Effectiveness of Trauma-Focused Cognitive Behavioral Therapy in a community-based program.
Trauma-Focused Cognitive Behavioral Therapy (TF-CBT) is a widely used treatment model for trauma-exposed children and adolescents (Cohen, Mannarino, & Deblinger, 2006). The Healthy Coping Program (HCP) was a multi-site community based intervention carried out in a diverse Canadian city. A randomized, waitlist-control design was used to evaluate the effectiveness of TF-CBT with trauma-exposed school-aged children (Muller & DiPaolo, 2008). A total of 113 children referred for clinical services and their caregivers completed the Trauma Symptom Checklist for Children (Briere, 1996) and the Trauma Symptom Checklist for Young Children (Briere, 2005). Data were collected pre-waitlist, pre-assessment, pre-therapy, post-therapy, and six months after the completion of TF-CBT. The passage of time alone in the absence of clinical services was ineffective in reducing children's posttraumatic symptoms. In contrast, children and caregivers reported significant reductions in children's posttraumatic stress (PTS) following assessment and treatment. The reduction in PTS was maintained at six month follow-up. Findings of the current study support the use of the TF-CBT model in community-based settings in a diverse metropolis. Clinical implications are discussed.


Organization and expression of hepatitis B sequences cloned from hepatocellular carcinoma tissue DNA.
We have constructed a phage lambda library of liver DNA fragments from West African patient who died of liver failure due to advanced hepatocellular carcinoma. Four hepatitis B virus (HBV) DNA-carrying recombinants have been isolated, one clone (lambda IA22) being analyzed in greatest detail. It contains approximately 3.8 kb of HBV DNA without detectable deletions or rearrangements. One site of integration lies close to the nick in free viral DNA. The restriction map of the HBV sequences is close to those published for the ay subtype. Coconvection of mouse Ltk- cells with lambda IA22 and cloned thymidine kinase gene results in the expression of gene S and the excretion of hepatitis B surface antigen (HBsAg) particles into the culture supernatant.


Successful Stepwise Development of Patient Research Partnership: 14 Years' Experience of Actions and Consequences in Outcome Measures in Rheumatology (OMERACT).
There is increasing interest in making patient participation an integral component of medical research. However, practical guidance on optimizing this engagement in healthcare is scarce. Since 2002, patient involvement has been one of the key features of the Outcome Measures in Rheumatology (OMERACT) international consensus effort. Based on a review of cumulative data from qualitative studies and internal surveys among OMERACT participants, we explored the potential benefits and challenges of involving patient research partners in conferences and working group activities. We supplemented our review with personal experiences and reflections regarding patient participation in the OMERACT process. We found that between 2002 and 2016, 67 patients have attended OMERACT conferences, of whom 28 had sustained involvement; many other patients contributed to OMERACT working groups. Their participation provided face validity to the OMERACT process and expanded the research agenda. Essential facilitators have been the financial commitment to guarantee sustainable involvement of patients at these conferences, procedures for recruitment, selection and support, and dedicated time allocated in the program for patient issues. Current challenges include the representativeness of the patient panel, risk of pseudo-professionalization, and disparity in patients' and researchers' perception of involvement. In conclusion, OMERACT has embedded long-term patient involvement in the consensus-building process on the measurement of core health outcomes. This integrative process continues to evolve iteratively. We believe that the practical points raised here can improve participatory research implementation.


Local delivery of novel MRTF/SRF inhibitors prevents scar tissue formation in a preclinical model of fibrosis.
The myocardin-related transcription factor/serum response factor (MRTF/SRF) pathway represents a promising therapeutic target to prevent fibrosis. We have tested the effects of new pharmacological inhibitors of MRTF/SRF signalling in a preclinical model of fibrosis. CCG-222740, a novel MRTF/SRF inhibitor, markedly decreased SRF reporter gene activity and showed a greater inhibitory effect on MRTF/SRF target genes than the previously described MRTF-A inhibitor CCG-203971. CCG-222740 was also five times more potent, with an IC50 of 5 μM, in a fibroblast-mediated collagen contraction assay, was less cytotoxic, and a more potent inhibitor of alpha-smooth muscle actin protein expression than CCG-203971. Local delivery of CCG-222740 and CCG-203971 in a validated and clinically relevant rabbit model of scar tissue formation after glaucoma filtration surgery increased the long-term success of the surgery by 67% (P < 0.0005) and 33% (P < 0.01), respectively, and significantly decreased fibrosis and scarring histologically. Unlike mitomycin-C, neither CCG-222740 nor CCG-203971 caused any detectable epithelial toxicity or systemic side effects with very low drug levels measured in the aqueous, vitreous, and serum. We conclude that inhibitors of MRTF/SRF-regulated gene transcription such as CCG-222740, potentially represent a new therapeutic strategy to prevent scar tissue formation in the eye and other tissues.


A novel anaerobic electrochemical membrane bioreactor (AnEMBR) with conductive hollow-fiber membrane for treatment of low-organic strength solutions.
A new anaerobic treatment system that combined a microbial electrolysis cell (MEC) with membrane filtration using electrically conductive, porous, nickel-based hollow-fiber membranes (Ni-HFMs) was developed to treat low organic strength solution and recover energy in the form of biogas. This new system is called an anaerobic electrochemical membrane bioreactor (AnEMBR). The Ni-HFM served the dual function as the cathode for hydrogen evolution reaction (HER) and the membrane for filtration of the effluent. The AnEMBR system was operated for 70 days with synthetic acetate solution having a chemical oxygen demand (COD) of 320 mg/L. Removal of COD was >95% at all applied voltages tested. Up to 71% of the substrate energy was recovered at an applied voltage of 0.7 V as methane rich biogas (83% CH4; <1% H2) due to biological conversion of the hydrogen evolved at the cathode to methane. A combination of factors (hydrogen bubble formation, low cathode potential and localized high pH at the cathode surface) contributed to reduced membrane fouling in the AnEMBR compared to the control reactor (open circuit voltage). The net energy required to operate the AnEMBR system at an applied voltage of 0.7 V was significantly less (0.27 kWh/m3) than that typically needed for wastewater treatment using aerobic membrane bioreactors (1-2 kWh/m3).


In-situ atomic layer deposition of tri-methylaluminum and water on pristine single-crystal (In)GaAs surfaces: electronic and electric structures.
The electronic structure of single-crystal (In)GaAs deposited with tri-methylaluminum (TMA) and water via atomic layer deposition (ALD) is presented with high-resolution synchrotron radiation core-level photoemission and capacitance-voltage (CV) characteristics. The interaction of the precursor atoms with (In)GaAs is confined at the topmost surface layer. The Ga-vacant site on the GaAs(111)A-2 × 2 surface is filled with Al, thereby effectively passivating the As dangling bonds. The As-As dimers on the GaAs(001)-2 × 4 surface are entirely passivated by one cycle of TMA and water. The presumed layerwise deposition fails to happen in GaAs(001)-4 × 6. In In0.20Ga0.80As(001)-2 × 4, the edge row As atoms are partially bonded with the Al, and one released methyl then bonds with the In. It is suggested that the unpassivated surface and subsurface atoms cause large frequency dispersions in CV characteristics under the gate bias. We also found that the (In)GaAs surface is immune to water in ALD. However, the momentary exposure of it to air (less than one minute) introduces significant signals of native oxides. This indicates the necessity of in situ works of high κ/(In)GaAs-related experiments in order to know the precise interfacial atomic bonding and thus know the electronic characteristics. The electric CV measurements of the ALD-Al2O3 on these (In)GaAs surfaces are correlated with their electronic properties.


Heart Failure in Relation to Tumor-Targeted Therapies and Immunotherapies.
Tumor-targeted therapies such as trastuzumab have led to significant improvements in survival of human epidermal growth factor receptor 2 (HER2)-positive breast cancer. However, these therapies have also been associated with significant left ventricular dysfunction. The incidence of trastuzumab-induced heart failure has decreased significantly since the initial reports, in large part due to improved screening, closer monitoring for early changes in left ventricular function, and a significant decrease in the concurrent administration of anthracyclines. The mechanism of trastuzumab cardiotoxicity is still not well understood, but current knowledge suggests that ErbB2 inhibition in cardiac myocytes plays a key role. In addition to trastuzumab and other HER2-targeted agents, vascular endothelial growth factor inhibitors, proteasome inhibitors, and immune checkpoint inhibitors are all additional classes of drugs used with great success in the treatment of solid tumors and hematologic malignancies. Yet these, too, have been associated with cardiac toxicity that ranges from a mild asymptomatic decrease in ejection fraction to fulminant myocarditis. In this review, we summarize the cardiotoxic effects of tumor-targeted and immunotherapies with a focus on HER2 antagonists.


Influence of exopolysaccharides on the electrophoretic properties of the model cyanobacterium Synechocystis.
The influence of extracellular polymeric substances (EPS) on cell electrokinetics was investigated in the model cyanobacterium Synechocystis, in wild-type strains and in ten EPS-depleted mutants. The charge density and the softness of the EPS polyelectrolyte layer were calculated from the dependence of the electrophoretic mobility values of the cells with the ionic strength of the surrounding fluid. Electrophoretic mobility data showed that the eleven Synechocystis strains investigated behave as soft particles and cannot be adequately described by classical electrokinetic models of rigid particles. EPS surrounding the cells, especially those released in the growth medium, significantly increased the softness of the cell surface. Furthermore, the anionic nature of EPS resulted in negative surface charge densities, which appeared to be strongly dependent on the composition of the suspending fluid, as documented by a strong reduction of their absolute values in the presence of calcium cations. These finding stresses the importance of the physicochemical properties of EPS and cell surfaces of cyanobacteria, for both cell-to-medium and cell-to-cell communications. In turn, these results emphasize that, whenever possible, natural waters should be used for meaningful ecotoxicological analyses of potential toxics.


