Contribution of red blood cells to the saturable mechanism of heparin clearance.
The pattern of clearance of intravenously administered heparin suggests the participation of a saturable mechanism which has been ascribed to heparin binding to the endothelium. Another potential site of heparin binding which may contribute to the saturable mechanism is the red blood cells. We have examined this possibility using unfractionated heparin together with a chemically-modified low affinity heparin labelled with tritium as tracer. Unfractionated heparin and the radioactive tracer were added to pooled plasma at a fixed concentration. The heparinized plasma was mixed with red cells to reconstitute whole blood. After centrifugation, the radioactivity, APTT, anti-Xa and anti-IIa activities were measured in the resultant plasmas. No evidence of significant binding to red cells could be detected even when the hematocrits of the reconstituted whole blood were increased. These results are at variance with those of a previously published report. We conclude that red cell binding does not contribute to the saturable mechanism of heparin clearance.


Striatal dopaminergic neurons as a potential target for GDNF based ischemic stroke therapy.
Glial cell-line-derived neurotrophic factor (GDNF) is a well-known regulatory neurotrophic factor on dopaminergic neurons. Several pathologies have been documented so far in case of any impairment in the dopaminergic system. This study aimed to investigate the potential protective role of lentiviral GNDF delivery on the small population of tyrosine hydroxylase (TH) positive dopamine producing striatal neurons after ischemic stroke. Fourteen C57BL/6J male mice (8-10 weeks) were intracerebrally treated with lentiviral GDNF (Lv-GDNF) or vehicle. Ten days after injections, cerebral ischemia was induced by blockage of the middle cerebral artery. Animals were terminated 72 h after ischemia, and their brains were taken for histological and molecular investigations. Following confirmation of GDNF overexpression, TH immunostaining and immunoblotting were used to evaluate the role of GDNF on dopaminergic neurons. Next, Fluro Jade C staining was implemented to examine the degree of neuronal degeneration at the damaged parenchyma. Neither the amount of TH positive dopaminergic neurons nor the expression of TH changed in the Lv-GDNF treated animals comparing to the vehicle group. On the other hand, GDNF exposure caused a significant increase in the expression of Nurr1, an essential transcription factor for dopaminergic neurons and Gap43, growth and plasticity promoting protein, in the ischemic striatum. Treatment with Lv-GDNF gave rise to a significant reduction in the number of degenerated neurons. Finally, enhanced GDNF expression also induced expression of an important stress-related transcription factor NF-κB as well as the nitric oxide synthase enzymes iNOS and nNOS in the contralesional hemisphere.


Evans Syndrome as First Manifestation of Primary Immunodeficiency in Clinical Practice.
Evans syndrome (ES) is a rare immune disorder in children, manifested by simultaneous or sequential autoimmune cytopenias (ACs) of unknown cause and having a chronic course with periods of exacerbation and remission. Some primary immunodeficiencies (PIDs) may present with autoimmune manifestations without infections, masking suspicion of them. The PIDs that can typically manifest as ES are autoimmune lymphoproliferative syndrome and common variable immunodeficiency (CVID). Review of clinical charts and laboratory results of pediatric patients followed-up in the outpatient clinic of PID with a diagnosis of ES and humoral immunodeficiency. Three pediatric patients, a boy and 2 girls, presented with corticosteroid-dependent ES. In the diagnostic approach, autoimmune lymphoproliferative syndrome was ruled out, and during follow-up, patients showed laboratory signs of humoral immune deficiency and were diagnosed with CVID. After initiating the recommended treatment for CVID with AC, patients improved without new exacerbations. These cases highlight the importance of detection of possible PID in the context of ES and the establishment of CVID treatment to control AC.


Rapid viral rebound after analytical treatment interruption in patients with very small HIV reservoir and minimal on-going viral transcription.
Viral remission after analytical treatment interruption (ATI), termed post-treatment control, has been described in a small proportion of HIV-positive patients. This phenomenon has been separately associated to both low levels of HIV-1 proviral DNA as well as cell-associated RNA. We investigated whether the combination of both parameters could help predict delayed viral rebound after treatment interruption (TI). We conducted an open single-arm ATI study in four Belgian HIV reference centres from January 2016 to July 2018. Eligible participants were adults who had fewer than 50 HIV-1 RNA copies/mL for more than two years, more than 500 CD4 cells/µL for more than three months, and were in general good health. Consenting participants who had fewer than 66 copies total HIV-1 DNA (t-DNA) and fewer than 10 copies cell-associated HIV-1 unspliced RNA (US-RNA) per million peripheral blood mononuclear cells (PBMCs), interrupted therapy and were monitored closely. Antiretroviral therapy (ART) was resumed after two consecutive viral loads exceeding 1000 copies or one exceeding 10,000 copies/mL. The primary outcome was the proportion of participants with fewer than 50 HIV-1 RNA copies/mL 48 weeks after TI. Secondary outcomes were time to viral rebound, the frequency of serious adverse events (AEs) and evolution of t-DNA and US-RNA after TI. All 16 consenting participants who interrupted therapy experienced rapid viral rebound two to eight weeks after TI. No serious AEs were observed. Levels of t-DNA and US-RNA increased after TI but returned to pre-ATI levels after treatment restart. None of the studied demographic, clinical and biological parameters were predictive of time of viral rebound. The combination of low levels of t-DNA and US-RNA in PBMCs, corresponding respectively to a small and transcriptionally silent viral reservoir, is not predictive of viral remission after TI in patients on ART.


Rapid change of methicillin-resistant Staphylococcus aureus clones in a Chinese tertiary care hospital over a 15-year period.
The incidence of methicillin-resistant Staphylococcus aureus (MRSA) has been increasing yearly at Peking Union Medical College Hospital (PUMCH). In order to understand the molecular evolution of MRSA at PUMCH, a total of 466 nonduplicate S. aureus isolates, including 302 MRSA and 164 methicillin-susceptible (MSSA) isolates recovered from 1994 to 2008 were characterized by staphylococcal cassette chromosome mec (SCCmec) typing, spa typing, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST). The 302 MRSA isolates were classified into 12 spa types and 9 sequence types (STs). During the years from 1994 to 2000, the most predominant MRSA clone was ST239-MRSA-III-spa t037. Since 2000, ST239-MRSA-III-spa t030 has rapidly replaced t037 and become the major clone. Another clone, ST5-MRSA-II-spa t002 emerged in 2002 and constantly existed at a low prevalence rate. The 164 MSSA isolates were classified into 62 spa types and 40 STs. ST398 was the most common MLST type for MSSA, followed by ST59, ST7, ST15, and ST1. Several MSSA genotypes, including ST398, ST1, ST121, and ST59, were identical to well-known epidemic community-acquired MRSA (CA-MRSA) isolates. MLST eBURST analysis revealed that the ST5, ST59, and ST965 clones coexisted in both MRSA and MSSA, which suggested that these MRSA clones might have evolved from MSSA by the acquisition of SCCmec. Two pvl-positive ST59-MRSA-IV isolates were identified as CA-MRSA according to the clinical data. Overall, our data showed that the ST239-MRSA-III-spa t037 clone was replaced by the emerging ST239-MRSA-III-spa t030 clone, indicating a rapid change of MRSA at a tertiary care hospital in China over a 15-year period.


Meta-analysis of randomized educational and behavioral interventions in type 2 diabetes.
This meta-analysis was conducted to assess the effect of educational and behavioral interventions on body weight and glycemic control in type 2 diabetes. Studies selected for analysis were published randomized controlled trials that evaluated educational and behavioral interventions (no drug interventions) in type 2 diabetes (sample size > or = 10). These criteria were applied to searches of electronic databases and relevant bibliographies. Data were independently abstracted by 2 reviewers and adjudicated by consensus. Of the 63 articles that met the inclusion criteria, 18 provided enough information for pooled estimates of glycohemoglobin (total Ghb, HbA1, or HbA1C). These 18 studies yielded 2720 participants (sample sizes of 18 to 749). Interventions ranged from 1 to 19 months; follow-up ranged from 1 to 26 months. Glycohemoglobin was reduced by a mean of 0.43%. When results were stratified by quality score, glycohemoglobin was -0.50% and -0.38% for studies with high and low quality scores, respectively. When weighting studies by sample size, fasting blood glucose was reduced by 24 mg/dL and weight by 3 lbs. Previous educational and behavioral interventions in type 2 diabetes have produced modest improvements in glycemic control. Future research should refine such interventions and improve methodology.


Tailored self-assembled nanocolloidal Huygens scatterers in the visible.
Existing nanocolloidal optical resonators exhibiting strong magnetic resonances often suffer from multi-step low yield synthesis methods as well as a limited tunability, particularly in terms of spectral superposition of electric and magnetic resonances, which is the cornerstone for achieving Huygens scatterers. To overcome these drawbacks, we have synthesized clusters of gold nanoparticles using an emulsion-based formulation approach. This fabrication technique involved emulsification of an aqueous suspension of gold nanoparticles in an oil phase, followed by controlled ripening of the emulsion. The structural control of the as synthesized clusters, of mean radius 120 nm and produced in large numbers, is demonstrated with microscopy and X-ray scattering techniques. Using a polarization-resolved multi-angle light scattering setup, we conduct a comprehensive angular and spectroscopic determination of their optical resonant scattering in the visible wavelength range. We thus report on the clear experimental evidence of strong optical magnetic resonances and directional forward scattering patterns. The clusters behave as strong Huygens sources. Our findings crucially show that the electric and magnetic resonances as well as the scattering patterns can be tuned by adjusting the inner cluster structure, modifying a simple parameter of the fabrication method. This experimental approach allows for the large scale production of nanoresonators with potential uses for Huygens metasurfaces.


