The role of integrin-linked kinase in the molecular architecture of focal adhesions.
Integrin-mediated focal adhesions (FAs) are large, multi-protein complexes that link the actin cytoskeleton to the extracellular matrix and take part in adhesion-mediated signaling. These adhesions are highly complex and diverse at the molecular level; thus, assigning particular structural or signaling functions to specific components is highly challenging. Here, we combined functional, structural and biophysical approaches to assess the role of a major FA component, namely, integrin-linked kinase (ILK), in adhesion formation. We show here that ILK plays a key role in the formation of focal complexes, early forms of integrin adhesions, and confirm its involvement in the assembly of fibronectin-bound fibrillar adhesions. Examination of ILK-null fibroblasts by cryo-electron tomography pointed to major structural changes in their FAs, manifested as disarray of the associated actin filaments and an increase in the packing density of FA-related particles. Interestingly, adhesion of the mutant cells to the substrate required a higher ligand density than in control cells. These data indicate that ILK has a key role in integrin adhesion assembly and sub-structure, and in the regulation of the FA-associated cytoskeleton.


Computational approaches for analyzing information flow in biological networks.
The advancements in "omics" (proteomics, genomics, lipidomics, and metabolomics) technologies have yielded large inventories of genes, transcripts, proteins, and metabolites. The challenge is to find out how these entities work together to regulate the processes by which cells respond to external and internal signals. Mathematical and computational modeling of signaling networks has a key role in this task, and network analysis provides insights into biological systems and has applications for medicine. Here, we review experimental and theoretical progress and future challenges toward this goal. We focus on how networks are reconstructed from data, how these networks are structured to control the flow of biological information, and how the design features of the networks specify biological decisions.


Nuclear cyclophilins affect spliceosome assembly and function in vitro.
Cyclophilins are ubiquitously expressed proteins that bind to prolines and can catalyse cis/trans isomerization of proline residues. There are 17 annotated members of the cyclophilin family in humans, ubiquitously expressed and localized variously to the cytoplasm, nucleus or mitochondria. Surprisingly, all eight of the nuclear localized cyclophilins are found associated with spliceosomal complexes. However, their particular functions within this context are unknown. We have therefore adapted three established assays for in vitro pre-mRNA splicing to probe the functional roles of nuclear cyclophilins in the context of the human spliceosome. We find that four of the eight spliceosom-associated cyclophilins exert strong effects on splicing in vitro. These effects are dose-dependent and, remarkably, uniquely characteristic of each cyclophilin. Using both qualitative and quantitative means, we show that at least half of the nuclear cyclophilins can act as regulatory factors of spliceosome function in vitro. The present work provides the first quantifiable evidence that nuclear cyclophilins are splicing factors and provides a novel approach for future work into small molecule-based modulation of pre-mRNA splicing.


Chronic pneumonia in calves after experimental infection with Mycoplasma bovis strain 1067: characterization of lung pathology, persistence of variable surface protein antigens and local immune response.
Mycoplasma bovis is associated with pneumonia in calves characterized by the development of chronic caseonecrotic lesions with the agent persisting within the lesion. The purposes of this study were to characterize the morphology of lung lesions, examine the presence of M. bovis variable surface protein (Vsp) antigens and study the local immune responses in calves after infection with M. bovis strain 1067. Lung tissue samples from eight calves euthanased three weeks after experimental infection with M. bovis were examined by bacteriology and pathology. Lung lesions were evaluated by immunohistochemical (IHC) staining for wide spectrum cytokeratin and for M. bovis Vsp antigens and pMB67 antigen. IHC identification and quantitative evaluation of CD4+ and CD8+ T lymphocytes and immunoglobulin (IgG1, IgG2, IgM, IgA)-containing plasma cells was performed. Additionally, expression of major histocompatibility complex class II (MHC class II) was studied by IHC. Suppurative pneumonic lesions were found in all calves. In two calves with caseonecrotic pneumonia, necrotic foci were surrounded by epithelial cells resembling bronchial or bronchiolar epithelium. In all calves, M. bovis Vsp antigens were constantly present in the cytoplasm of macrophages and were also present extracellularly at the periphery of necrotic foci. There was a considerable increase in numbers of IgG1- and IgG2-positive plasma cells among which IgG1-containing plasma cells clearly predominated. Statistical evaluation of the numbers of CD4+ and CD8+ T cells, however, did not reveal statistically significant differences between inoculated and control calves. In M. bovis infected calves, hyperplasia of bronchus-associated lymphoid tissue (BALT) was characterized by strong MHC class II expression of lymphoid cells, but only few of the macrophages demarcating the caseonecrotic foci were positive for MHC class II. The results from this study show that infection of calves with M. bovis results in various lung lesions including caseonecrotic pneumonia originating from bronchioli and bronchi. There is long-term persistence of M. bovis as demonstrated by bacteriology and immunohistochemistry for M. bovis antigens, i.e. Vsp antigens and pMB67. The persistence of the pathogen and its ability to evade the specific immune response may in part result from local downregulation of antigen presenting mechanisms and an ineffective humoral immune response with prevalence of IgG1 antibodies that, compared to IgG2 antibodies, are poor opsonins.


Genetic diversity as an objective in multi-objective evolutionary algorithms.
A key feature of an efficient and reliable multi-objective evolutionary algorithm is the ability to maintain genetic diversity within a population of solutions. In this paper, we present a new diversity-preserving mechanism, the Genetic Diversity Evaluation Method (GeDEM), which considers a distance-based measure of genetic diversity as a real objective in fitness assignment. This provides a dual selection pressure towards the exploitation of current non-dominated solutions and the exploration of the search space. We also introduce a new multi-objective evolutionary algorithm, the Genetic Diversity Evolutionary Algorithm (GDEA), strictly designed around GeDEM and then we compare it with other state-of-the-art algorithms on a well-established suite of test problems. Experimental results clearly indicate that the performance of GDEA is top-level.


IMMUNOLOGY. An interactive reference framework for modeling a dynamic immune system.
Immune cells function in an interacting hierarchy that coordinates the activities of various cell types according to genetic and environmental contexts. We developed graphical approaches to construct an extensible immune reference map from mass cytometry data of cells from different organs, incorporating landmark cell populations as flags on the map to compare cells from distinct samples. The maps recapitulated canonical cellular phenotypes and revealed reproducible, tissue-specific deviations. The approach revealed influences of genetic variation and circadian rhythms on immune system structure, enabled direct comparisons of murine and human blood cell phenotypes, and even enabled archival fluorescence-based flow cytometry data to be mapped onto the reference framework. This foundational reference map provides a working definition of systemic immune organization to which new data can be integrated to reveal deviations driven by genetics, environment, or pathology.


Role of physical performance component evaluations in occupational therapy functional assessment.
In the 1970s, occupational therapy assessments tended to focus on physical performance component (PPC) evaluations such as strength and range of motion; assessment of occupational performance played a secondary role. Given changes within and outside the profession of occupational therapy, it is time for the assessment of occupation and role performance to assume a primary role, while PPC evaluation should shift to a secondary role. Although PPC evaluations may be less prominent than in the past, they will continue to play an important role in occupational therapy practice and research. Therapists need to be critical consumers of PPC measurement tools because many require further development to validate their use for occupational therapy purposes.


Establishment of three new genera in the family Geminiviridae: Becurtovirus, Eragrovirus and Turncurtovirus.
The family Geminiviridae includes plant-infecting circular single-stranded DNA viruses that have geminate particle morphology. Members of this family infect both monocotyledonous and dicotyledonous plants and have a nearly global distribution. With the advent of new molecular tools and low-cost sequencing, there has been a significant increase in the discovery of new geminiviruses in various cultivated and non-cultivated plants. In this communication, we highlight the establishment of three new genera (Becurtovirus, Eragrovirus and Turncurtovirus) to accommodate various recently discovered geminiviruses that are highly divergent and, in some cases, have unique genome architectures. The genus Becurtovirus has two viral species, Beet curly top Iran virus (28 isolates; leafhopper vector Circulifer haematoceps) and Spinach curly top Arizona virus (1 isolate; unknown vector), whereas the genera Eragrovirus and Turncurtovirus each have a single assigned species: Eragrostis curvula streak virus (6 isolates; unknown vector) and Turnip curly top virus (20 isolates; leafhopper vector Circulifer haematoceps), respectively. Based on analysis of all of the genome sequences available in public databases for each of the three new genera, we provide guidelines and protocols for species and strain classification within these three new genera.


CD133 positive cells isolated from A549 cell line exhibited high liver metastatic potential.
Lung cancer remains a major cause of cancer-related lethality because of high incidence and recurrence in spite of significant advances in staging and therapy. In present study, we identified a subpopulation of cells isolated from the A549 cell line with marker CD133. In vivo results showed that A549 CD133+ cells displayed high liver metastatic potential. Severe liver cell damage with tumor cell invasion revealed by pathological examination and these changes were consistent with the results of serological tests where the plasma GPT and GOT level are significantly higher than that of the control group. Compared with A549 cells, A549 CD133+ cells expressed high levels of VEGF and exhibited high migration and invasion capability. In conclusion, we first reported that A549 CD133+ cells exhibited characteristic of high liver metastatic potential which makes it be a suitable model for further study of liver metastasis of lung adenocarcinoma and provide a potential platform for anti-metastatic drug discovery or evaluation.


The course of phosphorus in the reaction of N-acetyl-L-glutamate kinase, determined from the structures of crystalline complexes, including a complex with an AlF(4)(-) transition state mimic.
N-Acetyl-L-glutamate kinase (NAGK), the structural paradigm of the enzymes of the amino acid kinase family, catalyzes the phosphorylation of the gamma-COO(-) group of N-acetyl-L-glutamate (NAG) by ATP. We determine here the crystal structures of NAGK complexes with MgADP, NAG and the transition-state analog AlF(4)(-); with MgADP and NAG; and with ADP and SO(4)(2-). Comparison of these structures with that of the MgAMPPNP-NAG complex allows to delineate three successive steps during phosphoryl transfer: at the beginning, when the attacking and leaving O atoms and the P atom are imperfectly aligned and the distance between the attacking O atom and the P atom is 2.8A; midway, at the bipyramidal intermediate, with nearly perfect alignment and a distance of 2.3A; and, when the transfer is completed. The transfer occurs in line and is strongly associative, with Lys8 and Lys217 stabilizing the transition state and the leaving group, respectively, and with Lys61, in contrast with an earlier proposal, not being involved. Three water molecules found in all the complexes play, together with Asp162 and the Mg, crucial structural roles. Two glycine-rich loops (beta1-alphaA and beta2-alphaB) are also very important, moving in the different complexes in concert with the ligands, to which they are hydrogen-bonded, either locking them in place for reaction or stabilizing the transition state. The active site is too narrow to accommodate the substrates without compressing the reacting groups, and this compressive strain appears a crucial component of the catalytic mechanism of NAGK, and possibly of other enzymes of the amino acid kinase family such as carbamate kinase. Initial binding of the two substrates would require a different enzyme conformation with a wider active site, and the energy of substrate binding would be used to change the conformation of the active center, causing substrate strain towards the transition state.


