The effects of routine cage-changing on cardiovascular and behavioral parameters in male Sprague-Dawley rats.
The objective of this study was to determine whether the blood pressure and heart rate of adult male Sprague-Dawley rats are affected by the routine animal husbandry procedure of moving animals to clean cages. Cardiovascular parameters were obtained by using radiotelemetry; behavior in the home cage also was evaluated. Each rat had a radiotelemetry transmitter implanted in the peritoneal cavity, with the attached catheter placed in the femoral artery. After a 7- to 9-day recovery period, half of the rats were moved to clean cages with fresh wood-chip bedding; the other animals were left undisturbed. Systolic, diastolic, and mean arterial blood pressures; heart rate; and cage behavior (movement, rearing, grooming) increased promptly and significantly when animals were placed in clean cages. These cardiovascular and behavioral responses lasted for 45 to 60 min. Those animals not moved to clean cages but present in the animal room when this procedure was done did not show significant increases in blood pressure, heart rate, or activity. When rats were moved to clean cages that contained new bedding plus a small quantity of the soiled bedding from their previous cage, the cardiovascular and behavioral responses were similar to those of animals exposed to completely fresh bedding. The responses of rats being moved to new cages did not diminish between the first and fourth weekly cage change. Rats whose cages were not changed for 2 weeks showed small, but significant, increases in cardiovascular and behavioral responses above the responses in animals with weekly cage changes. We conclude that ordinary animal husbandry procedures such as moving rats to a clean cage can induce transient, but significant, cardiovascular and behavioral changes. Investigators and animal care staff should recognize that such routine procedures could confound experiments conducted shortly thereafter.


The Role of Thyroid Hormone in the Innate and Adaptive Immune Response during Infection.
In the past decades, there has been growing evidence for a functional interaction between the thyroid hormone and the immune system. This article provides an overview of the mechanisms by which thyroid hormones affect the innate and adaptive immune response during infection. The influence of thyroid hormone on the most important players of the innate [neutrophils, macrophages, natural killer (NK) cells, and dendritic cells (DCs)] and adaptive immune system (B- and T-lymphocytes) is reviewed here based on both clinical and preclinical studies. The effects of modulation of the immune system by drugs, such as monoclonal antibodies, tyrosine kinase inhibitors, and interferons on thyroid function, are beyond the scope of this article. Thyroid hormones regulate the activity of neutrophils which is reflected by higher numbers of neutrophils outside the bloodstream and enhanced activity of the respiratory burst following stimulation with thyroid hormone. Hyperthyroidism affects neutrophil function to a larger extent than hypothyroidism. In addition to neutrophil function, macrophage function is strongly affected by thyroid hormones, with triiodothyronine having a pro-inflammatory effect in these cells. NK cell proliferation and cytotoxic activity are also dependent on thyroid hormone levels. Finally, thyroid hormones enhance DC proliferation and maturation. In the adaptive immune system, a hyperthyroid state leads to increased activation of lymphocytes. This effect of thyroid hormone is mediated by various factors including NF-κB and protein kinase C signaling pathways and the β-adrenergic receptor. In general, a hyperthyroid state leads to a more activated immune system whereas hypothyroidism leads to a less activated immune system. © 2020 American Physiological Society. Compr Physiol 10:1277-1287, 2020.


The Jak/STAT pathway: A focus on pain in rheumatoid arthritis.
Pain is a manifestation of rheumatoid arthritis (RA) that is mediated by inflammatory and non-inflammatory mechanisms and negatively affects quality of life. Recent findings from a Phase 3 clinical trial showed that patients with RA who were treated with a Janus kinase 1 (Jak1) and Janus kinase 2 (Jak2) inhibitor achieved significantly greater improvements in pain than those treated with a tumor necrosis factor blocker; both treatments resulted in similar changes in standard clinical measures and markers of inflammation. These findings suggest that Jak1 and Jak2 inhibition may relieve pain in RA caused by inflammatory and non-inflammatory mechanisms and are consistent with the overarching involvement of the Jak-signal transducer and activator of transcription (Jak/STAT) pathway in mediating the action, expression, and regulation of a multitude of pro- and anti-inflammatory cytokines. In this review, we provide an overview of pain in RA, the underlying importance of cytokines regulated directly or indirectly by the Jak/STAT pathway, and therapeutic targeting of the Jak/STAT pathway in RA. As highlighted herein, multiple cytokines directly or indirectly regulated by the Jak/STAT pathway play important roles in mediating various mechanisms underlying pain in RA. Having a better understanding of these mechanisms may help clinicians make treatment decisions that optimize the control of inflammation and pain.


Room to Breathe: The Impact of Oxygen Rationing on Health Outcomes in SARS-CoV2.
As the primary surge of coronavirus disease 2019 (COVID-19) wanes in many countries, it is important to reconsider best practice. More cases, probably the majority of cases, are yet to come. Hopefully, during this next phase, we will have more time, more resources, and more experience from which to affect better outcomes. Here, we examine the compromised oxygen strategy that many nations followed. We explore the evidence related to such strategies and discuss the potential mortality impact of delaying oxygen treatment in COVID-19 pneumonia.


Cytomegalovirus infection: a quantitative prospective study of three hundred twenty consecutive renal transplants.
The quantitative effects of cytomegalovirus (CMV) infection on morbidity and mortality rates were examined in 320 renal transplant cases. With the use of virus cultures and CMV antibody measurements, all patients were studied, regardless of symptoms, from a time before transplantation to at least 1 year, 11 months after transplantation for a maximum of 5 years, 9 months. The posttransplant risk factors of CMV infection--patient age, type of donor (living-related or cadaver), antigen match between donor and recipient, presence of diabetes, and the presence of pretransplant CMV antibody--were evaluated for their relative effects on patient survival, graft survival, fever, and leukopenia. CMV infection was a significant risk factor for these four events. CMV infection occurred in 181 patients after transplantation and accounted for 25% of the deaths, 20% of the graft failures, 30% of the occurrences of fever, and 35% of the occurrences of leukopenia. Unexpectedly, female recipients were at higher risk than men for the adverse effects of CMV infection. Young patients and those receiving their second transplant were at higher risk of graft loss if they had associated CMV infection. CMV infection was most reliably predicted by the presence of pretransplant antibody, indicating that reactivation of endogenous virus was responsible for most infections. The presence of pretransplant antibody offered a small amount of protection against fever, but no protection against death, graft failure, or leukopenia. Simultaneous episodes of CMV infection and transplant rejection, both common posttransplant events, most often occurred by chance.


Headache in adolescents in Dutch general practice.
We set out to describe the occurrence of headache in adolescents in general practice, the diagnoses made, the referrals to specialised care and the role of psychosocial problems in idiopathic headache. Data from the Dutch national survey of morbidity and interventions in general practice were used. The practice population (103 practices) comprised 37,187 children aged 12-18 years. In 650 episodes headache was a reason for encounter. The incidence of headache as presented to the GP was 69.9 per 1000 person years. The incidence in girls was twice as high as in boys. Half of the episodes with headache complaints were related to an infectious disease. The incidence rate of idiopathic headache was 18.0 per 1000 person years. Eight percent of all headache episodes were diagnosed as a psychosocial problem. Of all episodes, 25% percent were diagnosed as idiopathic headache (i.e. headache as symptom diagnosis, tension headache or migraine). Of these episodes, 21% were associated with an underlying psychosocial problem. Referrals were made by the GP in three percent of all headache episodes and four percent of idiopathic headache episodes. Headache in adolescents is a common reason for visiting a general practice. The symptom is often associated with an infectious disease. The GP is aware of underlying psychosocial problems. Only few children are referred for specialist care.


A highly optimized grid deployment: the metagenomic analysis example.
Computational resources and computationally expensive processes are two topics that are not growing at the same ratio. The availability of large amounts of computing resources in Grid infrastructures does not mean that efficiency is not an important issue. It is necessary to analyze the whole process to improve partitioning and submission schemas, especially in the most critical experiments. This is the case of metagenomic analysis, and this text shows the work done in order to optimize a Grid deployment, which has led to a reduction of the response time and the failure rates. Metagenomic studies aim at processing samples of multiple specimens to extract the genes and proteins that belong to the different species. In many cases, the sequencing of the DNA of many microorganisms is hindered by the impossibility of growing significant samples of isolated specimens. Many bacteria cannot survive alone, and require the interaction with other organisms. In such cases, the information of the DNA available belongs to different kinds of organisms. One important stage in Metagenomic analysis consists on the extraction of fragments followed by the comparison and analysis of their function stage. By the comparison to existing chains, whose function is well known, fragments can be classified. This process is computationally intensive and requires of several iterations of alignment and phylogeny classification steps. Source samples reach several millions of sequences, which could reach up to thousands of nucleotides each. These sequences are compared to a selected part of the "Non-redundant" database which only implies the information from eukaryotic species. From this first analysis, a refining process is performed and alignment analysis is restarted from the results. This process implies several CPU years. The article describes and analyzes the difficulties to fragment, automate and check the above operations in current Grid production environments. This environment has been tuned-up from an experimental study which has tested the most efficient and reliable resources, the optimal job size, and the data transference and database reindexation overhead. The environment should re-submit faulty jobs, detect endless tasks and ensure that the results are correctly retrieved and workflow synchronised. The paper will give an outline on the structure of the system, and the preparation steps performed to deal with this experiment.


Hypertropic cardiomyopathy and hyperthyroidism in the cat.
In a 21/2-year period, hypertrophic cardiomyopathy was found at necropsy of 23 cats that died (13 cats) or were euthanatized (10) because of problems associated with hyperthyroidism. Of these, 4 (17%) also had evidence of cardiac failure (pulmonary edema or pleural effusion). The mean body weight of the cats with hyperthyroidism and hypertrophic cardiomyopathy was significantly less (P less than 0.001) than that of clinically normal cats and cats with primary cardiomyopathy (congestive or restrictive) or excessive moderator band cardiomyopathy. In addition, the ratio of heart weight to body weight was significantly greater (P less than 0.001) in the 23 hyperthyroid cats than in the normal cats and cats with primary cardiomyopathy. Twenty (87%) of the cats had symmetric hypertrophy of the ventricular septum and left ventricular free wall, whereas the remaining 3 cats had disproportionate thickening of the ventricular septum, compared with the free wall, similar to what is found in cats with asymmetric hypertrophic cardiomyopathy. Histologic cardiac abnormalities included large, hyperchromatic nuclei, interstitial fibrosis, endocardial fibroplasia, fibrosis of the atrioventricular node, and marked disorganization of cardiac muscle cells. The study showed that hypertrophic cardiomyopathy develops in most hyperthyroid cats, some of which also develop congestive heart failure. Although the signs of heart disease in primary myocardial disease and thyrotoxic disease are similar, the characteristic signalment and clinical signs of hyperthyroidism should lead one to suspect the association of hypertrophic cardiomyopathy with the hyperthyroidism.


