NTP 3-month toxicity studies of estragole (CAS No. 140-67-0) administered by gavage to F344/N rats and B6C3F1 mice.
Estragole is a natural organic compound that is used as an additive, flavoring agent, or fragrance in a variety of food, cleaning, and cosmetic products; as an herbal medicine; as an antimicrobial agent against acid-tolerant food microflora; and to produce synthetic anise oil. Estragole was nominated for toxicity testing by the National Institute of Environmental Health Sciences to characterize its toxicity when administered by gavage to F344/N rats and B6C3F1 mice and to determine how similar its effects might be to those of the structurally related compound, methyleugenol. Male and female F344/N rats and B6C3F1 mice were given estragole (greater than 99% pure) in corn oil by gavage for 3 months. Genetic toxicology studies were conducted in Salmonella typhimurium and mouse peripheral blood erythrocytes. Core and special study (rats only) groups of 10 male and 10 female rats and mice were administered 37.5, 75, 150, 300, or 600 mg estragole/kg body weight in corn oil by gavage, 5 days per week. The core study groups were given estragole for 3 months and the special study groups for 30 days. All core study rats survived the 3-month exposure period. Mean body weights of the 300 and 600 mg/kg groups were 73% to 92%, respectively, of those of the vehicle control groups. A staining pattern on the ventral surface anterior to the genitalia beginning at week 9 in the 300 and 600 mg/kg groups was attributed to residue of estragole or metabolites in the urine. Alterations in the erythron related to estragole administration occurred in male and female rats; male rats demonstrated a stronger response. The changes in the erythron were characterized as a microcytic, normochromic, nonresponsive anemia. There were decreases in serum iron concentration in the 300 mg/kg females and 600 mg/kg males and females. The average percent saturation of total iron binding capacity was decreased in the 600 mg/kg males and females. Dose-related increases in platelet counts occurred in most of the dosed groups of rats; the effect appeared to be stronger in males. The increase could be consistent with a reactive thrombocytosis. Increases in the serum alanine aminotransferase and sorbitol dehydrogenase activities suggested a hepatocellular effect (increased leakage) and were consistent with the morphological liver changes observed. There were dose-related increases in serum bile salt concentration in most treated male rats at all time points; females were less affected. Absolute and relative liver weights were significantly increased in 300 and 600 mg/kg males and in 75 mg/kg or greater females. Relative kidney weights were significantly increased in all dosed groups of male rats and in female rats given 75 mg/kg or greater. Absolute and relative testis weights of 300 and 600 mg/kg males were significantly decreased. Two 600 mg/kg male rats had multiple cholangiocarcinomas in the liver and a third had an hepatocellular adenoma. All 600 mg/kg males exhibited cholangiofibrosis. All 75 mg/kg or greater males and all 150 mg/kg or greater females had hepatocellular hypertrophy. Incidences of bile duct hyperplasia, oval cell hyperplasia, and chronic periportal inflammation were significantly increased in all dosed groups. Incidences of basophilic and mixed cell foci were significantly increased in 150 mg/kg or greater males and females. Incidences of eosinophilic focus were significantly increased in 300 and 600 mg/kg males and 600 mg/kg females. Incidences of cellular infiltration of the periportal region by histiocytes increased significantly in all dosed groups of males and in 150 mg/kg or greater females. Incidences of bone marrow hyperplasia were significantly increased in 75, 300, and 600 mg/kg male rats. Incidences of renal tubule papillary mineralization were significantly increased in 300 mg/kg males and females and 600 mg/kg males. Incidences of cortical renal tubule pigmentation were significantly increased in 150 mg/kg or greater males, and the incidence of renal tubule regeneration was significantly increased in 600 mg/kg females. Incidences of degeneration of the olfactory epithelium in the nose were significantly increased in 300 and 600 mg/kg rats. Incidences of hypertrophied chromophobe cells in the pars distalis of the pituitary gland were significantly increased in 300 and 600 mg/kg males. Cytoplasmic alteration of the submandibular salivary gland occurred in all 75 mg/kg or greater rats. Incidences of atrophy of the gastric glands in the stomach were significantly increased in 150 mg/kg or greater rats. Bilateral degeneration of the germinal epithelium in the testes and bilateral hypospermia of the epididymis occurred in all 300 and 600 mg/kg males. In the special study, serum gastrin concentration and stomach pH were significantly increased in rats exposed to 600 mg/kg for 30 days. Gastric gland atrophy was significantly increased in the stomach of 300 and 600 mg/kg rats. Hepatic 7-pentoxyresorufin-O-deethylase activity was significantly increased in all exposed groups except 37.5 mg/kg females, and the increases were generally dose related. In the mouse core study, a 600 mg/kg male died during week 9, and all 600 mg/kg female mice died during week 1; the female deaths were attributed to liver necrosis caused by estragole exposure. Mean body weights of 300 and 600 mg/kg males and 75 mg/kg or greater females were 79% to 89% those of the vehicle control groups. Liver weights were generally increased in 75 mg/kg or greater males and in 300 mg/kg females. Relative thymus weights were significantly increased in all dosed groups of female mice. The incidences of hepatocellular hypertrophy and hepatocellular degeneration were significantly increased in 300 and 600 mg/kg male mice and 150 and 300 mg/kg female mice. Incidences of oval cell hyperplasia were significantly increased in 300 and 600 mg/kg males and in 75 mg/kg or greater females. Liver necrosis occurred in all 600 mg/kg female mice, along with a significant increase in the incidence of diffuse fatty change. In addition, 600 mg/kg females exhibited significant increases in the incidences of degeneration of the gastric glands of the glandular stomach, as well as squamous hyperplasia, mineralization, and ulcer in the forestomach. Degeneration of the olfactory epithelium in the nose occurred in all 300 and 600 mg/kg mice. Estragole was not mutagenic in Salmonella typhimurium strains TA98, TA100, TA1535, or TA1537 when tested in the presence or absence of exogenous metabolic activation enzymes. No increases in the frequencies of micronucleated normochromatic erythrocytes were observed in peripheral blood samples from male and female mice in the 3-month study. Under the conditions of these 3-month studies, estragole showed carcinogenic activity based on the occurrence of two cholangiocarcinomas and one hepatocellular adenoma in the liver of three of 10 male F344/N rats in the high dose group. Because rats and mice were exposed for only 3 months, these studies do not access the full carcinogenic potential of estragole. Nonneoplastic effects were observed in the liver, glandular stomach, nose, kidney, and salivary gland of male and female rats and in the testes, epididymides, and pituitary gland of male rats. Nonneoplastic effects were also observed in the liver and nose of male and female mice and in the stomach of female mice.


