and FAK [Y576] phosphorylation. However, PP2 failed to alter thrombin-induced morphological changes and exhibited only partial inhibition of FAK site-specific tyrosine phosphorylation. These observations highlight the differential mechanisms of focal adhesion protein complex remodeling and FAK activation by S1P and thrombin and link differential FA remodeling to EC barrier regulation.


Music therapy-induced changes in salivary cortisol level are predictive of cardiovascular mortality in patients under maintenance hemodialysis.
Music therapy has been applied in hemodialysis (HD) patients for relieving mental stress. Whether the stress-relieving effect by music therapy is predictive of clinical outcome in HD patients is still unclear. We recruited a convenience sample of 99 patients on maintenance HD and randomly assigned them to the experimental (n=49) or control (n=50) group. The experimental group received relaxing music therapy for 1 week, whereas the control group received no music therapy. In the experimental group, we compared cardiovascular mortality in the patients with and without cortisol changes. The salivary cortisol level was lowered after 1 week of music therapy in the experimental group (-2.41±3.08 vs 1.66±2.11 pg/mL, P<0.05), as well as the frequency of the adverse reaction score (-3.35±5.76 vs -0.81±4.59, P<0.05), the severity of adverse reactions score (-1.93±2.73 vs 0.33±2.71, P<0.05), and hemodialysis stressor scale (HSS) score (-6.00±4.68 vs -0.877±7.08, P<0.05). The difference in salivary cortisol correlated positively with HD stress score scales (r=0.231, P<0.05), systolic blood pressure (r=0.264, P<0.05), and respiratory rates (r=0.369, P<0.05) and negatively with finger temperature (r=-0.235, P<0.05) in the total study population. The 5-year cardiovascular survival in the experimental group was higher in patients whose salivary cortisol lowered by <0.6 pg/mL than that in patients whose salivary cortisol lowered by >0.6 pg/mL (83.8% vs 63.6%, P<0.05). Providing music during HD is an effective complementary therapy to relieve the frequency and severity of adverse reactions, as well as to lower salivary cortisol levels. Differences in salivary cortisol after music therapy may predict cardiovascular mortality in patients under maintenance HD.


Double-blind placebo-controlled food challenges in children with alleged cow's milk allergy: prevention of unnecessary elimination diets and determination of eliciting doses.
Children with cow's milk allergy (CMA) need a cow's milk protein (CMP) free diet to prevent allergic reactions. For this, reliable allergy-information on the label of food products is essential to avoid products containing the allergen. On the other hand, both overzealous labeling and misdiagnosis that result in unnecessary elimination diets, can lead to potentially hazardous health situations. Our objective was to evaluate if excluding CMA by double-blind placebo-controlled food challenge (DBPCFC) prevents unnecessary elimination diets in the long term. Secondly, to determine the minimum eliciting dose (MED) for an acute allergic reaction to CMP in DBPCFC positive children. All children with suspected CMA under our care (Oct'05-Jun'09) were prospectively enrolled in a DBPCFC. Placebo and verum feedings were administered on two randomly assigned separate days. The MED was determined by noting the 'lowest observed adverse effect level' (LOAEL) in DBPCFC-positive children. Based on the outcomes of the DBPCFC a dietary advice was given. Parents were contacted by phone several months later about the diet of their child. 116 children were available for analysis. In 76 children CMA was rejected. In 60 of them CMP was successfully reintroduced, in 2 the parents refused introduction, in another 3 the parents stopped reintroduction. In 9 children CMA symptoms reappeared. In 40 children CMA was confirmed. Infants aged ≤ 12 months in our study group have a higher cumulative distribution of MED than older children. Excluding CMA by DBPCFC successfully stopped unnecessary elimination diets in the long term in most children. The MEDs form potential useful information for offering dietary advice to patients and their caretakers.


An investigation of attention, executive, and psychomotor aspects of cognitive fatigability.
Self-perceived mental fatigue is a common presenting symptom in many neurological diseases. Discriminating objective fatigability from self-perceived mental fatigue might facilitate neuropsychological diagnosis and treatment programs. However clinically valid neuropsychological instruments suitable for assessment of fatigability are still lacking. The prime aim of the study was to investigate aspects of cognitive fatigability and to identify properties of neuropsychological tests suitable to assess fatigability in patients with persistent cognitive complaints after mild brain injury. Another aim was to investigate whether cognitive fatigability captured by neuropsychological measures is influenced by depression or sleep disturbances. Twenty-four patients with persistent cognitive symptoms after mild traumatic brain injury (mTBI), (aged 18-51 years) and 31 healthy controls (aged 20-49 years) underwent neuropsychological testing measuring three cognitive fatigability domains: Attention fatigability was assessed using the Ruff 2 & 7 Selective Attention Test, executive fatigability using the Color Word Test (Stroop), and psychomotor fatigability using the Digit Symbol Substitution Test from the Wechsler Adult Intelligence Scale-Third Edition (WAIS-III). Subjective fatigue was measured using the Fatigue Severity Scale and a questionnaire of everyday consequences of fatigue. Depression was screened using the Hospital Anxiety and Depression Scale and sleep disturbances using the Pittsburgh Sleep Quality Index. The patients reported significantly more mental fatigue and performed worse on tests of psychomotor and executive fatigability than the healthy controls. Furthermore, the cognitive fatigability measures were not influenced by depression or sleep disturbances, as was the case in self-reported fatigue. Tests demanding executive or simultaneous processing of several neuropsychological functions seem most sensitive in order to capture cognitive fatigability. Clinical tests that can capture fatigability enable a deeper understanding of how fatigability might contribute to cognitive complaints and problems in maintaining daily activities.


Bacillus rhizoplanae sp. nov. from maize roots.
A Gram-stain-positive, aerobic and endospore-forming bacterial strain, isolated from the root surface of maize (Zea mays) was taxonomically studied. It could be clearly shown that, based on 16S rRNA gene sequence similarity comparisons, strain JJ-63T is a member of the genus Bacillus, most closely related to the type strain of Bacillus pseudomycoides (98.61%), followed by Bacillus cereus (98.47 %). Detailed phylogenetic analysis based on the 16S rRNA gene and the 87 proteins conserved within the phylum Firmicutes placed the strain into the Cereus clade. The average nucleotide identity, average amino acid identity and digital DNA-DNA hybridization values against the type strain of B. pseudomycoides were 80.97, 81.45 and 26.30 %, respectively. The quinone system of strain JJ-63T consisted exclusively of menaquinone MK-7. The polar lipid profile consisted of the major components diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol and an unidentified glycolipid. Major fatty acids were iso- and anteiso-branched with the major compounds iso-C15 : 0 and iso-C17 : 0. Also, the characteristic compounds C13 : 0 iso and C16 : 1 cis10 were found. Physiological and biochemical characteristics allowed a further phenotypic differentiation of strain JJ-63T from the most closely related species. For this reason, JJ-63T represents a novel species of the genus Bacillus, for which the name Bacillus rhizoplanae sp. nov. is proposed, with JJ-63T (=LMG 32091T=CCM 9090T=DSM 111827T= CIP 111899T) as the type strain.


From rhetoric to reality: consumer engagement in 16 multi-stakeholder alliances.
A key component of the Aligning Forces for Quality (AF4Q) program was engaging consumers in their health and healthcare. We examined the extent to which the alliances embraced 4 areas of consumer engagement: self-management, consumer friendliness of reports of healthcare provider quality, involvement of consumers in alliance governance, and the integration of consumers into quality improvement teams. We used a largely qualitative approach. The evaluation team conducted 1100 in-depth interviews with alliance stakeholders. Two authors reviewed the consumer engagement data for each alliance to assess its level of embrace in the 4 consumer engagement areas. For consumer friendliness of public reporting websites, we also assessed alliance public reports for reading level, technical language, and evaluable displays. Population-level effects were also examined for self-management and public reporting. Consumer engagement was new to most alliances, and few had staff with consumer engagement expertise or existing consumer constituencies. For each area of consumer engagement, some alliances enthusiastically embraced the work, other alliances made a concerted but limited effort to develop programs, and a third group of alliances did the minimum work required. Integrating consumers into governance was the area most often embraced, followed by making public reports consumer friendly. Two alliances strongly embraced both self-management and integrating patients into quality improvement efforts. The AF4Q program did not have greater population level effects from self-management or public reporting than were those observed in a national comparison sample. The AF4Q program sparked a few alliances to develop robust consumer engagement programming, while most alliances tried consumer engagement efforts for the first time and developed an appreciation for integrating consumer perspectives into their work.


Clinical use and molecular mechanisms of action of extract of Ginkgo biloba leaves in cardiovascular diseases.
Ginkgo biloba is one of the oldest living tree species that has been referred to as a living fossil. Extract from Ginkgo biloba leaves (GBE) is among the most commonly used herbal drugs and is popularized for its alleged tonic effect and possible curative and restorative properties. There is an increasing evidence of the potential role of GBE in treating cardiovascular diseases. We examined the history of GBE usage and reviewed the literature on its effects on the cardiovascular system. In the extensive studies involving cell cultures and animal models, GBE has been shown to exert its action through diverse mechanisms. GBE has been reported to have antioxidatant properties, to modify vasomotor function, to reduce adhesion of blood cells to endothelium, to inhibit activation of platelets and smooth muscle cells, to affect ion channels, and to alter signal transduction. In addition, relevant clinical trials with CBE are being carried out, particularly in the treatment of arterial and venous insufficiency and in the prevention of thrombosis. Finally, the controversial clinical findings and the possible adverse interactions between GBE and other drugs are discussed. This review underscores the potential benefits of Ginkgo biloba in cardiovascular diseases, highlights the gaps in our current research, and suggests the necessity for more rigorous systematic investigation of cardiovascular properties of CBE.


Agro-ecological compensation of watershed based on emergy.
The paper aims at providing a framework to calculate the agro-eco-compensation standard based on the systems agro-ecological concept of embodied energy as emergy. We have proposed a calculation system of eco-compensation standard for sustainable agricultural development based on the convertibility between emergy and price. According to our calculation, the total energy of agricultural production in Yongding River Basin was 3.45 E + 16 Sej/ha (the unit emergy value, expressed in solar emergy joules per unit) in terms of the renewable resources, nonrenewable resources, material inputs, and service costs. The energy of renewable and nonrenewable resources was 1.59 E + 16 and 1.86E + 16 Sej/ha, respectively. The ESI (environmental sustainability index) of the study area was 0.1056, indicating that its agricultural production was in a seriously unsustainable condition. To realize the sustainable agricultural production in the watershed, the downstream governments should pay $21.81 M (¥135 million) approximately to upstream governments for water and soil conservation. The results of the present study suggested that the emergy-based calculation method of agro-eco-compensation standard is feasible to a certain degree.


