Pharmacodynamic model of interleukin-21 effects on red blood cells in cynomolgus monkeys.
Interleukin-21 (IL-21) is a novel cytokine that is currently under clinical investigations as a potential anti-cancer agent. Like many other anti-cancer agents, including other interleukins, IL-21 is seen to produce a broad range of biological effects that may be related to both efficacy and safety of treatment. The present analysis investigates the observed pharmacodynamics effects on red blood cells following various treatment schedules of human IL-21 administrated to cynomolgus monkeys. These effects are described by a novel non-linear mixed-effects model that enabled separation of drug effects and sampling effects, the latter believed to be due partly to blood loss and partly to stress induced haemolysis in connection with blood sampling. Two different studies with a total of 9 different treatment groups of cynomolgus monkeys were used for model development. In conclusion, the model describes the IL-21 induced drop in red blood cells to be (1) caused by removal rather than suppression of production, consistent with increased reticulocyte concentration, and (2) considerably delayed compared to dosing, i.e. not related to the drop in red blood cells observed immediately post dose. It is believed that the structural model presented here can be used for other types of drug induced loss of red blood cells, whereas the mechanism for sampling related blood loss is relevant for investigations of anaemia in all pharmacological studies with smaller animals.


Microdialysis for monitoring the process of functional tissue culture.
Continuous monitoring is important during tissue culture. However, there are still technical difficulties in monitoring the internal status of cells or tissues. In this paper, microdialysis is adopted to monitor functional tissue growth in a bioreactor. Explanted bovine caudal intervertebral disc (IVD) was used as the test tissue. A microdialysis membrane probe of 100 kDa molecular weight cut-off was employed and in situ calibration methods with phenol red and fluorescent 40 kDa dextran were developed to measure the relative recovery of the solute of interest, and membrane fouling, respectively. Tissue metabolism was monitored successfully. At the same time soluble macromolecules were picked up by the probe and were detected and quantified by Fast Protein Liquid Chromatography (FPLC) and/or Sodium Dodecyl Sulphate Polyacrylamide Gel Electrophoresis (SDS-PAGE). These proteins were believed to be associated with biofunction of engineered tissue. Monitoring of phenol red content in the dialysate indicated that there was no significant fouling of the membrane probe during a 7-day culture period and the Relative Recovery of macromolecules of interests remained roughly 9%. We concluded that microdialysis could be used to sample a wide range of molecular species released during cell metabolism and extracellular matrix turnover, which were direct or indirect indications of cell and tissue functions. The application of the developed system could be extended to monitor tissue repair in vivo, and the development of the engineered tissue.


Multiple-channel spectrally encoded imaging.
Spectrally encoded endoscopy (SEE) uses miniature diffractive optics to encode space with wavelength, allowing video-rate three-dimensional imaging through sub-millimeter, flexible endoscopic probes. Here we present a new approach for SEE in which the illumination and the collection channels are separated in space, and spectral encoding is present only in the collection channel. Bench-top experiments using spatially incoherent white light illumination reveal significant improvement in image quality and considerable reduction of speckle noise compared to conventional techniques, and show that the new system is capable of high sensitivity fluorescence imaging of single cells. The presented new approach would allow improved functionality and usability of SEE.


Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR.
The ongoing outbreak of the recently emerged novel coronavirus (2019-nCoV) poses a challenge for public health laboratories as virus isolates are unavailable while there is growing evidence that the outbreak is more widespread than initially thought, and international spread through travellers does already occur. We aimed to develop and deploy robust diagnostic methodology for use in public health laboratory settings without having virus material available. Here we present a validated diagnostic workflow for 2019-nCoV, its design relying on close genetic relatedness of 2019-nCoV with SARS coronavirus, making use of synthetic nucleic acid technology. The workflow reliably detects 2019-nCoV, and further discriminates 2019-nCoV from SARS-CoV. Through coordination between academic and public laboratories, we confirmed assay exclusivity based on 297 original clinical specimens containing a full spectrum of human respiratory viruses. Control material is made available through European Virus Archive - Global (EVAg), a European Union infrastructure project. The present study demonstrates the enormous response capacity achieved through coordination of academic and public laboratories in national and European research networks.


The role of smoking and diet in explaining educational inequalities in lung cancer incidence.
Studies in many countries have reported higher lung cancer incidence and mortality in individuals with lower socioeconomic status. To investigate the role of smoking in these inequalities, we used data from 391,251 participants in the European Prospective Investigation into Cancer and Nutrition study, a cohort of individuals in 10 European countries. We collected information on smoking (history and quantity), fruit and vegetable consumption, and education through questionnaires at study entry and gathered data on lung cancer incidence for a mean of 8.4 years. Socioeconomic status was defined as the highest attained level of education, and participants were grouped by sex and region of residence (Northern Europe, Germany, or Southern Europe). Relative indices of inequality (RIIs) of lung cancer risk unadjusted and adjusted for smoking were estimated using Cox regression models. Additional analyses were performed by histological type. During the study period, 939 men and 692 women developed lung cancer. Inequalities in lung cancer risk (RII(men) = 3.62, 95% confidence interval [CI] = 2.77 to 4.73, 117 vs 52 per 100,000 person-years for lowest vs highest education level; RII(women) = 2.39, 95% CI = 1.77 to 3.21, 46 vs 25 per 100,000 person-years) decreased after adjustment for smoking but remained statistically significant (RII(men) = 2.29, 95% CI = 1.75 to 3.01; RII(women) = 1.59, 95% CI = 1.18 to 2.13). Large RIIs were observed among men and women in Northern European countries and among men in Germany, but inequalities in lung cancer risk were reverse (RIIs < 1) among women in Southern European countries. Inequalities differed by histological type. Adjustment for smoking reduced inequalities similarly for all histological types and among men and women in all regions. In all analysis, further adjustment for fruit and vegetable consumption did not change the estimates. Self-reported smoking consistently explains approximately 50% of the inequalities in lung cancer risk due to differences in education.


Altered carnitine homeostasis is associated with decreased mitochondrial function and altered nitric oxide signaling in lambs with pulmonary hypertension.
Utilizing aortopulmonary vascular graft placement in the fetal lamb, we have developed a model (shunt) of pulmonary hypertension that mimics congenital heart disease with increased pulmonary blood flow. Our previous studies have identified a progressive development of endothelial dysfunction in shunt lambs that is dependent, at least in part, on decreased nitric oxide (NO) signaling. The purpose of this study was to evaluate the possible role of a disruption in carnitine metabolism in shunt lambs and to determine the effect on NO signaling. Our data indicate that at 2 wk of age, shunt lambs have significantly reduced expression (P < 0.05) of the key enzymes in carnitine metabolism: carnitine palmitoyltransferases 1 and 2 as well as carnitine acetyltransferase (CrAT). In addition, we found that CrAT activity was inhibited due to increased nitration. Furthermore, free carnitine levels were significantly decreased whereas acylcarnitine levels were significantly higher in shunt lambs (P < 0.05). We also found that alterations in carnitine metabolism resulted in mitochondrial dysfunction, since shunt lambs had significantly decreased pyruvate, increased lactate, and a reduced pyruvate/lactate ratio. In pulmonary arterial endothelial cells cultured from juvenile lambs, we found that mild uncoupling of the mitochondria led to a decrease in cellular ATP levels and a reduction in both endothelial NO synthase-heat shock protein 90 (eNOS-HSP90) interactions and NO signaling. Similarly, in shunt lambs we found a loss of eNOS-HSP90 interactions that correlated with a progressive decrease in NO signaling. Our data suggest that mitochondrial dysfunction may play a role in the development of endothelial dysfunction and pulmonary hypertension and increased pulmonary blood flow.


Mutations of SPG4 are responsible for a loss of function of spastin, an abundant neuronal protein localized in the nucleus.
Mutations of spastin are responsible for the most common autosomal dominant form of hereditary spastic paraplegia (AD-HSP), a disease characterized by axonal degeneration of corticospinal tracts and posterior columns. Generation of polyclonal antibodies specific to spastin has revealed two isoforms of 75 and 80 kDa in both human and mouse tissues with a tissue-specific variability of the isoform ratio. Spastin is an abundant protein in neural tissues and immunolabeling experiments have shown that spastin is expressed in neurons but not in glial cells. These data indicate that axonal degeneration linked to spastin mutations is caused by a primary defect of neurons. Protein and transcript analyses of patients carrying either nonsense or frameshift spastin mutations revealed neither truncated protein nor mutated transcripts, providing evidence that these mutations are responsible for a loss of spastin function. Identifying agents able to induce the expression of the non-mutated spastin allele should represent an attractive therapeutic strategy in this disease.


The Role of Occlusion in the Dental Implant and Peri-implant Condition: A Review.
Dental implants have become a widely used dental treatment approach. It is important to identify factors that can be detrimental to dental implants and the peri-implant complex. There is controversy regarding whether occlusion plays a role in the implant and peri-implant condition. The present study aims to review the scientific literature regarding this topic. Animal and human studies, and previous reviews on the topic are included and presented. There is a wide heterogeneity among study designs. Several articles demonstrated that occlusion and occlusion overload could detrimentally affect the peri-implant condition, while other articles did not support these results. More studies are needed to help understand the mechanisms by which occlusion might play a role in the peri-implant condition.


Amino acid residue 67 (isoleucine) of HLA-DRB is associated with POHS.
To investigate whether presumed ocular histoplasmosis syndrome (POHS) in The Netherlands is associated with HLA-DR2 and HLA-B7, as previously shown in the United States. Twenty-four Dutch patients with POHS were included in this study. DNA isolated from peripheral blood leukocytes was typed for HLA by a sequence-based method. Associations were statistically determined. The frequencies of HLA alleles in bone marrow of donors listed on the European donor registry was used to represent the distribution in the normal population. Patients were included in the study only when no cells were present in the vitreous at any time and when fundus photographs fit the diagnosis made according to the following criteria: presence of peripapillary atrophy, presence of punched out chorioretinal lesions (histospots), and presence of a submacular scar. After the fundus photographs were judged, the patients were divided into two groups. Group1 contained patients who met all three diagnostic criteria (complete POHS), and group 2 contained patients who met one or two of the criteria (incomplete POHS). Group 1 consisted of 14 patients and group 2 of 10 patients. An association between POHS and HLA-DR2 and -B7 was present, compared with the normal Dutch control subjects. Although significant, the association between the frequency of HLA-DR2 and -B7 of all patients with POHS was less striking than the findings in patients with POHS in the United States. The association with DR2 in patients with incomplete POHS (group 2) was significantly different from that in the group with complete POHS (group 1). According to the defined criteria the association of POHS with HLA-B7 and -DR2 was confined to the incomplete POHS group and was not found in the complete POHS group. Furthermore, analysis of DR at the amino acid level, rather than at the allele level (DR2) showed that amino acid 67 of the DRB1 alleles had the most significant HLA association with POHS, independent of the two groups. POHS in Dutch patients was associated with HLA-B7 and -DR2, but more striking was the presence of isoleucine at position 67 of the HLA-DR molecule.