Use of a decision aid including information on overdetection to support informed choice about breast cancer screening: a randomised controlled trial.
Mammography screening can reduce breast cancer mortality. However, most women are unaware that inconsequential disease can also be detected by screening, leading to overdiagnosis and overtreatment. We aimed to investigate whether including information about overdetection of breast cancer in a decision aid would help women aged around 50 years to make an informed choice about breast screening. We did a community-based, parallel-group, randomised controlled trial in New South Wales, Australia, using a random cohort of women aged 48-50 years. Recruitment to the study was done by telephone; women were eligible if they had not had mammography in the past 2 years and did not have a personal or strong family history of breast cancer. With a computer program, we randomly assigned 879 participants to either the intervention decision aid (comprising evidence-based explanatory and quantitative information on overdetection, breast cancer mortality reduction, and false positives) or a control decision aid (including information on breast cancer mortality reduction and false positives). Participants and interviewers were masked to group assignment. The primary outcome was informed choice (defined as adequate knowledge and consistency between attitudes and screening intentions), which we assessed by telephone interview about 3 weeks after random allocation. The primary outcome was analysed in all women who completed the relevant follow-up interview questions fully. This trial is registered with the Australian New Zealand Clinical Trials Registry, number ACTRN12613001035718. Between January, 2014, and July, 2014, 440 women were allocated to the intervention group and 439 were assigned to the control group. 21 women in the intervention group and 20 controls were lost to follow-up; a further ten women assigned to the intervention and 11 controls did not answer all questions on attitudes. Therefore, 409 women in the intervention group and 408 controls were analysed for the primary outcome. 99 (24%) of 409 women in the intervention group made an informed choice compared with 63 (15%) of 408 in the control group (difference 9%, 95% CI 3-14; p=0·0017). Compared with controls, more women in the intervention group met the threshold for adequate overall knowledge (122/419 [29%] vs 71/419 [17%]; difference 12%, 95% CI 6-18; p<0·0001), fewer women expressed positive attitudes towards screening (282/409 [69%] vs 340/408 [83%]; 14%, 9-20; p<0·0001), and fewer women intended to be screened (308/419 [74%] vs 363/419 [87%]; 13%, 8-19; p<0·0001). When conceptual knowledge alone was considered, 203 (50%) of 409 women in the intervention group made an informed choice compared with 79 (19%) of 408 in the control group (p<0·0001). Information on overdetection of breast cancer provided within a decision aid increased the number of women making an informed choice about breast screening. Becoming better informed might mean women are less likely to choose screening. Australian National Health and Medical Research Council.


Ameliorative Effect of Trans-Sinapic Acid and its Protective Role in Cerebral Hypoxia in Aluminium Chloride Induced Dementia of Alzheimer's Type.
Trans-Sinapic Acid is a bioactive compound. Recent studies showed that it has a significant potential to attenuate various chemically induced Neurodegenerative toxicities. The present study investigates the potential of trans-Sinapic Acid as neuromodulator and its effect on release of Monoamine Oxidase (MAO-A, MAO-B), TNF-α, Acetylcholine esterase Enzyme, in cognitive dysfunctions associated with experimental dementia. Experiment: Aluminium chloride was administered at a dose of 175mg/kg, p.o. for a period of 25 days in rats and then divided into different groups, i.e. Treatment group, negative control and two groups of trans- Sinapic Acid, (at a dose of 30 and 60mg/kg, p.o.), where these groups treated and observed until the 35th day of experimental trial. Morris water Maze (MWM) and Photoactometer was used to access learning, memory and ambulatory movements on 5th, 16th, 26th and 36th day of experiment. Later, the animals were sacrificed for biochemical and histopahological studies. The oxidative stress was measured by estimating the levels of Glutathion (GSH), Superoxide dismutase (SOD), Nitrite, Catalase. Brain acetylcholine esterase (Ache) activity and Monoamine oxidase (MAO-A, MAO-B) were also estimated. The Brain level of TNF-α was measured as a marker of inflammation. Aluminium chloride (AlCl3) produced a marked decline in MWM performance and ambulatory movements' of animals, reflecting impairment of memory and learning. Trans-Sinapic Acid treatment significantly modulates AlCl3 induced memory deficits, biochemical and pathological alterations. The findings demonstrate that the memory restorative ability of trans-Sinapic Acid may be attributed to its anti-cholinesterase, anti-oxidative and anti-inflammatory potential.


Fasciolosis.
Fasciolosis is a zoonosis affecting ruminants, caused by the liver flukes Fasciola (F.) hepatica, and F. gigantica, which infect at least 2.4 million people worldwide. This disease may occur in cluster or family infections or after travel in high-risk areas such as the Nile Delta in Egypt, Iran, Turkey, South-East Asia, Mexico, the Caribbean and the Andean Altiplano. In Europe, fasciolosis occurs more frequently in Portugal, Spain and France, although autochthonous infections have also been reported from Ireland and Germany. Infectious metacercariae are ingested with contaminated water or raw or undercooked vegetables. During their larval stage immature flukes migrate through the liver producing an acute febrile syndrome some weeks after infection, followed by a chronic-latent stage which may last for years or decades. Acute fasciolosis is characterized by fever, high eosinophilia and hepatosplenomegaly. At this stage ova are usually not yet produced. Diagnosis relies on the detection of specific antibodies and/or antigens in serum. Typical imaging features include multiple, ill-defined, fleeting hypodense or hypoechoic areas in the liver. Intraabdominal bleeding due to fluke's penetration of the bowel wall or liver capsule, may occur. In chronic latent fasciolosis the diagnosis is achieved by specific serology tests, detection of eggs in bile and parasitological examinations of multiple enriched stools samples. Ultrasonography may sometimes reveal a dilated and thickened common bile duct or crescent-like parasites in the gallbladder or bile ducts. Some patients exhibit sludge which typically does not sediment. Triclabendazole at a single dose of 10 mg/kg body weight is the treatment of choice.


Frequency of kdr mutations in the voltage-sensitive sodium channel (VSSC) gene in Aedes aegypti from Yogyakarta and implications for Wolbachia-infected mosquito trials.
In the inner city of Yogyakarta, Indonesia, insecticide resistance is expected in the main dengue vector, Aedes aegypti, because of the intensive local application of pyrethroid insecticides. However, detailed information about the nature of resistance in this species is required to assist the release of Wolbachia mosquitoes in a dengue control program, so that we can ensure that insecticide resistance in the strain of Ae. aegypti being released matches that of the background population. High-resolution melt genotyping was used to screen for kdr mutations associated with pyrethroid resistance in the voltage-sensitive sodium channel (VSSC) gene in Ae. aegypti of some areas in the inner city of Yogyakarta. The results show that the V1016G mutation predominated, with individuals homozygous for the 1016G allele at a frequency of 82.1% and the mutant allele G at a frequency of 92%. Two patterns of co-occurrence of mutations were detected in this study, homozygous individuals V1016G/S989P; and heterozygous individuals V1016G/F1534C/S989P. We found the simultaneous occurrence of kdr mutations V1016G and F1534C at all collection sites, but not within individual mosquitoes. Homozygous mutants at locus 1016 were homozygous wild-type at locus 1534 and vice versa, and heterozygous V1016G were also heterozygous for F1534C. The most common tri-locus genotype co-occurrences were homozygous mutant 1016GG and homozygous wild-type FF1534, combined with homozygous mutant 989PP (GG/FF/PP) at a frequency of 38.28%. Given the relatively small differences in frequency of resistance alleles across the city area, locality variations in resistance should have minor implications for the success of Wolbachia mosquito trials being undertaken in the Yogyakarta area.


Metabolic changes in the knockout mouse for Canavan's disease: implications for patients with Canavan's disease.
Canavan's disease is an autosomal recessive disorder caused by aspartoacylase deficiency, which leads to accumulation of N-acetylaspartic acid in the brain and blood and an elevated level of N-acetylaspartic acid in the urine. The brain of patients with Canavan's disease shows spongy degeneration. How the enzyme deficiency and elevated N-acetylaspartic acid cause the pathophysiology observed in Canavan's disease is not obvious. The creation of a knockout mouse for Canavan's disease is being used as a tool to investigate metabolic pathways in the mouse and correlate them with the patients with Canavan's disease. The level of glutamate is lower in the knockout mouse brain than in the wild-type mouse brain, similar to what we have found in children with Canavan's disease, and so are the levels of gamma-aminobutyric acid (GABA). The level of aspartate is higher in the Canavan's disease mouse brain. The activity of aspartate aminotransferase, an enzyme involved in the malate-aspartate shuttle, is lower in the Canavan's disease mouse brain. The lower weight of the Canavan's disease mouse was in direct proportion to low total-body fat and bone mineral density. These changes might be similar to what is seen in patients with Canavan's disease and could have therapeutic implications.


Investigating the molecular mechanisms of learning and memory using Caenorhabditis elegans.
Learning is an essential biological process for survival since it facilitates behavioural plasticity in response to environmental changes. This process is mediated by a wide variety of genes, mostly expressed in the nervous system. Many studies have extensively explored the molecular and cellular mechanisms underlying learning and memory. This review will focus on the advances gained through the study of the nematode Caenorhabditis elegans. C. elegans provides an excellent system to study learning because of its genetic tractability, in addition to its invariant, compact nervous system (~300 neurons) that is well-characterised at the structural level. Importantly, despite its compact nature, the nematode nervous system possesses a high level of conservation with mammalian systems. These features allow the study of genes within specific sensory-, inter- and motor neurons, facilitating the interrogation of signalling pathways that mediate learning via defined neural circuits. This review will detail how learning and memory can be studied in C. elegans through behavioural paradigms that target distinct sensory modalities. We will also summarise recent studies describing mechanisms through which key molecular and cellular pathways are proposed to affect associative and non-associative forms of learning.


