Open arterial revascularization of the critically ischemic foot using arterial homograft.
Revascularization alternatives for patients with critical limb ischemia and without adequate autogenous vein remain challenging. We reviewed our experience with the use of arterial homograft as a conduit for limb salvage in patients with limb ischemia and active lower extremity infections. A retrospective review of patients who underwent open arterial revascularization of the lower extremity with cryopreserved femoral artery homograft for the treatment of symptomatic critical limb ischemia (i.e., foot ulceration, infection, or gangrene) during an 18-month period was performed. Relevant clinical variables and treatment outcomes were analyzed. Clinical success was defined as limb salvage for one year, patency of the reconstruction, and wound healing. Thirteen patients (5 men; average age 71 +/- 83 years, range 51-87 years) were treated during this study period. Treatment indications included 10 (77%) foot ulcerations, 2 (15%) critically ischemic limbs without ulceration, and 1 (8%) infected polytetrafluoroethylene bypass graft with acute occlusion and limb ischemia. A femoral below-the-knee popliteal bypass was performed in 4 (1%), femoral to anterior tibial artery in 4 (31%), femoral to posterior tibial artery in 3 (23%), and femoral to peroneal artery in 2 (15%). All 13 limbs were preserved. Minor amputations were performed in 6 patients, 2 underwent toe amputations and 4 patients had a trans-metatarsal amputation. The cumulative patency rate at 6, 9, and 18 months was 92.3%, 70.3%, and 58.6%, respectively. Open arterial revascularization with arterial femoral homograft is an acceptable treatment method in patients with critical limb ischemia and active infection in whom autogenous vein is not available or the use of a synthetic conduit is not possible.


Serum eosinophil peroxidase (EPO) levels in asthmatic patients.
Eosinophil granular proteins are a useful eosinophilic activation marker in asthmatic patients. In this study, the eosinophil peroxidase (EPO) levels were assessed in different stages of bronchial asthma, in 123 patients suffering from asthma, classified as mild (n =49), moderate (n = 49), and severe (n = 25), according to the International Consensus Report of Diagnosis and Treatment of Asthma, as well as in 27 healthy controls, with the aim of evaluating the importance of this protein as a severity marker in bronchial asthma, and its possible correlation with parameters such as anamnesis, respiratory function tests, and peripheral blood eosinophil count, and also with some allergologic diagnostic tests, both in vivo and in vitro. The geometric mean serum level of EPO was 9.3 +/- 11.3 ng/ml (median +/- SD) in controls, 28 +/- 37.8 ng/ml in the asthmatic patients. Depending on the asthma severity, the EPO levels were 25 +/- 30.5; 29 +/- 37.1, and 41 +/- 47.3 ng/ml in mild, moderate, and severe asthmatics, respectively, being the significant differences between the group of patients with mild and severe asthma (P < 0.001). The number of eosinophils (eos) in peripheral blood was 157 +/- 20 eos/mm3 in the controls, 334/35 eos/mm3 in mild asthmatics, 510 +/- 87 eos/mm3 in moderate asthmatics, and 658 +/- 72 eos/mm3 in severe asthmatics, with significant differences between all groups (from P < 0.05 to P < 0.001). Both the serum levels of EPO and the number of eosinophils were greater in patients with active asthma patients with inactive asthma (P < 0.001). Significant negative correlations (P < 0.001) were found between serum levels of EPO and FEV1 (rs = 0.30), MEF25-75 (rs = -0.33) and MEF50 (rs = -0.34), and a good positive correlation (rs = 0.80, P < 0.001) was found between EPO levels and the number of eosinophils in peripheral blood. We also found a significant positive correlation between eosinophil number and clinical score (rs = 0.54, P < 0.001) and between EPO levels and the mentioned score (rs = 0.46, P < 0.001).


Superficial temporal artery and vein as recipient vessels for scalp and facial reconstruction: radiographic support for underused vessels.
The choice of the recipient vessels for microsurgical reconstruction is based on proximity to the defect, vessel caliber, anatomic reliability, and ease of dissection. Traditionally in head and neck reconstruction, the superficial temporal artery and vein (STA/V) have been reserved as secondary recipient vessels. Others, including the facial artery (FA) and vein have been the preferred choice for facial reconstruction. A retrospective analysis of all consecutive head and neck reconstructions using the STA/V by the senior author (E.S.G.) over a 5-year period was performed. Perioperative data were reviewed for all the patients. In a subset of patients, radiographic analysis was used to delineate STA and FA anatomy including vessel diameter and distances to standard anatomic landmarks. A total of 31 patients had 32 microsurgical reconstructions using the STA/V as recipient vessels. Radiographic analysis revealed no significant difference between the STA and FA diameters. The distances from the STA to the upper and middle face were significantly shorter relative to the FA, 64 versus 102 mm (p < 0.0001) and 72 versus 80 mm (p < 0.04), respectively. The distances from the lower face to the STA and FA were 56 and 30 mm, respectively (p < 0.0001). The STA/V can be used as first choice recipient vessels in head and neck reconstruction. Key features of these vessels include proximity to defect, acceptable caliber, predictable anatomic location, and relative ease of dissection. We recommend that the STA/V be considered recipient vessels of choice for reconstruction of defects of the face and scalp.


Health-related quality of life following blind rehabilitation.
The purpose of this study was to investigate the effect of residential blind rehabilitation on patients' vision-targeted health-related quality of life (HRQOL) and general physical and mental function. The National Eye Institute 25-item Visual Function Questionnaire (NEI VFQ) plus appendix questions, the 12-item Short-Form Health Survey (SF-12), Hope Scale and Coopersmith self-esteem inventory were administered to 206 legally blind veterans prior to their entering a residential (in-patient) blind rehabilitation program and again to 185 and 176 of the original cohort at 2 and 6 months after completion of the rehabilitation program, respectively. Data on visual acuity, visual field extent, contrast sensitivity and scanning ability were also collected. The duration of the in-patient rehabilitation programs ranged from 11 to 109 days. Questionnaire scores were compared pre-rehabilitation and post-rehabilitation. Following rehabilitation there was a significant improvement in nine of 11 NEI VFQ subscales and in a composite score at both the 2- and 6-month post-rehabilitation intervals. Mental health (SF-12) and self esteem also improved significantly although physical health ratings declined over the course of the study (approximately 10 months). Residential blind rehabilitation appears to improve patients' self-reported vision-targeted HRQOL, self-esteem and mental health aspects of generic HRQOL.


Genetic variation on chromosome 9p21 shows association with the ischaemic stroke subtype large-vessel disease in a Swedish sample aged ≤ 70.
The aim of this study was to investigate whether we could replicate a recent finding of an association between genetic variants on chromosome 9p21 and the ischaemic stroke (IS) subtype large-vessel disease (LVD). The Sahlgrenska Academy Study on Ischemic Stroke comprises 844 patients with IS, who suffered IS before reaching the age of 70, and 668 healthy controls. IS subtype was defined according to the TOAST criteria, and 111 patients were categorized as LVD. Seven single-nucleotide polymorphisms (SNPs) on 9p21 were analyzed. Functional outcome was assessed 3 months after IS using the modified Rankin scale. The SNP rs7857345 showed a significant association with the subtype LVD independent of traditional vascular risk factors. In addition, an association between rs7857345 and functional outcome after LVD was observed. In this relatively young sample of patients with IS, genetic variation on 9p21 is associated with LVD.


Flaxseed consumption influences endogenous hormone concentrations in postmenopausal women.
Lignans, similar in structure to endogenous sex steroid hormones, may act in vivo to alter hormone metabolism and subsequent cancer risk. The objective of this study was to examine effects of dietary intake of a lignan-rich plant food (flaxseed) on serum concentrations of endogenous hormones and binding proteins (estrone, estrone sulfate, 17 beta-estradiol, sex hormone-binding globulin, progesterone, prolactin, dehydroepiandrosterone sulfate, dehydroepiandrosterone, androstenedione, testosterone, and free testosterone) in postmenopausal women. This randomized, crossover trial consisted of three seven-week feeding periods, during which 28 postmenopausal women, aged 52-82 yr, consumed their habitual diets plus 0, 5, or 10 g of ground flaxseed. Serum samples collected during the last week of each feeding period were analyzed for serum hormones using standard diagnostic kits. The flaxseed diets significantly reduced serum concentrations of 17 beta-estradiol by 3.26 pg/ml (12.06 pmol/l) and estrone sulfate by 0.09 ng/ml (0.42 nmol/l) and increased prolactin by 1.92 micrograms/l (0.05 IU/ml). Serum concentrations of androstenedione, estrone, sex hormone-binding globulin, progesterone, testosterone, free testosterone, dehydroepiandrosterone, and dehydroepiandrosterone sulfate were not altered with flaxseed feeding. In this group of postmenopausal women, consuming flaxseed in addition to their habitual diets influenced their endogenous hormone metabolism by decreasing serum 17 beta-estradiol and estrone sulfate and increasing serum prolactin concentrations.


Delivery of biannual ultrasound surveillance for individuals with cirrhosis and cured hepatitis C in the UK.
Previous studies show the uptake of biannual ultrasound (US) surveillance in patients with cirrhosis is suboptimal. Here, our goal was to understand in broader terms how surveillance is being delivered to cirrhosis patients with cured hepatitis C in the UK. Hepatitis C cirrhosis patients achieving a sustained viral response (SVR) to antiviral therapies were identified from the national Hepatitis-C-Research-UK resource. Data on (i) liver/abdominal US examinations, (ii) HCC diagnoses, and (iii) HCC curative treatment were obtained through record-linkage to national health registries. The rate of US uptake was calculated by dividing the number of US episodes by follow-up time. A total of 1908 cirrhosis patients from 31 liver centres were followed for 3.8 (IQR: 3.4-4.9) years. Overall, 10 396 liver/abdominal USs were identified. The proportion with biannual US was 19% in the first 3 years after SVR and 9% for all follow-up years. Higher uptake of biannual US was associated with attending a liver transplant centre; older age and cirrhosis decompensation. Funnel plot analysis indicated significant inter-centre variability in biannual US uptake, with 6/29 centres outside control limits. Incident HCC occurred in 133 patients, of which 49/133 (37%) were treated with curative intent. The number of US episodes in the two years prior to HCC diagnosis was significantly associated with higher odds of curative-intent treatment (aOR: 1.53; 95% CI: 1.12-2,09; p = .007). This study provides novel data on the cascade of care for HCC in the UK. Our findings suggest biannual US is poorly targeted, inefficient and is not being delivered equitably to all patients.