Current update and future directions on gut microbiome and nephrolithiasis.
The incidence of nephrolithiasis is increasing worldwide. Understanding how gut microbiome influences oxalate homeostasis has the potential to offer new strategies to prevent nephrolithiasis. The literature was reviewed to gather the evidence on the association between gut microbiome, hyperoxaluria and nephrolithiasis, and to identify the therapeutic interventions focused on the gut microbiome that could decrease hyperoxaluria and prevent nephrolithiasis. Gut microbiome is constituted by a plethora of microbiota including Oxalobacter formigenes (Oxf) and lactobacilli. Oxf can degrade dietary oxalate and induce enteral oxalate secretion. Animal studies suggested an association between oral Oxf supplementation and a decrease in hyperoxaluria. However, human studies have showed inconsistent results. Oral supplementation of lactobacilli did not show benefit in decreasing the hyperoxaluria. Antibiotic exposure, by affecting the gut microbiome, has been associated with an increase in nephrolithiasis. In vivo studies suggest fecal transplantation as a potential treatment option for reducing nephrolithiasis, but needs further evaluation in clinical studies. The current evidence suggests an association between gut microbiome and nephrolithiasis. However, the strategies focused on modulating gut microbiome for decreasing hyperoxaluria and preventing nephrolithiasis need further research. Judicious use of antibiotics in those predisposed to nephrolithiasis offers a preventative strategy for decreasing nephrolithiasis.


Microleakage of four Class II resin composite insertion techniques at intraoral temperature.
Twenty recently extracted human molars had standardized Class II slot cavities prepared on the mesial and distal surfaces with the gingival floor located on dentin. The teeth were embedded in a stone template, warmed to 37 degrees C, and restored immediately using one of two incremental placement techniques with visible light-cured composite (VLC), or one of two directed shrinkage placement techniques using an autocure/VLC composite combination. Teeth were stored at 37 degrees C for 2 weeks, thermocycled, stained, sectioned in the middle of the preparation, and evaluated for dye penetration. Reevaluation of the microleakage following removal of the resin composite from the preparation revealed greater microleakage at the gingival margin than did the initial sectioning technique. There were no differences among any of the groups. Enamel margins exhibited minimal leakage and no differences among the groups.


Structural basis of the allosteric inhibitor interaction on the HIV-1 reverse transcriptase RNase H domain.
HIV-1 reverse transcriptase (RT) has been an attractive target for the development of antiretroviral agents. Although this enzyme is bi-functional, having both DNA polymerase and ribonuclease H (RNH) activities, there is no clinically approved inhibitor of the RNH activity. Here, we characterize the structural basis and molecular interaction of an allosteric site inhibitor, BHMP07, with the wild-type (WT) RNH fragment. Solution NMR experiments for inhibitor titration on WT RNH showed relatively wide chemical shift perturbations, suggesting a long-range conformational effect on the inhibitor interaction. Comparisons of the inhibitor-induced NMR chemical shift changes of RNH with those of RNH dimer, in the presence and absence of Mg(2+) , were performed to determine and verify the interaction site. The NMR results, with assistance of molecular docking, indicate that BHMP07 preferentially binds to a site that is located between the RNH active site and the region encompassing helices B and D (the 'substrate-handle region'). The interaction site is consistent with the previous proposed site, identified using a chimeric RNH (p15-EC) [Gong et al. (2011) Chem Biol Drug Des 77, 39-47], but with slight differences that reflect the characteristics of the amino acid sequences in p15-EC compared to the WT RNH.


Therapies for neovascular age-related macular degeneration: current approaches and pharmacologic agents in development.
As one of the leading causes of blindness, age-related macular degeneration (AMD) has remained at the epicenter of clinical research in ophthalmology. During the past decade, focus of researchers has ranged from understanding the role of vascular endothelial growth factor (VEGF) in the angiogenic cascades to developing new therapies for retinal vascular diseases. Anti-VEGF agents such as ranibizumab and aflibercept are becoming increasingly well-established therapies and have replaced earlier approaches such as laser photocoagulation or photodynamic therapy. Many other new therapeutic agents, which are in the early phase clinical trials, have shown promising results. The purpose of this paper is to briefly review the available treatment modalities for neovascular AMD and then focus on promising new therapies that are currently in various stages of development.


Nanoarchitectures for Metal-Organic Framework-Derived Nanoporous Carbons toward Supercapacitor Applications.
The future advances of supercapacitors depend on the development of novel carbon materials with optimized porous structures, high surface area, high conductivity, and high electrochemical stability. Traditionally, nanoporous carbons (NPCs) have been prepared by a variety of methods, such as templated synthesis, carbonization of polymer precursors, physical and chemical activation, etc. Inorganic solid materials such as mesoporous silica and zeolites have been successfully utilized as templates to prepare NPCs. However, the hard-templating methods typically involve several synthetic steps, such as preparation of the original templates, formation of carbon frameworks, and removal of the original templates. Therefore, these methods are not favorable for large-scale production. Metal-organic frameworks (MOFs) with high surface areas and large pore volumes have been studied over the years, and recently, enormous efforts have been made to utilize MOFs for electrochemical applications. However, their low conductivity and poor stability still present major challenges toward their practical applications in supercapacitors. MOFs can be used as precursors for the preparation of NPCs with high porosity. Their parent MOFs can be prepared with endless combinations of organic and inorganic constituents by simple coordination chemistry, and it is possible to control their porous architectures, pore volumes, surface areas, etc. These unique properties of MOF-derived NPCs make them highly attractive for many technological applications. Compared with carbonaceous materials prepared using conventional precursors, MOF-derived carbons have significant advantages in terms of a simple synthesis with inherent diversity affording precise control over porous architectures, pore volumes, and surface areas. In this Account, we will summarize our recent research developments on the preparation of three-dimensional (3-D) MOF-derived carbons for supercapacitor applications. This Account will be divided into three main sections: (1) useful background on carbon materials for supercapacitor applications, (2) the importance of MOF-derived carbons, and (3) potential future developments of MOF-derived carbons for supercapacitors. This Account focuses mostly on carbons derived from two types of MOFs, namely, zeolite imidazolate framework-8 (ZIF-8) and ZIF-67. By using examples from our previous works, we will show the uniqueness of these carbons for achieving high performance by control of the chemical reactions/conditions as well proper utilization in asymmetric/symmetric supercapacitor configurations. This Account will promote further developments of MOF-derived multifunctional carbon materials with controlled porous architectures for optimization of their electrochemical performance toward supercapacitor applications.


T-cell autoimmunity in primary biliary cirrhosis.
1. Primary biliary cirrhosis is a chronic cholestatic liver disease with an autoimmune aetiology. The classical histopathological lesion, of portal tract biliary epithelial cell damage, is accompanied by a T-cell-rich mononuclear cell infiltrate and upregulation of cell surface markers suggestive of local T-cell activation and cytokine release. This suggests that T-cell mediated mechanisms play an important role in tissue damage in primary biliary cirrhosis. 2. CD4+ T-cells specific for the E2 component of human pyruvate dehydrogenase complex (PDC-E2), a highly conserved enzyme which plays a critical role in intermediate metabolism, are present in the peripheral repertoire of the majority of patients with primary biliary cirrhosis. These cells are almost entirely absent from normal and chronic liver disease control subjects. The observations that peripheral blood PDC-E2-specific cells are most commonly seen in early stage disease, when active bile duct damage is occurring, and that PDC-E2- specific cells can be found in the portal tract infiltrate at times when this damage is occurring, suggest that these autoreactive cells may have a role to play in the aetiology of primary biliary cirrhosis. 3. T-cells specific for the whole PDC and its E1 component are seen in significant numbers of normal control subjects as well as patients with primary biliary cirrhosis. Retention of potentially autoreactive cells in the normal T-cell repertoire has been reported for a number of other autoantigens. 4. T-cell epitopes appear to be widely distributed throughout PDC-E2. This is in contrast to the B-cell epitopes which are highly restricted to the inner lipoyl binding domain of the protein.


Why is there variation in test ordering practices for patients presenting to the emergency department with undifferentiated chest pain? A qualitative study.
Up to one-third of laboratory tests ordered in the ED for adults presenting with undifferentiated chest pain are generally not indicated by current Australian guidelines. This study set out to undertake a qualitative investigation of clinician perceptions to identify the reasons for variations in pathology requesting. For this study, we draw on data from semistructured interviews (n=38) conducted in the EDs and laboratories across three hospitals as part of a larger study on the test result management process from test request to result follow-up. Thematic analysis was conducted to determine what aspects of the clinical routines and environment might contribute to variations in pathology requesting. Informed by the findings from the analysis, targeted questions were developed and further focus groups (n=5) were held with clinicians, hospital management and electronic medical record (eMR) analysts to investigate in more detail the reasons for requesting outside of guidelines. Participants cited four main reasons for ordering outside of guidelines. Clinicians requested tests outside of guidelines and the ED scope of practice to facilitate the patient journey along the broader continuum of care, including admission to hospital or transfer to another site. Clinicians were also faced with multiple and inconsistent guidelines regarding appropriate test selection. Limited access to in-house specialty and diagnostic services also influenced ordering patterns in smaller non-referral hospitals. Finally, certain features of the current electronic ordering framework within the eMR facilitated overordering and failed to impose any real restrictions on ordering inappropriately or outside of scope of practice. Beyond the standardisation of pathology requesting advice across electronic decision support, order sets and guidelines, attempts to address issues related to the appropriateness and variation of laboratory test ordering should consider local and systemic factors which also shape the ordering process.