CT120A Acts as an Oncogene in Head and Neck Squamous Cell Carcinoma.
Squamous cell carcinoma of the head and neck (HNSCC) is among the most frequent cancers worldwide. The etiology and pathogenesis of HNSCC are influenced by multiple genetic factors in addition to environmental and lifestyle-related factors. However, the mechanism underlying the HNSCC is still far from clear. The membrane associated gene CT120 was previously identified from chromosome 17p13.3 as a lung cancer-associated gene. Its function as an activator of the Erk and Akt signaling pathways in human lung cancer cell lines suggested that CT120 has an oncogenic function. However, there is no data in the literature on the role of CT120 in any other cancer type. Therefore, the aim of this study was to determine the expression rate and probable function of CT120 in HNSCC. Tumor tissues from 50 patients were analyzed by real-time quantitative RT-PCR to investigate the expression rate and by direct sequencing to differentiate the CT120A and CT120B variants. CT120 overexpression was observed in 58% of tumors compared to non-cancerous tissue samples and this up-regulation was directly associated with the upregulation of the CT120A variant and with the stage of the disease (p=0.001). Our results indicate that the CT120 gene may function in the development of HNSCC.


Comparison of PCR-based methods for the simultaneous detection of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae in clinical samples.
Several in-house PCR-based assays have been described for the detection of bacterial meningitis caused by Neisseria meningitidis, Streptococcus pneumoniae, and Haemophilus influenzae from clinical samples. PCR-based methods targeting different bacterial genes are frequently used by different laboratories worldwide, but no standard method has ever been established. The aim of our study was to compare different in-house and a commercial PCR-based tests for the detection of bacterial pathogens causing meningitis and invasive disease in humans. A total of 110 isolates and 134 clinical samples (99 cerebrospinal fluid and 35 blood samples) collected from suspected cases of invasive disease were analyzed. Specific sets of primers frequently used for PCR-diagnosis of the three pathogens were used and compared with the results achieved using the multiplex approach described here. Several different gene targets were used for each microorganism, namely ctrA, crgA and nspA for N. meningitidis, ply for S. pneumoniae, P6 and bexA for H. influenzae. All used methods were fast, specific and sensitive, while some of the targets used for the in-house PCR assay detected lower concentrations of genomic DNA than the commercial method. An additional PCR reaction is described for the differentiation of capsulated and non-capsulated H. influenzae strains, the while commercial method only detects capsulated strains. The in-house PCR methods here compared showed to be rapid, sensitive, highly specific, and cheaper than commercial methods. The in-house PCR methods could be easily adopted by public laboratories of developing countries for diagnostic purposes. The best results were achieved using primers targeting the genes nspA, ply, and P6 which were able to detect the lowest DNA concentrations for each specific target.


Post-treatment resistance analysis of hepatitis C virus from phase II and III clinical trials of ledipasvir/sofosbuvir.
Ledipasvir/sofosbuvir combination treatment in phase III clinical trials resulted in sustained viral suppression in 94-99% of patients. This study characterized drug resistance in treatment failures, which may help to inform retreatment options. We performed NS5A and NS5B deep sequencing of hepatitis C virus (HCV) from patients infected with genotype (GT) 1 who participated in ledipasvir/sofosbuvir phase II and III clinical trials. Fifty-one of 2144 (2.4%) (42 GT1a and 9 GT1b) treated patients met the criteria for resistance analysis due to virologic failure following the end of treatment. The majority of patients with virologic failure (38 of 51; 74.5%) had detectable ledipasvir-specific resistance-associated substitutions (RASs) at the time of virologic failure (1% deep sequencing cut-off). The percent of patients with NS5A RASs at virologic failure were 37.5%, 66.7%, 94.7% and 100% in patients treated for 6, 8, 12 and 24weeks, respectively. The common substitutions detected at failure were Q30R/H, and/or Y93H/N in GT1a and Y93H in GT1b. At failure, 35.3% (18/51) of virologic failure patients' viruses had two or more NS5A RASs and the majority of patients harbored NS5A RASs conferring a 100-1000-fold (n=10) or >1000-fold (n=23) reduced susceptibility to ledipasvir. One patient in a phase II study with a known ledipasvir RAS at baseline (L31M) developed the S282T sofosbuvir (NS5B) RAS at failure. In GT1 HCV-infected patients treated with ledipasvir/sofosbuvir±ribavirin, virologic failure was rare. Ledipasvir resistance in NS5A was selected or enhanced in most patients with virologic failure, one of whom also developed resistance to sofosbuvir. Clinical studies have shown that combination treatment with ledipasvir/sofosbuvir efficiently cures most patients with genotype 1 hepatitis C infection. For the few patients failing treatment, we show that resistance to ledipasvir was observed in most patients, whereas resistance to sofosbuvir was less common. This has important implications for the selection of optimal retreatment strategies for these patients.


Corexit 9500 Enhances Oil Biodegradation and Changes Active Bacterial Community Structure of Oil-Enriched Microcosms.
To better understand the impacts of Corexit 9500 on the structure and activity levels of hydrocarbon-degrading microbial communities, we analyzed next-generation 16S rRNA gene sequencing libraries of hydrocarbon enrichments grown at 5 and 25°C using both DNA and RNA extracts as the sequencing templates. Oil biodegradation patterns in both 5 and 25°C enrichments were consistent with those reported in the literature (i.e., aliphatics were degraded faster than aromatics). Slight increases in biodegradation were observed in the presence of Corexit at both temperatures. Differences in community structure were observed between treatment conditions in the DNA-based libraries. The 25°C consortia were dominated by Vibrio, Idiomarina, Marinobacter, Alcanivorax, and Thalassospira species, while the 5°C consortia were dominated by several species of the genera Flavobacterium, Alcanivorax, and Oleispira Most of these genera have been linked to hydrocarbon degradation and have been observed after oil spills. Colwellia and Cycloclasticus, known aromatic degraders, were also found in these enrichments. The addition of Corexit did not have an effect on the active bacterial community structure of the 5°C consortia, while at 25°C, a decrease in the relative abundance of Marinobacter was observed. At 25°C, Thalassospira, Marinobacter, and Idiomarina were present at higher relative abundances in the RNA than DNA libraries, suggesting that they were active in degradation. Similarly, Oleispira was greatly stimulated by the addition of oil at 5°C.IMPORTANCE While dispersants such as Corexit 9500 can be used to treat oil spills, there is still debate on the effectiveness on enhancing oil biodegradation and its potential toxic effect on oil-degrading microbial communities. The results of this study provide some insights on the microbial dynamics of hydrocarbon-degrading bacterial populations in the presence of Corexit 9500. Operational taxonomic unit (OTU) analyses indicated that several OTUs were inhibited by the addition of Corexit. Conversely, a number of OTUs were stimulated by the addition of the dispersant, many of which were identified as known hydrocarbon-degrading bacteria. The results highlight the value of using RNA-based methods to further understand the impact of dispersant on the overall activity of different hydrocarbon-degrading bacterial groups.