The Effects of Stimulator, Waveform, and Current Direction on Intracortical Inhibition and Facilitation: A TMS Comparison Study.
Background: Cortical function is dependent on the balance between excitatory and inhibitory influences. In the human motor cortex, surrogates of these interactions can be measured in vivo, non-invasively with double-pulse transcranial magnetic stimulation (TMS). To compare results from data acquired with different available setups and bring data together, it is inevitable to determine whether different TMS setups lead to comparable or differential results. Objective: We assessed and compared short intracortical inhibition (SICI) and intracortical facilitation (ICF) testing four different experimental conditions. Methods: SICI and ICF were studied with different stimulators (Magstim BiStim2 or MagVenture MagPro X100), waveforms (monophasic or biphasic), current directions (anterior-posterior or posterior-anterior) at interstimulus intervals (ISIs) of 1, 3, 10, 15 ms. Results: We were not able to detect differences for SICI and ICF, when comparing the tested conditions, except for 3 ms SICI in which the anterior-posterior current direction led to stronger modulation. Correlation analysis suggested comparability for 3 ms SICI for the Magstim monophasic posterior-anterior condition with both tested MagVenture conditions. Conclusions: 3 ms SICI data sets obtained with two different, commonly used stimulators (Magstim BiStim2 or MagVenture MagPro X100) with conventionally used stimulation parameters are largely comparable. This may allow the combination of data sets in an open science view.


αB-crystallin polydispersity is a consequence of unbiased quaternary dynamics.
The inherent heterogeneity of many protein assemblies complicates characterization of their structure and dynamics, as most biophysical techniques require homogeneous preparations of isolated components. For this reason, quantitative studies of the molecular chaperone αB-crystallin, which populates a range of interconverting oligomeric states, have been difficult, and the physicochemical basis for its polydispersity has remained unknown. Here, we perform mass spectrometry experiments to study αB-crystallin and extract detailed information as to its oligomeric distribution and exchange of subunits under a range of conditions. This allows a determination of the thermodynamic and kinetic parameters that govern the polydisperse ensemble and enables the construction of a simple energy profile for oligomerization. We find that the quaternary structure and dynamics of the protein can be explained using a simple model with just two oligomer-independent interactions (i.e., interactions that are energetically identical in all oligomers from 10mers to 40mers) between constituent monomers. As such, the distribution of oligomers is governed purely by the dynamics of individual monomers. This provides a new means for understanding the polydispersity of αB-crystallin and a framework for interrogating other heterogeneous protein assemblies.


Mechanisms contributing to intracellular pH homeostasis in an immortalised human chondrocyte cell line.
The maintenance of chondrocyte pH is an important parameter controlling cartilage matrix turnover rates. Previous studies have shown that, to varying degrees, chondrocytes rely on Na(+)/H(+) exchange to regulate pH. HCO(3)(-)-dependent buffering and HCO(3)(-)-dependent acid-extrusion systems seem to play relatively minor roles. This situation may reflect minimal carbonic anhydrase activity in cartilage cells. In the present study, the pH regulation of the human chondrocyte cell line, C-20/A4 has been characterised. Intracellular pH (pH(i)) was measured using the H(+)-sensitive fluoroprobe BCECF. In solutions lacking HCO(3)(-)/CO(2), pH(i) was approximately 7.5, and the recovery from intracellular acidification was predominantly mediated by a Na(+)-dependent, amiloride- and HOE 694-sensitive process. A small additional component which was sensitive to chloro-7-nitrobenz-2-oxa-1,3-diazole, an inhibitor of the V-type H(+)-ATPase, was also apparent. In solutions containing HCO(3)(-)/CO(2), pH(i) was approximately 7.2. Comparison of buffering capacity in the two conditions showed that this variable was not significantly augmented in HCO(3)(-)/CO(2)-containing media. The recovery from intracellular acidification was more rapid in the presence of HCO(3)(-)/CO(2), although under these conditions it was again largely dependent on Na(+) ions and inhibited by amiloride and HOE 694. A small component was inhibited by SITS, although this effect did not reach the level of statistical significance. These findings indicate that HCO(3)(-)-dependent processes play only a minimal role in pH regulation in C-20/A4 chondrocytes. pH regulation instead relies heavily on the Na(+)/H(+) exchanger together with a H(+)-ATPase. The absence of extrinsic (HCO(3)(-)/CO(2)) buffering is likely to reflect the low levels of carbonic anhydrase in these cells. In addition to providing fundamental information about a widely-used cell line, these findings support the contention that the unusual nature of pH regulation in chondrocytes reflects the paucity of carbonic anhydrase activity in these cells.


Human NK cells are alerted to induction of p53 in cancer cells by upregulation of the NKG2D ligands ULBP1 and ULBP2.
Natural killer (NK) cells are immune cells sensing and eliminating foreign, stressed, transformed, and senescent cells through specialized surface receptors, such as NKG2D, that interacts with several virus- or stress-inducible ligands, including ULBP1 and -2, which are expressed on target cell surfaces. For example, induction of DNA damage or cellular senescence pathways in tumor cells led to upregulation of NKG2D ligands that activate NK cells. Although, both pathways activate p53, the relationship of p53 activation to upregulation of NKG2D ligands has not been addressed. In this study, we report that induction of wild-type p53, but not mutant p53, strongly upregulated mRNA and cell surface expression of ULBP1 and -2, whereas expression of other NK cell ligands was not affected. We defined intronic p53-responsive elements in these two novel p53 target genes. Coculture of wild-type p53-induced human tumor cells with primary human NK cells enhanced NKG2D-dependent degranulation and IFN-γ production by NK cells. Accordingly, treatment of certain wild-type p53-expressing tumor cell lines with the p53-reactivating small molecular compound RITA resulted in upregulation of ULBP2 mRNA and cell surface protein expression. Taken together, our findings define the involvement of p53 in the regulation of specific NKG2D ligands that enhance NK cell-mediated target recognition. One implication of our work is that activating p53 after adoptive transfer of NK cells might constitute an effective combinatorial strategy of NK cell-based immunochemotherapy in cancers in which wild-type p53 function is preserved.


Multicentre clinical trials of benzimidazole-carbamates in human cystic echinococcosis (phase 2).
A multicentre study which constituted the second phase of trials of the efficacy of albendazole and mebendazole in human cystic echinococcosis was coordinated by WHO. A total of 112 patients from four clinical centres in Beirut, Paris, Rome and Sofia completed standardized dosage of regimens of each drug and 68 patients were followed up for at least 12 months after treatment. Albendazole was more effective than mebendazole and adverse reactions were comparable with both treatment regimens. At least 12 months is needed after treatment for an objective evaluation of the efficacy of benzimidazoles. At present, treatment with albendazole or mebendazole should be reserved for inoperable cases of cystic echinococcosis (under strict medical supervision) and individualized according to the patient's response and the occurrence and severity of adverse reactions.


Digenean Holostephanus (Trematoda: Digenea: Cyathocotylidae) metacercariae in common carp (Cyprinus carpio Linnaeus, 1758) muscle: zoonotic potential and sensitivity to physico-chemical treatments.
Metacercariae of various species within the genus Holostephanus Szidat, 1936 (Trematoda: Digenea: Cyathocotylidae) occur in muscles of both farmed and wild fish, including common carp (Cyprinus carpio Linnaeus, 1758). The life cycle includes a snail as first intermediate host, fish as second intermediate host and birds or mammals as final hosts. We studied the zoonotic potential and the viability of Holostephanus metacercariae from common carp following exposure to various physical and chemical treatments. Muscle tissue samples of common carp specimens from a fish farm in the north-eastern part of Hungary were examined and metacercariae recovered. The zoonotic potential was evaluated experimentally by using small mammals as models (albino mice, n = 2; and Syrian hamsters, n = 4) infected per os with Holostephanus cysts. Parallelly, Metagonimus metacercariae were used as positive controls. We could not confirm the zoonotic potential of Holostephanus metacercariae as they did not survive in the mammalian intestine whereas Metagonimus metacercariae developed to the adult stage. We assessed the viability of metacercariae isolated from common carp specimens during exposure to different physical treatments (temperatures of -18°C, +20°C, +40°C and +60°C) and chemical agents (5% and 10% acetic acid and 10% sodium chloride (NaCl)). Metacercariae lost viability by freezing at -18°C (2 h), heating at 60°C (20 min), incubation in 5% and 10% acetic acid (5 min) and 10% NaCl (2 h). These methods served as models to investigate the effectiveness of food preparation techniques (such as cold and hot smoking, freezing, salting and pickling) on the survival of metacercariae.


Real-Time Predictions of Reservoir Size and Rebound Time during Antiretroviral Therapy Interruption Trials for HIV.
Monitoring the efficacy of novel reservoir-reducing treatments for HIV is challenging. The limited ability to sample and quantify latent infection means that supervised antiretroviral therapy (ART) interruption studies are generally required. Here we introduce a set of mathematical and statistical modeling tools to aid in the design and interpretation of ART-interruption trials. We show how the likely size of the remaining reservoir can be updated in real-time as patients continue off treatment, by combining the output of laboratory assays with insights from models of reservoir dynamics and rebound. We design an optimal schedule for viral load sampling during interruption, whereby the frequency of follow-up can be decreased as patients continue off ART without rebound. While this scheme can minimize costs when the chance of rebound between visits is low, we find that the reservoir will be almost completely reseeded before rebound is detected unless sampling occurs at least every two weeks and the most sensitive viral load assays are used. We use simulated data to predict the clinical trial size needed to estimate treatment effects in the face of highly variable patient outcomes and imperfect reservoir assays. Our findings suggest that large numbers of patients-between 40 and 150-will be necessary to reliably estimate the reservoir-reducing potential of a new therapy and to compare this across interventions. As an example, we apply these methods to the two "Boston patients", recipients of allogeneic hematopoietic stem cell transplants who experienced large reductions in latent infection and underwent ART-interruption. We argue that the timing of viral rebound was not particularly surprising given the information available before treatment cessation. Additionally, we show how other clinical data can be used to estimate the relative contribution that remaining HIV+ cells in the recipient versus newly infected cells from the donor made to the residual reservoir that eventually caused rebound. Together, these tools will aid HIV researchers in the evaluating new potentially-curative strategies that target the latent reservoir.