[Public health center based educational approach for families of the mentally disabled in Japan].
In order to evaluate a public health center based educational approach for families of the mentally disabled, questionnaires were sent to all public health centers in Japan. Out of 852 public health centers, 742 responded satisfactorily. At 68.6% of the 742 public health centers, some form of educational programs for families of the mentally disabled have been conducted. In a structured educational program, public health nurses and mental health counselors participated as the main staff. Training seminars for the staff were conducted by prefectural mental health centers or Zenkaren, The National Federation of Families with the Mentally Ill. In order to develop an educational program at the public health centers, the following three elements are required: 1) information transmitted through training seminars and publications, 2) trained staff members including mental health welfare counselors and public health nurses specialized in mental health, and 3) a sufficient budget. Further development of the program is considered crucial for a functional improvement of the public health centers.


The couple's decision-making in IVF: one or two embryos at transfer?
The aim of this study was to evaluate the decision-making process and factors that contribute to the decision of IVF participants to choose one or two embryos at transfer. Two hundred and seventy-four IVF patients equally distributed in males and females were personally interviewed using a semi-structured questionnaire which included 82 items. In the whole study population, previous childbirth [odds ratio (OR) 2.1; 95% confidence interval (CI) 1.9-3.6], and spare embryos to freeze (OR 23.6; 95% CI 11.2-54.5) emerged as the most important variables in patients who had one embryo transferred, while previous IVF treatments (OR 0.3; 95% CI 0.1-0.6) and the assumed increased pregnancy chance (OR 0.1; 95% CI 0.05-0.3) were the most important decision-making factors among those who had two embryos. The women were more satisfied with the information (83 versus 71%; P = 0.02), and more aware of the risks with twin pregnancies (77 versus 66%; P = 0.03) than the males. The women were also more concerned about their age. Knowledge about risks of multiple pregnancies was higher in females (77%) than in males (66%, P = 0.03). The results of this study indicate that despite good information about the risks for complications with multiple pregnancies, many patients wish to have two embryos transferred. Spare embryos to freeze, improvement of pregnancy rate in single embryo transfer and young age of the woman are predictive of choosing single embryo transfer. However, the final decision must always be made in agreement with the physician.


6-C-(E-phenylethenyl)-naringenin suppresses colorectal cancer growth by inhibiting cyclooxygenase-1.
Recent clinical trials raised concerns regarding the cardiovascular toxicity of selective cyclooxygenase-2 (COX-2) inhibitors and cyclooxygenase-1 (COX-1) is now being reconsidered as a target for chemoprevention. Our aims were to determine whether selective COX-1 inhibition could delay or prevent cancer development and also clarify the underlying mechanisms. Data clearly showed that COX-1 was required for maintenance of malignant characteristics of colon cancer cells or tumor promoter-induced transformation of preneoplastic cells. We also successfully applied a ligand-docking computational method to identify a novel selective COX-1 inhibitor, 6-C-(E-phenylethenyl)-naringenin (designated herein as 6CEPN). 6CEPN could bind to COX-1 and specifically inhibited its activity both in vitro and ex vivo. In colorectal cancer cells, it potently suppressed anchorage-independent growth by inhibiting COX-1 activity. 6CEPN also effectively suppressed tumor growth in a 28-day colon cancer xenograft model without any obvious systemic toxicity. Taken together, COX-1 plays a critical role in human colorectal carcinogenesis, and this specific COX-1 inhibitor merits further investigation as a potential preventive agent against colorectal cancer.


[Telemedicine: a novel service in pediatric emergency care].
Telemedicine is a field that deals with monitoring and caring for patients who are physically apart from the caregivers, so they cannot communicate without technological means. The use of telemedicine has been recognized by a number of official organizations worldwide. This article reviews a unique service, "pediatrician online", provided by Clalit HMO, through Medtrix Ltd, with Schneider Children's Hospital specialists' supervision. With this service, parents can receive medical consultation from pediatricians beyond community clinic hours. The service is provided by telephone, computer or a mobile application, and when possible video calls are made. To describe and analyze four years of telemedicine experience of the "pediatrician online" service. A retrospective study was performed including analysis of all consultation calls conducted between November 2009 (the beginning of the project) and December 2013. Every call to the service was recorded and documented. Furthermore, analysis was conducted of follow-up calls to 4% of the applicants in order to monitor medical outcome and satisfaction. During the first four years approximately half a million consultation calls were provided. The average time from referral to obtaining medical advice was eight minutes. The duration of the consultation was four and a half minutes on average. Most of the calls were related to young children under 4 years (74%), dealing with common pediatric problems - fever (23%), respiratory complaints (17%), and gastrointestinal problems (12%). One-third of the applicants intended to go to the emergency department before they called the service, but only about one-sixth were referred at the end of the consultation. Among those referred, only about 60% actually went to the emergency department. The use of video-calls has been increasing, and reached an average of 30% of the applications. Satisfaction with the service was very high according to surveys, scoring 9.8 out of ten. Eighty-two percent of patients sampled reported medical improvement on the next day. There is a very high demand for consulting pediatricians beyond clinic hours. The use of telemedicine, as provided by the "pediatrician online" service, addresses this need safely and effectively with high patient satisfaction. Discussion: The use of telemedicine is expected to expand. In order to ensure its proper use while recognizing the capabilities and limitations, controlled studies should be conducted, exploring the outcome of different approaches in various situations.


Electromyographic reflex responses to mechanical force, manually assisted spinal manipulative therapy.
Surface electromyographic reflex responses associated with mechanical force, manually assisted (MFMA) spinal manipulative therapy were analyzed in this prospective clinical investigation of 20 consecutive patients with low back pain. To characterize and determine the magnitude of electromyographic reflex responses in human paraspinal muscles during high loading rate mechanical force, manually assisted spinal manipulative therapy of the thoracolumbar spine and sacroiliac joints. Spinal manipulative therapy has been investigated for its effectiveness in the treatment of patients with low back pain, but its physiologic mechanisms are not well understood. Noteworthy is the fact that spinal manipulative therapy has been demonstrated to produce consistent reflex responses in the back musculature; however, no study has examined the extent of reflex responses in patients with low back pain. Twenty patients (10 male and 10 female, mean age 43.0 years) underwent standard physical examination on presentation to an outpatient chiropractic clinic. After repeated isometric trunk extension strength tests, short duration (<5 msec), localized posteroanterior manipulative thrusts were delivered to the sacroiliac joints, and L5, L4, L2, T12, and T8 spinous processes and transverse processes. Surface, linear-enveloped electromyographic (sEMG) recordings were obtained from electrodes located bilaterally over the L5 and L3 erector spinae musculature. Force-time and sEMG time histories were recorded simultaneously to quantify the association between spinal manipulative therapy mechanical and electromyographic response. A total of 1600 sEMG recordings were analyzed from 20 spinal manipulative therapy treatments, and comparisons were made between segmental level, segmental contact point (spinous vs. transverse processes), and magnitude of the reflex response (peak-peak [p-p] ratio and relative mean sEMG). Positive sEMG responses were defined as >2.5 p-p baseline sEMG output (>3.5% relative mean sEMG output). SEMG threshold was further assessed for correlation of patient self-reported pain and disability. Consistent, but relatively localized, reflex responses occurred in response to the localized, brief duration MFMA thrusts delivered to the thoracolumbar spine and SI joints. The time to peak tension (sEMG magnitude) ranged from 50 to 200 msec, and the reflex response times ranged from 2 to 4 msec, the latter consistent with intraspinal conduction times. Overall, the 20 treatments produced systematic and significantly different L5 and L3 sEMG responses, particularly for thrusts delivered to the lumbosacral spine. Thrusts applied over the transverse processes produced more positive sEMG responses (25.4%) in comparison with thrusts applied over the spinous processes (20.6%). Left side thrusts and right side thrusts over the transverse processes elicited positive contralateral L5 and L3 sEMG responses. When the data were examined across both treatment level and electrode site (L5 or L3, L or R), 95% of patients showed positive sEMG response to MFMA thrusts. Patients with frequent to constant low back pain symptoms tended to have a more marked sEMG response in comparison with patients with occasional to intermittent low back pain. This is the first study demonstrating neuromuscular reflex responses associated with MFMA spinal manipulative therapy in patients with low back pain. Noteworthy was the finding that such mechanical stimulation of both the paraspinal musculature (transverse processes) and spinous processes produced consistent, generally localized sEMG responses. Identification of neuromuscular characteristics, together with a comprehensive assessment of patient clinical status, may provide for clarification of the significance of spinal manipulative therapy in eliciting putative conservative therapeutic benefits in patients with pain of musculoskeletal origin.


Development and psychometric evaluation of the public health surveillance well-being scale.
To develop and psychometrically evaluate the brief Public Health Surveillance Well-Being Scale (PHS-WB) that captures mental, physical, and social components of well-being. Using data from 5,399 HealthStyles survey respondents, we conducted bi-factor, item response theory, and differential item functioning analyses to examine the psychometric properties of a pool of 34 well-being items. Based on the statistical results and content considerations, we developed a brief 10-item well-being scale and assessed its construct validity through comparisons of demographic subgroups and correlations with measures of related constructs. Based on the bi-factor analyses, the items grouped into both an overall factor and individual domain-specific factors. The PHS-WB scale demonstrated good internal consistency (alpha = 0.87) and correlated highly with scores for the entire item pool (r = 0.94). The well-being scale scores differed as expected across demographic groups and correlated with global and domain-specific measures of similar constructs, supporting its construct validity. The 10-item PHS-WB scale demonstrates good psychometric properties, and its high correlation with the item pool suggests minimal loss of information with the use of fewer items. The brief PHS-WB allows for well-being assessment on national surveys or in other situations where a longer form may not be feasible.