Tobacco smoking affects bacterial acquisition and colonization in oral biofilms.
Recent evidence suggests that smoking affects the composition of the disease-associated subgingival biofilm, yet little is known about its effects during the formation of this biofilm. The present investigation was undertaken to examine the contributions of smoking to the composition and proinflammatory characteristics of the biofilm during de novo plaque formation. Marginal and subgingival plaque and gingival crevicular fluid samples were collected from 15 current smokers and from 15 individuals who had never smoked (nonsmokers) following 1, 2, 4, and 7 days of undisturbed plaque formation. 16S rRNA gene cloning and sequencing were used for bacterial identification, and multiplex bead-based flow cytometry was used to quantify the levels of 27 immune mediators. Smokers demonstrated a highly diverse, relatively unstable initial colonization of both marginal and subgingival biofilms, with lower niche saturation than that seen in nonsmokers. Periodontal pathogens belonging to the genera Fusobacterium, Cardiobacterium, Synergistes, and Selenomonas, as well as respiratory pathogens belonging to the genera Haemophilus and Pseudomonas, colonized the early biofilms of smokers and continued to persist over the observation period, suggesting that smoking favors early acquisition and colonization of pathogens in oral biofilms. Smokers also demonstrated an early proinflammatory response to this colonization, which persisted over 7 days. Further, a positive correlation between proinflammatory cytokine levels and commensal bacteria was observed in smokers but not in nonsmokers. Taken together, the data suggest that smoking influences both the composition of the nascent biofilm and the host response to this colonization.


Identification of Lactobacillus from the saliva of adult patients with caries using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
Matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) mass spectrometry (MS) has been presented as a superior method for the detection of microorganisms in body fluid samples (e.g., blood, saliva, pus, etc.) However, the performance of MALDI-TOF MS in routine identification of caries-related Lactobacillus isolates from saliva of adult patients with caries has not been determined. In the present study, we introduced a new MALDI-TOF MS system for identification of lactobacilli. Saliva samples were collected from 120 subjects with caries. Bacteria were isolated and cultured, and each isolate was identified by both 16S rRNA sequencing and MALDI-TOF MS. The identification results obtained by MALDI-TOF MS were concordant at the genus level with those of conventional 16S rRNA-based sequencing for 88.6% of lactobacilli (62/70) and 95.5% of non-lactobacilli (21/22). Up to 96 results could be obtained in parallel on a single MALDI target, suggesting that this is a reliable high-throughput approach for routine identification of lactobacilli. However, additional reference strains are necessary to increase the sensitivity and specificity of species-level identification.


Challenges and Strategies for Improving the Regenerative Effects of Mesenchymal Stromal Cell-Based Therapies.
Cell-based therapies have the potential to revolutionize current treatments for diseases with high prevalence and related economic and social burden. Unfortunately, clinical trials have made only modest improvements in restoring normal function to degenerating tissues. This limitation is due, at least in part, to the death of transplanted cells within a few hours after transplant due to a combination of mechanical, cellular, and host factors. In particular, mechanical stress during implantation, extracellular matrix loss upon delivery, nutrient and oxygen deprivation at the recipient site, and host inflammatory response are detrimental factors limiting long-term transplanted cell survival. The beneficial effect of cell therapy for regenerative medicine ultimately depends on the number of administered cells reaching the target tissue, their viability, and their promotion of tissue regeneration. Therefore, strategies aiming at improving viable cell engraftment are crucial for regenerative medicine. Here we review the major factors that hamper successful cell engraftment and the strategies that have been studied to enhance the beneficial effects of cell therapy. Moreover, we provide a perspective on whether mesenchymal stromal cell-derived extracellular vesicle delivery, as a cell-free regenerative approach, may circumvent current cell therapy limitations.


The effectiveness of benzoate in the management of seizures in nonketotic hyperglycinemia.
Patients with nonketotic hyperglycinemia generally have intractable seizures that are poorly responsive to anticonvulsant medication. No effective treatment has been consistently reported. In three patients with nonketotic hyperglycinemia, the oral administration of sodium benzoate in dosages designed to lower the cerebrospinal fluid concentration of glycine was followed by an abrupt change from frequent major seizures before treatment to no seizures, or only occasional minor ones, after treatment. This attenuation of seizures was associated with a decrease in the concentrations of glycine in both plasma and cerebrospinal fluid. There was no evident change in psychomotor development.


The hypophyseal-adrenal axis in the protein-calorie malnourished rat.
Male rats were fed an 8% low protein diet for 30 consecutive days beginning at 20 days of age. On day 50, the effect of the diet was evaluated on the hypophyseal-adrenal axis and the observations compared with those noted in age-matched controls fed a standard laboratory diet. The malnourished animals had significantly larger adrenal gland weights, relative to body size, and higher levels of serum corticosterone. Additionally, the number of ACTH-secreting cells in the pituitary glands of these rats was 63.8% greater than that present in the controls. These results suggest that ACTH secretion and adrenocortical physiology are stimulated under conditions of experimentally induced protein-calorie malnutrition.


Canine malignant lymphomas: comparison of morphologic and immunologic parameters.
Twenty-three canine malignant lymphomas were studied to correlate morphologic and immunologic properties of the neoplastic cells. Morphotologic characterization included histologic classification of cell type and growth pattern, anatomic distribution of lesions, and transmission electron microscopic examination. Parameters examined to indicate B- or T-cell nature of lymphoma cells included demonstration of mitogen responsiveness, cell-surface Ig, spontaneous rosette formation with human red blood cells, and scanning electron microscope (SEM) examination of cell-surface features. Results indicated that the cells from histiocytic lymphomas were lymphocytes rather than histiocytes or macrophages. Most cells from lymphomas examined possessed cell-surface ig, indicating B-cell nature. The cell types represented by the different Ig-positive lymphomas were compatible with maturation arrest at different stages in normal lymphocyte differentiation. For the two most common histiologic cell types, nitogen responsiveness and the presence of cell-surface Ig indicated that diffuse, poorly differentiated lymphocytic cases were biologically heterogenous, whereas nodular histiocytic lymphomas were biologically homogenous. Most canine lymphomas had a multicentric anatomic distribution; however, one thymic and two alimentary forms were observed. Lymphomas with a nodular pattern in lymph nodes had multifocal splenic involvement centered on small arteries, whereas lymphomas with a diffuse pattern in lymph nodes had diffuse involvement of splenic white pulp. The cells of Ig-positive and Ig-negative neoplasms examined by SEM were predominantly of the smooth-cell type.


Catecholamines 101.
This review of clinical catecholamine neurochemistry is based on the Streeten Memorial Lecture at the 19th annual meeting of the American Autonomic Society and lectures at a satellite of the 6th Congress of the International Society of Autonomic Neuroscience. Here I provide historical perspective, describe sources and meanings of plasma levels of catecholamines and their metabolites, present a model of a sympathetic noradrenergic neuron that conveys how particular aspects of sympathetic nervous function affect plasma levels of catecholamines and their metabolites, and apply the model to understand plasma neurochemical patterns associated with some drugs and disease states.


In vitro dose-dependent effects of matrix metalloproteinases on ECM hydrogel biodegradation.
Matrix metalloproteinases (MMPs) cause proteolysis of extracellular matrix (ECM) in tissues affected by stroke. However, little is known about how MMPs degrade ECM hydrogels implanted into stroke cavities to regenerate lost tissue. To establish a structure-function relationship between different doses of individual MMPs and isolate their effects in a controlled setting, an in vitro degradation assay quantified retained urinary bladder matrix (UBM) hydrogel mass as a measure of degradation across time. A rheological characterization indicated that lower ECM concentrations (<4 mg/mL) did not cure completely at 37 °C and had a high fraction of mobile proteins that were easily washed-out. Hydrolysis by dH2O caused a steady 2% daily decrease in hydrogel mass over 14 days. An acceleration of degradation to 6% occurred with phosphate buffered saline and artificial cerebrospinal fluid. MMPs induced a dose-dependent increase and within 14 days almost completely (>95%) degraded the hydrogel. MMP-9 exerted the most significant biodegradation, compared to MMP-3 and -2. To model the in vivo exposure of hydrogel to MMPs, mixtures of MMP-2, -3, and -9, present in the cavity at 14-, 28-, or 90-days post-stroke, revealed that 14- and 28-days mixtures achieved an equivalent biodegradation, but a 90-days mixture exhibited a slower degradation. These results revealed that hydrolysis, in addition to proteolysis, exerts a major influence on the degradation of hydrogels. Understanding the mechanisms of ECM hydrogel biodegradation is essential to determine the therapeutic window for bioscaffold implantation after a stroke, and they are also key to determine optimal degradation kinetics to support tissue regeneration. STATEMENT OF SIGNIFICANCE: After implantation into a stroke cavity, extracellular matrix (ECM) hydrogel promotes tissue regeneration through the degradation of the bioscaffold. However, the process of degradation of an ECM hydrogel remains poorly understood. We here demonstrated in vitro under highly controlled conditions that hydrogel degradation is very dependent on its protein concentration. Lower protein concentration hydrogels were weaker in rheological measurements and particularly susceptible to hydrolysis. The proteolytic degradation of tissue ECM after a stroke is caused by matrix metalloproteinases (MMPs). A dose-dependent MMP-driven biodegradation of ECM hydrogel exceeded the effects of hydrolysis. These results highlight the importance of in vitro testing of putative causes of degradation to gain a better understanding of how these factors affect in vivo biodegradation.


Primary mesenteric liposarcoma: An unusual presentation of a rare condition.
Primary mesenteric liposarcoma is a rare neoplasm. Here, we report the case of a 50 year old Indian man with a pleomorphic liposarcoma of the mesentery. The clinical findings pointed towards a retroperitoneal growth but imaging resolved the issue. A laparotomy was performed and a 20 cm × 20 cm multilobulated tumour arising from the mesentery and weighing 1.8 kilograms was excised with a segment of jejunum. The cut surface had a variegated appearance with regions of necrosis. The histological features were suggestive of pleomorphic liposarcoma. Tumour cells were positive for S-100 and negative for SMA/Desmin. Complete resection is curative although the role of chemotherapy remains to be established.


Circulating endothelial cells. Biomarker of vascular disease.
Recent research has recognised new populations of non-hematopoietic cells in the blood. One of these, circulating endothelial cells (CECs), often defined by the expression of membrane glycoprotein CD146, are rarely found in the blood in health, but raised numbers are present in a wide variety of human conditions, including inflammatory, immune, infectious, neoplastic and cardiovascular disease, and seem likely to be evidence of profound vascular insult. An additional population are endothelial progenitor cells, defined by the co-expression of endothelial and immaturity cell surface molecules and also by the ability to form colonies in vitro. Although increased numbers of CECs correlate with other markers of vascular disease, questions remain regarding the precise definition, cell biology and origin of CECs. For example, they may be damaged, necrotic or apopototic, or alive, and could possess procoagulant and/or proinflammatory properties. However, since these cells seem to be representative of in situ endothelium, their phenotype may provide useful information. Indeed, whatever their phenotype, there is growing evidence that CECs may well be a novel biomarker, the measurement of which will have utility in various clinical settings related to vascular injury. Despite this promise, progress is impeded by the diversity of methodologies used to detect these cells. Accordingly, results are sometimes inconclusive and even conflicting. Nevertheless, increased CECs predict adverse cardiovascular events in acute coronary syndromes, suggesting they may move from being simply a research index to having a role in the clinic. The objective of the present communication is to condense existing data on CECs, briefly compare them with progenitor cells, and summarise possible mechanism(s) by which they may contribute to vascular pathology.