Rheum turkestanicum rhizomes possess anti-hypertriglyceridemic, but not hypoglycemic or hepatoprotective effect in experimental diabetes.
Rheum turkestanicum (R. turkestanicum) rhizomes have been used in Iranain traditional medicine as an anti-diabetic agent. The purpose of the present investigation was to evaluate the anti-diabetic and antioxidant activities of R. turkestanicum rhizome extract in streptozotocin-induced diabetic rats. Diabetes was induced by a single intraperitoneal injection of 55 mg/kg streptozotocin in male Wistar rats. Diabetic rats received the decoction extract of R. turkestanicum rhizomes at the doses of 200, 400 and 600 mg/kg daily by gavage for 3 weeks. Serum glucose and lipid levels were measured in all groups before diabetes induction and at the end of week 3. Oxidative stress was evaluated in the liver by measurement of malondialdehyde levels and total thiol concentration at the end of the experiment. A significant increase in serum glucose and triglyceride levels was observed in diabetic rats, which was accompanied by increased malondialdehyde levels and decreased total thiol concentration in the liver after 3 weeks. Treatment of diabetic rats with R. turkestanicum rhizome extract at the doses of 200, 400 and 600 mg/kg over a 3-week period did not change serum glucose, hepatic malondialdehyde and total thiol levels in diabetic rats. However, treatment with R. turkestanicum extract significantly decreased serum triglyceride levels in a dose-dependent manner at the end of the experiment. R. turkestanicum rhizome extract possess anti-hypertriglyceridemic, but not hypoglycemic or hepatoprotective effect in diabetic rats. Therefore, R. turkestanicum rhizome should be consumed with more caution by diabetic patients.


Essential role for the TRF2 telomere protein in adult skin homeostasis.
TRF2 is a component of shelterin, the protein complex that protects the ends of mammalian chromosomes. TRF2 is essential for telomere capping owing to its roles in suppressing an ATM-dependent DNA damage response (DDR) at chromosome ends and inhibiting end-to-end chromosome fusions. Mice deficient for TRF2 are early embryonic lethal. However, the role of TRF2 in later stages of development and in the adult organism remains largely unaddressed, with the exception of liver, where TRF2 was found to be dispensable for maintaining tissue function. Here, we study the impact of TRF2 conditional deletion in stratified epithelia by generating the TRF2(∆/∆) -K5-Cre mouse model, which targets TRF2 deletion to the skin from embryonic day E11.5. In marked contrast to TRF2 deletion in the liver, TRF2(∆/∆) -K5-Cre mice show lethality in utero reaching 100% lethality perinataly. At the molecular and cellular level, TRF2 deletion provokes induction of an acute DDR at telomeres, leading to activation of p53 signaling pathways and to programed cell death since the time of Cre expression at E11.5. Unexpectedly, neither inhibition of the NHEJ pathway by abrogation of 53BP1 nor inhibition of DDR by p53 deficiency rescued these severe phenotypes. Instead, TRF2 deletion provokes an extensive epidermal cell death accompanied by severe inflammation already at E16.5 embryos, which are independent of p53. These results are in contrast with conditional deletion of TRF1 and TPP1 in the skin, where p53 deficiency rescued the associated skin phenotypes, highlighting the comparatively more essential role of TRF2 in skin homeostasis.


Tolerance to musculoskeletal allografts with transient lymphocyte chimerism in miniature swine.
Although transplantation of musculoskeletal allografts in humans is technically feasible, the adverse effects of long-term immunosuppression subject the patient to high risks for correcting a non-life-threatening condition. Achieving immunologic tolerance to musculoskeletal allografts, without the need for chronic immunosuppression, could expand the clinical application of limb tissue allografting. Tolerance to musculoskeletal allografts has been accomplished previously in miniature swine in our laboratory. Although stable, mixed chimerism has been suggested as the mechanism underlying long-term tolerance in a rat limb model, the mechanism of this tolerance induction has not been established. This report explores the possible relationship between hematopoietic chimerism and tolerance to musculoskeletal allografts in swine. Twelve miniature swine underwent vascularized musculoskeletal allograft transplantation from histocompatibility complex (MHC) matched, minor antigen-mismatched donors. Eight animals received a 12-day coprse of cyclosporine, one of which was excluded due to subtherapeutic levels. Four recipients were not immunosuppressed. Serial biopsies to assess graft viability and flow cytometry to assess chimerism were performed. Donor and third-party skin grafts were placed on recipients with surviving allografts greater than 100 days to validate tolerance. Both groups developed early peripheral chimerism, but this chimerism became undetectable by postoperative day 19 in the cyclosporine group and by day 13 in the control group. Animals receiving cyclosporine developed permanent tolerance to their allografts, whereas those not receiving cyclosporine rejected their allografts in 6-9 weeks. Animals demonstrating tolerance to their bone allografts also demonstrated prolonged donor skin graft survival. Induction of tolerance to musculoskeletal allografts can be achieved in the MHC matched swine. Although hematopoietic chimerism is present in the immediate postoperative period, persistent, long-term chimerism does not seem to be necessary for maintenance of such tolerance.


Prevention of chemotherapy-induced neutropenia by special honey intake.
Febrile neutropenia is a serious side effect of chemotherapy. Colony-stimulating factors (CSFs) are used for primary and secondary treatment in patients with grade 4 neutropenia. The use of CSFs is expensive and accompanied by side effects. In the current study, Life-Mel Honey (LMH) was administered to prevent neutropenia and to reduce the need for CSFs in patients treated with chemotherapy. Thirty cancer patients receiving chemotherapy for primary or metastatic disease were included. All patients had grade 4 neutropenia and were treated with CSFs. The patients repeated the same chemotherapy schedule, with the addition of LMH for 5 d. Blood count was performed weekly. There was no recurrence of neutropenia after LMH intake and no need for treatment with CSFs in 12 (40%) of patients. Eighteen (60%) patients with LMH developed neutropenia grade 4 and were treated with CSFs (p=0.007). Hemoglobin levels remained >11 g/dL during LMH intake in 19 (64%) patients. Only three (10%) patients had thrombocytopenia. Eight (32%) patients reported improvement in quality of life. The use of LMH in patients who are at high risk of developing neutropenia as a result of chemotherapy decreases the risk of pancytopenia and the need for CSFs. LMH is inexpensive, has no side effects, and is easy to administer.


Expressional alterations in functional ultra-conserved non-coding RNAs in response to all-trans retinoic acid--induced differentiation in neuroblastoma cells.
Ultra-conserved regions (UCRs) are segments of the genome (≥ 200 bp) that exhibit 100% DNA sequence conservation between human, mouse and rat. Transcribed UCRs (T-UCRs) have been shown to be differentially expressed in cancers versus normal tissue, indicating a possible role in carcinogenesis. All-trans-retinoic acid (ATRA) causes some neuroblastoma (NB) cell lines to undergo differentiation and leads to a significant decrease in the oncogenic transcription factor MYCN. Here, we examine the impact of ATRA treatment on T-UCR expression and investigate the biological significance of these changes. We designed a custom tiling microarray to profile the expression of 481 T-UCRs in sense and anti-sense orientation (962 potential transcripts) in untreated and ATRA-treated neuroblastoma cell lines (SH-SY5Y, SK-N-BE, LAN-5). Following identification of significantly differentially expressed T-UCRs, we carried out siRNA knockdown and gene expression microarray analysis to investigate putative functional roles for selected T-UCRs. Following ATRA-induced differentiation, 32 T-UCRs were differentially expressed (16 up-regulated, 16 down-regulated) across all three cell lines. Further insight into the possible role of T-UC.300A, an independent transcript whose expression is down-regulated following ATRA was achieved by siRNA knockdown, resulting in the decreased viability and invasiveness of ATRA-responsive cell lines. Gene expression microarray analysis following knockdown of T-UC.300A revealed a number of genes whose expression was altered by changing T-UC.300A levels and that might play a role in the increased proliferation and invasion of NB cells prior to ATRA-treatment. Our results indicate that significant numbers of T-UCRs have altered expression levels in response to ATRA. While the precise roles that T-UCRs might play in cancer or in normal development are largely unknown and an important area for future study, our findings strongly indicate that the function of non-coding RNA T-UC.300A is connected with proliferation, invasion and the inhibition of differentiation of neuroblastoma cell lines prior to ATRA treatment.


Body composition measurements and risk of hematological malignancies: A population-based cohort study during 20 years of follow-up.
High body mass index (BMI) is associated with development of hematological malignancies (HMs). However, although BMI is a well-established measurement of excess weight, it does not fully reflect body composition and can sometimes misclassify individuals. This study aimed at investigating what body composition measurements had highest association with development of HM. Body composition measurements on 27,557 individuals recorded by healthcare professionals as part of the Malmö Diet and Cancer study conducted in Sweden between 1991-1996 were matched with data from national registers on cancer incidence and causes of death. Cox regression models adjusted for age and sex were used to test the association between one standard deviation increments in body composition measurements and risk of HM. During a median follow-up of 20 years, 564 persons developed an HM. Several body composition measurements were associated with risk of developing an HM, but the strongest association was found for multiple myeloma (MM). Waist circumference (HR 1.31, p = 0.04) and waist-hip ratio (HR 1.61, p = 0.05) had higher risk estimates than BMI (HR 1.18, p = 0.07) for MM. In conclusion, our study shows that measurements of abdominal adiposity better predict the risk of developing HM, particularly MM, compared to BMI.