Does the Incidence and Mortality of Pulmonary Thromboembolism Change Over the Years?
In the last 20 years, with the use of computed tomography (CT) angiography, the number of patients diagnosed with pulmonary thromboembolism (PTE) has increased. At the same time, data show that pulmonary embolism mortality has also reduced in this duration. In this study, we analyzed records of patients with PTE (using ICD's) in the hospital automation system from 2001 to 2013. Data regarding age, sex, date of diagnosis, diagnosis of cancer, hemodynamic status, initial and maintenance treatment, hospital length of stay, and hospital mortality were recorded. Primary endpoints of the study were hospital length of stay and all-cause hospital mortality. The total number of patients included in the study was 1185. The median age was 61 years in 2001 and 71 years in 2013. The number of patients who were diagnosed using CT increased from 10% to 92.8%. Between 2001 and 2013, the number of patients diagnosed with PTE increased, and of all patients with PTE, 13.7% was diagnosed in 2009. The hospital length of stay of 13 days declined to 9 days. The use of a vena cava filter in 2007 was 1.1% and that in 2013 was 4.6%. Mortality rate was 15%, however hospital mortality did not significantly differ over the years but varied between 9.4% and 18.8%. Increased use of thrombolytics in patients with massive PTE has been observed over the years. Massive PTE ratio in 2006 was 8.5% and thrombolytic use was 5.8%, however in 2013, these ratios were 2.6%, 6% respectively (p=0.017). Finally, despite the increased number of patients diagnosed with PTE over the years, the mortality rate was not observed to have changed from 2001 to 2013.


Chronic hyperandrogenemia and western-style diet beginning at puberty reduces fertility and increases metabolic dysfunction during pregnancy in young adult, female macaques.
What are the impacts of elevated testosterone (T) and an obesogenic western-style diet (WSD), either independently or together, on fertility and metabolic adaptations of pregnancy in primates? Testosterone increases the time to achieve pregnancy, while a WSD reduces overall fertility, and the combination of testosterone and WSD additionally impairs glucose tolerance and causes pregnancy loss. Both hyperandrogenemia and obesity are hallmarks of polycystic ovary syndrome, which is a leading cause of infertility among women worldwide. Female macaques receiving T and WSD beginning at puberty show increased metabolic, ovarian and uterine dysfunction in the non-pregnant state by 3 years of treatment. The same cohort of female rhesus macaques continued treatments from the time of puberty (2.5 years) to 4 years, including this fertility trial. There were four groups (n = 9-10/group): controls (C), T-treated (T; average total serum level 1.35 ng/ml), WSD-treated, and combined T and WSD-treated (T + WSD) females. Females, which were typically having menstrual cycles, were paired for 4 days with a proven male breeder following the late follicular rise in circulating estradiol (≥100 pg/ml). The presence of sperm in the reproductive tract was used to confirm mating. Animals went through up to three successive rounds of mating until they became pregnant, as confirmed by a rise in circulating mCG during the late luteal phase and ultrasound evidence of a gestational sac at Day 30 post-mating (GD30). Placental vascular parameters were also measured at GD30. Metabolic measurements consisted of fasting levels of blood glucose and insulin at approximately GD30, 60, 90 and 115, as well as an intravenous (iv) glucose tolerance test (GTT) at GD115. While all animals in the C and T groups eventually became pregnant, T-treated females on average had a greater interval to achieve pregnancy (P < 0.05). However, only ~70% of animals in the WSD and T + WSD groups became pregnant (P < 0.004). One pregnancy in T + WSD group resulted in an anembryonic pregnancy which miscarried around GD60, while another T + WSD female conceived with a rare identical twin pregnancy which required cessation due to impending fetal loss at GD106. Thus, the number of viable fetuses was less in the T + WSD group, compared to C, T or WSD. Placental blood volume at GD30 was reduced in all treatments compared to the C group (P < 0.05). Maternal P4 levels were elevated in the WSD (P < 0.03) group and E2 levels were elevated in T + WSD animals (P < 0.05). An increase in serum A4 levels throughout gestation was observed in all groups (P < 0.03) except WSD (P = 0.3). All groups displayed increased insulin resistance with pregnancy, as measured from the ivGTT during pregnancy. However, only the T + WSD group had a significant increase in fasting glucose levels and glucose clearance during the GTT indicating a worsened glucose tolerance. WSD treatment decreased female fetuses third trimester weights, but there was an interaction between WSD and T to increase female fetal weight when normalized to maternal weight. N/A. The small number of pregnancies in the WSD and T + WSD groups hampers the ability to make definitive conclusions on effects during gestation. Also, the high fertility rate in the controls indicates the cohort was at their breeding prime age, which may impair the ability to observe subtle fertility defects. The low number of fetuses used for male and female analysis requires additional studies. The current findings strongly suggest that both hyperandrogenemia and obesity have detrimental effects on fertility and gestation in primates, which may be directly relevant to women with polycystic ovary syndrome. All ONPRC Cores and Units were supported by NIH Grant P51 OD011092 awarded to ONPRC. Research reported in this publication was supported by the Eunice Kennedy Shriver National Institute of Child Health & Human Development (NICHD) of the National Institutes of Health (NIH) under Award Number P50HD071836 (to R.L.S.). The authors have no competing conflict of interests to disclose.


Cardiovascular abnormalities in Folr1 knockout mice and folate rescue.
Periconceptional folic acid supplementation is widely believed to aid in the prevention of neural tube defects (NTDs), orofacial clefts, and congenital heart defects. Folate-binding proteins or receptors serve to bind folic acid and 5-methyltetrahydrofolate, representing one of the two major mechanisms of cellular folate uptake. We herein describe abnormal cardiovascular development in mouse fetuses lacking a functional folate-binding protein gene (Folr1). We also performed a dose-response study with folinic acid and determined the impact of maternal folate supplementation on Folr1 nullizygous cardiac development. Partially rescued preterm Folr1(-/-) (formerly referred to as Folbp1) fetuses were found to have outflow tract defects, aortic arch artery abnormalities, and isolated dextracardia. Maternal supplementation with folinic acid rescued the embryonic lethality and the observed cardiovascular phenotypes in a dose-dependant manner. Maternal genotype exhibited significant impact on the rescue efficiency, suggesting an important role of in utero folate status in embryonic development. Abnormal heart looping was observed during early development of Folr1(-/-) embryos partially rescued by maternal folinic acid supplementation. Migration pattern of cardiac neural crest cells, genetic signals in pharyngeal arches, and the secondary heart field were also found to be affected in the mutant embryos. Our observations suggest that the beneficial effect of folic acid for congenital heart defects might be mediated via its impact on neural crest cells and by gene regulation of signaling pathways involved in the development of the pharyngeal arches and the secondary heart field.


Combination strategy targeting the hypoxia inducible factor-1 alpha with mammalian target of rapamycin and histone deacetylase inhibitors.
The hypoxia-inducible factor-1 alpha (HIF-alpha) is a key regulator of tumor angiogenesis. Mammalian target of rapamycin (mTOR) and histone deacetylase (HDAC) inhibitors suppress tumor-induced angiogenesis by reducing tumor HIF-1 alpha protein expression. Thus, we hypothesized that combination treatment of rapamycin and the HDAC inhibitor LBH589 has greater antiangiogenic and antitumor activity compared with single agents. To evaluate the effect of LBH589 and rapamycin on HIF-1 alpha in human prostate PC3, renal C2 carcinoma cell lines, and endothelial cells (human umbilical vein endothelial cells), we did Western blot analysis. To determine the antitumor activity of LBH589 and rapamycin, cell proliferation assays and xenograft experiments were conducted. Western blotting showed that combination treatment of human umbilical vein endothelial cells, C2 and PC3, significantly reduced HIF-1 alpha protein expression compared with single agents. Treatment with rapamycin resulted in inhibition of the downstream signals of the mTOR pathway and increased phosphorylation of Akt in C2 cells, whereas the constitutively activated Akt in PC3 cells was not modulated. LBH589 decreased both constitutively expressed and rapamycin-induced phosphorylated Akt levels in PC3 and C2 cell lines. In clonogenic assays, the combination treatment had a greater inhibitory effect in PC3 cells (93 +/- 1.4%) compared with single agents (66 +/- 9% rapamycin and 43 +/- 4% LBH589). Combination of rapamycin and LBH589 significantly inhibited PC3 and C2 in vivo tumor growth and angiogenesis as measured by tumor weight and microvessel density. Combination treatment of mTOR and HDAC inhibitors represents a rational therapeutic strategy targeting HIF-1 alpha that warrants clinical testing.


Incidence of morbidity from penetrating palate trauma.
The true rate of neurologic sequelae and infection from penetrating palatal trauma in children is unknown, which leads to significant variation in testing and treatment. To (1) determine the incidence of stroke and infection in well-appearing children with penetrating palatal trauma and (2) describe patterns of testing and treatment for uncomplicated palatal trauma. We assembled a retrospective cohort of children aged 9 months to 18 years with palatal trauma seen in the emergency department (ED) at a tertiary care pediatric hospital. Patients met the following definition: well-appearing with normal neurologic examination and a palate laceration but no findings requiring immediate operative care. Stroke was defined as any abnormal neurologic examination secondary to palatal trauma. Infection was defined as cellulitis or abscess secondary to palatal injury. All abnormal computed tomographic angiography (CTA) findings, except for free air, were considered positive and potentially significant. We identified 1656 potential subjects. A total of 995 of 1656 subjects were screened, and 205 of 995 met the case definition. A total of 122 of 205 had follow-up through at least 1 week after injury. The incidence of stroke in our study population was 0% (95% confidence interval [CI]: 0-2.5). One of 116 patients developed infection, for an incidence of 0.9% (95% CI: 0-5.3). A total of 90 of 205 (44%) subjects had CTA scans; the results of 9 (10%) were positive. No patients with positive CTA findings required operative care. No patients received anticoagulant medications. The incidence of morbidity from penetrating palatal trauma in the well-appearing child is extremely low. Diagnostic evaluation in the ED did not prompt clinical interventions other than antibiotics.


Inverse correlation between promoter strength and excision activity in class 1 integrons.
Class 1 integrons are widespread genetic elements that allow bacteria to capture and express gene cassettes that are usually promoterless. These integrons play a major role in the dissemination of antibiotic resistance among Gram-negative bacteria. They typically consist of a gene (intI) encoding an integrase (that catalyzes the gene cassette movement by site-specific recombination), a recombination site (attI1), and a promoter (Pc) responsible for the expression of inserted gene cassettes. The Pc promoter can occasionally be combined with a second promoter designated P2, and several Pc variants with different strengths have been described, although their relative distribution is not known. The Pc promoter in class 1 integrons is located within the intI1 coding sequence. The Pc polymorphism affects the amino acid sequence of IntI1 and the effect of this feature on the integrase recombination activity has not previously been investigated. We therefore conducted an extensive in silico study of class 1 integron sequences in order to assess the distribution of Pc variants. We also measured these promoters' strength by means of transcriptional reporter gene fusion experiments and estimated the excision and integration activities of the different IntI1 variants. We found that there are currently 13 Pc variants, leading to 10 IntI1 variants, that have a highly uneven distribution. There are five main Pc-P2 combinations, corresponding to five promoter strengths, and three main integrases displaying similar integration activity but very different excision efficiency. Promoter strength correlates with integrase excision activity: the weaker the promoter, the stronger the integrase. The tight relationship between the aptitude of class 1 integrons to recombine cassettes and express gene cassettes may be a key to understanding the short-term evolution of integrons. Dissemination of integron-driven drug resistance is therefore more complex than previously thought.