Pregnant African American women's attitudes toward perinatal depression prevention.
Depression during the perinatal period is common, debilitating, and consequential for women and their children, particularly among low income African American women. Viable approaches to prevention of depression have emerged. Yet little is known about women's preferences for approaches to preventing depression. A sample of 60 pregnant, low-income African American women seeking routine prenatal care was presented with standardized descriptions of three approaches to depression prevention (pharmacotherapy and two psychosocial approaches) and measures of preferences, perceived credibility, and personal reactions to each approach. Women also completed measures of perceived barriers and facilitators and current depression. Consistent with expectations, both of the psychosocial approaches were rated as more strongly preferred, more credible, and associated with more positive personal reactions relative to the pharmacotherapy approach. Depression did not alter women's preferences among the approaches. Contrary to prediction, women with clinically significant levels of depression did not find the pharmacotherapy approach to be more credible or to have more favorable personal reactions to it than women with low depression symptom levels. Exploration of women's perceptions of barriers revealed the importance of logistics, beliefs, and stigma barriers whereas women reported that concern about depression being impairing and ease of pragmatics would both facilitate engagement with preventive interventions. The findings suggest the need to examine the role of preferences in tests of the effectiveness of approaches to the prevention of perinatal depression in order to enhance service delivery among low income African American women.


Olfactory disorders and quality of life--an updated review.
Olfactory disorders are common and affect about one-fifth of the general population. The main causes of olfactory loss are post viral upper respiratory infection, nasal/sinus disease, and head trauma and are therefore very frequent among patients in ear, nose, and throat clinics. We have systematically reviewed the impact of quantitative, qualitative, and congenital olfactory disorders on daily life domains as well as on general quality of life and depression. From the extensive body of literature, it can be concluded that loss of the sense of smell leads to disturbances in important areas, mainly in food enjoyment, detecting harmful food and smoke, and to some extent in social situations and working life. Most patients seem to deal well and manage those restrictions. However, a smaller proportion has considerable problems and expresses a noticeable reduction in general quality of life and enhanced depression. The impact of coping strategies is discussed.


Mortality and survival among a cohort of drug injectors in Glasgow, 1982-1994.
There has been much speculation about the nature and extent of mortality among drug injectors in Glasgow. In order to determine injectors' mortality rate and compare this rate to the general population, identifier information from 459 drug injectors who received treatment for drug misuse in Glasgow between 1982 and 1994 was linked to the Scottish Mortality Register. The average duration of follow-up from cohort entry was 5.5 years and 10.2 years from commencement of drug injection. By the end of 1994, 53 cohort members had died. The average annual mortality rate of 1.8% was the same as that observed in a London cohort followed-up from 1969 to 1991. However, the excess mortality ratio (EMR) of 22.0 was almost double the London rate (11.9) because of the much lower average age of mortality (26.3 vs. 38.2 years). There was no significant time trend in EMR. Kaplan-Meier hazard analyzes show that younger patients and those who were HIV positive had significantly elevated mortality rates. The main cause of death was overdose, although it is unclear how many were accidental and how many intentional. Three of the six fatalities among HIV positive injectors were AIDS related. This study enables the first realistic assessment of the hypothesis that drug-related deaths in Glasgow are especially high. In relation to other populations of drug injectors, the annual mortality rate is comparable, although the average age of mortality is much lower in Glasgow. Consequently, in comparison to the general population, the mortality rate of drug injectors is higher in Glasgow compared to other cities.


Second re-irradiation: A delicate balance between safety and efficacy.
Except for straightforward palliative indications such as painful bone metastases, re-irradiation is often characterized by a narrow therapeutic window and the potential for decreased efficacy and increased toxicity, especially if the cumulative total dose from both courses is high. Second re-irradiations tend to pose even bigger challenges and are thus offered in a restrictive manner to highly selected patients on a case-by-case basis. Normal tissue dose constraints are still an area of active investigation. Nevertheless, examples of potentially useful indications have been published. The present review briefly summarizes areas of uncertainty and opportunities for future research. If evidence-based concepts with acceptable side effect profiles can be developed, an increasing number of patients may benefit from additional radiotherapy to previously exposed target volumes.


Nondestructive interferometric measurement of the delta and alpha of clad optical fibers.
It is shown that interferograms generated by passing light perpendicular to the axis of a fiber can be used to obtain the maximum refractive index difference and the index profile shape to better than +/-10% accuracy in regions of the core where alpha is constant. This technique avoids the time consuming sample preparation required for the slab method, the propagation problems associated with the near-field technique, and the surface quality problem associated with the reflection technique.


Bisubstrate analogue inhibitors of 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase: synthesis and biochemical and crystallographic studies.
6-Hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) catalyzes the transfer of pyrophosphate from ATP to 6-hydroxymethyl-7,8-dihydropterin (HP), leading to the biosynthesis of folate cofactors. Like other enzymes in the folate pathway, HPPK is an ideal target for the development of antimicrobial agents because the enzyme is essential for microorganisms but is absent from human and animals. Three bisubstrate analogues have been synthesized for HPPK and characterized by biochemical and X-ray crystallographic analyses. All three bisubstrate analogues consist of a pterin, an adenosine moiety, and a link composed of 2-4 phosphoryl groups. P(1)-(6-Hydroxymethylpterin)-P(2)-(5'-adenosyl)diphosphate (HP(2)A, 5) shows little affinity and inhibitory activity for E. coli HPPK. P(1)-(6-Hydroxymethylpterin)-P(3)-(5'-adenosyl)triphosphate (HP(3)A, 6) shows moderate affinity and inhibitory activity with K(d) = 4.25 microM in the presence of Mg(2+) and IC(50) = 1.27 microM. P(1)-(6-Hydroxymethylpterin)-P(4)-(5'-adenosyl)tetraphosphate (HP(4)A, 7) shows the highest affinity and inhibitory activity with K(d) = 0.47 microM in the presence of Mg(2+) and IC(50) = 0.44 microM. The affinity of MgHP(4)A for HPPK is approximately 116 and 76 times higher than that of MgADP and 6-hydroxymethylpterin, respectively. The crystal structure of HPPK in complex with 7 (HPPK.MgHP(4)A) has been determined at 1.85 A resolution with a crystallographic R factor of 0.185. The crystal structure shows that 7 occupies both HP- and ATP-binding sites and induces significant conformational changes in HPPK. The biochemical and structural studies of the bisubstrate analogues indicate that the bisubstrate analogue approach can produce more potent inhibitors for HPPK and the minimum length of the link for a bisubstrate analogue is approximately 7 A.


Study of Optimal Perimetric Testing in Children (OPTIC): evaluation of kinetic approaches in childhood neuro-ophthalmic disease.
We compared feasibility, quality and outcomes of visual field (VF) testing in children with neuro-ophthalmic disease between the discontinued 'gold-standard' Goldmann and Octopus perimeters. Children with neuro-ophthalmic disease, attending Great Ormond Street Hospital, London, were assessed using standardised protocols by one examiner in a single sitting, using Goldmann and Octopus kinetic perimetry. Outputs were classified to compare severity of loss and defect type. Test quality was assessed using both qualitative and quantitative methods. Thirty children (40% female) aged 5-15 years participated. Goldmann perimetry was completed in full by 90.0% vs 72.4% for Octopus. Inability to plot the blind spot was the most common reason for not completing testing. Over 75% completed a test in ≤20 min. Duration was similar between perimeters (paired t-test, mean difference: 0.48min (-1.2, 2.2), p=0.559). The lowest quality tests were for Octopus perimetry in children <8 years, without significant differences between perimeters in older children (McNemar's test, χ2=1.0, p=0.317). There was broad agreement between Goldmann and Octopus outputs (good quality, n=21, Bland-Altman, mean difference for isopters I4e (-514.3 deg2 (-817.4, -211.2), p=0.814), I2e (-575.5 deg2 (-900.1, -250.9), p=0.450) and blind spot (20.8 deg2 (5.7, 35.8), p=0.451)). However, VF severity grades and defect type matched in only 57% and 69% of tests, respectively. Octopus perimetry underestimated severe VF defects. Informative perimetry is feasible in children ≥8 years with neuro-ophthalmic conditions, with either Goldmann or Octopus perimeters. However, meaningful differences exist between the two approaches with implications for consistency in longitudinal assessments.


Inhaled nitric oxide versus intravenous vasodilators in severe pulmonary hypertension after cardiac surgery.
Inhaled nitric oxide (iNO) is superior to i.v. vasodilators for treatment of pulmonary hypertension (PH) after cardiac surgery, but iNO is a potentially toxic gas, and patient subsets who benefit from iNO are not yet clearly defined. We administered iNO 40 ppm, prostaglandin E1 (PGE1) 0.1 microg x kg(-1) min(-1), and nitroglycerin (NTG) 3 to 5 microg x kg(-1) min(-1), in a randomized crossover study to 14 adult patients with severe PH after cardiac surgery. iNO, PGE1, and NTG were of similar efficacy in reducing pulmonary vascular resistance (P = 0.003). iNO induced selective pulmonary vasodilation, while PGE1 and NTG had significant concomitant systemic vasodilatory effects. iNO led to an increase in cardiac index (CI) (P = 0.012), and PGE1 increased CI (P = 0.006) and right ventricular (RV) ejection fraction (P = 0.015), while NTG had no effect on CI and RV performance. After study completion, patients continued with PGE1 administration with favorable in-hospital outcome. We conclude that PH per se, even if severe, does not necessarily imply postoperative RV dysfunction, and selective pulmonary vasodilation with iNO may not be superior to PGE1 with regard to CI and RV performance. In a prospective, randomized crossover study of inhaled nitric oxide (iNO) versus IV vasodilators, performed in adult patients with severe pulmonary hypertension but preserved right ventricular function after cardiac surgery, iNO was not superior to IV prostaglandin E1 with regard to cardiac index and right ventricular performance. Considering the potential toxicity of iNO, better definition of patient subsets with a positive benefit/risk ratio is warranted.


Effect of RNAi-mediated silencing of Livin gene on biological properties of colon cancer cell line LoVo.
This study aimed to investigate the effect of RNAi-mediated silencing of the Livin gene on biological properties of the colon cancer cell line LoVo. Interference vectors pSilencer4.1-Ll and pSilencer4.1-L2 targeting the Livin gene were constructed and transfected into LoVo cells. The expression of the Livin gene was determined by RT-PCR and Western blotting. The apoptosis, cell cycle, colony formation, proliferation of LoVo cells, as well as their sensitivity to cisplatin, were detected by flow cytometry, colony formation assay and MTT. Livin mRNA and protein expression in LoVo cells could be effectively silenced by pSilencer4.1-Ll but not pSilencer4.1-L2. In the pSilencer4.1-Ll transfection group, the apoptosis rate of LoVo cells was significantly higher than in the control group (24.2 ± 3.2 vs 8.1 ± 1.4%, P < 0.01), and after 72 h, cell proliferation was clearly decreased (about 70% inhibition). Compared with the control group, the colony formation rate in pSilencer4.1-Ll transfection group was obviously decreased (15 ± 4.6 vs 85 ± 5.8%, P < 0.01), with increased proportion of S phase cells (45.7 ± 4.9 vs 28.0 ± 3.0%, P < 0.01), decreased proportion of G1 phase cells (43.0 ± 5.2 vs 62.8 ± 5.1%, P < 0.01), and increased sensitivity to cisplatin (apoptosis rate increased from 43.4 ± 6.9 to 65.3 ± 6.2%, P < 0.01). pSilencer4.1-Ll can effectively silence Livin gene expression in LoVo colon cancer cells, inhibit cell proliferation and colony formation, induce apoptosis, and enhance sensitivity to cisplatin.