Prospective evaluation of gastric neurostimulation for diabetic gastroparesis in Canada.
The efficacy of gastric neurostimulation therapy for diabetic gastroparesis (GP) in a 'real-life' Canadian setting has not been assessed. To assess changes in health-related quality of life (QoL), weekly vomiting frequency (WVF), total symptom score (TSS) and health care utilization 12 months before and after gastric neurostimulator implantation in a diabetic GP cohort. Medication-refractory diabetic GP patients (n=7, four female, mean age 42 years) were prospectively recruited from 2008 to 2012. QoL scores were self-administered and obtained at baseline, 24 and 48 weeks postimplantion. WVF and TSS were assessed similarly. Health care usage, measured as hospitalization frequency and medication cost, was obtained six and 12 months before and after implant. Changes from baseline to six and 12 months for all outcomes were compared. The mean ( ± SD) QoL according to EuroQol was significantly better at 24 weeks after the baseline measurement (baseline 29 ± 5, 24 weeks 52 ± 7; P = 0.03). The mean improvement in TSS was significantly better at one year postintervention (baseline score 35 ± 5 versus 12 months 27 ± 3; P = 0.03). Changes in Short-Form 36 Health Survey and WVF were not significant. Days of GP-related hospitalization were highly variable but decreased from a median of 71 days (range 0 to 227 days) to 29 days (range two to 334 days) one year before and after surgery, respectively (P = 0.735). Outpatient medication costs did not decrease to a significant extent. Gastric neurostimulation for diabetic GP appeared to show some beneficial palliative effects overall in the present small open-label series, but the effect is highly variable among patients, and placebo effect cannot be ruled out.


Patient-Reported Health-Related Quality of Life Is a Predictor of Outcomes in Ambulatory Heart Failure Patients Treated With Left Ventricular Assist Device Compared With Medical Management: Results From the ROADMAP Study (Risk Assessment and Comparative Effectiveness of Left Ventricular Assist Device and Medical Management).
The prospective observational ROADMAP study (Risk Assessment and Comparative Effectiveness of Left Ventricular Assist Device and Medical Management) demonstrated that ambulatory advanced heart failure patients selected for left ventricular assist device (LVAD) were more likely to be alive at 1 year on original therapy with ≥75-m improvement in 6-minute walk distance compared with patients assigned to optimal medical management. Whether baseline health-related quality of life (hrQoL) resulted in a heterogeneity of this treatment benefit is unknown. Patient-reported hrQoL was assessed with EuroQol questionnaire and visual analogue scale (VAS). We aimed to identify predictors of event-free survival and survival with acceptable hrQoL (VAS≥60). LVAD patients had significant improvement in 3 of 5 EuroQol dimensions (P<0.05), but no significant changes were observed with optimal medical management. Among patients with baseline VAS<55, survival on original treatment was lower for optimal medical management patients compared with those assigned to LVAD (58±7% versus 82±5%; P=0.004). No such difference was seen if baseline VAS was ≥55 (70±7% versus 75±9%; P=0.79). Survival on original therapy with acceptable quality of life was also more likely with LVAD versus optimal medical management if baseline VAS was <55, whereas outcomes in patients with higher baseline VAS scores were similar regardless of treatment assignment (P=0.046 for treatment arm and baseline VAS interaction). LVAD therapy resulted in improvement of patient health status in heart failure patients with low self-reported hrQoL, but not in patients with acceptable quality of life at the time of LVAD implantation. Patient-reported hrQoL should be integrated into decision making concerning the use and timing of LVAD therapy in heart failure patients who are symptom limited but remain ambulatory. URL: http://www.ClinicalTrials.gov. Unique identifier: NCT01452802.


Efficacy of Cancer Immunotherapy: An Umbrella Review of Meta-analyses of Randomized Controlled Trials.
We conducted a systematic review for evidence of the clinical efficacy of cancer immunotherapies. We searched PubMed from inception to 14 February 2018 for meta-analyses of randomized controlled trials (RCTs) of cancer immunotherapies. Re-analyses were performed to estimate the summary effect size under random-effects, the 95% confidence interval (CI), heterogeneity, and the 95% prediction interval, and we determined the strength of the evidence. We examined publication bias and excess significance bias. 63 articles corresponding to 247 meta-analyses were eligible. Nine meta-analyses were classified to have convincing evidence, and 75 were classified as suggestive evidence. The clinical benefit of immunotherapy was supported by convincing evidence in the following settings: anti-PD-1/PD-L1 monoclonal antibody (mAb) therapy for treating advanced melanoma and non-small cell lung cancer (NSCLC), the combination of rituximab and chemotherapy for treating chronic lymphocytic leukemia and B-cell non-Hodgkin's lymphoma, adoptive cell immunotherapy for NSCLC, and the combination of interferon α and chemotherapy for metastatic melanoma. A further meta-analysis of 16 RCTs showed that anti-PD-1/PD-L1 mAb therapy had a benefit in patients with solid tumors (overall survival; hazard ratio = 0.73, 95% CI: 0.68-0.79; p < 0.001), supported by convincing evidence. In the future, rigorous approaches are needed when interpreting meta-analyses to gain better insight into the true efficacy of cancer immunotherapy.


Phosphomolybdic acid as an efficient hole injection material in perovskite optoelectronic devices.
Efficient perovskite devices consist of a perovskite film sandwiched between charge selective layers, in order to avoid non-radiative recombination. A common metal oxide used as a p-type or hole transport layer is molybdenum oxide. MoO3 is of particular interest for its very large work function, which allows it to be used both as an interfacial charge transfer material and a dopant for organic semiconductors. However, high quality and high work function MoO3 is typically thermally evaporated in a vacuum. An alternative solution-processable high work function material is phosphomolybdic acid (PMA), which is stable, commercially available and environmentally friendly. In this Communication, we show the first application of PMA in efficient vacuum processed perovskite devices. We found that the direct growth of perovskite films onto PMA leads to strong charge carrier recombination, hindering the solar cell photovoltage. Using an energetically suitable selective transport layer placed between PMA and the perovskite film, solar cells with efficiency >13% as well as LEDs with promising quantum efficiency can be obtained.


Cellular electron cryo tomography and in situ sub-volume averaging reveal the context of microtubule-based processes.
Electron cryo-tomography (cryoET) is currently the only technique that allows the direct observation of proteins in their native cellular environment. Sub-volume averaging of electron tomograms offers a route to increase the signal-to-noise of repetitive biological structures, such improving the information content and interpretability of tomograms. We discuss the potential for sub-volume averaging in highlighting and investigating specific processes in situ, focusing on microtubule structure and viral infection. We show that (i) in situ sub-volume averaging from single tomograms can guide and complement segmentation of biological features, (ii) the in situ determination of the structure of individual viruses is possible as they infect a cell, and (iii) novel, transient processes can be imaged with high levels of detail.


Deletion of the hypoxia-response element in the vascular endothelial growth factor promoter causes motor neuron degeneration.
Hypoxia stimulates angiogenesis through the binding of hypoxia-inducible factors to the hypoxia-response element in the vascular endothelial growth factor (Vegf) promotor. Here, we report that deletion of the hypoxia-response element in the Vegf promotor reduced hypoxic Vegf expression in the spinal cord and caused adult-onset progressive motor neuron degeneration, reminiscent of amyotrophic lateral sclerosis. The neurodegeneration seemed to be due to reduced neural vascular perfusion. In addition, Vegf165 promoted survival of motor neurons during hypoxia through binding to Vegf receptor 2 and neuropilin 1. Acute ischemia is known to cause nonselective neuronal death. Our results indicate that chronic vascular insufficiency and, possibly, insufficient Vegf-dependent neuroprotection lead to the select degeneration of motor neurons.


Home visiting programs for HIV-affected families: a comparison of service quality between volunteer-driven and paraprofessional models.
Home visiting is a popular component of programs for HIV-affected children in sub-Saharan Africa, but its implementation varies widely. While some home visitors are lay volunteers, other programs invest in more highly trained paraprofessional staff. This paper describes a study investigating whether additional investment in paraprofessional staffing translated into higher quality service delivery in one program context. Beneficiary children and caregivers at sites in KwaZulu-Natal, South Africa were interviewed after 2 years of program enrollment and asked to report about their experiences with home visiting. Analysis focused on intervention exposure, including visit intensity, duration and the kinds of emotional, informational and tangible support provided. Few beneficiaries reported receiving home visits in program models primarily driven by lay volunteers; when visits did occur, they were shorter and more infrequent. Paraprofessional-driven programs not only provided significantly more home visits, but also provided greater interaction with the child, communication on a larger variety of topics, and more tangible support to caregivers. These results suggest that programs that invest in compensation and extensive training for home visitors are better able to serve and retain beneficiaries, and they support a move toward establishing a professional workforce of home visitors to support vulnerable children and families in South Africa.


Soy food and isoflavone intake in relation to semen quality parameters among men from an infertility clinic.
High isoflavone intake has been related to decreased fertility in animal studies, but data in humans are scarce. Thus, we examined the association of soy foods and isoflavones intake with semen quality parameters. The intake of 15 soy-based foods in the previous 3 months was assessed for 99 male partners of subfertile couples who presented for semen analyses to the Massachusetts General Hospital Fertility Center. Linear and quantile regression were used to determine the association of soy foods and isoflavones intake with semen quality parameters while adjusting for personal characteristics. There was an inverse association between soy food intake and sperm concentration that remained significant after accounting for age, abstinence time, body mass index, caffeine and alcohol intake and smoking. In the multivariate-adjusted analyses, men in the highest category of soy food intake had 41 million sperm/ml less than men who did not consume soy foods (95% confidence interval = -74, -8; P, trend = 0.02). Results for individual soy isoflavones were similar to the results for soy foods and were strongest for glycitein, but did not reach statistical significance. The inverse relation between soy food intake and sperm concentration was more pronounced in the high end of the distribution (90th and 75th percentile) and among overweight or obese men. Soy food and soy isoflavone intake were unrelated to sperm motility, sperm morphology or ejaculate volume. These data suggest that higher intake of soy foods and soy isoflavones is associated with lower sperm concentration.


Scaffold functions of 14-3-3 adaptors in B cell immunoglobulin class switch DNA recombination.
Class switch DNA recombination (CSR) of the immunoglobulin heavy chain (IgH) locus crucially diversifies antibody biological effector functions. CSR involves the induction of activation-induced cytidine deaminase (AID) expression and AID targeting to switch (S) regions by 14-3-3 adaptors. 14-3-3 adaptors specifically bind to 5'-AGCT-3' repeats, which make up for the core of all IgH locus S regions. They selectively target the upstream and downstream S regions that are set to undergo S-S DNA recombination. We hypothesized that 14-3-3 adaptors function as scaffolds to stabilize CSR enzymatic elements on S regions. Here we demonstrate that all seven 14-3-3β, 14-3-3ε, 14-3-3γ, 14-3-3η, 14-3-3σ, 14-3-3τ and 14-3-3ζ adaptors directly interacted with AID, PKA-Cα (catalytic subunit) and PKA-RIα (regulatory inhibitory subunit) and uracil DNA glycosylase (Ung). 14-3-3 adaptors, however, did not interact with AID C-terminal truncation mutant AIDΔ(180-198) or AIDF193A and AIDL196A point-mutants (which have been shown not to bind to S region DNA and fail to mediate CSR). 14-3-3 adaptors colocalized with AID and replication protein A (RPA) in B cells undergoing CSR. 14-3-3 and AID binding to S region DNA was disrupted by viral protein R (Vpr), an accessory protein of human immunodeficiency virus type-1 (HIV-1), which inhibited CSR without altering AID expression or germline IH-CH transcription. Accordingly, we demonstrated that 14-3-3 directly interact with Vpr, which in turn, also interact with AID, PKA-Cα and Ung. Altogether, our findings suggest that 14-3-3 adaptors play important scaffold functions and nucleate the assembly of multiple CSR factors on S regions. They also show that such assembly can be disrupted by a viral protein, thereby allowing us to hypothesize that small molecule compounds that specifically block 14-3-3 interactions with AID, PKA and/or Ung can be used to inhibit unwanted CSR.