Infection control preparedness for human infection with influenza A H7N9 in Hong Kong.
OBJECTIVE To assess the effectiveness of infection control preparedness for human infection with influenza A H7N9 in Hong Kong. DESIGN A descriptive study of responses to the emergence of influenza A H7N9. SETTING A university-affiliated teaching hospital. PARTICIPANTS Healthcare workers (HCWs) with unprotected exposure (not wearing N95 respirator during aerosol-generating procedure) to a patient with influenza A H7N9. METHODS A bundle approach including active and enhanced surveillance, early airborne infection isolation, rapid molecular diagnostic testing, and extensive contact tracing for HCWs with unprotected exposure was implemented. Seventy HCWs with unprotected exposure to an index case were interviewed especially regarding their patient care activities. RESULTS From April 1, 2013, through May 31, 2014, a total of 126 (0.08%) of 163,456 admitted patients were tested for the H7 gene by reverse transcription-polymerase chain reaction per protocol. Two confirmed cases were identified. Seventy (53.8%) of 130 HCWs had unprotected exposure to an index case, whereas 41 (58.6%) and 58 (82.9%) of 70 HCWs wore surgical masks and practiced hand hygiene after patient care, respectively. Sixteen (22.9%) of 70 HCWs were involved in high-risk patient contacts. More HCWs with high-risk patient contacts received oseltamivir prophylaxis (P=0.088) and significantly more had paired sera collected for H7 antibody testing (P<0.001). Ten (14.3%) of 70 HCWs developed influenza-like illness during medical surveillance, but none had positive results by reverse transcription-polymerase chain reaction. Paired sera was available from 33 of 70 HCWs with unprotected exposure, and none showed seroconversion against H7N9. CONCLUSIONS Despite the delay in airborne precautions implementation, no patient-to-HCW transmission of influenza A H7N9 was demonstrated.


Treatment of Hailey-Hailey disease with tacrolimus ointment and clobetasol propionate foam.
Hailey-Hailey disease, or familial benign chronic pemphigus, is a chronic disease without a known cure. Current therapeutic strategies attempt to suppress Hailey-Hailey outbreaks and allow the patient to live comfortably with this condition. We have found that applying topical tacrolimus 0.1% ointment (Protopic) twice a day to affected areas is an excellent way to control Hailey-Hailey disease. In addition to effectively controlling Hailey-Hailey outbreaks, tacrolimus is a relatively safe and noninvasive mode of treatment, without significant side effects. We recommend intermittent therapy with clobetasol propionate 0.05% foam (Olux Foam) for patients who break through suppressive therapy with tacrolimus a few times per year. In patients with frequent outbreaks of Hailey-Hailey disease despite suppressive therapy with tacrolimus, we recommend alternating the tacrolimus with clobetasol propionate 0.05% foam every 6 weeks.


Improved longitudinal magneto-optic Kerr effect signal contrast from nanomagnets with dielectric coatings.
We report on the use of dielectric coatings to improve the contrast of longitudinal magneto-optic Kerr effect signals from submicron magnetic structures. Electron-beam lithography was used to define disks in 22 nm thick Ni films deposited on Si substrates. The structures were measured in four configurations: as-deposited, through a fused silica prism using index-matching fluid, coated with ZnS, and using a prism on top of the ZnS layer. The modified samples show up to 20 times improvement in the MOKE contrast due to admittance matching to the magnetic material and suppression of the substrate reflectance. The behavior is successfully predicted by a model that includes the magneto-optic response of the nickel layer and accounts for the fraction of the beam intercepted by the magnetic structure.


Effective and ineffective management of incontinence: issues around illness trajectory and health care.
In this article, findings from a study that investigated the impact of incontinence on individuals and the impact of effective and ineffective health interventions for the management of incontinence on patient careers and health care are reported. Twenty-seven participants whose incontinence was judged by health professionals as being successfully managed or unsuccessfully managed were interviewed. The participants were 19 women and 8 men (mean age: 61 years). Twenty-two participants experienced urinary incontinence, 4 people experienced both urinary and fecal incontinence, and 1 woman experienced fecal constipation. The findings have been set in the context of the management of chronic conditions and provide illuminating evidence that may be useful when reviewing health care and health services. People who were actively involved with their care and who were involved with decision making felt that their incontinence was being effectively managed. Multiple referrals and the marshaling and targeting of health professionals and services also appeared to be associated with effective management of incontinence.


Helping patients decide about back surgery: a randomized trial of an interactive video program.
A randomized trial of 100 patients with low back pain who were potential surgical candidates. To determine whether an interactive videodisc with a booklet is superior to a booklet alone for informing patients about back surgery. Substantial geographic variation has been observed in lumbar spine surgery. Informed patient preferences should play an important role in decisions about surgery. However, little is known about optimal strategies for informing patients. Subjects were randomized to receive an interactive videodisc (with a booklet) or a booklet alone. A knowledge test administered at baseline and follow-up was used to measure improvement in knowledge about treatment options for lumbar spine problems. Patients' reactions to the videodisc and booklet and preferences for treatment were also assessed. The patients' knowledge improved after exposure to either intervention. Multivariate analyses adjusted for baseline score, age, education, gender, and diagnosis showed a significant advantage for the videodisc with booklet over the booklet alone. The videodisc-booklet group showed significantly greater gains in knowledge among subjects with the worst baseline knowledge scores. A larger proportion of subjects in the videodisc-booklet group rated the materials easy to understand (93% vs.- 72%,P = 0.04), containing the right amount of information (93% vs.- 80%,P = 0.3), and adequate to assist in choice of treatment (75% vs.- 51%,P = 0.2). Those who viewed the videodisc expressed a somewhat lower preference for surgery than those who received the booklet alone (23% vs.- 42%,P = 0.4). Both the booklet alone and the combination of videodisc and booklet improved knowledge. The combination produced greater knowledge gains than the booklet alone for the subgroup with the least knowledge at baseline. Patients preferred the combination and had a slightly lower preference for surgery if they had viewed the video presentation. For some patients, the video may enhance involvement in clinical decisions.


The 2.0 A structure of malarial purine phosphoribosyltransferase in complex with a transition-state analogue inhibitor.
Malaria is a leading cause of worldwide mortality from infectious disease. Plasmodium falciparum proliferation in human erythrocytes requires purine salvage by hypoxanthine-guanine-xanthine phosphoribosyltransferase (HGXPRTase). The enzyme is a target for the development of novel antimalarials. Design and synthesis of transition-state analogue inhibitors permitted cocrystallization with the malarial enzyme and refinement of the complex to 2.0 A resolution. Catalytic site contacts in the malarial enzyme are similar to those of human hypoxanthine-guanine phosphoribosyltransferase (HGPRTase) despite distinct substrate specificity. The crystal structure of malarial HGXPRTase with bound inhibitor, pyrophosphate, and two Mg(2+) ions reveals features unique to the transition-state analogue complex. Substrate-assisted catalysis occurs by ribooxocarbenium stabilization from the O5' lone pair and a pyrophosphate oxygen. A dissociative reaction coordinate path is implicated in which the primary reaction coordinate motion is the ribosyl C1' in motion between relatively immobile purine base and (Mg)(2)-pyrophosphate. Several short hydrogen bonds form in the complex of the enzyme and inhibitor. The proton NMR spectrum of the transition-state analogue complex of malarial HGXPRTase contains two downfield signals at 14.3 and 15.3 ppm. Despite the structural similarity to the human enzyme, the NMR spectra of the complexes reveal differences in hydrogen bonding between the transition-state analogue complexes of the human and malarial HG(X)PRTases. The X-ray crystal structures and NMR spectra reveal chemical and structural features that suggest a strategy for the design of malaria-specific transition-state inhibitors.


Identification of ABC Transporter Interaction of a Novel Cyanoquinoline Radiotracer and Implications for Tumour Imaging by Positron Emission Tomography.
The epidermal growth factor receptor (EGFR) is overexpressed in many cancers including lung, ovarian, breast, head and neck and brain. Mutation of this receptor has been shown to play a crucial role in the response of non-small cell lung carcinoma (NSCLC) to EGFR-targeted therapies. It is envisaged that imaging of EGFR using positron emission tomography (PET) could aid in selection of patients for treatment with novel inhibitors. We recognised multi-drug resistant phenotype as a threat to development of successful imaging agents. In this report, we describe discovery of a novel cyanoquinoline radiotracer that lacks ABC transporter activity. Cellular retention of the prototype cyanoquinoline [18F](2E)-N-{4-[(3-chloro-4-fluorophenyl)amino]-3-cyano-7-ethoxyquinolin-6-yl}-4-({[1-(2-fluoroethyl)-1H-1,2,3-triazol-4-yl]methyl}amino)-but-2-enamide ([18F]FED6) and [18F](2E)-N-{4-[(3-chloro-4-fluorophenyl)amino]-3-cyano-7-ethoxyquinolin-6-yl}-4-[({1-[(2R,5S)-3-fluoro-4,5-dihydroxy-6-(hydroxymethyl)oxan-2-yl]-1H-1,2,3-triazol-4-yl}methyl)amino]but-2-enamide ([18F]FED20) were evaluated to establish potential for imaging specificity. The substrate specificity of a number of cyanoquinolines towards ABC transporters was investigated in cell lines proficient or deficient in ABCB1 or ABCG2. FED6 demonstrated substrate specificity for both ABCG2 and ABCB1, a property that was not observed for all cyanoquinolines tested, suggesting scope for designing novel probes. ABC transporter activity was confirmed by attenuating the activity of transporters with drug inhibitors or siRNA. We synthesized a more hydrophilic compound [18F]FED20 to overcome ABC transporter activity. FED20 lacked substrate specificity for both ABCB1 and ABCG2, and maintained a strong affinity for EGFR. Furthermore, FED20 showed higher inhibitory affinity for active mutant EGFR versus wild-type or resistant mutant EGFR; this property resulted in higher [18F]FED20 cellular retention in active mutant EGFR expressing NSCLC. [18F]FED20 binds EGFR but is devoid of ABC transporter activity, thus, has potential for EGFR imaging.