Retooling the Remediation Process: An Overview of One School's Process Overhaul.
This article describes the process of review and modification of a Doctor of Pharmacy didactic course remediation process. A highlight of the new process includes early intervention with a Learning Specialist, creating the opportunity for students to address learning issues earlier in the courses. Review of past remediation processes, student success, and pertinent literature related to remediation processes to allow for a new remediation process that supports students' learning and success. Creation of a new 2-phase process, including an early intervention and remediation process. The process includes a Learning Specialist position to focus on learning and barriers to academics. Remediation opportunities focus on proficiency in specific areas for each course. The new remediation policy provides greater support to students through the Learning Specialist. The new policy helps students connect with resources earlier in the program and provides multiple opportunities for assisting students during the semester. Over the past 4 years, the Learning Specialist has met with an average of 73 individual students per semester, while only an average of 25 students in the didactic portion of the curriculum (year 1-3) require remediation each semester.


Network connectivity is shown to change in C57BL/6 mice during a continuing immune response subsequent to tetanus toxoid hyperimmunization.
We have already demonstrated (Stojanovic et al., 2009) a connection between tetanus toxoid (TTd) hyperimmunization and the induction of anti-phospholipid syndrome (APS) in BALB/c mice. Here we show that C57BL/6 mice subjected to an identical procedure do not exhibit any like pathology attributable to anti-phospholipid antibodies; we explain that this absence results from idiotypic connectivity. Six groups of C57BL/6 mice were hyperimmunized with TTd in aluminum hydroxide or glycerol, with or without pretreatments. Pretreated mice had been injected with polyclonal or nonspecific immune stimulators, such as complete Freund's adjuvant (CFA) or glycerol. The epitope specificity of induced antibodies was tested by indirect ELISA using a tetanus toxoid immunogen and these autoantigens: phospholipids, gangliosides, laminin. Idiotypic connectivity was tested by competitive ELISA and gauged from the degree to which the interaction of idiotypic/anti-idiotypic complementary antibodies was inhibited in the presence of immunized sera antibodies. Higher idiotypic connectivity was noted amongst pretreated mice. There was a positive correlation between higher connectivity and autoantibody levels that acted to favor the participation of natural autoantibodies in the inhibitory process. We conclude that idiotypic connectivity plays a protective role in immunization-induced autoimmunity.


Extra-glissonian Approach for Total Laparoscopic Left Hepatectomy: A Prospective Cohort Study.
Laparoscopic liver resection under hemihepatic vascular inflow control has advantages over Pringle's maneuver, especially in patients with cirrhosis. From January 2016 to August 2016, 7 patients who underwent total laparoscopic left hepatectomy under hemihepatic vascular inflow occlusion using the extra-glissonian approach were included in this study. All were hepatitis B carriers and 4 had cirrhosis. The mean operation time was 247 minutes. The mean transection time was 110 minutes. No patient needed additional Pringle's maneuver. The mean intraoperative blood loss was 74 ml and no patient required blood transfusion. No open conversion happened. Postoperatively, no patient developed complications and there was no perioperative mortality. The mean resection margin was 2 cm. The mean hospital stay was 6 days. Upon a mean follow-up of 9 months, no patient developed tumor recurrence. The technique of total laparoscopic left hepatectomy using extra-glissonian approach was safe and feasible. The early surgical outcomes were good.


Detection of Pneumocystis carinii DNA by filtration of air.
The high incidence of pneumonia caused by Pneumocystis carinii in immunosuppressed patients makes it the most important parasite in non-tropical geographical regions. It has recently been shown to be a fungus, but several aspects of this organism are still poorly understood. A major question of clinical relevance is the nature of transmission and, thereby, the related problem of prevention. The mode of P. carinii transmission is thought to be air, but this is based on circumstantial evidence, the transmissive stage has not been identified. We attempted to capture P. carinii by filtration of air in the vicinity of cages containing P. carinii infected Wistar rats. Using nested polymerase chain reaction amplification of the thymidylate synthase gene to demonstrate P. carinii we were able to demonstrate P. carinii DNA on such filters. This strongly supports the suggested mechanism of transmission of Pneumocystis by means of airborne spores and suggests an approach to their isolation and characterization.


Association of the ADH2*2 allele with reduced ethanol consumption in Jewish men in Israel: a pilot study.
This study provides preliminary evidence on the associations between alcohol consumption patterns and polymorphisms of the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) enzymes in a Jewish population. Two groups of Jewish men were studied--one group (n = 92) representative of the free-living population of Jerusalem and generally light consumers of ethanol and the other group (n = 53) composed of treatment-enrolled heroin dependent individuals in the same city, most with a history of heavy daily drinking. All participants were interviewed regarding sociodemographic background, present and past alcohol consumption patterns, and familial characteristics including alcohol problems among first-degree relatives. Polymorphisms of the ADH2, ADH3 and ALDH loci were determined for all participants. The less common allele of the ADH2 locus (ADH2*2 allele frequency approximately 20% in Ashkenazic and non-Ashkenazic members of both groups) was related to a reduced mean level of peak weekly alcohol intake in the two groups. In multiple regression models adjusting for family history of alcohol problems and other factors, the ADH2*2 allele accounted for 20% and 30% of the explained alcohol intake variance in these two groups, respectively. Results from a logistic regression indicated that the ADH2*2 allele was also related to infrequent drinking in both groups. Evidence for an independent association between the ADH3 polymorphism and alcohol consumption patterns was not found. The ALDH gene was not polymorphic in this population. This report describes for the first time an association between alcohol consumption patterns and a polymorphism at the ADH2 locus in a Jewish population. The relatively high frequency of the ADH2*2 allele may contribute to the seemingly lower levels of alcohol consumption and heightened sensitivity to alcohol observed among Jews.


Polarization-insensitive wavelength conversion of 40 Gb/s NRZ-DPSK signals in a silicon polarization diversity circuit.
Polarization insensitive wavelength conversion of a 40 Gb/s non-return-to-zero (NRZ) differential phase-shift keying (DPSK) data signal is demonstrated using four-wave mixing (FWM) in a silicon nanowire circuit. Polarization independence is achieved using a diversity circuit based on polarization rotators and splitters, which is fabricated by a simple process on the silicon-on-insulator (SOI) platform. Error-free performance is achieved with only 0.5 dB of power penalty compared to the wavelength conversion of a signal with well optimized input polarization. Additionally, data transmission over 161 km standard single-mode fiber (SSMF) is demonstrated at 40 Gb/s using optical phase conjugation (OPC) in the proposed circuit.


Potential impact of recombination on sitewise approaches for detecting positive natural selection.
Current sitewise methods for detecting positive selection on gene sequences (the de facto standard being the CODEML method (Yang et al., 2000)) assume no recombination. This paper presents simulation results indicating that violation of this assumption can lead to false positive detection of sites undergoing positive selection. Through the use of population-scaled mutation and recombination rates, simulations can be performed that permit the generation of appropriate null distributions corresponding to neutral expectations in the presence of recombination, thereby allowing for a more accurate estimation of positive selection.


mGlu receptors and drug addiction.
Historically, brain catecholamine systems have been the primary focus of studies examining the neural substrates of drug addiction. In the past two decades, however, a wealth of evidence has accumulated indicating a pivotal role for glutamatergic neurotransmission in mediating addictive behaviors as well as long-term neuroplasticity associated with chronic drug use. As a result, there has been increased interest in developing glutamate-based therapies for the treatment of addictive disorders. Metabotropic glutamate (mGlu) receptors are classified into subcategories designated as Group I (mGlu1 and mGlu5), Group II (mGlu2 and mGlu3), and Group III (mGlu4, mGlu6, mGlu7, and mGlu8), and have received a great deal of attention due to their mediation of slower modulatory excitatory neurotransmission. Pharmacological ligands targeting these receptors have demonstrated reduced incidences of excitotoxicity or severe adverse side effects as compared to those targeting ionotropic glutamate (iGlu) receptors. Behavioral genetic and pharmacological studies have explored the role of individual mGlu receptor subtypes in regulating various addiction-related behaviours and several mGlu receptor ligands have been the subject of clinical testing for other medical conditions.


The Relationship Between Female Reproductive Functions and Vitamin D.
Nonclassical target organs recently defined for vitamin D, a major regulator of calcium phosphorus homeostasis and bone health, include reproductive ones. This compilation study focuses on the potential effects of vitamin D on female reproductive functions. Vitamin D receptor enzymes that metabolize vitamin D are expressed in both central and peripheral reproductive organs. Most studies suggest that vitamin D may be directly or indirectly related to gonadal functions. Vitamin D's effects on reproductive functions may be indirectly related to diseases such as polycystic ovary syndrome (PCOS), uterine leiomyomas, and endometriosis. In case of vitamin D deficiency during infertility treatment, vitamin D supplementation can be recommended especially for women who have PCOS, insulin resistance, or low anti-Mullerian hormone levels. Supplementation, however, should take into account possible toxic effects of high-dose vitamin D. To be able to recommend measuring vitamin D as a routine screening test and to better understand the effects of vitamin D and its supplementation on female reproductive functions, larger randomized controlled prospective studies are needed.


[H1- and H2-receptor sensitivity to 3-(beta-aminoethyl)-1,2,4-triazole derivatives].
Histamine-like derivatives of 3-(beta-aminoethyl)-1,2,4-trizole were synthetized and subjected to a pharmacological study. The sensitivity of H1- and H2-receptors to these substances was defined by the following tests, viz. in the isolated uterus of a rat, on the isolated atrium of a guinea pig and measurements of the blood pressure nn a cat. The derivative 3-(beta-aminoethyl)-5-amino-1,2,4-triazole (IEM-813) affects predominantly H1-receptors, while 3-(aminoethyl)-5-amino-1,2,4,triazole (IEM-759) has the closest affinity for H2-receptors, The authors presume that corresponding zones of histamine recpetors differ in their lyophilic and hydrophilic properties.


