Alphavirus mutator variants present host-specific defects and attenuation in mammalian and insect models.
Arboviruses cycle through both vertebrates and invertebrates, which requires them to adapt to disparate hosts while maintaining genetic integrity during genome replication. To study the genetic mechanisms and determinants of these processes, we use chikungunya virus (CHIKV), a re-emerging human pathogen transmitted by the Aedes mosquito. We previously isolated a high fidelity (or antimutator) polymerase variant, C483Y, which had decreased fitness in both mammalian and mosquito hosts, suggesting this residue may be a key molecular determinant. To further investigate effects of position 483 on RNA-dependent RNA-polymerase (RdRp) fidelity, we substituted every amino acid at this position. We isolated novel mutators with decreased replication fidelity and higher mutation frequencies, allowing us to examine the fitness of error-prone arbovirus variants. Although CHIKV mutators displayed no major replication defects in mammalian cell culture, they had reduced specific infectivity and were attenuated in vivo. Unexpectedly, mutator phenotypes were suppressed in mosquito cells and the variants exhibited significant defects in RNA synthesis. Consequently, these replication defects resulted in strong selection for reversion during infection of mosquitoes. Since residue 483 is conserved among alphaviruses, we examined the analogous mutations in Sindbis virus (SINV), which also reduced polymerase fidelity and generated replication defects in mosquito cells. However, replication defects were mosquito cell-specific and were not observed in Drosophila S2 cells, allowing us to evaluate the potential attenuation of mutators in insect models where pressure for reversion was absent. Indeed, the SINV mutator variant was attenuated in fruit flies. These findings confirm that residue 483 is a determinant regulating alphavirus polymerase fidelity and demonstrate proof of principle that arboviruses can be attenuated in mammalian and insect hosts by reducing fidelity.


The tuberculosis-associated immune reconstitution inflammatory syndrome: recent advances in clinical and pathogenesis research.
Antiretroviral therapy (ART) is an essential, life-saving intervention for HIV infection. However, ART initiation is frequently complicated by the tuberculosis-associated immune reconstitution inflammatory syndrome (TB-IRIS) in TB endemic settings. Here, we summarize the current understanding highlighting the recent evidence. The incidence of paradoxical TB-IRIS is estimated at 18% (95% CI 16-21%), higher than previously reported and may be over 50% in high-risk groups. Early ART initiation in TB patients increases TB-IRIS risk by greater than two-fold, but is critical in TB patients with CD4 counts less than 50 cells/μl because it improves survival. There remains no validated diagnostic test for TB-IRIS, and biomarkers recently proposed are not routinely used. Prednisone initiated alongside ART in selected patients with CD4 less than 100 cells/μl reduced the risk of paradoxical TB-IRIS by 30% in a recent randomized-controlled trial (RCT) and was not associated with significant adverse effects. Effective also for treating paradoxical TB-IRIS, corticosteroids remain the only therapeutic intervention for TB-IRIS supported by RCT trial data. TB-IRIS pathogenesis studies implicate high antigen burden, innate immune cell cytotoxicity, inflammasome activation and dysregulated matrix metalloproteinases in the development of the condition. Specific biomarkers would aid in identifying high-risk patients for interventions and a diagnostic test is needed. Clinicians should consider prednisone for TB-IRIS prevention in selected patients. Future research should focus on improving diagnosis and investigating novel therapeutic interventions, especially for patients in whom corticosteroid therapy is contraindicated.


Molecular mechanism of K65 acetylation-induced attenuation of Ubc9 and the NDSM interaction.
The negatively charged amino acid-dependent sumoylation motif (NDSM) carries an additional stretch of acidic residues downstream of the consensus Ψ-K-x-E/D sumoylation motif. We have previously shown that acetylation of the SUMO E2 conjugase enzyme, Ubc9, at K65 downregulates its binding to the NDSM and renders a selective decrease in sumoylation of substrates with the NDSM motif. Here, we provide detailed structural, thermodynamic, and kinetics results of the interactions between Ubc9 and its K65 acetylated variant (Ac-Ubc9K65) with three NDSMs derived from Elk1, CBP, and Calpain2 to rationalize the mechanism beneath this reduced binding. Our nuclear magnetic resonance (NMR) data rule out a direct interaction between the NDSM and the K65 residue of Ubc9. Similarly, we found that NDSM binding was entropy-driven and unlikely to be affected by the negative charge by K65 acetylation. Moreover our NMR, mutagenesis and molecular dynamics simulation studies defined the sequence of the NDSM as Ψ-K-x-E/D-x1-x2-(x3/E/D)-(x4/E/D)-xn and determined that K74 and K76 were critical Ubc9 residues interacting with the negatively charged residues of the NDSM.


HER3 genomic gain and sensitivity to gefitinib in advanced non-small-cell lung cancer patients.
In non-small-cell lung cancer (NSCLC), sensitivity to tyrosine kinase inhibitors (TKIs) is associated with activating mutations and genomic gain of the epidermal growth factor receptor (EGFR). Preclinical data suggested that HER3 overexpression increases sensitivity to TKIs. A total of 82 NSCLC patients treated with gefitinib (250 mg), and previously evaluated for EGFR and HER2 status by fluorescence in situ hybridisation (FISH) and DNA sequencing, and for Phospho-Akt status by immunohistochemistry, were investigated for HER3 genomic gain by FISH. Patients with high polysomy and gene amplification were considered as HER3 FISH positive (+). HER3 FISH+ pattern was significantly associated with female gender (P=0.02) and never smoking history (P=0.02). Patients with HER3+ tumours (26.8%) had a significantly longer time to progression (3.7 vs 2.7, P=0.04) than patients with HER3- tumours, but not a significantly better response rate or survival. Patients with EGFR+/HER3+ tumours had higher objective response rate (36.4 vs 9.9%, P=0.03) and time to progression (7.7 vs 2.7 months, P=0.03) than patients with EGFR- and/or HER3- tumours, but no significantly longer survival. No difference in response was observed according to HER3 status in patients with EGFR+ tumours. Patients with HER2+/HER3+ tumours had similar outcome as patients with HER2- and/or HER3- tumours. Significantly different clinical end points were not observed between patients with HER3+/P-Akt+ and HER3- and/or P-Akt- tumours. Genomic gain for HER3 is not a marker for response or resistance to TKI therapy in advanced NSCLC patients.


Comparative outcomes of penetrating and component endothelial cell corneal allografts in outbred sheep.
Lamellar (component cell) corneal transplantation is replacing penetrating keratoplasty for some corneal disorders in humans, but the relative risks of immunological graft rejection for the two procedures remain uncertain. A model of component endothelial cell keratoplasty (endokeratoplasty) was developed in outbred sheep. Clinical and histological graft outcomes after endokeratoplasty were then compared with contemporaneous penetrating corneal allografts. No topical or systemic immunosuppression was administered to any recipient sheep. Endothelial cell allografts (n = 10) took significantly longer to achieve perfect transparency following surgery than did penetrating corneal grafts (n = 7) (day 10 vs. day 4; p = 0.003; two-tailed Mann-Whitney U test). The median day to rejection of penetrating grafts was postoperative day 18; for endothelial cell grafts, it was day 48 (p = 0.04; two-tailed Mann-Whitney U test). The clinical courses of the two procedures were therefore quite different. Penetrating grafts gained clarity quickly but exhibited rapid graft neovascularization. Clinical rejection was preceded by inflammation in the anterior segment. Endothelial cell grafts exhibited a fluctuating, more indolent course of opacification, although all did eventually fail. Histological analysis confirmed immunological rejection in all failed grafts, but with different patterns of leukocytic infiltration in endokeratoplasties compared with penetrating keratoplasties. Inflammatory cells in endothelial cell grafts were generally fewer in number and were more often found in the posterior stroma. We conclude that, in the absence of immunosuppression, all endothelial cell allografts do undergo immunological rejection, albeit at a slower rate than penetrating grafts.


Sulfasalazine, a therapeutic agent for ulcerative colitis, inhibits the growth of CD44v9(+) cancer stem cells in ulcerative colitis-related cancer.
Sulfasalazine reduces the risk of ulcerative colitis (UC)-related cancer through its anti-inflammatory effect and induction of oxidative stress in cancer cells by inhibiting the glutamate-cystine transporter, which is closely associated with the cancer stem cell surface marker CD44v9. This study aimed to quantify the effects of sulfasalazine on CD44v9 expression and pathological factors in colorectal cancers (CRCs) arising from UC. Twenty-six patients with UC-related cancer were classified into groups according to the length of sulfasalazine treatment as follows: (1) long-term (LT) (≥ 5 years) and (2) short-term (ST) (< 5 years). Using immunohistochemistry, we compared CD44v9 and Ki-67 expression and pathological characteristics of each group's tumors. In vitro assay was performed to investigate the effect of sulfasalazine on epithelial differentiation and proliferation of CD44(+) cancer cells. Immunohistochemical analysis revealed that CD44v9 expression tended to be lower in the LT group (LT:ST=15.4%:46.2%, P=0.20), and Ki-67/CD44v9 double-stained cells were significantly lower in the LT group (LT:ST=0%:6.9%, P=0.01). Pathologically, the frequency of well-differentiated adenocarcinomas was higher in the LT group (LT:ST=84.6%:38.5%, P=0.04). In vitro assay revealed that sulfasalazine promoted the expression of epithelial differentiation markers (E-cadherin and CDX2) and inhibited the proliferation of CD44(+) cancer cells. Long-term sulfasalazine administration reduced proliferative CD44v9(+) cells and increased the degree of differentiation of adenocarcinomas. These findings indicate the importance of CD44v9(+) cells in UC-related cancer progression and suggest that sulfasalazine may serve as a novel therapeutic agent that targets CD44v9(+) cells.