Short report: in vivo sensitivity of Plasmodium falciparum to halofantrine in southern central Vietnam.
Drug-resistant Plasmodium falciparum is present in Vietnam. We assessed the in vivo sensitivity of P. falciparum to halofantrine in two villages in the southern part of central Vietnam. Halofantrine (8 mg/kg x 3 doses) was administered to 37 patients with either P. falciparum (n = 32) or mixed P. falciparum/P. vivax malaria (n = 5). End points were parasite sensitivity or resistance (RI/RII/RIII) determined by parasite clearance, persistence, or recurrence during 28 days of follow-up. By day 28, 31 (93.9%) of 33 (95% confidence interval = 79.8-99.2%) patients were sensitive. Two patients had recurrent P. falciparum parasitemia on days 14 and 21. Halofantrine effectively treated uncomplicated P. falciparum malaria in these Vietnamese patients.


Galactolipid deficiency and abnormal chloroplast development in the Arabidopsis MGD synthase 1 mutant.
The lipid monogalactosyl diacylglycerol (MGD) is a major structural component of photosynthetic membranes in chloroplasts. Its formation is catalyzed by the enzyme MGD synthase. In many plants, MGD derives from two different biosynthetic pathways: the prokaryotic pathway, which operates entirely within the plastid, and the eukaryotic pathway, which involves steps in the endoplasmic reticulum. Here, we describe the identification and characterization of an Arabidopsis mutant with a defective MGD synthase gene (MGD1). The mutant was identified in a screen of T-DNA lines for individuals with defects in chloroplast biogenesis. It has a yellow-green phenotype that correlates with a approximately 50% deficiency in total chlorophyll per plant. A single T-DNA insertion is located adjacent to the transcription initiation site of the MGD1 gene, and the abundance of MGD1 mRNA is reduced by 75% compared with wild type. Correlation between steady-state MGD1 transcript levels and MGD synthase activity (also reduced by 75% in mgd1) suggests that MGD1 is the most important MGD synthase in green tissues. The amount of MGD in mutant leaves is reduced by 42% compared with wild type. MGD from the mutant contains 23% less 16:3 fatty acid and 10% more 18:3 fatty acid. Because 16:3 is a characteristic feature of MGD from the prokaryotic pathway, it is possible that MGD1 operates with some preference in the prokaryotic pathway. Finally, the MGD-deficiency of mgd1 is correlated with striking defects in chloroplast ultrastructure, strongly suggesting a unique role for MGD in the structural organization of plastidic membranes.


Multilineage stem cells in the adult: a perivascular legacy?
Mesenchymal stem cells proliferate extensively in cultures of unselected, total cell isolates from multiple fetal and adult organs. Perivascular cells, principally pericytes surrounding capillaries and microvessels, but also adventitial cells located around larger arteries and veins, have been recently identified as possible originators of mesenchymal stem cells, first by phenotypic analogies and eventually following stringent cell sorting. While it is clear that purified perivascular cells exhibit multiple mesodermal developmental potentials and become indistinguishable from conventionally derived mesenchymal stem cells after in vitro culture, the possible roles played by these blood vessel-bound cells in organogenesis and adult tissue repair remain elusive. Unsolved questions regarding the identity of mesenchymal stem cells have not compromised the consideration of these cells as outstanding candidates for cell therapies. Better knowledge of the lineage affiliation, tissue distribution and molecular identity of mesenchymal stem cells will contribute to the development of more efficient, safer therapeutic cells.


A critical review and development of a conceptual model of exclusion from social relations for older people.
Social exclusion is complex and dynamic, and it leads to the non-realization of social, economic, political or cultural rights or participation within a society. This critical review takes stock of the literature on exclusion of social relations. Social relations are defined as comprising social resources, social connections and social networks. An evidence review group undertook a critical review which integrates, interprets and synthesizes information across studies to develop a conceptual model of exclusion from social relations. The resulting model is a subjective interpretation of the literature and is intended to be the starting point for further evaluations. The conceptual model identifies individual risks for exclusion from social relations (personal attributes, biological and neurological risk, retirement, socio-economic status, exclusion from material resources and migration). It incorporates the evaluation of social relations, and the influence of psychosocial resources and socio-emotional processes, sociocultural, social-structural, environmental and policy contextual influences on exclusion from social relations. It includes distal outcomes of exclusion from social relations, that is, individual well-being, health and functioning, social opportunities and social cohesion. The dynamic relationships between elements of the model are also reported. We conclude that the model provides a subjective interpretation of the data and an excellent starting point for further phases of conceptual development and systematic evaluation(s). Future research needs to consider the use of sophisticated analytical tools and an interdisciplinary approach in order to understand the underlying biological and ecopsychosocial associations that contribute to individual and dynamic differences in the experience of exclusion from social relations.


International Veterinary Epilepsy Task Force consensus proposal: medical treatment of canine epilepsy in Europe.
In Europe, the number of antiepileptic drugs (AEDs) licensed for dogs has grown considerably over the last years. Nevertheless, the same questions remain, which include, 1) when to start treatment, 2) which drug is best used initially, 3) which adjunctive AED can be advised if treatment with the initial drug is unsatisfactory, and 4) when treatment changes should be considered. In this consensus proposal, an overview is given on the aim of AED treatment, when to start long-term treatment in canine epilepsy and which veterinary AEDs are currently in use for dogs. The consensus proposal for drug treatment protocols, 1) is based on current published evidence-based literature, 2) considers the current legal framework of the cascade regulation for the prescription of veterinary drugs in Europe, and 3) reflects the authors' experience. With this paper it is aimed to provide a consensus for the management of canine idiopathic epilepsy. Furthermore, for the management of structural epilepsy AEDs are inevitable in addition to treating the underlying cause, if possible.


YAP1 mediates gastric adenocarcinoma peritoneal metastases that are attenuated by YAP1 inhibition.
Peritoneal carcinomatosis (PC; malignant ascites or implants) occurs in approximately 45% of advanced gastric adenocarcinoma (GAC) patients and associated with a poor survival. The molecular events leading to PC are unknown. The yes-associated protein 1 (YAP1) oncogene has emerged in many tumour types, but its clinical significance in PC is unclear. Here, we investigated the role of YAP1 in PC and its potential as a therapeutic target. Patient-derived PC cells, patient-derived xenograft (PDX) and patient-derived orthotopic (PDO) models were used to study the function of YAP1 in vitro and in vivo. Immunofluorescence and immunohistochemical staining, RNA sequencing (RNA-Seq) and single-cell RNA-Seq (sc-RNA-Seq) were used to elucidate the expression of YAP1 and PC cell heterogeneity. LentiCRISPR/Cas9 knockout of YAP1 and a YAP1 inhibitor were used to dissect its role in PC metastases. YAP1 was highly upregulated in PC tumour cells, conferred cancer stem cell (CSC) properties and appeared to be a metastatic driver. Dual staining of YAP1/EpCAM and sc-RNA-Seq revealed that PC tumour cells were highly heterogeneous, YAP1high PC cells had CSC-like properties and easily formed PDX/PDO tumours but also formed PC in mice, while genetic knockout YAP1 significantly slowed tumour growth and eliminated PC in PDO model. Additionally, pharmacologic inhibition of YAP1 specifically reduced CSC-like properties and suppressed tumour growth in YAP1high PC cells especially in combination with cytotoxics in vivo PDX model. YAP1 is essential for PC that is attenuated by YAP1 inhibition. Our data provide a strong rationale to target YAP1 in clinic for GAC patients with PC.


On the in situ impedance measurement with pu-probes--simulation of the measurement setup.
High-quality numerical simulations in room acoustics require a detailed knowledge of the acoustic reflection characteristics of the materials in the room, in order to realistically model the interferences between multiple sound reflections at the room boundaries. While different standardized measurement methods exist for the determination of the absorption coefficient and reflection factor these methods can generally not be applied in situ. Thus time-consuming laboratory measurements and the supply of material samples are required. Driven by the obvious demand for a reliable in situ measurement technique, a pu-probe based method has emerged during the last years, which derives the reflection factor based on the simultaneous measurement of sound pressure and velocity. However, previous investigations of the setup and publications by other authors have shown that the measurement results are affected by various uncertainty factors. The present study aims at the identification, separation, and quantitative assessment of the uncertainty factors related to reflection and diffraction effects at the loudspeaker, sensor, and the absorber geometry. Therefore, a purely simulative approach will be used that replicates the actual measurement situation in every detail, including the geometries of sensor, loudspeaker, and absorber. The simulation setup is validated by measurements and is used to systematically separate the different uncertainty factors.