Non-dystrophic myotonia: prospective study of objective and patient reported outcomes.
Non-dystrophic myotonias are rare diseases caused by mutations in skeletal muscle chloride and sodium ion channels with considerable phenotypic overlap between diseases. Few prospective studies have evaluated the sensitivity of symptoms and signs of myotonia in a large cohort of patients. We performed a prospective observational study of 95 participants with definite or clinically suspected non-dystrophic myotonia recruited from six sites in the USA, UK and Canada between March 2006 and March 2009. We used the common infrastructure and data elements provided by the NIH-funded Rare Disease Clinical Research Network. Outcomes included a standardized symptom interview and physical exam; the Short Form-36 and the Individualized Neuromuscular Quality of Life instruments; electrophysiological short and prolonged exercise tests; manual muscle testing; and a modified get-up-and-go test. Thirty-two participants had chloride channel mutations, 34 had sodium channel mutations, nine had myotonic dystrophy type 2, one had myotonic dystrophy type 1, and 17 had no identified mutation. Phenotype comparisons were restricted to those with sodium channel mutations, chloride channel mutations, and myotonic dystrophy type 2. Muscle stiffness was the most prominent symptom overall, seen in 66.7% to 100% of participants. In comparison with chloride channel mutations, participants with sodium mutations had an earlier age of onset of stiffness (5 years versus 10 years), frequent eye closure myotonia (73.5% versus 25%), more impairment on the Individualized Neuromuscular Quality of Life summary score (20.0 versus 9.44), and paradoxical eye closure myotonia (50% versus 0%). Handgrip myotonia was seen in three-quarters of participants, with warm up of myotonia in 75% chloride channel mutations, but also 35.3% of sodium channel mutations. The short exercise test showed ≥10% decrement in the compound muscle action potential amplitude in 59.3% of chloride channel participants compared with 27.6% of sodium channel participants, which increased post-cooling to 57.6% in sodium channel mutations. In evaluation of patients with clinical and electrical myotonia, despite considerable phenotypic overlap, the presence of eye closure myotonia, paradoxical myotonia, and an increase in short exercise test sensitivity post-cooling suggest sodium channel mutations. Outcomes designed to measure stiffness or the electrophysiological correlates of stiffness may prove useful for future clinical trials, regardless of underlying mutation, and include patient-reported stiffness, bedside manoeuvres to evaluate myotonia, muscle specific quality of life instruments and short exercise testing.


Simple sequence repeat (SSR) markers linked to E1, E3, E4, and E7 maturity genes in soybean.
Soybean near isogenic lines (NILs), contrasting for maturity and photoperiod sensitivity loci, were genotyped with approximately 430 mapped simple sequence repeats (SSRs), also known as microsatellite markers. By analysis of allele distributions across the NILs, it was possible to confirm the map location of the Dt1 indeterminate growth locus, to refine the SSR mapping of the T tawny pubescence locus, to map E1 and E3 maturity loci with molecular markers, and to map the E4 and E7 maturity loci for the first time. Molecular markers flanking these loci are now available for marker-assisted breeding for these traits. Analysis of map locations identified a putative homologous relationship among four chromosomal regions; one in the middle of linkage group (LG) C2 carrying E1 and E7, one on LG I carrying E4, one at the top of LG C2, at which there is a reproductive period quantitative trait locus (QTL), and the fourth on LG B1. Other evidence suggests that homology also exists between the E1 + E7 region on LG C2 and a region on LG L linked to a pod maturity QTL. Homology relationships predict possible locations in the soybean genome of additional maturity loci, as well as which maturity loci may share a common evolutionary origin and similar mechanism(s) of action.


Black soybean extract protects against TMT-induced cognitive defects in mice.
To find a neuroactive compound with a potent inhibitory effect on acetylcholinesterase (AChE) and in vivo anti-amnesic activity from natural resources, we evaluated anthocyanins and nonanthocyanins from black soybean extract. Nonanthocyanins from black soybean extract were the most potent and dose-dependent AChE inhibitors. Intracellular reactive oxygen species accumulation resulting from H₂O₂ treatment was significantly decreased compared with cells treated with H₂O₂ only. Nonanthocyanins were also neuroprotective against H₂O₂ treated neurotoxicity by 3-[4,5-dimethythiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) and lactate dehydrogenase (LDH) assay. Finally, nonanthocyanins from black soybean in the preadministration group attenuated trimethyltin (TMT)-induced memory injury in both in vivo tests. AChE, prepared from mice brain tissues, was inhibited by nonanthocyanins from black soybean in a dose-dependent manner. Malondialdehyde generation in the brain homogenates of mice treated with nonanthocyanins from black soybean was decreased. We concluded that nonanthocyanins from black soybean had an efficacious in vitro AChE inhibitory activity, and protected against H₂O₂-induced neurotoxicity. In addition, our findings suggest that nonanthocyanins from black soybean may improve the TMT-induced learning and memory deficit because of AChE inhibition of mice brain tissue. Consequently, these results demonstrate that the nonanthocyanins from black soybean could possess a wide range of beneficial activities for neurodegenerative disorders.


Genetic segregation of airway disease traits despite redundancy of calcium-activated chloride channel family members.
Complex airway diseases such as asthma and chronic obstructive pulmonary disease exhibit stereotyped traits (especially airway hyperreactivity and mucous cell metaplasia) that are variably expressed in each patient. Here, we used a mouse model for virus-induced long-term expression of these traits to determine whether individual traits can be genetically segregated and thereby linked to separate determinants. We showed that an F2 intercross population derived from susceptible and nonsusceptible mouse strains can manifest individual phenotypic extremes that exhibit one or the other disease trait. Functional genomic analysis of these extremes further indicated that a member of the calcium-activated chloride channel (CLCA) gene family designated mClca3 was inducible with mucous cell metaplasia but not airway hyperreactivity. In confirmation of this finding, we found that mClca3 gene transfer to mouse airway epithelium was sufficient to induce mucous cell metaplasia but not airway hyperreactivity. However, newly developed mClca3(-/-) mice exhibited the same degree of mucous cell metaplasia and airway hyperreactivity as wild-type mice. Bioinformatic analysis of the Clca locus led to the identification of mClca5, and gene transfer indicated that mClca5 also selectively drives mucous cell metaplasia. Thus, in addition to the capacity of CLCA family members to exhibit diverse functional activities, there is also preserved function so that more than one family member mediates mucous cell metaplasia. Nonetheless, Clca expression appears to be a selective determinant of mucous cell metaplasia so that shared homologies between CLCA family members may still represent a useful target for focused therapeutic intervention in hypersecretory airway disease.


Cell Therapy for Chronic TBI: Interim Analysis of the Randomized Controlled STEMTRA Trial.
To determine if chronic motor deficits secondary to traumatic brain injury (TBI) can be improved by implantation of allogeneic modified bone marrow-derived mesenchymal stromal/stem cells (SB623). This 6-month interim analysis of the 1-year double-blind, randomized, surgical sham-controlled, phase 2 STEMTRA trial (NCT02416492) evaluated safety and efficacy of the stereotactic intracranial implantation of SB623 in patients with stable chronic motor deficits secondary to TBI. Patients in this multi-center trial (N = 63) underwent randomization in a 1:1:1:1 ratio to 2.5 × 106, 5.0 × 106, 10 × 106 SB623 cells or control. Safety was assessed in patients who underwent surgery (N = 61), and efficacy in the modified intent-to-treat population of randomized patients who underwent surgery (N = 61; SB623 = 46, control = 15). The primary efficacy endpoint of significant improvement from baseline of Fugl-Meyer Motor Scale score at 6 months for SB623-treated patients was achieved. SB623-treated patients improved by (LS mean [SE]) +8.3 (1.4) vs +2.3 (2.5) for control at 6 months, the LS mean difference was 6.0 (95% CI: 0.3-11.8); p = 0.040. Secondary efficacy endpoints improved from baseline, but were not statistically significant vs control at 6 months. There were no dose-limiting toxicities or deaths, and 100% of SB623-treated patients experienced treatment-emergent adverse events vs 93.3% of control patients (p = 0.25). SB623 cell implantation appeared to be safe and well tolerated, and patients implanted with SB623 experienced significant improvement from baseline motor status at 6 months compared to controls. This study provides Class I evidence that implantation of SB623 was well tolerated and associated with improvement in motor status.


Snake envenomation in dogs in New South Wales.
To obtain baseline data on the prevalence of elapid snake envenomation in dogs presented to veterinary practices in New South Wales and to assess attitudes of veterinarians to this clinical entity. A mailed questionnaire, sent to all veterinary clinics within New South Wales, was utilised to collect epidemiological information regarding elapid snake envenomation in dogs. A response rate of 68% was obtained and a yearly prevalence of snake envenomation in dogs across New South Wales veterinary clinics was estimated as 0.31%. The most common species reported to be responsible for envenomation within NSW was the Red Bellied Black snake (Pseudechis porphyriacus) followed by the Brown snake (Pseudonaja textilis) and then Tiger snake (Notechis scutatus). The reported envenomation syndromes caused by these common snake species were perceived to be similar for Brown and Tiger snakes but differed for Red Bellied Black snakes. Diagnosis of snake envenomation was based predominantly on the recognition of clinical signs. Specific diagnostic tests, such as venom detection kits, were used infrequently. The most common treatment was reported to be a combination of intravenous fluid therapy and antivenom, and monitoring of response to this treatment was usually through assessment of clinical signs. Survival after antivenom administration was reported to be highest for Red Bellied Black snake species. Survival was perceived to be associated with time between envenomation and presentation to the veterinary clinic and with antivenom administration. Current attitudes and perceptions of veterinarians have been defined. Diagnosis of species-specific snake envenomation is shown to be made on the basis of clinical signs which are, however, reported as similar for each species. Clearer definition of these envenomation syndromes and identification of accessible diagnostic testing procedures are needed.