Genetic variants associated with carboplatin-induced cytotoxicity in cell lines derived from Africans.
To gain a better understanding of the genetic variants associated with carboplatin-induced cytotoxicity in individuals of African descent, we present a step-wise approach integrating genotypes, gene expression, and sensitivity of HapMap cell lines to carboplatin. Cell lines derived from 30 trios of African descent (YRI) were used to develop a preclinical model to identify genetic variants and gene expression that contribute to carboplatin-induced cytotoxicity. Cytotoxicity was determined as cell growth inhibition at increasing concentrations of carboplatin for 72 h. Gene expression of 89 HapMap YRI cell lines was determined using the Affymetrix GeneChip Human Exon 1.0 ST Array. Single nucleotide polymorphism genotype and the percent survival at different treatment concentrations along with carboplatin IC50 were linked through whole genome association. A second association test was done between single nucleotide polymorphism genotype and gene expression, and linear regression was then used to capture those genes whose expression correlated to drug sensitivity phenotypes. This approach allows us to identify genetic variants that significantly associate with sensitivity to the cytotoxic effects of carboplatin through their effect on gene expression. We found a gene (GPC5) whose expression is important in all carboplatin treatment concentrations as well as many genes unique to either low (e.g., MAPK1) or high (e.g., BRAF, MYC, and BCL2L1) concentrations of drug. Our whole genome approach enables us to evaluate the contribution of genetic and gene expression variation to a wide range of cellular phenotypes. The identification of concentration specific genetic signatures allows for potential integration of pharmacokinetics, pharmacodynamics, and pharmacogenetics in tailoring chemotherapy.


Real time visualization and characterization of platelet deposition under flow onto clinically relevant opaque surfaces.
Although the thrombogenic nature of the surfaces of cardiovascular devices is an important aspect of blood biocompatibility, few studies have examined platelet deposition onto opaque materials used for these devices in real time. This is particularly true for the metallic surfaces used in current ventricular assist devices (VADs). Using hemoglobin depleted red blood cells (RBC ghosts) and long working distance optics to visualize platelet deposition, we sought to perform such an evaluation. Fluorescently labeled platelets mixed with human RBC ghosts were perfused across six opaque materials (a titanium alloy (Ti6Al4V), silicon carbide (SiC), alumina (Al2O3, 2-methacryloyloxyethyl phosphorylcholine polymer coated Ti6Al4V (MPC-Ti6Al4V), yttria partially stabilized zirconia (YZTP), and zirconia toughened alumina (ZTA)) for 5 min at wall shear rates of 400 and 1000 s(-1). Ti6Al4V had significantly increased platelet deposition relative to MPC-Ti6Al4V, Al2 O3 , YZTP, and ZTA at both wall shear rates (p < 0.01). For all test surfaces, increasing the wall shear rate produced a trend of decreased platelet adhesion. The described system can be a utilized as a tool for comparative analysis of candidate blood-contacting materials with acute blood contact.


Dual dopamine/serotonin releasers: potential treatment agents for stimulant addiction.
"Agonist therapy" for cocaine and methamphetamine addiction involves administration of stimulant-like medications (e.g., monoamine releasers) to reduce withdrawal symptoms and prevent relapse. A significant problem with this strategy is that many candidate medications possess abuse liability because of activation of mesolimbic dopamine (DA) neurons in the brain. One way to reduce DA-mediated abuse liability of candidate drugs is to add in serotonin (5-HT) releasing properties, since substantial evidence shows that 5-HT neurons provide an inhibitory influence over mesolimbic DA neurons. This article addresses several key issues related to the development of dual DA/5-HT releasers for the treatment of substance use disorders. First, the authors briefly summarize the evidence supporting a dual deficit in DA and 5-HT function during withdrawal from chronic cocaine or alcohol abuse. Second, the authors discuss data demonstrating that 5HT release can dampen DA-mediated stimulant effects, and the "antistimulant" role of 5-HT-sub(2C) receptors is considered. Next, the mechanisms underlying potential adverse effects of 5-HT releasers are described. Finally, the authors discuss recently published data with PAL-287, a novel nonamphetamine DA/5-HT releasing agent that suppresses cocaine self-administration but lacks positive reinforcing properties. It is concluded that DA/5-HT releasers could be useful therapeutic adjuncts for the treatment of cocaine and alcohol addictions, as well as for obesity, attention-deficit disorder, and depression.


Dynamics of protofibril elongation and association involved in Aβ42 peptide aggregation in Alzheimer's disease.
The aggregates of a protein called, 'Aβ' found in brains of Alzheimer's patients are strongly believed to be the cause for neuronal death and cognitive decline. Among the different forms of Aβ aggregates, smaller aggregates called 'soluble oligomers' are increasingly believed to be the primary neurotoxic species responsible for early synaptic dysfunction. Since it is well known that the Aβ aggregation is a nucleation dependent process, it is widely believed that the toxic oligomers are intermediates to fibril formation, or what we call the 'on-pathway' products. Modeling of Aβ aggregation has been of intense investigation during the last decade. However, precise understanding of the process, pre-nucleation events in particular, are not yet known. Most of these models are based on curve-fitting and overlook the molecular-level biophysics involved in the aggregation pathway. Hence, such models are not reusable, and fail to predict the system dynamics in the presence of other competing pathways. In this paper, we present a molecular-level simulation model for understanding the dynamics of the amyloid-β (Aβ) peptide aggregation process involved in Alzheimer's disease (AD). The proposed chemical kinetic theory based approach is generic and can model most nucleation-dependent protein aggregation systems that cause a variety of neurodegenerative diseases. We discuss the challenges in estimating all the rate constants involved in the aggregation process towards fibril formation and propose a divide and conquer strategy by dissecting the pathway into three biophysically distinct stages: 1) pre-nucleation stage 2) post-nucleation stage and 3) protofibril elongation stage. We next focus on estimating the rate constants involved in the protofibril elongation stages for Aβ42 supported by in vitro experimental data. This elongation stage is further characterized by elongation due to oligomer additions and lateral association of protofibrils (13) and to properly validate the rate constants involved in these phases we have presented three distinct reaction models. We also present a novel scheme for mapping the fluorescence sensitivity and dynamic light scattering based in vitro experimental plots to estimates of concentration variation with time. Finally, we discuss how these rate constants will be incorporated into the overall simulation of the aggregation process to identify the parameters involved in the complete Aβ pathway in a bid to understand its dynamics. We have presented an instance of the top-down modeling paradigm where the biophysical system is approximated by a set of reactions for each of the stages that have been modeled. In this paper, we have only reported the kinetic rate constants of the fibril elongation stage that were validated by in vitro biophysical analyses. The kinetic parameters reported in the paper should be at least accurate upto the first two decimal places of the estimate. We sincerely believe that our top-down models and kinetic parameters will be able to accurately model the biophysical phenomenon of Aβ protein aggregation and identify the nucleation mass and rate constants of all the stages involved in the pathway. Our model is also reusable and will serve as the basis for making computational predictions on the system dynamics with the incorporation of other competing pathways introduced by lipids and fatty acids.


Forced phage uncorking: viral DNA ejection triggered by a mechanically sensitive switch.
The foremost event of bacteriophage infection is the ejection of genomic material into the host bacterium after virus binding to surface receptor sites. How ejection is triggered is yet unknown. Here we show, in single mature T7 phage particles, that tapping the capsid wall with an oscillating atomic-force-microscope cantilever triggers rapid DNA ejection via the tail complex. The triggering rate increases exponentially as a function of force, following transition-state theory, across an activation barrier of 23 kcal mol-1 at 1.2 nm along the reaction coordinate. The conformation of the ejected DNA molecule revealed that it had been exposed to a propulsive force. This force, arising from intra-capsid pressure, assists in initiating the ejection process and the transfer of DNA across spatial dimensions beyond that of the virion. Chemical immobilization of the tail fibers also resulted in enhanced DNA ejection, suggesting that the triggering process might involve a conformational switch that can be mechanically activated either by external forces or via the tail-fiber complex.


Comparative evaluation of selected diagnostic assays for the detection of IgG and IgM antibody to Orientia tsutsugamushi in Thailand.
We compared the performance of 2 commercially available dipstick assays, 2 enzyme-linked immunosorbent assays (ELISAs), and an indirect immunofluorescent antibody (IFA) assay for the diagnosis of scrub typhus, using the indirect immunoperoxidase (IIP) test as the reference standard. The dipstick assays were the Integrated Diagnostics (Baltimore, MD) Dip-S-Ticks Scrub Recombinant (r56) dipstick test (INDX assay) and the PanBio (Brisbane, Australia) Scrub Typhus IgM and IgG Rapid Immunochromatographic test (PanBio assay). One of the ELISAs used pooled cell lysates of Karp, Kato, and Gilliam strain Orientia tsutsugamushi as antigen (pooled-antigen ELISA), and the other used a recombinant r56 protein as the antigen (recombinant ELISA). With a panel of 123 positive and 227 negative sera, sensitivity and specificity of the assays were as follows: INDX assay, IgG, 60% and 95%, IgM, 60% and 97%; PanBio assay, IgG, 94% and 96%, IgM, 83% and 93%; IFA (1:400 cutoff), IgG, 91% and 96%, IgM, 85% and 98%; pooled-antigen ELISA, IgG (1:1600 cutoff), 97% and 89%, IgM (1:400 cutoff), 94% and 91%; recombinant ELISA, IgG (1:1600 cutoff), 97% and 92%, IgM (1:400 cutoff), 93% and 94%. Because of its excellent performance and use of a standardized, commercially available antigen, the recombinant ELISA is suitable for use in a diagnostic laboratory, where it may be able to replace the IFA and IIP assays. In contrast, the PanBio dipstick assay was easy to perform and did not require sophisticated equipment, making it suitable for use in rural areas where more sophisticated diagnostic tests such as the ELISA and IFA may not be available.


The Caenorhabditis elegans JIP3 protein UNC-16 functions as an adaptor to link kinesin-1 with cytoplasmic dynein.
Kinesin-1 is a microtubule plus-end-directed motor that transports various cargos along the axon. Previous studies have elucidated the physical and genetic interactions between kinesin-1 and cytoplasmic dynein, a microtubule minus-end-directed motor, in neuronal cells. However, the physiological importance of kinesin-1 in the dynein-dependent retrograde transport of cargo molecules remains obscure. Here, we show that Caenorhabditis elegans kinesin-1 forms a complex with dynein via its interaction with UNC-16, which binds to the dynein light intermediate (DLI) chain. Both kinesin-1 and UNC-16 are required for localization of DLI-1 at the plus ends of nerve process microtubules. In addition, retrograde transport of APL-1 depends on kinesin-1, UNC-16, and dynein. These results suggest that kinesin-1 mediates the anterograde transport of dynein using UNC-16 as a scaffold and that dynein in turn mediates the retrograde transport of cargo molecules in vivo. Thus, UNC-16 functions as an adaptor for kinesin-1-mediated transport of dynein.