Quasielastic light scattering study of amyloid beta-protein fibril formation.
Quasielastic light scattering spectroscopy (QLS) is an optical method for the determination of diffusion coefficients of particles in solution. Here we discuss the principles of QLS and explain how the distribution of particle sizes can be reconstructed from the measured correlation function of scattered light. Non-invasive observation of the temporal evolution of particle sizes provides a powerful tool for studying protein assembly. We illustrate practical applications of QLS with examples from studies of fibril formation of the amyloid beta-protein.


Predictors of survival in neonates with critical aortic stenosis.
Failure of infants with critical aortic stenosis to survive after adequate valvotomy despite a left ventricular size that appears to be adequate indicates that additional preoperative anatomic features may contribute to mortality. Discriminant analysis was used to determine which of several echocardiographically measured left heart structures were independent predictors of survival after valvotomy for neonatal critical aortic stenosis. It was possible to predict outcome after classic valvotomy (two-ventricle-type repair) with 95% accuracy based on mitral valve area, long-axis dimension of the left ventricle relative to the long-axis dimension of the heart, diameter of the aortic root, and body surface area. Left ventricular volume was not a major determinant in this study, in part because patients who had initial valvotomy had been preselected in favor of an adequately sized left ventricle. Patients with multiple small left ventricular structures were found to have significantly improved survival after initial Norwood operation. In contrast, balloon valvotomy with subsequent Norwood procedure was usually unsuccessful. The adverse effects of small inflow, outflow, and/or cavity size of the left ventricle are cumulative. The accuracy of prediction of outcome based only on preoperative anatomy indicates that adequacy of valvotomy is not generally a limiting factor for survival in this group of patients. It is possible to identify subjects whose chance of survival is better after a Norwood procedure rather than valvotomy, even if left ventricular volume is not critically small.


A first online intervention to increase patients' perceived ability to act in situations of abuse in health care: reports of a Swedish pre-post study.
Efforts to counteract abuse in health care, defined as patient-experienced abuse, have mainly focused on interventions among caregivers. This study is the first to test an online intervention focusing on how patients can counteract such abuse. The intervention aimed at increasing patients' intention and perceived ability to act in future situations where they risk experiencing abuse. Participants were recruited through a nephrology clinic in Sweden. The intervention consisted of an online program that aimed to stimulate patients to think of possible actions in situations in which they risk experiencing abuse. The program comprised stories and exercises in text and comic form. The participants filled out a questionnaire immediately before and after going through the program, as well as during follow-up four to eight weeks later. Forty-eight patients (39 %) participated in the study and spent, on average, 41 min responding to questions and going through the program. Both men and women, of various ages and educational backgrounds, participated. An increase in participants' self-reported ability to identify opportunities to act in a given situation was seen immediately afterwards, as well as during follow up. The current study suggests that it is feasible and most likely useful to a variety of patients to work with the provided material that has the aim of counteracting abuse in health care. It would be of interest to further develop ways of using comics and to test similar interventions in other health care settings.


Diabetic Retinopathy and Macular Edema Quality-of-Life Item Banks: Development and Initial Evaluation Using Computerized Adaptive Testing.
The purpose of this study was to assess the psychometric properties of diabetic retinopathy (DR) and diabetic macular edema (DME) quality-of-life (QoL) item banks and determine the utility of the final calibrated item banks by simulating a computerized adaptive testing (CAT) application. In this clinical, cross-sectional study, 514 participants with DR/DME (mean age ± SD, 60.4 ± 12.6 years; 64% male) answered 314 items grouped under nine QoL item pools: Visual Symptoms (SY); Ocular Comfort Symptoms (OS); Activity Limitation (AL); Mobility (MB); Emotional (EM); Health Concerns (HC); Social (SC); Convenience (CV); and Economic (EC). The psychometric properties of the item pools were assessed using Rasch analysis, and CAT simulations determined the average number of items administered at high and moderate precision levels. The SY, MB, EM, and HC item pools required minor amendments, mainly involving removal of six poorly worded, highly misfitting items. AL and CV required substantial modification to resolve multidimensionality, which resulted in two new item banks: Driving (DV) and Lighting (LT). Due to unresolvable psychometric issues, the OS, SC, and EC item pools were not pursued further. This iterative process resulted in eight operational item banks that underwent CAT simulations. Correlations between CAT and the full item banks were high (range, 0.88-0.99). On average, only 3.6 and 7.2 items were required to gain measurement at moderate and high precision, respectively. Our eight psychometrically robust and efficient DR/DME item banks will enable researchers and clinicians to accurately assess the impact and effectiveness of treatment therapies for DR/DME in all areas of QoL.


Misleading one detail: a preventable mode of diagnostic error?
Despite advances in our understanding of cognitive biases in clinical practice, little is known about correction or prevention of diagnostic errors. The presence of a single misleading detail may lead clinicians down a cognitive and actual path toward an incorrect diagnosis. In a large teaching hospital, we surveyed 51 attending doctors in internal medicine, presenting each with 10 clinical vignettes and soliciting their diagnosis of the condition leading to the presentation. Each of the 10 clinical cases included a single misleading detail. This survey elicited a wrong diagnosis in 90% of cases, which was reduced to 30% when omitting the misleading detail from the vignette. Diagnostic accuracy did not improve by warning doctors about potentially misleading information. Asking doctors to identify a leading diagnostic detail and then to formulate an alternative diagnosis after omission of the detail, significantly reduced diagnostic error rate by nearly 50%. Systematic re-examination of leading diagnostic clues may help to reduce errors in diagnosis.