Identifying mRNA targets of microRNA dysregulated in cancer: with application to clear cell Renal Cell Carcinoma.
MicroRNA regulate mRNA levels in a tissue specific way, either by inducing degradation of the transcript or by inhibiting translation or transcription. Putative mRNA targets of microRNA identified from seed sequence matches are available in many databases. However, such matches have a high false positive rate and cannot identify tissue specificity of regulation. We describe a simple method to identify direct mRNA targets of microRNA dysregulated in cancers from expression level measurements in patient matched tumor/normal samples. The word "direct" is used here in a strict sense to: a) represent mRNA which have an exact seed sequence match to the microRNA in their 3'UTR, b) the seed sequence match is strictly conserved across mouse, human, rat and dog genomes, c) the mRNA and microRNA expression levels can distinguish tumor from normal with high significance and d) the microRNA/mRNA expression levels are strongly and significantly anti-correlated in tumor and/or normal samples. We apply and validate the method using clear cell Renal Cell Carcinoma (ccRCC) and matched normal kidney samples, limiting our analysis to mRNA targets which undergo degradation of the mRNA transcript because of a perfect seed sequence match. Dysregulated microRNA and mRNA are first identified by comparing their expression levels in tumor vs normal samples. Putative dysregulated microRNA/mRNA pairs are identified from these using seed sequence matches, requiring that the seed sequence be conserved in human/dog/rat/mouse genomes. These are further pruned by requiring a strong anti-correlation signature in tumor and/or normal samples. The method revealed many new regulations in ccRCC. For instance, loss of miR-149, miR-200c and mir-141 causes gain of function of oncogenes (KCNMA1, LOX), VEGFA and SEMA6A respectively and increased levels of miR-142-3p, miR-185, mir-34a, miR-224, miR-21 cause loss of function of tumor suppressors LRRC2, PTPN13, SFRP1, ERBB4, and (SLC12A1, TCF21) respectively. We also found strong anti-correlation between VEGFA and the miR-200 family of microRNA: miR-200a*, 200b, 200c and miR-141. Several identified microRNA/mRNA pairs were validated on an independent set of matched ccRCC/normal samples. The regulation of SEMA6A by miR-141 was verified by a transfection assay. We describe a simple and reliable method to identify direct gene targets of microRNA in any cancer. The constraints we impose (strong dysregulation signature for microRNA and mRNA levels between tumor/normal samples, evolutionary conservation of seed sequence and strong anti-correlation of expression levels) remove spurious matches and identify a subset of robust, tissue specific, functional mRNA targets of dysregulated microRNA.


Probe current determination in analytical TEM/STEM and its application to the characterization of large area EDS detectors.
A simple procedure, which enables accurate measurement of transmission electron microscopy (TEM)/STEM probe currents using an energy loss spectrometer drift tube is described. The currents obtained are compared with those measured on the fluorescent screen to enable the losses due to secondary and backscattered electrons to be determined. The current values obtained from the drift tube allow the correction of fluorescent screen current densities to yield true current. They also enable CCD conversion efficiencies to be obtained, which in turn allows images to be calibrated in terms of electron fluence. Using probes of known current in conjunction with a NiO reference specimen enables the X-ray detector solid angle to be determined. The NiO specimen also allows a wide range of other EDS detector parameters to be obtained, including the presence of ice and carbon contamination. A range of performance characteristics are reported for two large area EDS detector systems. Many of the measurements reported herein have been automated via the use of freely available scripts for DigitalMicrograph.


Deleted in breast cancer 1 (DBC1) protein regulates hepatic gluconeogenesis.
Liver gluconeogenesis is essential to provide energy to glycolytic tissues during fasting periods. However, aberrant up-regulation of this metabolic pathway contributes to the progression of glucose intolerance in individuals with diabetes. Phosphoenolpyruvate carboxykinase (PEPCK) expression plays a critical role in the modulation of gluconeogenesis. Several pathways contribute to the regulation of PEPCK, including the nuclear receptor Rev-erbα and the histone deacetylase SIRT1. Deleted in breast cancer 1 (DBC1) is a nuclear protein that binds to and regulates both Rev-erbα and SIRT1 and, therefore, is a candidate to participate in the regulation of PEPCK. In this work, we provide evidence that DBC1 regulates glucose metabolism and the expression of PEPCK. We show that DBC1 levels decrease early in the fasting state. Also, DBC1 KO mice display higher gluconeogenesis in a normal and a high-fat diet. DBC1 absence leads to an increase in PEPCK mRNA and protein expression. Conversely, overexpression of DBC1 results in a decrease in PEPCK mRNA and protein levels. DBC1 regulates the levels of Rev-erbα, and manipulation of Rev-erbα activity or levels prevents the effect of DBC1 on PEPCK. In addition, Rev-erbα levels decrease in the first hours of fasting. Finally, knockdown of the deacetylase SIRT1 eliminates the effect of DBC1 knockdown on Rev-erbα levels and PEPCK expression, suggesting that the mechanism of PEPCK regulation is, at least in part, dependent on the activity of this enzyme. Our results point to DBC1 as a novel regulator of gluconeogenesis.


Hydrolase secretion is a consequence of membrane recycling.
Acanthamoeba releases lysosomal hydrolases continuously into the culture medium. This release is specific for lysosomal hydrolases, but not other cellular proteins, and is energy dependent. The secreted hydrolases can be separated into two groups on the basis of their secretion kinetics: one is secreted at approximately 15% of the cellular activity per hour and the other at approximately 5%. Intracellularly the lysosomal hydrolases are restricted almost exclusively to secondary lysosomes where the hydrolases demonstrate a differential pH-dependent binding to membrane. Hydrolase secretion is not the result of secondary lysosomes' fusing with the plasma membrane since soluble and particulate lysosomal contents are not released at the same rate. Together the data suggest that the secreted hydrolases are trapped in shuttle vesicles that cycle membrane from secondary lysosomes to the cell surface. The inner membrane and content of these vesicles undergo a marked pH shift when, following fragmentation from lysosomes, these vesicles fuse with plasma membrane. This rapid pH shift and the differential pH-dependent membrane binding of hydrolases appear to account for the heterogeneous hydrolase secretion kinetics.


Twelve-Week Rifapentine Plus Isoniazid Versus 9-Month Isoniazid for the Treatment of Latent Tuberculosis in Renal Transplant Candidates.
Renal transplant candidates (RTC) with latent tuberculosis infection (LTBI) are at significant risk for tuberculosis reactivation. Twelve-week rifapentine (RPT)/isoniazid (INH) is effective for LTBI but clinical experience in RTC is scarce. We conducted a retrospective study of RTC with LTBI treated with either 12-week RPT/INH or 9-month INH from March 1, 2012, through February 28, 2014. We evaluated both groups for differences in rates of treatment completion, monthly follow-up visit compliance, transaminase elevations, and adverse reactions leading to discontinuation of LTBI treatment. The utility of weekly reminders was also evaluated in the 12-week regimen. Direct observed therapy was not performed in our study. Of 153 patients, 43 (28%) and 110 (72%) were started on 12-week RPT/INH and 9-month INH, respectively. The treatment completion and monthly follow-up visit compliance rates were higher in the 12-week RPT/INH group (40 [93%] vs 52 [47%], P < 0.001) and (11/40 [28%] vs 13/104 [13%], P = 0.03), respectively. Transaminase elevations were not observed in the RPT/INH group, but occurred in 6 (5%) of the INH group. There were no differences in adverse reactions leading to discontinuation of LTBI treatment. Twelve-week RPT/INH appears to be an excellent choice for LTBI in RTC. It has a higher treatment completion rate and causes less transaminase elevations, and weekly reminders may be an alternative when direct observed therapy is not feasible.


Self-regulating genomic island encoding tandem regulators confers chromatic acclimation to marine Synechococcus.
The evolutionary success of marine Synechococcus, the second-most abundant phototrophic group in the marine environment, is partly attributable to this group's ability to use the entire visible spectrum of light for photosynthesis. This group possesses a remarkable diversity of light-harvesting pigments, and most of the group's members are orange and pink because of their use of phycourobilin and phycoerythrobilin chromophores, which are attached to antennae proteins called phycoerythrins. Many strains can alter phycoerythrin chromophore ratios to optimize photon capture in changing blue-green environments using type IV chromatic acclimation (CA4). Although CA4 is common in most marine Synechococcus lineages, the regulation of this process remains unexplored. Here, we show that a widely distributed genomic island encoding tandem master regulators named FciA (for type four chromatic acclimation island) and FciB plays a central role in controlling CA4. FciA and FciB have diametric effects on CA4. Interruption of fciA causes a constitutive green light phenotype, and interruption of fciB causes a constitutive blue light phenotype. These proteins regulate all of the molecular responses occurring during CA4, and the proteins' activity is apparently regulated posttranscriptionally, although their cellular ratio appears to be critical for establishing the set point for the blue-green switch in ecologically relevant light environments. Surprisingly, FciA and FciB coregulate only three genes within the Synechococcus genome, all located within the same genomic island as fciA and fciB These findings, along with the widespread distribution of strains possessing this island, suggest that horizontal transfer of a small, self-regulating DNA region has conferred CA4 capability to marine Synechococcus throughout many oceanic areas.


Protective effect of thymol on high fat diet induced diabetic nephropathy in C57BL/6J mice.
Obesity is one of several factors implicated in chronic kidney disease (CKD). Thymol, a monoterpene phenolic compound found in the oils of thyme with multiple biological properties especially antidiabetic activity. The present study was undertaken to evaluate the thymol against diabetic nephropathy by high fat diet (HFD)-induced diabetic C57BL/6J mice. After 10 weeks of continuous dietary intervention, HFD (fat- 35.2%) to mice presented characteristic features of progressive nephropathy by significant increased in kidney weight, blood, and urinary parameters, glomerulosclerosis, oxidative stress, hyperlipidemia and subsequent renal injuries. After intragastric administration of thymol (40 mg/kg BW) daily for the subsequent 5 weeks significantly decreased the blood, urinary parameters and kidney weight. Thymol inhibited the activation of transforming growth factor-β1 (TGF-β1) and vascular endothelial growth factor (VEGF). Also, significantly increased the antioxidants and suppresses the lipid peroxidation markers in erythrocytes and kidney tissue compared to the diabetic mice. Thymol downregulated the expression level of sterol regulatory element binding protein-1c (SREBP-1c) and reduced the lipid accumulation in renal. Histopathological study of kidney tissues showed that extracellular mesangial matrix expansion, glomerulosclerosis in diabetic mice were suppressed by thymol. Further, our results indicate that administration of thymol afforded remarkable protection against HFD-induced diabetic nephropathy.