Promoting experiential learning through the use of high-fidelity human patient simulators in midwifery: A qualitative study.
The need to use innovative teaching and learning strategies in the nursing pedagogy is important in the 21st century. The challenges of clinical sites and opportunities for nursing students to gain clinical experience are a growing concern for many nurse educators. High-fidelity human patient simulators (HFHPS) are computerised mannequins that replicate a real-life patient, and when integrated into classroom teaching they allow students to become fully immersed into an almost real-life scenario. The aim of this study was to describe how HFHPS can promote experiential learning following the management of postpartum haemorrhage as a midwifery clinical emergency. A descriptive qualitative research approach was carried out in this study. The research setting was a local university in KwaZulu-Natal. The total population included all (N = 43) fourth-year baccalaureate of nursing undergraduate student midwives who participated as observers and/or role-players of a scenario role-play. An all-inclusive sampling was performed. There were 43 student midwives involved in the simulation teaching session with 6 of these students actively participating in each role-play at a time, while the remaining 37 observed. This occurred in two separate sessions and all the student midwives were involved in a debriefing session. These student midwives were then followed up and asked to participate in a focus group. The data in this article came from two separate focus groups which comprised 20 student midwives in total. Data were analysed using content analysis. Four categories emerged from the data, namely HFHPS offers a unique opportunity for student midwives to manage complex real-life emergencies; promotes reflection by allowing student midwives to reflect or review their roles, decisions and skills; allows student midwives to learn from their own experiences and encourages student midwives to try out what they learnt in a real-life situation. High-fidelity human patient simulators can be used in a complex case scenario to promote experiential learning of a clinical emergency.


Recruitment of chromatin remodelling factors during gene activation via the glucocorticoid receptor N-terminal domain.
We have shown that yeast mutants with defects in the Ada adaptor proteins are defective in hormone-dependent gene activation by ectopically expressed human glucocorticoid receptor (GR). Others have shown that the Ada2 protein is required for physical interactions between some activation domains and TBP (TATA-binding protein), whereas the Gcn5 (Ada4) protein has a histone acetyltransferase (HAT) activity. Although all HAT enzymes are able to acetylate histone substrates, some also acetylate non-histone proteins. Taken together, these observations suggest that the Ada proteins have the ability to effect different steps in the process of gene activation. It has recently been shown that the Ada proteins are present in two distinct protein complexes, the Ada complex and a larger SAGA complex. Our recent work has focused on determining (1) which of the Ada-containing complexes mediates gene activation by GR, (2) whether the HAT activity encoded by GCN5 is required for GR-dependent gene activation, (3) whether the Ada proteins contribute to GR-mediated activation at the level of chromatin remodelling and (4) how the role of these HAT complexes is integrated with other chromatin remodelling activities during GR-mediated gene activation. Our results suggest a model in which GR recruits the SAGA complex and that this contributes to chromatin remodelling via a mechanism involving the acetylation of histones. Furthermore, recruitment of the SWI/SNF remodelling complex also has a role in GR-mediated activation that is independent of the role of SAGA. These complexes are similar to analogous mammalian complexes and therefore these results are likely to be relevant to the human system.


Effects of Rising Temperature on the Growth, Stoichiometry, and Palatability of Aquatic Plants.
Global warming is expected to strengthen herbivore-plant interactions leading to enhanced top-down control of plants. However, latitudinal gradients in plant quality as food for herbivores suggest lower palatability at higher temperatures, but the underlying mechanisms are still unclear. If plant palatability would decline with temperature rise, then this may question the expectation that warming leads to enhanced top-down control. Therefore, experiments that directly test plant palatability and the traits underlying palatability along a temperature gradient are needed. Here we experimentally tested the impact of temperature on aquatic plant growth, plant chemical traits (including stoichiometry) and plant palatability. We cultured three aquatic plant species at three temperatures (15, 20, and 25°C), measured growth parameters, determined chemical traits and performed feeding trial assays using the generalist consumer Lymnaea stagnalis (pond snail). We found that rising temperature significantly increased the growth of all three aquatic plants. Plant nitrogen (N) and phosphorus (P) content significantly decreased, and carbon (C):N and C:P stoichiometry increased as temperature increased, for both Potamogeton lucens and Vallisneria spiralis, but not for Elodea nuttallii. By performing the palatability test, we found that rising temperatures significantly decreased plant palatability in P. lucens, which could be explained by changes in the underlying chemical plant traits. In contrast, the palatability of E. nuttallii and V. spiralis was not affected by temperature. Overall, P. lucens and V. spiralis were always more palatable than E. nuttallii. We conclude that warming generally stimulates aquatic plant growth, whereas the effects on chemical plant traits and plant palatability are species-specific. These results suggest that the outcome of the impact of temperature rise on macrophyte stoichiometry and palatability from single-species studies may not be broadly applicable. In contrast, the plant species tested consistently differed in palatability, regardless of temperature, suggesting that palatability may be more strongly linked to species identity than to intraspecific variation in plant stoichiometry.


Major and minor form of hereditary hyperekplexia.
Hyperekplexia is a hereditary neurological disorder characterized by excessive startle responses. Within the disorder two clinical forms can be distinguished. The major form is characterized by continuous generalized stiffness in the first year of life and an exaggerated startle reflex, accompanied by temporary generalized stiffness and falls, whereas in the minor form only excessive startle and hypnic jerks have been described. Mutations in the gene encoding the alpha-1 subunit of the glycine receptor (GLRA1) are responsible for the major form of hyperekplexia but no mutation was detected in patients with the minor form in the large Dutch pedigree originally described by Suhren and colleagues. Here we describe the genetic analysis of the GLRA1 gene of two English families in which both forms of hyperekplexia were present. Mutation analysis revealed no genetic defect in the GLRA1 gene in patients carrying either the minor or major forms. This is further evidence that the minor form of hyperekplexia is seldom due to a genetic defect in the GLRA1 gene.


Examining the factor structures of the five facet mindfulness questionnaire and the self-compassion scale.
The five facet mindfulness questionnaire (FFMQ; Baer, Smith, Hopkins, Krietemeyer, & Toney, 2006) and the self-compassion scale (SCS; Neff, 2003) are widely used measures of mindfulness and self-compassion in mindfulness-based intervention research. The psychometric properties of the FFMQ and the SCS need to be independently replicated in community samples and relevant clinical samples to support their use. Our primary aim was to establish the factor structures of the FFMQ and SCS in individuals with recurrent depression in remission, since mindfulness-based cognitive therapy (MBCT) was developed as a treatment for preventing depressive relapse. In order to determine the consistency across populations, we examined the factor structures of the FFMQ and SCS in 3 samples: (1) a convenience sample of adults, (2) a sample of adults who practice meditation, and (3) a sample of adults who suffer from recurrent depression and were recruited to take part in a trial of MBCT. Confirmatory factor analyses (CFAs) showed that a 4-factor hierarchical model of the FFMQ best fits the community sample and the clinical sample but that a 5-factor hierarchical model of the FFMQ best fits the meditator sample. CFA did not endorse the SCS 6-factor hierarchical structure in any of the 3 samples. Clinicians and researchers should be aware of the psychometric properties of the FFMQ to measure mindfulness when comparing meditators and nonmeditators. Further research is needed to develop a more psychometrically robust measure of self-compassion.


Post-partum hemorrhage and malpractice claims: what can we learn from the findings of placental examination and endometrial curettage? A retrospective analysis of surgical pathology reports.
This study reviews the surgical pathology reports of post-partum hemorrhages to support clinicians in malpractice litigation and, potentially, to enhance pregnancy-related diagnoses. This work is a retrospective study of surgical pathology reports of term pregnancies between January 2000 and January 2012 selected from the Surgical Pathology database of the I.R.C.C.S Azienda Ospedaliera Universitaria San Martino-IST (Istituto Nazionale per la Ricerca sul Cancro, Genoa). Ninety-five revision reports were identified (0.22% "placenta accrete," 0.46% "non-accreta placental tissue retention," and 0.31% "no placental fragments retention"). Secondary post-partum hemorrhages occurred in 0.3%, and primary PPH occurred in 0.05%, regardless of the group examined. Both types of PPH were most often associated with vaginal deliveries (58%). The most frequent endometrial finding was post-partum endometritis (43%). The entire placenta was submitted to the pathologist in 22/95 cases (23%). Hypermaturity and/or villous immaturity were the main histological patterns. This review supports the hypothesis that the pathological placenta abnormalities observed, rather than underlying myometrium abnormalities, may underlie the contractile failure and the incomplete removal of the placenta. For these reasons, the authors emphasized the importance of investigating the placenta in cases of complicated deliveries not associated with PPH to support clinicians in malpractice claims.


Mechanical Detection and Imaging of Hyperbolic Phonon Polaritons in Hexagonal Boron Nitride.
Mid-infrared nanoimaging and spectroscopy of two-dimensional (2D) materials have been limited so far to scattering-type scanning near-field optical microscopy (s-SNOM) experiments, where light from the sample is scattered by a metallic-coated atomic force microscope (AFM) tip interacting with the material at the nanoscale. These experiments have recently allowed imaging of plasmon polaritons in graphene as well as hyperbolic phonon polaritons in hexagonal boron nitride (hBN). Here we show that the high mechanical sensitivity of an AFM cantilever can be exploited for imaging hyperbolic phonon polaritons in hBN. In our imaging process, the lattice vibrations of hBN micrometer-sized flakes are locally enhanced by the launched phonon polaritons. These enhanced vibrations are coupled to the AFM tip in contact with the sample surface and recorded during scanning. Imaging resolution of Δ/20 is shown (Δ being the polaritonic fringes' separation distance), comparable to the best resolution in s-SNOM. Importantly, this detection mechanism is free from light background, and it is in fact the first photonless detection of phonon polaritons.


Allogeneic stem cell transplantation for myelofibrosis in 2012.
Myelofibrosis (MF) is a heterogeneous disease for which long-term, effective medical therapeutic options are currently limited. The role of allogeneic haematopoietic stem cell transplant (AHSCT) in this population, many of whom are elderly, often provides a challenge with regard to the identification of suitable candidates, timing of transplantation in the disease course and choice of conditioning regimen. This review summarizes key findings from published data concerning AHSCT in MF and attempts to provide a state of the art approach to MF-AHSCT in 2012. In addition, we postulate on how the era of JAK inhibition might impact on transplantation for MF.