GIP: an open-source computational pipeline for mapping genomic instability from protists to cancer cells.
Genome instability has been recognized as a key driver for microbial and cancer adaptation and thus plays a central role in many diseases. Genome instability encompasses different types of genomic alterations, yet most available genome analysis software are limited to just one type of mutation. To overcome this limitation and better understand the role of genetic changes in enhancing pathogenicity we established GIP, a novel, powerful bioinformatic pipeline for comparative genome analysis. Here, we show its application to whole genome sequencing datasets of Leishmania, Plasmodium, Candida and cancer. Applying GIP on available data sets validated our pipeline and demonstrated the power of our tool to drive biological discovery. Applied to Plasmodium vivax genomes, our pipeline uncovered the convergent amplification of erythrocyte binding proteins and identified a nullisomic strain. Re-analyzing genomes of drug adapted Candida albicans strains revealed correlated copy number variations of functionally related genes, strongly supporting a mechanism of epistatic adaptation through interacting gene-dosage changes. Our results illustrate how GIP can be used for the identification of aneuploidy, gene copy number variations, changes in nucleic acid sequences, and chromosomal rearrangements. Altogether, GIP can shed light on the genetic bases of cell adaptation and drive disease biomarker discovery.


Heteropoly blue doped polymer nanoparticles: an efficient theranostic agent for targeted photoacoustic imaging and near-infrared photothermal therapy in vivo.
With the development of personalized medicine, the research of theranostic agents with good biocompatibility, stability, and targeting properties remains meaningful. Herein, we report a two-step process to construct heteropoly blue (HPB) doped polymer nanoparticles (HPB/P4VP-b-PEO NPs) with efficient near-infrared (NIR) light absorption and photothermal conversion efficiency of ∼23% to simultaneously perform photoacoustic imaging and photothermal therapy in vivo. After intravenous injection into the 4T1 xenograft model, in vivo photoacoustic imaging confirmed the targeted property of HPB/P4VP-b-PEO NPs due to the enhanced EPR effect. The photoacoustic signal in the tumor at 24 h p.i. was more than ∼10 times that of the pre-injection group. By virtue of their EPR effect, HPB/P4VP-b-PEO NPs achieved a good photothermal therapeutic efficacy in vivo.


[Hyperbaric oxygenation in the comprehensive therapy of patients with rheumatoid arthritis (clinico-immunologic study)].
For 35 of 50 patients with rheumatoid arthritis traditional drug therapy was a minor success for a long time. Without any modifications of the drug therapy every patient went through a course of hyperbaric oxygenation (HBO): 21 sessions under 1.7 ata for 40 min. Good clinical results both immediate and remote have been obtained. The effect of HBO on the immune system of the patients has intensified the suppressive function of T-lymphocytes (especially with systemic symptoms of the disease), normalized cell-bound immunity and decreased the serum concentration in immune complexes.


Chronic exposure to KATP channel openers results in attenuated glucose sensing in hypothalamic GT1-7 neurons.
Individuals with Type 1 diabetes (T1D) are often exposed to recurrent episodes of hypoglycaemia. This reduces hormonal and behavioural responses that normally counteract low glucose in order to maintain glucose homeostasis, with altered responsiveness of glucose sensing hypothalamic neurons implicated. Although the molecular mechanisms are unknown, pharmacological studies implicate hypothalamic ATP-sensitive potassium channel (KATP) activity, with KATP openers (KCOs) amplifying, through cell hyperpolarization, the response to hypoglycaemia. Although initial findings, using acute hypothalamic KCO delivery, in rats were promising, chronic exposure to the KCO NN414 worsened the responses to subsequent hypoglycaemic challenge. To investigate this further we used GT1-7 cells to explore how NN414 affected glucose-sensing behaviour, the metabolic response of cells to hypoglycaemia and KATP activity. GT1-7 cells exposed to 3 or 24 h NN414 exhibited an attenuated hyperpolarization to subsequent hypoglycaemic challenge or NN414, which correlated with diminished KATP activity. The reduced sensitivity to hypoglycaemia was apparent 24 h after NN414 removal, even though intrinsic KATP activity recovered. The NN414-modified glucose responsiveness was not associated with adaptations in glucose uptake, metabolism or oxidation. KATP inactivation by NN414 was prevented by the concurrent presence of tolbutamide, which maintains KATP closure. Single channel recordings indicate that NN414 alters KATP intrinsic gating inducing a stable closed or inactivated state. These data indicate that exposure of hypothalamic glucose sensing cells to chronic NN414 drives a sustained conformational change to KATP, probably by binding to SUR1, that results in loss of channel sensitivity to intrinsic metabolic factors such as MgADP and small molecule agonists.


Integrated high pressure microhydraulic actuation and control for surgical instruments.
To reduce the surgical trauma to the patient, minimally invasive surgery is gaining considerable importance since the eighties. More recently, robot assisted minimally invasive surgery was introduced to enhance the surgeon's performance in these procedures. This resulted in an intensive research on the design, fabrication and control of surgical robots over the last decades. A new development in the field of surgical tool manipulators is presented in this article: a flexible manipulator with distributed degrees of freedom powered by microhydraulic actuators. The tool consists of successive flexible segments, each with two bending degrees of freedom. To actuate these compliant segments, dedicated fluidic actuators are incorporated, together with compact hydraulic valves which control the actuator motion. Especially the development of microvalves for this application was challenging, and are the main focus of this paper. The valves distribute the hydraulic power from one common high pressure supply to a series of artificial muscle actuators. Tests show that the angular stroke of the each segment of this medical instrument is 90°.


Identification of osteocyte-selective proteins.
Since little is known regarding osteocytes, cells embedded within the mineralized bone matrix, a proteomics approach was used to discover proteins more highly expressed in osteocytes than in osteoblasts to determine osteocyte-specific function. Two proteomic profiles obtained by two different proteomic approaches using total cell lysates from the osteocyte cell line MLO-Y4 and the osteoblast cell line MC3T3 revealed unique differences. Three protein clusters, one related to glycolysis (Phosphoglycerate kinase 1, fructose-bisphosphate aldolase A, hypoxia up-regulated 1 [ORP150], triosephosphate isomerase), one to protein folding (Mitochondrial Stress-70 protein, ORP150, Endoplasmin), and one to actin cytoskeleton regulation (Macrophage-capping protein [CapG], destrin, forms of lamin A and vimentin) were identified. Higher protein expression of ORP-150, Cap G, and destrin in MLO-Y4 cells compared with MC3T3 cells was validated by gene expression, Western blotting, and in vivo expression. These proteins were shown to be selective in osteocytes in vivo using immuno-staining of mouse ulnae. Destrin was most highly expressed in embedding osteoid osteocytes, GapG in embedded osteocytes, and ORP150 in deeply embedded osteocytes. In summary, the proteomic approach has yielded important information regarding molecular mechanisms used by osteocytes for embedding in matrix, the formation of dendritic processes, and protection within a hypoxic environment.