Kinky thresholds revisited: opportunity costs differ in the NE and SW quadrants.
Historically, a kinked threshold line on the cost-effectiveness plane at the origin was suggested due to differences in willingness to pay (WTP) for health gain with trade-offs in the north-east (NE) quadrant versus willingness to accept (WTA) cost reductions for health loss with trade-offs in the south-west (SW) quadrant. Empirically, WTA is greater than WTP for equivalent units of health, a finding supported by loss aversion under prospect theory. More recently, appropriate threshold values for health effects have been shown to require an endogenous consideration of the opportunity cost of alternative actions in budget-constrained health systems, but also allocative and displacement inefficiency observed in health system practice. Allocative and displacement inefficiency arise in health systems where the least cost-effective program in contraction has a higher incremental cost-effectiveness ratio (ICER = m) than the most cost-effective program in expansion (ICER = n) and displaced services (ICER = d), respectively. The health shadow price derived by Pekarsky, [Formula: see text] reflects the opportunity cost of best alternative adoption and financing actions in reimbursing new technology with expected incremental costs and net effect allowing for allocative (n < m), and displacement, inefficiency (d < m). This provides an appropriate threshold value for the NE quadrant. In this paper, I show that for trade-offs in the SW quadrant, where new strategies have lower expected net cost while lower expected net effect than current practice, the opportunity cost is contraction of the least cost-effective program, with threshold ICER m. That is, in the SW quadrant, the cost reduction per unit of decreased effect should be compared with the appropriate opportunity cost, best alternative generation of funding. Consequently, appropriate consideration of opportunity cost produces a kink in the threshold at the origin, with the health shadow price in the NE quadrant and ICER of the least cost-effective program in contraction (m) in the SW quadrant having the same general shape as that previously suggested by WTP versus WTA. The extent of this kink depends on the degree of allocative and displacement inefficiency, with no kink in the threshold line strictly only appropriate with complete allocative and displacement efficiency, that is n = d = m.


Activation of the GDNF-inducible transcription factor (GIF) gene promoter by glucocorticoid and progesterone.
Steroid hormones, especially glucocorticoids, exert physiologic effects on dopaminergic neurotransmission and have been implicated in several dopamine-mediated neuropsychiatric conditions. D(2) dopamine receptor gene expression is regulated by the zinc finger-type nuclear protein GDNF-inducible transcription factor (GIF). In this study, we sought to investigate if steroids could regulate transcription of the GIF gene itself. Transient co-transfection of the D(2) expressing neuroblastoma cell line NB41A3 with GIF promoter-luciferase constructs along with expression vectors for steroid hormone receptors showed that activation of glucocorticoid receptors but not estrogen receptors up-regulates transcription from the GIF promoter 5.0-fold. Progesterone receptors, which share the same consensus DNA recognition sequence as glucocorticoid receptors, also activated the GIF promoter. Serial 5'-deletion mutants of the GIF gene upstream region localized the glucocorticoid-responsive segment between nucleotides -128 and -66 relative to the transcription start site. This region contains a putative glucocorticoid-responsive element/progesterone-responsive element (GRE/PRE). Additionally, this fragment of the GIF gene 5'-upstream region activated the heterologous herpes simplex virus thymidine kinase (TK) promoter, which is known to be glucocorticoid and progesterone responsive. Furthermore, glucocorticoid receptor activation up-regulated endogenous GIF gene mRNA expression in NB41A3 cells. These observations demonstrate a molecular basis for glucocorticoid and progesterone-induced up-regulation of GIF gene transcription and provide a mechanism for the modulation of dopamine-mediated behaviors by these hormones.


Reducing the incidence of oxyhaemoglobin desaturation during rapid sequence intubation in a paediatric emergency department.
Rapid sequence intubation (RSI) is the standard for definitive airway management in emergency medicine. In a video-based study of RSI in a paediatric emergency department (ED), we reported a high degree of process variation and frequent adverse effects, including oxyhaemoglobin desaturation (SpO2<90%). This report describes a multidisciplinary initiative to improve the performance and safety of RSI in a paediatric ED. We conducted a local improvement initiative in a high-volume academic paediatric ED. We simultaneously tested: (1) an RSI checklist, (2) a pilot/copilot model for checklist execution, (3) the use of a video laryngoscope and (4) the restriction of laryngoscopy to specific providers. Data were collected primarily by video review during the testing period and the historical period (2009-2010, baseline). We generated statistical process control charts (G-charts) to measure change in the performance of six key processes, attempt failure and the occurrence of oxyhaemoglobin desaturation during RSI. We iteratively revised the four interventions through multiple plan-do-study-act cycles within the Model for Improvement. There were 75 cases of RSI during the testing period (July 2012-September 2013). Special cause variation occurred on the G-charts for three of six key processes, attempt failure and desaturation, indicating significant improvement. The frequency of desaturation was 50% lower in the testing period than the historical (16% vs 33%). When all six key processes were performed, only 6% of patients experienced desaturation. Following the simultaneous introduction of four interventions in a paediatric ED, RSI was performed more reliably, successfully and safely.


EuroQol Protocols for Time Trade-Off Valuation of Health Outcomes.
The time trade-off (TTO) valuation technique is widely used to determine utility values of health outcomes to inform quality-adjusted life-year (QALY) calculations for use in economic evaluation. Protocols for implementing TTO vary in aspects such as the trade-off framework, iteration procedure and its administration model and method, training of respondents and interviewers, and quality control of data collection. The most widely studied and utilized TTO valuation protocols are the Measurement and Valuation of Health (MVH) protocol, the Paris protocol and the EuroQol Valuation Technology (EQ-VT) protocol, all developed by members of the EuroQol Group. The MVH protocol and its successor, the Paris protocol, were developed for valuation of EQ-5D-3L health states. Both protocols were designed for a trained interviewer to elicit preferences from a respondent using the conventional TTO framework with a fixed time horizon of 10 years and an iteration procedure combining bisection and titration. Developed for valuation of EQ-5D-5L health states, the EQ-VT protocol adopted a composite TTO framework and made use of computer technology to facilitate data collection. Training and monitoring of interviewers and respondents is a pivotal component of the EQ-VT protocol. Research is underway aiming to further improve the EuroQol protocols, which form an important basis for the current practice of health technology assessment in many countries.


Contrast-enhanced ultrasound to visualize hemodynamic changes after rodent spinal cord injury.
OBJECTIVE Traumatic spinal cord injury (tSCI) causes an almost complete loss of blood flow at the site of injury (primary injury) as well as significant hypoperfusion in the penumbra of the injury. Hypoperfusion in the penumbra progresses after injury to the spinal cord and is likely to be a major contributor to progressive cell death of spinal cord tissue that was initially viable (secondary injury). Neuroprotective treatment strategies seek to limit secondary injury. Clinical monitoring of the temporal and spatial patterns of blood flow within the contused spinal cord is currently not feasible. The purpose of the current study was to determine whether ultrafast contrast-enhanced ultrasound (CEUS) Doppler allows for detection of local hemodynamic changes within an injured rodent spinal cord in real time. METHODS A novel ultrafast CEUS Doppler technique was developed utilizing a research ultrasound platform combined with a 15-MHz linear array transducer. Ultrafast plane-wave acquisitions enabled the separation of higher-velocity blood flow in macrocirculation from low-velocity flow within the microcirculation (tissue perfusion). An FDA-approved contrast agent (microbubbles) was used for visualization of local blood flow in real time. CEUS Doppler acquisition protocols were developed to characterize tissue perfusion both during contrast inflow and during the steady-state plateau. A compression injury of the thoracic spinal cord of adult rats was induced using iris forceps. RESULTS High-frequency ultrasound enabled visualization of spinal cord vessels such as anterior spinal arteries as well as central arteries (mean diameter [± SEM] 145.8 ± 10.0 µm; 76.2 ± 4.5 µm, respectively). In the intact spinal cord, ultrafast CEUS Doppler confirmed higher perfusion of the gray matter compared to white matter. Immediately after compression injury of the thoracic rodent spinal cord, spinal cord vessels were disrupted in an area of 1.93 ± 1.14 mm2. Ultrafast CEUS Doppler revealed a topographical map of local tissue hypoperfusion with remarkable spatial resolution. Critical loss of perfusion, defined as less than 40% perfusion compared to the surrounding spared tissue, was seen within an area of 2.21 ± 0.6 mm2. CONCLUSIONS In our current report, we introduce ultrafast CEUS Doppler for monitoring of spinal vascular structure and function in real time. Development and clinical implementation of this type of imaging could have a significant impact on the care of patients with tSCI.


Structure, expression, and regulation of the murine renin genes.
It has long been known that the renin-angiotensin system plays an integral role in the regulation of blood pressure and electrolyte and fluid balance in mammals. The advent of molecular biologic techniques has afforded new insights into the genes regulating blood pressure. Laboratory mice and rats have been used as experimental models to examine the structural organization and expression of the renin gene. It is now well established that some mice, unlike rats and humans, contain a duplicated copy of the renin locus, which accounts for the high level of renin activity long known to be found in the submandibular gland of some mice. Indeed it is this fortuitous observation that facilitated the isolation of the first complementary DNA clones for renin and ultimately the many species-specific probes now available to analyze mammalian tissues for evidence of primary renin expression. The use of complementary DNAs as probes for primary renin expression helped confirm and further clarify earlier studies demonstrating the presence of renin activity in a number of extrarenal tissues. Although expression in some of these tissues is evolutionarily conserved, their significance has still been elusive. In this report we review the impact of molecular biology on our current understanding of renin gene structure and organization, tissue- and cell-specific expression and regulation, and the changes in renin expression throughout ontogeny. In addition, we describe how new developments in gene transfer technology have added important tools to our arsenal for examining renin gene regulation and how these technologies can be used to develop new tools for renin and hypertension research.


Refined annotation of the Arabidopsis genome by complete expressed sequence tag mapping.
Expressed sequence tags (ESTs) currently encompass more entries in the public databases than any other form of sequence data. Thus, EST data sets provide a vast resource for gene identification and expression profiling. We have mapped the complete set of 176,915 publicly available Arabidopsis EST sequences onto the Arabidopsis genome using GeneSeqer, a spliced alignment program incorporating sequence similarity and splice site scoring. About 96% of the available ESTs could be properly aligned with a genomic locus, with the remaining ESTs deriving from organelle genomes and non-Arabidopsis sources or displaying insufficient sequence quality for alignment. The mapping provides verified sets of EST clusters for evaluation of EST clustering programs. Analysis of the spliced alignments suggests corrections to current gene structure annotation and provides examples of alternative and non-canonical pre-mRNA splicing. All results of this study were parsed into a database and are accessible via a flexible Web interface at http://www.plantgdb.org/AtGDB/.


Trends in Nonfatal Agricultural Injury in Maine and New Hampshire: Results From a Low-Cost Passive Surveillance System.
Agriculture is a dangerous industry, and although data on fatal injuries exist, less is known about nonfatal injuries. The purpose of this study is to describe trends in agricultural morbidity in Maine and New Hampshire from 2008 to 2010 using a newly established passive surveillance system. This passive system is supplied by injury cases gathered from prehospital care reports and hospital data. Demographics and specifics of the event were recorded for each incident case. The average age of injured people in Maine and New Hampshire was 41.7. Women constituted 43.8% of all agricultural injuries. Machinery- (n = 303) and animal- (n = 523) related injuries accounted for most agricultural incidents. Of all injured women, over 60% sustained injuries due to animal-related causes. Agricultural injuries were spread across the two states, with clustering in southern New Hampshire and south central Maine, with additional injuries in the Aroostook County area, which is located in the northeast part of the state. Seasonal variation in agricultural injuries was evident with peaks in the summer months. There was some overlap between the agricultural and logging industry for tree-related work. Our methods are able to capture traumatic injury in agriculture in sufficient detail to prioritize interventions and to evaluate outcomes. The system is low-cost and has the potential to be sustained over a long period. Differences in rates of animal- and machinery-related injuries suggest the need for state-specific safety prioritization.