Pediatric astrocytomas with monomorphous pilomyxoid features and a less favorable outcome.
Among tumors classified as pilocytic astrocytoma (PA) in the Johns Hopkins Hospital Department of Pathology files, we identified 18 cases with a distinctive monomorphous pilomyxoid histological pattern and a higher recurrence rate than that of PA with classical histological features (classical PA). The majority of the tumors occurred in infants and young children and involved the hypothalamic/chiasmatic region. The tumors were histologically similar to PA, but they were more monomorphous and more myxoid. Rosenthal fibers were not seen and only 1 of 18 tumors had eosinophilic granular bodies. At the end of the follow-up period, 6 patients were dead and 12 were alive with evidence of disease. Progression free survival (PFS) at 1 year was 38.7%. In comparison, we identified a control group of 13 classical PAs in the same age range and location as the study group. In this group, PFS at 1 year was 69.2%, which was significantly better than that for pilomyxoid tumors (p = 0.04). There was no CSF dissemination or death due to tumor progression among patients with classical PA. Eight of these patients are alive with recurrent disease, and 4 have no evidence of disease. While the monomorphous pilomyxoid tumors have some resemblance to classical PA, our results suggest that the former is a more aggressive variant or a separate entity that needs to be recognized for prognostic purposes.


Structural rearrangement within an enveloped virus upon binding to the host cell.
We have made the surprising discovery that the interactions of herpes simplex virus with its initial cell attachment receptor induce a rapid and highly efficient structural change in the tegument, the region of the virion situated between the membrane and the capsid. It has been known for nearly a decade that viruses can trigger host signaling pathways when they bind to receptors on the cell surface; however, until now there has been no evidence that a signal can be sent in reverse--from the "outside in"--across a viral membrane. Evidence for this signaling event was found during studies of UL16, a tegument protein that is conserved among all the herpesviruses. Previous work has demonstrated that UL16 is bound to capsids isolated from the cytoplasm of infected cells, but this interaction is destabilized during subsequent egress steps, leading to release of the extracellular virion. Pretreatment with N-ethylmaleimide, a small, membrane-permeating compound that covalently modifies free cysteines, restabilizes the interaction, thereby permitting the capsid-UL16 complex to be isolated following disruption of virions with NP-40. In the experiments described here, we found that the natural signal for release of UL16 from capsids is sent when virions merely bind to cells at 4 degrees C. The internal change was also observed upon binding to immobilized heparin in a manner that requires viral glycoprotein C. This represents the first example of signaling across a viral envelope following receptor binding.


Therapy for oral squamous cell carcinoma by tegafur and streptococcal agent OK-432 in combination with radiotherapy: association of the therapeutic effect with differentiation and apoptosis in the cancer cells.
Twenty patients with oral squamous cell carcinoma having mainly stage II or III lesions without distant metastasis, were treated with tegafur and streptococcal agent, OK-432, in combination with radiotherapy. As a consequence, 16 cases among the treated 20 cases showed complete remission by this therapy alone. Especially, we have found that the squamous cell carcinoma arising in non-keratinizing oral epithelium rather than in keratinizing oral epithelium has better response to this therapy. Among the 16 cases with complete remission (CR) by the current therapy, 10 cases were histopathologically diagnosed as well-differentiated squamous cell carcinoma and six cases as moderately differentiated squamous cell carcinoma. When we examined immunohistochemically the expression of various antigens such as proliferating cell nuclear antigen (PCNA), p53 and LeY or the presence of DNA fragmentation by nick-end labelling in the biopsy materials taken at the first visit to our clinic from 20 patients treated with the current therapy, the CR group showed a significantly increased LeY expres-sion level ( p< 0.05) and DNA fragmentation rate (p< 0.05) as compared with the partial response (PR, n= 3) + no change (NC, n= 1) group. On the other hand, the CR group with respect to PCNA expression level was significantly decreased as compared with the PR + NC group ( p< 0.05). From these findings, it can be considered that the therapy for oral squamous cell carcinoma by UFT and OK-432 in combination with radiotherapy is very effective, which may be associated with differentiation or apoptosis in oral squamous carcinoma cells. In addition, we present the clinical findings and results of immunohistochemical staining for the biopsy materials obtained from four CR cases treated with the current therapeutic method.


Octopamine neuromodulation regulates Gr32a-linked aggression and courtship pathways in Drosophila males.
Chemosensory pheromonal information regulates aggression and reproduction in many species, but how pheromonal signals are transduced to reliably produce behavior is not well understood. Here we demonstrate that the pheromonal signals detected by Gr32a-expressing chemosensory neurons to enhance male aggression are filtered through octopamine (OA, invertebrate equivalent of norepinephrine) neurons. Using behavioral assays, we find males lacking both octopamine and Gr32a gustatory receptors exhibit parallel delays in the onset of aggression and reductions in aggression. Physiological and anatomical experiments identify Gr32a to octopamine neuron synaptic and functional connections in the suboesophageal ganglion. Refining the Gr32a-expressing population indicates that mouth Gr32a neurons promote male aggression and form synaptic contacts with OA neurons. By restricting the monoamine neuron target population, we show that three previously identified OA-Fru(M) neurons involved in behavioral choice are among the Gr32a-OA connections. Our findings demonstrate that octopaminergic neuromodulatory neurons function as early as a second-order step in this chemosensory-driven male social behavior pathway.


Sol-gel microencapsulation of odorants and flavors: opening the route to sustainable fragrances and aromas.
The sol-gel approach to encapsulate fragrance and aroma chemicals in porous silica-based materials has the potential to afford odorant and flavoured materials with novel and unique beneficial properties. Amorphous sol-gel SiO2 is non-toxic and safe, whereas the sol-gel entrapment of delicate chemicals in its inner pores results in pronounced chemical and physical stabilization of the entrapped actives, thereby broadening the practical utilization of chemically unstable essential oils. Reviewing progress in the fabrication of diverse odorant and flavoured sol-gels, we show how different synthetic strategies afford advanced materials suitable for practical application with important health and environmental benefits.


[Effect of allapinin after a single intravenous administration to patients with chronic ventricular extrasystole].
Clinical assessment of a new antiarrhythmic drug, allapinin, was carried out in 42 patients with chronic ventricular extrasystoles. Single intravenous doses were effective where they exceeded 0.32 mg/kg. Doses above 0.42 mg/kg were regularly associated with side effects, such as dizziness or, less commonly, diplopia. The 0.39 mg/kg dose produced an antiarrhythmic effect in 61% of patients, whose PQ interval was lengthened by 23%, and the QRS complex, by 18%, while QTc duration remained unchanged. Heart rate and arterial blood pressure were virtually unchanged, either. The peak of antiarrhythmic effect fell to the second postadministration hour, and the effect persisted within an average of 7.6 hours. Therefore, allapinin appears to be a potent quinidine-like agent.


[Enzyme immunoassay in diagnosis of opisthorchiasis. Communication 2. Enzyme immunoassay test system for diagnosis of acute opisthorchiasis].
A standard, highly specific and sensitive enzyme immunoassay test system has been developed for the diagnosis of acute opisthorchiasis, which is based on indirect solid-phase enzyme immunoassay for determination IgM antibodies to the antigen derived from Opisthorchis felineus marita extracts. The sensitivity of the test system is 87%, its specificity is 96.8%.


Identification of enzyme-substrate and enzyme-product complexes in the catalytic mechanism of glucoamylase from Aspergillus awamori.
Intermediates in the catalytic mechanism of Aspergillus awamori glucoamylase (GA) were identified by studying pre-steady-state and steady-state kinetics of the wild-type GA/maltose and Trp120 -->Phe GA/maltotriose reactions in H2O and D2O. Pre-steady-state fluorescence signal analysis was carried out to ascertain the relative intrinsic fluorescence of the enzyme intermediates. A three-step minimal pathway for oligosaccharide hydrolysis represented by E + Gx (k1) reversible (k-1) EGX (k2)reversible(k-2) EP (kcat)--> E + P is proposed. The first step, represented by the association constant K1 (k1/k-1), depicts the fast formation of enzyme-substrate complex and is the primary factor in fluorescence quenching. A 2.7-fold increase in K1 with D2O as solvent is observed with both enzymes due to the cumulative effect of deuterium on complex hydrogen bonding at the active site. The second step further quenches the enzyme fluorescence and is identified as the hydrolytic step, forming an enzyme-product complex. Both k2 and k-2 values show similar 2-fold decreases in D2O for both enzymes, consistent with the microscopic reversibility of the hydrolytic reaction. The solvent isotopic effect on the hydrolytic step is likely due to either abstraction of an exchangeable proton from the general acid Glu179 or directed addition of water to the oxocarbonium ion intermediate by the general base Glu400. No significant isotope effect was observed on the steady-state kcat value for wild-type GA with maltose, indicating a ronhydrolytic step as rate-limiting. The third step, a posthydrolytic rate-determining step, is the product release as evident from steady-state kinetics with wild-type and Trp120-->Phe GAs using alpha-D-glucosyl fluoride.


HIV-Infected Patient Knowledge, Attitudes, and Beliefs Regarding the Affordable Care Act.
We evaluated patient knowledge, attitudes, and beliefs regarding changes present with the Affordable Care Act (ACA). HIV-infected adults attending an academic medical center HIV clinic in Omaha, Nebraska were asked to complete a self-administered survey between November 2013 and March 2014. Information collected included demographics, knowledge regarding healthcare reform policies, as well as attitudes and beliefs regarding the potential impact of the ACA on patient access to healthcare. Basic descriptive statistics were used to assess demographic characteristics of respondents and outcomes of interest. Chi-square tests were used for comparisons of interest among participants; some trends were evaluated with Cochran-Armitage trend tests. Four hundred and six patients completed the questionnaire. Of the respondents 90% were between the ages of 27 and 64, 61% were white, 27% had no health insurance, and 21% reported that they felt they had or will eventually benefit from the ACA. The proportion who responded "I don't know" to this question decreased over the study period (p=0.036). Overall, 57% reported they do not believe that they are informed enough to make decisions about the ACA. In answering four knowledge-based questions, only 3% answered all of them correctly. Knowledge about the ACA was significantly associated with perception of benefit (p=0.018). HIV-infected patients are not well informed about the ACA and few perceive that they will benefit from healthcare reform. Targeted education and outreach are necessary to reduce the knowledge gap for this population that stands to benefit greatly from the ACA.