Disrupted functional connectivity in primary progressive apraxia of speech.
Apraxia of speech is a motor speech disorder thought to result from impaired planning or programming of articulatory movements. It can be the initial or only manifestation of a degenerative disease, termed primary progressive apraxia of speech (PPAOS). The aim of this study was to use task-free functional magnetic resonance imaging (fMRI) to assess large-scale brain network pathophysiology in PPAOS. Twenty-two PPAOS participants were identified from a prospective cohort of degenerative speech and language disorders patients. All participants had a comprehensive, standardized evaluation including an evaluation by a speech-language pathologist, examination by a behavioral neurologist and a multimodal imaging protocol which included a task-free fMRI sequence. PPAOS participants were age and sex matched to amyloid-negative, cognitively normal participants with a 1:2 ratio. We chose a set of hypothesis driven, predefined intrinsic connectivity networks (ICNs) from a large, out of sample independent component analysis and then used them to initialize a spatiotemporal dual regression to estimate participant level connectivity within these ICNs. Specifically, we evaluated connectivity within the speech and language, face and hand sensorimotor, left working memory, salience, superior parietal, supramarginal, insular and deep gray ICNs in a multivariate manner. The spatial maps for each ICN were then compared between PPAOS and control participants. We used clinical measures of apraxia of speech severity to assess for clinical-connectivity correlations for regions found to differ between PPAOS and control participants. Compared to controls, PPAOS participants had reduced connectivity of the right supplementary motor area and left posterior temporal gyrus to the rest of the speech and language ICN. The connectivity of the right supplementary motor area correlated negatively with an articulatory error score. PPAOS participants also had reduced connectivity of the left supplementary motor area to the face sensorimotor ICN, between the left lateral prefrontal cortex and the salience ICN and between the left temporal-occipital junction and the left working memory ICN. The latter connectivity correlated with the apraxia of speech severity rating scale, although the finding did not survive correction for multiple comparisons. Increased connectivity was noted in PPAOS participants between the dorsal posterior cingulate and the left working memory ICN. Our results support the importance of the supplementary motor area in the pathophysiology of PPAOS, which appears to be disconnected from speech and language regions. Supplementary motor area connectivity may serve as a biomarker of degenerative apraxia of speech severity.


The evaluation of the diuretic action of parenteral formulations of metolazone.
This study was design to compare the diuretic and natriuretic effects of the 2 parenteral formulations of metolazone and the combination of these 2 formulations of metolazone with the parenteral administration of furosemide. Metolazone is an anthracrene acid derivate and manifests a dual diuretic effect on the proximal and distal tubule with a minimal kaluretic effect. It is currently only marketed in an orally administrable formulation, and this has limited its utility in critically ill patients. Metolazone given orally and furosemide given orally or parenterally are frequently administrated together when furosemide alone is clinically inadequate at producing a desired diuresis. Sprague Dawley male rats (400 to 450 g) were divided into groups to receive a parenteral formulation of metolazone or furosemide administrated separately intraperitoneally (IP) or administrated IP in combination with one another. Tris buffer-administered IP was used as a control vehicle comparator. The urine volume voided over the following 24 hours was collected, measured and analyzed for sodium content. Vehicle (Tris buffer) caused 9 +/- 1 mL/d output of urine with a sodium [Na+] concentration of 194 +/- 41 micromol/L (n=6 per group). Metolazone 2 mg/kg resulted in 16 +/- 3 mL/d urine output and sodium [Na+] of 278 +/- 76 micromol/L (n=6 per group). Furosemide 2, 4, and 6 mg/kg resulted in a volume of urine 9 +/- 1, 14 +/- 2 and 17 +/- 2 mL/d and [Na+] micromol/L of 194 +/- 41, 206 +/- 108, and 229 +/- 91, respectively. Metolazone 4 mg/kg combined with furosemide 4 mg/kg resulted in a urine volume of 21 +/- 1 mL/d and [Na+] of 326 +/- 108 micromol/L. Combining metolazone and furosemide can cause an increase in urine volume and sodium excretion. Metolazone administrated parenterally in combination with the parenteral administration of furosemide appears to have an important clinical potential.


Sec61p is required for ERAD-L: genetic dissection of the translocation and ERAD-L functions of Sec61P using novel derivatives of CPY.
Misfolded proteins in the endoplasmic reticulum (ER) are exported to the cytosol for degradation by the proteasome in a process known as ER-associated degradation (ERAD). CPY* is a well characterized ERAD substrate whose degradation is dependent upon the Hrd1 complex. However, although the functions of some of the components of this complex are known, the nature of the protein dislocation channel remains obscure. Sec61p has been suggested as an obvious candidate because of its role as a protein-conducting channel through which polypeptides are initially translocated into the ER. However, it has not yet been possible to functionally dissect any role for Sec61p in dislocation from its essential function in translocation. By changing the translocation properties of a series of novel ERAD substrates, we are able to separate these two events and find that functional Sec61p is essential for the ERAD-L pathway.


Three novel mammalian toll-like receptors: gene structure, expression, and evolution.
We describe three novel genes, encoding members of the Toll-like receptor (Tlr) family (TLR7, TLR8, and TLR9). These Tlr family members, unlike others reported to date, were identified within a genomic database. TLR7 and TLR8 each have three exons, two of which have coding function, and lie in close proximity to one another at Xp22, alongside a pseudogene. The remaining gene (TLR9) resides at 3p21.3 (in linkage with the MyD88 gene), and is expressed in at least two splice forms, one of which is monoexonic and one of which is biexonic, the latter encoding a protein with 57 additional amino acids at the N-terminus. The novel Tlrs comprise a cluster as nearest phylogenetic neighbors. Combining all sequence data related to Toll-like receptors, we have drawn several inferences concerning the phylogeny of vertebrate and invertebrate Tlrs. According to our best estimates, mammalian TLRs 1 and 6 diverged from a common mammalian ancestral gene 95 million years ago. TLR4, which encodes the endotoxin sensor in present-day mammals, emerged as a distinct entity 180 million years ago. TLRs 3 and 5 diverged from a common ancestral gene approximately 150 million years ago, as did Tlr7 and Tlr8. Very likely, fewer Tlrs existed during early vertebrate evolution: at most three or four were transmitted with the primordial vertebrate line. Phylogenetic data that we have adduced in the course of this work also suggest the existence of a Drosophila equivalent of MyD88, and indicate that the plasma membrane protein SIGIRR is close functional relative of MyD88 in mammals. Finally, a single present-day representative of the Toll-like proteins in Drosophila has striking cytoplasmic domain homology to mammalian Tlrs within the cluster that embraces TLRs 1, 2, 4, and 6. This would suggest that an ancestral (pre-vertebrate) Tlr may have adopted a pro-inflammatory function 500 million years ago.


Effect of lateral hinge fractures for bone union and clinical outcomes following opening-wedge distal tibial tubercle osteotomy in comparison with opening-wedge high tibial osteotomy.
Although the effects of lateral hinge fractures (LHF) on bone union and clinical outcomes after opening-wedge high tibial osteotomy (OWHTO) have been established, the effects of LHF after opening-wedge distal tibial tubercle osteotomy (OWDTO) are unclear. We hypothesised that LHF after OWDTO would be associated with delayed bone union and result in poorer clinical outcomes than expected for LHF after OWHTO. This study enrolled 100 patients, with 50 OWDTO patients (18 men; mean age, 63.2 years) and 50 OWHTO patients compared based on the propensity score matched analysis. The effect of LHF on bone union was compared between the groups. Clinical outcomes were assessed using the Lysholm score and the Knee Injury and Osteoarthritis Outcome Score (KOOS) at the mean follow-up of 28 months. There was no between-group difference in the incidence rate of LHF. However, the rate of bone union at the anterior flange in the presence of an LHF was significantly lower in the OWDTO (26%) than in the OWHTO (80%) 3 months postoperatively (p < 0.05), but no difference was observed 12 months postoperatively. The Lysholm score was significantly lower for patients with LHF following OWDTO than for OWDTO patients without LHF or OWHTO patients with/without LHF 3 and 12 months postoperatively (p < 0.001); Lysholm score and KOOS were not different at the final follow-up. LHF after OWDTO was associated with delayed bone union and poor clinical outcomes until 12 months. This information can guide decisions regarding the indications and the management of patients after OWDTO. IV.


Sildenafil induced priapism.
A 53 year-old Japanese man was referred to our hospital for persistent priapism, which had been induced by 200 mg (usual dose 25-50 mg) of sildenafil citrate (Viagra) three days earlier. He had a history of erectile dysfunction and had undergone penile injection therapy elsewhere; however, he had not used injection therapy this time. He obtained sildenafil personally without a doctor's prescription. He had not taken any other drugs that affect the metabolism of sildenafil, nor did he have any medical complications that might induce priapism. Since needle aspiration and irrigation were ineffective as first line therapy, surgical treatment was indicated to relieve the condition; a incision of tunica albuginea of both corpora cavernosa was made, and vigorous irrigation of saline washed out the blood clots. This is the first case report of priapism induced by sildenafil. Although this drug can be obtained through private commerce, it should be used under professional guidance as its abuse may lead to severe morbidity.


DNA Damage Tolerance Pathway Choice Through Uls1 Modulation of Srs2 SUMOylation in Saccharomyces cerevisiae.
DNA damage tolerance and homologous recombination pathways function to bypass replication-blocking lesions and ensure completion of DNA replication. However, inappropriate activation of these pathways may lead to increased mutagenesis or formation of deleterious recombination intermediates, often leading to cell death or cancer formation in higher organisms. Post-translational modifications of PCNA regulate the choice of repair pathways at replication forks. Its monoubiquitination favors translesion synthesis, while polyubiquitination stimulates template switching. Srs2 helicase binds to small ubiquitin-related modifier (SUMO)-modified PCNA to suppress a subset of Rad51-dependent homologous recombination. Conversely, SUMOylation of Srs2 attenuates its interaction with PCNA Sgs1 helicase and Mus81 endonuclease are crucial for disentanglement of repair intermediates at the replication fork. Deletion of both genes is lethal and can be rescued by inactivation of Rad51-dependent homologous recombination. Here we show that Saccharomyces cerevisiae Uls1, a member of the Swi2/Snf2 family of ATPases and a SUMO-targeted ubiquitin ligase, physically interacts with both PCNA and Srs2, and promotes Srs2 binding to PCNA by downregulating Srs2-SUMO levels at replication forks. We also identify deletion of ULS1 as a suppressor of mus81Δ sgs1Δ synthetic lethality and hypothesize that uls1Δ mutation results in a partial inactivation of the homologous recombination pathway, detrimental in cells devoid of both Sgs1 and Mus81 We thus propose that Uls1 contributes to the pathway where intermediates generated at replication forks are dismantled by Srs2 bound to SUMO-PCNA. Upon ULS1 deletion, accumulating Srs2-SUMO-unable to bind PCNA-takes part in an alternative PCNA-independent recombination repair salvage pathway(s).