Enhanced Mucosal Defense and Reduced Tumor Burden in Mice with the Compromised Negative Regulator IRAK-M.
Aberrant inflammation is a hallmark of inflammatory bowel disease (IBD) and colorectal cancer. IRAK-M is a critical negative regulator of TLR signaling and overzealous inflammation. Here we utilize data from human studies and Irak-m-/- mice to elucidate the role of IRAK-M in the modulation of gastrointestinal immune system homeostasis. In human patients, IRAK-M expression is up-regulated during IBD and colorectal cancer. Further functional studies in mice revealed that Irak-m-/- animals are protected against colitis and colitis associated tumorigenesis. Mechanistically, our data revealed that the gastrointestinal immune system of Irak-m-/- mice is highly efficient at eliminating microbial translocation following epithelial barrier damage. This attenuation of pathogenesis is associated with expanded areas of gastrointestinal associated lymphoid tissue (GALT), increased neutrophil migration, and enhanced T-cell recruitment. Further evaluation of Irak-m-/- mice revealed a splice variant that robustly activates NF-κB signaling. Together, these data identify IRAK-M as a potential target for future therapeutic intervention.


Cardiac myocyte diversity and a fibroblast network in the junctional region of the zebrafish heart revealed by transmission and serial block-face scanning electron microscopy.
The zebrafish has emerged as an important model of heart development and regeneration. While the structural characteristics of the developing and adult zebrafish ventricle have been previously studied, little attention has been paid to the nature of the interface between the compact and spongy myocardium. Here we describe how these two distinct layers are structurally and functionally integrated. We demonstrate by transmission electron microscopy that this interface is complex and composed primarily of a junctional region occupied by collagen, as well as a population of fibroblasts that form a highly complex network. We also describe a continuum of uniquely flattened transitional cardiac myocytes that form a circumferential plate upon which the radially-oriented luminal trabeculae are anchored. In addition, we have uncovered within the transitional ring a subpopulation of markedly electron dense cardiac myocytes. At discrete intervals the transitional cardiac myocytes form contact bridges across the junctional space that are stabilized through localized desmosomes and fascia adherentes junctions with adjacent compact cardiac myocytes. Finally using serial block-face scanning electron microscopy, segmentation and volume reconstruction, we confirm the three-dimensional nature of the junctional region as well as the presence of the sheet-like fibroblast network. These ultrastructural studies demonstrate the previously unrecognized complexity with which the compact and spongy layers are structurally integrated, and provide a new basis for understanding development and regeneration in the zebrafish heart.


Memory CD8+ T cells mediate antibacterial immunity via CCL3 activation of TNF/ROI+ phagocytes.
Cytolysis, interferon gamma and tumor necrosis factor (TNF) alpha secretion are major effector mechanisms of memory CD8+ T cells that are believed to be required for immunological protection in vivo. By using mutants of the intracellular bacterium Listeria monocytogenes, we found that none of these effector activities is sufficient to protect against secondary infection with wild-type (WT) bacteria. We demonstrated that CCL3 derived from reactivated memory CD8+ T cells is required for efficient killing of WT bacteria. CCL3 induces a rapid TNF-alpha secretion by innate inflammatory mononuclear phagocytic cells (MPCs), which further promotes the production of radical oxygen intermediates (ROIs) by both MPCs and neutrophils. ROI generation is the final bactericidal mechanism involved in L. monocytogenes clearance. These results therefore uncover two levels of regulation of the antibacterial secondary protective response: (a) an antigen-dependent phase in which memory CD8+ T cells are reactivated and control the activation of the innate immune system, and (b) an antigen-independent phase in which the MPCs coordinate innate immunity and promote the bactericidal effector activities. In this context, CCL3-secreting memory CD8+ T cells are able to mediate "bystander" killing of an unrelated pathogen upon antigen-specific reactivation, a mechanism that may be important for the design of therapeutic vaccines.


Spiers Memorial Lecture. Surface-enhanced Raman spectroscopy: from single particle/molecule spectroscopy to ångstrom-scale spatial resolution and femtosecond time resolution.
Four decades on, surface-enhanced Raman spectroscopy (SERS) continues to be a vibrant field of research that is growing (approximately) exponentially in scope and applicability while pushing at the ultimate limits of sensitivity, spatial resolution, and time resolution. This introductory paper discusses some aspects related to all four of the themes for this Faraday Discussion. First, the wavelength-scanned SERS excitation spectroscopy (WS-SERES) of single nanosphere oligomers (viz., dimers, trimers, etc.), the distance dependence of SERS, the magnitude of the chemical enhancement mechanism, and the progress toward developing surface-enhanced femtosecond stimulated Raman spectroscopy (SE-FSRS) are discussed. Second, our efforts to develop a continuous, minimally invasive, in vivo glucose sensor based on SERS are highlighted. Third, some aspects of our recent work in single molecule SERS and the translation of that effort to ångstrom-scale spatial resolution in ultrahigh vacuum tip-enhanced Raman spectroscopy (UHV-TERS) and single molecule electrochemistry using electrochemical (EC)-TERS will be presented. Finally, we provide an overview of analytical SERS with our viewpoints on SERS substrates, approaches to address the analyte generality problem (i.e. target molecules that do not spontaneously adsorb and/or have Raman cross sections <10-29 cm2 sr-1), SERS for catalysis, and deep UV-SERS.


Comparison of fMRI motion correction software tools.
Motion correction of fMRI data is a widely used step prior to data analysis. In this study, a comparison of the motion correction tools provided by several leading fMRI analysis software packages was performed, including AFNI, AIR, BrainVoyager, FSL, and SPM2. Comparisons were performed using data from typical human studies as well as phantom data. The identical reconstruction, preprocessing, and analysis steps were used on every data set, except that motion correction was performed using various configurations from each software package. Each package was studied using default parameters, as well as parameters optimized for speed and accuracy. Forty subjects performed a Go/No-go task (an event-related design that investigates inhibitory motor response) and an N-back task (a block-design paradigm investigating working memory). The human data were analyzed by extracting a set of general linear model (GLM)-derived activation results and comparing the effect of motion correction on thresholded activation cluster size and maximum t value. In addition, a series of simulated phantom data sets were created with known activation locations, magnitudes, and realistic motion. Results from the phantom data indicate that AFNI and SPM2 yield the most accurate motion estimation parameters, while AFNI's interpolation algorithm introduces the least smoothing. AFNI is also the fastest of the packages tested. However, these advantages did not produce noticeably better activation results in motion-corrected data from typical human fMRI experiments. Although differences in performance between packages were apparent in the human data, no single software package produced dramatically better results than the others. The "accurate" parameters showed virtually no improvement in cluster t values compared to the standard parameters. While the "fast" parameters did not result in a substantial increase in speed, they did not degrade the cluster results very much either. The phantom and human data indicate that motion correction can be a valuable step in the data processing chain, yielding improvements of up to 20% in the magnitude and up to 100% in the cluster size of detected activations, but the choice of software package does not substantially affect this improvement.


The Personality Assessment Inventory as a proxy for the Psychopathy Checklist Revised: testing the incremental validity and cross-sample robustness of the Antisocial Features Scale.
The Personality Assessment Inventory's (PAI's) ability to predict psychopathic personality features, as assessed by the Psychopathy Checklist-Revised (PCL-R), was examined. To investigate whether the PAI Antisocial Features (ANT) Scale and subscales possessed incremental validity beyond other theoretically relevant PAI scales, optimized regression equations were derived in a sample of 281 Canadian federal offenders. ANT, or ANT-Antisocial Behavior (ANT-A), demonstrated unique variance in regression analyses predicting PCL-R total and Factor 2 (Lifestyle Impulsivity and Social Deviance) scores, but only the Dominance (DOM) Scale was retained in models predicting Factor 1 (Interpersonal and Affective Deficits). Attempts to cross-validate the regression equations derived from the first sample on a sample of 85 U.S. sex offenders resulted in considerable validity shrinkage, with the ANT Scale in isolation performing comparably to or better than the statistical models for PCL-R total and Factor 2 scores. Results offer limited evidence of convergent validity between the PAI and the PCL-R.


Climate and dengue transmission: evidence and implications.
Climate influences dengue ecology by affecting vector dynamics, agent development, and mosquito/human interactions. Although these relationships are known, the impact climate change will have on transmission is unclear. Climate-driven statistical and process-based models are being used to refine our knowledge of these relationships and predict the effects of projected climate change on dengue fever occurrence, but results have been inconsistent. We sought to identify major climatic influences on dengue virus ecology and to evaluate the ability of climate-based dengue models to describe associations between climate and dengue, simulate outbreaks, and project the impacts of climate change. We reviewed the evidence for direct and indirect relationships between climate and dengue generated from laboratory studies, field studies, and statistical analyses of associations between vectors, dengue fever incidence, and climate conditions. We assessed the potential contribution of climate-driven, process-based dengue models and provide suggestions to improve their performance. Relationships between climate variables and factors that influence dengue transmission are complex. A climate variable may increase dengue transmission potential through one aspect of the system while simultaneously decreasing transmission potential through another. This complexity may at least partly explain inconsistencies in statistical associations between dengue and climate. Process-based models can account for the complex dynamics but often omit important aspects of dengue ecology, notably virus development and host-species interactions. Synthesizing and applying current knowledge of climatic effects on all aspects of dengue virus ecology will help direct future research and enable better projections of climate change effects on dengue incidence.


Multiportal Combined Transorbital and Transnasal Endoscopic Resection of Fibrous Dysplasia.
Introduction Historically, access to the anterior skull base was achieved with open procedures. The paradigms to this approach were challenged with the advent of minimally disruptive endoscopic surgical techniques and supporting technology. The next step in the evolution of minimally disruptive surgery was the combination of multiportal endoscopic surgery. Results The patient was an 18-year-old man who presented with right-sided proptosis. Further diagnostic tests revealed a fibrous dysplasia (FD) occupying the skull base and orbit. The lesion was successfully resected. Conclusions The location of the tumor in this case was challenging, in which surgeons at some centers would have opted to have performed as an open procedure instead of endoscopically. The combined transnasal/transorbital approach is an uncommonly used technique that we have used to remove this tumor successfully. The patient also had a unique disease (FD) in a unique location that was treated without complications. This case report highlights how surgeons may use an expanded armamentarium in dealing with complex pathologies.