Measurement of natriuretic peptides at the point of care in the emergency and ambulatory setting: current status and future perspectives.
The measurement of natriuretic peptides (NPs), B-type NP or N-terminal pro-B-type NP, can be an important tool in the diagnosis of acute heart failure in patients presenting to an Emergency Department (ED) with acute dyspnea, according to international guidelines. Studies and subsequent meta-analyses are mixed on the absolute value of routine NP assessment of ED patients. However, levels of NPs are likely to be used also to guide treatment and to assess risk of adverse outcomes in other patients at risk of developing heart failure, including those with pulmonary embolism or diabetes, or receiving chemotherapy. Natriuretic peptide levels, like other biomarkers, can now be measured at the point of care (POC). We have reviewed the current status of NP measurement together with the potential contribution of POC measurement of NPs to clinical care delivery in the emergency and other settings. Several POC systems for measuring NP levels are now available: these produce test results within 15 minutes and appear sufficiently sensitive and robust to be used routinely in diagnostic evaluations. Point-of-care systems could be used to assess NP levels in the ED and community outpatient settings to monitor the risk of acute heart failure. Furthermore, the use of protocol-driven POC testing of NP within the time frame of a patient consultation in the ED may facilitate and accelerate the throughput and disposition of at-risk patients. Appropriately designed clinical trials will be needed to confirm these potential benefits. It is also important that processes of care delivery are redesigned to take full advantage of the faster turnaround times provided by POC technology.


Dual role of isocitrate lyase 1 in the glyoxylate and methylcitrate cycles in Mycobacterium tuberculosis.
The role of isocitrate lyase (ICL) in the glyoxylate cycle and its necessity for persistence and virulence of Mycobacterium tuberculosis has been well described. Recent reports have alluded to an additional role for this enzyme in M. tuberculosis metabolism, specifically for growth on propionate. A product of beta-oxidation of odd-chain fatty acids is propionyl-CoA. Clearance of propionyl-CoA and the by-products of its metabolism via the methylcitrate cycle is vital due to their potentially toxic effects. Although the genome of M. tuberculosis encodes orthologues of two of the three enzymes of the methylcitrate cycle, methylcitrate synthase and methylcitrate dehydratase, it does not appear to contain a distinct 2-methylisocitrate lyase (MCL). Detailed structural analysis of the MCL from Escherichia coli suggested that the differences in substrate specificity between MCLs and ICLs could be attributed to three conserved amino acid substitutions in the active site, suggesting an MCL signature. However, here we provide enzymatic evidence that shows that despite the absence of the MCL signature, ICL1 from M. tuberculosis can clearly function as a MCL. Furthermore, the crystal structure of ICL1 with pyruvate and succinate bound demonstrates that the active site can accommodate the additional methyl group without significant changes to the structure.


Selective functional vulnerability of cortical neurons following transient MCA-occlusion in the cat.
Simultaneous recordings of several cortical neurons were obtained before, during and after transient 15 min occlusion of the middle cerebral artery in cats. With the use of a multiple electrode array consisting of 4-7 platinum/iridium microelectrodes, the cortical pericellular blood flow was concurrently measured by means of the hydrogen clearance technique. Hydrogen clearance measurements revealed a homogeneous blood flow distribution throughout all phases of the experiment in the area covered by the different microelectrodes. Considering only the results of experiments with low residual blood flow during ischemia (less than 0.1 ml/g/min), single unit activity ceased immediately after occlusion and remained so during the ischemic period. The recovery time of action potentials after reperfusion ranged from 10 min to 3 hours depending on the examined neuron. Lower values for discharge rates of the individual cells were generally observed after reoccurrence, although some units exhibited temporarily an even higher spike frequency. Furthermore, the spike form usually changed in that the hyperpolarizing afterpotentials were enlarged after recirculation. However, some cells with a nearly unchanged spike form were found as well. The results indicate that the recovery of cell function largely depends on the individual neuron which supports the idea of a selective functional vulnerability of cortical neurons in response to ischemia.


Motion Tracking System for Robust Non-Contact Blood Perfusion Sensor.
We propose a motion-robust laser Doppler flowmetry (LDF) system that can be used as a non-contact blood perfusion sensor for medical diagnosis. Endoscopic LDF systems are typically limited in their usefulness in clinical contexts by the need for the natural organs to be immobilized, as serious motion artifacts due to the axial surface displacement can interfere with blood perfusion measurements. In our system, the focusing lens moves to track the motion of the target using a low-frequency reference signal in the optical data, enabling the suppression of these motion artifacts in the axial direction. This paper reports feasibility tests on a prototype of this system using a microfluidic phantom as a measurement target moving in the direction of the optical axis. The frequency spectra detected and the perfusion values calculated from those spectra show that the motion tracking system is capable of suppressing motion artifacts in perfusion readings. We compared the prototype LDF system's measurements with and without motion feedback, and found that motion tracking improves the fidelity of the perfusion signal by as much as 87%.


A Study of Practice Behavior for Endotracheal Intubation Site for Children With Congenital Heart Disease Undergoing Surgery: Impact of Endotracheal Intubation Site on Perioperative Outcomes-An Analysis of the Society of Thoracic Surgeons Congenital Cardiac Anesthesia Society Database.
In adults undergoing cardiopulmonary bypass surgery, oral intubation is typically preferred over nasal intubation due to reduced risk of sinusitis and infection. In children, nasal intubation is more common and sometimes preferred due to perceived benefits of less postoperative sedation and a lower risk for accidental extubation. This study sought to describe the practice of nasal intubation in the pediatric population undergoing cardiopulmonary bypass surgery and assess the risks/benefits of a nasal route against an oral one. Patients <18 years of age in the Society of Thoracic Surgeons Congenital Heart Surgery Database between January 2010 and December 2015 were included. Patients with a preoperative endotracheal tube, tracheostomy, or known airway anomalies were excluded. Multivariable modeling was used to assess the association between route of tracheal intubation and a composite measure of infection risk (wound infection, mediastinitis, septicemia, pneumonia, and endocarditis). Covariates were included to adjust for important patient characteristics (eg, weight, age, comorbidities), case complexity, and center effects. Secondary outcomes included length of intubation, hospital length of stay, and airway complications including accidental extubations. We also performed a subanalysis in children <12 months of age in high-volume centers (>100 cases/y) examining how infection risk may change with age at the time of surgery. Nasal intubation was used in 41% of operations in neonates, 38% in infants, 15% in school-aged children, and 2% in adolescents. Nasal intubation appeared protective for accidental extubation only in neonates (P = .02). Multivariable analysis in infants and neonates showed that the nasal route of intubation was not associated with the infection composite (relative risk [RR], 0.84; 95% CI, 0.59-1.18) or a shorter length of stay (RR, 0.992; 95% CI, 0.947-1.039), but was associated with a shorter intubation length (RR, 0.929; 95% CI, 0.869-0.992). Restricting to high-volume centers showed a significant interaction between age and intubation route with a risk change for infection occurring between approximately 6-12 months of age (P = .003). While older children undergoing nasal intubation trend similar to the adult population with an increased risk of infection, nasal intubation in neonates and infants does not appear to carry a similar risk. Nasal intubation in neonates and infants may also be associated with a shorter intubation length but not a shorter length of stay. Prospective studies are required to better understand these complex associations.


Bulged-out nucleotides in an antisense RNA are required for rapid target RNA binding in vitro and inhibition in vivo.
Naturally occurring antisense RNAs in prokaryotes are generally short, highly structured and untranslated. Stem-loops are always present, and loop regions serve as primary recognition structures in most cases. Single-stranded tails or internal unstructured regions are required for initiation of stable pairing between antisense and target RNA. Most antisense RNAs contain bulged-out nucleotides or small internal loops in upper stem regions. Here we investigated the role of the bulged-out nucleotides of CopA (the copy number regulator of plasmid R1) in determining the binding properties of this antisense RNA to its target in vitro and the efficiency of a translational inhibition in vivo. The introduction of perfect helicity in the region of the two bulges in CopA decreased pairing rate constants by up to 180-fold, increased equilibrium dissociation constants of the 'kissing intermediate' up to 14-fold, and severely impaired inhibition of repA expression. A previously described loop size mutant of CopA showed decreased pairing rates, but, in contrast to the bulge-less mutant CopAs, shows a decreased dissociation constant of the 'kissing complex'. We conclude that removal of the specific bulges/internal loops within the stem-loop II of CopA impairs the inhibitor, and that creation of an internal loop at a different position does not restore activity, emphasizing the optimal folding of wild-type CopA. The accompanying paper shows that an additional function of bulges can be protection from RNase III cleavage.


The use of focus groups in evaluating quality of life components among elderly Chinese people.
In Taiwan, to measure the quality of life (QOL) of elderly Chinese, one must rely on instruments developed in other Chinese or Western populations and not specifically for the elderly. The purpose of this study is to understand the components of QOL for elderly Chinese from Taiwan living in residential homes or in their communities. Forty-four elderly men and women divided into six focus groups were interviewed on video tape and the resultant recording was analyzed qualitatively by six independent researchers. The study yielded 15 QOL domains grouped into six dimensions: physical health (physical well-being, impact of illness, medical care), psychological health (mood states, life attitude and retrospection, philosophy of living, self-efficacy), social function (connectedness, exercise and leisure activities, social activities and services), living environment (living environment and arrangements, institutional factors), economic status, and religion and death (religion, death). For elderly Chinese in Taiwan, positive and negative life domains are equally important in the perception of life quality; person-environment interaction is a major consideration in the evaluation of QOL; family ties are an important component of QOL; traditional Chinese beliefs exert a positive influence on perceived QOL; and social functioning and vitality have a different meaning in Chinese compared to Western cultures.