Commentary: Psychiatric advance directives as tools for enhancing treatment of the mentally ill.
In this issue of the Journal, Swanson et al. explore the demand for and utilization of psychiatric advance directives (PADs), and discuss some of the barriers to their greater acceptance. This author suggests that PADs may be more widely accepted and utilized if they are viewed as tools for maximizing individual autonomy, with an emphasis on optimizing rather than merely restricting care.


[Participation in the IEC Workshop on Adolescent Sexual Health--a challenge to protect reproductive health/right in Latin America and Caribbean Region].
In order to explore the possibility of new global activity in nursing/midwifery to be performed by the WHO Collaborating Center for Nursing Development in Primary Health Care at St. Luke's College of Nursing, the center sent two faculty members to attend the IEC Workshop on Adolescent Sexual Health held in November 1996 in Mexico. The two representatives took part as resource persons and reported to the workshop on the activities of nurses and midwives for adolescent sexual health in Japan. During the session, they obtained information on the actual situation of adolescent sexual health in Latin America and Caribbean region and the novel strategies which were carried out in cooperation between governmental and non governmental organizations. The workshop also provided a good opportunity for international communication and information exchange with health workers in the region about adolescent sexual health. By participating in the workshop, we obtained first-hand information on various aspects of cooperation in international health.


Prospective evaluation of unexplained chronic liver transaminase abnormalities in asymptomatic and symptomatic patients.
It is currently recommend to perform a liver biopsy for patients with chronically elevated liver function tests (LFT) of unknown etiology (marker negative). The necessity and benefits of these recommendations are unknown. The aims of this study were to determine the prevalence of marker-negative LFT in patients referred for evaluation of chronically elevated LFT; to determine the prevalence of diseases that may be associated with marker-negative abnormal LFT; and to assess whether a liver biopsy alters the management of such patients. We conducted a prospective observational study of 1124 adults referred for evaluation of chronically elevated LFT. Patients who consented to a liver biopsy were eligible. Marker-negative abnormal LFT was defined as the absence of accepted serum markers for infectious, metabolic, autoimmune, or hereditary liver disease, the absence of a history of alcohol or hepatotoxic drug use, and the absence of signs of chronic liver disease. Eighty-one of 1124 eligible patients were marker-negative. Liver biopsies in the 81 marker-negative patients revealed: normal histology (eight), steatosis (41), steatohepatitis (26), fibrosis (four), and cirrhosis (two). All 73 abnormal liver biopsies had some degree of steatosis. There were no significant associations between histological findings and the presence of obesity (p = 0.13), hyperlipidemia (p = 0.4), or diabetes (p = 0.9). There were no significant associations when classifying patients by gender or by symptoms. In the setting of marker-negative elevated LFT, the most likely histological diagnosis is fatty metamorphosis of the liver with occasional associated fibrosis.


Two-photon absorption-induced photoacoustic imaging of Rhodamine B dyed polyethylene spheres using a femtosecond laser.
In the present paper we demonstrate the possibility to image dyed solids, i.e. Rhodamine B dyed polyethylene spheres, by means of two-photon absorption-induced photoacoustic scanning microscopy. A two-photon luminescence image is recorded simultaneously with the photoacoustic image and we show that location and size of the photoacoustic and luminescence image match. In the experiments photoacoustic signals and luminescence signals are generated by pulses from a femtosecond laser. Photoacoustic signals are acquired with a hydrophone; luminescence signals with a spectrometer or an avalanche photo diode. In addition we derive the expected dependencies between excitation intensity and photoacoustic signal for single-photon absorption, two-photon absorption and for the combination of both. In order to verify our setup and evaluation method the theoretical predictions are compared with experimental results for liquid and solid specimens, i.e. a carbon fiber, Rhodamine B solution, silicon, and Rhodamine B dyed microspheres. The results suggest that the photoacoustic signals from the Rhodamine B dyed microspheres do indeed stem from two-photon absorption.


[Screening for hearing disorders. Current status and bottlenecks].
In The Netherlands a nation-wide screening program on hearing for infants is available. This program started around the late sixties and is now incorporated in the Health Care System and added to the tasks of the Well Baby Clinics. A modification of the method as developed by the Ewings is used. This method is relatively easy to learn. It is, however, essential that it is executed accurately and with a maximum of care. Therefore the testing itself as well as the assessment of the results is strictly standardized and a lot of attention is given to the training program. The Dutch Foundation for the Deaf and Hearing-Impaired Child co-ordinates the screening program. About 75% of the 9 months old babies are screened and some 5% of the children has to be referred. The Ewing-test has proved to be effective. It detects both severe and moderate sensory-neural hearing-losses as well as conductive hearing-losses. However, the program faces us with a few difficulties: the structure is effective, though time-consuming; the referral-procedure is not always satisfactory. Due to different factors, some groups of children, sometimes at risk, are not covered by the screening program or any other program.


[Sarcoma of the endometrium. Epidemiology and prognostic factors. Apropos of a study of 22 cases].
Endometrial sarcomas (sarcomas of the endometrial stroma and mixed mullerian tumors) are rare tumors with a bad prognosis. Because of our experience in treating these tumors we have been able to make an analysis of epidemiology and the prognostic factors. We have treated 22 patients. The mean age was 61 years. There were 15 stage I, 3 stage II, 2 stage III, and 2 stage IV. 21/22 patients had a surgical resection of the primary tumor, 19/22 had pelvic irradiation after surgery with vaginal radiotherapy given for 14 patients. The actuarial survival rates at 3 and 5 years were 51 and 34%. 5/22 patients had pelvic recurrence, 6/22 had distant metastasis. In our study the prognostic factors were the stage and the histological subtype. Pelvic radiotherapy seems to improve the rates for survival. The main points for discussion are the need to have a definite histological diagnosis done after initial surgery. A CT scan of the lungs and a lymphangiogram must be performed before treatment. After surgery it appears necessary to give pelvic and vaginal radiotherapy. At the present time chemotherapy did not appear to improve survival.


PAKa, a putative PAK family member, is required for cytokinesis and the regulation of the cytoskeleton in Dictyostelium discoideum cells during chemotaxis.
We have identified a Dictyostelium discoideum gene encoding a serine/threonine kinase, PAKa, a putative member of the Ste20/PAK family of p21-activated kinases, with a kinase domain and a long NH(2)-terminal regulatory domain containing an acidic segment, a polyproline domain, and a CRIB domain. PAKa colocalizes with myosin II to the cleavage furrow of dividing cells and the posterior of polarized, chemotaxing cells via its NH(2)-terminal domain. paka null cells are defective in completing cytokinesis in suspension. PAKa is also required for maintaining the direction of cell movement, suppressing lateral pseudopod extension, and proper retraction of the posterior of chemotaxing cells. paka null cells are defective in myosin II assembly, as the myosin II cap in the posterior of chemotaxing cells and myosin II assembly into cytoskeleton upon cAMP stimulation are absent in these cells, while constitutively active PAKa leads to an upregulation of myosin II assembly. PAKa kinase activity against histone 2B is transiently stimulated and PAKa incorporates into the cytoskeleton with kinetics similar to those of myosin II assembly in response to chemoattractant signaling. However, PAKa does not phosphorylate myosin II. We suggest that PAKa is a major regulator of myosin II assembly, but does so by negatively regulating myosin II heavy chain kinase.


Head and neck position for direct laryngoscopy.
The sniffing position (SP) has traditionally been considered the optimal head position for direct laryngoscopy (DL). Its superiority over other head positions, however, has been questioned during the last decade. We reviewed the scarce literature on the subject to examine the evidence either in favor or against the routine use of the SP. A standard definition for the position should be used (e.g., 35° neck flexion and 15° head extension) to avoid confusion about what constitutes a proper SP and to compare the results from different studies. Although several theories were proposed to explain the superiority of the SP, the three axes alignment theory is still considered a valid anatomical explanation. Although head elevation is needed to achieve the desired neck flexion, the elevation height may vary from one patient to another depending on head and neck anatomy and size of the chest. In infants and small children, for example, no head elevation is needed because the size and shape of the head allow axes approximation in the head-flat position. Horizontal alignment of the external auditory meatus with the sternum, in both obese and non-obese patients, indicates, and can be used as a marker for, proper positioning. Analysis of the available literature supports the use of the SP for DL. To achieve a proper SP in obese patients, the "ramped" (or the back-up) position should be used. The SP does not guarantee adequate exposure in all patients, because many other anatomical factors control the final degree of visualization. However, it should be the starting head position for DL because it provides the best chance at adequate exposure. Attention to details during positioning and avoidance of minor technical errors are essential to achieve the proper position. DL should be a dynamic procedure and position adjustment should be instituted in case poor visualization is encountered in the SP.


Autoreactive T cells in type 1 diabetes.
Type 1 diabetes (T1D) is a chronic autoimmune disease that causes severe loss of pancreatic β cells. Autoreactive T cells are key mediators of β cell destruction. Studies of organ donors with T1D that have examined T cells in pancreas, the diabetogenic insulitis lesion, and lymphoid tissues have revealed a broad repertoire of target antigens and T cell receptor (TCR) usage, with initial evidence of public TCR sequences that are shared by individuals with T1D. Neoepitopes derived from post-translational modifications of native antigens are emerging as novel targets that are more likely to evade self-tolerance. Further studies will determine whether T cell responses to neoepitopes are major disease drivers that could impact prediction, prevention, and therapy. This Review provides an overview of recent progress in our knowledge of autoreactive T cells that has emerged from experimental and clinical research as well as pathology investigations.