The V-ATPase proteolipid cylinder promotes the lipid-mixing stage of SNARE-dependent fusion of yeast vacuoles.
The V-ATPase V(0) sector associates with the peripheral V(1) sector to form a proton pump. V(0) alone has an additional function, facilitating membrane fusion in the endocytic and late exocytic pathways. V(0) contains a hexameric proteolipid cylinder, which might support fusion as proposed in proteinaceous pore models. To test this, we randomly mutagenized proteolipids. We recovered alleles that preserve proton translocation, normal SNARE activation and trans-SNARE pairing but that impair lipid and content mixing. Critical residues were found in all subunits of the proteolipid ring. They concentrate within the bilayer, close to the ring subunit interfaces. The fusion-impairing proteolipid substitutions stabilize the interaction of V(0) with V(1). Deletion of the vacuolar v-SNARE Nyv1 has the same effect, suggesting that both types of mutations similarly alter the conformation of V(0). Also covalent linkage of subunits in the proteolipid cylinder blocks vacuole fusion. We propose that a SNARE-dependent conformational change in V(0) proteolipids might stimulate fusion by creating a hydrophobic crevice that promotes lipid reorientation and formation of a lipidic fusion pore.


Seasonal water stress tolerance and habitat associations within four neotropical tree genera.
We investigated the relationship between habitat association and physiological performance in four congeneric species pairs exhibiting contrasting distributions between seasonally flooded and terra firme habitats in lowland tropical rain forests of French Guiana, including Virola and Iryanthera (Myristicaceae), Symphonia (Clusiaceae), and Eperua (Caesalpiniaceae). We analyzed 10-year data sets of mapped and measured saplings (stems >150 cm in height and <10 cm diameter at breast height [dbh]) and trees (stems > or =10 cm dbh) across 37.5 ha of permanent plots covering a 300-ha zone, within which seasonally flooded areas (where the water table never descends below 1 m) have been mapped. Additionally, we tested the response of growth, survival, and leaf functional traits of these species to drought and flood stress in a controlled experiment. We tested for habitat preference using a modification of the torus translation method. Strong contrasting associations of the species pairs of Iryanthera, Virola, and Symphonia were observed at the sapling stage, and these associations strengthened for the tree stage. Neither species of Eperua was significantly associated with flooded habitats at the sapling stage, but E. falcata was significantly and positively associated with flooded forests at the tree stage, and trees of E. grandiflora were found almost exclusively in nonflooded habitats. Differential performance provided limited explanatory support for the observed habitat associations, with only congeners of Iryanthera exhibiting divergent sapling survival and tree growth. Seedlings of species associated with flooded forest tended to have higher photosynthetic capacity than their congeners at field capacity. In addition, they tended to have the largest reductions in leaf gas exchange and growth rate in response to experimental drought stress and the least reductions in response to experimental inundation. The corroboration of habitat association with differences in functional traits and, to a lesser extent, measures of performance provides an explanation for the regional coexistence of these species pairs. We suggest that specialization to seasonally flooded habitats may explain patterns of adaptive radiation in many tropical tree genera and thereby provide a substantial contribution to regional tree diversity.


Nebulized Recombinant Tissue Plasminogen Activator (rt-PA) for Acute COVID-19-Induced Respiratory Failure: An Exploratory Proof-of-Concept Trial.
Acute lung injury in COVID-19 results in diffuse alveolar damage with disruption of the alveolar-capillary barrier, coagulation activation, alveolar fibrin deposition and pulmonary capillary thrombi. Nebulized recombinant tissue plasminogen activator (rt-PA) has the potential to facilitate localized thrombolysis in the alveolar compartment and improve oxygenation. In this proof-of-concept safety study, adults with COVID-19-induced respiratory failure and a <300 mmHg PaO2/FiO2 (P/F) ratio requiring invasive mechanical ventilation (IMV) or non-invasive respiratory support (NIRS) received nebulized rt-PA in two cohorts (C1 and C2), alongside standard of care, between 23 April-30 July 2020 and 21 January-19 February 2021, respectively. Matched historical controls (MHC; n = 18) were used in C1 to explore efficacy. Safety co-primary endpoints were treatment-related bleeds and <1.0-1.5 g/L fibrinogen reduction. A variable dosing strategy with clinical efficacy endpoint and minimal safety concerns was determined in C1 for use in C2; patients were stratified by ventilation type to receive 40-60 mg rt-PA daily for ≤14 days. Nine patients in C1 (IMV, 6/9; NIRS, 3/9) and 26 in C2 (IMV, 12/26; NIRS, 14/26) received nebulized rt-PA for a mean (SD) of 6.7 (4.6) and 9.1(4.6) days, respectively. Four bleeds (one severe, three mild) in three patients were considered treatment related. There were no significant fibrinogen reductions. Greater improvements in mean P/F ratio from baseline to study end were observed in C1 compared with MHC (C1; 154 to 299 vs. MHC; 154 to 212). In C2, there was no difference in the baseline P/F ratio of NIRS and IMV patients. However, a larger improvement in the P/F ratio occurred in NIRS patients (NIRS; 126 to 240 vs. IMV; 120 to 188) and fewer treatment days were required (NIRS; 7.86 vs. IMV; 10.5). Nebulized rt-PA appears to be well-tolerated, with a trend towards improved oxygenation, particularly in the NIRS group. Randomized clinical trials are required to demonstrate the clinical effect significance and magnitude.


Pediatric obstructive sleep apnea syndrome and anesthetic management.
Sleep-related breathing disorders require special attention in children who spend a considerable time sleeping. Obstructive sleep apnea syndrome is characterized by episodes of upper airway obstruction during sleep. Symptoms include hyperactivity, enuresis, headache, failure to thrive, and increased respiratory effort and total sleep time. The most common cause is adenotonsillar hypertrophy. Coexisting diseases are obesity, neuromuscular and craniofacial anomalies, and Down's syndrome. Early diagnosis is important to minimize neurocognitive, cardiac and developmental complications. Polysomnography is the gold standard for diagnosis. Although the features of pediatric obstructive sleep apnea syndrome are distinctly different from that in adults, it may predispose to the adult type of the syndrome. As therapy concerns several surgical approaches as well as conservative techniques, anesthetic management calls for particular attention. Pre- and postoperative sedation must be performed cautiously and patients must be watched closely with respect to airway obstruction and hypoventilation. Difficult intubation must always be considered.


Efficient site-specific integration in Plasmodium falciparum chromosomes mediated by mycobacteriophage Bxb1 integrase.
Here we report an efficient, site-specific system of genetic integration into Plasmodium falciparum malaria parasite chromosomes. This is mediated by mycobacteriophage Bxb1 integrase, which catalyzes recombination between an incoming attP and a chromosomal attB site. We developed P. falciparum lines with the attB site integrated into the glutaredoxin-like cg6 gene. Transfection of these attB(+) lines with a dual-plasmid system, expressing a transgene on an attP-containing plasmid together with a drug resistance gene and the integrase on a separate plasmid, produced recombinant parasites within 2 to 4 weeks that were genetically uniform for single-copy plasmid integration. Integrase-mediated recombination resulted in proper targeting of parasite proteins to intra-erythrocytic compartments, including the apicoplast, a plastid-like organelle. Recombinant attB x attP parasites were genetically stable in the absence of drug and were phenotypically homogeneous. This system can be exploited for rapid genetic integration and complementation analyses at any stage of the P. falciparum life cycle, and it illustrates the utility of Bxb1-based integrative recombination for genetic studies of intracellular eukaryotic organisms.


Beyond co-localization: inferring spatial interactions between sub-cellular structures from microscopy images.
Sub-cellular structures interact in numerous direct and indirect ways in order to fulfill cellular functions. While direct molecular interactions crucially depend on spatial proximity, other interactions typically result in spatial correlations between the interacting structures. Such correlations are the target of microscopy-based co-localization analysis, which can provide hints of potential interactions. Two complementary approaches to co-localization analysis can be distinguished: intensity correlation methods capitalize on pattern discovery, whereas object-based methods emphasize detection power. We first reinvestigate the classical co-localization measure in the context of spatial point pattern analysis. This allows us to unravel the set of implicit assumptions inherent to this measure and to identify potential confounding factors commonly ignored. We generalize object-based co-localization analysis to a statistical framework involving spatial point processes. In this framework, interactions are understood as position co-dependencies in the observed localization patterns. The framework is based on a model of effective pairwise interaction potentials and the specification of a null hypothesis for the expected pattern in the absence of interaction. Inferred interaction potentials thus reflect all significant effects that are not explained by the null hypothesis. Our model enables the use of a wealth of well-known statistical methods for analyzing experimental data, as demonstrated on synthetic data and in a case study considering virus entry into live cells. We show that the classical co-localization measure typically under-exploits the information contained in our data. We establish a connection between co-localization and spatial interaction of sub-cellular structures by formulating the object-based interaction analysis problem in a spatial statistics framework based on nearest-neighbor distance distributions. We provide generic procedures for inferring interaction strengths and quantifying their relative statistical significance from sets of discrete objects as provided by image analysis methods. Within our framework, an interaction potential can either refer to a phenomenological or a mechanistic model of a physico-chemical interaction process. This increased flexibility in designing and testing different hypothetical interaction models can be used to quantify the parameters of a specific interaction model or may catalyze the discovery of functional relations.


Age-related changes in the cellular composition and epithelial organization of the mouse trachea.
We report here senescent changes in the structure and organization of the mucociliary pseudostratified epithelium of the mouse trachea and main stem bronchi. We confirm previous reports of the gradual appearance of age-related, gland-like structures (ARGLS) in the submucosa, especially in the intercartilage regions and carina. Immunohistochemistry shows these structures contain ciliated and secretory cells and Krt5+ basal cells, but not the myoepithelial cells or ciliated ducts typical of normal submucosal glands. Data suggest they arise de novo by budding from the surface epithelium rather than by delayed growth of rudimentary or cryptic submucosal glands. In old mice the surface epithelium contains fewer cells per unit length than in young mice and the proportion of Krt5+, p63+ basal cells is reduced in both males and females. However, there appears to be no significant difference in the ability of basal stem cells isolated from individual young and old mice to form clonal tracheospheres in culture or in the ability of the epithelium to repair after damage by inhaled sulfur dioxide. Gene expression analysis by Affymetrix microarray and quantitative PCR, as well as immunohistochemistry and flow sorting studies, are consistent with low-grade chronic inflammation in the tracheas of old versus young mice and an increase in the number of immune cells. The significance of these changes for ARGL formation are not clear since several treatments that induce acute inflammation in young mice did not result in budding of the surface epithelium.