Efficient temporal shaping of ultrashort pulses with birefringent crystals.
We report on a simple and robust technique to temporally shape ultrashort pulses. A number of birefringent crystals with appropriate crystal length and orientation form a crystal set. When a short pulse propagates through the crystal set, the pulse is divided into numerous pulses, producing a desired temporal shape. Flexibility in the final pulse shape is achieved through varying initial pulse duration, divided-pulse number, the polarization-mode delay, and energy distribution of the divided pulses. The energy efficiency of the technique is near 100% for a pulse train of alternating polarizations, and 50% for a linearly polarized pulse train.


Evaluation of Oral Cavity DNA Extraction Methods on Bacterial and Fungal Microbiota.
The objective of this study was to evaluate the most effective method of DNA extraction of oral mouthwash samples for use in microbiome studies that utilize next generation sequencing (NGS). Eight enzymatic and mechanical DNA extraction methods were tested. Extracted DNA was amplified using barcoded primers targeting the V6 variable region of the bacterial 16S rRNA gene and the ITS1 region of the fungal ribosomal gene cluster and sequenced using the Illumina NGS platform. Sequenced reads were analyzed using QIIME and R. The eight methods yielded significantly different quantities of DNA (p < 0.001), with the phenol-chloroform extraction method producing the highest total yield. There were no significant differences in observed bacterial or fungal Shannon diversity (p = 0.64, p = 0.93 respectively) by extraction method. Bray-Curtis beta-diversity did not demonstrate statistically significant differences between the eight extraction methods based on bacterial (R2 = 0.086, p = 1.00) and fungal (R2 = 0.039, p = 1.00) assays. No differences were seen between methods with or without bead-beating. These data indicate that choice of DNA extraction method affect total DNA recovery without significantly affecting the observed microbiome.


Genome Analysis of Rhodococcus Sp. DSSKP-R-001: A Highly Effective β-Estradiol-Degrading Bacterium.
We screened bacteria that use E2 as its sole source of carbon and energy for growth and identified them as Rhodococcus, and we named them DSSKP-R-001. For a better understanding of the metabolic potential of the strain, whole genome sequencing of Rhodococcus DSSKP-R-001 and annotation of the functional genes were performed. The genomic sketches included a predicted protein-coding gene of approximately 5.4 Mbp with G + C content of 68.72% and 5180. The genome of Rhodococcus strain DSSKP-R-001 consists of three replicons: one chromosome and two plasmids of 5.2, 0.09, and 0.09, respectively. The results showed that there were ten steroid-degrading enzymes distributed in the whole genome of the strain. The existence and expression of estradiol-degrading enzymes were verified by PCR and RTPCR. Finally, comparative genomics was used to compare multiple strains of Rhodococcus. It was found that Rhodococcus DSSKP-R-001 had the highest similarity to Rhodococcus sp. P14 and there were 2070 core genes shared with Rhodococcus sp. P14, Rhodococcus jostii RHA1, Rhodococcus opacus B4, and Rhodococcus equi 103S, showing evolutionary homology. In summary, this study provides a comprehensive understanding of the role of Rhodococcus DSSKP-R-001 in estradiol-efficient degradation of these assays for Rhodococcus. DSSKP-R-001 in bioremediation and evolution within Rhodococcus has important meaning.


Infrared mass spectrometric imaging below the diffraction limit.
Matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS)1 is an established technique for the analysis of biological macromolecules. Its relative insensitivity to pollutants makes MALDI-MS very suitable for the direct analysis of biological samples. As such, it has facilitated great advances in the field of biomolecular imaging mass spectrometry. Traditionally, MALDI-MS imaging is performed in a scanning microprobe methodology.(2-4) However, in a recent study we have demonstrated an alternative methodology; the so-called microscope mode,(5) where the requirement for a highly focused ionization beam is removed. Spatial details from within the desorption area are conserved during the flight of the ions through the mass analyzer, and a magnified ion image is projected onto a 2D-detector. In this paper, we demonstrate how imaging mass spectrometry benefits from the microscope mode approach. For the first time, high-lateral resolution ion images were recorded using infrared MALDI at 2.94 microm wavelength. The ion optical resolution achieved was well below the theoretical limit of (light-) diffraction for the setup used, which is impossible to achieve in the conventional scanning microprobe approach.


Dietary carbohydrate composition is associated with polycystic ovary syndrome: a case-control study.
The present study aimed to investigate the association between dietary carbohydrate components and polycystic ovary syndrome (PCOS) in Iran. In this case-control study, the diagnosis of PCOS was made based on the Rotterdam criteria in hospital clinics. Dietary assessments were performed using a validated semi-quantitative food frequency questionnaire. In total, 281 women with incident PCOS and 472 age-matched controls were assessed. Participants were interviewed through the clinics in Tehran, Iran, from February 2012 until March 2014. Average dietary glycaemic index (GI) and glycaemic load (GL) were calculated using GI of Iranian Foods Table and international tables of GI and GL values. We also assessed total dietary carbohydrate, refined grains, whole grains and fibre intakes. Participation rates were 97.5% among cases and 96.3% among controls. Mean (SD) dietary GI values among the controls and cases were 51.8 (4.7) and 59.7 (5.9) (P = 0.02) and GL values were 155.34 (35.2) and 173.6 (39.1) (P < 0.001), respectively. The multivariate adjusted odds ratio (OR) comparing the highest tertile of dietary GI and GL with the lowest tertile were 2.18 [95% confidence interval (CI) = 1.29-3.81; P-test for trend = 0.012] and 2.39 (95% CI = 1.23-3.01; P-test for trend = 0.001), respectively, with a significant trend. Fibre intake was inversely associated with PCOS (OR = 0.73; 95% CI = 0.49-0.91; P-test for trend = 0.013). The findings of the present study suggest that high dietary GI and GL and low fibre intake are significantly associated with PCOS.


Chemotherapy-induced ileal crypt apoptosis and the ileal microbiome shape immunosurveillance and prognosis of proximal colon cancer.
The prognosis of colon cancer (CC) is dictated by tumor-infiltrating lymphocytes, including follicular helper T (TFH) cells and the efficacy of chemotherapy-induced immune responses. It remains unclear whether gut microbes contribute to the elicitation of TFH cell-driven responses. Here, we show that the ileal microbiota dictates tolerogenic versus immunogenic cell death of ileal intestinal epithelial cells (IECs) and the accumulation of TFH cells in patients with CC and mice. Suppression of IEC apoptosis led to compromised chemotherapy-induced immunosurveillance against CC in mice. Protective immune responses against CC were associated with residence of Bacteroides fragilis and Erysipelotrichaceae in the ileum. In the presence of these commensals, apoptotic ileal IECs elicited PD-1+ TFH cells in an interleukin-1R1- and interleukin-12-dependent manner. The ileal microbiome governed the efficacy of chemotherapy and PD-1 blockade in CC independently of microsatellite instability. These findings demonstrate that immunogenic ileal apoptosis contributes to the prognosis of chemotherapy-treated CC.


Evaluation of two enzyme-linked immunosorbent assays for detecting Salmonella enterica subsp. enterica Serovar Dublin antibodies in bulk milk.
Two enzyme-linked immunosorbent assays (ELISAs) for the detecting Salmonella enterica subsp. enterica serovar Dublin antibodies in bulk milk were developed and evaluated for potential use in control programs. The ELISAs were based on either lipopolysacharide (LPS ELISA) or flagellar antigen (GP ELISA). Sensitivity was determined with 79 case herds with a wide range of clinical signs. Specificity was determined with 125 Dutch and 200 Swedish control herds. The relation between antibodies in bulk milk, antibodies in serum, and the level of milk production of individual cows was studied with 61 case herds. The optimal optical density (OD) values of the LPS ELISA and the GP ELISA were determined to be 0.2 and 0.5, respectively. The sensitivities of the LPS ELISA and the GP ELISA were 54 and 63%, respectively, with a specificity of 98% for both ELISAs with samples from the Dutch control herds. The specificities for samples from the Swedish herds were 100% for the LPS ELISA and 95% for the GP ELISA. The sensitivity of the combination of tests was 65% when samples were run in parallel, and the specificity was 100% when samples were run in series, irrespective of whether the samples came from Dutch or Swedish control herds. The variance (R(2)) in the OD value for bulk milk samples could be explained by the percentage of seropositive lactating cows in a herd with the LPS ELISA for 51% of the samples and with the GP ELISA for 72%. The variance in the OD value was best explained by the combination of the percentage of seropositive lactating cows in the herd and the mean log(10) serum antibody titer for that herd (R(2) = 62% for the LPS ELISA and R(2) = 75% for the GP ELISA). Case herds more often tested negative by the ELISA with bulk milk when the percentage of seropositive lactating cows was less than 5%. It is concluded that both ELISAs with bulk milk can be used in control programs to distinguish between infected and noninfected herds. Specificity can be increased by using the two tests in combination. Sensitivity was relatively low for both single tests and both tests combined.


The Reproductive Ecology of Industrial Societies, Part II : The Association between Wealth and Fertility.
Studies of the association between wealth and fertility in industrial populations have a rich history in the evolutionary literature, and they have been used to argue both for and against a behavioral ecological approach to explaining human variability. We consider that there are strong arguments in favor of measuring fertility (and proxies thereof) in industrial populations, not least because of the wide availability of large-scale secondary databases. Such data sources bring challenges as well as advantages, however. The purpose of this article is to illustrate these by examining the association between wealth and reproductive success in the United States, using the National Longitudinal Study of Youth 1979. We conduct a broad-based exploratory analysis of the relationship between wealth and fertility, employing both cross-sectional and longitudinal approaches, and multiple measures of both wealth (income and net worth) and fertility (lifetime reproductive success and transitions to first, second and third births). We highlight the kinds of decisions that have to be made regarding sample selection, along with the selection and construction of explanatory variables and control measures. Based on our analyses, we find a positive effect of both income and net worth on fertility for men, which is more pronounced for white men and for transitions to first and second births. Income tends to have a negative effect on fertility for women, while net worth is more likely to positively predict fertility. Different reproductive strategies among different groups within the same population highlight the complexity of the reproductive ecology of industrial societies. These results differ in a number of respects from other analyses using the same database. We suggest this reflects the impossibility of producing a definitive analysis, rather than a failure to identify the "correct" analytical strategy. Finally, we discuss how these findings inform us about (mal)adaptive decision-making.