[Changes in target volumes definition by using MRI for prostate bed radiotherapy planning--preliminary results].
Magnetic resonance imaging (MRI) is used quite routinely in radiotherapy treatment planning in the primary radiotherapy of prostate cancer as it provides more contrast imaging of soft tissues in the small pelvis than planning CT, thanks to which it allows more exact delineation of target volumes and thus the saving of organs at risk We tried to verify whether it is possible to use MRI by analogy in the planning of prostate bed radiotherapy. Twentyone patients indicated for prostate bed radiotherapy were considered in this study. Here we present the preliminary results of 10 of them. Four patients were indicated for adjuvant, 6 for salvage radiotherapy. All the patients underwent, besides standard planning CT, MRI in the same position. Target volumes and organs at risk were delineated into CT,T1 and T2 MRI images - clinical target volume (CTV), planning target volume (PTV), urinary bladder and rectum. Based on the merging of images, the volumes delineated in MRI were copied into planning CT, where the evaluation was done. We evaluated the volumes of each structure, agreement in contouring with the help of the rate of union and intersection of the volumes and with Cohen's kappa, and 3D differences between volumes of CTV on CT, T1 and T2 MRI. Statistically, volumes of CTV and PTV are not significantly different. The volume of the rectum is significantly smaller on T1 and also T2 MRI images. The index of agreement (union/intersection) is statistically significantly different from 1 for CTV and PTV as well. Cohen's kappa indicates moderate agreement for CTV CT and T1, T1 and T2 MRI, fair agreement for CTV CT and T2 MRI, and substantial agreement for PTV. In the superior and superolateral direction, the CTV volume on MRI in the central plane is smaller on T1 and T2 images. In the area of seminal vesicles (SV) the cranial border is similar on CT and MRI. In the superoposterior direction, the volume of CTV is smaller on CT than on T1 and T2 MRI, which means, that seminal vesicles are delineated larger in the posterior direction on MRI (about 0.24cm on T1; by about 0.20cm on T2 images). In the posterior direction, there are no differences in CTV on CT and T1 while on T2 the CTV is larger (a difference of 0.29 cm). In the posterolateral direction, CTV is smaller on T1 MRI than on CT on both sides, on the right as well as on the left. Preliminary results suggest that clinical target volume defined with the help of MRI is shifted compared with CTV defined on planning CT. The agreement of CTV delineation by one radiation oncologist is moderate to fair and is similar to interobserver variability in the contouring of the prostate bed in the planning CT. MRI provides more contrast imaging of the anterior rectal wall, where we have confirmed the most differences in contouring. Moreover, it provides better imaging of local recurrences and seminal vesicles, where the most differences in our group of patients were seen in comparison with planning CT.


Comparability of a single-trial passive avoidance learning task in the young chick across different laboratories.
The discrepant results noted by the La Trobe/Monash, Sussex, Open, and Berkeley University memory research groups employing the passive avoidance task (PAT) with the chick indicate that some of these differences may be due to differences in training procedures between the various groups. The procedures employed by each group were replicated as closely as possible and compared using the same strain of chick. Higher levels of pecking and lower training latencies were observed in the La Trobe/Monash chicks. Improved training latency was observed following a change in the day of experimentation with Sussex chicks, and in white light in Open chicks. A powerful reinforcing effect was observed when pretraining was conducted with a wet lure across multiple pretraining trials, indicating the importance of this difference between groups. The observed impact on chick performance of these variables indicates that differences between PAT procedures may have important interactive effects with aversive learning. The results of data gathered using different procedures may only be superficially comparable, and further investigation of the effects of these variables is indicated.


The Tyrosine Sulfate Domain of Fibromodulin Binds Collagen and Enhances Fibril Formation.
Small leucine-rich proteoglycans interact with other extracellular matrix proteins and are important regulators of matrix assembly. Fibromodulin has a key role in connective tissues, binding collagen through two identified binding sites in its leucine-rich repeat domain and regulating collagen fibril formation in vitro and in vivo Some nine tyrosine residues in the fibromodulin N-terminal domain are O-sulfated, a posttranslational modification often involved in protein interactions. The N-terminal domain mimics heparin, binding proteins with clustered basic amino acid residues. Because heparin affects collagen fibril formation, we investigated whether tyrosine sulfate is involved in fibromodulin interactions with collagen. Using full-length fibromodulin and its N-terminal tyrosine-sulfated domain purified from tissue, as well as recombinant fibromodulin fragments, we found that the N-terminal domain binds collagen. The tyrosine-sulfated domain and the leucine-rich repeat domain both bound to three specific sites along the collagen type I molecule, at the N terminus and at 100 and 220 nm from the N terminus. The N-terminal domain shortened the collagen fibril formation lag phase and tyrosine sulfation was required for this effect. The isolated leucine-rich repeat domain inhibited the fibril formation rate, and full-length fibromodulin showed a combination of these effects. The fibrils formed in the presence of fibromodulin or its fragments showed more organized structure. Fibromodulin and its tyrosine sulfate domain remained bound on the formed fiber. Taken together, this suggests a novel, regulatory function for tyrosine sulfation in collagen interaction and control of fibril formation.


Description of Clavibacter zhangzhiyongii sp. nov., a phytopathogenic actinobacterium isolated from barley seeds, causing leaf brown spot and decline.
Clavibacter michiganensis is a Gram-stain-positive bacterium with eight subspecies, five of which have been redefined as different species on the basis of their genome sequence data. On the basis of the results of phylogenetic analysis of dnaA gene sequences, strains of members of the genus Clavibacter isolated from barley have been grouped in a separate clade from other species and subspecies of the genus Clavibacter. In this study, the biochemical, physiological, fatty acids and genetic characteristics of strains DM1T and DM3, which represented the barley isolates, were examined. On the basis of results from multi-locus sequence typing and other biochemical and physiological features, including colony colour, carbon source utilisation and enzyme activities, DM1T and DM3 are categorically differentiated from the aforementioned eight species and subspecies of the genus Clavibacter. Moreover, the results of genomic analysis reveal that the DNA G+C contents of DM1T and DM3 are 73.7 and 73.5 %, respectively, and the average nucleotide identity (ANI) values between DM1T and DM3 and other species and subspecies range from 90.4 to 92.0 %. The ANI value between DM1T and DM3 is 98.0 %. These results indicate that DM1T and DM3 are distinct from other known species and subspecies of the genus Clavibacter. Therefore, we propose a novel species, C. zhangzhiyongii, with DM1T (=CFCC 16553 T=LMG 31970T) as the type strain.


Physiological responses of the CAM epiphyte Tillandsia usneoides L. (Bromeliaceae) to variations in light and water supply.
In an effort to understand the mechanisms that sustain rootless atmospheric plants, the modulation of Crassulacean acid metabolism (CAM) in response to variations in irradiance and water supply was investigated in the epiphyte Tillandsia usneoides. Plants were acclimated to three light regimes, i.e. high, intermediate and low, with integrated photon flux densities (PFD) of 14.40, 8.64 and 4.32 mol m-2 d-1 equivalent to an instantaneous PFD of 200, 100, and 50 mumol m-2 s-1, respectively. Daily watering was then withdrawn from half of the plants at each PFD for 7 d prior to sampling. In response to the three PFD treatments, chlorophyll content increased in plants acclimated to lower irradiances. Light response curves using non-invasive measurements of chlorophyll fluorescence demonstrated that photosystem II efficiency (phi PSII) was maintained in high PFD acclimated plants, as they exhibited a larger capacity for non-photochemical dissipation (NPQ) of excess light energy than low PFD acclimated plants. Net CO2 uptake increased in response to higher PFD, reflecting enhanced carboxylation capacity in terms of phosphoenolpyruvate carboxylase (PEPc) and ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) activities. After water was withdrawn, nocturnal net CO2 uptake and accumulated levels of acidity declined in all PFD treatments, concomitant with increased respiratory recycling of malate. Examining the strategies employed by epiphytes such as T. usneodies to tolerate extreme light and water regimes has demonstrated the importance of physiological mechanisms that allow flexible carboxylation capacity and continued carbon cycling to maintain photosynthetic integrity.


Therapeutic modulation of lipoprotein-associated phospholipase A2 (Lp-PLA2).
Lipoprotein-associated phospholipase A2 (Lp-PLA2) is a calcium-independent phospholipase A2 that circulates in plasma in association with lipoprotein particles, whereas in atherosclerotic plaques it is co-localized with macrophages. Lp-PLA2 generates two proinflammatory mediators, lysophosphatidylcholine and oxidized nonesterified fatty acids, which play a role in the development of atherosclerotic lesions and formation of a necrotic core, leading to more vulnerable plaques. Epidemiologic studies demonstrate that increased circulating levels of Lp-PLA2 predict an increased risk of myocardial infarction, stroke and cardiovascular mortality. Furthermore, histologic examination of diseased human coronary arteries reveals intense presence of the enzyme in atherosclerotic plaques that are prone to rupture. These considerations suggest Lp-PLA2 as a promising therapeutic target in cardiovascular disease. Plasma levels of Lp-PLA2 are increased in various types of hyperlipidemias, while hypolipidemic drugs reduce plasma Lp-PLA2 activity and mass along with the improvement of plasma lipid profile. A selective inhibitor of Lp-PLA2 activity, darapladib, has been developed and studies in animal models and humans have shown that it effectively and safely reduces Lp-PLA2 activity in plasma and in atherosclerotic plaques. Furthermore, in animal models darapladib decreases plaque area and necrotic core area whereas in humans it prevents the expansion of necrotic core volume. Whether the results obtained from the use of darapladib in studies in vitro, as well as in preclinical and clinical studies would translate into benefits on cardiovascular event outcomes, awaits to be proved in 2 ongoing phase 3 trials.