Transcriptional responses underlying the hormetic and detrimental effects of the plant secondary metabolite gossypol on the generalist herbivore Helicoverpa armigera.
Hormesis is a biphasic biological response characterized by the stimulatory effect at relatively low amounts of chemical compounds which are otherwise detrimental at higher concentrations. A hormetic response in larval growth rates has been observed in cotton-feeding insects in response to increasing concentrations of gossypol, a toxic metabolite found in the pigment glands of some plants in the family Malvaceae. We investigated the developmental effect of gossypol in the cotton bollworm, Helicoverpa armigera, an important heliothine pest species, by exposing larvae to different doses of this metabolite in their diet. In addition, we sought to determine the underlying transcriptional responses to different gossypol doses. Larval weight gain, pupal weight and larval development time were measured in feeding experiments and a hormetic response was seen for the first two characters. On the basis of net larval weight gain responses to gossypol, three concentrations (0%, 0.016% and 0.16%) were selected for transcript profiling in the gut and the rest of the body in a two-color double reference design microarray experiment. Hormesis could be observed at the transcript level, since at the low gossypol dose, genes involved in energy acquisition such as β-fructofuranosidases were up-regulated in the gut, and genes involved in cell adhesion were down-regulated in the body. Genes with products predicted to be integral to the membrane or associated with the proteasome core complex were significantly affected by the detrimental dose treatment in the body. Oxidoreductase activity-related genes were observed to be significantly altered in both tissues at the highest gossypol dose. This study represents the first transcriptional profiling approach investigating the effects of different concentrations of gossypol in a lepidopteran species. H. armigera's transcriptional response to gossypol feeding is tissue- and dose-dependent and involves diverse detoxifying mechanisms not only to alleviate direct effects of gossypol but also indirect damage such as pH disturbance and oxygen radical formation. Genes discovered through this transcriptional approach may be additional candidates for understanding gossypol detoxification and coping with gossypol-induced stress. In a generalist herbivore that has evolved transcriptionally-regulated responses to a variety of different plant compounds, hormesis may be due to a lower induction threshold of growth-promoting, stress-coping responses and a higher induction threshold of detoxification pathways that are costly and cause collateral damage to the cell.


Sensititre MYCOTB MIC plate for testing Mycobacterium tuberculosis susceptibility to first- and second-line drugs.
For Mycobacterium tuberculosis, phenotypic methods for drug susceptibility testing of second-line drugs are poorly standardized and technically challenging. The Sensititre MYCOTB MIC plate (MYCOTB) is a microtiter plate containing lyophilized antibiotics and configured for determination of MICs to first- and second-line antituberculosis drugs. To evaluate the performance of MYCOTB for M. tuberculosis drug susceptibility testing using the Middlebrook 7H10 agar proportion method (APM) as the comparator, we conducted a two-site study using archived M. tuberculosis isolates from Uganda and the Republic of Korea. Thawed isolates were subcultured, and dilutions were inoculated into MYCOTB wells and onto 7H10 agar. MYCOTB results were read at days 7, 10, 14, and 21; APM results were read at 21 days. A total of 222 isolates provided results on both platforms. By APM, 106/222 (47.7%) of isolates were resistant to at least isoniazid and rifampin. Agreement between MYCOTB and APM with respect to susceptibility or resistance was ≥92% for 7 of 12 drugs when a strict definition was used and ≥96% for 10 of 12 drugs when agreement was defined by allowing a ± one-well range of dilutions around the APM critical concentration. For ethambutol, agreement was 80% to 81%. For moxifloxacin, agreement was 83% to 85%; incorporating existing DNA sequencing information for discrepant analysis raised agreement to 91% to 96%. For MYCOTB, the median time to plate interpretation was 10 days and interreader agreement was ≥95% for all drugs. MYCOTB provided reliable results for M. tuberculosis susceptibility testing of first- and second-line drugs except ethambutol, and results were available sooner than those determined by APM.


New genetic variants improve personalized breast cancer diagnosis.
Recent large-scale genome-wide association studies (GWAS) have identified a number of new genetic variants associated with breast cancer. However, the degree to which these genetic variants improve breast cancer diagnosis in concert with mammography remains unknown. We conducted a case-control study and collected mammography features and 77 genetic variants which reflect the state of the art GWAS findings on breast cancer. A naïve Bayes model was developed on the mammography features and these genetic variants. We observed that the incorporation of the genetic variants significantly improved breast cancer diagnosis based on mammographic findings.


Chronic spinal pain: a randomized clinical trial comparing medication, acupuncture, and spinal manipulation.
A randomized controlled clinical trial was conducted. To compare medication, needle acupuncture, and spinal manipulation for managing chronic (>13 weeks duration) spinal pain because the value of medicinal and popular forms of alternative care for chronic spinal pain syndromes is uncertain. Between February 1999 and October 2001, 115 patients without contraindication for the three treatment regimens were enrolled at the public hospital's multidisciplinary spinal pain unit. One of three separate intervention protocols was used: medication, needle acupuncture, or chiropractic spinal manipulation. Patients were assessed before treatment by a sports medical physician for exclusion criteria and by a research assistant using the Oswestry Back Pain Disability Index (Oswestry), the Neck Disability Index (NDI), the Short-Form-36 Health Survey questionnaire (SF-36), visual analog scales (VAS) of pain intensity and ranges of movement. These instruments were administered again at 2, 5, and 9 weeks after the beginning of treatment. Randomization proved to be successful. The highest proportion of early (asymptomatic status) recovery was found for manipulation (27.3%), followed by acupuncture (9.4%) and medication (5%). Manipulation achieved the best overall results, with improvements of 50% (P = 0.01) on the Oswestry scale, 38% (P = 0.08) on the NDI, 47% (P < 0.001) on the SF-36, and 50% (P < 0.01) on the VAS for back pain, 38% (P < 0.001) for lumbar standing flexion, 20% (P < 0.001) for lumbar sitting flexion, 25% (P = 0.1) for cervical sitting flexion, and 18% (P = 0.02) for cervical sitting extension. However, on the VAS for neck pain, acupuncture showed a better result than manipulation (50% vs 42%). The consistency of the results provides, despite some discussed shortcomings of this study, evidence that in patients with chronic spinal pain, manipulation, if not contraindicated, results in greater short-term improvement than acupuncture or medication. However, the data do not strongly support the use of only manipulation, only acupuncture, or only nonsteroidal antiinflammatory drugs for the treatment of chronic spinal pain. The results from this exploratory study need confirmation from future larger studies.


Mixed cryoglobulinemia.
Mixed cryoglobulinemia (MC), type II and type III, refers to the presence of circulating cryoprecipitable immune complexes in the serum and manifests clinically by a classical triad of purpura, weakness and arthralgias. It is considered to be a rare disorder, but its true prevalence remains unknown. The disease is more common in Southern Europe than in Northern Europe or Northern America. The prevalence of 'essential' MC is reported as approximately 1:100,000 (with a female-to-male ratio 3:1), but this term is now used to refer to a minority of MC patients only. MC is characterized by variable organ involvement including skin lesions (orthostatic purpura, ulcers), chronic hepatitis, membranoproliferative glomerulonephritis, peripheral neuropathy, diffuse vasculitis, and, less frequently, interstitial lung involvement and endocrine disorders. Some patients may develop lymphatic and hepatic malignancies, usually as a late complication. MC may be associated with numerous infectious or immunological diseases. When isolated, MC may represent a distinct disease, the so-called 'essential' MC. The etiopathogenesis of MC is not completely understood. Hepatitis C virus (HCV) infection is suggested to play a causative role, with the contribution of genetic and/or environmental factors. Moreover, MC may be associated with other infectious agents or immunological disorders, such as human immunodeficiency virus (HIV) infection or primary Sjögren's syndrome. Diagnosis is based on clinical and laboratory findings. Circulating mixed cryoglobulins, low C4 levels and orthostatic skin purpura are the hallmarks of the disease. Leukocytoclastic vasculitis involving medium- and, more often, small-sized blood vessels is the typical pathological finding, easily detectable by means of skin biopsy of recent vasculitic lesions. Differential diagnoses include a wide range of systemic, infectious and neoplastic disorders, mainly autoimmune hepatitis, Sjögren's syndrome, polyarthritis, and B-cell lymphomas. The first-line treatment of MC should focus on eradication of HCV by combined interferon-ribavirin treatment. Pathogenetic treatments (immunosuppressors, corticosteroids, and/or plasmapheresis) should be tailored to each patient according to the progression and severity of the clinical manifestations. Long-term monitoring is recommended in all MC patients to assure timely diagnosis and treatment of the life-threatening complications. The overall prognosis is poorer in patients with renal disease, liver failure, lymphoproliferative disease and malignancies.


Solid phase extraction of N-linked glycopeptides using hydrazide tip.
Glycoproteome contains valuable information where biomarkers may be discovered for disease diagnosis and monitoring. Nowadays, with the ever-increasing performances of mass spectrometers, the emphasis is shifting to the sample preparation for better throughput and reproducibility. Therefore, to facilitate high throughput N-linked glycopeptide isolation, in this study, a novel hydrazide tip was devised and an integrated workflow of N-linked glycopeptide isolation using hydrazide tips was presented. With the use of bovine fetuin as a standard glycoprotein, the incubation time was determined for each major step of glycopeptide isolation. With the use of commercially available human serum, multiple parallel isolations of glycopeptides were performed using hydrazide tips with a liquid handling robotic system. We demonstrated that, with the hydrazide tips, the processing time was significantly decreased from 3 to 4 days to less than 8 h with excellent reproducibility. The hydrazide pipet tips have great potential in achieving automation of N-linked glycopeptide isolation for high-throughput sample preparation when used in combination with liquid handling robotic systems.


Two-trace model for spike-timing-dependent synaptic plasticity.
We present an effective model for timing-dependent synaptic plasticity (STDP) in terms of two interacting traces, corresponding to the fraction of activated NMDA receptors and the [Formula: see text] concentration in the dendritic spine of the postsynaptic neuron. This model intends to bridge the worlds of existing simplistic phenomenological rules and highly detailed models, thus constituting a practical tool for the study of the interplay of neural activity and synaptic plasticity in extended spiking neural networks. For isolated pairs of pre- and postsynaptic spikes, the standard pairwise STDP rule is reproduced, with appropriate parameters determining the respective weights and timescales for the causal and the anticausal contributions. The model contains otherwise only three free parameters, which can be adjusted to reproduce triplet nonlinearities in hippocampal culture and cortical slices. We also investigate the transition from time-dependent to rate-dependent plasticity occurring for both correlated and uncorrelated spike patterns.