It takes more than one for parenting: How do maternal temperament and child's conduct problems relate to maternal parenting behavior?
The current study examined how individual differences in maternal temperament and child problem behaviors correlate with observed maternal positivity and negativity toward the child. The sample consisted of 153 mothers of 3-to-7 year old children. Mothers reported their own temperament (surgency, orienting sensitivity, effortful control and negative affect) and their children's problem behaviors. Maternal behavior was videotaped in a set of structured interaction tasks with the child during a lab visit. Results indicated that children's problem behaviors were related to less maternal positivity and more negativity. In addition, observed maternal negativity was associated with less maternal effortful control and more negative affect. In contrast, maternal temperament was unrelated to observed maternal positivity toward the child. Furthermore, maternal temperament was related to mothers' positivity and negativity but only for children high in problem behaviors. The findings implicate that child problem behaviors may interact with maternal temperament in explaining variance in caregiving positivity and negativity.


Validation of a clinical algorithm to identify neonates with severe illness during routine household visits in rural Bangladesh.
To validate a clinical algorithm for community health workers (CHWs) during routine household surveillance for neonatal illness in rural Bangladesh. Surveillance was conducted in the intervention arm of a trial of newborn interventions. CHWs assessed 7587 neonates on postnatal days 0, 2, 5 and 8 and identified neonates with very severe disease (VSD) using an 11-sign algorithm. A nested prospective study was conducted to validate the algorithm (n=395). Physicians evaluated neonates to determine whether newborns with VSD needed referral. The authors calculated algorithm sensitivity and specificity in identifying (1) neonates needing referral and (2) mortality during the first 10 days of life. The 11-sign algorithm had sensitivity of 50.0% (95% CI 24.7% to 75.3%) and specificity of 98.4% (96.6% to 99.4%) for identifying neonates needing referral-level care. A simplified 6-sign algorithm had sensitivity of 81.3% (54.4% to 96.0%) and specificity of 96.0% (93.6% to 97.8%) for identifying referral need and sensitivity of 58.0% (45.5% to 69.8%) and specificity of 93.2% (92.5% to 93.7%) for screening mortality. Compared to our 6-sign algorithm, the Young Infant Study 7-sign (YIS7) algorithm with minor modifications had similar sensitivity and specificity. Community-based surveillance for neonatal illness by CHWs using a simple 6-sign clinical algorithm is a promising strategy to effectively identify neonates at risk of mortality and needing referral to hospital. The YIS7 algorithm was also validated with high sensitivity and specificity at community level, and is recommended for routine household surveillance for newborn illness. ClinicalTrials.gov no. NCT00198627.


Direct examination of the relevance for folding, binding and electron transfer of a conserved protein folding intermediate.
Near the minimum free energy basin of proteins where the native ensemble resides, partly unfolded conformations of slightly higher energy can be significantly populated under native conditions. It has been speculated that they play roles in molecular recognition and catalysis, but they might represent contemporary features of the evolutionary process without functional relevance. Obtaining conclusive evidence on these alternatives is difficult because it requires comparing the performance of a given protein when populating and when not populating one such intermediate, in otherwise identical conditions. Wild type apoflavodoxin populates under native conditions a partly unfolded conformation (10% of molecules) whose unstructured region includes the binding sites for the FMN cofactor and for redox partner proteins. We recently engineered a thermostable variant where the intermediate is no longer detectable. Using the wild type and variant, we assess the relevance of the intermediate comparing folding kinetics, cofactor binding kinetics, cofactor affinity, X-ray structure, intrinsic dynamics, redox potential of the apoflavodoxin-cofactor complex (Fld), its affinity for partner protein FNR, and electron transfer rate within the Fld/FNR physiological complex. Our data strongly suggest the intermediate state, conserved in long-chain apoflavodoxins, is not required for the correct assembly of flavodoxin nor does it contribute to shape its electron transfer properties. This analysis can be applied to evaluate other native basin intermediates.


A system for the analysis of seat support surfaces using surface shape control and simultaneous measurement of applied pressures.
A system for the design and analysis of seat support and buttock tissue interfaces has been developed. It has the ability to control the seating surface shape while measuring the pressure applied to the buttocks by the surface. Pressures are measured over an 11 x 12 rectangular array of support elements using silicon pressure sensors mounted in a swiveling head atop each support element. Control of surface shape is mediated by selective linear translation of the support elements along their respective vertical axes. Closed-loop control of the system allows for the dynamic formulation of a support surface on the basis of programmable criteria. The system is intended to function as a research tool to facilitate the study of the relationships between support surface shape and interface pressure, and support surface shape and soft tissue distortion. The purpose of this paper is to present the system instrumentation and the rationale behind its design and development. The paper also presents the results of several tests to evaluate the accuracy and performance of the system. This evaluation included a pilot study on 10 able-bodied subjects. The results of these system evaluations indicate that the system is capable of making repeatable and precise measurements of pressure and surface element position and can formulate support surface shapes that satisfy specified optimization criteria.


Effectiveness of a mobile smoking cessation service in reaching elderly smokers and predictors of quitting.
Different smoking cessation programmes have been developed in the last decade but utilization by the elderly is low. We evaluated a pilot mobile smoking cessation service for the Chinese elderly in Hong Kong and identified predictors of quitting. The Mobile Smoking Cessation Programme (MSCP) targeted elderly smokers (aged 60 or above) and provided service in a place that was convenient to the elderly. Trained counsellors provided individual counselling and 4 week's free supply of nicotine replacement therapy (NRT). Follow up was arranged at 1 month by face-to-face and at 3 and 6 months by telephone plus urinary cotinine validation. A structured record sheet was used for data collection. The service was evaluated in terms of process, outcome and cost. 102 governmental and non-governmental social service units and private residential homes for the elderly participated in the MSCP. We held 90 health talks with 3266 elderly (1140 smokers and 2126 non-smokers) attended. Of the 1140 smokers, 365 (32%) received intensive smoking cessation service. By intention-to-treat, the validated 7 day point prevalence quit rate was 20.3% (95% confidence interval: 16.2%-24.8%). Smoking less than 11 cigarettes per day and being adherent to NRT for 4 weeks or more were significant predictors of quitting. The average cost per contact was US$54 (smokers only); per smoker with counselling: US$168; per self-reported quitter: US$594; and per cotinine validated quitter: US$827. This mobile smoking cessation programme was acceptable to elderly Chinese smokers, with quit rate comparable to other comprehensive programmes in the West. A mobile clinic is a promising model to reach the elderly and probably other hard to reach smokers.


The KLHL12-Cullin-3 ubiquitin ligase negatively regulates the Wnt-beta-catenin pathway by targeting Dishevelled for degradation.
Dishevelled is a conserved protein that interprets signals received by Frizzled receptors. Using a tandem-affinity purification strategy and mass spectrometry we have identified proteins associated with Dishevelled, including a Cullin-3 ubiquitin ligase complex containing the Broad Complex, Tramtrack and Bric à Brac (BTB) protein Kelch-like 12 (KLHL12). This E3 ubiquitin ligase complex is recruited to Dishevelled in a Wnt-dependent manner that promotes its poly-ubiquitination and degradation. Functional analyses demonstrate that regulation of Dishevelled by this ubiquitin ligase antagonizes the Wnt-beta-catenin pathway in cultured cells, as well as in Xenopus and zebrafish embryos. Considered with evidence that the distinct Cullin-1 based SCF(beta-TrCP)complex regulates beta-catenin stability, our data on the stability of Dishevelled demonstrates that two distinct ubiquitin ligase complexes regulate the Wnt-beta-catenin pathway.


Effect of Ni ion release on the cells in contact with NiTi alloys.
Nickel-titanium alloys have been used in medical applications for several years; however, biocompatibility of the material remains controversial. In the present study, the human umbilical vein endothelial cells (HUVEC) were cultured in contact with the nitinol used in two different heat treatment surface modifications-helium and hydrogen. The amount of Ni ions released from these alloys in contact with HUVEC was measured in media and in the cells by ICP-MS. An increased release of Ni ions was detected in He alloy compared with H2 alloy modification with an elevation with the metal exposition duration (24 h vs. 72 h). The cells contained the Ni ions in both selected alloy modifications with the lower levels in H2 alloys. To evaluate the potential of multiple metal applications, similar values were observed in media and in cell suspension for all surface modification combinations. The model analysis of effect of metal ion release on distant cells in the body showed that the concentration is interestingly similar to concentrations in cells in direct contact with the metal alloy. The cells are able to regulate the concentration of Ni ions within the cell. According to our best knowledge, the study for the first time describes the presence of Ni ions released from nitinol directly in the cells. In the case of the H2 modification, the lowest levels of Ni ions were detected both in medium and in the cells, which likely increases the biocompatibility of the nitinol alloy.


Geographic Variation in Daily Temporal Activity Patterns of a Neotropical Marsupial (Gracilinanus agilis).
The temporal activity of animals is an outcome of both biotic and abiotic factors, which may vary along the geographic range of the species. Therefore, studies conducted with a species in different localities with distinct features could elucidate how animals deal with such factors. In this study, we used live traps equipped with timing devices to investigate the temporal activity patterns of the didelphid Gracilinanus agilis in two dry-woodland areas of the Brazilian savanna (Cerrado). These areas were located about 660 km apart, one in Central Brazil and the other in Southeastern Brazil. We compared such patterns considering both reproductive and non-reproductive periods, and how it varies as a function of temperature on a seasonal basis. In Central Brazil, we found a constant, and temperature-independent activity during the night in both reproductive and non-reproductive periods. On the other hand, in Southeastern Brazil, we detected a constant activity during the reproductive period, but in the non-reproductive period G. agilis presented a peak of activity between two and four hours after sunset. Moreover, in this latter we found a relation between temporal activity and temperature during the autumn and spring. These differences in temporal activity between areas, observed during the non-reproductive period, might be associated with the higher seasonal variability in temperature, and lower mean temperatures in the Southeastern site in comparison to the Central one. In Southeastern Brazil, the decrease in temperature during the non-reproductive season possibly forced G. agilis to be active only at certain hours of the night. However, likely due to the reproductive activities (intensive foraging and searching for mates) this marsupial showed constant, temperature-independent activity during the night in the reproductive period at both sites.