Bee-eaters ( Merops orientalis) respond to what a predator can see.
Two sets of experiments are reported that show that the small green bee-eater ( Merops orientalis, a small tropical bird) can appreciate what a predator can or cannot see. Bee-eaters avoid entering the nest in the presence of a potential nest predator. In the first set of experiments bee-eaters entered the nest more frequently when the predator was unable to see the nest from its position, as compared to an approximately equidistant position from which the nest could be seen. In the second set of experiments bee-eaters entered the nest more frequently when the predator was looking away from the nest. The angle of gaze from the nest was associated significantly positively with the probability of entering the nest whereas the angle from the bird was not. Birds showed considerable flexibility as well as individual variation in the possible methods of judging the predator's position and direction of gaze.


Streptozotocin-induced deficits in sex behavior and neuroendocrine function in male rats.
The effects of streptozotocin-induced (STZ) diabetes on neuroendocrine and sexual function were evaluated in adult male rats. Adult male rats were injected with STZ (50 mg/kg) or vehicle and tested for copulatory behavior 7, 14, and 21 days later. The rats were killed 1 month after STZ or vehicle treatment for measurement of plasma hormone levels, hypothalamic catecholamine turnover, LHRH content, and in vitro pituitary function. The STZ rats showed significant deficits in mount, intromission, and ejaculatory behaviors. Plasma levels of testosterone, LH, FSH, and PRL were all significantly reduced in the STZ compared to the control rats, but in vitro LH secretion was enhanced after STZ treatment. In vitro PRL secretion and the inhibitory response to dopamine did not differ between the two groups. The levels of LHRH were reduced in the medial basal hypothalamus (MBH), but LHRH levels in the median eminence (ME) and anterior hypothalamus (AH) were unchanged after STZ treatment. Norepinephrine turnover was reduced in the ME, MBH, and AH of the STZ rats, while dopamine turnover was unchanged in the ME, increased in the MBH, and reduced in the AH of the STZ rats compared to those in the vehicle-treated controls. These results suggest that changes in pituitary and testicular function in rats made diabetic by STZ treatment are secondary to changes in hypothalamic catecholamine metabolism. Changes in copulatory behavior could be due to both reductions in plasma testosterone levels as well as changes in central neurotransmitter metabolism.


Heritability of lenticular myopia in English Springer spaniels.
We determined whether naturally-occurring lenticular myopia in English Springer spaniels (ESS) has a genetic component. Streak retinoscopy was performed on 226 related ESS 30 minutes after the onset of pharmacologic mydriasis and cycloplegia. A pedigree was constructed to determine relationships between affected offspring and parents. Estimation of heritability was done in a Bayesian analysis (facilitated by the MCMCglmm package of R) of refractive error in a model, including terms for sex and coat color. Myopia was defined as ≤-0.5 diopters (D) spherical equivalent. The median refractive error for ESS was 0.25 D (range, -3.5 to +4.5 D). Median age was 0.2 years (range, 0.1-15 years). The prevalence of myopia in related ESS was 19% (42/226). The ESS had a strong correlation (r = 0.95) for refractive error between the two eyes. Moderate heritability was present for refractive error with a mean value of 0.29 (95% highest probability density, 0.07-0.50). The distribution of refractive error, and subsequently lenticular myopia, has a moderate genetic component in ESS. Further investigation of genes responsible for regulation of the development of refractive ocular components in canines is warranted.


HGF/MET pathway aberrations as diagnostic, prognostic, and predictive biomarkers in human cancers.
Cancer is a major cause of death worldwide. MET tyrosine kinase receptor [MET, c-MET, hepatocyte growth factor (HGF) receptor] pathway activation is associated with the appearance of several hallmarks of cancer. The HGF/MET pathway has emerged as an important actionable target across many solid tumors; therefore, biomarker discovery becomes essential in order to guide clinical intervention and patient stratification with the aim of moving towards personalized medicine. The focus of this review is on how the aberrant activation of the HGF/MET pathway in tumor tissue or the circulation can provide diagnostic and prognostic biomarkers and predictive biomarkers of drug response. Many meta-analyses have shown that aberrant activation of the MET pathway in tumor tissue, including MET gene overexpression, gene amplification, exon 14 skipping and other activating mutations, is almost invariably associated with shorter survival and poor prognosis. Most meta-analyses have been performed in non-small cell lung cancer (NSCLC), breast, head and neck cancers as well as colorectal, gastric, pancreatic and other gastrointestinal cancers. Furthermore, several studies have shown the predictive value of MET biomarkers in the identification of patients who gain the most benefit from HGF/MET targeted therapies administered as single or combination therapies. The highest predictive values have been observed for response to foretinib and savolitinib in renal cancer, as well as tivantinib in NSCLC and colorectal cancer. However, some studies, especially those based on MET expression, have failed to show much value in these stratifications. This may be rooted in lack of standardization of methodologies, in particular in scoring systems applied in immunohistochemistry determinations or absence of oncogenic addiction of cancer cells to the MET pathway, despite detection of overexpression. Measurements of amplification and mutation aberrations are less likely to suffer from these pitfalls. Increased levels of MET soluble ectodomain (sMET) in circulation have also been associated with poor prognosis; however, the evidence is not as strong as it is with tissue-based biomarkers. As a diagnostic biomarker, sMET has shown its value in distinguishing cancer patients from healthy individuals in prostate and bladder cancers and in melanoma. On the other hand, increased circulating HGF has also been presented as a valuable prognostic and diagnostic biomarker in many cancers; however, there is controversy on the predictive value of HGF as a biomarker. Other biomarkers such as circulating tumor DNA (ctDNA) and tumor HGF levels have also been briefly covered. In conclusion, HGF/MET aberrations can provide valuable diagnostic, prognostic and predictive biomarkers and represent vital assets for personalized cancer therapy.