Improvement in delivery of type 2 diabetes services differs by mode of care: a retrospective longitudinal analysis in the Aboriginal and Torres Strait Islander Primary Health Care setting.
Addressing evidence-practice gaps in primary care remains a significant public health challenge and is likely to require action at different levels of the health system. Whilst Continuous Quality Improvement (CQI) is associated with improvements in overall delivery, little is known about delivery of different types of care processes, and their relative improvement during CQI. We used data from over 15,000 clinical audit records of clients with Type 2 diabetes collected as part of a wide-scale CQI program implemented between 2005 and 2014 in 162 Aboriginal and Torres Strait Islander health centres. We abstracted data from clinical records on 15 service items recommended in clinical guidelines and categorised these items into five modes of care on the basis of the mechanism through which care is delivered: laboratory tests; generalist-delivered physical checks; specialist-delivered checks; education/counselling for nutrition and physical activity and education/counselling for high risk substance use. We calculated delivery for each patient for each of mode of care by determining the proportion of recommended services delivered for that mode. We used multilevel regression models to quantify variation attributable to health centre or client level factors and to identify factors associated with greater adherence to clinical guidelines for each mode of care. Clients on average received 43 to 60 % of recommended care in 2005/6. Different modes of care showed different patterns of improvement. Generalist-delivered physical checks (delivered by a non-specialist) showed a steady year on year increase, delivery of laboratory tests showed improvement only in the later years of the study, and delivery of counselling/education interventions showed early improvement which then plateaued. Health centres participating in CQI had increased odds of top quartile service delivery for all modes compared to baseline, but effects differed by mode. Health centre factors explained 20-52 % of the variation across jurisdictions and health centres for different modes of care. Levels of adherence to clinical guidelines and patterns of improvement during participation in a CQI program differed for different modes of care. Policy and funding decisions may have had important effects on the level and nature of improvements achieved.


Tumor-specific immunity induced by cryoablation in a murine renal cell carcinoma model.
To evaluate tumor-specific immunity and define the mechanisms involved in the cryoimmunologic response, we compared the tumor control efficacy and immunologic responses of cryoablation with those of surgical excision in a tumor rechallenge model. Sixty BALB/c mice with RENCA tumors that were generated in the left flank area underwent cryoablation or radical excision. The mice successfully treated were rechallenged with RENCA or an undifferentiated colon carcinoma cell line, CT26, in the contralateral right flank area. The recurrence rate after tumor rechallenge in each group was then observed. To assess the immunologic response of each treatment modality, fluorescent-activated cell sorting (FACS) analysis and a cytotoxicity assay using (51)Cr release were performed. After reinoculation of the RENCA cells, the rate of tumor growth was significantly higher in the surgical excision group than in the cryoablation group (94.4% vs. 11.1%, p=0.001). In the cryoablation group, the tumor growth rate was significantly increased after rechallenge of CT26 cells compared with RENCA (94.1% vs. 11.1%, p=0.001). The cryoablation group showed an elevated CD3, CD4, CD8 T, and natural killer cell count in the FACS analysis and also showed significantly increased cytotoxicity in the (51)Cr release assay compared with the excision group. These results showed that cryoablation, compared to surgical resection, was more effective in preventing tumor growth after rechallenge with RENCA cells and that this response was tumor-specific, because the CT26 cells did not have the same effect.


Maximizing Iodine Contrast-to-Noise Ratios in Abdominal CT Imaging through Use of Energy Domain Noise Reduction and Virtual Monoenergetic Dual-Energy CT.
To determine the iodine contrast-to-noise ratio (CNR) for abdominal computed tomography (CT) when using energy domain noise reduction and virtual monoenergetic dual-energy (DE) CT images and to compare the CNR to that attained with single-energy CT at 80, 100, 120, and 140 kV. This HIPAA-compliant study was approved by the institutional review board with waiver of informed consent. A syringe filled with diluted iodine contrast material was placed into 30-, 35-, and 45-cm-wide water phantoms and scanned with a dual-source CT scanner in both DE and single-energy modes with matched scanner output. Virtual monoenergetic images were generated, with energies ranging from 40 to 110 keV in 10-keV steps. A previously developed energy domain noise reduction algorithm was applied to reduce image noise by exploiting information redundancies in the energy domain. Image noise and iodine CNR were calculated. To show the potential clinical benefit of this technique, it was retrospectively applied to a clinical DE CT study of the liver in a 59-year-old male patient by using conventional and iterative reconstruction techniques. Image noise and CNR were compared for virtual monoenergetic images with and without energy domain noise reduction at each virtual monoenergetic energy (in kiloelectron volts) and phantom size by using a paired t test. CNR of virtual monoenergetic images was also compared with that of single-energy images acquired with 80, 100, 120, and 140 kV. Noise reduction of up to 59% (28.7 of 65.7) was achieved for DE virtual monoenergetic images by using an energy domain noise reduction technique. For the commercial virtual monoenergetic images, the maximum iodine CNR was achieved at 70 keV and was 18.6, 16.6, and 10.8 for the 30-, 35-, and 45-cm phantoms. After energy domain noise reduction, maximum iodine CNR was achieved at 40 keV and increased to 30.6, 25.4, and 16.5. These CNRs represented improvement of up to 64% (12.0 of 18.6) with the energy domain noise reduction technique. For single-energy CT at the optimal tube potential, iodine CNR was 29.1 (80 kV), 21.2 (80 kV), and 11.5 (100 kV). For patient images, 39% (24 of 61) noise reduction and 67% (0.74 of 1.10) CNR improvement were observed with the energy domain noise reduction technique when compared with standard filtered back-projection images. Iodine CNR for adult abdominal CT may be maximized with energy domain noise reduction and virtual monoenergetic DE CT.


Quantitative localisation of synaptic and extrasynaptic GABAA receptor subunits on hippocampal pyramidal cells by freeze-fracture replica immunolabelling.
Hippocampal CA1 pyramidal cells, which receive γ-aminobutyric acid (GABA)ergic input from at least 18 types of presynaptic neuron, express 14 subunits of the pentameric GABA(A) receptor. The relative contribution of any subunit to synaptic and extrasynaptic receptors influences the dynamics of GABA and drug actions. Synaptic receptors mediate phasic GABA-evoked conductance and extrasynaptic receptors contribute to a tonic conductance. We used freeze-fracture replica-immunogold labelling, a sensitive quantitative immunocytochemical method, to detect synaptic and extrasynaptic pools of the alpha1, alpha2 and beta3 subunits. Antibodies to the cytoplasmic loop of the subunits showed immunogold particles concentrated on distinct clusters of intramembrane particles (IMPs) on the cytoplasmic face of the plasma membrane on the somata, dendrites and axon initial segments, with an abrupt decrease in labelling at the edge of the IMP cluster. Neuroligin-2, a GABAergic synapse-specific adhesion molecule, co-labels all beta3 subunit-rich IMP clusters, therefore we considered them synapses. Double-labelling for two subunits showed that virtually all somatic synapses contain the alpha1, alpha2 and beta3 subunits. The extrasynaptic plasma membrane of the somata, dendrites and dendritic spines showed low-density immunolabelling. Synaptic labelling densities on somata for the alpha1, alpha2 and beta3 subunits were 78-132, 94 and 79 times higher than on the extrasynaptic membranes, respectively. As GABAergic synapses occupy 0.72% of the soma surface, the fraction of synaptic labelling was 33-48 (alpha1), 40 (alpha2) and 36 (beta3)% of the total somatic surface immunolabelling. Assuming similar antibody access to all receptors, about 60% of these subunits are in extrasynaptic receptors.


Theoretical stability maps for guiding preparation of emulsions stabilized by protein-polysaccharide interfacial complexes.
The purpose of this study was to evaluate the usefulness of a simple theoretical model at predicting the stability of emulsions containing lipid droplets and polyelectrolytes. The influence of droplet concentration, mean droplet diameter, droplet charge and polyelectrolyte concentration on the aggregation stability of the emulsions was examined. Emulsions stabilized by a globular protein (beta-lactoglobulin) were prepared with different oil droplet (0.5-10 wt %) and pectin (0-0.5 wt %) concentrations at pH 7 (where lipid droplets and pectin molecules were both anionic) and pH 3.5 (where lipid droplets were cationic and pectin molecules anionic). The particle charge, size, and creaming stability of the emulsions were then measured, and stability maps were constructed at pH 3.5 and 7. At pH 7, there was no evidence of pectin adsorption to droplet surfaces, and the emulsions were stable to bridging flocculation, but depletion flocculation occurred when the pectin concentration exceeded about 0.1 wt % (independent of droplet concentration). At pH 3.5, pectin adsorbed to the droplet surfaces, and the emulsions were unstable to bridging flocculation at intermediate pectin concentrations (dependent on droplet concentration) and unstable to depletion flocculation at high pectin concentrations. At certain droplet and pectin concentrations, stable emulsions could be formed consisting of protein-coated lipid droplets surrounded by a pectin layer. The information gained from this study would be useful for optimizing the production of emulsions stabilized by protein-polysaccharide interfacial complexes.


Effects of caffeine and stress on biomarkers of cardiovascular disease in healthy men and women with a family history of hypertension.
The connection between caffeine and its potentially detrimental effects on blood markers of cardiovascular disease (CVD) are controversial. Most studies have focused on cholesterol as a putative mediator of the caffeine-CVD relationship. Other blood markers such as C-reactive protein (CRP) and fibrinogen have been understudied. We examined the effects of caffeine and psychological stress on these CVD markers in healthy, young men and women with a confirmed family history of hypertension. A total of 52 normotensive, healthy adults (26 men and 26 women) aged 18-29 years (21.4 ± 0.3) participated in a laboratory session to examine stress reactivity following caffeine consumption. All participants had normal cholesterol levels. Blood pressure (BP), heart rate, serum cortisol and CRP and plasma fibrinogen were collected. Men and women administered caffeine displayed an additional increase in systolic BP and cortisol response to the stressor (p < 0.05). Stress interacted with caffeine and sex to alter cortisol, fibrinogen and systolic BP but not CRP levels. These results may shed light on sex-specific pathways that associate caffeine with CVD.