Atrial fibrillation, the arrhythmia of the elderly, causes and associated conditions.
Atrial fibrillation (AF) is a common clinical problem, particularly in the elderly, and in patients with organic heart disease. A small percentage of patients, have a potentially reversible cause. Atrial fibrillation is in most patients (approximately 70%) associated with chronic organic heart disease including valvular heart disease, coronary artery disease, hypertension, particularly if left ventricular hypertrophy is present, hypertrophic cardiomyopathy, dilated cardiomyopathy and congenital heart disease and most commonly in adults, atrial septal defect. As in many chronic conditions, determining whether AF is the result or is unrelated to the underlying heart disease, remains unclear. The list of possible etiologies also include cardiac amyloidosis, hemochromatosis and endomyocardial fibrosis. Other heart diseases, such as mitral valve prolapse (with or without mitral regurgitation), calcification of the mitral annulus, atrial myxoma, pheochomocytoma and idiopathic dilated right atrium, present a higher incidence of AF. The relationship between these findings and the arrhythmia are still unclear. Atrial fibrillation may occur in the absence of detectable organic heart disease, the so-called "lone AF", in about 30% of cases. The term "lone AF" or "idiopathic AF" implies the absence of any detectable etiology including hyperthyroidism, chronic obstructive lung disease, overt sinus node dysfunction, and overt or concealed preexcitation (Wolf-Parkinson-White syndrome), only to mention a few of other rare causes of AF. In every instance of recently discovered AF, thyrotoxicosis should be ruled out. The autonomous nervous system may contribute to the occurrence of AF in some patients. Atrial fibrillation occurs commonly in patients with valvular heart disease, particularly when it involves the mitral valve. The occurrence of AF is unrelated to the severity of mitral stenosis but is more common in patients with enlarged left atrium and congestive heart failure. In patients with coronary artery disease, Af occurs predominantly in older patients, males and patients with left ventricular dysfunction. Important predictive factors of AF include hypertension, left ventricular hypertrophy and diabetes. However, the relation between AF and hypertension remains unclear. The risk of the development of AF, in an individual patient, is often difficult to assess but increasing age, presence of valvular heart disease and congestive heart failure, increase the risk of AF.


Benthic microbial diversity trends in response to heavy metals in an oxygen-deficient eutrophic bay of the Humboldt current system offshore the Atacama Desert.
Mejillones Bay is a coastal ecosystem situated in an oxygen-deficient upwelling area impacted by mining activities in the coastal desert region of northern Chile, where conspicuous microbial life develops in the sediments. Herein, heavy metal (loid)s (HMs) such as Cu, Pb, As, Zn, Al, Fe, Cd, Mo, Ni and V as well as benthic microbial communities were studied using spectrometry and iTag-16 S rRNA sequencing. Samples were taken from two contrasting sedimentary localities in the Bay named Punta Rieles (PR) and Punta Chacaya (PC) within 10-50 m water-depth gradient. PR sediments were organic matter rich (21.1% of TOM at 50 m) and overlaid with low-oxygen waters (<0.06 ml O2/L bottom layer) compared with PC. In general, HMs like Al, Ni, Cd, As and Pb tended to increase in concentration with depth in PR, while the opposite pattern was observed in PC. In addition, PR presented a higher number of unique families (72) compared to PC (35). Among the top ten microbial families, Desulfobulbaceae (4.6% vs. 3.2%), Flavobacteriaceae (2.8% vs. 2.3%) and Anaerolineaceae (3.3% vs. 2.3%) dominated in PR, meanwhile Actinomarinales_Unclassified (8.1% vs. 4.2%) and Sandaracinaceae (4.4% vs. 2.0%) were more abundant in PC. Multivariate analyses confirmed that water depth-related variation was a good proxy for oxygen conditions and metal concentrations, explaining the structure of benthic microbial assemblages. Cd, Ni, As and Pb showed uniformly positive associations with communities that represented the keystone taxa in the co-occurrence network, including Anaerolineaceae, Thiotrichaceae, Desulfobulbaceae, Desulfarculaceae and Bacteroidales_unclassified communities. Collectively, these findings provide new insights for establishing the ecological interconnections of benthic microorganisms in response to metal contamination in a coastal upwelling environment.


Evaluation of salivary immunoglobulin A levels in tobacco smokers and patients with recurrent aphthous ulcers.
The aim of the present study was to analyze the influence of smoking on the salivary immunoglobulin response in smokers and to evaluate the salivary immunoglobulin A in patients with recurrent aphthous ulcers. The study included total of 80 subjects, of whom 40 were having history of chronic smoking habit, 20 were clinically diagnosed cases of recurrent aphthous ulcer and 20 were in the control group. Sample of unstimulated saliva was collected, centrifuged and analyzed for the level of salivary immunoglobulin A with turbidimetric immunoassay. For all the tests, a P- value of < 0.05 was considered for statistical significance. The mean salivary immunoglobulin A level in control group was 0.20 Grams/litre and in smokers the mean salivary immunoglobulin A level was 0.13 Grams / Litre. In patients with recurrent aphthous ulcers mean salivary immunoglobulin A level was 0.31 Grams / Litre. The mean salivary immunoglobulin A levels showed a decreasing trend from controls to smokers. These results were highly significant for values between control groups to smokers. The mean salivary immunoglobulin A levels demonstrated a progressive decrease from controls to smokers. This investigative procedure although non-specific, can be used as a diagnostic marker in smokers and patients with recurrent aphthous ulcers.


Signal transduction through the Ras/Erk pathway is essential for the mycoestrogen zearalenone-induced cell-cycle progression in MCF-7 cells.
Zearalenone is a naturally occurring estrogenic contaminant of moldy feeds and is present in high concentrations in dairy products and cereals. Zearalenone was postulated to contribute to the overall estrogen load of women, but the mechanisms of its action are not known. We demonstrated that zearalenone could stimulate the growth of estrogen receptor-positive human breast carcinoma cell line MCF-7. In addition, zearalenone functioned as an antiapoptotic agent by increasing the survival of MCF-7 cell cultures undergoing apoptosis caused by serum withdrawal. Treatment of these cells with 100 nM zearalenone induced cell-cycle transit after increases in the expression of c-myc mRNA and cyclins D1, A, and B1 and downregulation of p27(Kip-1). G(1)/G(2)-phase kinase activity and phosphorylation of the retinoblastoma gene product was also evident. Flow cytometric analysis demonstrated entry of cells into the S and G(2)/M phases of the cell cycle, and phosphorylation of histone H3 occurred 36 h after zearalenone treatment. Ectopic expression of a dominant-negative p21(ras) completely abolished the zearalenone-induced DNA synthesis in these cells, and the specific inhibitor PD98059 for mitogen/extracellular-regulated protein kinase kinase arrested S-phase entry induced by zearalenone. These data suggest that the mitogen-activated protein kinase signaling cascade is required for zearalenone's effects on cell-cycle progression in MCF-7 cells. Given the presence of this mycotoxin in cereals, milk, and meat, the possibility that zearalenone is a potential promoter of breast cancer tumorigenesis should be investigated further. Mol. Carcinog. 30:88-98, 2001.


Isolated posterior cruciate ligament reconstruction. Long-term results.
From 1973 to 1987, 28 patients seen at our institution sustained isolated posterior cruciate ligament tears. Of these 28 patients, 25 were reevaluated at an average followup of 7 years and 1 month after secondary reconstruction of the posterior cruciate ligament using the semitendinosus and gracilis tendons alone or with an extraarticular procedure. Subjectively, 22 of 25 patients related no restrictions regarding activities of daily living, with 14 of 25 patients being able to return to their previous competitive level in sports. Objective evaluation after reconstruction revealed no change in the preoperative and postoperative posterior drawer examination in 13 of 25 patients, a finding confirmed by KT-1000 arthrometer measurements. Radiographic evaluation revealed degenerative changes predominantly involving the medial and patellofemoral compartments in 15 of 25 patients. Despite optimistic subjective reporting, this long-term retrospective study reveals that this procedure inconsistently limits posterior instability and therefore cannot be recommended.


Recent developments in nosocomial infections and their control.
Hospital epidemiology has advanced as a discipline largely because of contributions from government and certifying agencies, and from recognition that nosocomial infections are unique and require specialized attention. A modern infection control program should consist of surveillance, outbreak investigation, consultative services, education, and research activities. Guidance for program development may be obtained from an institution such as the Centers for Disease Control. This agency has provided standardized definitions as well as guidelines for care. An effective infection control program can improve the understanding and prevention of hospital-acquired infections.


Development of macaque secondary follicles exposed to neutral red prior to 3-dimensional culture.
Neutral red (NR) may assist identification of preantral follicles in pieces of cortical tissue prior to cryopreservation in cancer patients requesting fertility preservation. This study is the first to analyze this effect by follicle growth rate after long-term culture in primates. Ovarian cortex was obtained from adult rhesus macaques, was cut into fragments, and was incubated with NR. Secondary follicles were readily visualized following NR staining and then were encapsulated into alginate beads and cultured individually for 4 weeks in αMEM media supplemented with 10 ng/ml FSH at 5% O2. The survival rates of secondary follicles during culture were similar between those derived from control tissue (71 ± 13%) and those treated with NR (68 ± 9%). The proportion of surviving follicles that formed an antrum were also similar in both groups (70 ± 17% control; 48 ± 24% NR-treated). Follicle diameters were not different between control follicles (184 ± 5μm) and those stained with NR (181 ± 7 μm) on the day of isolation. The percentages of surviving follicles within three cohorts based on their diameters at week 4 of culture were similar between the control group and NR-stained tissue group, fast-grow follicles (24 ± 6% vs. 13 ± 10%), slow-grow follicles (66 ± 5% vs. 60 ± 9%), or no-grow (10 ± 9% vs. 27 ± 6%), respectively. There were no differences in follicle diameters between groups during the culture period. Pre-exposure of secondary follicles to NR diminished their capacity to produce both estradiol and androstenedione by week 4 of culture, when follicles are exhibiting an antrum. Inhibitory effects of NR on steroid production by slow-grow follicles was less pronounced. NR does not affect secondary follicle survival, growth, and antrum formation during long-term culture, but steroid hormone production by fast-grow follicles is compromised. NR can be used as a non-invasive tool for in situ identification of viable secondary follicles in ovarian cortex before tissue cryopreservation without affecting follicle survival and growth in vitro. Whether maturation or developmental competence of oocytes derived from antral follicles in 3D culture that were previously isolated from NR-stained tissue is normal or compromised remains to be determined. Likewise, the functional consequences of pre-exposure to NR prior to ovarian cortical tissue cryopreservation and transplantation are unknown.