Drosophila E2f2 promotes the conversion from genomic DNA replication to gene amplification in ovarian follicle cells.
Drosophila contains two members of the E2F transcription factor family (E2f and E2f2), which controls the expression of genes that regulate the G1-S transition of the cell cycle. Previous genetic analyses have indicated that E2f is an essential gene that stimulates DNA replication. We show that loss of E2f2 is viable, but causes partial female sterility associated with changes in the mode of DNA replication in the follicle cells that surround the developing oocyte. Late in wild-type oogenesis, polyploid follicle cells terminate a program of asynchronous endocycles in which the euchromatin is entirely replicated, and then confine DNA synthesis to the synchronous amplification of specific loci, including two clusters of chorion genes that encode eggshell proteins. E2f2 mutant follicle cells terminate endocycles on schedule, but then fail to confine DNA synthesis to sites of gene amplification and inappropriately begin genomic DNA replication. This ectopic DNA synthesis does not represent a continuation of the endocycle program, as the cells do not complete an entire additional S phase. E2f2 mutant females display a 50% reduction in chorion gene amplification, and lay poorly viable eggs with a defective chorion. The replication proteins ORC2, CDC45L and ORC5, which in wild-type follicle cell nuclei localize to sites of gene amplification, are distributed throughout the entire follicle cell nucleus in E2f2 mutants, consistent with their use at many genomic replication origins rather than only at sites of gene amplification. RT-PCR analyses of RNA purified from E2f2 mutant follicle cells indicate an increase in the level of Orc5 mRNA relative to wild type. These data indicate that E2f2 functions to inhibit widespread genomic DNA synthesis in late stage follicle cells, and may do so by repressing the expression of specific components of the replication machinery.


Jules Bernard Luys in Charcot's penumbra.
Jules Bernard Luys (1828-1897) is a relatively unknown figure in 19th century French neuropsychiatry. Although greatly influenced by Jean-Martin Charcot (1825-1893), Luys worked in the shadow of the 'master of La Salpêtrière' for about a quarter of a century. When he arrived at this institution in 1862, he used microscopy and photomicrography to identify pathological lesions underlying locomotor ataxia and progressive muscular atrophy. He later made substantial contributions to our knowledge of normal human brain anatomy, including the elucidation of thalamic organization and the discovery of the subthalamic nucleus. Luys's name has long been attached to the latter structure (corps de Luys), which is at the center of our current thinking about the functional organization of basal ganglia and the physiopathology of Parkinson's disease. As head of the Maison de santé d'Ivry, Luys developed a highly original view of the functional organization of the normal human brain, while improving our understanding of the neuropathological and clinical aspects of mental illnesses. In 1886, Luys left La Salpêtrière and became chief physician at La Charité hospital. Following Charcot, whom he considered as the father of scientific hypnotism, Luys devoted the last part of his career to hysteria and hypnosis. However, Luys ventured too deeply into the minefield of hysteria. He initiated experiments as unconventional as the distant action of medication, and became one of the most highly caricatured examples of the fascination that hysteria exerted upon neurologists as well as laypersons at the end of the 19th century.


A sensor fusion method for tracking vertical velocity and height based on inertial and barometric altimeter measurements.
A sensor fusion method was developed for vertical channel stabilization by fusing inertial measurements from an Inertial Measurement Unit (IMU) and pressure altitude measurements from a barometric altimeter integrated in the same device (baro-IMU). An Extended Kalman Filter (EKF) estimated the quaternion from the sensor frame to the navigation frame; the sensed specific force was rotated into the navigation frame and compensated for gravity, yielding the vertical linear acceleration; finally, a complementary filter driven by the vertical linear acceleration and the measured pressure altitude produced estimates of height and vertical velocity. A method was also developed to condition the measured pressure altitude using a whitening filter, which helped to remove the short-term correlation due to environment-dependent pressure changes from raw pressure altitude. The sensor fusion method was implemented to work on-line using data from a wireless baro-IMU and tested for the capability of tracking low-frequency small-amplitude vertical human-like motions that can be critical for stand-alone inertial sensor measurements. Validation tests were performed in different experimental conditions, namely no motion, free-fall motion, forced circular motion and squatting. Accurate on-line tracking of height and vertical velocity was achieved, giving confidence to the use of the sensor fusion method for tracking typical vertical human motions: velocity Root Mean Square Error (RMSE) was in the range 0.04-0.24 m/s; height RMSE was in the range 5-68 cm, with statistically significant performance gains when the whitening filter was used by the sensor fusion method to track relatively high-frequency vertical motions.


Is the Addition of Sublingual Melatonin to Omeprazole Superior to Omeprazole Alone in the Management of Gastroesophageal Reflux Disease Symptoms: A Clinical Trial.
Proton pump inhibitors are frequently used to treat gastroesophageal reflux disease, but their effect is restricted. The present study aimed to investigate whether the addition of sublingual melatonin to omeprazole was effective in the treatment of gastro gastroesophageal reflux disease symptoms. This was a randomized double-blind clinical trial. A total of 78 patients with gastro gastroesophageal reflux disease were randomly allocated to either omeprazole 20 mg/d plus sublingual melatonin (3 mg/d) or omeprazole 20 mg/d plus placebo for 4 weeks. The selected patients had histories of heartburn and regurgitation and a score ≤32 on the Frequency Scale for the Symptoms of gastroesophageal reflux disease (FSSG). The outcome measures for the assessment of treatment efficacy were heartburn, epigastric pain and the Frequency Scale for the Symptoms of gastroesophageal reflux disease score. Safety and quality of life were evaluated in the patients as the secondary outcomes too. Seventy-two out of 78 eligible patients completed this trial (35 in the melatonin group and 37 in the placebo group). Heartburn, epigastric pain, and Frequency Scale for the Symptoms of gastroesophageal reflux disease score declined significantly in the melatonin group compared to the placebo group (P = .04, P = .03, and P = .0001, respectively). Moreover, the quality of life score was significantly higher in the melatonin group compared with the placebo group (P = .0001). Adverse events were similarly observed in the 2 groups (P = .55), and there were no serious adverse events. The combination of sublingual melatonin (3 mg/day) with omeprazole (20 mg/day) may be more effective than omeprazole (20 mg/day) alone in the treatment of gastroesophageal reflux disease.


Theoretical biology in the third millennium.
During the 20th century our understanding of genetics and the processes of gene expression have undergone revolutionary change. Improved technology has identified the components of the living cell, and knowledge of the genetic code allows us to visualize the pathway from genotype to phenotype. We can now sequence entire genes, and improved cloning techniques enable us to transfer genes between organisms, giving a better understanding of their function. Due to the improved power of analytical tools databases of sequence information are growing at an exponential rate. Soon complete sequences of genomes and the three-dimensional structure of all proteins may be known. The question we face in the new millennium is how to apply this data in a meaningful way. Since the genes carry the specification of an organism, and because they also record evolutionary changes, we need to design a theoretical framework that can take account of the flow of information through biological systems.


Investigation of neurokinin-2 and -3 receptors in the human and pig bladder.
To investigate the role of neurokinin (NK)-2 and -3 receptors in mediating the contraction of detrusor muscle strips from human and pig, to determine whether the pig is a good model for the study of tachykinin receptors in the human bladder, as the biological actions of tachykinins, e.g. substance P and NKA are mediated via three distinct receptor subtypes, NK-1, -2 and -3. Strips of detrusor muscle were obtained from the bladder dome and neck of female pigs and from patients undergoing cystectomy. Cumulative concentration-response curves to NKA were obtained in the absence and presence of either the NK-2 receptor-selective antagonist SR48968 or the NK-3 receptor-selective antagonist SB223412. NKA produced concentration-dependent contractions in the human and pig detrusor muscle; the curves were shifted to the right by SR48968, with high affinity (pKB 8.9, 8.3 and 8.0 in the human, pig dome and pig neck, respectively), whereas SB223412 had a minimal effect (pKB 5.8, 5.8 and 6.3, respectively). These data confirm that the NK-2 receptor subtype mediates NKA-induced contraction of the human and pig detrusor muscle. The NK-3 receptor appears to have no role in detrusor contraction of either species. The results also provide evidence that the NK-2 receptor in human and pig are the same, and the latter may be an appropriate species to study tachykinin-induced contractions in human bladder.


nab-Paclitaxel-Based Therapy in Underserved Patient Populations: The ABOUND.PS2 Study in Patients With NSCLC and a Performance Status of 2.
The phase II ABOUND.PS2 study (NCT02289456) assessed safety/tolerability of a first-line modified nab-paclitaxel/carboplatin regimen for patients with advanced non-small cell lung cancer (NSCLC) and Eastern Cooperative Oncology Group (ECOG) performance status (PS) 2. Chemotherapy-naive patients with stage IIIB/IV NSCLC and ECOG PS 2 received four cycles of nab-paclitaxel 100 mg/m2 days 1 and 8 plus carboplatin area under the curve 5 day 1 q3w (induction). Patients without progression received nab-paclitaxel monotherapy (100 mg/m2 days 1 and 8 q3w) until progression/unacceptable toxicity. Primary endpoint: percentage of patients discontinuing induction due to treatment-emergent adverse events (TEAEs). 11/40 treated patients (27.5%; 95% CI, 14.60-43.89) discontinued chemotherapy induction due to TEAEs; 16/40 (40.0%) continued nab-paclitaxel monotherapy. Median progression-free and overall survival were 4.4 (95% CI, 2.99-7.00) and 7.7 (95% CI, 4.93-13.17) months. Grade 3/4 TEAEs during induction included neutropenia (22.5%), anemia (17.5%), thrombocytopenia (5.0%), and peripheral neuropathy (2.5%). This nab-paclitaxel-based regimen was tolerable in patients with advanced NSCLC and ECOG PS 2, with efficacy comparable to historical chemotherapy data.


A classification approach to coreference in discharge summaries: 2011 i2b2 challenge.
To create a highly accurate coreference system in discharge summaries for the 2011 i2b2 challenge. The coreference categories include Person, Problem, Treatment, and Test. An integrated coreference resolution system was developed by exploiting Person attributes, contextual semantic clues, and world knowledge. It includes three subsystems: Person coreference system based on three Person attributes, Problem/Treatment/Test system based on numerous contextual semantic extractors and world knowledge, and Pronoun system based on a multi-class support vector machine classifier. The three Person attributes are patient, relative and hospital personnel. Contextual semantic extractors include anatomy, position, medication, indicator, temporal, spatial, section, modifier, equipment, operation, and assertion. The world knowledge is extracted from external resources such as Wikipedia. Micro-averaged precision, recall and F-measure in MUC, BCubed and CEAF were used to evaluate results. The system achieved an overall micro-averaged precision, recall and F-measure of 0.906, 0.925, and 0.915, respectively, on test data (from four hospitals) released by the challenge organizers. It achieved a precision, recall and F-measure of 0.905, 0.920 and 0.913, respectively, on test data without Pittsburgh data. We ranked the first out of 20 competing teams. Among the four sub-tasks on Person, Problem, Treatment, and Test, the highest F-measure was seen for Person coreference. This system achieved encouraging results. The Person system can determine whether personal pronouns and proper names are coreferent or not. The Problem/Treatment/Test system benefits from both world knowledge in evaluating the similarity of two mentions and contextual semantic extractors in identifying semantic clues. The Pronoun system can automatically detect whether a Pronoun mention is coreferent to that of the other four types. This study demonstrates that it is feasible to accomplish the coreference task in discharge summaries.