Age at diagnosis of diabetes in Appalachia.
Appalachia is a region of the United States noted for the poverty and poor health outcomes of its residents. Residents of the poorest Appalachian counties have a high prevalence of diabetes and risk factors (obesity, low income, low education, etc.) for type 2 diabetes. However, diabetes prevalence exceeds what these risk factors alone explain. Based on this, the history of poor health outcomes in Appalachia, and personally observed high rates of childhood obesity and lack of concern about prediabetes, we speculated that people in Appalachia with diagnosed diabetes might tend to be diagnosed younger than their non-Appalachian counterparts. We used data from the Behavioral Risk Factor Surveillance System (2006-2008). We compared age at diagnosis among counties by Appalachian Regional Commission-defined level of economic development. To account for risk differences, we constructed a model for average age at diagnosis of diabetes, adjusting for county economic development, obesity, income, sedentary lifestyle, and other covariates. After adjustment for risk factors for diabetes, people in distressed or at-risk counties (the least economically developed) had their diabetes diagnosed two to three years younger than comparable people in non-Appalachian counties. No significant differences between non-Appalachian counties and Appalachian counties at higher levels of economic development remained after adjusting. People in distressed and at-risk counties have poor access to care, and are unlikely to develop diabetes at the same age as their non-Appalachian counterparts but be diagnosed sooner. Therefore, people in distressed and at-risk counties are likely developing diabetes at younger ages. We recommend that steps to reduce health disparities between the poorest Appalachian counties and non-Appalachian counties be considered.


Loss of miR-223 and JNK Signaling Contribute to Elevated Stathmin in Malignant Pleural Mesothelioma.
Malignant pleural mesothelioma (MPM) is often fatal, and studies have revealed that aberrant miRNAs contribute to MPM development and aggressiveness. Here, a screen of miRNAs identified reduced levels of miR-223 in MPM patient specimens. Interestingly, miR-223 targets Stathmin (STMN1), a microtubule regulator that has been associated with MPM. However, whether miR-223 regulates STMN1 in MPM and the functions of miR-223 and STMN1 in this disease are yet to be determined. STMN1 is also regulated by c-Jun N-terminal kinase (JNK) signaling, but whether this occurs in MPM and whether miR-223 plays a role are unknown. The relationship between STMN1, miR-223, and JNK was assessed using MPM cell lines, cells from pleural effusions, and MPM tissue. Evidence indicates that miR-223 is decreased in all MPM tissue compared with normal/healthy tissue. Conversely, STMN1 expression was higher in MPM cell lines when compared with primary mesothelial cell controls. Following overexpression of miR-223 in MPM cell lines, STMN1 levels were reduced, cell motility was inhibited, and tubulin acetylation induced. Knockdown of STMN1 using siRNAs led to inhibition of MPM cell proliferation and motility. Finally, miR-223 levels increased while STMN1 was reduced following the re-expression of the JNK isoforms in JNK-null murine embryonic fibroblasts, and STMN1 was reduced in MPM cell lines following the activation of JNK signaling. miR-223 regulates STMN1 in MPM, and both are in turn regulated by the JNK signaling pathway. As such, miR-223 and STMN1 play an important role in regulating MPM cell motility and may be therapeutic targets.


Enhanced conditioning of adverse memories in the mouse modified swim test is associated with neuroinflammatory changes - Effects that are susceptible to antidepressants.
Deficient learning and memory are well-established pathophysiologic features of depression, however, mechanisms of the enhanced learning of aversive experiences associated with this disorder are poorly understood. Currently, neurobiological mechanisms of enhanced retention of aversive memories during depression, and, in particular, their relation to neuroinflammation are unclear. As the association between major depressive disorder and inflammation has been recognized for some time, we aimed to address whether neuroinflammatory changes are involved in enhanced learning of adversity in a depressive state. To study this question, we used a recently described mouse model of enhanced contextual conditioning of aversive memories, the modified forced swim model (modFST). In this model, the classic two-day forced swim is followed by an additional delayed session on Day 5, where increased floating behaviour and upregulated glycogen synthase kinase-3 (GSK-3) are context-dependent. Here, increased time spent floating on Day 5, a parameter of enhanced learning of the adverse context, was accompanied by hypercorticosteronemia, increased gene expression of GSK-3α, GSK-3β, c-Fos, cyclooxygenase-1 (COX-1) and pro-inflammatory cytokines interleukin-1 beta (IL-1β), tumor necrosis factor (TNF), and elevated concentrations of protein carbonyl, a marker of oxidative stress, in the prefrontal cortex and hippocampus. There were significant correlations between cytokine levels and GSK-3β gene expression. Two-week administration of compounds with antidepressant properties, imipramine (7 mg/kg/day) or thiamine (vitamin B1; 200 mg/kg/day) ameliorated most of the modFST-induced changes. Thus, enhanced learning of adverse memories is associated with pro-inflammatory changes that should be considered for optimizing pharmacotherapy of depression associated with enhanced learning of aversive memories.


Cumulative contextual and individual disadvantages over the life course and adult functional somatic symptoms in Sweden.
Disadvantage, originating in one's residential context or in one's past life course, has been shown to impact on health in adulthood. There is however little research on the accumulated health impact of both neighbourhood and individual conditions over the life course. This study aims to examine whether the accumulation of contextual and individual disadvantages from adolescence to middle-age predicts functional somatic symptoms (FSS) in middle-age, taking baseline health into account. The sample is the age 16, 21, 30 and 42 surveys of the prospective Northern Swedish Cohort, with analytical sample size n = 910 (85% of the original cohort). FSS at age 16 and 42, and cumulative socioeconomic disadvantage, social adversity and material adversity between 16 and 42 years were operationalized from questionnaires, and cumulative neighbourhood disadvantage between 16 and 42 years from register data. Results showed accumulation of disadvantages jointly explained 9-12% of FSS variance. In the total sample, cumulative neighbourhood and socioeconomic disadvantage significantly predicted FSS at age 42 in the total sample. In women, neighbourhood disadvantage but not socioeconomic disadvantage contributed significantly, whereas in men, socioeconomic but not neighbourhood disadvantage contributed significantly. In all analyses, associations were largely explained by the parallel accumulation of social and material adversities, but not by symptoms at baseline. In conclusion, the accumulation of diverse forms of disadvantages together plays an important role for somatic complaints in adulthood, independently of baseline health.


Effects of N-acetylcysteine on lead-exposed PC-12 cells.
The neurotoxicity of lead has been well established through numerous studies. However, the cellular processes of lead neurotoxicity, as well as techniques to prevent or reverse cellular damage after lead exposure, remain unknown. If oxidative stress plays a primary role in lead-induced neurotoxicity, antioxidants should assist in reviving lead-exposed cells. The present study explores N-acetylcysteine (NAC) as an antioxidant agent in PC-12 cells after lead exposure. Selective oxidative stress parameters, including glutathione (GSH), glutathione disulfide (GSSG), and malondialdehyde (MDA), were measured in PC-12 cells exposed to various concentrations of lead acetate. Administering NAC after lead exposure improved cell survival as measured by Trypan Blue exclusion. NAC treatment also increased the GSH/GSSG ratio compared to the lead-only group, and reduced MDA to near control levels. These results imply that NAC protects cells from lead-induced oxidative damage by boosting the PC-12 cells' antioxidant defense mechanisms.


The emergence of cardiovascular disease during urbanisation of Africans.
To review the available data on risk factors for cardiovascular disease (CVD), the influence of urbanisation of Africans on these risk factors, and to examine why stroke emerges as a higher risk than ischaemic heart disease (IHD) in the health transition of black South Africans. A review of published data on mortality from and risk factors of CVD in South Africans. South Africa. South African population groups and communities. The available data on the contribution of stroke and IHD to CVD mortality in South Africa are briefly reviewed, followed by a comparison of published data on the prevalence and/or levels of CVD risk factors in the different South African population groups. The impact of urbanisation of black South Africans on these risk factors is assessed by comparing rural and urban Africans who participated in the Transition and Health during Urbanisation of South Africans (THUSA) study. The mortality rates from CVD confirmed that stroke is a major public health problem amongst black South Africans, possibly because of an increase in hypertension, obesity, smoking habit and hyperfibrinogenaemia during various stages of urbanisation. The available data further suggest that black South Africans may be protected against IHD because of favourable serum lipid profiles (low cholesterol and high ratios of high-density lipoprotein cholesterol) and low homocysteine values. However, increases in total fat and animal protein intake of affluent black South Africans, who can afford Western diets, are associated with increases in body mass indices of men and women and in total serum cholesterol. These exposures may increase IHD risk in the future.


Low-Dose Gamma Knife Radiosurgery for Acromegaly.
Remission rate is associated with higher dose of Gamma Knife Radiosurgery (GKRS; Gamma Knife: Elekta AB, Stockholm, Sweden) for acromegaly, but the dose ≥25 Gy is not always feasible when the functioning adenoma is close to optic apparatus. To evaluate the efficacy and safety of low-dose (<25 Gy) GKRS in the treatment of patients with acromegaly. Single-center retrospective review of acromegaly cases treated with GKRS between June 1994 and December 2016. A total of 76 patients with the diagnosis of acromegaly who were treated with low-dose GKRS were selected for inclusion. Patients were treated with a median margin dose, isodose line, and treatment volume of 15.8 Gy, 57.5%, and 4.8 mL, respectively. Any identifiable portion of the optic apparatus was limited to a radiation dose of 10 Gy. All patients underwent full endocrine, ophthalmological, and imaging evaluation prior to and after GKRS treatments, and results of these were analyzed. Biochemical remission was achieved in 33 (43.4%) patients. Actuarial remission rates were 20.3%, 49.9%, and 76.3% at 4, 8, and 12 yr, respectively. Absence of cavernous sinus invasion (P = .042) and lower baseline insulin-like growth factor-1 levels (P = .019) were significant predictors of remission. New hormone deficiencies were found in 9 (11.8%) patients. Actuarial hormone deficiency rates were 3%, 14%, and 22.2% at 4, 8, and 10 yr, respectively. Two (2.6%) patients who achieved initial remission experienced recurrence. No optic complications were encountered. Reasonable remission and new hormone deficiency rates can be achieved with low-dose GKRS for acromegaly. These rates may be comparable to those with standard GKRS margin doses.