A new principle of pulse detection based on terahertz wave plethysmography.
This study presents findings in the terahertz (THz) frequency spectrum for non-contact cardiac sensing applications. Cardiac pulse information is simultaneously extracted using THz waves based on the established principles in electronics and optics. The first fundamental principle is micro-Doppler motion effect. This motion based method, primarily using coherent phase information from the radar receiver, has been widely exploited in microwave frequency bands and has recently found popularity in millimeter waves (mmWave) for breathe rate and heart rate detection. The second fundamental principle is reflectance based optical measurement using infrared or visible light. The variation in the light reflection is proportional to the volumetric change of the heart, often referred as photoplethysmography (PPG). Herein, we introduce the concept of terahertz-wave-plethysmography (TPG), which detects blood volume changes in the upper dermis tissue layer by measuring the reflectance of THz waves, similar to the existing remote PPG (rPPG) principle. The TPG principle is justified by scientific deduction, electromagnetic wave simulations and carefully designed experimental demonstrations. Additionally, pulse measurements from various peripheral body parts of interest (BOI), palm, inner elbow, temple, fingertip and forehead, are demonstrated using a wideband THz sensing system developed by the Terahertz Electronics Lab at Arizona State University, Tempe. Among the BOIs under test, it is found that the measurements from forehead BOI gives the best accuracy with mean heart rate (HR) estimation error 1.51 beats per minute (BPM) and standard deviation 1.08 BPM. The results validate the feasibility of TPG for direct pulse monitoring. A comparative study on pulse sensitivity is conducted between TPG and rPPG. The results indicate that the TPG contains more pulsatile information from the forehead BOI than that in the rPPG signals in regular office lighting condition and thus generate better heart rate estimation statistic in the form of empirical cumulative distribution function of HR estimation error. Last but not least, TPG penetrability test for covered skin is demonstrated using two types of garment materials commonly used in daily life.


Body mass, mercury exposure, biochemistry and untargeted metabolomics of incubating common eiders (Somateria mollissima) in three Baltic colonies.
The Baltic/Wadden Sea Flyway of common eiders has declined over the past three decades. Multiple factors such as contaminant exposure, global warming, hunting, white-tailed eagle predation, decreased agricultural eutrophication and infectious diseases have been suggested to explain the decline. We collected information on body mass, mercury (Hg) concentration, biochemistry and untargeted metabolomics of incubating birds in two colonies in the Danish Straits (Hov Røn, n = 100; Agersø, n = 29) and in one colony in the Baltic proper (Christiansø, n = 23) to look into their metabolisms and energy balance. Body mass was available from early and late incubation for Hov Røn and Christiansø, showing a significant decline (25-30%) in both colonies with late body mass at Christiansø being the lowest. Whole blood concentrations of total mercury Hg were significantly higher in birds at Christiansø in the east compared to Hov Røn in the west. All birds in the three colonies had Hg concentrations in the range of ≤1.0 μg/g ww, which indicates that the risk of effects on reproduction is in the no to low risk category for wild birds. Among the biochemical measures, glucose, fructosamine, amylase, albumin and protein decreased significantly from early to late incubation at Hov Røn and Christiansø, reflecting long-term fastening as supported by the decline in body mass. Untargeted metabolomics performed on Christiansø eiders revealed presence of 8,433 plasma metabolites. Of these, 3,179 metabolites changed significantly (log2-fold change ≥1, p ≤ 0.05) from the early to late incubation. For example, smaller peptides and vitamin B2 (riboflavin) were significantly down-regulated while 11-deoxycorticosterone and palmitoylcarnitine were significantly upregulated. These results show that cumulative stress including fasting during incubation affect the eiders' biochemical profile and energy metabolism and that this may be most pronounced for the Christiansø colony in the Baltic proper. This amplify the events of temperature increases and food web changes caused by global warming that eventually accelerate the loss in body weight. Future studies should examine the relationship between body condition, temperature and reproductive outcomes and include mapping of food web contaminant, energy and nutrient content to better understand, manage and conserve the populations.


5-HT receptors: implications for the neuropharmacology of alcohol and alcoholism.
The involvement of serotonergic mechanisms in the neuropharmacology of alcohol was appreciated before it was recognized that there were multiple subtypes of serotonin (5-hydroxytryptamine; 5-HT) receptors. Thus, it was known that manipulations of the central serotonergic system could lead to a modification of the rate of tolerance development to alcohol (Frankel et al., 1975) or to a modulation of alcohol intake (Myers and Martin, 1973; Myers and Melchior, 1975) before Peroutka and Snyder (1979) first suggested that there were at least two subtypes of 5-HT receptors. Since these early reports were written, there has been a wealth of studies which have continued to support a role for 5-HT in the regulation of alcohol intake (See McBride et al., 1993b; Sellers et al., 1992, for reviews). Simultaneously, a tremendous expansion in the number of known 5-HT receptor subtypes has occurred (See Peroutka, 1988). However, there have not been, to our knowledge, any papers which have examined the possible role of specific 5-HT receptor subtypes in the regulation of alcohol's central effects. The present review addresses this deficiency in the literature. This review will focus on three major areas: the pharmacological regulation of alcohol intake; differences in 5-HT receptor subtypes among alcohol-preferring and -nonpreferring rat strains; and alterations in 5-HT receptor subtypes following chronic exposure to alcohol.


Formation of extracellular C14-C18 2-D-hydroxy fatty acids by species of Saccharomycopsis.
Eighteen of 19 strains of Saccharomycopsis fibuligera and one of two strains of S. capsularis produced mixtures of C(14)-C(18) 2-D-hydroxy acids in liquid culture medium. The mixture of these acids showed antimicrobial activity against Vibrio tyrogenus but not against the other microorganisms tested. Candida lactosa, a recently described species, was shown to be an isolates of S. fibuligera.


Contemporary external beam radiotherapy boost or high dose-rate brachytherapy boost for cervical cancer: a propensity-score-matched, nationwide, population-based cohort study.
To estimate the survival effects of contemporary external beam radiotherapy (EBRT) boost modalities (intensity-modulated radiation therapy or volumetric modulated arc therapy) and high dose-rate brachytherapy (HDR-BT) boost in patients with cervical cancer (CC). Patients who had been diagnosed as having CC were recruited from the Taiwan Cancer Registry Database. Propensity score matching was performed, and Cox proportional-hazards model curves were used to analyze the all-cause mortality of patients who received standard whole-pelvis irradiation with different boost modalities. The matching process yielded a final cohort of 1,630 patients (815 in the EBRT boost and HDR-BT boost groups, respectively) eligible for further analysis. The multivariate Cox regression analyses indicated that the adjusted hazard ratio (95% confidence intervals) for EBRT boost compared with HDR-BT boost was 1.62 (1.43-1.84). Multivariable analysis revealed that the independent poor prognostic factors of all-cause mortality among patients with CC were adenocarcinoma, no chemotherapy, Charlson comorbidity index score ≥ 1, age ≥ 60 years, and advanced International Federation of Gynecology and Obstetrics stage. HDR-BT boost may be more beneficial than contemporary EBRT boost in selected patients with CC.


Associations of Dietary Intake Levels with Ante-natal Depression in Pregnant Women.
This study was conducted to identify the association of dietary intake levels with ante-natal depression among pregnant women. Secondary data analysis was done using Can Pro software (version 3.0) to evaluate the diet intake levels with 24 hour recall diary method with 130 pregnant women visiting a general hospital. Statistical analysis included descriptive statistics, t-test, Mann-Whitney U test and adjusted logistic regression using the SPSS/WIN program. Whole dietary intake levels of pregnant women were insufficient according to Recommended Nutrient Intake (RI) with the except of phosphorus intake. There were significant differences in RI by gestation period and high risk pregnancies. Non-depressed women showed more intake in niacin (z=1.33, p=.018) and zinc (t=3.99, p=.048) than depressed women did. Niacin was a significant determinant of ante-natal depression (Exp (B)=5.88, p=.47, 95% CI [1.02~23.83]). Dietary intake assessment would be necessary during pregnancies for the screening of ante-natal depression. For ante-natal depression care, a tailored dietary intervention should be applied for women with insufficient nutrient intake.


Analysis of the N gene hypersensitive response induced by a fluorescently tagged tobacco mosaic virus.
The hypersensitive response (HR) triggered on Nicotiana edwardsonii by tobacco mosaic virus was studied using a modified viral genome that directed expression of the green fluorescent protein. Inoculated plants were initially incubated at 32 degrees C to inhibit the N gene-mediated HR. Transfer to 20 degrees C initiated the HR, and fluorescent infection foci were monitored for early HR-associated events. Membrane damage, which preceded visible cell collapse by more than 3 h, was accompanied by a transient restriction of the xylem within infection sites. Following cell collapse and the rapid desiccation of tissue undergoing the HR, isolated, infected cells were detected at the margin of necrotic lesions. These virus-infected cells were able to reinitiate infection on transfer to 32 degrees C, however, if maintained at 20 degrees C they eventually died. The results indicate that the tobacco mosaic virus-induced HR is a two-phase process with an early stage culminating in rapid cell collapse and tissue desiccation followed by a more extended period during which the remaining infected cells are eliminated.


Pre-pupation behaviour of the aphid parasitoid Aphidius ervi (Haliday) and its consequences for pre-imaginal learning.
Olfactory learning may occur at different stages of insect ontogeny. In parasitoid wasps, it has been mostly shown at adult emergence, whilst it remains controversial at pre-imaginal stages. We followed larval growth of the parasitoid wasp, Aphidius ervi Haliday, inside the host aphid, Acyrthosiphom pisum Harris, and characterised in detail the behaviour of third instar larvae. We found that just before cocoon spinning begins, the third instar larva bites a hole through the ventral side of the mummified aphid exoskeleton. We then evaluated whether this period of exposure to the external environment represented a sensitive stage for olfactory learning. In our first experiment, the third instar larvae were allowed to spin their cocoon on the host plant (Vicia faba L.) surface or on a plastic plate covering the portion of the host plant exposed to the ventral opening. Recently emerged adults of the first group showed a preference for plant volatiles in a glass Y-olfactometer, whereas no preference was found in adults of the second group. In a second experiment, during the period in which the aphid carcass remains open or is being sealed by cocoon spinning, third instar larvae were exposed for 24 h to either vanilla odours or water vapours as control. In this experiment, half of the parasitoid larvae were later excised from the mummy to avoid further exposure to vanilla. Adult parasitoids exposed to vanilla during the larval ventral opening of the mummy showed a significant preference for vanilla odours in the olfactometer, regardless of excision from the mummy. The larval behaviour described and the results of the manipulations performed are discussed as evidences for the acquisition of olfactory memory during the larval stage and its persistence through metamorphosis.