Alternative view of self-diffusion and shear viscosity.
By performing an elementary transformation, the conventional velocity autocorrelation function expression for the temperature and density dependent self-diffusion constant D(T,rho) has been reformulated to emphasize how initial particle momentum biases final mean displacement. Using collective flow variables, an analogous expression has been derived for 1/eta(T,rho), the inverse of shear viscosity. The Stokes-Einstein relation for liquids declares that D and T/eta should have a fixed ratio as T and rho vary, but experiment reveals substantial violations for deeply supercooled liquids. Upon analyzing the self-diffusion and viscous flow processes in terms of configuration space inherent structures and kinetic transitions between their basins, one possible mechanism for this violation emerges. This stems from the fact that interbasin transitions become increasingly Markovian as T declines, and though self-diffusion is possible in a purely Markovian regime, shear viscosity in the present formulation intrinsically relies on successive correlated transitions.


Patchy particles made by colloidal fusion.
Patches on the surfaces of colloidal particles provide directional information that enables the self-assembly of the particles into higher-order structures. Although computational tools can make quantitative predictions and can generate design rules that link the patch motif of a particle to its internal microstructure and to the emergent properties of the self-assembled materials, the experimental realization of model systems of particles with surface patches (or 'patchy' particles) remains a challenge. Synthetic patchy colloidal particles are often poor geometric approximations of the digital building blocks used in simulations and can only rarely be manufactured in sufficiently high yields to be routinely used as experimental model systems. Here we introduce a method, which we refer to as colloidal fusion, for fabricating functional patchy particles in a tunable and scalable manner. Using coordination dynamics and wetting forces, we engineer hybrid liquid-solid clusters that evolve into particles with a range of patchy surface morphologies on addition of a plasticizer. We are able to predict and control the evolutionary pathway by considering surface-energy minimization, leading to two main branches of product: first, spherical particles with liquid surface patches, capable of forming curable bonds with neighbouring particles to assemble robust supracolloidal structures; and second, particles with a faceted liquid compartment, which can be cured and purified to yield colloidal polyhedra. These findings outline a scalable strategy for the synthesis of patchy particles, first by designing their surface patterns by computer simulation, and then by recreating them in the laboratory with high fidelity.


Impact of pressure profile and duration of recruitment maneuvers on morphofunctional and biochemical variables in experimental lung injury.
To investigate the effects of the rate of airway pressure increase and duration of recruitment maneuvers on lung function and activation of inflammation, fibrogenesis, and apoptosis in experimental acute lung injury. Prospective, randomized, controlled experimental study. University research laboratory. Thirty-five Wistar rats submitted to acute lung injury induced by cecal ligation and puncture. After 48 hrs, animals were randomly distributed into five groups (seven animals each): 1) nonrecruited (NR); 2) recruitment maneuvers (RMs) with continuous positive airway pressure (CPAP) for 15 secs (CPAP15); 3) RMs with CPAP for 30 secs (CPAP30); 4) RMs with stepwise increase in airway pressure (STEP) to targeted maximum within 15 secs (STEP15); and 5) RMs with STEP within 30 secs (STEP30). To perform STEP RMs, the ventilator was switched to a CPAP mode and positive end-expiratory pressure level was increased stepwise. At each step, airway pressure was held constant. RMs were targeted to 30 cm H2O. Animals were then ventilated for 1 hr with tidal volume of 6 mL/kg and positive end-expiratory pressure of 5 cm H2O. Blood gases, lung mechanics, histology (light and electronic microscopy), interleukin-6, caspase 3, and type 3 procollagen mRNA expressions in lung tissue. All RMs improved oxygenation and lung static elastance and reduced alveolar collapse compared to NR. STEP30 resulted in optimal performance, with: 1) improved lung static elastance vs. NR, CPAP15, and STEP15; 2) reduced alveolar-capillary membrane detachment and type 2 epithelial and endothelial cell injury scores vs. CPAP15 (p < .05); and 3) reduced gene expression of interleukin-6, type 3 procollagen, and caspase 3 in lung tissue vs. other RMs. Longer-duration RMs with slower airway pressure increase efficiently improved lung function, while minimizing the biological impact on lungs.


Comparison of semiautomated tangential VMAT with 3DCRT for breast or chest wall and regional nodes.
Radiotherapy to the breast after surgery sometimes requires adjoining nodes to be included in the treatment volume. In these cases, the traditional approach has been a complex 3-Dimensional Conformal Radiotherapy (3DCRT) beam arrangement which can result in significant dose heterogeneity at the beam junctions. A Volumetric Modulated Arc Therapy (VMAT) beam arrangement has previously been proposed for breast cases, where the chest wall/breast is treated with a limited angle (partial arc) tangential VMAT technique (Virén et al. [2015] Radiat Oncol. 10:79). In our study, this approach is extended to breast and chest wall cases with adjoining nodes by adding a separate conventional VMAT arc field specifically limited to the superior nodes. This VMAT method was implemented using a semiautomated approach on 27 patients, and the resultant plan compared to a monoisocentric 3DCRT plan. Plan statistics, Dose-Volume Histogram (DVH) analysis and Radiation Oncologist (RO) preference were assessed. When compared to the 3DCRT technique, the VMAT planning method was found to result in better target volume coverage, high doses to organs at risk (OAR) were reduced but greater OAR volumes received low doses. Having said that, the volume receiving low doses with this tangential VMAT technique was less than that of other VMAT planning methods described in the literature, and the integral dose was less than the 3DCRT method. The VMAT technique also resulted in more robust junction doses that the 3DCRT method. RO review found that the VMAT technique was preferred in 81% of cases. Specifically, the VMAT plans were preferred in all categories of patients except left chest wall cases where the intermammary nodes were also treated. The VMAT technique described here is a useful addition to the treatment options available for breast/chest wall and nodal patients.


Studies on the expression of an H-2K/human growth hormone fusion gene in giant transgenic mice.
Transgenic mice carrying the H-2K/human growth hormone (hGH) fusion gene were produced by microinjecting into the pronucleus of fertilized eggs DNA molecules containing 2 kb of the 5' flanking sequences (including promoter) of the class I H-2Kb gene joined to the coding sequences of the hGH gene. Thirteen transgenic mice were obtained which all contained detectable levels of hGH hormone in their blood. Nine grew larger than their control litter-mates. Endogenous H-2Kb and exogenous hGH mRNA levels were analysed by S1 nuclease digestion experiments. hGH transcripts were found in all the tissues examined and the pattern of expression paralleled that of endogenous H-2K gene expression, being high in liver and lymphoid organs and low in muscle and brain. Thus 2 kb of the 5' promoter/regulatory region of the H-2K gene are sufficient to ensure regulated expression of hGH in transgenic mice. This promoter may therefore be of use to target the expression of different exogenous genes in most tissues of transgenic mice and to study the biological role of the corresponding proteins in different cellular environments.


The effect of variation in definition on the diagnosis of multidirectional instability of the shoulder.
There currently is a wide variation in the definition of multidirectional instability of the shoulder in the literature. The purpose of this study was to determine if these variations influence the distribution of the diagnoses in a cohort of patients with shoulder instability. A cohort of 168 patients who underwent shoulder surgery for instability of any type was studied. Statistical analysis was performed in two steps. First, the instability of the shoulder in each patient was classified with the use of four existing systems, and the number of patients classified as having multidirectional instability was compared among the classification systems. Second, the definition of multidirectional instability was modified so that the result of laxity testing was the criterion for making the diagnosis, and the changes in the distribution of patients with a diagnosis of multidirectional instability were analyzed. Classification with the four existing systems resulted in significant differences in the number of patients diagnosed as having multidirectional instability, with two (1.2%), seven (4.2%), thirteen (7.7%), and fourteen patients (8.3%) so diagnosed (p < 0.05). Modification of the definition of multidirectional instability so that it was based on laxity testing resulted in a wide variation in the number of patients diagnosed as having multidirectional instability; these numbers ranged from fourteen (8.3%) to 139 (82.7%) (p < 0.05). This study demonstrated that variations in the criteria used for the diagnosis of multidirectional instability significantly affect the distribution of patients with that diagnosis. The use of laxity testing tends to result in an overestimation of the number of patients with this condition. This observation is important because the results of studies may vary if patients with traumatic instability are considered to have multidirectional instability on the basis of laxity testing. Investigators studying patients with multidirectional instability should carefully define the inclusion criteria that they used.


A Monte Carlo-based framework enhances the discovery and interpretation of regulatory sequence motifs.
Discovery of functionally significant short, statistically overrepresented subsequence patterns (motifs) in a set of sequences is a challenging problem in bioinformatics. Oftentimes, not all sequences in the set contain a motif. These non-motif-containing sequences complicate the algorithmic discovery of motifs. Filtering the non-motif-containing sequences from the larger set of sequences while simultaneously determining the identity of the motif is, therefore, desirable and a non-trivial problem in motif discovery research. We describe MotifCatcher, a framework that extends the sensitivity of existing motif-finding tools by employing random sampling to effectively remove non-motif-containing sequences from the motif search. We developed two implementations of our algorithm; each built around a commonly used motif-finding tool, and applied our algorithm to three diverse chromatin immunoprecipitation (ChIP) data sets. In each case, the motif finder with the MotifCatcher extension demonstrated improved sensitivity over the motif finder alone. Our approach organizes candidate functionally significant discovered motifs into a tree, which allowed us to make additional insights. In all cases, we were able to support our findings with experimental work from the literature. Our framework demonstrates that additional processing at the sequence entry level can significantly improve the performance of existing motif-finding tools. For each biological data set tested, we were able to propose novel biological hypotheses supported by experimental work from the literature. Specifically, in Escherichia coli, we suggested binding site motifs for 6 non-traditional LexA protein binding sites; in Saccharomyces cerevisiae, we hypothesize 2 disparate mechanisms for novel binding sites of the Cse4p protein; and in Halobacterium sp. NRC-1, we discoverd subtle differences in a general transcription factor (GTF) binding site motif across several data sets. We suggest that small differences in our discovered motif could confer specificity for one or more homologous GTF proteins. We offer a free implementation of the MotifCatcher software package at http://www.bme.ucdavis.edu/facciotti/resources_data/software/.