Expert Statement on the Single-Agent Use of Inhaled Bronchodilator in the Treatment of Stable Mild-Moderate Chronic Obstructive Pulmonary Disease.
To describe the evidence- and experience-based expert consensus on the use of single-agent bronchodilators in patients with stable mild-moderate chronic obstructive pulmonary disease (COPD). Using Delphi methodology, a panel of 7 respiratory medicine experts was established, who, in the first nominal group meeting defined the scope, users, and document sections. The panel drew up 14 questions on the use of single-agent bronchodilators in patients with mild-moderate stable COPD to be answered with a systematic review of the literature. The results of the review were discussed in a second nominal group meeting and 17 statements were generated. Agreement/disagreement with the statements was tested among16 different experts including respiratory medicine experts and primary care physicians. Statements were scored from1 (total disagreement) to10 (total agreement). Agreement was considered if at least 70% voted ≥7. The level of evidence and grade of recommendation of the systematic literature review was assessed using the Oxford Centre for Evidence-based Medicine levels. A total of 12 of the 17 statements were selected. Specific statements were generated on different profiles of patients with stable mild-moderate COPD in whom single-agent bronchodilators could be prescribed. These statements on the use of single-agent bronchodilators might improve the outcomes and prognosis of patients with stable mild-moderate COPD.


Early Placebo Improvement Is a Marker for Subsequent Placebo Response in Long-Acting Injectable Antipsychotic Trials for Schizophrenia: Combined Analysis of 4 RCTs.
Placebo effects remain largely unexplored in clinical trials of long-acting injectable (LAI) antipsychotics for schizophrenia. This study aims to characterize patients showing improvements after placebo injections and to search for criteria for the prediction of subsequent response based on the magnitude of score changes after the first week of treatment. Data from 450 patients with schizophrenia (DSM-IV) who received placebo injections in 4 double-blind randomized controlled trials evaluating efficacy of LAI paliperidone palmitate obtained through the Yale University Open Data Access (YODA) project were analyzed. These 4 studies were conducted from October 2003 to March 2008. Multiple logistic regression analyses were conducted to examine associations between placebo response and demographic and clinical characteristics. The predictive power of improvement at week 1 for response at week 9 was investigated; sensitivity and specificity of incremental 5% cutoff points between a 5% and 25% reduction in Positive and Negative Syndrome Scale (PANSS) total score at week 1 were calculated. Percent reduction in the PANSS total score at week 1 and a lower PANSS G12 item score (ie, better in judgment and insight) at baseline were significantly associated with placebo response at week 9 (odds ratio [OR] = 1.063; 95% CI, 1.040-1.087, P < .001; and OR= 0.739; 95% CI, 0.553-0.986, P = .040, respectively, in the per-protocol analysis). Cutoffs of a 10% (accuracy = 0.724 in the per-protocol analysis) and 15% (accuracy = 0.722 in the last-observation-carried-forward analysis) reduction in the PANSS total score at week 1 showed the highest predictive power. The appreciation that longer-term response following placebo injections can be predicted by a 10%-15% PANSS total score reduction at week 1 could guide the design of future clinical trials of LAI antipsychotics in schizophrenia to identify and exclude potential placebo responders early during the course of the study.


Dynorphin increases extracellular levels of excitatory amino acids in the brain through a non-opioid mechanism.
Administration of dynorphin A-(1-17) (Dyn 1-17), through a microdialysis probe stereotaxically placed into rat hippocampus, caused marked increases in the extracellular levels of glutamate and aspartate. The degree and duration of elevation of these excitatory amino acids (EAA) induced by Dyn 1-17 were dose dependent but were not modified by the centrally active opioid receptor antagonist nalmefene. At comparable doses, Dyn 2-17, which is inactive at the opioid receptor, produced similar alterations in EAA as Dyn 1-17, whereas Dyn 1-8 caused significantly smaller changes of glutamate. Dynorphin and EAAs have each been implicated as pathophysiological factors in brain or spinal cord injuries, with dynorphin's actions shown to involve both opioid and non-opioid components. The present observations indicate a direct potential linkage between dynorphin and excitotoxin mechanisms of CNS injury and provide further support for the concept that dynorphin's pathophysiologic effects may include non-opioid actions of this peptide.


Proteogenomic basis for ecological divergence of closely related bacteria in natural acidophilic microbial communities.
Bacterial species concepts are controversial. More widely accepted is the need to understand how differences in gene content and sequence lead to ecological divergence. To address this relationship in ecosystem context, we investigated links between genotype and ecology of two genotypic groups of Leptospirillum group II bacteria in comprehensively characterized, natural acidophilic biofilm communities. These groups share 99.7% 16S rRNA gene sequence identity and 95% average amino acid identity between their orthologs. One genotypic group predominates during early colonization, and the other group typically proliferates in later successional stages, forming distinct patches tens to hundreds of micrometers in diameter. Among early colonizing populations, we observed dominance of five genotypes that differed from each other by the extent of recombination with the late colonizing type. Our analyses suggest that the specific recombinant variant within the early colonizing group is selected for by environmental parameters such as temperature, consistent with recombination as a mechanism for ecological fine tuning. Evolutionary signatures, and strain-resolved expression patterns measured via mass spectrometry-based proteomics, indicate increased cobalamin biosynthesis, (de)methylation, and glycine cleavage in the late colonizer. This may suggest environmental changes within the biofilm during development, accompanied by redirection of compatible solutes from osmoprotectants toward metabolism. Across 27 communities, comparative proteogenomic analyses show that differential regulation of shared genes and expression of a small subset of the approximately 15% of genes unique to each genotype are involved in niche partitioning. In summary, the results show how subtle genetic variations can lead to distinct ecological strategies.


Alpha-linolenic acid: an omega-3 fatty acid with neuroprotective properties-ready for use in the stroke clinic?
Alpha-linolenic acid (ALA) is plant-based essential omega-3 polyunsaturated fatty acids that must be obtained through the diet. This could explain in part why the severe deficiency in omega-3 intake pointed by numerous epidemiologic studies may increase the brain's vulnerability representing an important risk factor in the development and/or deterioration of certain cardio- and neuropathologies. The roles of ALA in neurological disorders remain unclear, especially in stroke that is a leading cause of death. We and others have identified ALA as a potential nutraceutical to protect the brain from stroke, characterized by its pleiotropic effects in neuroprotection, vasodilation of brain arteries, and neuroplasticity. This review highlights how chronic administration of ALA protects against rodent models of hypoxic-ischemic injury and exerts an anti-depressant-like activity, effects that likely involve multiple mechanisms in brain, and may be applied in stroke prevention. One major effect may be through an increase in mature brain-derived neurotrophic factor (BDNF), a widely expressed protein in brain that plays critical roles in neuronal maintenance, and learning and memory. Understanding the precise roles of ALA in neurological disorders will provide the underpinnings for the development of new therapies for patients and families who could be devastated by these disorders.


Early Intervention Project: can its claims be substantiated and its effects replicated?
A comprehensive report to the National Institute of Health on the diagnosis, etiology, epidemiology, and treatment of autism indicated that early intervention has the potential of being an effective intervention (Bristol et al., 1996). In spite of this positive outlook, several research and methodological questions remain regarding time of treatment initiation, intensity of treatment and duration of treatment, random assignment, comparative treatment designs, and treatment integrity. Against this backdrop we consider the claims made by the Early Intervention Project (EIP; Lovaas, 1987, 1993; McEachin, Smith, & Lovaas, 1993). The EIP claims to produce recovery from autism in 47% of the cases and to greatly reduce its severity in an additional 42% of cases. This article evaluates the EIP against threats to internal and external validity and is found to suffer from a number of methodological problems. Based on rebuttals to criticisms of their program, the EIP authors seem unwilling to admit any methodological flaws in the sampling, design, and analysis of data of the EIP. It is recommended that parents and fair hearing officers adopt an attitude of healthy skepticism before proceeding to an unqualified endorsement of the EIP as a treatment for autism.


Gene therapy for chronic relapsing experimental allergic encephalomyelitis using cells expressing a novel soluble p75 dimeric TNF receptor.
In a murine relapsing experimental allergic encephalomyelitis (EAE) model, gene therapy to block TNF was investigated with the use of a retroviral dimeric p75 TNF receptor (dTNFR) construct. To effectively produce these TNF inhibitors in vivo, a conditionally immortalized syngeneic fibroblast line was established, using a temperature-sensitive SV40 large T Ag-expressing retrovirus. These cells were subsequently infected with a retrovirus expressing soluble dTNFR. CNS-injected cells could be detected 3 mo after transplantation and were shown to produce the transgene product by immunocytochemistry and ELISA of tissue fluids. These levels of dTNFR protein were biologically active and could significantly ameliorate both acute and relapsing EAE. This cell-based gene-vector approach is ideal for delivering proteins to the CNS and has particular relevance to the control of inflammatory CNS disease.


School closure as an influenza mitigation strategy: how variations in legal authority and plan criteria can alter the impact.
States' pandemic influenza plans and school closure statutes are intended to guide state and local officials, but most faced a great deal of uncertainty during the 2009 influenza H1N1 epidemic. Questions remained about whether, when, and for how long to close schools and about which agencies and officials had legal authority over school closures. This study began with analysis of states' school-closure statutes and pandemic influenza plans to identify the variations among them. An agent-based model of one state was used to represent as constants a population's demographics, commuting patterns, work and school attendance, and community mixing patterns while repeated simulations explored the effects of variations in school closure authority, duration, closure thresholds, and reopening criteria. The results show no basis on which to justify statewide rather than school-specific or community-specific authority for school closures. Nor do these simulations offer evidence to require school closures promptly at the earliest stage of an epidemic. More important are criteria based on monitoring of local case incidence and on authority to sustain closure periods sufficiently to achieve epidemic mitigation. This agent-based simulation suggests several ways to improve statutes and influenza plans. First, school closure should remain available to state and local authorities as an influenza mitigation strategy. Second, influenza plans need not necessarily specify the threshold for school closures but should clearly define provisions for early and ongoing local monitoring. Finally, school closure authority may be exercised at the statewide or local level, so long as decisions are informed by monitoring incidence in local communities and schools.