Short communication: antiretroviral therapy resistance mutations present in the HIV type 1 subtype C pol and env regions from therapy-naive patients in Zambia.
The prevalence of antiretroviral therapy (ART) resistance mutations present in HIV-1 subtype C pol and env regions of the proviral DNA was analyzed and compared from therapy-naive individuals before (Cohort A) and after (Cohort B) the availability of free ART in Zambia. Mutations present in sequences published in a previous study from Zambian ART-naive individuals infected with subtype C were analyzed using current parameters for the classification of ART drug resistance and compared with Cohorts A and B. No statistically significant differences were observed when comparing mutations present in the pol and env of these cohorts. However, an increase in the number of minor, borderline, or partial resistance mutations as well as the presence of major resistance mutations were observed in Cohort B. These results suggest there is an increasing trend of drug resistance-associated mutations that could be a result of the availability of free ART in Zambia. Moreover, the high prevalence of resistance mutations observed for maraviroc and vicriviroc in both cohorts may suggest a limited efficacy of entry inhibitors on HIV-1 subtype C.


Haemosiderin-laden sputum macrophages for diagnosis in pulmonary veno-occlusive disease.
Pulmonary veno-occlusive disease (PVOD) is a rare condition of pulmonary arterial hypertension (PAH), in which post-capillary veins are affected. Since the therapeutic approach in PVOD differs from other forms of PAH, it is crucial to establish the diagnosis. Due to the fact that affected patients are often hemodynamically unstable, minimal invasive procedures are necessary for the diagnostic work-up. Chronic alveolar haemorrhage has been observed during bronchoalveolar lavage in PVOD cases. This study therefore investigates whether signs of alveolar haemorrhage can also be found in the sputum of these patients. Six patients suffering from PVOD were included in this analysis. As controls, patients with idiopathic PAH (n = 11), chronic thromboembolic PH (n = 9) and with sclerodermia-associated PH (n = 10) were assessed. Sputum from every patient was obtained by a non-invasive manner. The amount of haemosiderin-laden macrophages was determined using the Golde score. There were statistically significant more haemosiderin-laden macrophages in the sputum of patients suffering from PVOD as compared to the other groups (P<0.05). Assuming a cut-off of 200 on the Golde score, all of the 6 PVOD patients surpassed this value compared with only 1 out of the 30 cases with precapillary PH. Thus, sensitivity and specificity with respect to the diagnosis of PVOD was 100% and 97%, respectively. The content of haemosiderin-laden macrophages in the sputum of patients suffering from PVOD is significantly higher as compared to other forms of PH and may be useful in the non-invasive diagnostic work-up of these patients.


Crouzon Syndrome: Relationship of Eye Movements to Pattern Strabismus.
To characterize conjugate eye movements in Crouzon syndrome (CS) patients with and without strabismus. Smooth pursuit, saccades, horizontal optokinetic nystagmus (OKN), and horizontal vestibulo-ocular reflex (VOR) were recorded using binocular video-oculography (VOG) in 10 children with CS (5 orthotropic, 5 strabismic) and 12 age-matched controls. Hess-Lancaster plots were generated from Orbit 1.8 using rectus muscle pulley locations from computed tomography imaging. Two-dimensional eye scan paths from VOG recordings were compared with the Hess-Lancaster plots. Targeted saccades were normometric on average but variable, and followed the main sequence in both CS groups. Smooth pursuit gains were normal for both CS groups; however, SP gains of the fixating eye in subjects with strabismus were significantly lower. Optokinetic nystagmus gains were reduced in both CS groups (P < 0.02) but were lower in subjects with strabismus. Shifting misalignments of binocular eye position in primary and eccentric gazes were associated with reduction in OKN gain in both CS groups. Vestibulo-ocular reflex gains for both CS groups were largely normal despite the presence of an off-axis vertical component. Normal gains for saccades and smooth pursuit in CS patients with strabismus are consistent with accurate execution of both movements despite extorsion of the globe. Vestibulo-ocular reflex in CS patients with strabismus had an off-axis vertical component consistent with extorted muscle pulleys. Optokinetic nystagmus is reduced in CS without strabismus owing to binocular position disparities and in CS with strabismus is likely due to cortical suppression associated with cross-axis orientation and exotropia.


In vitro eugenics.
A series of recent scientific results suggest that, in the not-too-distant future, it will be possible to create viable human gametes from human stem cells. This paper discusses the potential of this technology to make possible what I call 'in vitro eugenics': the deliberate breeding of human beings in vitro by fusing sperm and egg derived from different stem-cell lines to create an embryo and then deriving new gametes from stem cells derived from that embryo. Repeated iterations of this process would allow scientists to proceed through multiple human generations in the laboratory. In vitro eugenics might be used to study the heredity of genetic disorders and to produce cell lines of a desired character for medical applications. More controversially, it might also function as a powerful technology of 'human enhancement' by allowing researchers to use all the techniques of selective breeding to produce individuals with a desired genotype.


MicroRNA-31 Regulates Chemosensitivity in Malignant Pleural Mesothelioma.
Malignant pleural mesothelioma (MPM) is associated with an extremely poor prognosis, and most patients initially are or rapidly become unresponsive to platinum-based chemotherapy. MicroRNA-31 (miR-31) is encoded on a genomic fragile site, 9p21.3, which is reportedly lost in many MPM tumors. Based on previous findings in a variety of other cancers, we hypothesized that miR-31 alters chemosensitivity and that miR-31 reconstitution may influence sensitivity to chemotherapeutics in MPM. Reintroduction of miR-31 into miR-31 null NCI-H2452 cells significantly enhanced clonogenic resistance to cisplatin and carboplatin. Although miR-31 re-expression increased chemoresistance, paradoxically, a higher relative intracellular accumulation of platinum was detected. This was coupled to a significantly decreased intranuclear concentration of platinum. Linked with a downregulation of OCT1, a bipotential transcriptional regulator with multiple miR-31 target binding sites, we subsequently identified an indirect miR-31-mediated upregulation of ABCB9, a transporter associated with drug accumulation in lysosomes, and increased uptake of platinum to lysosomes. However, when overexpressed directly, ABCB9 promoted cellular chemosensitivity, suggesting that miR-31 promotes chemoresistance largely via an ABCB9-independent mechanism. Overall, our data suggest that miR-31 loss from MPM tumors promotes chemosensitivity and may be prognostically beneficial in the context of therapeutic sensitivity.


Allelic functional variation of serotonin transporter expression is a susceptibility factor for late onset Alzheimer's disease.
We examined a deletion/insertion promoter polymorphism of the serotonin transporter gene, which confers an approximately 40% reduction in expression of the protein, in 196 subjects with late onset Alzheimer's disease (AD) and 271 controls. The frequency of the 484 bp low activity allele was elevated in the subjects with AD (p = 0.004), and an excess of the low activity genotype (30%) was also found in comparison with the controls (20%) (chi 2 = 7.16; p = 0.03). This association was unrelated to the age of the subjects or controls, or to epsilon 4 alleles of the ApoE gene. The odds ratio for the effect of the homozygous low activity genotype was 1.7 (95% CI 1.08-2.67), with a population attributable risk of 33% (95% CI 5-54%). These findings indicate that the low activity allele of the serotonin transporter is a risk factor for late onset AD.


Accuracy of a commercial multiplex PCR for the diagnosis of bacterial vaginosis.
Bacterial vaginosis (BV) is a common clinical condition characterized by odorous vaginal discharge, vaginal itching and/or burning. BV can occur when vaginal lactobacilli are depleted and replaced by diverse anaerobic bacteria. We evaluated a commercial multiplex PCR (ATRiDA) for the diagnosis of BV. Cervicovaginal samples were included from women reporting urogenital symptoms and from women notified for sexually transmitted infections (STI) - who were not (necessarily) symptomatic. Clinical BV diagnoses were obtained from electronic patient files. The ATRiDA test measures the loads of Gardnerella vaginalis, Atopobium vaginae and Lactobacillus species in relation to overall bacterial load. The ATRiDA test outcome was compared to the clinical BV diagnosis and to vaginal microbiota composition, determined by 16SrRNA gene sequencing. We included samples from 185 women reporting urogenital symptoms, of whom 81 had BV and 93 women who were notified for an STI, of whom 16 had BV. Overall, compared to the clinical BV diagnosis, the ATRiDA test demonstrated high sensitivity (96.9 %) and moderate specificity (70.2 %). The negative predictive value was high (>97.3). The positive predictive value differed by study group and was highest in women reporting urogenital symptoms (78.2 %). Sequencing showed that 54 % of women who had an ATRiDA BV-positive test outcome, but who were not clinically diagnosed with BV, had diverse anaerobic vaginal microbiota (asymptomatic vaginal dysbiosis). The ATRiDA test is a sensitive method for the detection of BV but, given the high occurrence of asymptomatic vaginal dysbiosis, a positive test outcome should be interpreted together with clinical symptoms.


Distinct classes and subclasses of antibodies to hemolysin co-regulated protein 1 and O-polysaccharide and correlation with clinical characteristics of melioidosis patients.
Melioidosis is a tropical infectious disease caused by Burkholderia pseudomallei that results in high mortality. Hemolysin co-regulated protein 1 (Hcp1) and O-polysaccharide (OPS) are vaccine candidates and potential diagnostic antigens. The correlation of classes/subclasses of antibodies against these antigens with clinical characteristics of melioidosis patients is unknown. Antibodies in plasma samples from melioidosis patients and healthy donors were quantified by ELISA and compared with clinical features. In melioidosis patients, Hcp1 induced high IgG levels. OPS induced high IgG and IgA levels. The area under receiver operating characteristic curve (AUROCC) to discriminate melioidosis cases from healthy donors was highest for anti-Hcp1 IgG (0.92) compared to anti-Hcp1 IgA or IgM. In contrast, AUROCC for anti-OPS for IgG (0.91) and IgA (0.92) were comparable. Anti-Hcp1 IgG1 and anti-OPS IgG2 had the greatest AUROCCs (0.87 and 0.95, respectively) compared to other IgG subclasses for each antigen. Survivors had significantly higher anti-Hcp1 IgG3 levels than non-survivors. Male melioidosis patients with diabetes had higher anti-OPS IgA levels than males without diabetes. Thus, diverse and specific antibody responses are associated with distinct clinical characteristics in melioidosis, confirming the diagnostic utility of these responses and providing new insights into immune mechanisms.