Bioinformatics analysis identification of AKT3 and RAC1 as key genes in postmenopausal osteoporosis.
Postmenopausal osteoporosis (PMO) is an aging-associated disease that manifests as degradation of bone tissue microstructure leading to decreased bone mass and increased bone fragility. Differentiation of peripheral blood mononuclear cells into osteoclasts is an important process in the development of PMO and identification of key genes that drive differentiation is essential to reveal the mechanism of PMO. The present study combined bioinformatics analysis of a Gene Expression Omnibus dataset of PMO and drug (bisphosphonate) target prediction using the STITCH database to identify hub genes in patients with PMO. Next, the expression of candidate hub genes was assessed in osteoclasts differentiated from THP-1 cells and small interfering RNA assays were performed to assess the function of selected hub genes. The present study identified 10 hub genes including WNT1, AKT3, disheveled segment polarity protein 1, cyclin D1, H2B clustered histone 17, JUN, EGFR, RAC1, actinin α1 (ACTN1) and ACTN2. Among these, AKT3 and RAC1 were highly upregulated during osteoclast differentiation, and knockdown of AKT3 and RAC1 using small interfering RNA enhanced the inhibitory effect of bisphosphonates on osteoclast differentiation and apoptosis of monocytes as assessed by tartrate-resistant acid phosphatase staining and flow cytometry examining Annexin V-FITC/PI staining, respectively. In conclusion, AKT3 and RAC1 were key for development of PMO and inhibiting AKT3 and RAC1 may improve the therapeutic efficacy of bisphosphonates.


Multiple virus infections in the honey bee and genome divergence of honey bee viruses.
Using uniplex RT-PCR we screened honey bee colonies for the presence of several bee viruses, including black queen cell virus (BQCV), deformed wing virus (DWV), Kashmir bee virus (KBV), and sacbrood virus (SBV), and described the detection of mixed virus infections in bees from these colonies. We report for the first time that individual bees can harbor four viruses simultaneously. We also developed a multiplex RT-PCR assay for the simultaneous detection of multiple bee viruses. The feasibility and specificity of the multiplex RT-PCR assay suggests that this assay is an effective tool for simultaneous examination of mixed virus infections in bee colonies and would be useful for the diagnosis and surveillance of honey bee viral diseases in the field and laboratory. Phylogenetic analysis of putative helicase and RNA-dependent RNA polymerase (RdRp) encoded by viruses reveal that DWV and SBV fall into a same clade, whereas KBV and BQCV belong to a distinct lineage with other picorna-like viruses that infect plants, insects and vertebrates. Results from field surveys of these viruses indicate that mixed infections of BQCV, DWV, KBV, and SBV in the honey bee probably arise due to broad geographic distribution of viruses.


From early intervention in psychosis to youth mental health reform: a review of the evolution and transformation of mental health services for young people.
The objective of this review is to report on recent developments in youth mental health incorporating all levels of severity of mental disorders encouraged by progress in the field of early intervention in psychotic disorders, research in deficiencies in the current system and social advocacy. The authors have briefly reviewed the relevant current state of knowledge, challenges and the service and research response across four countries (Australia, Ireland, the UK and Canada) currently active in the youth mental health field. Here we present information on response to principal challenges associated with improving youth mental services in each country. Australia has developed a model comprised of a distinct front-line youth mental health service (Headspace) to be implemented across the country and initially stimulated by success in early intervention in psychosis; in Ireland, Headstrong has been driven primarily through advocacy and philanthropy resulting in front-line services (Jigsaw) which are being implemented across different jurisdictions; in the UK, a limited regional response has addressed mostly problems with transition from child-adolescent to adult mental health services; and in Canada, a national multi-site research initiative involving transformation of youth mental health services has been launched with public and philanthropic funding, with the expectation that results of this study will inform implementation of a transformed model of service across the country including indigenous peoples. There is evidence that several countries are now engaged in transformation of youth mental health services and in evaluation of these initiatives.


Problems and prediction in survival-data analysis.
Twenty-one years after its appearance, Cox's 1972 paper on 'Regression models and life tables' continues to be one of the most frequently cited publications in the scientific and medical literature. The proportional-hazards model and partial-likelihood technique have been applied to thousands of data sets, not always appropriately, and have motivated hundreds of theoretical studies, not always relevant. Recent developments are reviewed and continuing problems discussed, especially with respect to predictive inference. The accuracy of model-based predictions, and how they compare with consultants' judgements are investigated by means of an example.


High-grade C-reactive protein elevation correlates with accelerated atherogenesis in patients with rheumatoid arthritis.
Patients with rheumatoid arthritis (RA) are at greater risk of developing cardiovascular events compared with individuals without RA. Increased risk for cardiovascular disease in these patients is a consequence of atherosclerosis. Case-control studies have shown that increased intima-media thickness (IMT) of the common carotid artery is an indicator of generalized atherosclerosis. Some investigators have suggested that the development of atherosclerosis in RA may be related to the magnitude and chronicity of the systemic inflammation. We examined the relationship between carotid IMT to C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR), which are the most commonly assessed markers of inflammatory response in patients with RA. Retrospective review of CRP and ESR values in 47 patients with longterm actively treated (at least 5 years) RA without clinically evident atherosclerosis or its complications, who had been studied for carotid IMT with high resolution B-mode ultrasound. No correlation between ESR and carotid IMT was observed. However, a correlation was found between the maximum CRP values and the carotid IMT (p = 0.009). The distribution of patients in 4 quartiles according to the average CRP values showed significant differences in the carotid IMT (p = 0.03). Those exhibiting the highest mean CRP values (quartile 4) had greater carotid IMT. There was no correlation between CRP at the time of disease diagnosis or at the time of the ultrasound study and the carotid IMT. Our study confirms that the magnitude and chronicity of the inflammatory response measured by CRP correlates directly with the presence of atherosclerosis in patients with RA.


Identification and cloning of a novel plasmid-encoded enterotoxin of enteroinvasive Escherichia coli and Shigella strains.
We have employed a molecular genetic approach to characterize the nature of enteroinvasive Escherichia coli (EIEC) enterotoxic activity, as previously observed in Ussing chambers (A. Fasano, B.A. Kay, R.G. Russell, D.R. Maneval, Jr., and M.M. Levine, Infect. Immun. 58:3717-3723, 1990). The screening of TnphoA mutants of EIEC yielded a single insertion mutant which had significantly reduced levels of enterotoxic activity in the Ussing chamber assay. DNA flanking the insertion was used as a probe to screen for EIEC cosmid clones which conferred secretogenic activity. Such screening resulted in the identification of two overlapping cosmid clones which elicited significant changes in mucosal short-circuit current (Isc). Subcloning and nucleotide sequence analysis of a DNA fragment from one of the cosmid clones led to the identification of a single open reading frame which conferred this enterotoxic activity. By DNA hybridization, this gene (designated sen for shigella enterotoxin) was found in 75% of EIEC strains and 83% of Shigella strains and was localized to the inv plasmid of Shigella flexneri 2457T. By PCR, a sen gene with 99.7% nucleotide identity was cloned and sequenced from 2457T. A deletion in the EIEC sen gene was constructed by allelic exchange, resulting in significantly lower rises in Isc than were elicited by the wild-type parent; however, significant enterotoxic activity remained in the sen deletion mutant. To purify the Sen protein, the gene was cloned into the multiple cloning site of the expression vector pKK223-3. Purification of the sen gene product yielded a protein with a molecular mass of 63 kDa which elicited rises in Isc in the Ussing chamber. We believe that the sen gene product may constitute all or part of a novel enterotoxin in EIEC and Shigella spp.


Clinical and microscopic predictors of Entamoeba histolytica intestinal infection in travelers and migrants diagnosed with Entamoeba histolytica/dispar infection.
Amebiasis is a protozoal infection caused by Entamoeba histolytica, while the morphologically indistinguishable E. dispar is considered as non-pathogenic. Polymerase chain reaction (PCR) assays are necessary to differentiate both species. The most common clinical presentations of E. histolytica disease are amebic colitis and amebic liver abscess, but asymptomatic infection is also possible. We assessed the frequency and pattern of clinical symptoms and microscopic features in travelers/migrants associated with E. histolytica intestinal infection and compared them to those found in individuals with E. dispar infection. We conducted a retrospective study at the travel clinic of the Institute of Tropical Medicine, Antwerp, Belgium on travelers/migrants found from 2006 to 2016 positive for Entamoeba histolytica/dispar through antigen detection and/or through microscopy confirmed by PCR. All files of individuals with a positive PCR for E. histolytica (= cases) and a random selection of an equal number of Entamoeba dispar carriers (= controls) were reviewed. We calculated the sensitivity, specificity and likelihood ratios (LRs) of clinical symptoms (blood in stool, mucus in stool, watery diarrhea, abdominal cramps, fever or any of these 5 symptoms) and of microscopic features (presence of trophozoites in direct and in sodium acetate-acetic acid-formalin (SAF)-fixed stool smears) to discriminate between E. histolytica and E. dispar infection. Of all stool samples positive for Entamoeba histolytica/dispar for which PCR was performed (n = 810), 30 (3.7%) were true E. histolytica infections, of which 39% were asymptomatic. Sensitivity, specificity and positive LRs were 30%, 100% and 300 (p 0.007) for presence of blood in stool; 22%, 100% and 222 (p 0.03) for mucus in stool; 44%, 90% and 4.7 (p 0.009) for cramps and 14%, 97% and 4.8 (p = 0.02) for trophozoites in direct smears. For watery diarrhea, fever and for trophozoites in SAF fixated smears results were non-significant. E. histolytica infection was demonstrated in a small proportion of travelers/migrants with evidence of Entamoeba histolytica/dispar infection. In this group, history of blood and mucus in stool and cramps had good to strong confirming power (LR+) for actual E. histolytica infection. Trophozoites were also predictive for true E. histolytica infection but in direct smears only.


Toward Control? The Prospects and Challenges of Typhoid Conjugate Vaccine Introduction.
With a newly World Health Organization (WHO)-prequalified typhoid conjugate vaccine (TCV), Gavi funding for eligible countries, and a WHO policy recommendation for TCV use, now is the time for countries to introduce TCVs as part of an integrated typhoid control program, particularly in light of the increasing burden of antimicrobial resistance. Continued vaccine development efforts will lead to secure supply of low-cost vaccines, and ongoing vaccine studies will provide critical vaccine performance data and inform optimal deployment strategies, in both routine use and in outbreak settings. TCV programs should include thoughtful communication planning and community engagement to counter vaccine hesitancy.