Tris(ethanol-κO)tris-(picrato-κO,O)lanthanum(III) tri-2-pyridylamine solvate.
The title compound, [La(C(6)H(2)N(3)O(7))(3)(C(2)H(6)O)(3)]·C(15)H(12)N(4), has two mol-ecular building blocks, namely the neutral mononuclear adduct of lanthanum picrate with ethanol [i.e. La(pic)(3):EtOH (1:3); La(pic)(3) = lanthanum picrate and EtOH = ethanol] and the oligodentate aromatic nitro-gen base tri-2-pyridylamine (tpa). The asymmetric unit contains two formula units. The compound was prepared during an investigation of the stereochemistry of lanthanoid picrate complexes with O-donor ligands. The metal-ligand adduct adopts a nine-coordinate tricapped trigonal-prismatic metal atom environment. The stereochemical arrangement of the ligands about the metal core is typical of a fac-isomer with stoichiometry M(bidentate)(3)(monodentate)(3). Face-to-face hydrogen bonds are found between the tpa mol-ecule and the ethanol ligands. One ethanol ligand is disordered over two positions, with site occupancy factors of ca 0.7 and 0.3. The oxygen atoms of a nitro group are also disordered over two positions; the site occupancy factors are ca 0.6 and 0.4.


Multidetector CT findings of an extraordinary fenestration of the internal jugular vein.
Fenestration of the internal jugular vein is a rare malformation. Herein, the authors describe an extreme fenestration of the left internal jugular vein. This anomaly was found incidentally in a 47-year-old male patient undergoing multidetector computed tomography (MDCT) imaging and MDCT angiography evaluation for vertebral artery injury due to cervical trauma. MDCT angiography showed the presence of an extremely large fenestration in the left internal jugular vein.


The stability of Cα peptide radicals: why glycyl radical enzymes?
The conformational space of dipeptide models derived from glycine, alanine, phenylalanine, proline, tyrosine, and cysteine has been searched extensively and compared with the corresponding C(α) dipeptide radicals at the G3(MP2)-RAD level of theory. The results indicate that the (least-substituted) glycine dipeptide radical is the thermochemically most stable of these species. Analysis of the structural parameters indicates that this is due to repulsive interactions between the C(α) substituents and peptide units in the radical. A comparison of the conformational preferences of dipeptide radicals and their closed-shell parents also indicates that radical stability is a strongly conformation-dependent property.


Mycobacterium tuberculosis expresses ftsE gene through multiple transcripts.
Bacterial FtsE gene codes for the ATP-binding protein, FtsE, which in complex with the transmembrane protein, FtsX, participates in diverse cellular processes. Therefore, regulated expression of FtsE and FtsX might be critical to the human pathogen, Mycobacterium tuberculosis, under stress conditions. Although ftsX gene of M. tuberculosis (MtftsX) is known to be transcribed from a promoter inside the upstream gene, ftsE, the transcriptional status of ftsE gene of M. tuberculosis (MtftsE) remains unknown. Therefore, the authors initiated transcriptional analyses of MtftsE, using total RNA from M. tuberculosis cells that were grown under stress conditions, which the pathogen is exposed to, in granuloma in tuberculosis patients. Primer extension experiments showed the presence of putative transcripts, T1, T2, T3, and T4. T1 originated from the intergenic region between the upstream gene, MRA_3135, and MtftsE. T2 and T3 were found initiated from within MRA_3135. T4 was transcribed from a region upstream of MRA_3135. RT-PCR confirmed co-transcription of MRA_3135 and MtftsE. The cloned putative promoter regions for T1, T2, and T3 elicited transcriptional activity in Mycobacterium smegmatis transformants. T1, T2, and T3, but no new transcript, were present in the M. tuberculosis cells that were grown under the stress conditions, which the pathogen is exposed to in granuloma in tuberculosis patients. It showed lack of modulation of MtftsE transcripts under the stress conditions tested, indicating that ftsE may not have a stress response-specific function in M. tuberculosis.


Contrasts between habitat generalists and specialists: an empirical extension to the basic metacommunity framework.
Emergence of the metacommunity concept has made a substantial contribution to better understanding of the community composition and dynamics in a regional context. However, long-term field data for testing of available metacommunity models are still scarce, and the extent to which these models apply to the real world remains unknown. Tests conducted so far have largely sought to fit data on the entire regional set of species to one of several metacommunity models, implicitly assuming that all species operate similarly over the same set of sites. However, species differ in their habitat use. These differences can, in the most general terms, be expressed as a gradient of habitat specialization (ranging from habitat specialists to habitat generalists). We postulate that such differences in habitat specialization will have implications for metacommunity dynamics. Specifically, we predict that specialists respond more to local processes and generalists respond to regional spatial processes. We tested these predictions using natural microcosm communities for which long-term (nine-year) environmental and population dynamics data were available. We used redundancy analysis to determine the proportion of variation explained by environmental and spatial factors. We repeated this analysis to explain variation in the entire regional set of species, in generalist species only, and in specialists only. We further used ANOVA to test for differences in the proportions of explained variation. We found that habitat specialists responded primarily to environmental factors and habitat generalists responded mainly to spatial factors. Thus, from the metacommunity perspective, the dynamics of habitat specialists are best explained by a combination of species sorting and mass effects, while that of habitat generalists are best explained by patch dynamics and neutral models. Consequently, we infer that a natural metacommunity can exhibit complicated dynamics, with some groups of species (e.g., habitat specialists) governed according to environmental processes and other groups (e.g., habitat generalists) governed mainly by dispersal processes.


A cohort study of coagulation parameters and the use of blood products in surgery of the thoracic and thoracoabdominal aorta.
We sought to study the hemostatic profile and the use of blood products in patients undergoing thoracic and thoracoabdominal aortic aneurysm repair. This is a cohort study comparing three groups of patients: 7 undergoing elective acute normovolemic hemodilution (ANH); 15 undergoing elective procedures without ANH (non-ANH); and 8 undergoing repair of ruptured thoracoabdominal aneurysms. A control group of 10 patients was used for comparison of preoperative hemostasis. The parameters studied were platelet concentration, partial thromboplastin time (PTT), and international normalized ratio (before and after surgery), packed red blood cells, fresh frozen plasma, platelets, cryoprecipitates, donor exposure, and use of desmopressin and epsilon-aminocaproic acid. Analysis of variance and multiple stepwise regression analysis were used. Before operation the patients with aneurysms had PTTs prolonged compared with control subjects (P <.05). After operation the ANH group had higher platelet counts than the ruptured group (P =.001) and higher platelet counts (P =.05) and lower PTTs (P =.001) than the non-ANH group. The ANH group was transfused fewer platelets than the non-ANH group (P =.001) and less of every blood product than the ruptured group (P =.05); statistically significant differences were not observed for packed red blood cells, fresh frozen plasma, or platelets. The ANH group was exposed to an average of 65 donors fewer than the ruptured group (P <.001) and 34 fewer than the non-ANH group (P <.05). These differences could not be explained by baseline coagulation status or by the intraoperative use of desmopressin or epsilon-aminocaproic acid. The coagulation abnormality identified before surgery is that of higher PTT values, suggesting a disturbance of the extrinsic coagulation pathway. Blood losses, donor exposure, and platelet use were highest in the ruptured group and lowest in the ANH group. After surgery the ruptured group exhibited the worst coagulation parameters, and the ANH group exhibited the best with higher platelet count and lower PTT values than the other groups. The ANH technique appears to be an useful adjunct in the anesthetic management of these patients.


Leydig cell numbers, daily sperm production, and serum gonadotropin levels in aging men.
Age-related changes in Leydig cell number, daily sperm production, and circulating hormone levels were studied in 15 men between 20 and 48 yr of age and 15 men between 50 and 76 yr of age. Testes and blood samples were obtained at autopsy less than 15 h after death due to trauma or heart attack. Leydig cell number was determined by quantitative histometric estimation of the proportion of glutaraldehyde-perfused, decapsulated testicular parenchyma occupied by Leydig cell nuclei of measured average volume in both testes of each subject. Daily sperm production was determined by phase contrast cytometry of round spermatid nuclei in homogenates of both fixed testes from each individual. LH, FSH, PRL, and testosterone in serum from the heart or large veins were quantified by RIA. No relationship was detected between any of the testicular or hormonal values and postmortem time. The average total number of Leydig cells was reduced by 44% in the older men and was negatively correlated with age (p = -0.62) in all men. Mean serum LH was elevated more than 2-fold in the older men and was positively correlated with age (p = +0.53) in all men. Neither serum testosterone nor serum PRL changed significantly with age. Daily sperm production was more than 50% lower in older men and was negatively correlated with age (p = -0.76) in all men. Serum FSH was more than 3-fold higher in the older men and was positively correlated with age (p = +0.72) in all men. The highest FSH levels were found in men with the lowest rates of sperm production, and FSH and daily sperm production were inversely correlated even after the effects of age were removed. These findings show that the response of the human testis to aging is variable and that the predictive value of age for most testicular characteristics is weak at the level of individual men. Nevertheless, age accounts for more than a third of the variation in Leydig cell number, and it explains more than half the variation in daily sperm production. Hence, age is the largest single contributing factor yet identified in the search for explanations underlying the variation in testicular structure and function found in groups of normal men.


Variants in KCNJ11 and BAD do not predict response to ketogenic dietary therapies for epilepsy.
In the absence of specific metabolic disorders, predictors of response to ketogenic dietary therapies (KDT) are unknown. We aimed to determine whether variants in established candidate genes KCNJ11 and BAD influence response to KDT. We sequenced KCNJ11 and BAD in individuals without previously-known glucose transporter type 1 deficiency syndrome or other metabolic disorders, who received KDT for epilepsy. Hospital records were used to obtain demographic and clinical data. Two response phenotypes were used: ≥ 50% seizure reduction and seizure-freedom at 3-month follow-up. Case/control association tests were conducted with KCNJ11 and BAD variants with minor allele frequency (MAF)>0.01, using PLINK. Response to KDT in individuals with variants with MAF<0.01 was evaluated. 303 Individuals had KCNJ11 and 246 individuals had BAD sequencing data and diet response data. Six SNPs in KCNJ11 and two in BAD had MAF>0.01. Eight variants in KCNJ11 and seven in BAD (of which three were previously-unreported) had MAF<0.01. No significant results were obtained from association analyses, with either KDT response phenotype. P-values were similar when accounting for ethnicity using a stratified Cochran-Mantel-Haenszel test. There did not seem to be a consistent effect of rare variants on response to KDT, although the cohort size was too small to assess significance. Common variants in KCNJ11 and BAD do not predict response to KDT for epilepsy. We can exclude, with 80% power, association from variants with a MAF of >0.05 and effect size >3. A larger sample size is needed to detect associations from rare variants or those with smaller effect sizes.