A preliminary analysis of the DNA and diet of the extinct Beothuk: a systematic approach to ancient human DNA.
We have used a systematic protocol for extracting, quantitating, sexing and validating ancient human mitochondrial and nuclear DNA of one male and one female Beothuk, a Native American population from Newfoundland, which became extinct approximately 180 years ago. They carried mtDNA haplotypes, which fall within haplogroups X and C, consistent with Northeastern Native populations today. In addition we have sexed the male using a novel-sexing assay and confirmed the authenticity of his Y chromosome with the presence of the Native American specific Y-QM3 single nucleotide polymorphism (SNP). This is the first ancient nuclear SNP typed from a Native population in the Americas. In addition, using the same teeth we conducted a stable isotopes analysis of collagen and dentine to show that both individuals relied on marine sources (fresh and salt water fish, seals) with no hierarchy seen between them, and that their water sources were pooled or stored water. Both mtDNA sequence data and Y SNP data hint at possible gene flow or a common ancestral population for both the Beothuk and the current day Mikmaq, but more importantly the data do not lend credence to the proposed idea that the Beothuk (specifically, Nonosabasut) were of admixed (European-Native American) descent. We also analyzed patterns of DNA damage in the clones of authentic mtDNA sequences; there is no tendency for DNA damage to occur preferentially at previously defined mutational hotspots, suggesting that such mutational hotspots are not hypervariable because they are more prone to damage.


Treatment of bilateral refractive amblyopia in children three to less than 10 years of age.
To determine the amount and time course of binocular visual acuity improvement during treatment of bilateral refractive amblyopia in children three to less than 10 years of age. Prospective, multicenter, noncomparative intervention. One hundred and thirteen children (mean age, 5.1 years) with previously untreated bilateral refractive amblyopia were enrolled at 27 community- and university-based sites and were provided with optimal spectacle correction. Bilateral refractive amblyopia was defined as 20/40 to 20/400 best-corrected binocular visual acuity in the presence of 4.00 diopters (D) or more of hypermetropia by spherical equivalent, 2.00 D or more of astigmatism, or both in each eye. Best-corrected binocular and monocular visual acuities were measured at baseline and at five, 13, 26, and 52 weeks. The primary study outcome was binocular acuity at one year. Mean binocular visual acuity improved from 0.50 logarithm of the minimum angle of resolution (logMAR) units (20/63) at baseline to 0.11 logMAR units (20/25) at one year (mean improvement, 3.9 lines; 95% confidence interval [CI], 3.5 to 4.2). Mean improvement at one year for the 84 children with baseline binocular acuity of 20/40 to 20/80 was 3.4 lines (95% CI, 3.2 to 3.7) and for the 16 children with baseline binocular acuity of 20/100 to 20/320 was 6.3 lines (95% CI, 5.1 to 7.5). The cumulative probability of binocular visual acuity of 20/25 or better was 21% at five weeks, 46% at 13 weeks, 59% at 26 weeks, and 74% at 52 weeks. Treatment of bilateral refractive amblyopia with spectacle correction improves binocular visual acuity in children three to less than 10 years of age, with most improving to 20/25 or better within one year.


Effect of freeboard extension on co-combustion of coal and olive cake in a fluidized bed combustor.
In this study, flue gas emissions and combustion efficiencies during combustion and co-combustion of olive cake and coal were investigated in a bubbling fluidized bed. Temperature distributions along the combustion column and flue gas concentrations of O(2), CO, SO(2) and NO(x) were measured during combustion experiments. Two sets of experiments were performed to examine the effect of fuel composition, excess air ratio and freeboard extension on flue gas emissions and combustion efficiency. The results of the experiments showed that coal combustion occurs at lower parts of the combustion column whereas olive cake combustion takes place more in the freeboard region. As olive cake percentage in the fuel mixture increased, CO emissions increased, SO(2) and NO(x) emissions decreased. Additionally, flue gas emissions could be lowered with the freeboard extension while burning biomass or biomass/coal mixtures. Noticeable decrease in CO emissions and slight increase in combustion efficiencies were observed with a column height of 1900 mm instead of 900 mm.


LKB1 deletion causes early changes in atrial channel expression and electrophysiology prior to atrial fibrillation.
Liver kinase B1 (LKB1) is a protein kinase that activates the metabolic regulator AMP-activated protein kinase (AMPK) and other related kinases. Deletion of LKB1 in mice leads to cardiomyopathy and atrial fibrillation (AF). However, the specific role of the LKB1 pathway in early atrial biology remains unknown. Thus, we investigated whether LKB1 deletion altered atrial channel expression and electrophysiological function in a cardiomyocyte-specific knockout mouse model. We performed a systematic comparison of αMHC-Cre LKB1(fl/fl) and littermate LKB1(fl/fl) male mice. This included analysis of gene expression, histology, and echocardiography, as well as cellular and tissue-level electrophysiology using patch-clamp recordings in vitro, optical mapping ex vivo, and ECG recordings in vivo. At postnatal day 1, atrial depolarization was prolonged, and Nav1.5 and Cx40 expression were markedly down-regulated in MHC-Cre LKB1(fl/fl) mice. Inward sodium current density was significantly decreased in MHC-Cre LKB1(fl/fl) neonatal atrial myocytes. Subsequently, additional alterations in atrial channel expression, atrial fibrosis, and spontaneous onset of AF developed by 2 weeks of age. In adult mice, abnormalities of interatrial conduction and bi-atrial electrical coupling were observed, likely promoting the perpetuation of AF. Mice with AMPK-inactivated hearts demonstrated modest overlap in channel expression with MHC-Cre LKB1(fl/fl) hearts, but retained normal structure, electrophysiological function and contractility. Deletion of LKB1 causes early defects in atrial channel expression, action potential generation and conduction, which precede widespread atrial remodelling, fibrosis and AF. LKB1 is critical for normal atrial growth and electrophysiological function.


Evaluation of ERG and SPINK1 by Immunohistochemical Staining and Clinicopathological Outcomes in a Multi-Institutional Radical Prostatectomy Cohort of 1067 Patients.
Distinguishing between patients with early stage, screen detected prostate cancer who must be treated from those that can be safely watched has become a major issue in prostate cancer care. Identification of molecular subtypes of prostate cancer has opened the opportunity for testing whether biomarkers that characterize these subtypes can be used as biomarkers of prognosis. Two established molecular subtypes are identified by high expression of the ERG oncoprotein, due to structural DNA alterations that encode for fusion transcripts in approximately ½ of prostate cancers, and over-expression of SPINK1, which is purportedly found only in ERG-negative tumors. We used a multi-institutional prostate cancer tissue microarray constructed from radical prostatectomy samples with associated detailed clinical data and with rigorous selection of recurrent and non-recurrent cases to test the prognostic value of immunohistochemistry staining results for the ERG and SPINK1 proteins. In univariate analysis, ERG positive cases (419/1067; 39%) were associated with lower patient age, pre-operative serum PSA levels, lower Gleason scores (≤ 3+4=7) and improved recurrence free survival (RFS). On multivariate analysis, ERG status was not correlated with RFS, disease specific survival (DSS) or overall survival (OS). High-level SPINK1 protein expression (33/1067 cases; 3%) was associated with improved RFS on univariate and multivariate Cox regression analysis. Over-expression of either protein was not associated with clinical outcome. While expression of ERG and SPINK1 proteins was inversely correlated, it was not mutually exclusive since 3 (0.28%) cases showed high expression of both. While ERG and SPINK1 appear to identify discrete molecular subtypes of prostate cancer, only high expression of SPINK1 was associated with improved clinical outcome. However, by themselves, neither ERG nor SPINK1 appear to be useful biomarkers for prognostication of early stage prostate cancer.


Anomericity of T-2 toxin-glucoside: masked mycotoxin in cereal crops.
T-2 toxin is a trichothecene mycotoxin produced when Fusarium fungi infect grains, especially oats and wheat. Ingestion of T-2 toxin contaminated grain can cause diarrhea, hemorrhaging, and feed refusal in livestock. Cereal crops infected with mycotoxin-producing fungi form toxin glycosides, sometimes called masked mycotoxins, which are a potential food safety concern because they are not detectable by standard approaches and may be converted back to the parent toxin during digestion or food processing. The work reported here addresses four aspects of T-2 toxin-glucosides: phytotoxicity, stability after ingestion, antibody detection, and the anomericity of the naturally occurring T-2 toxin-glucoside found in cereal plants. T-2 toxin-β-glucoside was chemically synthesized and compared to T-2 toxin-α-glucoside prepared with Blastobotrys muscicola cultures and the T-2 toxin-glucoside found in naturally contaminated oats and wheat. The anomeric forms were separated chromatographically and differ in both NMR and mass spectrometry. Both anomers were significantly degraded to T-2 toxin and HT-2 toxin under conditions that mimic human digestion, but with different kinetics and metabolic end products. The naturally occurring T-2 toxin-glucoside from plants was found to be identical to T-2 toxin-α-glucoside prepared with B. muscicola. An antibody test for the detection of T-2 toxin was not effective for the detection of T-2 toxin-α-glucoside. This anomer was produced in sufficient quantity to assess its animal toxicity.


Glucose metabolism and insulin sensitivity in transgenic mice overexpressing leptin with lethal yellow agouti mutation: usefulness of leptin for the treatment of obesity-associated diabetes.
Leptin acts as an adipocyte-derived blood-borne satiety factor that can increase glucose metabolism. To elucidate the therapeutic implications of leptin for obesity-associated diabetes, we crossed transgenic skinny mice overexpressing leptin (Tg/+), which we have developed recently, and lethal yellow KKAy mice (Ay/+), a genetic model for obesity-diabetes syndrome, and examined the metabolic phenotypes of F1 animals. At 6 weeks of age, plasma leptin concentrations in Tg/+ mice with the Ay allele (Tg/+:Ay/+) were significantly higher than those in Ay/+ mice. Although no significant differences in body weight were noted among Tg/+:Ay/+ mice, Ay/+ mice, and their wild-type lean littermates (+/+), glucose and insulin tolerance tests revealed increased glucose tolerance and insulin sensitivity in Tg/+:Ay/+ compared with Ay/+ mice. However, at 12 weeks of age, when plasma leptin concentrations in Ay/+ mice were comparable to those in Tg/+:Ay/+ mice, Tg/+:Ay/+ mice developed obesity-diabetes syndrome similar to that of Ay/+ mice. Body weights of 12-week-old Tg/+:Ay/+ and Ay/+ mice were reduced to those of +/+ mice by a 3-week food restriction; when plasma leptin concentrations remained high in Tg/+:Ay/+ mice but were markedly reduced in Ay/+ and +/+ mice, glucose tolerance and insulin sensitivity in Tg/+:Ay/+ mice were markedly improved as compared with Ay/+ and +/+ mice. The present study demonstrates that hyperleptinemia can delay the onset of impaired glucose metabolism and accelerate the recovery from diabetes during caloric restriction in Tg/+:Ay/+ mice, thereby suggesting the potential usefulness of leptin in combination with a long-term caloric restriction for the treatment of obesity-associated diabetes.