Comparison of the oxidative respiratory burst and mitogen-induced leukocyte responses of camels, goats, sheep, and cows.
The reports from the Ministry of Agriculture and Fisheries suggest that camels suffer less compared to goats, sheep, and cows from a number of common infectious diseases in Oman. However, there is no immunological evidence to substantiate this claim. This present study is, therefore, an attempt to study the immunological responses of camels, goats, sheep, and cows by comparing their oxidative respiratory burst of peripheral blood leukocytes (PBLs) as a marker of innate immunity occurring during phagocytosis and the mitogenic responses of their peripheral blood mononuclear leukocytes (PBMLs) as a marker of their adaptive immune response. Ten female adult animals (n = 10) were selected from each species (goats, sheep, and cows). The goats, sheep, and cows were maintained at the Agricultural Experiment Station, while camels were kept at the Royal Camel Corps (RCC). Blood samples were collected from the jugular vein in 7 mL of heparin and ethylenediaminetetraacetic acid vacutainer tubes. The oxidative respiratory burst of PBLs was measured using a chemiluminescence (CL) assay. Reactants consisted of 75 mL of whole blood diluted (1:50), 75 mL of luminol/isoluminol, and 75 mL of zymosan opsonized with non-heat inactivated serum/heat-inactivated serum or non-opsonized zymosan. CL responses were measured as relative light units and expressed as the mean count per minute and peak CL values. The mitogenic response of PBMLs to concanavalin A (Con-A), phytohemagglutinin (PHA), and pokeweed mitogen (PWM) was tested using a WST-8 assay and read spectrophotometrically at 450 nm. The present findings showed that camel PBLs generate significantly higher CL responses, both intracellularly as well as extracellularly, with zymosan opsonized with autologous serum. Camel PBLs demonstrated a significantly higher (p = 0.001) response when stimulated with zymosan opsonized with heat-inactivated serum compared to those of goat, sheep, and cow lymphocytes from camels exhibited significantly higher (p = 0.001) stimulation indices (SI) with Con-A, PHA, and PWM. The present study suggests that camels are capable of mounting both superior innate as well as adaptive immune responses and provide immunological evidence supporting the belief of some authors, who have proposed that camels are less susceptible to a number of common infectious diseases than other domesticated ruminants.


Geographic and socio-economic variation in markers of indoor air pollution in Nepal: evidence from nationally-representative data.
In low-income countries such as Nepal, indoor air pollution (IAP), generated by the indoor burning of biomass fuels, is the top-fourth risk factor driving overall morbidity and mortality. We present the first assessment of geographic and socio-economic determinants of the markers of IAP (specifically fuel types, cooking practices, and indoor smoking) in a nationally-representative sample of Nepalese households. Household level data on 11,040 households, obtained from the 2016 Nepal Demographic and Health Survey, were analyzed. Binary logistic regression analyses were conducted to assess the use of fuel types, indoor cooking practices, indoor smoking and IAP with respect to socio-economic indicators and geographic location of the household. More than 80% of the households had at least one marker of IAP: 66% of the household used unclean fuel, 45% did not have a separate kitchen to cook in, and 43% had indoor smoking. In adjusted binary logistic regression, female and educational attainment of household's head favored cleaner indoor environment, i.e., using clean fuel, cooking in a separate kitchen, not smoking indoors, and subsequently no indoor pollution. In contrast, households belonging to lower wealth quintile and rural areas did not favor a cleaner indoor environment. Households in Province 2, compared to Province 1, were particularly prone to indoor pollution due to unclean fuel use, no separate kitchen to cook in, and smoking indoors. Most of the districts had a high burden of IAP and its markers. Fuel choice and clean indoor practices are dependent on household socio-economic status. The geographical disparity in the distribution of markers of IAP calls for public health interventions targeting households that are poor and located in rural areas.


Constraint Handling Guided by Landscape Analysis in Combinatorial and Continuous Search Spaces.
The notion and characterisation of fitness landscapes has helped us understand the performance of heuristic algorithms on complex optimisation problems. Many practical problems, however, are constrained, and when significant areas of the search space are infeasible, researchers have intuitively resorted to a variety of constraint-handling techniques intended to help the algorithm manoeuvre through infeasible areas and toward feasible regions of better fitness. It is clear that providing constraint-related feedback to the algorithm to influence its choice of solutions overlays the violation landscape with the fitness landscape in unpredictable ways whose effects on the algorithm cannot be directly measured. In this work, we apply metrics of violation landscapes to continuous and combinatorial problems to characterise them. We relate this information to the relative performance of six well-known constraint-handling techniques to demonstrate how some properties of constrained landscapes favour particular constraint-handling approaches. For the problems with sampled feasible solutions, a bi-objective approach was the best performing approach overall, but other techniques performed better on problems with the most disjoint feasible areas. For the problems with no measurable feasibility, a feasibility ranking approach was the best performing approach overall, but other techniques performed better when the correlation between fitness values and the level of constraint violation was high.


Subcellular localization and immunological detection of proteins encoded by the vir locus of Bordetella pertussis.
The DNA sequence of the central regulatory locus vir of Bordetella pertussis predicts that three gene products, BvgA, BvgB, and BvgC, are encoded. Features of the predicted primary structures of these proteins and their homology to other two-component systems suggest that BvgA is located in the cytoplasm, BvgB is located in the periplasm, and BvgC spans the inner membrane. We have used gene fusions to the phoA and lacZ genes of Escherichia coli to investigate the subcellular localization and membrane topology of these proteins. PhoA fusion proteins were also purified and used to raise antibodies that allowed visualization of the vir-encoded polypeptides by Western immunoblotting. Our results have largely confirmed the predictions of the DNA sequence, with the exception that BvgB and BvgC were found to constitute one larger protein that was homologous to the sensor class of two-component systems. We propose that this protein be named BvgS (for sensor) and that its gene be named bvgS.


Distribution of different fibre types in human skeletal muscles: a method for the detection of neurogenic disorders.
Human skeletal muscles are composed of two distinguishable types of fibres, which in healthy muscles appear to be randomly arranged. Large groups of one fibre type are commonly regarded as evidence of a neuropathological process affecting the peripheral nerves or the nerve cells in the spinal cord. An objective method that detects non-random arrangements as a sign of a neurogenic disorder, particularly in its early stages, could improve diagnosis. The randomness, or otherwise, of the fibre type arrangement is here considered in terms of the numbers of fibres surrounded entirely by others of the same type (enclosed fibres). The distribution of the number of enclosed fibres is studied for a free-sampling model using Monte Carlo methods. The negative binomial distribution is shown to fit closely, where the parameters can be expressed in terms of the number of fibres and the fibre type proportion in a sample area. This result permits the calculation of significance levels for a sample area and the combination of information in several sample areas. Finally, the method is applied to whole cross-sections of 24 male human autopsied muscles.