Characterization of T cell differentiation in the murine gut.
Gut intraepithelial CD8 T lymphocytes (T-IEL) are distinct from thymus-derived cells and are thought to derive locally from cryptopatch (CP) precursors. The intermediate stages of differentiation between CP and mature T-IEL were not identified, and the local differentiation process was not characterized. We identified and characterized six phenotypically distinct lineage-negative populations in the CP and the gut epithelium: (a) we determined the kinetics of their generation from bone marrow precursors; (b) we quantified CD3-epsilon, recombination activating gene (Rag)-1, and pre-Talpha mRNAs expression at single cell level; (c) we characterized TCR-beta, -gamma, and -alpha locus rearrangements; and (d) we studied the impact of different mutations on the local differentiation. These data allowed us to establish a sequence of T cell precursor differentiation in the gut. We also observed that the gut differentiation varied from that of the thymus by a very low frequency of pre-Talpha chain mRNA expression, a different kinetics of Rag-1 mRNA expression, and a much higher impact of CD3 epsilon/delta and pre-Talpha deficiencies. Finally, only 3% of CP cells were clearly involved in T cell differentiation, suggesting that these structures may have additional physiological roles in the gut.


A single-nucleotide polymorphism in the human ENTPD1 gene encoding CD39 is associated with worsened graft-versus-host disease in a humanized mouse model.
Regulatory T cells (Tregs) protect against graft-versus-host disease (GVHD), a life-threatening complication of allogeneic hematopoietic stem cell transplantation. The ectoenzyme CD39 is important for increasing the immunosuppressive function of Tregs. The rs10748643 (A → G) single-nucleotide polymorphism (SNP) in intron 1 of the human ENTPD1 gene is associated with increased proportions of CD39+ Tregs. This study aimed to determine whether the rs10748643 SNP corresponded to increased proportions of CD39+ Tregs in an Australian donor population, and whether this SNP influences clinical GVHD in a humanized mouse model. Donors were genotyped for the rs10748643 SNP by Sanger sequencing, and the proportion of CD39+ T cells in donor peripheral blood was determined by flow cytometry. Donors encoding the G allele (donorsAG/GG ) demonstrated higher proportions of CD39+ CD3+ CD4+ CD25+ CD127lo Tregs, but not CD39+ CD3+ CD8+ T cells or CD39+ CD3+ CD4+ conventional T cells, compared with donors homozygous for the A allele (donorsAA ). NOD-SCID-IL2Rγnull mice were injected with human peripheral blood mononuclear cells from either donorsAA (hCD39AA mice) or donorsAG/GG (hCD39AG/GG mice). hCD39AG/GG mice demonstrated significantly greater weight loss and GVHD clinical scores, and significantly reduced survival, compared with hCD39AA mice. hCD39AG/GG mice showed significantly higher hCD4+ :hCD8+ T-cell ratios than hCD39AA mice, but displayed similar proportions of CD3+ hCD4+ hCD25+ hCD127lo Tregs and hCD39+ Tregs. However, the proportion of human Tregs corresponded to survival in hCD39AA mice, but not in hCD39AG/GG mice. This study demonstrates that donors encoding the G allele show higher percentages of CD39+ Tregs, but cause worsened GVHD in humanized mice compared with donors homozygous for the A allele.


Serum Albumin Levels and Economic Status in Japanese Older Adults.
Low serum albumin levels are associated with aging and medical conditions such as cancer, liver dysfunction, inflammation, and malnutrition and might be an independent predictor of long-term mortality in healthy older populations. We tested the hypothesis that economic status is associated with serum albumin levels and explained by nutritional and health status in Japanese older adults. We performed a cross-sectional analysis using data from the Japan Gerontological Evaluation study (JAGES). The study participants were 6528 functionally independent residents (3189 men and 3339 women) aged ≥65 years living in four municipalities in Aichi prefecture. We used household income as an indicator of economic status. Multiple linear regression was used to compare serum albumin levels in relation to household income, which was classified as low, middle, and high. Additionally, mediation by nutritional and health-related factors was analyzed in multivariable models. With the middle-income group as reference, participants with low incomes had a significantly lower serum albumin level, even after adjustment for sex, age, residential area, education, marital status, and household structure. The estimated mean difference was -0.17 g/L (95% confidence interval, -0.33 to -0.01 g/L). The relation between serum albumin level and low income became statistically insignificant when "body mass index", "consumption of meat or fish", "self-rated health", "presence of medical conditions", "hyperlipidemia", or "respiratory disease "was included in the model. Serum albumin levels were lower in Japanese older adults with low economic status. The decrease in albumin levels appears to be mediated by nutrition and health-related factors with low household incomes. Future studies are needed to reveal the existence of other pathways.


Assessment of phylogenetic structure in genome size--gene content correlations.
Gene content and gene-coding percentage can be predicted from genome size in newly sequenced organisms. Here, we investigate whether these predictions are influenced by phylogenetic relationships between the involved species. Combining a highly resolved phylogenetic tree with a large compilation of gene content data, our results reveal the presence of significant phylogenetic structure in the correlations between genome size and gene content in both bacteria and eukaryotes. The variation in log(gene content) explained by log(genome size) in combination with phylogeny was found to be 97% in bacteria and 55% in eukaryotes. Further, in bacteria, gene-coding percentages are only significantly correlated to genome size if phylogenetic information is taken into account in the analyses. These findings support the usage of phylogenetic correlation models for gene content predictions.


Serological Differentiation of the Citrus Tristeza Virus Isolates Causing Stem Pitting in Sweet Orange.
Citrus tristeza virus (CTV) complex comprises a number of isolates or strains producing several economically important disease syndromes in commercial Citrus spp. The stem pitting syndrome is the most important, and causes substantial losses in many citrus-producing regions of the world. In an attempt to develop a serological tool to rapidly differentiate stem pitting isolates of CTV, we evaluated many combinations of trapping and detecting antibodies in an indirect double-antibody sandwich (I-DAS) enzyme-linked immunosorbent assay (ELISA). Two combinations of trapping and detecting antibodies were found suitable for differentiating stem pitting isolates in extracts of infected sweet orange plants. One used a polyclonal serum raised against bacterially expressed CTV coat protein (CP) for trapping and a conformational monoclonal antibody 3E10 for detection, and the other used two polyclonal antisera generated against bacterially expressed CTV CP. Seventy-six CTV isolates from 20 countries, including 35 that cause stem pitting in sweet orange plants, were analyzed in I-DAS-ELISA using different combinations of polyclonal and monoclonal antibodies for trapping and as intermediate detecting antibodies. The ELISA format developed produces a strong positive signal for CTV isolates that cause stem pitting in sweet orange plants and a negative ELISA signal for CTV isolates that do not cause stem pitting. When combined with data on a universal ELISA format, i.e., reacting with a broad range of CTV isolates, these selective ELISA formats allowed reliable serological differentiation of CTV isolates that caused stem pitting in infected sweet orange plants.


Single cycle and transient force measurements in dynamic atomic force microscopy.
The monitoring of the deflection of a micro-cantilever, as the end of a sharp probe mounted at its end, i.e. the tip, interacts with a surface, forms the foundation of atomic force microscopy AFM. In a nutshell, developments in the field are driven by the requirement of obtaining ever increasing throughput and sensitivity, and enhancing the versatility of the instrument to simultaneously map the topography and quantify nanoscale processes and properties. In the most common dynamic mode of operation, the motion of the driven cantilever is monitored at a single point on its longitudinal axis. Here, we show that from this single point a waveform is obtained that contains all the details about conservative and dissipative interactions. Then a formalism that accounts for multiple arbitrary flexural modes is developed for an indirectly driven cantilever. The formalism is shown to allow recovery of the details of the interaction even in the presence of complex and relevant hysteretic forces when the cantilever oscillates in the steady state. In a different approach, we develop a formalism that monitors the wave profile of the cantilever, i.e. the waveform at five different points on its longitudinal axis. With this formalism the interaction can be reconstructed during a single oscillation cycle even in the transient state of oscillation. Finally, we discuss the potential and advantages of the proposed methods and future technical challenges. Other standard and state of the art techniques and methods are also discussed and compared with the ones presented here. This work should also provide insight into the current high throughput-high sensitivity developments dealing with multifrequency dynamic AFM where information is recovered from multiple eigenmodes.


Optimizing Contact Precautions to Curb the Spread of Antibiotic-resistant Bacteria in Hospitals: A Multicenter Cohort Study to Identify Patient Characteristics and Healthcare Personnel Interactions Associated With Transmission of Methicillin-resistant Staphylococcus aureus.
Healthcare personnel (HCP) acquire antibiotic-resistant bacteria on their gloves and gowns when caring for intensive care unit (ICU) patients. Yet, contact precautions for patients with methicillin-resistant Staphylococcus aureus (MRSA) remains controversial despite existing guidelines. We sought to understand which patients are more likely to transfer MRSA to HCP and to identify which HCP interactions are more likely to lead to glove or gown contamination. This was a prospective, multicenter cohort study of cultured HCP gloves and gowns for MRSA. Samples were obtained from patients' anterior nares, perianal area, and skin of the chest and arm to assess bacterial burden. Among 402 MRSA-colonized patients with 3982 interactions, we found that HCP gloves and gowns were contaminated with MRSA 14.3% and 5.9% of the time, respectively. Contamination of either gloves or gowns occurred in 16.2% of interactions. Contamination was highest among occupational/physical therapists (odds ratio [OR], 6.96; 95% confidence interval [CI], 3.51, 13.79), respiratory therapists (OR, 5.34; 95% CI, 3.04, 9.39), and when any HCP touched the patient (OR, 2.59; 95% CI, 1.04, 6.51). Touching the endotracheal tube (OR, 1.75; 95% CI, 1.38, 2.19), bedding (OR, 1.43; 95% CI, 1.20, 1.70), and bathing (OR, 1.32; 95% CI, 1.01, 1.75) increased the odds of contamination. We found an association between increasing bacterial burden on the patient and HCP glove or gown contamination. Gloves and gowns are frequently contaminated with MRSA in the ICU. Hospitals may consider using fewer precautions for low-risk interactions and more for high-risk interactions and personnel.