Amphibian immune defenses against chytridiomycosis: impacts of changing environments.
Eco-immunology is the field of study that attempts to understand the functions of the immune system in the context of the host's environment. Amphibians are currently suffering devastating declines and extinctions in nearly all parts of the world due to the emerging infectious disease chytridiomycosis caused by the chytrid fungus, Batrachochytrium dendrobatidis. Because chytridiomycosis is a skin infection and remains confined to the skin, immune defenses of the skin are critical for survival. Skin defenses include secreted antimicrobial peptides and immunoglobulins as well as antifungal metabolites produced by symbiotic skin bacteria. Low temperatures, toxic chemicals, and stress inhibit the immune system and may impair natural defenses against B. dendrobatidis. Tadpoles' mouth parts can be infected by B. dendrobatidis. Damage to the mouth parts can impair growth, and the affected tadpoles maintain the pathogen in the environment even when adults have dispersed. Newly metamorphosing frogs appear to be especially vulnerable to infection and to the lethal effects of this pathogen because the immune system undergoes a dramatic reorganization at metamorphosis, and postmetamorphic defenses are not yet mature. Here we review our current understanding of amphibian immune defenses against B. dendrobatidis and the ability of the pathogen to resist those defenses. We also briefly review what is known about the impacts of temperature, environmental chemicals, and stress on the host-pathogen interactions and suggest future directions for research.


Parenchymal trafficking of pleural mesothelial cells in idiopathic pulmonary fibrosis.
Idiopathic pulmonary fibrosis (IPF) is characterised by myofibroblast proliferation leading to architectural destruction. Neither the origin nor the continued proliferation of myofibroblasts is well understood. Explanted human IPF lungs were stained by immunohistochemistry for calretinin, a marker of pleural mesothelial cells (PMCs). Chronic obstructive pulmonary disease (COPD) and cystic fibrosis (CF) lungs acted as controls. The number of PMCs per 100 nucleated cells and per photomicrograph was estimated along with the Ashcroft score of fibrosis. Mouse PMCs expressing green fluorescent protein (GFP) or labelled with nanoparticles were injected into the pleural space of mice given intranasal transforming growth factor (TGF)-β1. Mouse lungs were lavaged and examined for the presence of GFP, smooth muscle α-actin (α-SMA) and calretinin. Calretinin-positive PMCs were found throughout IPF lungs, but not in COPD or CF lungs. The number of PMCs correlated with the Ashcroft score. In mice, nanoparticle-laden PMCs were recoverable by bronchoalveolar lavage, depending on the TGF-β1 dose. Fluorescent staining showed α-SMA expression in GFP-expressing PMCs, with co-localisation of GFP and α-SMA. PMCs can traffic through the lung and show myofibroblast phenotypic markers. PMCs are present in IPF lungs, and their number correlates with IPF severity. Since IPF presumably begins subpleurally, PMCs could play a pathogenetic role via mesothelial-mesenchymal transition.


SIRT4 accelerates Ang II-induced pathological cardiac hypertrophy by inhibiting manganese superoxide dismutase activity.
Oxidative stress contributes to the development of cardiac hypertrophy and heart failure. One of the mitochondrial sirtuins, Sirt4, is highly expressed in the heart, but its function remains unknown. The aim of the present study was to investigate the role of Sirt4 in the pathogenesis of pathological cardiac hypertrophy and the molecular mechanism by which Sirt4 regulates mitochondrial oxidative stress. Male C57BL/6 Sirt4 knockout mice, transgenic (Tg) mice exhibiting cardiac-specific overexpression of Sirt4 (Sirt4-Tg) and their respective controls were treated with angiotensin II (Ang II, 1.1 mg/kg/day). At 4 weeks, hypertrophic growth of cardiomyocytes, fibrosis and cardiac function were analysed. Sirt4 deficiency conferred resistance to Ang II infusion by significantly suppressing hypertrophic growth, and the deposition of fibrosis. In Sirt4-Tg mice, aggravated hypertrophy and reduced cardiac function were observed compared with non-Tg mice following Ang II treatment. Mechanistically, Sirt4 inhibited the binding of manganese superoxide dismutase (MnSOD) to Sirt3, another member of the mitochondrial sirtuins, and increased MnSOD acetylation levels to reduce its activity, resulting in elevated reactive oxygen species (ROS) accumulation upon Ang II stimulation. Furthermore, inhibition of ROS with manganese 5, 10, 15, 20-tetrakis-(4-benzoic acid) porphyrin, a mimetic of SOD, blocked the Sirt4-mediated aggravation of the hypertrophic response in Ang II-treated Sirt4-Tg mice. Sirt4 promotes hypertrophic growth, the generation of fibrosis and cardiac dysfunction by increasing ROS levels upon pathological stimulation. These findings reveal a role of Sirt4 in pathological cardiac hypertrophy, providing a new potential therapeutic strategy for this disease.


Induction of cytokine expression by leukemia inhibitory factor.
Biological effects of cytokines are in part determined by their interactions in the regulation of cytokine production. This study analyzes the effects of leukemia inhibitory factor (LIF) on cytokine expression in different cell lineages. Recombinant human LIF increases levels of IL-1 beta, IL-6, and IL-8 mRNA in human articular chondrocytes as demonstrated by Northern blotting. These cytokine mRNAs are detectable as early as 1.3 h after stimulation and reach their maximum after 5 h. The LIF effects are dose dependent and of similar magnitude to those of IL-1. By metabolic labeling and immunoprecipitation it is shown that LIF induces synthesis and secretion of IL-6. IL-6 bioactivity in conditioned media, as measured by the B9 hybridoma proliferation assay, is increased by LIF. Effects of LIF on cytokine expression are not confined to connective tissue cells. By PCR it is shown that human blood monocytes express IL-6 mRNA after stimulation with LIF. An increase in IL-6 mRNA levels is detectable 2 h after stimulation, and this starts to decline by 5 h. The response is of shorter duration as compared with IL-1 beta. In addition to increased mRNA expression, LIF also stimulates release of biologically active IL-6 from blood monocytes. In synoviocytes and neuronal as well as epithelial cell lines, LIF increases IL-1 beta and IL-6 gene expression. In summary, LIF induces cytokine expression in a wide variety of tissues. These results suggest that through the induction of cytokines, LIF can modulate inflammation, immune responses, and connective tissue metabolism, and act as a pathogenetic mediator in different disease states.


Hypoxia inducible factors in cancer stem cells.
Oxygen is an essential regulator of cellular metabolism, survival, and proliferation. Cellular responses to oxygen levels are monitored, in part, by the transcriptional activity of the hypoxia inducible factors (HIFs). Under hypoxia, HIFs regulate a variety of pro-angiogenic and pro-glycolysis pathways. In solid cancers, regions of hypoxia are commonly present throughout the tissue because of the chaotic vascular architecture and regions of necrosis. In these regions, the hypoxic state fluctuates in a spatial and temporal manner. Transient hypoxic cycling causes an increase in the activity of the HIF proteins above what is typical for non-pathologic tissue. The extent of hypoxia strongly correlates to poor patient survival, therapeutic resistance and an aggressive tumour phenotype, but the full contribution of hypoxia and the HIFs to tumour biology is an area of active investigation. Recent reports link resistance to conventional therapies and the metastatic potential to a stem-like tumour population, termed cancer stem cells (CSCs). We and others have shown that within brain tumours CSCs reside in two niches, a perivascular location and the surrounding necrotic tissue. Restricted oxygen conditions increase the CSC fraction and promote acquisition of a stem-like state. Cancer stem cells are critically dependant on the HIFs for survival, self-renewal, and tumour growth. These observations and those from normal stem cell biology provide a new mechanistic explanation for the contribution of hypoxia to malignancy. Further, the presence of hypoxia in tumours may present challenges for therapy because of the promotion of CSC phenotypes even upon successful killing of CSCs. The current experimental evidence suggests that CSCs are plastic cell states governed by microenvironmental conditions, such as hypoxia, that may be critical for the development of new therapies targeted to disrupt the microenvironment.


New Directions for Preventing Dating Violence in Adolescence: The Study of Gender Models.
Dating violence is a huge transcultural and alarming phenomenon, directly linked with endless discrimination against women. The latest research on dating violence in adolescence shows how dating violence is persistent and common in the adolescent period as well and pinpoints the origin of gender violence from first adolescent relationships. This element takes us to considerate how recent gender violence studies and policies, increased also thanks to international efforts on this issue, are not bringing expected results, especially among young people. This mini-review aims to analyze the main characteristics of current gender studies and policies on dating violence, focusing on percentages with a woman-centered approach, which stresses the consequences of gender violence. Other gender studies, that consider gender as a relational product, stress the importance of integrating the analysis of gender models as a key instrument to understand the main causes of dating violence, providing new elements to develop effective policies against dating violence. Indeed, gender models of femininity and masculinity are based on a binary system, which is also a reciprocal recognition and identity system: gender models define female and male characteristics, roles, stereotypes, and expectation, being complementary and foreclosing at the same time. Recent studies on gender relationships, especially among the youth, allows us to propose a new dialog between dating violence studies and gender model studies, underling the need of a complete and complex understanding of gender structure, and of its tensions and contradictions, to put an end to gender and dating violence, through effective programs.


Identification of the minimal replicon and the origin of replication of the crenarchaeal plasmid pRN1.
We have determined the minimal replicon of the crenarchaeal plasmid pRN1. It consists of 3097 base pairs amounting to 58% of the genome of pRN1. The minimal replicon comprises replication operon orf56/orf904 coding for a transcriptional repressor and the replication protein of pRN1. An upstream region of 64 bp that contains the promoter of the replication operon is essential as well as 166 bp of sequence downstream of the orf904 gene. This region contains a putative transcriptional terminator and a 100 nucleotides long stem-loop structure. Only the latter structure was shown to be required for replication. In addition replication was sustained when the stem-loop was displaced to another part of the pRN1 sequence. By mutational analysis we also find that the integrity of the stem-loop structure is required to maintain the replication of pRN1-derived constructs. As similar stem-loop structures are also present in other members of the pRN family, we suggest that this conserved structural element could be the origin of replication for the pRN plasmids. Further bioinformatic analysis revealed that the domain structure of the replication protein and the presence of a similar stem-loop structure as the putative replication origin are also found in several bacteriophages.