Induction chemotherapy with mitomycin, epirubicin, cisplatin, fluorouracil, and leucovorin followed by radiotherapy in the treatment of locoregionally advanced nasopharyngeal carcinoma.
Survival in advanced nasopharyngeal carcinoma (NPC) is compromised by distant metastasis. Because mitomycin is active against hypoxic and G0 cells, which may help to eradicate micrometastasis, we investigated the effect of mitomycin-containing cisplatin-based induction chemotherapy. Recruited for this study were American Joint Committee on Cancer (AJCC) 1992 staging system stage IV NPC patients with the following adverse features: obvious intracranial invasion, supraclavicular or bilateral neck lymph node metastasis, large neck node (> 6 cm), or elevated serum lactate dehydrogenase (LDH) level. Patients were given three cycles of chemotherapy before radiotherapy. The chemotherapy comprised a 3-week cycle of mitomycin, epirubicin, and cisplatin on day 1 and fluorouracil and leucovorin on day 8 (MEPFL). From January 1994 to December 1997, 111 patients were recruited. The median follow-up period was 43 months. The actuarial 5-year overall survival rate was 70% (95% confidence interval [CI], 60% to 80%; n = 111). For patients having completed radiotherapy (n = 100), the 5-year locoregional control rate was 70% (95% CI, 55% to 84%) and the distant metastasis-free rate was 81% (95% CI, 73% to 89%). The 5-year distant metastasis-free rate of N3a and N3b disease of AJCC 1997 staging system were 79% (95% CI, 62% to 95%) and 74% (95% CI, 60% to 89%), respectively. By Cox multivariate analysis, high pretreatment serum LDH level (P = .04) and neck nodal enlargement before radiotherapy (P = .001) were adverse prognostic factors of survival. The good 5-year survival of N3 disease supports the effectiveness of induction MEPFL in the primary treatment of advanced NPC. Further investigation to incorporate concurrent chemoradiotherapy is warranted.


Long-Term Trends and Role of Climate in the Population Dynamics of Eurasian Reindeer.
Temperature is increasing in Arctic and sub-Arctic regions at a higher rate than anywhere else in the world. The frequency and nature of precipitation events are also predicted to change in the future. These changes in climate are expected, together with increasing human pressures, to have significant impacts on Arctic and sub-Arctic species and ecosystems. Due to the key role that reindeer play in those ecosystems, it is essential to understand how climate will affect the region's most important species. Our study assesses the role of climate on the dynamics of fourteen Eurasian reindeer (Rangifer tarandus) populations, using for the first time data on reindeer abundance collected over a 70-year period, including both wild and semi-domesticated reindeer, and covering more than half of the species' total range. We analyzed trends in population dynamics, investigated synchrony among population growth rates, and assessed the effects of climate on population growth rates. Trends in the population dynamics were remarkably heterogeneous. Synchrony was apparent only among some populations and was not correlated with distance among population ranges. Proxies of climate variability mostly failed to explain population growth rates and synchrony. For both wild and semi-domesticated populations, local weather, biotic pressures, loss of habitat and human disturbances appear to have been more important drivers of reindeer population dynamics than climate. In semi-domesticated populations, management strategies may have masked the effects of climate. Conservation efforts should aim to mitigate human disturbances, which could exacerbate the potentially negative effects of climate change on reindeer populations in the future. Special protection and support should be granted to those semi-domesticated populations that suffered the most because of the collapse of the Soviet Union, in order to protect the livelihood of indigenous peoples that depend on the species, and the multi-faceted role that reindeer exert in Arctic ecosystems.


Nocardioides opuntiae sp. nov., isolated from soil of a cactus.
A novel high G+C actinobacterium, designated strain OS1-21(T), was isolated from the rhizosphere soil of a cactus (Opuntia fiscus-indica var. sanboten) and the taxonomic status was investigated using a polyphasic approach. Cells of strain OS1-21(T) were aerobic, Gram-stain-positive, non-endospore-forming, non-motile rods; colonies of the cells were circular, translucent, smooth and moderate yellow in colour. LL-Diaminopimelic acid was the diagnostic diamino acid in cell-wall peptidoglycan. The predominant menaquinone was MK-8(H4). The major fatty acids were iso-C(16 : 0), iso-C(16 : 0) 2-OH, 10-methyl C(17 : 0), 10-methyl C(18 : 0) and C(17 : 1)cis9. The polar lipids contained diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and two unknown phospholipids. The DNA G+C content was 73.7 mol%. Based on 16S rRNA gene sequence analysis, the closest phylogenetic neighbours were Nocardioides panacihumi Gsoil 616(T) (98.7% sequence similarity) and Nocardioides terrae VA15(T) (97.8%), followed by Nocardioides marinus CL-DD14(T) (97.1%). DNA-DNA relatedness values of strain OS1-21(T) with the type strains of the closest phylogenetic neighbours were low (<16.0%). Combined data of polyphasic taxonomic analyses revealed that the organism could be assigned to a novel species of the genus Nocardioides, for which the name Nocardioides opuntiae sp. nov. is proposed. The type strain is OS1-21(T) ( = KCTC 19804(T) = NBRC 107915(T)).


Genome-wide catalogue of chromosomal aberrations in barrett's esophagus and esophageal adenocarcinoma: a high-density single nucleotide polymorphism array analysis.
To better understand the molecular mechanisms behind esophageal adenocarcinoma (EAC) tumorigenesis, we used high-density single nucleotide polymorphism arrays to profile chromosomal aberrations at each of the four sequential progression stages, Barrett's metaplasia (BM), low-grade dysplasia (LGD), high-grade dysplasia (HGD), and EAC, in 101 patients. We observed a significant trend toward increasing loss of chromosomes with higher progression stage. For BM, LGD, HGD, and EAC, respectively, the average numbers of chromosome arms with loss per sample were 0.30, 3.21, 7.70, and 11.90 (P for trend = 4.82 x 10(-7)), and the mean percentages of single nucleotide polymorphisms with allele loss were 0.1%, 1.8%, 6.6%, and 17.2% (P for trend = 2.64 x 10(-6)). In LGD, loss of 3p14.2 (68.4%) and 16q23.1 (47.4%) was limited to narrow regions within the FHIT (3p14.2) and WWOX (16q23.1) genes, whereas loss of 9p21 (68.4%) occurred in larger regions. A significant increase in the loss of other chromosomal regions was seen in HGD and EAC. Loss of 17p (47.6%) was one of the most frequent events in EAC. Many recurrent small regions of chromosomal loss disrupted single genes, including FHIT, WWOX, RUNX1, KIF26B, MGC48628, PDE4D, C20orf133, GMDS, DMD, and PARK2, most of which are common fragile site regions in the human genome. However, RUNX1 at 21q22 seemed to be a potential tumor suppressor gene in EAC. Amplifications were less frequent than losses and mostly occurred in EAC. 8q24 (containing Myc) and 8p23.1 (containing CTSB) were the two most frequently amplified regions. In addition, a significant trend toward increasing amplification was associated with higher progression stage.


Activation and function of the MAPKs and their substrates, the MAPK-activated protein kinases.
The mitogen-activated protein kinases (MAPKs) regulate diverse cellular programs by relaying extracellular signals to intracellular responses. In mammals, there are more than a dozen MAPK enzymes that coordinately regulate cell proliferation, differentiation, motility, and survival. The best known are the conventional MAPKs, which include the extracellular signal-regulated kinases 1 and 2 (ERK1/2), c-Jun amino-terminal kinases 1 to 3 (JNK1 to -3), p38 (α, β, γ, and δ), and ERK5 families. There are additional, atypical MAPK enzymes, including ERK3/4, ERK7/8, and Nemo-like kinase (NLK), which have distinct regulation and functions. Together, the MAPKs regulate a large number of substrates, including members of a family of protein Ser/Thr kinases termed MAPK-activated protein kinases (MAPKAPKs). The MAPKAPKs are related enzymes that respond to extracellular stimulation through direct MAPK-dependent activation loop phosphorylation and kinase activation. There are five MAPKAPK subfamilies: the p90 ribosomal S6 kinase (RSK), the mitogen- and stress-activated kinase (MSK), the MAPK-interacting kinase (MNK), the MAPK-activated protein kinase 2/3 (MK2/3), and MK5 (also known as p38-regulated/activated protein kinase [PRAK]). These enzymes have diverse biological functions, including regulation of nucleosome and gene expression, mRNA stability and translation, and cell proliferation and survival. Here we review the mechanisms of MAPKAPK activation by the different MAPKs and discuss their physiological roles based on established substrates and recent discoveries.


Gatekeeping versus promiscuity in the early stages of the andrimid biosynthetic assembly line.
The antibiotic andrimid, a nanomolar inhibitor of bacterial acetyl coenzyme A carboxylase, is generated on an unusual polyketide/nonribosomal peptide enzyme assembly line in that all thiolation (T) domains/small-molecule building stations are on separate proteins. In addition, a transglutaminase homologue is used to condense andrimid building blocks together on the andrimid assembly line. The first two modules of the andrimid assembly line yields an octatrienoyl-beta-Phe-thioester tethered to the AdmI T domain, with amide bond formation carried out by a free-standing transglutaminase homologue AdmF. Analysis of the aminomutase AdmH reveals its specific conversion from l-Phe to (S)-beta-Phe, which in turn is activated by AdmJ and ATP to form (S)-beta-Phe-aminoacyl-AMP. AdmJ then transfers the (S)-beta-Phe moiety to one of the free-standing T domains, AdmI, but not AdmA, which instead gets loaded with an octatrienoyl group by other enzymes. AdmF, the amide synthase, will accept a variety of acyl groups in place of the octatrienoyl donor if presented on either AdmA or AdmI. AdmF will also use either stereoisomer of phenylalanine or beta-Phe when presented on AdmA and AdmI, but not when placed on noncognate T domains. Further, we show the polyketide synthase proteins responsible for the polyunsaturated acyl cap can be bypassed in vitro with N-acetylcysteamine as a low-molecular-weight acyl donor to AdmF and also in vivo in an Escherichia coli strain bearing the andrimid biosynthetic gene cluster with a knockout in admA.