Parents' experiences of caring for a child with a cleft lip and/or palate: a review of the literature.
This review brings together for the first time the existing quantitative and qualitative research evidence about the experiences of parents caring for a child with a cleft. It summarizes salient themes on the emotional, social and service-related experiences of parents and critiques the literature to date, comparing it with wider, selected literature from the field of children's long-term conditions, including disability. The review suggests that there are similarities and differences between the literatures, in terms of research focus and approach. Similarities are found across children's conditions in the perspectives of parents on emotional, social and service-related aspects, although much of the cleft literature is focused on the early stages of children's lives. However, the quality of cleft research to date about parents' experiences has also been variable, with a narrow emphasis on cross-sectional, deficit-orientated psychological approaches focused mainly on mothers. Despite a substantial literature, little qualitative research has examined parents' perspectives in-depth, particularly about their child's treatment journey. This contrasts with the wider children's literature, which has traditionally drawn not only on psychological approaches but also on the broader perspectives of sociology, social policy, nursing and health services research, using both qualitative and quantitative methods, often in integrated ways. Such approaches have been able to highlight a greater range of experiences from both mothers and fathers, about caring for a child with a long-term condition and views about treatment. The review identifies a lack of comparable research in the cleft field to examine parents' experiences and needs at different stages of their children's lives. Above all, research is needed to investigate how both mothers and fathers might experience the long-term and complex treatment journey as children become older and to elicit their views about decision making for cleft treatments, particularly elective surgeries.


One step engineering of the small-subunit ribosomal RNA using CRISPR/Cas9.
Bacteria are indispensable for the study of fundamental molecular biology processes due to their relatively simple gene and genome architecture. The ability to engineer bacterial chromosomes is quintessential for understanding gene functions. Here we demonstrate the engineering of the small-ribosomal subunit (16S) RNA of Mycoplasma mycoides, by combining the CRISPR/Cas9 system and the yeast recombination machinery. We cloned the entire genome of M. mycoides in yeast and used constitutively expressed Cas9 together with in vitro transcribed guide-RNAs to introduce engineered 16S rRNA genes. By testing the function of the engineered 16S rRNA genes through genome transplantation, we observed surprising resilience of this gene to addition of genetic elements or helix substitutions with phylogenetically-distant bacteria. While this system could be further used to study the function of the 16S rRNA, one could envision the "simple" M. mycoides genome being used in this setting to study other genetic structures and functions to answer fundamental questions of life.


Hepatoprotective effects of Arctium lappa on carbon tetrachloride- and acetaminophen-induced liver damage.
The root of Arctium lappa Linne (A. lappa) (Compositae), a perennial herb, has been cultivated for a long time as a popular vegetable. In order to investigate the hepatoprotective effects of A. lappa, male ICR mice were injected with carbon tetrachloride (CCl4, 32 microl/kg, i.p.) or acetaminophen (600 mg/kg, i.p.). A. lappa suppressed the SGOT and SGPT elevations induced by CCl4 or acetaminophen in a dose-dependent manner and alleviated the severity of liver damage based on histopathological observations. In an attempt to elucidate the possible mechanism(s) of this hepatoprotective effect, glutathione (GSH), cytochrome P-450 (P-450) and malondialdehyde (MDA) contents were studied. A. lappa reversed the decrease in GSH and P-450 induced by CCl4 and acetaminophen. It was also found that A. lappa decreased the malondialdehyde (MDA) content in CCl4 or acetaminophen-intoxicated mice. From these results, it was suggested that A. lappa could protect the liver cells from CCl4 or acetaminophen-induced liver damages, perhaps by its antioxidative effect on hepatocytes, hence eliminating the deleterious effects of toxic metabolites from CCl4 or acetaminophen.


Discontinuation of prophylaxis against Mycobacterium avium complex disease in HIV-infected patients who have a response to antiretroviral therapy. Terry Beirn Community Programs for Clinical Research on AIDS.
Several agents are effective in preventing Mycobacterium avium complex disease in patients with advanced human immunodeficiency virus (HIV) infection. However, there is uncertainty about whether prophylaxis should be continued in patients whose CD4+ cell counts have increased substantially with antiviral therapy. We conducted a multicenter, double-blind, randomized trial of treatment with azithromycin (1200 mg weekly) as compared with placebo in HIV-infected patients whose CD4+ cell counts had increased from less than 50 to more than 100 per cubic millimeter in response to antiretroviral therapy. The primary end point was M. avium complex disease or bacterial pneumonia. A total of 520 patients entered the study; the median CD4+ cell count at entry was 230 per cubic millimeter. In 48 percent of the patients, the HIV RNA value was below the level of quantification. The median prior nadir CD4+ cell count was 23 per cubic millimeter, and 65 percent of the patients had had an acquired immunodeficiency syndrome-defining illness. During follow-up over a median period of 12 months, there were no episodes of confirmed M. avium complex disease in either group (95 percent confidence interval for the rate of disease in each group, 0 to 1.5 episodes per 100 person-years). Three patients in the azithromycin group (1.2 percent) and five in the placebo group (1.9 percent) had bacterial pneumonia (relative risk in the azithromycin group, 0.60; 95 percent confidence interval, 0.14 to 2.50; P=0.48). Neither the rate of progression of HIV disease nor the mortality rate differed significantly between the two groups. Adverse effects led to discontinuation of the study drug in 19 patients assigned to receive azithromycin (7.4 percent) and in 3 assigned to receive placebo (1.1 percent; relative risk, 6.6; P=0.002). Azithromycin prophylaxis can safely be withheld in HIV-infected patients whose CD4+ cell counts have increased to more than 100 cells per cubic millimeter in response to antiretroviral therapy.


Annual seroprevalence of Yersinia pestis in coyotes as predictors of interannual variation in reports of human plague cases in Arizona, United States.
Although several health departments collect coyote blood samples for plague surveillance, the association between reported human cases and coyote seroprevalence rates remains anecdotal. Using data from an endemic region of the United States, we sought to quantify this association. From 1974 to 1998, about 2,276 coyote blood samples from four Arizona counties were tested for serological evidence of exposure to Yersinia pestis, the causative agent of plague. Using a titer threshold presumed to be indicative of recent infection (serum titers of ≥1:256), we found a statistically significant relationship between years with >17% sero-positive coyotes and years with two or more human cases reported. Moreover, when the annual coyote seroprevalence rates were dichotomized at 17%, 84% of the years were correctly classified using four biologically relevant meteorological variables in a linear regression. This is the first time a statistically significant temporal association between human plague cases and coyote seroprevalence rates has been shown. However, issues with data resolution and surveillance effort that potentially limit the public health utility of using coyote seroprevalence rates are discussed.


Structure and scattering properties of Ni80Nb20-MgO water-window multilayer mirrors.
Ni80Nb20-MgO multilayers with d spacing that varies from 2.50 to 3.07 nm were prepared by pulsed laser deposition under conditions of ultrahigh vacuum (UHV) and argon. The morphological and atomic structure in the multilayers was determined by hard-x-ray scattering. It was found that the interface roughness in both cases, UHV and argon deposition, is <0.4 nm, whereas the lateral and longitudinal correlation lengths in the case of argon deposition, 5.0 and 1.0 nm, respectively, are an order of magnitude lower. This is due to a reduction in kinetic energy of the condensing species in argon by orders of magnitude due to multiple collisions, which reduces the lateral relaxation probability. Hence the soft-x-ray reflectance of [Ni80Nb20-MgO]10 multilayers deposited in argon was determined at 413 eV (3.00 nm), middle of the water window. The reflectance has a peak at approximately 35.2 degrees with a half-width of 3.5 degrees and 0.19% maximum value. These results agree well with the simulation results performed by use of the structural parameters obtained from hard-x-ray scattering. The atomic structure determined by high-angle x-ray diffraction shows that both Ni80Nb20 and MgO are amorphous in the as-deposited condition.


Symbiosis between methanogenic archaea and delta-proteobacteria as the origin of eukaryotes: the syntrophic hypothesis
We present a novel hypothesis for the origin of the eukaryotic cell, or eukaryogenesis, based on a metabolic symbiosis (syntrophy) between a methanogenic archaeon (methanobacterial-like) and a delta-proteobacterium (an ancestral sulfate-reducing myxobacterium). This syntrophic symbiosis was originally mediated by interspecies H2 transfer in anaerobic, possibly moderately thermophilic, environments. During eukaryogenesis, progressive cellular and genomic cointegration of both types of prokaryotic partners occurred. Initially, the establishment of permanent consortia, accompanied by extensive membrane development and close cell-cell interactions, led to a highly evolved symbiotic structure already endowed with some primitive eukaryotic features, such as a complex membrane system defining a protonuclear space (corresponding to the archaeal cytoplasm), and a protoplasmic region (derived from fusion of the surrounding bacterial cells). Simultaneously, bacterial-to-archaeal preferential gene transfer and eventual replacement took place. Bacterial genome extinction was thus accomplished by gradual transfer to the archaeal host, where genes adapted to a new genetic environment. Emerging eukaryotes would have inherited archaeal genome organization and dynamics and, consequently, most DNA-processing information systems. Conversely, primordial genes for social and developmental behavior would have been provided by the ancient myxobacterial symbiont. Metabolism would have been issued mainly from the versatile bacterial organotrophy, and progressively, methanogenesis was lost.