GW in the Gaussian and Plane Waves Scheme with Application to Linear Acenes.
We present an implementation of G0W0 and eigenvalue-self-consistent GW (evGW) in the Gaussian and plane waves scheme for molecules. We calculate the correlation self-energy for imaginary frequencies employing the resolution of the identity. The correlation self-energy for real frequencies is then evaluated by analytic continuation. This technique allows an efficient parallel implementation and application to systems with several hundreds of atoms. Various benchmark calculations are presented. In particular, the convergence with respect to the most important numerical parameters is assessed for the benzene molecule. Comparisons with respect to other G0W0 implementations are reported for a set of molecules, while the performance of the method has been measured for water clusters containing up to 480 atoms in a cc-TZVP basis. Additionally, G0W0 has been applied for studying the influence of the ligands on the gap of small CdSe nanoparticles. evGW has been employed to calculate the HOMO-LUMO gaps of linear acenes, linear chains formed of connected benzene rings. Distinct differences between the closed and the open-shell (broken-symmetry) evGW HOMO-LUMO gaps for long acenes are found. In future experiments, a comparison of measured HOMO-LUMO gaps and our calculated evGW values may be helpful to determine the electronic ground state of long acenes.


Sudden cardiac death and the 'athlete's heart'.
To review the current literature pertaining to the cardiovascular adaptations to exercise and the impact on the physical examination, the electrocardiogram, and the echocardiogram, and to distinguish those physiologic changes from pathologic conditions associated with sudden cardiac death in athletes. Specific cardiovascular adaptations occur in response to regular physical exercise. The extent of these changes depend on the type and duration of exercise as well as the gender of the athlete. These cardiac adaptations are morphologically different from those conditions associated with sudden cardiac death. In the athlete younger than 35 years, hypertrophic cardiomyopathy is the most common cardiac condition associated with sudden death. Right ventricular dysplasia, idiopathic left ventricular hypertrophy, coronary anomalies, premature atherosclerosis, and Marfan syndrome compose the majority of the remaining causes of sudden cardiac death in athletes. In the athlete older than 35 years, coronary atherosclerosis is the leading cause of sudden death followed by those conditions responsible for sudden death in the younger athlete. Despite this, regular exercise before or following a myocardial infarction provides a protective effect with respect to overall mortality, cardiovascular mortality, and fatal reinfarction rates. Cardiovascular adaptation to regular physical exercise leads to morphologic changes in the myocardium that influence the cardiac examination, the electrocardiogram, and the echocardiogram. Knowledge and recognition of those changes can allow the clinician to distinguish normal physiologic changes from cardiac abnormality. Proper detection of athletes at high risk for sudden cardiac death and abstention from vigorous physical activity in these athletes may prevent sudden death.


Genome-wide mega-analysis identifies 16 loci and highlights diverse biological mechanisms in the common epilepsies.
The epilepsies affect around 65 million people worldwide and have a substantial missing heritability component. We report a genome-wide mega-analysis involving 15,212 individuals with epilepsy and 29,677 controls, which reveals 16 genome-wide significant loci, of which 11 are novel. Using various prioritization criteria, we pinpoint the 21 most likely epilepsy genes at these loci, with the majority in genetic generalized epilepsies. These genes have diverse biological functions, including coding for ion-channel subunits, transcription factors and a vitamin-B6 metabolism enzyme. Converging evidence shows that the common variants associated with epilepsy play a role in epigenetic regulation of gene expression in the brain. The results show an enrichment for monogenic epilepsy genes as well as known targets of antiepileptic drugs. Using SNP-based heritability analyses we disentangle both the unique and overlapping genetic basis to seven different epilepsy subtypes. Together, these findings provide leads for epilepsy therapies based on underlying pathophysiology.


Complex synaptic and intrinsic interactions disrupt input/output functions in the hippocampus of Scn1b knockout mice.
Mutations in the SCN1B gene have been linked to severe developmental epileptic encephalopathies including Dravet syndrome. Scn1b k nock o ut (KO) mice model SCN1B loss of function disorders, demonstrating seizures, developmental delays, and early death. SCN1B encodes the protein β1, an ion channel auxiliary subunit that also has roles in cell adhesion, neurite outgrowth, and gene expression. The goal of this project is to better understand of how loss of β1 alters information processing in the brain, resulting in seizures and associated cognitive dysfunction. Using slice electrophysiology in the CA1 region of the hippocampus from male and female Scn1b KO mice and w ild-type (WT) littermates, we found that processing of physiologically relevant patterned S chaffer c ollateral (SC) stimulation produces larger, prolonged depolarizations and increased spiking in KO neurons compared to WTs. KO neurons exhibit enhanced intrinsic excitability, firing more action potentials with current injection. Interestingly, SC stimulation produces smaller, more facilitating excitatory and inhibitory postsynaptic currents in KO pyramidal neurons, but larger postsynaptic potentials with the same stimulation. We also found reduced intrinsic firing of parvalbumin-expressing interneurons and disrupted recruitment of both parvalbumin- and somatostatin-expressing interneurons in response to patterned synaptic stimulation. Neuronal information processing relies on the interplay between synaptic properties, intrinsic properties that amplify or suppress incoming synaptic signals, and firing properties that produce cellular output. We found changes at each of these levels in Scn1b KO pyramidal neurons, resulting in fundamentally altered information processing in the hippocampus that likely contributes to the complex phenotypes of SCN1B -linked epileptic encephalopathies. Genetic developmental epileptic encephalopathies have limited treatment options, in part due to our lack of understanding of how genetic changes result in dysfunction at the cellular and circuit levels. SCN1B is a gene linked to Dravet syndrome and other epileptic encephalopathies, and Scn1b knockout mice phenocopy the human disease, allowing us to study underlying neurophysiological changes. Here we found changes at all levels of neuronal information processing in brains lacking β1, including intrinsic excitability, synaptic properties, and synaptic integration, resulting in greatly enhanced input/output functions of the hippocampus. Our study shows that loss of β1 results in a complex array of cellular and network changes that fundamentally alters information processing in the hippocampus.


[Chronic active hepatitis and cirrhosis induced by wild germander. 3 cases].
We report 3 cases of chronic liver injury that were observed after prolonged treatment with wild germander, a herbal medicine recently prohibited by French Ministry of Health, following several reports suggesting its hepatotoxicity. Chronic active hepatitis was found in 2 cases, and active cirrhosis in 1 case. The onset of hepatitis occurred after 6 to 7 months of treatment. Serum anti-nuclear and anti-smooth muscle antibodies were present in 2 patients. In 2 cases, wild-germander involvement was recognized several months after appearance of liver injury. Following treatment discontinuation, outcome was favorable in the 3 patients. These observations suggest that diagnosis of acute or chronic liver injury of unknown origin should always include the search for herbal medicine treatment.


Clinical application of C-reactive protein across the spectrum of acute coronary syndromes.
High-sensitivity C-reactive protein (hsCRP) is associated with adverse cardiovascular outcomes in acute coronary syndromes (ACS). The ability to formulate recommendations regarding clinical use of hsCRP is limited by a paucity of data regarding several key issues. The purpose of this study was to evaluate hsCRP across the spectrum of ACS. hsCRP was measured on admission in 3225 patients with ACS. hsCRP concentrations were compared in patients who suffered an adverse cardiac outcome within 10 months of study entry and in patients who had no adverse event. Because of heterogeneity in the relationship between hsCRP and clinical outcomes, evaluation was limited to patients from whom samples were collected within 48 h of symptom onset. Patients in the highest quartile of hsCRP compared to those in the lowest quartile were at increased risk of death at 30 days [adjusted hazard ratio (adjHR) 4.6, P <0.001] and 10 months (adjHR 3.9, P <0.001). In patients with unstable angina/non-ST-elevation myocardial infarction (STEMI), hsCRP >3 mg/L was associated with increased 10-month mortality (adjHR 2.3, P = 0.002), whereas in STEMI a relationship with mortality was seen at hsCRP >10 mg/L (adjHR 3.0, P = 0.008). Increased concentrations of hsCRP were strongly associated with the development of heart failure at 30 days (adjHR 8.2, P = 0.001) and 10 months (adjHR 2.6, P = 0.014). Increased baseline concentrations of hsCRP are strongly associated with mortality and heart failure across the ACS spectrum. hsCRP measurement should be performed early after presentation and index diagnosis-specific cutpoints should be used.


Skeletal muscle morphology in sarcopenia defined using the EWGSOP criteria: findings from the Hertfordshire Sarcopenia Study (HSS).
Sarcopenia is defined as the loss of muscle mass and function with age and is associated with decline in mobility, frailty, falls and mortality. There is considerable interest in understanding the underlying mechanisms. Our aim was to characterise muscle morphology changes associated with sarcopenia among community dwelling older men. One hundred and five men aged 68-76 years were recruited to the Hertfordshire Sarcopenia Study (HSS) for detailed characterisation of muscle including measures of muscle mass, strength and function. Muscle tissue was obtained from a biopsy of the vastus lateralis for 99 men and was processed for immunohistochemical studies to determine myofibre distribution and area, capillarisation and satellite cell (SC) density. Six (6 %) men had sarcopenia as defined by the European Working Group on Sarcopenia in Older People (EWGSOP) criteria. These men had lower SC density (1.7 cells/mm(2) vs 3.8 cells/mm(2), p = 0.06) and lower SC/fibre ratio (0.02 vs 0.06, p = 0.06) than men without sarcopenia. Although men with sarcopenia tended to have smaller myofibres and lower capillary to fibre ratio, these relationships were not statistically significant. We have shown that there may be altered muscle morphology parameters in older men with sarcopenia. These results have the potential to help identify cell and molecular targets for therapeutic intervention. This work now requires extension to larger studies which also include women.