Physiological characterization of adaptive clones in evolving populations of the yeast, Saccharomyces cerevisiae.
Populations of a diploid strain of S. cerevisiae were grown in glucose-limited continuous culture for more than 260 generations. A series of seven sequential adaptive changes were identified by monitoring the frequency of cycloheximide resistance in these populations. Samples were taken from the continuous cultures following each adaptive shift and characterized physiologically to determine (1) the range of phenotypes that can be selected in a precisely defined constant environment and (2) the order and predictability of the occurrence of the adaptive mutations in evolving populations. The clones were characterized with respect to the growth parameters, maximum growth rate, saturation coefficient and yield, as well as for changes in cell size and geometry and rate of glucose uptake. The maximum growth rates of the seven adaptive clones were very similar, but in contrast the saturation coefficients differed substantially. Surprisingly, not all clones showed reductions in the saturation coefficients, in comparison to the immediately preceding clones, as would be predicted from classical continuous culture kinetics. In addition, yield estimates first increased and then decreased for later isolated adaptive clones. In general, the results suggest epistatic interactions between the adaptive clones, consistent with earlier published results. The rate of glucose uptake, as measured by 14C-xylose uptake, increased dramatically after the selection and fixation of seven adaptive clones. Progressive decreases in cell volume and changes in cell geometry, resulting in increased surface area to volume ratios, were also observed in the adaptive clones, but these changes were not always seen in other haploid and diploid yeast populations evolving under the same conditions. Such changes may be easily explainable in terms of the characteristics of the glucose-limited environment. The significance of the results to the evolution of microorganisms under nutrient-limiting conditions is discussed.


The impact of extent of resection on malignant transformation of pure oligodendrogliomas.
Recent evidence suggests that a greater extent of resection (EOR) extends malignant progression-free survival among patients with low-grade gliomas (LGGs). These studies, however, rely on the combined analysis of oligodendrogliomas, astrocytomas, and mixed oligoastrocytomas-3 histological subtypes with distinct genetic and molecular compositions. To assess the value of EOR in a homogeneous LGG patient population and delineate its impact on LGG transformation, the authors examined its effect on newly diagnosed supratentorial oligodendrogliomas. The authors identified 93 newly diagnosed adult patients with WHO Grade II oligodendrogliomas treated with microsurgical resection at Barrow Neurological Institute. Clinical, laboratory, and radiographic data were collected retrospectively, including 1p/19q codeletion status and volumetric analysis based on T2-weighted MRI. The median preoperative and postoperative tumor volumes and EOR were 29.0 cm(3) (range 1.3-222.7 cm(3)), 5.2 cm(3) (range 0-156.1 cm(3)), and 85% (range 6%-100%), respectively. Median follow-up was 75.4 months, and there were 14 deaths (15%). Progression and malignant progression were identified in 31 (33%) and 20 (22%) cases, respectively. A greater EOR was associated with longer overall survival (p = 0.005) and progression-free survival (p = 0.004); however, a greater EOR did not prolong the interval to malignant progression, even when controlling for 1p/19q codeletion. A greater EOR is associated with an improved survival profile for patients with WHO Grade II oligodendrogliomas. However, for this particular LGG patient population, the interval to tumor transformation is not influenced by cytoreduction. These data raise the possibility that the capacity for microsurgical resection to modulate malignant progression is mediated through biological mechanisms specific to nonoligodendroglioma LGG histologies.


Recovery of phenotypes obtained by adaptive evolution through inverse metabolic engineering.
In a previous study, system level analysis of adaptively evolved yeast mutants showing improved galactose utilization revealed relevant mutations. The governing mutations were suggested to be in the Ras/PKA signaling pathway and ergosterol metabolism. Here, site-directed mutants having one of the mutations RAS2(Lys77), RAS2(Tyr112), and ERG5(Pro370) were constructed and evaluated. The mutants were also combined with overexpression of PGM2, earlier proved as a beneficial target for galactose utilization. The constructed strains were analyzed for their gross phenotype, transcriptome and targeted metabolites, and the results were compared to those obtained from reference strains and the evolved strains. The RAS2(Lys77) mutation resulted in the highest specific galactose uptake rate among all of the strains with an increased maximum specific growth rate on galactose. The RAS2(Tyr112) mutation also improved the specific galactose uptake rate and also resulted in many transcriptional changes, including ergosterol metabolism. The ERG5(Pro370) mutation only showed a small improvement, but when it was combined with PGM2 overexpression, the phenotype was almost the same as that of the evolved mutants. Combination of the RAS2 mutations with PGM2 overexpression also led to a complete recovery of the adaptive phenotype in galactose utilization. Recovery of the gross phenotype by the reconstructed mutants was achieved with much fewer changes in the genome and transcriptome than for the evolved mutants. Our study demonstrates how the identification of specific mutations by systems biology can direct new metabolic engineering strategies for improving galactose utilization by yeast.


Schizotypy and nicotine, alcohol, and cannabis use in a non-psychiatric sample.
Schizotypy is a multidimensional personality construct that is characterized by perceptual abnormalities, social withdrawal, mild suspiciousness, and odd thinking patterns. This study examined the relationship between four dimensions of self-reported schizotypy and substance use involving nicotine, alcohol, and cannabis, in undergraduate students. Results showed that higher levels of disorganized schizotypy, or odd thinking and behavior, were associated with greater indices of use of all three substances. Furthermore, higher cognitive-perceptual schizotypy was selectively associated with cannabis use. Results confirm findings of recent research that has discovered associations among schizotypy and substance use, highlighting links between behavioral traits and use of nicotine, alcohol, and cannabis. This study is one of the first to examine a wide range of schizotypy domains, and to show selective effects of the disorganized domain of schizotypy.


Interactions between species introduce spurious associations in microbiome studies.
Microbiota contribute to many dimensions of host phenotype, including disease. To link specific microbes to specific phenotypes, microbiome-wide association studies compare microbial abundances between two groups of samples. Abundance differences, however, reflect not only direct associations with the phenotype, but also indirect effects due to microbial interactions. We found that microbial interactions could easily generate a large number of spurious associations that provide no mechanistic insight. Using techniques from statistical physics, we developed a method to remove indirect associations and applied it to the largest dataset on pediatric inflammatory bowel disease. Our method corrected the inflation of p-values in standard association tests and showed that only a small subset of associations is directly linked to the disease. Direct associations had a much higher accuracy in separating cases from controls and pointed to immunomodulation, butyrate production, and the brain-gut axis as important factors in the inflammatory bowel disease.


Effect of folic Acid supplementation on the folate status of buccal mucosa and lymphocytes.
Folate deficiency may be associated with an increased risk of cancer at certain sites. There is a need to measure folate status and putative biomarkers of cancer risk in the same target tissue, or in surrogate tissues. A study was carried out to develop a method for the rapid measurement of folate in human buccal mucosa and lymphocytes and to evaluate the responsiveness of this measurement in both tissues to folic acid supplementation in healthy subjects, relative to conventional markers of folate status. Three hundred and twenty-three adults, ages between 20 and 60 years, were screened for RBC folate concentrations. Sixty-five subjects with red cell folate between 200 and 650 nmol/L participated in a randomized, double blind, placebo-controlled, folic acid (1.2 mg) intervention trial, lasting 12 weeks. As anticipated, a significant baseline correlation (r = 0.36, P < 0.01) was observed between red cell folate and plasma 5-methyltetrahydrofolate (5-MeTHF). Lymphocyte total folate was significantly associated with plasma 5-MeTHF (r = 0.28, P < 0.05) and plasma total homocysteine concentration (r = -0.34, P < 0.05). Buccal mucosa total folate showed no correlation with either red cell folate or 5-MeTHF, but was significantly associated with lymphocyte total folate (r = 0.35, P < 0.01). Supplementation elicited a significant increase in lymphocyte total folate (P < 0.01), and this was strongly associated with the increase in RBC total folate (P < 0.01) and plasma 5-MeTHF (P < 0.01). Buccal mucosa total folate was not influenced by folate supplementation. Methods have been developed for the rapid measurement of lymphocyte and buccal mucosal total folate. Lymphocyte folate is sensitive to folate intake and is reflected by plasma 5-MeTHF.


Altered expression of the voltage-gated calcium channel subunit α₂δ-1: a comparison between two experimental models of epilepsy and a sensory nerve ligation model of neuropathic pain.
The auxiliary α2δ-1 subunit of voltage-gated calcium channels is up-regulated in dorsal root ganglion neurons following peripheral somatosensory nerve damage, in several animal models of neuropathic pain. The α2δ-1 protein has a mainly presynaptic localization, where it is associated with the calcium channels involved in neurotransmitter release. Relevant to the present study, α2δ-1 has been shown to be the therapeutic target of the gabapentinoid drugs in their alleviation of neuropathic pain. These drugs are also used in the treatment of certain epilepsies. In this study we therefore examined whether the level or distribution of α2δ-1 was altered in the hippocampus following experimental induction of epileptic seizures in rats, using both the kainic acid model of human temporal lobe epilepsy, in which status epilepticus is induced, and the tetanus toxin model in which status epilepticus is not involved. The main finding of this study is that we did not identify somatic overexpression of α2δ-1 in hippocampal neurons in either of the epilepsy models, unlike the upregulation of α2δ-1 that occurs following peripheral nerve damage to both somatosensory and motor neurons. However, we did observe local reorganization of α2δ-1 immunostaining in the hippocampus only in the kainic acid model, where it was associated with areas of neuronal cell loss, as indicated by absence of NeuN immunostaining, dendritic loss, as identified by areas where microtubule-associated protein-2 immunostaining was missing, and reactive gliosis, determined by regions of strong OX42 staining.