Poly I:C induces development of diabetes mellitus in BB rat.
Polyinosinic polycytidilic acid (poly I:C), an inducer of alpha-interferon, accelerates the development of diabetes in diabetes-prone (DP) BioBreeding (BB) rats. This study investigates the effect of administering poly I:C to a diabetes-resistant (DR) strain of BB rats. We compared the incidence of diabetes, the degree of insulitis, the number of NK cells, helper-inducer cells, cytotoxic-suppressor cells, Ia+ T cells, RT6.1+ T cells, and NK cell bioactivity in DR rats treated with saline and with a 5 micrograms/g body wt (poly-5) dose and a 10 micrograms/g body wt (poly-10) dose of poly I:C. The incidence of diabetes was also compared with that of DP rats receiving poly-5. We found that both doses of poly I:C significantly induce the development of diabetes in the DR BB rat. However, treatment of DR rats with the higher dose induces a greater rate of development of diabetes and earlier onset of diabetes than the lower poly-5 dose. The rate of diabetes development and the mean age of onset were similar in poly-10-treated DR and poly-5-treated DP rats. A significant degree of insulitis occurred in all the poly I:C-treated DR rats, even those not developing diabetes. Peripheral blood NK cell number was greater in poly I:C than in saline-treated rats, after 2 wk of treatment and when killed. The percentage of OX19+ peripheral blood mononuclear cells expressing RT6.1 allotype or Ia antigen were similar in poly I:C- and saline-treated rats.(ABSTRACT TRUNCATED AT 250 WORDS)


Oligomerization of the extracellular domain of Boss enhances its binding to the Sevenless receptor and its antagonistic effect on R7 induction.
In the developing compound eye of Drosophila, neuronal differentiation of the R7 photoreceptor cell is induced by the interaction of the receptor tyrosine kinase Sevenless with its ligand Bride of sevenless (Boss), which is expressed on the neighboring R8 cell. Boss is an unusual ligand of a receptor tyrosine kinase: it is composed of a large extracellular domain, a transmembrane domain with seven membrane-spanning segments and a cytoplasmic tail. Expression of a monomeric, secreted form of the extracellular domain of Boss is not sufficient for Sevenless activation, and instead acts as a weak antagonist. Because oligomerization appears to be a critical step in the activation of receptor tyrosine kinases, we used oligomerized forms of the Boss extracellular domain to test their ability to bind to Sevenless in vivo and restore R7 induction in vivo. Oligomerization was achieved by fusion to the leucine zipper of the yeast transcription factor GCN4 or to the tetramerization helix of Lac repressor. Binding of these multivalent proteins to Sevenless could be detected in vitro by immunoprecipitation of cross-linked ligand/receptor complexes and in vivo by receptor-dependent ligand localization. However, neither R8-specific or ubiquitous expression of multivalent Exboss ligands rescued the boss phenotype. Instead, these ligands acted as competitive inhibitors for wild-type Boss protein and thereby suppressed R7 induction. Therefore the role of the transmembrane or cytoplasmic domains of Boss in the activation of the Sev receptor cannot be replaced by oligomerization.


Conversion of a 3-desoxysteroid to 3-desoxyestrogen by human placental aromatase.
Human placental aromatase is a cytochrome P-450 enzyme system which converts androgens to estrogens by three successive oxidative reactions. The first two steps have been shown to be hydroxylations at the androgen 19-carbon, but the third step remains unknown. A leading theory for the third step involves ferric peroxide attack on the 19-oxo group to produce a 19,19-hydroxyferric peroxide intermediate and subsequent collapse to estrogen. We had previously developed a nonenzymatic peroxide model reaction which was based on the above-mentioned theory, and we demonstrated the importance of 3-ketone enolization in facilitating aromatization. This study discusses the synthesis and nonenzymatic and enzymatic study of a 3-desoxy-2,4-diene-19-oxo androgen analogue. This compound was found to be a potent nonenzymatic model substrate and competitive inhibitor of aromatase (Ki = 73 nM). Furthermore, in an unprecedented event, this compound served as a substrate for aromatase, with conversion to the corresponding 3-desoxyestrogen.


Analysis of gene expression profiles in the offspring of rats following maternal exposure to xenoestrogens.
Many environmental chemicals are known endocrine disruptors (EDs). These have the potential to alter endocrine systems via various mechanisms that include binding to hormone receptors, thereby either mimicking or blocking the hormone actions and causing abnormal gene expression. Here, to elucidate the molecular mechanism(s) underlying the detrimental effects associated with the estrogenicity of these chemicals, we determined whether gene profiles were altered in rats exposed to 4-tert-octyphenol (OP) and diethylstilbestrol (DES) in utero. Pregnant rats were treated with a high dose of OP (600 mg/kg BW per day) or DES (500 microg/kg BW per day) at gestational days (GD) 17, 18 and 19. Both dams and neonates were euthanized at lactation day (LD) 5. The transcript profiles of uterine tissue were compared in treated versus control in both maternal and neonatal sites using cDNA microarray to determine the expression levels of approximately 13,000 genes and expressed sequence tags (ESTs). The expression levels of some known estrogen-responsive genes, i.e., complement component 3, epidermal growth factor receptor or c-fos oncogene and calbindin 3, as well as some other randomly selected genes, including general transcription factor IIa, transcription factor 4 and lymphocyte specific 1, were increased by OP and/or DES treatment in the uteri of both maternal and neonate groups. However, the magnitude of these alterations in gene expression differed markedly between dams and neonates, most likely reflecting the temporal susceptibility of the reproductive tract to estrogenic chemicals. Importantly, the altered gene patterns identified by microarray analysis were confirmed by RT-PCR and real-time RT-PCR. Fifteen primers were designed to amplify specific altered genes. These genes were selected for validation because of their markedly increased expression levels and they were classified on the basis of gene ontology. Overall, a high correlation was observed between microarray and real-time PCR data. Taken together, these results indicate that placental exposure to OP or DES may cause temporal changes in gene expression in the uteri of dams and neonates. Moreover, these findings may provide useful indicators of the adverse effects of EDs and prove particularly important in elucidating the effects of xenoestrogens on estrogen-responsive tissues, such as the developing reproductive tract.


Mental health interventions for traumatized asylum seekers and refugees: What do we know about their efficacy?
The prevalence of trauma-related problems among refugees and asylum seekers is extremely high due to adverse experiences associated with forced migration. Although the literature presents a considerable number of guidelines and theoretical frameworks for working with traumatized refugees and asylum seekers, the efficacy, feasibility and applicability of these interventions have little empirical evidence. The purpose of this article is to critically review the literature to provide a rationale for developing culturally sensitive, evidence-based interventions for refugees and asylum seekers. A literature review integrating research findings on interventions designed especially for traumatized asylum seekers and refugees was conducted. Retained studies had to use some quantitative measurements of post-traumatic stress and to have pre- and post-measurements to evaluate the efficacy of the intervention. Studies included in this review cover a wide variety of interventions, including trauma-focused interventions, group therapy, multidisciplinary interventions and pharmacological treatments. The majority of studies with traumatized refugees and asylum seekers reported positive outcomes of the intervention in reducing trauma-related symptoms. There is evidence to support the suitability of cognitive-behavioral therapy (CBT) and narrative exposure therapy (NET) in certain populations of refugees. Other intervention studies are limited by methodological considerations, such as lack of randomization, absence of control group and small samples. This review has again highlighted the shortage of guiding frameworks available to investigators and clinicians who are interested in tailoring interventions to work with refugees and asylum seekers. Theoretical, ethical and methodological considerations for future research are discussed.


The protective activity of immunostimulants against Listeria monocytogenes infection in mice.
The function of peritoneal macrophages induced by intraperitoneal (i.p.) injection of attenuated Streptococcus pyogenes (OK-432), Bacillus Calmette Guérin (BCG), protein-bound polysaccharide preparation isolated from Coriolus vesicolor (PSK) or Lactobacillus casei was examined. The PMA-triggered respiratory burst (production of O2- and H2O2) of macrophages induced by OK-432, BCG or Lac. casei was greater than that of resident or thioglycollate-stimulated macrophages and was similar to that of BCG-activated macrophages. PSK failed to enhance the production of O2- or H2O2 by macrophages. Alkaline phosphodiesterase (APD) activity was reduced in macrophages induced by OK-432, BCG or Lac. casei injection and in BCG-activated macrophages. The APD activity of macrophages obtained 7 and 13 days after i.p. injection of PSK was elevated, as with thioglycollate-stimulated macrophages. Listericidal activity in vitro was enhanced in macrophages obtained in 13 and 7 days, but suppressed in macrophages obtained 2 days after OK-432, BCG or Lac. casei injection. Lac. casei administered either 2 or 13 days before infection with Listeria monocytogenes was protective but OK-432, BCG (0.1 mg) and PSK were not, even though they were able to stimulate macrophage function.


Metabolic mechanisms in heart failure.
Although neurohumoral antagonism has successfully reduced heart failure morbidity and mortality, the residual disability and death rate remains unacceptably high. Though abnormalities of myocardial metabolism are associated with heart failure, recent data suggest that heart failure may itself promote metabolic changes such as insulin resistance, in part through neurohumoral activation. A detrimental self-perpetuating cycle (heart failure --> altered metabolism --> heart failure) that promotes the progression of heart failure may thus be postulated. Accordingly, we review the cellular mechanisms and pathophysiology of altered metabolism and insulin resistance in heart failure. It is hypothesized that the ensuing detrimental myocardial energetic perturbations result from neurohumoral activation, increased adverse free fatty acid metabolism, decreased protective glucose metabolism, and in some cases insulin resistance. The result is depletion of myocardial ATP, phosphocreatine, and creatine kinase with decreased efficiency of mechanical work. On the basis of the mechanisms outlined, appropriate therapies to mitigate aberrant metabolism include intense neurohumoral antagonism, limitation of diuretics, correction of hypokalemia, exercise, and diet. We also discuss more novel mechanistic-based therapies to ameliorate metabolism and insulin resistance in heart failure. For example, metabolic modulators may optimize myocardial substrate utilization to improve cardiac function and exercise performance beyond standard care. The ultimate success of metabolic-based therapy will be manifest by its capacity further to lessen the residual mortality in heart failure.


Understanding and tuning the properties of redox-accumulating manganese helicates.
The pentanuclear Mn complex [Mn5(μ3-O)(bpp)6]n+ can access six consecutive total oxidation levels from Mn(ii)5 to Mn(iii)5. The electronic structure and redox behavior of this cluster are studied computationally and the results are compared with experimental data. The tunability of the redox potential span and of the absolute potential position is explored through systematic modifications of the 3,5-bis(pyridin-2-yl)-pyrazole (Hbpp) ligand. By substitutions with electron-donating and electron-withdrawing groups, the position of the redox events can be shifted by ca. 1 eV, however no ligand modification is predicted to alter the total redox span. By changing the pyridyl groups to benzimidazole groups, yielding the 3,5-bis(benzimidazol-2-yl)pyrazole (H3bbp) ligand, it is predicted that higher oxidation states can be stabilized, from Mn(iii)4Mn(iv) up to Mn(iv)5. In this system, the redox span for the same number of redox equivalents accumulated is slightly smaller than that in the original system. The manganese system is compared with its pentanuclear iron analogue that was recently reported to be catalytically active in oxygen evolution (Okamura et al., Nature, 2016, 530, 465). The electronic and structural requirements for utilization of the stored oxidizing equivalents in water oxidation are discussed.