The micro-ribonucleic acid (miRNA) miR-206 targets the human estrogen receptor-alpha (ERalpha) and represses ERalpha messenger RNA and protein expression in breast cancer cell lines.
Micro-RNAs are small noncoding RNAs, which diminish the stability and/or translation of mRNAs. This study examined whether miR-206, previously shown to be elevated in estrogen receptor (ER)alpha-negative breast cancer, regulates the expression of ERalpha. Two putative miR-206 sites, (hERalpha1 and hERalpha2), were found in silico within the 3'-untranslated region of human ERalpha mRNA. Transfection of MCF-7 cells with pre-miR-206 or 2'-O-methyl antagomiR-206 specifically decreased or increased, respectively, ERalpha mRNA levels. Overexpression of pre-miR-206 reduced ERalpha and beta-actin protein levels, with no effect on ERbeta, E-cadherin, or glyceraldehyde-3-phosphate dehydrogenase. Reporter constructs containing the hERalpha1 or hERalpha2 binding sites inserted into the 3'-untranslated region of the luciferase mRNA conferred a 1.6- and 2.2-fold repression of luciferase activity, respectively, in HeLa cells. Both miR-206 sites responded accordingly to exogenous hsa-pre-miR-206 and 2'-O-methyl antagomiR-206, and both sites were rendered inactive by mutations that disrupted hybridization to the 5'-seed of miR-206. A C-->T single nucleotide polymorphism in the hERalpha1 site increased repression of luciferase activity to approximately 3.3-fold in HeLa cells. MiR-206 levels were higher in ERalpha-negative MB-MDA-231 cells than ERalpha-positive MCF-7 cells, but only the ERalpha1 site mediated significantly more repression in reporter constructs. MiR-206 expression was strongly inhibited by ERalpha agonists, but not by an ERbeta agonist or progesterone, indicating a mutually inhibitory feedback loop. These findings provide the first evidence for the posttranscriptional regulation of ERalpha by a micro-RNA in the context of breast cancer.


Downregulation of LSD1 suppresses the proliferation, tumorigenicity and invasion of papillary thyroid carcinoma K1 cells.
The present study aimed to evaluate the effects of lysine-specific demethylase 1 (LSD1) downregulation, induced by small interfering RNA (siRNA) transfection, on the proliferation, colony formation, migration and invasion of the papillary thyroid carcinoma K1 cell line. The siRNA targeting LSD1 and scrambled non-targeting siRNA were each transfected into papillary thyroid carcinoma K1 cells. Downregulation of LSD1 mRNA and protein level was evaluated by reverse transcription-quantitative polymerase chain reaction, and immunocytochemical (ICC) analysis and western blotting, respectively. A Cell Counting kit-8 assay was applied to estimate the effect of LSD1-siRNA on cell growth. Migration and invasion abilities were estimated by Transwell chamber assay. A soft agar colony formation assay was performed to estimate the effect of LSD1-siRNA on tumorigenicity in vitro. ICC data showed that LSD1 protein was strongly expressed in the blank and control K1 cells compared with the LSD1-siRNA cells (F=15.192, P<0.01). Compared with the control cells, cells transfected with siRNA targeting LSD1 exhibited significant downregulation of LSD1 mRNA (t=6.845, P<0.01) and protein (F=53.764, P<0.01) levels. siRNA targeting LSD1 also downregulated cell proliferation following transfection for 24, 48 and 72 h (t=4.777, P<0.001; t=3.302, P=0.003; and t=3.017, P=0.006, respectively). Compared with the control group, the amount of cell invasion was gradually reduced in the LSD1-siRNA group (t=12.301, P<0.01). The number of migrating cells was significantly higher in the negative control group compared with the LSD1-siRNA group (t=7.911, P<0.01), and the ability of colony formation in the LSD1-siRNA cells was notably reduced in the soft agar formation assay (t=3.612, P=0.005). siRNA targeting LSD1 efficiently inhibits the proliferation, colony formation, migration and invasion of papillary thyroid carcinoma K1 cells.


A systematic review of cytokines in chronic fatigue syndrome/myalgic encephalomyelitis/systemic exertion intolerance disease (CFS/ME/SEID).
Cytokines in Chronic Fatigue Syndrome/Myalgic Encephalomyelitis/Systemic Exertion Intolerance Disease (CFS/ME/SEID) patients compared with healthy controls have been extensively studied. However, the evidence regarding whether a baseline difference between CFS/ME/SEID patients and the normal population remains unclear. The aim of this study was to conduct a systematic review of the literature regarding cytokines in CFS/ME/SEID and whether there is a significant difference in cytokine levels between this patient group and the normal population. Pubmed, Scopus, Medline (EBSCOHost), and EMBASE databases were searched to source relevant studies for CFS/ME/SEID. The review included any studies examining cytokines in CFS/ME/SEID patients compared with healthy controls. Results of the literature search were summarised according to aspects of their study design and outcome measures, namely, cytokines. Quality assessment was also completed to summarise the level of evidence available. A total of 16,702 publications were returned using our search terms. After screening of papers according to our inclusion and exclusion criteria, 15 studies were included in the review. All the included studies were observational case control studies. Ten of the studies identified measured serum cytokines in CFS/ME/SEID patients, and four measured cytokines in other physiological fluids of CFS/ME/SEID patients. The overall quality assessment revealed most papers included in this systematic review to be consistent. Despite the availability of moderate quality studies, the findings of this review are inconclusive as to whether cytokines play any definitive role in CFS/ME/SEID, and consequently, they would not serve as reliable biomarkers. Therefore, in light of these results, it is recommended that further efforts toward a diagnostic test and treatment for CFS/ME/SEID continue to be developed in a range of research fields.


Role of ipsdienol, ipsenol, and cis-verbenol in chemical ecology of Ips avulsus, Ips calligraphus, and Ips grandicollis (Coleoptera: Curculionidae: Scolytinae).
Stressed or damaged pine (Pinus sp.) trees in the southeastern United States are often colonized simultaneously by three southern Ips species (Coleoptera: Curculionidae: Scolytinae): small southern pine engraver, Ips avulsus (Eichhoff); sixspined ips, Ips calligraphus (Germar); and eastern fivespined ips, Ips grandicollis (Eichhoff). All three species mediate colonization of host material with volatile pheromones. All of the southern Ips produce cis-verbenol, and either ipsdienol or ipsenol, and electrophysiological studies have demonstrated that all three southern Ips are able to detect all three compounds. This study examined the role of ipsdienol, ipsenol, and cis-verbenol in the chemical ecology of the southern Ips in Georgia and Louisiana. The most attractive blends of pheromones, with the fewest number of components, were ipsdienol plus ipsenol for I. avulsus, cis-verbenol plus ipsdienol for I. calligraphus, and either cis-verbenol plus ipsenol or ipsdienol plus ipsenol for I. grandicollis. Cross-attraction of I. grandicollis to the pheromone blend most attractive to I. avulsus was observed. Although the presence of heterospecific pheromone reduced the catches of all three species (i.e., the tertiary blend captured fewer beetles than the most attractive binary blends) in both states (significantly in two cases), high numbers of all three species were still captured in traps baited with all three compounds. These results suggest that the pheromones cis-verbenol, ipsdienol, and ipsenol can be combined for monitoring all three species of the southern Ips simultaneously.


Closure of VDAC causes oxidative stress and accelerates the Ca(2+)-induced mitochondrial permeability transition in rat liver mitochondria.
The electron transport chain of mitochondria is a major source of reactive oxygen species (ROS), which play a critical role in augmenting the Ca(2+)-induced mitochondrial permeability transition (MPT). Mitochondrial release of superoxide anions (O(2)(-)) from the intermembrane space (IMS) to the cytosol is mediated by voltage dependent anion channels (VDAC) in the outer membrane. Here, we examined whether closure of VDAC increases intramitochondrial oxidative stress by blocking efflux of O(2)(-) from the IMS and sensitizing to the Ca(2+)-induced MPT. Treatment of isolated rat liver mitochondria with 5microM G3139, an 18-mer phosphorothioate blocker of VDAC, accelerated onset of the MPT by 6.8+/-1.4min within a range of 100-250microM Ca(2+). G3139-mediated acceleration of the MPT was reversed by 20microM butylated hydroxytoluene, a water soluble antioxidant. Pre-treatment of mitochondria with G3139 also increased accumulation of O(2)(-) in mitochondria, as monitored by dihydroethidium fluorescence, and permeabilization of the mitochondrial outer membrane with digitonin reversed the effect of G3139 on O(2)(-) accumulation. Mathematical modeling of generation and turnover of O(2)(-) within the IMS indicated that closure of VDAC produces a 1.55-fold increase in the steady-state level of mitochondrial O(2)(-). In conclusion, closure of VDAC appears to impede the efflux of superoxide anions from the IMS, resulting in an increased steady-state level of O(2)(-), which causes an internal oxidative stress and sensitizes mitochondria toward the Ca(2+)-induced MPT.


Effects of in vitro repaglinide supplementation on improving sperm motility parameters, viability, and DNA integrity in normozoospermic and asthenozoospermic men.
Human sperm motility and hyperactivation (HA) are induced by different factors such as intracellular calcium concentration. Repaglinide is an antidiabetic drug that, via the blocking of ATP-sensitive potassium channels (K-ATP channels), depolarization of the β-cell membrane, and opening of the voltage-gated calcium channels leads to an increase in intracellular calcium. The present study aimed to examine the effects of repaglinide on in vitro sperm motility parameters, viability, and DNA integrity in normozoospermic and asthenozoospermic men. Semen samples were collected from two groups of normozoospermic donors and asthenozoospermic patients. The samples were washed free of seminal plasma and then treated with medium alone (control) or with 100 nM and 1µM concentrations of repaglinide. After 1 h of incubation, percent sperm motility and hyperactivation were assessed; after 2 h of incubation, sperm viability and DNA fragmentation rate were evaluated by the Eosin-Y and acridine orange staining, respectively. In both groups, repaglinide at a concentration of 100 nM and 1µM significantly improved percent sperm motility, hyperactivation, and vital sperms with normal DNA; in specimens from normozoospermic men, the 1µM concentration had a noticeable effect on progressive motility; in samples from asthenozoospermic men, the highest hyperactivation rate was seen at a concentration of 100 nM as compared with the 1µM concentration and controls (p<0.05). Our results suggest that repaglinide can improve sperm motility, hyperactivity, viability, and DNA integrity in both normozoospermic and asthenozoospermic men.