WD40 domain divergence is important for functional differences between the fission yeast Tup11 and Tup12 co-repressor proteins.
We have previously demonstrated that subsets of Ssn6/Tup target genes have distinct requirements for the Schizosaccharomyces pombe homologs of the Tup1/Groucho/TLE co-repressor proteins, Tup11 and Tup12. The very high level of divergence in the histone interacting repression domains of the two proteins suggested that determinants distinguishing Tup11 and Tup12 might be located in this domain. Here we have combined phylogenetic and structural analysis as well as phenotypic characterization, under stress conditions that specifically require Tup12, to identify and characterize the domains involved in Tup12-specific action. The results indicate that divergence in the repression domain is not generally relevant for Tup12-specific function. Instead, we show that the more highly conserved C-terminal WD40 repeat domain of Tup12 is important for Tup12-specific function. Surface amino acid residues specific for the WD40 repeat domain of Tup12 proteins in different fission yeasts are clustered in blade 3 of the propeller-like structure that is characteristic of WD40 repeat domains. The Tup11 and Tup12 proteins in fission yeasts thus provide an excellent model system for studying the functional divergence of WD40 repeat domains.


Monitoring Age-Group Swimmers Over a Training Macrocycle: Energetics, Technique, and Anthropometrics.
Zacca, R, Azevedo, R, Chainok, P, Vilas-Boas, JP, Castro, FAdS, Pyne, DB, and Fernandes, RJ. Monitoring age-group swimmers over a training macrocycle: energetics, technique, and anthropometrics. J Strength Cond Res 34(3): 818-827, 2020-The aim of this study was to quantify changes and contributions of energetic, technique, and anthropometric profiles across the first training macrocycle (16-week) in a traditional 3-peak swimming season. Twenty-four age-group swimmers (10 boys and 14 girls age 14.4 ± 0.9 years) of equal maturational stage were monitored through a 400-m test in front crawl (T400). Energetic, technique, and anthropometric characteristics were compared before (experimental testing 1, E1) and after the preparatory (E2), specific (E3), and competitive (E4) training periods. Sex interaction was not significant for any variable. Multiple linear regressions and principal component analysis were used to identify the most influential variables and the relative contribution of each domain (energetics, technique, and anthropometrics) to changes in swimming performance of T400. The relative contributions for performance of T400 at E1, E2, E3, and E4 were 15, 12, 6, and 13% for energetics, 78, 85, 75, and 70% for technique, and 7, 3, 19 and 17% for anthropometrics, respectively. Technique played the main role during the first 16-week macrocycle in a competitive season, regardless of small fluctuations in the influence of energetics and anthropometrics. Changes and influence of energetics, technique, and anthropometric on age-group swimmers' performance could be described by the T400 swimming test, providing a comprehensive biophysical overview of the main contributors to swimming performance.


Fiber-optic Fabry-Perot sensors based on a combination of spatial-frequency division multiplexing and wavelength division multiplexing formed by chirped fiber Bragg grating pairs.
Effective multiplexing for a very large number of fiber-optic fiber-Bragg-grating-based Fabry-Perot (FBGFP) sensors is proposed that is based on wavelength division multiplexing (WDM) and spatial-frequency division multiplexing (SFDM). For WDM, FBGFP sensors are arranged in different wavelength domains formed by a series of chirped fiber Bragg gratings with different central wavelengths while the sensors with different cavity lengths within the same wavelength domain are multiplexed by use of SFDM because they have different spatial frequencies as a result of their different cavity lengths. In principle, a thousand FBGFP sensors could be multiplexed with such an approach. The experimental results show that a strain accuracy of better than +/-10 microepsilon has been achieved with little cross talk.


Mitochondrial optic neuropathies.
Mitochondrial optic neuropathies have a leading role in the field of mitochondrial medicine ever since 1988, when the first mutation in mitochondrial DNA was associated with Leber's hereditary optic neuropathy (LHON). Autosomal dominant optic atrophy (DOA) was subsequently associated in 2000 with mutations in the nuclear DNA affecting the OPA1 gene. LHON and DOA are both characterized by selective neurodegeneration of retinal ganglion cells (RGCs) triggered by mitochondrial dysfunction. This is centered on respiratory complex I impairment in LHON and defective mitochondrial dynamics in OPA1-related DOA, leading to distinct clinical phenotypes. LHON is a subacute, rapid, severe loss of central vision involving both eyes within weeks or months, with age of onset between 15 and 35 years old. DOA is a more slowly progressive optic neuropathy, usually apparent in early childhood. LHON is characterized by marked incomplete penetrance and a clear male predilection. The introduction of next-generation sequencing has greatly expanded the genetic causes for other rare forms of mitochondrial optic neuropathies, including recessive and X-linked, further emphasizing the exquisite sensitivity of RGCs to compromised mitochondrial function. All forms of mitochondrial optic neuropathies, including LHON and DOA, can manifest either as pure optic atrophy or as a more severe multisystemic syndrome. Mitochondrial optic neuropathies are currently at the forefront of a number of therapeutic programs, including gene therapy, with idebenone being the only approved drug for a mitochondrial disorder.


Proton magnetic resonance spectroscopy for detection of axonal injury in the splenium of the corpus callosum of brain-injured patients.
This study was conducted to determine whether proton magnetic resonance spectroscopy (MRS) is a sensitive method for detecting diffuse axonal injury, which is a primary sequela of traumatic brain injury (TBI). Diffuse axonal injury is characterized by selective damage to white matter tracts that is caused in part by the severe inertial strain created by rotational acceleration and deceleration, which is often associated with motor vehicle accidents. This axonal injury is typically difficult to detect by using conventional imaging techniques because it is microscopic in nature. The splenium was selected because it is a site vulnerable to shearing forces that produce diffuse axonal injury. The authors used proton MRS to evaluate the splenium, the posterior commissure of the corpus callosum, in normal control volunteers and in patients with TBI. Proton MRS provided an index of neuronal and axonal viability by measuring levels of N-acetyl aspartate (NAA). A majority of mildly brain injured patients, as well as those more severely injured, showed diminished NAA/creatine (Cr) levels in the splenium compared with normal control volunteers. The patients displaying lowered NAA/Cr in the splenium were also likely to exhibit lowered NAA/Cr in lobar white matter. Also, the levels of NAA/Cr in the splenium of normal volunteers were higher compared with those found in lobar white matter. Decreases in NAA/Cr levels in the splenium may be a marker for diffuse injury. A proton MRS examination may be particularly useful in evaluating mildly injured patients with unexplained neurological and cognitive deficits. It is concluded that MRS is a sensitive tool in detecting axonal injury.


Positron emission tomography for monitoring response to neoadjuvant therapy in patients with oesophageal and gastro-oesophageal junction carcinoma.
The aim of this review is to consolidate our knowledge on an important and rapidly expanding area of expertise. Numerous methods for predicting response (in terms of pathological response and survival) to neoadjuvant therapy (chemotherapy/chemo-radiotherapy) in oesophageal and junctional cancers have been proposed. This review concerns itself only with the use of positron emission tomography for such a purpose. At present there are no standardised criteria amongst PET trials as to what determines a response according to PET, what is the optimal time to perform PET in relation to the timing of neoadjuvant therapy, and what is the ideal method of quantifying PET tracer uptake. An electronic search was performed of PubMed, Ovid and Embase websites to identify studies, in the English language, using the search terms: PET; oesophageal; oesophago-gastric; survival; cancer; response; chemotherapy and chemo-radiotherapy. The reference lists were searched manually to identify further relevant studies. Twenty-two studies were identified, all using (18)FDG as the tracer, using PET to predict response in terms of pathological response and survival following neoadjuvant therapy (chemotherapy/chemo-radiotherapy). PET had a varying degree of success in predicting both pathological response and survival outcomes, with only one study using PET to influence management decisions. PET seems a promising technique, but large-scale conclusions are hindered by small study numbers, lack of criteria as to what constitutes a response and markedly differing PET imaging times. A large randomised trial concerning a homogeneous group of patients and tumours is required before PET might be used to influence management.


2H[19F] REDOR for distance measurements in biological solids using a double resonance spectrometer.
A new approach for distance measurements in biological solids employing 2H[19F] rotational echo double resonance was developed and validated on 2H,19F-D-alanine and an imidazopyridine based inhibitor of the gastric H+/K+-ATPase. The 2H-19F double resonance experiments presented here were performed without 1H decoupling using a double resonance NMR spectrometer. In this way, it was possible to benefit from the relatively longer distance range of fluorine without the need of specialized fluorine equipment. A distance of 2.5 +/- 0.3 A was measured in the alanine derivative, indicating a gauche conformation of the two labels. In the case of the imidazopyridine compound a lower distance limit of 5.2 A was determined and is in agreement with an extended conformation of the inhibitor. Several REDOR variants were compared, and their advantages and limitations discussed. Composite fluorine dephasing pulses were found to enhance the frequency bandwidth significantly, and to reduce the dependence of the performance of the experiment on the exact choice of the transmitter frequency.


Immunophenotypic differences between osteoclasts and macrophage polykaryons: immunohistological distinction and implications for osteoclast ontogeny and function.
The antigenic phenotype of human fetal osteoclasts was compared with that of human tissue macrophages and macrophage polykaryons in foreign body lesions using a large number of monoclonal antibodies directed against myeloid (granulocyte/mononuclear phagocyte) antigens. Osteoclasts expressed a restricted range of macrophage-associated antigens including CD13, CD15A, CD44, CD45, CD54, (ICAM-1), CD71 (transferrin receptor), and CD68. These antigens were also present on macrophages and macrophage polykaryons both of which also strongly expressed CD11a,b,c, CD18, (LFA family), CD14, CD31, CD36, CD37, CD39 and CD43 antigens. There was also weak and occasional expression of CD16 (FcRIII), CD25 (interleukin 2 receptor), CD32 (FcRII), CD35 (C3b receptor) and HLA-DR by macrophage polykaryons. The presence of some macrophage associated antigens on osteoclasts is consistent with their originating from cells of the mononuclear phagocyte system. The numerous differences in antigenic phenotype between osteoclasts and macrophage polykaryons, however, suggest that their pathways of development and differentiation are not identical. The differences discerned in antigenic phenotype should also permit distinction between these polykaryons (and possibly their mononuclear precursors) in normal and diseased tissues.