Coronary rehabilitation in the community.
Over a five-year period, 162 patients with coronary disease joined a physical rehabilitation course at a community sports centre gymnasium under the supervision of a general practitioner. One hundred and forty-seven patients had suffered a recent or old myocardial infarction and 15 suffered from angina pectoris. One hundred and thirty-eight patients (85 per cent) completed a three-month course of exercises, 16 (10 per cent) defaulted and eight (5 per cent) were withdrawn. One patient died at home during the three-month course. There were no changes in weight, blood pressure or blood fat measurements during the course but predicted maximum oxygen uptake increased by 26.9 per cent and the double product after effort (which is proportional to myocardial oxygen uptake) decreased by 13.6 per cent.We believe that the rehabilitation of patients in community sports centres is safe and effective and should be more widely practised.


Bill and Monica: memory, emotion and normativity in Clinton's Grand Jury testimony.
We examine links between factual recall, emotion and constructions of normativity in narrative accounts, using as an empirical case President Clinton's descriptions of his relationship with Monica Lewinsky. We analyse those accounts in the sequences of talk in which they occurred, under Grand Jury cross-examination. Clinton's accounts of Lewinsky were part of how he attended to issues alive in court concerning himself, including his possible exploitation and abuse of power in an asymmetrical relationship; his motives, sincerity, credibility and intentions; and, indirectly, his fitness for office as President. Analysis focuses on how Clinton's portrayal of Lewinsky accomplished a reflexive portrayal of himself, not as mendacious and exploitative, but as caring, responsible, sincere, rational and consistent, while reducing the scope and implications of their admitted sexual relationship. This study is linked to a broader discursive psychology of factual description, memory, mental and emotional states, and their relevance to the larger business of institutional settings.


Delineation of selective influences shaping the mutated expressed human Ig heavy chain repertoire.
After Ag exposure, somatic hypermutation and subsequent selection play significant roles in shaping the peripheral B cell repertoire. However, the disparate impact of each process has not been completely delineated. To address this, the mutational patterns of a large panel of productive V(H)DJ(H) rearrangements of individual human B cells were analyzed and compared with those of a previously reported panel of nonproductive V(H)DJ(H) rearrangements. The productive V(H) rearrangements exhibited a significantly lower mutational frequency and a significantly smaller number of replacement mutations than the nonproductively rearranged genes, suggesting that structural constraints of the Ig molecule and selective influences both impacted the repertoire, militating against replacement mutations. Positive selection favored a mean of four to six replacements in complementarity-determining region 1 (CDR1) and CDR2, and less than two replacements in the framework regions (FRs). In contrast, the negative impact of replacement mutations generated an increased number of silent mutations within both the CDRs and FRs of the productive repertoire accompanied by a net increase in the ratio of replacement to silent mutations in the CDRs compared with that in the FRs. Moreover, there was a negative influence on the distribution of amino acid changes resulting from mutations of highly mutable codons, such as AGY, TAY, GTA, and GCT, preferentially leading to conservative changes in the expressed Ig repertoire. The results are consistent with the conclusion that the expressed repertoire is limited, compared with the potential generated by the mutational machinery, by the dual requirements of avoiding autoreactivity and satisfying structural constraints of an intact Ig molecule.


Do models of care designed for terminally ill 'home alone' people improve their end-of-life experience? A patient perspective.
Palliative care patients who live alone report greater psychological distress, and are less likely to die at home than those living with a family carer. However, there is a lack of research on the value of models of care that specifically address this disadvantage. This article describes the experiences of terminally ill 'home alone' people using one of two models of care aimed at maintaining participants' need for independent living, focusing on the effect of these two models of care on their physical, social and emotional needs. Twenty six palliative care patients of Silver Chain Hospice Care, in Western Australia, were randomly assigned to either having a personal alarm or additional care-aide hours in their home. An in-depth qualitative study was conducted in two phases in 2010 using face-to-face interviews. The care-aide model of care resulted in benefits such as easing the burden of everyday living; supporting well-being; enhancing quality of life and preserving a sense of dignity; and reducing loneliness and isolation. The personal alarm model of care imparted a sense of security; provided peace of mind; and helped to deal with feelings of isolation. Participants in both groups felt that they could remain at home longer. By providing a safer, more secure environment through the use of a personal alarm or additional care-aide hours, patients were able to continue their activities of daily living, could build a sense of 'normality' into their lives, and they could live independently through support and dignity.


Training cognition in ADHD: current findings, borrowed concepts, and future directions.
With both its high prevalence and myriad of negative outcomes, Attention-Deficit/Hyperactivity Disorder (ADHD) demands a careful consideration of the efficacy of its treatment options. Although the benefits of medication have a robust empirical background, nonpharmaceutical interventions evoke particular interest, as they are often viewed more favorably by parents. This review pays special attention to the use of working memory and recent cognitive training attempts in ADHD, describing its cognitive, behavioral, and biological effects in relation to current neurological theory of the disorder. While these treatments have demonstrated positive effects on some measures, there are limitations, as studies have failed to demonstrate generalization to critical measures, such as teacher-rated classroom behaviors, and have provided limited but growing evidence of functionally significant improvements in behavior. There is also a clear lack of research on the effects of training on reward systems and self-control. These limitations may be addressed by broadening the scope and procedures of the training and incorporating research concepts from other fields of study. First, it is important to consider the developmental trajectories of brain regions in individuals with the disorder, as they may relate to the effectiveness of cognitive training. Notions from behavioral economics, including delay discounting and framing (i.e., context) manipulations that influence present orientation, also have applications in the study of cognitive training in ADHD. In considering these other domains, we may find new ways to conceptualize and enhance cognitive training in ADHD and, in turn, address current limitations of interventions that fall in this category.


Fall Risk and Its Associated Factors among Older Adults without Home-Help Services in a Swedish Municipality.
During preventive home visits, the purpose of this study was to identify the prevalence of fall risk and any associated factors. Participants (n = 1471) were cognitively sound community-dwelling older adults (≥ 70 years) without home-help service, living in a Swedish municipality. The Downton Fall Risk Index and nine single items were used. Tiredness/fatigue, age ≥ 80, inability to walk 1 hr, inability to climb stairs and worrying were significantly associated with fall risk. Preventive home visits incorporating fall-risk screening proved valuable, providing information for interventions aimed at preventing falls, maintaining independence, and facilitating health among community dwelling participants.


Differential regulation of IL-6 and TNF-alpha via calcineurin in human skeletal muscle cells.
Interleukin-6 increases in skeletal muscle during exercise, and evidence points to Ca2+ as an initiator of IL-6 production. However, the signalling pathway whereby this occurs is unknown. One candidate for Ca2+ -mediated IL-6 induction is calcineurin, an activator of NF-AT. Here we investigated whether skeletal myocytes produce IL-6 in a Ca2+/calcineurin-dependent manner, and whether TNF-alpha, an inducer of IL-6, is affected by these stimuli. Human skeletal muscle cell cultures were stimulated with ionomycin time-and dose-dependently to elevate intracellular Ca2+ levels, with or without addition of cyclosporin A (CSA); a calcineurin inhibitor. mRNA was extracted from myocytes and analysed for IL-6 and TNF-alpha gene expression. IL-6 mRNA increased time- and dose-dependently with ionomycin stimulation, an effect that was blunted by approximately 75% in the presence of CSA. In contrast, TNF-alpha gene expression was decreased by approximately 70% in response to ionomycin treatment, but increased in response to addition of CSA. These data demonstrate that IL-6 and TNF-alpha are regulated differentially in skeletal muscle cells in response to a Ca2+ stimulus. Blocking the calcineurin pathway resulted in inhibition of the IL-6 response to ionomycin, whereas TNF-alpha increased by addition of CSA, further indicating a differential regulation of IL-6 and TNF-alpha in human skeletal myocytes.


Molecular Shape Analysis-Guided Virtual Screening Platform for Adenosine Kinase Inhibitors.
We propose a new application of molecular shape descriptors in hierarchical selection during virtual screening (VS). Here, a structure-based pharmacophore and docking-guided VS protocol have been evolved to identify inhibitors against adenosine kinase (AK). The knowledge gained on the shape requirements has been extrapolated in classifying active and inactive molecules against this target. This classification enabled us to pick the appropriate ligand conformation in the binding site. We have suggested a set of hierarchical filters for VS, from a simple molecular shape analysis (MSA) descriptor-based recursive models to docking scores. This approach permits a systematic study to understand the importance of spatial requirements and limitations for inhibitors against AK. Finally, the guidelines on how to select compounds for AK to achieve success have been highlighted. The utility of this approach has been suggested by giving an example of database screening for plausible active compounds.


A randomised controlled trial of tidal irrigation vs corticosteroid injection in knee osteoarthritis: the KIVIS Study.
Patients with knee osteoarthritis (OA) often suffer pain that is not fully controlled by analgesics and often require intra-articular therapies. The aim of this study was to compare the benefits of intra-articular corticosteroid injections (CSIs) and tidal irrigation (TI) in patients with OA of the knee. We performed a dual-centre, single blind, randomised, parallel group trial comparing TI and CSI. Patients with knee OA were randomised to either irrigation using a 3.2mm arthroscope under local anaesthesia or an intra-articular injection of 40 mg triamcinolone acetonide and 1% lidocaine. Patients were followed for 6 months. The primary outcome measure was the Western Ontario and McMaster Universities OA Index total pain score (visual analogue scale, VAS). One hundred and fifty patients were recruited of whom 71 received TI and 79 CSI. In both treatment groups, over 80% of patients reported improvement at 2 and 4 weeks. After this time, the benefit of CSI decreased whereas that of TI was maintained: at 26 weeks the pain relief afforded by TI was significantly greater than that of CSI. At 26 weeks 29% of the CSI group reported improvement vs 64% of the TI group (P<0.001). Patients with a knee effusion responded better to both treatments, however, this was most apparent for CSI. Patients with less severe radiographic OA also obtained the greatest improvement from both treatments. Both procedures lead to significant short-term pain relief of at least 4 weeks, however, TI displayed a significantly greater duration of benefit. Patients with effusions and milder radiographic change obtained the best response to treatment.