Molecular cloning of sequences from a Drosophila RNA polymerase II locus by P element transposon tagging.
We have identified a lethal mutation in the D. melanogaster RNA polymerase II locus, RpIIC4, caused by insertion of a transposable element associated with the phenomenon of hybrid dysgenesis (P element). Using previously cloned P element sequences as a hybridization probe we have isolated a hybrid lambda phage clone carrying a 10 kb genomic DNA fragment containing a 1.3 kb P element insert and flanking sequences from the RpII locus. The non-P sequences in this clone (lambda DmRpII-1) hybridize to polytene chromosome band region 10C, the cytogenetic location of RpIIC4, and revertants which lose the lethal RNA polymerase II mutation also lose P element sequences from the locus. We have generated several additional P element insertions into the locus and shown that they can occur at two or more different sites. These experiments illustrate that mutagenesis by P element insertion and use of cloned P DNA to retrieve the DNA sequences into which insertion has occurred may be a general method for cloning genetically defined loci in Drosophila.


RBM4 promotes neuronal differentiation and neurite outgrowth by modulating Numb isoform expression.
RBM4 participates in cell differentiation by regulating tissue-specific alternative pre-mRNA splicing. RBM4 also has been implicated in neurogenesis in the mouse embryonic brain. Using mouse embryonal carcinoma P19 cells as a neural differentiation model, we observed a temporal correlation between RBM4 expression and a change in splicing isoforms of Numb, a cell-fate determination gene. Knockdown of RBM4 affected the inclusion/exclusion of exons 3 and 9 of Numb in P19 cells. RBM4-deficient embryonic mouse brain also exhibited aberrant splicing of Numb pre-mRNA. Using a splicing reporter minigene assay, we demonstrated that RBM4 promoted exon 3 inclusion and exon 9 exclusion. Moreover, we found that RBM4 depletion reduced the expression of the proneural gene Mash1, and such reduction was reversed by an RBM4-induced Numb isoform containing exon 3 but lacking exon 9. Accordingly, induction of ectopic RBM4 expression in neuronal progenitor cells increased Mash1 expression and promoted cell differentiation. Finally, we found that RBM4 was also essential for neurite outgrowth from cortical neurons in vitro. Neurite outgrowth defects of RBM4-depleted neurons were rescued by RBM4-induced exon 9-lacking Numb isoforms. Therefore our findings indicate that RBM4 modulates exon selection of Numb to generate isoforms that promote neuronal cell differentiation and neurite outgrowth.


Reinforcement frequency, but not gender, determines sensitivity to discriminative stimulus effects of morphine.
We demonstrated previously that several parameters of a morphine discrimination were significantly different in female versus male rats, using a fixed ratio (FR)-10 schedule of food reinforcement; however, this schedule produced a significant bias in reinforcement frequency between saline and morphine sessions in males but not in females. To determine whether this schedule-drug-sex interaction caused the sex difference in discriminability of morphine, female and male rats were trained to discriminate 3.0 mg/kg morphine from saline using a variable interval (VI) 15-s/VI 15-s (Phase I), a VI 7.5-s/VI 15-s (Phase II), and a VI 15-s/VI 15-s (Phase III) schedule of food reinforcement on morphine/saline levers, respectively. After a minimum of 40 training sessions in each phase, mean reinforcement rates in morphine sessions were highest, and the ED50 values for morphine discrimination were lowest, in Phase II. Thus, as predicted, the morphine dose-effect curves shifted to the left from Phase I to Phase II, and back to the right from Phase II to III, presumably due to the bias in reinforcement rate between saline and morphine sessions that was induced by manipulating the VI schedule on the morphine lever. However, there were no sex differences in the morphine versus saline reinforcement rate or in discrimination ED50 in any phase, suggesting that the sex difference observed in our initial study was probably due to the bias in reinforcement frequency (towards the saline condition) that occurred only in males under the FR-10 schedule. This study demonstrates the importance of considering group differences in schedule-drug interactions when comparing discriminative stimulus properties of drugs between groups.


Nutritional assessment in elderly care: a MUST!
Malnutrition affects over three million people in the UK with associated health costs exceeding £13 billion annually.[1] In hospital, malnutrition has been shown to increase complication rates, morbidity, mortality, hospital readmissions, and length of hospital stay.[2] To screen for malnutrition, a reliable and validated screening tool such as the malnutrition universal screening tool (MUST) should be used.[3] We believe that improved patient outcomes and significant savings to the trust can be achieved, not only by ensuring that every patient has a MUST score documented, but that it is calculated correctly and the appropriate interventions are implemented. We have carried out the audit three times (May, July, and November 2013). The study included the patients on the elderly care ward of Watford General Hospital (n=64, 62, and 63 respectively). MUST scores documented in nursing notes for each patient were noted. We re-calculated each MUST score ourselves for comparison. We went through patient notes and nursing information and noted which recommended nutritional interventions were being implemented. Our results highlighted several issues: 1) Patients did not consistently have a MUST score documented 2) MUST scores were calculated incorrectly. This was generally due to BMIs calculated incorrectly, and patients' weights from six months ago not being known 3) High MUST scores not being acted on appropriately. Our interventions have involved liaising with various teams within the hospital to maximise the efficacy of the MUST score. This has included encouraging the trust to provide regular training to nurses because of high nursing staff turnover. Following our audit, the dietitian department agreed to undertake weekly ward rounds to screen for patients at risk of malnutrition. Our interventions so far have resulted in increased proportion of MUST scores being calculated (73 to 97%), and increased rates of patients being referred to dietitians (62 to 86% in the second audit cycle).


Improving ethanol tolerance of Escherichia coli by rewiring its global regulator cAMP receptor protein (CRP).
A major challenge in bioethanol fermentation is the low tolerance of the microbial host towards the end product bioethanol. Here we report to improve the ethanol tolerance of E. coli from the transcriptional level by engineering its global transcription factor cAMP receptor protein (CRP), which is known to regulate over 400 genes in E. coli. Three ethanol tolerant CRP mutants (E1- E3) were identified from error-prone PCR libraries. The best ethanol-tolerant strain E2 (M59T) had the growth rate of 0.08 h(-1) in 62 g/L ethanol, higher than that of the control at 0.06 h(-1). The M59T mutation was then integrated into the genome to create variant iE2. When exposed to 150 g/l ethanol, the survival of iE2 after 15 min was about 12%, while that of BW25113 was <0.01%. Quantitative real-time reverse transcription PCR analysis (RT-PCR) on 444 CRP-regulated genes using OpenArray® technology revealed that 203 genes were differentially expressed in iE2 in the absence of ethanol, whereas 92 displayed differential expression when facing ethanol stress. These genes belong to various functional groups, including central intermediary metabolism (aceE, acnA, sdhD, sucA), iron ion transport (entH, entD, fecA, fecB), and general stress response (osmY, rpoS). Six up-regulated and twelve down-regulated common genes were found in both iE2 and E2 under ethanol stress, whereas over one hundred common genes showed differential expression in the absence of ethanol. Based on the RT-PCR results, entA, marA or bhsA was knocked out in iE2 and the resulting strains became more sensitive towards ethanol.


The adhesion molecules of L-selectin and ICAM-1 in thrombocytosis and thrombocytopenia.
Thrombopoiesis is regulated by a variety of cytokines. Intracellular adhesion molecules are inducible cell-surface glycoproteins that belong to the immunoglobulin superfamily. Cytokines, endothelium and adhesive molecules represent the point of crosstalk in normal and pathological hematopoiesis. With the hypothesis that circulating intracellular adhesion molecule-1 (ICAM-1) and lymhocyte adhesion molecule-1 (L-selectin) concentrations could be changed based on pathological thrombopoiesis resulting in quantitative platelet disorders, we evaluated ICAM-1 and L-selectin levels in patients with thrombocytosis, thrombocytopenia and healthy controls. The L-selectin levels were found to be significantly higher in the thrombocytopenia group compared to the control group. ICAM-1 levels were found to be significantly higher in both thrombocytopenia and thrombocytosis groups compared to control group. Our study corroborates our original hypothesis implying the roles of adhesion molecules in the challenging status of pathological thrombopoiesis.


Erosive arthritis in monoclonal gammopathy of uncertain significance: report of four cases.
We report 4 case histories in which an erosive arthritis was associated with the presence of a monoclonal gammopathy of uncertain significance. In all 4 cases, the appearance of paraprotein was noted either before or during the development of the arthritis. Two patients had a rather atypical oligoarthritis, while the others had a rheumatoid-like, symmetric polyarthritis. A synovial amyloid deposit was present in 2 patients, while mild mixed mononuclear infiltrates were the main pathologic finding in the others. In 2 patients, immunohistochemical investigation demonstrated deposits of immunoglobulin-derived material of the same isotype as the monoclonal component in the synovial tissue.


Dating of iron gall ink using the dissolution-diffusion method.
In many criminal and civil cases, some questioned documents are written with iron gall ink. Determining the date when an iron gall ink entry was written can be important to assess the authenticity of a document. A dissolution-diffusion method was successfully employed to draw aging curves of iron gall ink entries stored in controlled conditions over 40 months. Calibration curves were created to indicate the relationship between the average dissolution-diffusion rate of ink components and the age of ink entries stored under natural aging conditions. As preliminary findings of this study, the mixed solution of dimethyl formamide (DMF) and anhydrous ethanol was suitable to dissolve the dye of iron gall ink strokes made at different time. It was also determined that brands of iron gall inks, types of paper, and thickness of iron gall ink strokes had varying impacts on estimating the dates of iron gall ink strokes.


Is native quantitative thought concretized in linguistically privileged ways? A look at the global picture.
This work investigates whether reference in speech to certain quantities, namely 1, 2, and 3, is privileged linguistically due to our brain's native quantitative capacities. It is suggested that these small quantities are not privileged in specific ways suggested in the literature. The case that morphology privileges these quantities, apart from 1, is difficult to maintain in light of the cross-linguistic data surveyed. The grammatical expression of 2 is explained without appealing to innate quantitative reasoning and the grammatical expression of 3 is not truly characteristic of speech once language relatedness is considered. The case that 1, 2, and 3 are each privileged lexically is also difficult to maintain in the face of the global linguistic data. While native neurobiological architecture biases humans towards recognizing small quantities in precise ways, these biases do not yield clear patterns in numerical language worldwide.