'We were treated like we are nobody': a mixed-methods study of medical doctors' internship experiences in Kenya and Uganda.
Medical interns are an important workforce providing first-line healthcare services in hospitals. The internship year is important for doctors as they transition from theoretical learning with minimal hands-on work under supervision to clinical practice roles with considerable responsibility. However, this transition is considered stressful and commonly leads to burn-out due to challenging working conditions and an ongoing need for learning and assessment, which is worse in countries with resource constraints. In this study, we provide an overview of medical doctors' internship experiences in Kenya and Uganda. Using a convergent mixed-methods approach, we collected data from a survey of 854 medical interns and junior doctors and semistructured interviews with 54 junior doctors and 14 consultants. Data collection and analysis were guided by major themes identified from a previous global scoping review (well-being, educational environment and working environment and condition), using descriptive analysis and thematic analysis respectively for quantitative and qualitative data. Most medical interns are satisfied with their job but many reported suffering from stress, depression and burn-out, and working unreasonable hours due to staff shortages. They are also being affected by the challenging working environment characterised by a lack of adequate resources and a poor safety climate. Although the survey data suggested that most interns were satisfied with the supervision received, interviews revealed nuances where many interns faced challenging scenarios, for example, poor supervision, insufficient support due to consultants not being available or being 'treated like we are nobody'. We highlight challenges experienced by Kenyan and Ugandan medical interns spanning from burn-out, stress, challenging working environment, inadequate support and poor quality of supervision. We recommend that regulators, educators and hospital administrators should improve the resource availability and capacity of internship hospitals, prioritise individual doctors' well-being and provide standardised supervision, support systems and conducive learning environments.


Control of variceal bleeding by superior mesenteric artery vasopressin infusion.
Liver function and clotting tests were evaluated in 39 patients with variceal bleeding prior to superior mesenteric artery vasopressin infusion. In six patients with mild hepatic dysfunction (Child's class A), permanent control of hemorrhage was achieved in all six and all survived the hospitalization. In 21 patients with moderate dysfunction (Child's class B), permanent control of hemorrhage was achieved in 13 (62%) and temporary control for 24 hr or longer in the remaining eight (38%). Survival in class B was 67% (14 of 21). In only four of 12 patients with severe hepatic dysfunction (Child's class C) was control of hemorrhage achieved (33 percent). None of these patients survived. Therapeutic failure also was associated with clotting derangements and the initial bleeding rate. It is concluded that the effectiveness of vasopressin in variceal hemorrhage is a function of the underlying liver disease and derangements in clotting function.


Analysis of antibody induction upon immunization with distinct NTS-DBL1α-domains of PfEMP1 from rosetting Plasmodium falciparum parasites.
Rosette-formation of Plasmodium falciparum parasitized erythrocytes is of importance in the development of severe malaria. The parasite-derived molecule PfEMP1 (Plasmodium falciparum erythrocyte membrane protein 1), central to rosetting, is suggested to be included in a multimeric vaccine targeting severe disease. Three recombinant NTS-DBL1α-domains of PfEMP1 were generated in Escherichia coli, purified and used for immunization of rats and goats. Antibody titres were determined in ELISA assays and responses were compared in-between different individual animals and species. Reactivity with the parasites was tested in live pRBC using FACS. B-cell epitopes prediction was carried out in silico and compared to the results obtained by peptide microarray. Screening for serological cross-reactivity with heterologous NTS-DBL1α variants was carried out by ELISA, peptide array and FACS on pRBC of different laboratory strains and patient isolates. All three NTS-DBL1α-domains induced high titres of antibodies that were biologically active with no apparent difference between constructs covering slightly different parts of the DBL1α-sequence. The different animal species showed comparable titres of antibodies, while variations within individuals of the species could be observed.Mapping of the recognized epitopes revealed that most parts of the molecule were able to induce an antibody response with a tendency for the N and C terminal parts of the molecule for slightly higher recognition. Important differences to the epitopes predicted were found as some of the most conserved parts of the DBL1α-domain contained the main epitopes for antibody reactivity. ELISA assays and peptide microarray demonstrated substantial cross-reactivity to heterologous variants, while binding to native PfEMP1 was observed only in few combinations on the pRBC surface, underlining that mainly internal, conserved and not surface exposed parts of the DBL1α-domain are responsible for this observation. Biologically active antibodies can be induced consistently, with high titres, in different animal species and the antibodies elicited by different constructs react with similar epitopes. Induced antibodies recognize epitopes localized in all subdomains of the DBL1α-sequence. Cross-reactivity between NTS-DBL1α-variants is common in ELISA, but rare with live pRBC emphasizing that also internal, conserved areas of PfEMP1 carry important highly immunogenic epitopes of the molecule.


Hepatopulmonary syndrome is a frequent cause of dyspnea in the short telomere disorders.
Telomere syndromes have their most common manifestation in idiopathic pulmonary fibrosis and emphysema. The short telomere defect in these patients may manifest systemically as bone marrow failure and liver disease. We sought to understand the causes of dyspnea in telomerase and telomere gene mutation carriers who have no parenchymal lung disease. Clinical and pathologic data were reviewed as part of a Johns Hopkins-based natural history study of short telomere syndromes including dyskeratosis congenita. Hepatopulmonary syndrome (HPS) was diagnosed in nine of 42 cases (21%). Their age at presentation was significantly younger than that of cases initially presenting with pulmonary fibrosis and emphysema (median, 25 years vs 55 years; P < .001). Cases had evidence of intra- and extrapulmonary arteriovascular malformations that caused shunt physiology. Nodular regenerative hyperplasia was the most frequent histopathologic abnormality, and it was seen in the absence of cirrhosis. Dyspnea and portal hypertension were progressive, and the median time to death or liver transplantation was 6 years (range, 4-10 years; n = 6). In cases that underwent liver transplantation, dyspnea and hypoxia improved, but pulmonary fibrosis subsequently developed. This report identifies HPS as a frequent cause of dyspnea in telomerase and telomere gene mutation carriers. While it usually precedes the development of parenchymal lung disease, HPS may also co-occur with pulmonary fibrosis and emphysema. Recognizing this genetic diagnosis is critical for management, especially in the lung and liver transplantation setting.


Pharmacological enhancement of mGlu1 metabotropic glutamate receptors causes a prolonged symptomatic benefit in a mouse model of spinocerebellar ataxia type 1.
Spinocerebellar ataxia type 1 (SCA1) is a genetic disorder characterized by severe ataxia associated with progressive loss of cerebellar Purkinje cells. The mGlu1 metabotropic glutamate receptor plays a key role in mechanisms of activity-dependent synaptic plasticity in the cerebellum, and its dysfunction is linked to the pathophysiology of motor symptoms associated with SCA1. We used SCA1 heterozygous transgenic mice (Q154/Q2) as a model for testing the hypothesis that drugs that enhance mGlu1 receptor function may be good candidates for the medical treatment of SCA1. Symptomatic 30-week old SCA1 mice showed reduced mGlu1 receptor mRNA and protein levels in the cerebellum. Interestingly, these mice also showed an intense expression of mGlu5 receptors in cerebellar Purkinje cells, which normally lack these receptors. Systemic treatment of SCA1 mice with the mGlu1 receptor positive allosteric modulator (PAM), Ro0711401 (10 mg/kg, s.c.), caused a prolonged improvement of motor performance on the rotarod and the paw-print tests. A single injection of Ro0711401 improved motor symptoms for several days, and no tolerance developed to the drug. In contrast, the mGlu5 receptor PAM, VU0360172 (10 mg/kg, s.c.), caused only a short-lasting improvement of motor symptoms, whereas the mGlu1 receptor antagonist, JNJ16259685 (2.5 mg/kg, i.p.), further impaired motor performance in SCA1 mice. The prolonged symptomatic benefit caused by Ro0711401 outlasted the time of drug clearance from the cerebellum, and was associated with neuroadaptive changes in the cerebellum, such as a striking reduction of the ectopically expressed mGlu5 receptors in Purkinje cells, increases in levels of total and Ser880-phosphorylated GluA2 subunit of AMPA receptors, and changes in the length of spines in the distal dendrites of Purkinje cells. These data demonstrate that pharmacological enhancement of mGlu1 receptors causes a robust and sustained motor improvement in SCA1 mice, and lay the groundwork for the development of mGlu1 receptor PAMs as novel "cerebellum-specific", effective, and safe symptomatic drugs for the treatment of SCA1 in humans.


A framework for assessing the risk of resistance for anti-malarials in development.
Resistance is a constant challenge for anti-infective drug development. Since they kill sensitive organisms, anti-infective agents are bound to exert an evolutionary pressure toward the emergence and spread of resistance mechanisms, if such resistance can arise by stochastic mutation events. New classes of medicines under development must be designed or selected to stay ahead in this vicious circle of resistance control. This involves both circumventing existing resistance mechanisms and selecting molecules which are resilient against the development and spread of resistance. Cell-based screening methods have led to a renaissance of new classes of anti-malarial medicines, offering us the potential to select and modify molecules based on their resistance potential. To that end, a standardized in vitro methodology to assess quantitatively these characteristics in Plasmodium falciparum during the early phases of the drug development process has been developed and is presented here. It allows the identification of anti-malarial compounds with overt resistance risks and the prioritization of the most robust ones. The integration of this strategy in later stages of development, registration, and deployment is also discussed.


Advance directives and outcomes of surrogate decision making before death.
Recent discussions about health care reform have raised questions regarding the value of advance directives. We used data from survey proxies in the Health and Retirement Study involving adults 60 years of age or older who had died between 2000 and 2006 to determine the prevalence of the need for decision making and lost decision-making capacity and to test the association between preferences documented in advance directives and outcomes of surrogate decision making. Of 3746 subjects, 42.5% required decision making, of whom 70.3% lacked decision-making capacity and 67.6% of those subjects, in turn, had advance directives. Subjects who had living wills were more likely to want limited care (92.7%) or comfort care (96.2%) than all care possible (1.9%); 83.2% of subjects who requested limited care and 97.1% of subjects who requested comfort care received care consistent with their preferences. Among the 10 subjects who requested all care possible, only 5 received it; however, subjects who requested all care possible were far more likely to receive aggressive care as compared with those who did not request it (adjusted odds ratio, 22.62; 95% confidence interval [CI], 4.45 to 115.00). Subjects with living wills were less likely to receive all care possible (adjusted odds ratio, 0.33; 95% CI, 0.19 to 0.56) than were subjects without living wills. Subjects who had assigned a durable power of attorney for health care were less likely to die in a hospital (adjusted odds ratio, 0.72; 95% CI, 0.55 to 0.93) or receive all care possible (adjusted odds ratio, 0.54; 95% CI, 0.34 to 0.86) than were subjects who had not assigned a durable power of attorney for health care. Between 2000 and 2006, many elderly Americans needed decision making near the end of life at a time when most lacked the capacity to make decisions. Patients who had prepared advance directives received care that was strongly associated with their preferences. These findings support the continued use of advance directives.