Mechanisms of injury and emergency care of acute spinal cord injury in dogs and cats.
To review the literature in regards to the pathophysiology of acute spinal cord injury, and to describe current concepts in regards to patient assessment, diagnostic, and therapeutic measures with a special emphasis on emergency and critical care considerations. Acute spinal cord injury occurs in 2 phases. The primary injury occurs at the time of initial injury and may include intervertebral disk herniation, vertebral fracture or luxation, penetrating injury, and vascular anomalies such as fibrocartilaginous embolic myelopathy. Secondary injury occurs following primary injury and is multifactorial encompassing numerous biochemical and vascular events that result in progression of injury. The diagnosis is based on history and physical examination findings. A neurologic examination should be performed following initial patient assessment and stabilization. Further diagnostics to characterize acute spinal injury include radiographs and advanced imaging modalities such as myelography, computed tomography, or magnetic resonance imaging. Initial treatment should focus on addressing the patient's cardiovascular and respiratory system. Supportive measures to support systemic perfusion are vital to minimizing secondary injury. Specific therapy toward minimizing secondary injury in veterinary medicine remains controversial, especially in regards to the utilization of methylprednisolone. Other therapies are either in need of additional research or have failed to document clinical difference. The prognosis for acute spinal injury is varied and is dependent upon the presence of concurrent trauma, location, and type of primary injury sustained, and extent of neurologic impairment at the time of initial presentation. The etiology of the underlying trauma is of great importance in determining prognosis and outcome. Loss of deep pain is generally accepted as a poor prognostic indicator; however, even these patients can recover depending on their response to treatment.


Curcumin Treatment Improves Motor Behavior in α-Synuclein Transgenic Mice.
The curry spice curcumin plays a protective role in mouse models of neurodegenerative diseases, and can also directly modulate aggregation of α-synuclein protein in vitro, yet no studies have described the interaction of curcumin and α-synuclein in genetic synucleinopathy mouse models. Here we examined the effect of chronic and acute curcumin treatment in the Syn-GFP mouse line, which overexpresses wild-type human α-synuclein protein. We discovered that curcumin diet intervention significantly improved gait impairments and resulted in an increase in phosphorylated forms of α-synuclein at cortical presynaptic terminals. Acute curcumin treatment also caused an increase in phosphorylated α-synuclein in terminals, but had no direct effect on α-synuclein aggregation, as measured by in vivo multiphoton imaging and Proteinase-K digestion. Using LC-MS/MS, we detected ~5 ng/mL and ~12 ng/mL free curcumin in the plasma of chronic or acutely treated mice, with a glucuronidation rate of 94% and 97%, respectively. Despite the low plasma levels and extensive metabolism of curcumin, these results show that dietary curcumin intervention correlates with significant behavioral and molecular changes in a genetic synucleinopathy mouse model that mimics human disease.


A clinical practice guideline for physiotherapists treating patients with chronic obstructive pulmonary disease based on a systematic review of available evidence.
Update of a clinical practice guideline for the physiotherapy management of patients with chronic obstructive pulmonary disease supporting the clinical decision-making process. A systematic computerized literature search was performed on different modalities for improving physical exercise capacity, reducing exertional dyspnoea, improving airway clearance and encouraging changes in physical activity behaviour. Methodological quality was scored with the PEDro Scale. Scientific conclusions were graded according to the criteria of the; Dutch Evidence Based Guideline Development Platform'. These, together with practical considerations, were used to formulate recommendations for clinical practice. A total of 103 studies were included in the systematic review, consisting of five meta-analyses of randomized controlled trials, 84 randomized controlled trials and 14 uncontrolled studies. Twenty scientific conclusions supported six recommendations on physical exercise training. Nineteen scientific conclusions supported eight recommendations on interventions for reducing dyspnoea. Five scientific conclusions supported seven recommendations concerning treatment modalities to improve mucus clearance, and two scientific conclusions supported two recommendations on strategies for encouraging permanent changes in physical activity behaviour. Strong recommendations support the use of physical exercise training to improve health-related quality of life and functional exercise capacity. Future research should investigate whether additional interventions for reducing exertional dyspnoea have a place as adjuncts to physical exercise training in selected patients. In addition, treatment of impaired mucus clearance, especially during acute exacerbations, requires further research. With the advance of new technologies for objective measurements of physical activities in daily life more research is needed concerning interventions to initiate and maintain physical activity behaviour change during and after supervised physical exercise training programmes.


Stable nitrogen isotope ratios of bone collagen reflect marine and terrestrial components of prehistoric human diet.
The delta 15N values of bone collagen from Eskimos and from Northwest Coast Indians dependent on salmon fishing are about 10 per mil more positive than those from agriculturalists in historic times. Among prehistoric humans, two groups dependent on marine food sources show bone collagen delta 15N values that are 4 to 6 per mil more positive than those from two agricultural groups. The nitrogen isotope ratios of bone collagen from prehistoric inhabitants of the Bahamas are anomalously low for reasons that relate to the biogeochemical cycle of nitrogen in coral reefs.


Inducible clindamycin resistance and molecular epidemiologic trends of pediatric community-acquired methicillin-resistant Staphylococcus aureus in Dallas, Texas.
Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) infection occurs commonly in children. Clindamycin resistance may be inducible or constitutive, and the rates of inducible resistance in CA-MRSA that could produce clindamycin treatment failures vary worldwide. The double-disk test was performed in 197 erythromycin-resistant and clindamycin-susceptible CA-MRSA strains from children in Dallas, Texas, from 1999 to 2002 to determine inducible clindamycin resistance. Resistance mechanisms were studied by PCR; epidemiologic trends were studied by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST). Inducible resistance was demonstrated in 28 (93%+/-6%) of 30 tested isolates in 1999, 21 (64%, +/-11%) of 33 in 2000, 12 (23%+/-7%) of 52 in 2001, and 6 (7%+/-3%) of 82 in 2002. All noninducible strains had the msr(A) gene. Among inducible resistant strains, 31 had erm(B), 24 had erm(C), and 12 had erm(A) genes. Two distinct pulsed types were the most prevalent; one of them was the most common pulsed type in 1999, whereas in 2002 a different pulsed type was prevalent. MLST analyses determined that ST-8 was the most common type, with 76%+/-5% found in 2002. All but one of these clindamycin-susceptible, erythromycin-resistant ST-8 strains showed no induction of clindamycin resistance. We conclude that, among erythromycin-resistant, clindamycin-susceptible CA-MRSA strains isolated from children in Dallas, inducible methylase resistance became less common from 1999 to 2002 (P<0.001). The phenotype of strains was associated with their sequence type. Our results demonstrate a clonal shift in CA-MRSA in Dallas children from 1999 to 2002.