Oxymetholone-Induced Acute Renal Failure: A Case Report.
The prevalence of using anabolic steroids such as oxymetholone is increasing. This highlights the need for closely monitoring side effects of this drug. Acute renal failure (ARF) has been reported as a complication of rhabdomyolysis in anabolic steroids users. We present one 33-year-old man complaining of decreased urine volume, urine color change, and lower abdominal pain. He is engaged with a rare side effect of oxymetholone abuse. During assessments of potential medical issues associated with the intake of anabolic steroids, known side effects are known to be transient, but the need for appropriate interventions remains essential. Rhabdomyolysis due to drug use and the consequent acute kidney injury are among the lethal risks associated with anabolic steroid abuse. In most cases, the symptoms are extensive and often misleading. Therefore, detailed history taking, physical scrutiny, paraclinical testing, and early diagnosis are crucial for rhabdomyolysis patients.


The Effects of Hydrogen Gas Inhalation on Adverse Left Ventricular Remodeling After Percutaneous Coronary Intervention for ST-Elevated Myocardial Infarction - First Pilot Study in Humans.
Hydrogen gas inhalation (HI) reduced infarct size and mitigated adverse left ventricular (LV) remodeling in a rat model of acute myocardial infarction (AMI). We designed a prospective, open-label, rater-blinded clinical pilot study in patients experiencing ST-elevated MI (STEMI).Methods and Results:The 20 patients with an initial diagnosis of STEMI were assigned to either an HI group (1.3% H2with 26% oxygen) or a control group (26% oxygen). There were no HI-related severe adverse events. In the full analysis set, the cardiac salvage index as evaluated using cardiac magnetic resonance imaging at 7 days after primary percutaneous coronary intervention (PCI), showed no significant between-group difference (HI: 50.0±24.3%; control: 60.1±20.1%; P=0.43). However, the improvement from day 7 in the HI group was numerically greater than that in the control group in some of the surrogate outcomes at 6-month follow-up, including the LV stroke volume index (HI: 9.2±7.1 mL/m2; control: -1.4±7.2 mL/m2; P=0.03) and the LV ejection fraction (HI: 11.0%±9.3%; control: 1.7%±8.3%; P=0.11). The first clinical study has shown that HI during PCI is feasible and safe and may also promote LV reverse remodeling at 6 months after STEMI. The study was not powered to test efficacy and a further large-scale trial is warranted. (Clinical trials registration: UMIN00006825).


ROAMER: roadmap for mental health research in Europe.
Despite the high impact of mental disorders in society, European mental health research is at a critical situation with a relatively low level of funding, and few advances been achieved during the last decade. The development of coordinated research policies and integrated research networks in mental health is lagging behind other disciplines in Europe, resulting in lower degree of cooperation and scientific impact. To reduce more efficiently the burden of mental disorders in Europe, a concerted new research agenda is necessary. The ROAMER (Roadmap for Mental Health Research in Europe) project, funded under the European Commission's Seventh Framework Programme, aims to develop a comprehensive and integrated mental health research agenda within the perspective of the European Union (EU) Horizon 2020 programme, with a translational goal, covering basic, clinical and public health research. ROAMER covers six major domains: infrastructures and capacity building, biomedicine, psychological research and treatments, social and economic issues, public health and well-being. Within each of them, state-of-the-art and strength, weakness and gap analyses were conducted before building consensus on future research priorities. The process is inclusive and participatory, incorporating a wide diversity of European expert researchers as well as the views of service users, carers, professionals and policy and funding institutions.


The promise of deep brain stimulation in X-linked dystonia parkinsonism.
X-Linked dystonia parkinsonism (XDP) is a rapidly progressive and disabling neurodegenerative disease affecting mainly male Filipinos with origins from Panay Island. We reviewed all the past neurosurgical ablative procedures done for XDP patients listed in the Philippine XDP registry. From 1960 to 1982, six patients had undergone bilateral chemopallidotomies or bilateral thalomotomies staged over time. Half of these patients had significant improvement in their symptoms but five of the six patients (83%) developed postoperative morbidities, mainly speech impairment or hemiparesis. All the five reported GPi deep brain stimulation (DBS) cases for XDP were also reviewed, showing consistently immediate improvement of symptoms (61.5%-88.3% decrease in the Burke-Marsden-Fahn Dystonia Rating Scale) lasting up to a year with no adverse effects noted. We also present the first Philippine case of GPi DBS done in the youngest XDP patient to date. This present case showed dramatic improvement (88.3% decrease of the Burke-Marsden-Fahn Dystonia Rating Scale) of his dystonic symptoms, without incurring any persistent adverse effects. The results of these early cases of pallidal DBS for XDP show that DBS is generally a safe and effective procedure for alleviating the disabling symptoms of XDP in contrast to previous ablative surgeries performed on these patients.


In vitro evaluation of heparinized Cuprophan hemodialysis membranes.
Cuprophan hemodialysis membranes can be heparinized using N,N'-carbonyldiimidazole (CDI) as a coupling agent. In this study, the characteristics of heparinized Cuprophan membranes have been evaluated. After immobilization, heparin partially retained its biologic activity. An anticoagulant activity of 12.4 +/- 4.2 mU/cm2 was measured using a thrombin inactivation assay. Immobilized heparin also displayed an anti-complement activity. After contact with human serum; heparinized Cuprophan induced no generation of significant amounts of fluid phase terminal complement complex (TCC), whereas untreated Cuprophan induced the generation of substantial amounts of TCC. Heparinization did not affect the permeability of Cuprophan for model solutes with molecular weights up to 12,000 g/mol except for sulfobromophthalein sodium salt. The permeability of Cuprophan for sulfobromophthalein sodium salt was slightly decreased after heparinization. The ultrafiltration rate of Cuprophan increased by about 30% after heparinization, probably owing to an increased swelling of the membrane in water. Heparinized Cuprophan incubated in phosphate-buffered saline at 37 degrees C showed some release of heparin. These amounts of released heparin, however, were very low as compared to the amounts of heparin which are systemically administered during clinical hemodialysis treatment. It is concluded that Cuprophan membranes heparinized by means of the CDI-activation procedure are highly promising for application in hemodialyzers to be used for the treatment of patients with reduced or without systemic administration of heparin.


Process cost and facility considerations in the selection of primary cell culture clarification technology.
The bioreactor volume delineating the selection of primary clarification technology is not always easily defined. Development of a commercial scale process for the manufacture of therapeutic proteins requires scale-up from a few liters to thousands of liters. While the separation techniques used for protein purification are largely conserved across scales, the separation techniques for primary cell culture clarification vary with scale. Process models were developed to compare monoclonal antibody production costs using two cell culture clarification technologies. One process model was created for cell culture clarification by disc stack centrifugation with depth filtration. A second process model was created for clarification by multi-stage depth filtration. Analyses were performed to examine the influence of bioreactor volume, product titer, depth filter capacity, and facility utilization on overall operating costs. At bioreactor volumes <1,000 L, clarification using multi-stage depth filtration offers cost savings compared to clarification using centrifugation. For bioreactor volumes >5,000 L, clarification using centrifugation followed by depth filtration offers significant cost savings. For bioreactor volumes of ∼ 2,000 L, clarification costs are similar between depth filtration and centrifugation. At this scale, factors including facility utilization, available capital, ease of process development, implementation timelines, and process performance characterization play an important role in clarification technology selection. In the case study presented, a multi-product facility selected multi-stage depth filtration for cell culture clarification at the 500 and 2,000 L scales of operation. Facility implementation timelines, process development activities, equipment commissioning and validation, scale-up effects, and process robustness are examined.


Bundle branch reentrant ventricular tachycardia.
Bundle branch reentrant (BBR) tachycardia is an uncommon form of ventricular tachycardia (VT) incorporating both bundle branches into the reentry circuit. The arrhythmia is usually seen in patients with an acquired heart disease and significant conduction system impairment, although patients with structurally normal heart have been described. Surface ECG in sinus rhythm (SR) characteristically shows intraventricular conduction defects. Patients typically present with presyncope, syncope or sudden death because of VT with fast rates frequently above 200 beats per minute. The QRS morphology during VT is a typical bundle branch block pattern, usually left bundle branch block, and may be identical to that in SR. Prolonged His-ventricular (H-V) interval in SR is found in the majority of patients with BBR VT, although some patients may have the H-V interval within normal limits. The diagnosis of BBR VT is based on electrophysiological findings and pacing maneuvers that prove participation of the His- Purkinje system in the tachycardia mechanism. Radiofrequency catheter ablation of a bundle branch can cure BBR VT and is currently regarded as the first line therapy. The technique of choice is ablation of the right bundle. The reported incidence of clinically significant conduction system impairment requiring implantation of a permanent pacemaker varies from 0% to 30%. Long-term outcome depends on the underlying cardiac disease. Patients with poor systolic left ventricular function are at risk of sudden death or death from progressive heart failure despite successful BBR VT ablation and should be considered for an implantable cardiovertor-defibrillator.


Liver injury from herbals and dietary supplements in the U.S. Drug-Induced Liver Injury Network.
The Drug-Induced Liver Injury Network (DILIN) studies hepatotoxicity caused by conventional medications as well as herbals and dietary supplements (HDS). To characterize hepatotoxicity and its outcomes from HDS versus medications, patients with hepatotoxicity attributed to medications or HDS were enrolled prospectively between 2004 and 2013. The study took place among eight U.S. referral centers that are part of the DILIN. Consecutive patients with liver injury referred to a DILIN center were eligible. The final sample comprised 130 (15.5%) of all subjects enrolled (839) who were judged to have experienced liver injury caused by HDS. Hepatotoxicity caused by HDS was evaluated by expert opinion. Demographic and clinical characteristics and outcome assessments, including death and liver transplantation (LT), were ascertained. Cases were stratified and compared according to the type of agent implicated in liver injury; 45 had injury caused by bodybuilding HDS, 85 by nonbodybuilding HDS, and 709 by medications. Liver injury caused by HDS increased from 7% to 20% (P < 0.001) during the study period. Bodybuilding HDS caused prolonged jaundice (median, 91 days) in young men, but did not result in any fatalities or LT. The remaining HDS cases presented as hepatocellular injury, predominantly in middle-aged women, and, more frequently, led to death or transplantation, compared to injury from medications (13% vs. 3%; P < 0.05). The proportion of liver injury cases attributed to HDS in DILIN has increased significantly. Liver injury from nonbodybuilding HDS is more severe than from bodybuilding HDS or medications, as evidenced by differences in unfavorable outcomes (death and transplantation). (Hepatology 2014;60:1399-1408).