Effectiveness and success factors of educational inhaler technique interventions in asthma & COPD patients: a systematic review.
With the current wealth of new inhalers available and insurance policy driven inhaler switching, the need for insights in optimal education on inhaler use is more evident than ever. We aimed to systematically review educational inhalation technique interventions, to assess their overall effectiveness, and identify main drivers of success. Medline, Embase and CINAHL databases were searched for randomised controlled trials on educational inhalation technique interventions. Inclusion eligibility, quality appraisal (Cochrane's risk of bias tool) and data extraction were performed by two independent reviewers. Regression analyses were performed to identify characteristics contributing to inhaler technique improvement. Thirty-seven of the 39 interventions included (95%) indicated statistically significant improvement of inhaler technique. However, average follow-up time was relatively short (5 months), 28% lacked clinical relevant endpoints and all lacked cost-effectiveness estimates. Poor initial technique, number of inhalation procedure steps, setting (outpatient clinics performing best), and time elapsed since intervention (all, p < 0.05), were shown to have an impact on effectiveness of the intervention, explaining up to 91% of the effectiveness variation. Other factors, such as disease (asthma vs. chronic obstructive pulmonary disease), education group size (individual vs. group training) and inhaler type (dry powder inhalers vs. pressurised metered dose inhalers) did not play a significant role. Notably, there was a trend (p = 0.06) towards interventions in adults being more effective than those in children and the intervention effect seemed to wane over time. In conclusion, educational interventions to improve inhaler technique are effective on the short-term. Periodical intervention reinforcement and longer follow-up studies, including clinical relevant endpoints and cost-effectiveness, are recommended.


Microorganisms Associated with the Marine Sponge Scopalina hapalia: A Reservoir of Bioactive Molecules to Slow Down the Aging Process.
Aging research aims at developing therapies that delay normal aging processes and some related pathologies. Recently, many compounds and extracts from natural products have been shown to slow aging and/or extend lifespan. Marine sponges and their associated microorganisms have been found to produce a wide variety of bioactive secondary metabolites; however, those from the Southwest of the Indian Ocean are much less studied, especially regarding anti-aging activities. In this study, the microbial diversity of the marine sponge Scopalina hapalia was investigated by metagenomic analysis. Twenty-six bacterial and two archaeal phyla were recovered from the sponge, of which the Proteobacteria phylum was the most abundant. In addition, 30 isolates from S. hapalia were selected and cultivated for identification and secondary metabolites production. The selected isolates were affiliated to the genera Bacillus, Micromonospora, Rhodoccocus, Salinispora, Aspergillus, Chaetomium, Nigrospora and unidentified genera related to the family Thermoactinomycetaceae. Crude extracts from selected microbial cultures were found to be active against seven clinically relevant targets (elastase, tyrosinase, catalase, sirtuin 1, Cyclin-dependent kinase 7 (CDK7), Fyn kinase and proteasome). These results highlight the potential of microorganisms associated with a marine sponge from Mayotte to produce anti-aging compounds. Future work will focus on the isolation and the characterization of bioactive compounds.


Protective effects of Lactobacillus plantarum CCFM8610 against chronic cadmium toxicity in mice indicate routes of protection besides intestinal sequestration.
Our previous study confirmed the ability of Lactobacillus plantarum CCFM8610 to protect against acute cadmium (Cd) toxicity in mice. This study was designed to evaluate the protective effects of CCFM8610 against chronic Cd toxicity in mice and to gain insights into the protection mode of this strain. Experimental mice were divided into two groups and exposed to Cd for 8 weeks via drinking water or intraperitoneal injection. Both groups were further divided into four subgroups, control, Cd only, CCFM8610 only, and Cd plus CCFM8610. Levels of Cd were measured in the feces, liver, and kidneys, and alterations of several biomarkers of Cd toxicity were noted. The results showed that when Cd was introduced orally, cotreatment with Cd and CCFM8610 effectively decreased intestinal Cd absorption, reduced Cd accumulation in tissue, alleviated tissue oxidative stress, reversed hepatic and renal damage, and ameliorated the corresponding histopathological changes. When Cd was introduced intraperitoneally, administration of CCFM8610 did not have an impact on tissue Cd accumulation or reverse the activities of antioxidant enzymes. However, CCFM8610 still offered protection against oxidative stress and reversed the alterations of Cd toxicity biomarkers and tissue histopathology. These results suggest that CCFM8610 is effective against chronic cadmium toxicity in mice. Besides intestinal Cd sequestration, CCFM8610 treatment offers direct protection against Cd-induced oxidative stress. We also provide evidence that the latter is unlikely to be mediated via protection against Cd-induced alteration of antioxidant enzyme activities.


Phenolic glycolipid-I of Mycobacterium leprae induces general suppression of in vitro concanavalin A responses unrelated to leprosy type.
Using a costimulant assay, in vitro Con A responses of patients across the leprosy spectrum were found to be markedly suppressed by phenolic glycolipid-I (PGL-I), a unique antigen of M. leprae. The degree of inducible suppression as well as the number of leprosy patients showing suppression of mitogenic responses was higher with PGL-I as compared with integral M. leprae (p less than 0.05 to less than 0.01). Both untreated lepromatous (60%) as well as tuberculoid leprosy (67%) patients showed significant suppression ranging from 13 to 64% and 12 to 79%, respectively. Thus, PGL-I appears to have a universal suppressive effect on Con A responses and is unlikely to play a central role in determining the leprosy spectrum.


Effect of fasting on the intracellular metabolic partition of intravenously infused glucose in humans.
The effects of fasting on the pathways of insulin-stimulated glucose disposal were explored in three groups of seven normal subjects. Group 1 was submitted to a euglycemic hyperinsulinemic clamp ( approximately 100 microU/ml) after both a 12-h and a 4-day fast. The combined use of [3-(3)H]- and [U-(14)C]glucose allowed us to demonstrate that fasting inhibits, by approximately 50%, glucose disposal, glycolysis, glucose oxidation, and glycogen synthesis via the direct pathway. In group 2, in which the clamp glucose disposal during fasting was restored by hyperglycemia (155 +/- 15 mg/dl), fasting stimulated glycogen synthesis (+29 +/- 2%) and inhibited glycolysis (-32 +/- 3%) but only in its oxidative component (-40 +/- 3%). Results were similar in group 3 in which the clamp glucose disposal was restored by a pharmacological elevation of insulin ( approximately 2,800 microU/ml), but in this case, both glycogen synthesis and nonoxidative glycolysis participated in the rise in nonoxidative glucose disposal. In all groups, the reduction in total carbohydrate oxidation (indirect calorimetry) induced by fasting markedly exceeded the reduction in circulating glucose oxidation, suggesting that fasting also inhibits intracellular glycogen oxidation. Thus prior fasting favors glycogen retention by three mechanisms: 1) stimulation of glycogen synthesis via the direct pathway; 2) preferential inhibition of oxidative rather than nonoxidative glycolysis, thus allowing carbon conservation for glycogen synthesis via the indirect pathway; and 3) suppression of intracellular glycogen oxidation.


Mimicking Intermolecular Interactions of Tight Protein-Protein Complexes for Small-Molecule Antagonists.
Tight protein-protein interactions (Kd <100 nm) that occur over a large binding interface (>1000 Å2 ) are highly challenging to disrupt with small molecules. Historically, the design of small molecules to inhibit protein-protein interactions has focused on mimicking the position of interface protein ligand side chains. Here, we explore mimicry of the pairwise intermolecular interactions of the native protein ligand with residues of the protein receptor to enrich commercial libraries for small-molecule inhibitors of tight protein-protein interactions. We use the high-affinity interaction (Kd =1 nm) between the urokinase receptor (uPAR) and its ligand urokinase (uPA) to test our methods. We introduce three methods for rank-ordering small molecules docked to uPAR: 1) a new fingerprint approach that represents uPA's pairwise interaction energies with uPAR residues; 2) a pharmacophore approach to identify small molecules that mimic the position of uPA interface residues; and 3) a combined fingerprint and pharmacophore approach. Our work led to small molecules with novel chemotypes that inhibited a tight uPAR⋅uPA protein-protein interaction with single-digit micromolar IC50 values. We also report the extensive work that identified several of the hits as either lacking stability, thiol reactive, or redox active. This work suggests that mimicking the binding profile of the native ligand and the position of interface residues can be an effective strategy to enrich commercial libraries for small-molecule inhibitors of tight protein-protein interactions.


Validation of a Deep Learning Tool in the Detection of Intracranial Hemorrhage and Large Vessel Occlusion.
Purpose: Recently developed machine-learning algorithms have demonstrated strong performance in the detection of intracranial hemorrhage (ICH) and large vessel occlusion (LVO). However, their generalizability is often limited by geographic bias of studies. The aim of this study was to validate a commercially available deep learning-based tool in the detection of both ICH and LVO across multiple hospital sites and vendors throughout the U.S. Materials and Methods: This was a retrospective and multicenter study using anonymized data from two institutions. Eight hundred fourteen non-contrast CT cases and 378 CT angiography cases were analyzed to evaluate ICH and LVO, respectively. The tool's ability to detect and quantify ICH, LVO, and their various subtypes was assessed among multiple CT vendors and hospitals across the United States. Ground truth was based off imaging interpretations from two board-certified neuroradiologists. Results: There were 255 positive and 559 negative ICH cases. Accuracy was 95.6%, sensitivity was 91.4%, and specificity was 97.5% for the ICH tool. ICH was further stratified into the following subtypes: intraparenchymal, intraventricular, epidural/subdural, and subarachnoid with true positive rates of 92.9, 100, 94.3, and 89.9%, respectively. ICH true positive rates by volume [small (<5 mL), medium (5-25 mL), and large (>25 mL)] were 71.8, 100, and 100%, respectively. There were 156 positive and 222 negative LVO cases. The LVO tool demonstrated an accuracy of 98.1%, sensitivity of 98.1%, and specificity of 98.2%. A subset of 55 randomly selected cases were also assessed for LVO detection at various sites, including the distal internal carotid artery, middle cerebral artery M1 segment, proximal middle cerebral artery M2 segment, and distal middle cerebral artery M2 segment with an accuracy of 97.0%, sensitivity of 94.3%, and specificity of 97.4%. Conclusion: Deep learning tools can be effective in the detection of both ICH and LVO across a wide variety of hospital systems. While some limitations were identified, specifically in the detection of small ICH and distal M2 occlusion, this study highlights a deep learning tool that can assist radiologists in the detection of emergent findings in a variety of practice settings.


Body image, visual working memory and visual mental imagery.
Body dissatisfaction (BD) is a highly prevalent feature amongst females in society, with the majority of individuals regarding themselves to be overweight compared to their personal ideal, and very few self-describing as underweight. To date, explanations of this dramatic pattern have centred on extrinsic social and media factors, or intrinsic factors connected to individuals' knowledge and belief structures regarding eating and body shape, with little research examining links between BD and basic cognitive mechanisms. This paper reports a correlational study in which visual and executive cognitive processes that could potentially impact on BD were assessed. Visual memory span and self-rated visual imagery were found to be predictive of BD, alongside a measure of inhibition derived from the Stroop task. In contrast, spatial memory and global precedence were not related to BD. Results are interpreted with reference to the influential multi-component model of working memory.