Evaluation of hemostasis and endothelial function in patients with paroxysmal nocturnal hemoglobinuria receiving eculizumab.
Paroxysmal nocturnal hemoglobinuria (PNH) is associated with an increased risk of thrombosis through unknown mechanisms. We studied 23 patients with PNH, before and after five and 11 weeks of treatment with eculizumab. We examined markers of thrombin generation and reactional fibrinolysis (prothrombin fragment 1+2 (F1+2), D-dimers, and plasmin antiplasmin complexes (P-AP), and endothelial dysfunction tissue plasminogen activator (t-PA), plasminogen activator inhibitor (PAI-1), soluble thrombomodulin (sTM), intercellular adhesion molecule 1 (sICAM-1), vascular cell adhesion molecule (sVCAM-1), endothelial microparticles (EMPs), and tissue factor pathway inhibitor (TFPI). At baseline, vWF, sVCAM-1, the EMP count, and F1+2 and D-dimer levels were significantly elevated in the patients, including those with no history of clinical thrombosis. Treatment with eculizumab was associated with significant decreases in plasma markers of coagulation activation (F1+2, P=0.012, and D-dimers, P=0.01), and reactional fibrinolysis (P-AP, P=0.0002). Eculizumab treatment also significantly reduced plasma markers of endothelial cell activation (t-PA, P=0.0005, sVCAM-1, P<0.0001, and vWF, P=0.0047) and total (P=0.0008) and free (P=0.0013) TFPI plasma levels. Our results suggest a new understanding of the contribution of endothelial cell activation to the pathogenesis of thrombosis in PNH. The terminal complement inhibitor, eculizumab, induced a significant and sustained decrease in the activation of both the plasma hemostatic system and the vascular endothelium, likely contributing to the protective effect of eculizumab on thrombosis in this setting.


Decisions and delays within stroke patients' route to the hospital: a qualitative study.
We examine acute stroke patients' decisions and delays en route to the hospital after onset of symptoms. This was a qualitative study carried out in the West Midlands, United Kingdom. Semistructured interviews were conducted with 30 patients (6 accompanied by partners). Patients were asked about their previous experience of having had a stroke and their initial engagement with health services. "One sheet of paper" and thematic analyses were used. Three potential types of delay were identified from onset of symptoms to accessing stroke care in the hospital: primary delays caused by lack of recognition of symptoms or not dealing with symptoms immediately, secondary delays caused by initial contact with nonemergency services, and tertiary delays in which health service providers did not interpret the patients' presenting symptoms as suggestive of stroke. The main factors determining the speed of action by patients were the presence and influence of a bystander and the perceived seriousness of symptoms. Despite campaigns to increase public awareness of stroke symptoms, the behavior of both patients and health service providers apparently led to delays in the recognition of and response to stroke symptoms, potentially reducing access to optimum and timely acute specialist assessment and treatment for acute stroke.


Single shot, three-dimensional fluorescence microscopy with a spatially rotating point spread function.
A wide-field fluorescence microscope with a double-helix point spread function (PSF) is constructed to obtain the specimen's three-dimensional distribution with a single snapshot. Spiral-phase-based computer-generated holograms (CGHs) are adopted to make the depth-of-field of the microscope adjustable. The impact of system aberrations on the double-helix PSF at high numerical aperture is analyzed to reveal the necessity of the aberration correction. A modified cepstrum-based reconstruction scheme is promoted in accordance with properties of the new double-helix PSF. The extended depth-of-field images and the corresponding depth maps for both a simulated sample and a tilted section slice of bovine pulmonary artery endothelial (BPAE) cells are recovered, respectively, verifying that the depth-of-field is properly extended and the depth of the specimen can be estimated at a precision of 23.4nm. This three-dimensional fluorescence microscope with a framerate-rank time resolution is suitable for studying the fast developing process of thin and sparsely distributed micron-scale cells in extended depth-of-field.


A risk-management approach for effective integration of biomarkers in clinical trials: perspectives of an NCI, NCRI, and EORTC working group.
Clinical cancer research today often includes testing the value of biomarkers to direct treatment and for drug development. However, the practical challenges of integration of molecular information into clinical trial protocols are increasingly appreciated. Inherent difficulties include evidence gaps in available biomarker data, a paucity of robust assay methods, and the design of appropriate studies within the constraints of feasible trial operations, and finite resources. Scalable and proportionate approaches are needed to systematically cope with these challenges. Therefore, we assembled international experts from three clinical trials organisations to identify the common challenges and common solutions. We present a practical risk-assessment framework allowing targeting of scarce resources to crucial issues coupled with a library of useful resources and a simple actionable checklist of recommendations. We hope that these practical methods will be useful for running biomarker-driven trials and ultimately help to develop biomarkers that are ready for integration in routine practice.


Evaluation in humans of ELF-EMF exposure on chromogranin A, a marker of neuroendocrine tumors and stress.
Chromogranin A (CgA), which is a major protein in adrenal chromaffin cells and adrenergic neurons, is a clinically relevant endocrine and neuroendocrine tumor marker including pheochromocytomas, neuroblastomas, and related neurogenic tumors. In this study, we looked at the effect in humans of chronic daily exposure to a 50-Hz magnetic field. We examined in 15 men (38.0 ± 0.9 years) the effects of chronic daily exposure to a 50-Hz magnetic field for 1-20 yrs both at home and at work. EMDEX II dosimeters were used to record magnetic field all day long every 30 s. for 1 week. The weekly geometric mean of the individual exposures ranged from 0.1 to 2.6 μT. Blood samples were taken hourly between 20:00 h and 08:00 h. CgA patterns of exposed subjects were compared to age-matched controls. The results of exposed subjects were compared with those for 15 unexposed men who served as controls and whose individual exposure was ten times lower ranging from 0.004 to 0.092 μT. This work shows that in the control group the serum CgA levels exhibited a nighttime peak with a progressive decline of the serum concentrations and a nadir in the morning. Both the profile and the serum concentrations of CgA, a marker of neuroendocrine tumors and stress, did not appear to be impaired in the subjects chronically exposed over a long period (up to 20 yrs) to magnetic fields though a trend toward lower levels were found at the highest exposure (>0.3 μT). This does not rule out, however, that the potential deleterious risk of ELF-EMF on frail populations such as children and the elderly may be greater at low exposure and should hence be documented, at least for their residential exposure.


Progress in the development of subunit vaccines for gastrointestinal nematodes of ruminants.
The global increase in anthelmintic resistant nematodes of ruminants, together with consumer concerns about chemicals in food, necessitates the development of alternative methods of control for these pathogens. Subunit recombinant vaccines are ideally placed to fill this gap. Indeed, they are probably the only valid option for the long-term control of ruminant parasitic nematodes given the increasing ubiquity of multidrug resistance in a range of worm species across the world. The development of a subunit multicellular parasite vaccine to the point of practical application would be a groundbreaking step in the control of these important endemic infections of livestock. This review summarizes the current status of subunit vaccine development for a number of important gastrointestinal nematodes of cattle and sheep, with a focus on the limitations and problems encountered thus far, and suggestions as to how these hurdles might be overcome.


Incidence of symptomatic tachycardia in untreated patients with paroxysmal supraventricular tachycardia.
The purpose of this article is to investigate the occurrence of symptomatic paroxysmal supraventricular tachycardia (PSVT) in untreated patients and to assess factors that influenced its occurrence. We studied 34 patients with this arrhythmia during an observation period in which they received no antiarrhythmic drug therapy for up to 90 days. Recurrence of PSVT was documented by telephone transmission of the electrocardiogram. Each patient was allowed to have exactly one episode of tachycardia before being removed from the study. We measured how long patients remained free of their tachycardia (the tachycardia-free period) and heart rate during tachycardia. Twenty-nine of the 34 patients had an attack of symptomatic tachycardia within the 90-day observation period. The proportion of patients who had not had any symptomatic PSVT by each day of follow-up was calculated using the Kaplan-Meier method as follows: 75% by day 3, 50% by day 19, 25% by day 36, and 17% by day 90. Patients with any other heart or lung disease had significantly shorter tachycardia-free periods. The mean heart rate during spontaneous tachycardia was 203.5 +/- 34.9 beats per minute (range, 142 to 288 beats per minute). Patients with longer tachycardia-free periods had significantly faster heart rates during tachycardia.


GIS-based modelling for the estimation of municipal solid waste generation and collection.
In the present paper, an innovative model for the estimation of municipal solid waste generation and collection is proposed. This model is part of an extended solid waste management system and uses a spatial Geodatabase, integrated in a GIS environment. It takes into consideration several parameters of waste generation, such as population density, commercial activities, road characteristics and their influence on the location and allocation of waste bins. Ground-based analysis was applied for the estimation of the inter-relations between the aforementioned factors and the variations in waste generation between residential and commercial areas. Therefore, the proposed model follows a unified categorization approach for residential and commercial activities and focuses on the dominant factors that determine waste generation in the area under study. The most important result of the research work presented in the current paper is an accurate estimation of the optimal number of waste bins and their allocation. A new methodology and an appropriate algorithm have been developed for this purpose in order to facilitate routing and waste collection. By using these results, municipalities aware of social, economical and environmental factors, related to waste management, can achieve optimal usage of their resources and offer the best possible services to their citizens.


Severely dependent alcohol abusers may be vulnerable to alcohol cues in television programs.
The self-reported ability of 96 alcohol abusers to resist the urge to drink heavily was assessed after they viewed a videotape of a popular prime time television program complete with advertisements. Different versions of the videotape were used to evaluate the effects of a television program with and without alcohol scenes as crossed with the effects of three different types of commercials (i.e., beer, nonalcoholic beverages, food). Before and after viewing the videotape, subjects, who were led to believe that they were participating in two separate and unrelated sets of experimental procedures, completed several drinking questionnaires. Responses to one of the questionnaires provided an unobtrusive measure of self-reported ability to resist the urge to drink heavily. Results indicated that alcohol cues in a television program affected some alcohol abusers' perceived ability to resist the urge to drink heavily. In particular, those with higher alcohol dependence scores showed a decrease in confidence after viewing a television program with alcohol cues compared to subjects who watched the same program but without the alcohol scenes. The clinical implications of these findings are discussed. Until further research is forthcoming, given the artificial nature of the study setting, the results of this study must be viewed with some caution.


Determinants of Employment Outcome for the People with Schizophrenia Using the WHODAS 2.0.
Purpose Since the vocational outcomes of people with schizophrenia should be viewed in a holistic way, the second edition of the World Health Organization Disability Assessment Schedule (WHODAS 2.0) might provide an evaluation regarding employment potential. To determine whether the WHODAS 2.0 scores can be used to predict employment status, we examined the probabilistic cut-off values of the scores and analyzed the relationship between work status and demographic characteristics. Methods We selected 31,793 people aged between 18 and 65 with schizophrenia or schizoaffective disorder from the disability evaluation database in Taiwan and separated them into two groups based on employment status (employed and unemployed). We used logistic regression to explore the association between employment and demographic characteristics. Moreover, we conducted a receiver operating characteristic (ROC) analysis to determine the cut-off point to assist in determining employment potential based on the WHODAS 2.0 score. Results Among the 31,793 participants, 3367 were employed and 18,801 were unemployed. The unemployed participants accounted for a higher percentage of disability in each domain of the WHODAS. The ROC analysis revealed that the optimal cut-off point of the WHODAS score to distinguish the people who were employed and unemployed was 25.78 (area under curve = 0.80). Conclusions The present study indicated that work status can be determined by the total score across the six domains of the WHODAS score. Furthermore, the probability of employment may be determined initially by the cut-off point of the WHODAS score in order to economize evaluation time and prepare prevocational training for those with scores above 25.78.


Freedom and Responsibility in Synthetic Genomics: The Synthetic Yeast Project.
First introduced in 2011, the Synthetic Yeast Genome (Sc2.0) PROJECT is a large international synthetic genomics project that will culminate in the first eukaryotic cell (Saccharomyces cerevisiae) with a fully synthetic genome. With collaborators from across the globe and from a range of institutions spanning from do-it-yourself biology (DIYbio) to commercial enterprises, it is important that all scientists working on this project are cognizant of the ethical and policy issues associated with this field of research and operate under a common set of principles. In this commentary, we survey the current ethics and regulatory landscape of synthetic biology and present the Sc2.0 Statement of Ethics and Governance to which all members of the project adhere. This statement focuses on four aspects of the Sc2.0 PROJECT: societal benefit, intellectual property, safety, and self-governance. We propose that such project-level agreements are an important, valuable, and flexible model of self-regulation for similar global, large-scale synthetic biology projects in order to maximize the benefits and minimize potential harms.


Understanding the modifiable health systems barriers to hypertension management in Malaysia: a multi-method health systems appraisal approach.
The growing burden of non-communicable diseases in middle-income countries demands models of care that are appropriate to local contexts and acceptable to patients in order to be effective. We describe a multi-method health system appraisal to inform the design of an intervention that will be used in a cluster randomized controlled trial to improve hypertension control in Malaysia. A health systems appraisal was undertaken in the capital, Kuala Lumpur, and poorer-resourced rural sites in Peninsular Malaysia and Sabah. Building on two systematic reviews of barriers to hypertension control, a conceptual framework was developed that guided analysis of survey data, documentary review and semi-structured interviews with key informants, health professionals and patients. The analysis followed the patients as they move through the health system, exploring the main modifiable system-level barriers to effective hypertension management, and seeking to explain obstacles to improved access and health outcomes. The study highlighted the need for the proposed intervention to take account of how Malaysian patients seek treatment in both the public and private sectors, and from western and various traditional practitioners, with many patients choosing to seek care across different services. Patients typically choose private care if they can afford to, while others attend heavily subsidised public clinics. Public hypertension clinics are often overwhelmed by numbers of patients attending, so health workers have little time to engage effectively with patients. Treatment adherence is poor, with a widespread belief, stemming from concepts of traditional medicine, that hypertension is a transient disturbance rather than a permanent asymptomatic condition. Drug supplies can be erratic in rural areas. Hypertension awareness and education material are limited, and what exist are poorly developed and ineffective. Despite having a relatively well funded health system offering good access to care, Malaysia's health system still has significant barriers to effective hypertension management. The study uncovered major patient-related barriers to the detection and control of hypertension which will have an impact on the design and implementation of any hypertension intervention. Appropriate models of care must take account of the patient modifiable health systems barriers if they are to have any realistic chance of success; these findings are relevant to many countries seeking to effectively control hypertension despite resource constraints.


Recombinant nipah virus vaccines protect pigs against challenge.
Nipah virus (NiV), of the family Paramyxoviridae, was isolated in 1999 in Malaysia from a human fatality in an outbreak of severe human encephalitis, when human infections were linked to transmission of the virus from pigs. Consequently, a swine vaccine able to abolish virus shedding is of veterinary and human health interest. Canarypox virus-based vaccine vectors carrying the gene for NiV glycoprotein (ALVAC-G) or the fusion protein (ALVAC-F) were used to intramuscularly immunize four pigs per group, either with 10(8) PFU each or in combination. Pigs were boosted 14 days postvaccination and challenged with 2.5 x 10(5) PFU of NiV two weeks later. The combined ALVAC-F/G vaccine induced the highest levels of neutralization antibodies (2,560); despite the low neutralizing antibody levels in the F vaccinees (160), all vaccinated animals appeared to be protected against challenge. Virus was not isolated from the tissues of any of the vaccinated pigs postchallenge, and a real-time reverse transcription (RT)-PCR assay detected only small amounts of viral RNA in several samples. In challenge control pigs, virus was isolated from a number of tissues (10(4.4) PFU/g) or detected by real-time RT-PCR. Vaccination of the ALVAC-F/G vaccinees appeared to stimulate both type 1 and type 2 cytokine responses. Histopathological findings indicated that there was no enhancement of lesions in the vaccinees. No virus shedding was detected in vaccinated animals, in contrast to challenge control pigs, from which virus was isolated from the throat and nose (10(2.9) PFU/ml). Based on the data presented, the combined ALVAC-F/G vaccine appears to be a very promising vaccine candidate for swine.


Variations in potassium channel genes are associated with distinct trajectories of persistent breast pain after breast cancer surgery.
Persistent pain after breast cancer surgery is a common clinical problem. Given the role of potassium channels in modulating neuronal excitability, coupled with recently published genetic associations with preoperative breast pain, we hypothesized that variations in potassium channel genes will be associated with persistent postsurgical breast pain. In this study, associations between 10 potassium channel genes and persistent breast pain were evaluated. Using growth mixture modeling (GMM), 4 distinct latent classes of patients, who were assessed before and monthly for 6 months after breast cancer surgery, were identified previously (ie, No Pain, Mild Pain, Moderate Pain, Severe Pain). Genotyping was done using a custom array. Using logistic regression analyses, significant differences in a number of genotype or haplotype frequencies were found between: Mild Pain vs No Pain and Severe Pain vs No Pain classes. Seven single-nucleotide polymorphisms (SNPs) across 5 genes (ie, potassium voltage-gated channel, subfamily A, member 1 [KCNA1], potassium voltage-gated channel, subfamily D, member 2 [KCND2], potassium inwardly rectifying channel, subfamily J, members 3 and 6 (KCNJ3 and KCNJ6), potassium channel, subfamily K, member 9 [KCNK9]) were associated with membership in the Mild Pain class. In addition, 3 SNPs and 1 haplotype across 4 genes (ie, KCND2, KCNJ3, KCNJ6, KCNK9) were associated with membership in the Severe Pain class. These findings suggest that variations in potassium channel genes are associated with both mild and severe persistent breast pain after breast cancer surgery. Although findings from this study warrant replication, they provide intriguing preliminary information on potential therapeutic targets.


Binding site for coenzyme A revealed in the structure of pyruvate:ferredoxin oxidoreductase from Moorella thermoacetica.
Pyruvate:ferredoxin oxidoreductase (PFOR) is a microbial enzyme that uses thiamine pyrophosphate (TPP), three [4Fe-4S] clusters, and coenzyme A (CoA) in the reversible oxidation of pyruvate to generate acetyl-CoA and carbon dioxide. The two electrons that are generated as a result of pyruvate decarboxylation are used in the reduction of low potential ferredoxins, which provide reducing equivalents for central metabolism, including the Wood-Ljungdahl pathway. PFOR is a member of the 2-oxoacid:ferredoxin oxidoreductase (OFOR) superfamily, which plays major roles in both microbial redox reactions and carbon dioxide fixation. Here, we present a set of crystallographic snapshots of the best-studied member of this superfamily, the PFOR from Moorella thermoacetica (MtPFOR). These snapshots include the native structure, those of lactyl-TPP and acetyl-TPP reaction intermediates, and the first of an OFOR with CoA bound. These structural data reveal the binding site of CoA as domain III, the function of which in OFORs was previously unknown, and establish sequence motifs for CoA binding in the OFOR superfamily. MtPFOR structures further show that domain III undergoes a conformational change upon CoA binding that seals off the active site and positions the thiolate of CoA directly adjacent to the TPP cofactor. These structural findings provide a molecular basis for the experimental observation that CoA binding accelerates catalysis by 105-fold.


Boiler briquette coal versus raw coal: Part II--Energy, greenhouse gas, and air quality implications.
The objective of this paper is to conduct an integrated analysis of the energy, greenhouse gas, and air quality impacts of a new type of boiler briquette coal (BB-coal) in contrast to those of the raw coal from which the BB-coal was formulated (R-coal). The analysis is based on the source emissions data and other relevant data collected in the present study and employs approaches including the construction of carbon, energy, and sulfur balances. The results show that replacing R-coal with BB-coal as the fuel for boilers such as the one tested would have multiple benefits, including a 37% increase in boiler thermal efficiency, a 25% reduction in fuel demand, a 26% reduction in CO2 emission, a 17% reduction in CO emission, a 63% reduction in SO2 emission, a 97% reduction in fly ash and fly ash carbon emission, a 22% reduction in PM2.5 mass emission, and a 30% reduction in total emission of five toxic hazardous air pollutant (HAP) metals contained in PM10. These benefits can be achieved with no changes in boiler hardware and with a relatively small amount of tradeoffs: a 30% increase in PM10 mass emission and a 9-16% increase in fuel cost.


The DMM complex prevents spreading of DNA methylation from transposons to nearby genes in Neurospora crassa.
Transposable elements are common in genomes and must be controlled. Many organisms use DNA methylation to silence such selfish DNA, but the mechanisms that restrict the methylation to appropriate regions are largely unknown. We identified a JmjC domain protein in Neurospora, DNA METHYLATION MODULATOR-1 (DMM-1), that prevents aberrant spreading of DNA and histone H3K9 methylation from inactivated transposons into nearby genes. Mutation of a conserved residue within the JmjC Fe(II)-binding site abolished dmm-1 function, as did mutations in conserved cysteine-rich domains. Mutants defective only in dmm-1 mutants grow poorly, but growth is restored by reduction or elimination of DNA methylation using the drug 5-azacytosine or by mutation of the DNA methyltransferase gene dim-2. DMM-1 relies on an associated protein, DMM-2, which bears a DNA-binding motif, for localization and proper function. HP1 is required to recruit the DMM complex to the edges of methylated regions.


[Protective effects of CB1 receptor agonist WIN 55.212-2 in seizure activity in the model of temporal lobe epilepsy].
The influence of a low dose (1 microM, 2 microl) of nonselective agonist of cannabinoid CB1 receptor WIN 55.212-2 on seizure activity in different brain structures was investigated in waking guinea pigs. Changes in spontaneous local field potentials and seizure afterdischarges evoked by the electrical stimulation of the perforant path were recorded simultaneously in the hippocampus, entorhinal cortex, medial septal region, and amygdala after a preliminary intraventricular injection of WIN 55.212-2. It was found that WIN 55.212-2 blocked the stimulation-elicited seizures in 80% of tests. A repeated injection of the agonist within 30 days caused an increase in the amplitude of local field potentials and the power of the theta rhythm in all the structures under study. The infusion of kainic acid provoked the onset of status epilepticus in control animals, whereas guinea pigs injected with the agonist (daily, during 25-30 days) did not develop the status. Possible mechanisms of the protective influence of WIN 55.212-2 are discussed.


Genetic variants in Nogo receptor signaling pathways may be associated with early life adversity in schizophrenia susceptibility.
Schizophrenia is a severe neuropsychiatric disorder thought to result from abnormal brain development. Nogo, an oligodendrocyte bound molecule, signals by binding to the Nogo receptor (NgR) located on axonal membranes. The NgR co-receptors include p75 neurotrophin receptor or TNF receptor orphan Y (TROY). Nogo signaling is responsible for central nervous system myelin regulation and neurite outgrowth during neurodevelopment, and plasticity in the mature brain. We examined single nucleotide polymorphisms (SNPs) in NgR, p75, and TROY receptor genes and downstream signaling partner With No Lysine (K) (WNK1) and Myelin transcription factor 1-like (Myt1l) genes in an Australian case-control schizophrenia cohort (n = 268/group). High-throughput SNP genotyping was performed using the MassARRAY® genotyping assay. Analysis revealed a significant association between the Myt1l SNP rs2304008 and female schizophrenia subjects. The WNK1 SNP rs1468326 and the Myt1l SNP rs3748988 showed significant associations with schizophrenia in subjects with a maternal mental history and in subjects who experienced childhood trauma respectively. Following Bonferroni correction, all significance was lost. Despite the lack of positive findings in our population after correction for multiple testing, previous gene expression and association studies in schizophrenia suggest the implication of NgR signaling pathway genes in the etiology of schizophrenia remains topical and timely. Further investigations will be necessary to fully assess the role of these genes in the pathophysiology of schizophrenia. However these genes may prove useful in further understanding the mechanism by which negative experiences early in life can affect myelin-related processes in the context of schizophrenia.


Is it safe to use visible blue light-emitting diode phototherapy for neonatal jaundice in infants who are also treated with amiodarone?
Amiodarone is a useful treatment for neonatal cardiac arrhythmias. A minority of neonates with arrhythmias treated with amiodarone may also develop neonatal jaundice. Phototherapy is recommended to treat neonatal jaundice. However, amiodarone is known to cause skin reactions when patients are exposed to light. Therefore phototherapy may not be administered due to concerns about this interaction. Exchange transfusions are used as an alternative to phototherapy to avoid the interaction. However, exchange transfusions are associated with serious complications such as the transmission of bloodborne disease, seizures and haemodynamic instability. We report a literature review and outcome of a premature infant treated with amiodarone and visible blue light-emitting diode (LED) phototherapy. Phototherapy was used for 54 hours and no skin reactions were observed. This is because visible blue LED phototherapy lamps emit predominantly visible light. Therefore exposure to ultraviolet light that is known to cause amiodarone phototoxicity is minimised.


Association mapping in Salix viminalis L. (Salicaceae) - identification of candidate genes associated with growth and phenology.
Willow species (Salix) are important as short-rotation biomass crops for bioenergy, which creates a demand for faster genetic improvement and breeding through deployment of molecular marker-assisted selection (MAS). To find markers associated with important adaptive traits, such as growth and phenology, for use in MAS, we genetically dissected the trait variation of a Salix viminalis (L.) population of 323 accessions. The accessions were sampled throughout northern Europe and were established at two field sites in Pustnäs, Sweden, and at Woburn, UK, offering the opportunity to assess the impact of genotype-by-environment interactions (G × E) on trait-marker associations. Field measurements were recorded for growth and phenology traits. The accessions were genotyped using 1536 SNP markers developed from phenology candidate genes and from genes previously observed to be differentially expressed in contrasting environments. Association mapping between 1233 of these SNPs and the measured traits was performed taking into account population structure and threshold selection bias. At a false discovery rate (FDR) of 0.2, 29 SNPs were associated with bud burst, leaf senescence, number of shoots or shoot diameter. The percentage of accession variation (Radj2) explained by these associations ranged from 0.3% to 4.4%, suggesting that the studied traits are controlled by many loci of limited individual impact. Despite this, a SNP in the EARLY FLOWERING 3 gene was repeatedly associated (FDR < 0.2) with bud burst. The rare homozygous genotype exhibited 0.4-1.0 lower bud burst scores than the other genotype classes on a five-grade scale. Consequently, this marker could be promising for use in MAS and the gene deserves further study. Otherwise, associations were less consistent across sites, likely due to their small Radj2 estimates and to considerable G × E interactions indicated by multivariate association analyses and modest trait accession correlations across sites (0.32-0.61).


Regional Differences in Diffuse Idiopathic Skeletal Hyperostosis: A Retrospective Cohort Study from Sweden and Japan.
We retrospectively reviewed computed tomography (CT) records of patients in Japan and Sweden, which are both aging populations. To research the influence of ethnicity and region on diffuse idiopathic skeletal hyperostosis (DISH) prevalence. DISH can complicate non-surgical treatment of spinal fractures and often requires surgical intervention. We previously reported a prevalence of DISH in Japan that was higher than that reported in other studies. We retrospectively reviewed CT records of patients in Japan and Sweden, which have both aging populations. Patients undergoing whole body CT during trauma examinations at an acute outpatient clinic in Uppsala University Hospital in a 1-year period were eligible for inclusion. Excluded were those less than 40 and more than or equal to 90 years old, and those with previous spinal surgery. The prevalence of DISH by sex and age was determined according to radiographic criteria by Resnick. Results from Sweden were compared with the Japan data, which we previously reported. Age of the eligible subjects (265 men and 153 women) ranged from 40 to 89 years, with a mean age of 63.4 years. Among men, 86 (32.5%) were diagnosed with DISH, and the results by age (40s, 50s, 60s, 70s, and 80s) were: 6 (10.7%), 13 (22%), 35 (46.1%), 17 (34%), and 15 (62.5%) patients, respectively. Among women, 16 (10.5%) had DISH, and the results by age were as follows: 1 (2.6%), 1 (3.3%), 2 (6.7%), 6 (22.2%), and 6 (22.2%) patients, respectively. These results did not differ from those previously published for Japan (Fisher exact test, men: P = 1, 0.27, 0.12, 0.06, and 1, respectively; women: P = 0.49, 0.62, 0.5, 0.8, and 0.3, respectively). The presented cohort study revealed that ethnicity and region may not be notable factors of DISH prevalence, since patients from both Japan and Sweden had similar DISH prevalence. 3.


Accelerated collagen-induced arthritis in IFN-gamma receptor-deficient mice.
Collagen-induced arthritis (CIA) is a model for rheumatoid arthritis. Here, we describe experiments showing that IFN-gamma receptor knockout (IFN-gammaR alpha KO) mice of the DBA/1 strain develop CIA more readily than their wild-type counterparts. Symptoms of disease started 10 days earlier and the cumulative incidence of arthritis was significantly higher in the mutant mice than in wild-type mice. Similarly, accelerated onset of the disease was also found in wild-type DBA/1 mice treated with neutralizing mAbs against IFN-gamma. Histologic examination of the joints revealed a massive infiltration of the synovium with mononuclear cells and neutrophils, hyperplasia, and severe pannus formation in IFN-gammaR alpha KO mice when such inflammatory lesions were not yet detectable in wild-type mice. Serum levels of anti-collagen type II Abs, including total IgG and IgM, as well as IgG1, IgG2a, and IgG2b isotypes were found to be lower in the mutant mice. IL-2 and IL-4 remained undetectable in sera of both groups of mice, but did appear in the circulation after anti-CD3 Ab challenge. Significantly higher IL-2 and lower IL-4 serum levels were found in anti-CD3-challenged IFN-gammaR alpha KO mice than in wild-type counterparts, both at an early and at a later stage of the disease. These observations indicate that endogenous IFN-gamma counteracts development of collagen-induced arthritis and suggest that IFN-gamma does so by up-regulating IL-4 production and/or down-regulating IL-2 production. The data are in line with the concept of a pathogenic role of Th1-type cellular immunity in CIA in spite of a decreased Ab response to collagen type II.


Synergy between Nutritional Immunity and Independent Host Defenses Contributes to the Importance of the MntABC Manganese Transporter during Staphylococcus aureus Infection.
During infection, the host utilizes a diverse array of processes to combat invaders, including the restriction of availability of essential nutrients such as manganese. Similarly to many other pathogens, Staphylococcus aureus possesses two manganese importers, MntH and MntABC. Several infection models have revealed a critical role for MntABC during staphylococcal infection. However, culture-based studies have suggested parity between the two transporters when cells are resisting manganese starvation imposed by the manganese binding immune effector calprotectin. In this investigation, initial elemental analysis revealed that MntABC is the primary transporter responsible for obtaining manganese in culture in the presence of calprotectin. MntABC was also necessary to maintain wild-type levels of manganese-dependent superoxide dismutase activity in the presence of calprotectin. Building on this framework, we investigated if MntABC enabled S. aureus to resist the synergistic actions of nutritional immunity and other host defenses. This analysis revealed that MntABC critically contributes to staphylococcal growth when S. aureus is subjected to manganese limitations and exposed to oxidative stress. This transporter was also important for growth in manganese-limited environments when S. aureus was forced to consume glucose as an energy source, which occurs when it encounters nitric oxide. MntABC also expanded the pH range conducive for S. aureus growth under conditions of manganese scarcity. Collectively, the data presented in this work provide a robust molecular basis for the crucial role of MntABC in staphylococcal virulence. Further, this work highlights the importance of synergy between host defenses and the necessity of evaluating the contribution of virulence factors to pathogenesis in the presence of multiple stressors.


Omega-3 fatty acids as a treatment for non-alcoholic fatty liver disease in children: A systematic review and meta-analysis of randomized controlled trials.
The most typical chronic liver disease in children and adolescents is non-alcoholic fatty liver disease (NAFLD). The dietary addition of ω-3 polyunsaturated fatty acids (PUFAs) provides a promising therapy for children with NAFLD due to its convenience and safety; however, several studies suggested contradictory results for PUFA supplementation in children. Hence, we performed a systematic review and meta-analysis to evaluate the effectiveness of PUFA supplementation in children with NAFLD. Published randomized controlled trials (RCTs) that evaluated the effectiveness of the dietary addition of PUFA in children with NAFLD were considered. The primary result was the alteration in hepatic steatosis grade on ultrasound after treatment. The secondary outcomes included alanine aminotransferase (ALT), aspartate aminotransferase (AST), C-reactive protein (CRP) and components of metabolic syndrome. Predefined sensitivity analysis was also performed to explore possible explanations for heterogeneity in the evaluations. In total, 4 studies with 263 subjects were identified. PUFA supplementation was associated with significantly improved hepatic steatosis grade on ultrasound (risk difference: 25%, 95% CI: 12-38%), without heterogeneity (P = 0.27, I2 = 24%). Sensitivity analysis confirmed the robustness of our findings. PUFA supplementation could decrease AST levels after 6 months, but could only reduce ALT levels after 12 months. PUFA did not have a significant effect on most components of metabolic syndrome and the CRP level. ω-3 PUFA supplementation can improve liver steatosis and liver functions, and it is a potential food supplementation to treat NAFLD in children.


Hysteroscopy for treating subfertility associated with suspected major uterine cavity abnormalities.
Observational studies suggest higher pregnancy rates after the hysteroscopic removal of endometrial polyps, submucous fibroids, uterine septum or intrauterine adhesions, which are detectable in 10% to 15% of women seeking treatment for subfertility. To assess the effects of the hysteroscopic removal of endometrial polyps, submucous fibroids, uterine septum or intrauterine adhesions suspected on ultrasound, hysterosalpingography, diagnostic hysteroscopy or any combination of these methods in women with otherwise unexplained subfertility or prior to intrauterine insemination (IUI), in vitro fertilisation (IVF) or intracytoplasmic sperm injection (ICSI). We searched the Cochrane Menstrual Disorders and Subfertility Specialised Register (8 September 2014), the Cochrane Central Register of Controlled Trials (The Cochrane Library 2014, Issue 9), MEDLINE (1950 to 12 October 2014), EMBASE (inception to 12 October 2014), CINAHL (inception to 11 October 2014) and other electronic sources of trials including trial registers, sources of unpublished literature and reference lists. We handsearched the American Society for Reproductive Medicine (ASRM) conference abstracts and proceedings (from January 2013 to October 2014) and we contacted experts in the field. Randomised comparisons between operative hysteroscopy versus control in women with otherwise unexplained subfertility or undergoing IUI, IVF or ICSI and suspected major uterine cavity abnormalities diagnosed by ultrasonography, saline infusion/gel instillation sonography, hysterosalpingography, diagnostic hysteroscopy or any combination of these methods. Primary outcomes were live birth and hysteroscopy complications. Secondary outcomes were pregnancy and miscarriage. Two review authors independently assessed studies for inclusion and risk of bias, and extracted data. We contacted study authors for additional information. We retrieved 12 randomised trials possibly addressing the research questions. Only two studies (309 women) met the inclusion criteria. Neither reported the primary outcomes of live birth or procedure related complications. In women with otherwise unexplained subfertility and submucous fibroids there was no conclusive evidence of a difference between the intervention group treated with hysteroscopic myomectomy and the control group having regular fertility-oriented intercourse during 12 months for the outcome of clinical pregnancy. A large clinical benefit with hysteroscopic myomectomy cannot be excluded: if 21% of women with fibroids achieve a clinical pregnancy having timed intercourse only, the evidence suggests that 39% of women (95% CI 21% to 58%) will achieve a successful outcome following the hysteroscopic removal of the fibroids (odds ratio (OR) 2.44, 95% confidence interval (CI) 0.97 to 6.17, P = 0.06, 94 women, very low quality evidence). There is no evidence of a difference between the comparison groups for the outcome of miscarriage (OR 0.58, 95% CI 0.12 to 2.85, P = 0.50, 30 clinical pregnancies in 94 women, very low quality evidence). The hysteroscopic removal of polyps prior to IUI can increase the chance of a clinical pregnancy compared to simple diagnostic hysteroscopy and polyp biopsy: if 28% of women achieve a clinical pregnancy with a simple diagnostic hysteroscopy, the evidence suggests that 63% of women (95% CI 50% to 76%) will achieve a clinical pregnancy after the hysteroscopic removal of the endometrial polyps (OR 4.41, 95% CI 2.45 to 7.96, P < 0.00001, 204 women, moderate quality evidence). A large benefit with the hysteroscopic removal of submucous fibroids for improving the chance of clinical pregnancy in women with otherwise unexplained subfertility cannot be excluded. The hysteroscopic removal of endometrial polyps suspected on ultrasound in women prior to IUI may increase the clinical pregnancy rate. More randomised studies are needed to substantiate the effectiveness of the hysteroscopic removal of suspected endometrial polyps, submucous fibroids, uterine septum or intrauterine adhesions in women with unexplained subfertility or prior to IUI, IVF or ICSI.


Estrogen-related receptor alpha induces the expression of vascular endothelial growth factor in breast cancer cells.
Estrogen-related receptor alpha (ERRalpha) is an orphan member of the nuclear receptor family of transcription factors. In addition to its function as a metabolic regulator, ERRalpha has been implicated in the growth and progression of several malignancies. In the setting of breast cancer, not only is ERRalpha a putative negative prognostic factor, but we have recently found that knock-down of its expression retards tumor growth in a xenograft model of this disease. The specific aspects of ERRalpha function that are responsible for its actions in breast cancer, however, remain unclear. Using the coactivator PGC-1alpha as a protein ligand to regulate ERRalpha activity, we analyzed the effects of this receptor on gene expression in the ERalpha-positive MCF-7 cell line. This analysis led to the identification of a large number of potential ERRalpha target genes, many of which were subsequently validated in other breast cancer cell lines. Importantly, we demonstrate in this study that activation of ERRalpha in several different breast cancer cell lines leads to a significant increase in VEGF mRNA expression, an activity that translates into an increase in VEGF protein secretion. The induction of VEGF results from the interaction of ERRalpha with specific ERR-responsive elements within the VEGF promoter. These findings suggest that ERRalpha-dependent induction of VEGF may contribute to the overall negative phenotype observed in tumors in which ERRalpha is expressed and provide validation for its use as a therapeutic target in cancer.


Pharmacological treatment of acute spinal cord injury: how do we build on past success?
Most acute spinal cord injuries (SCI) do not involve complete transection of the spinal cord; typically, a rim of white matter survives. The potential for neurological recovery depends on optimal preservation of the ascending and descending white matter axons and their normal myelination. Pharmacologic strategies focus on the control of secondary injury processes, primarily lipid peroxidation (LP), and the salvage of as much white matter as possible. The first effective neuroprotective agent was methylprednisolone (MP), a glucocorticosteroid that in high doses improves neurological recovery in animals and humans following acute SCI. Tirilazad is a more targeted non-glucocorticoid LP inhibitor that has been shown to be neuroprotective and has fewer side effects than MP. Future SCI therapy is likely to encompass various neuroprotective agents, including inhibitors of LP, inhibitors of the nitric oxide-derived reactive oxygen species peroxynitrite, inhibitors of calpain (which is responsible for degrading the spinal cord cytoskeleton), and inhibitors of post-traumatic apoptosis of neurons and myelin-forming oligodendroglia. In addition, neuroprotective strategies will eventually be followed by neurorestorative agents that stimulate the plasticity of surviving neural pathways, and will be used in conjunction with other neurorestorative therapies like cell transplantation and gene therapy techniques.


Predicting Aspergillus fumigatus exposure from composting facilities using a dispersion model: A conditional calibration and validation.
Bioaerosols are released in elevated quantities from composting facilities and are associated with negative health effects, although dose-response relationships are unclear. Exposure levels are difficult to quantify as established sampling methods are costly, time-consuming and current data provide limited temporal and spatial information. Confidence in dispersion model outputs in this context would be advantageous to provide a more detailed exposure assessment. We present the calibration and validation of a recognised atmospheric dispersion model (ADMS) for bioaerosol exposure assessments. The model was calibrated by a trial and error optimisation of observed Aspergillus fumigatus concentrations at different locations around a composting site. Validation was performed using a second dataset of measured concentrations for a different site. The best fit between modelled and measured data was achieved when emissions were represented as a single area source, with a temperature of 29°C. Predicted bioaerosol concentrations were within an order of magnitude of measured values (1000-10,000CFU/m3) at the validation site, once minor adjustments were made to reflect local differences between the sites (r2>0.7 at 150, 300, 500 and 600m downwind of source). Results suggest that calibrated dispersion modelling can be applied to make reasonable predictions of bioaerosol exposures at multiple sites and may be used to inform site regulation and operational management.


Taxonomic similarity does not predict necessary sample size for ex situ conservation: a comparison among five genera.
Effectively conserving biodiversity with limited resources requires scientifically informed and efficient strategies. Guidance is particularly needed on how many living plants are necessary to conserve a threshold level of genetic diversity in ex situ collections. We investigated this question for 11 taxa across five genera. In this first study analysing and optimizing ex situ genetic diversity across multiple genera, we found that the percentage of extant genetic diversity currently conserved varies among taxa from 40% to 95%. Most taxa are well below genetic conservation targets. Resampling datasets showed that ideal collection sizes vary widely even within a genus: one taxon typically required at least 50% more individuals than another (though Quercus was an exception). Still, across taxa, the minimum collection size to achieve genetic conservation goals is within one order of magnitude. Current collections are also suboptimal: they could remain the same size yet capture twice the genetic diversity with an improved sampling design. We term this deficiency the 'genetic conservation gap'. Lastly, we show that minimum collection sizes are influenced by collection priorities regarding the genetic diversity target. In summary, current collections are insufficient (not reaching targets) and suboptimal (not efficiently designed), and we show how improvements can be made.