Variation of Age-Related Changes in Endurance Performance Between Modes of Locomotion in Men: An Analysis of Master World Records.
Age-related declines in sport performance are characteristic of all endurance and sprinting disciplines. However, it is not known if the mode of locomotion (ie, swimming, cycling or running) influences the age-related decline in sport performance in sprinting and endurance events. To examine the age-related decline in 3 different modes of locomotion (ie, swimming, cycling, and running) for endurance and sprint events, the world-best performances achieved for men in the age groups 18-39, 40-44, 45-49, 50-54, 55-59, 60-64, 65-69, 70-74, 75-79, and 80-84 y were compared in swimming (1500 and 50 m), cycling (1 h and 200 m), and running (10 and 100 m). Each performance was considered as an average speed (throughout the distance), and the age-related decline in performance was expressed as a percentage of the world record (regardless of age group) for that discipline. The age-related decline in 1-h track cycling is less pronounced than in 1500-m swimming and 10-km running after 60 y. In contrast, the age-related decline was similar among the 3 locomotion modes for the sprinting events. The data show that the maintenance of high performance in cycling persists longer into old age than in running and swimming.


Mutational Analysis of the QRRQ Motif in the Yeast Hig1 Type 2 Protein Rcf1 Reveals a Regulatory Role for the Cytochrome c Oxidase Complex.
The yeast Rcf1 protein is a member of the conserved family of proteins termed the hypoxia-induced gene (domain) 1 (Hig1 or HIGD1) family. Rcf1 interacts with components of the mitochondrial oxidative phosphorylation system, in particular the cytochrome bc1 (complex III)-cytochrome c oxidase (complex IV) supercomplex (termed III-IV) and the ADP/ATP carrier proteins. Rcf1 plays a role in the assembly and modulation of the activity of complex IV; however, the molecular basis for how Rcf1 influences the activity of complex IV is currently unknown. Hig1 type 2 isoforms, which include the Rcf1 protein, are characterized in part by the presence of a conserved motif, (Q/I)X3(R/H)XRX3Q, termed here the QRRQ motif. We show that mutation of conserved residues within the Rcf1 QRRQ motif alters the interactions between Rcf1 and partner proteins and results in the destabilization of complex IV and alteration of its enzymatic properties. Our findings indicate that Rcf1 does not serve as a stoichiometric component, i.e. as a subunit of complex IV, to support its activity. Rather, we propose that Rcf1 serves to dynamically interact with complex IV during its assembly process and, in doing so, regulates a late maturation step of complex IV. We speculate that the Rcf1/Hig1 proteins play a role in the incorporation and/or remodeling of lipids, in particular cardiolipin, into complex IV and. possibly, other mitochondrial proteins such as ADP/ATP carrier proteins.


A randomised controlled trial of energetic activity for depression in young people (READY): a multi-site feasibility trial protocol.
Prevalence of depression is increasing in young people, and there is a need to develop and evaluate behavioural interventions which may provide benefits equal to or greater than talking therapies or pharmacological alternatives. Exercise could be beneficial for young people living with depression, but robust, large-scale trials of effectiveness and the impact of exercise intensity are lacking. This study aims to test whether a randomised controlled trial (RCT) of an intervention targeting young people living with depression is feasible by determining whether it is possible to recruit and retain young people, develop and deliver the intervention as planned, and evaluate training and delivery. The design is a three-arm cluster randomised controlled feasibility trial with embedded process evaluation. Participants will be help-seeking young people, aged 13-17 years experiencing mild to moderate low mood or depression, referred from three counties in England. The intervention will be delivered by registered exercise professionals, supported by mental health support workers, twice a week for 12 weeks. The three arms will be high-intensity exercise, low-intensity exercise, and a social activity control. All arms will receive a 'healthy living' behaviour change session prior to each exercise session and the two exercise groups are energy matched. The outcomes are referral, recruitment, and retention rates; attendance at exercise sessions; adherence to and ability to reach intensity during exercise sessions; proportions of missing data; adverse events, all measured at baseline, 3, and 6 months; resource use; and reach and representativeness. UK National Health Service (NHS) policy is to provide young people with advice about using exercise to help depression but there is no evidence-based exercise intervention to either complement or as an alternative to medication or talking therapies. UK National Institute for Health and Care Excellence (NICE) guidelines suggest that exercise can be an effective treatment, but the evidence base is relatively weak. This feasibility trial will provide evidence about whether it is feasible to recruit and retain young people to a full RCT to assess the effectiveness and cost-effectiveness of an exercise intervention for depression. ISRCTN, ISRCTN66452702 . Registered 9 April 2020.


Shared associations of nonatherosclerotic, large-vessel, cerebrovascular arteriopathies: considering intracranial aneurysms, cervical artery dissection, moyamoya disease and fibromuscular dysplasia.
With ongoing advancements in noninvasive vascular imaging and high-throughput genomics, we have the opportunity to reclassify the cerebrocervical disorders by these shared associations, rather than their downstream events, and to better understand etiology, mechanism and preventive treatments going forward. The common nonatherosclerotic, large-vessel arteriopathies affecting the cerebrovasculature include intracranial aneurysms, cervical artery dissection, fibromuscular dysplasia and moyamoya disease. Together, these entities contribute to a high incidence of devastating cerebrovascular outcomes, including ischemic stroke and subarachnoid hemorrhage, leading to long-term physical and cognitive disability frequently in young otherwise healthy adults. In addition to well reported clinical overlap, these polygenic phenotypes share epidemiological characteristics, environmental risk and a common pathological weakening of the arterial wall. We reviewed both past and present studies relating these shared associations, including reported candidate gene analyses and genome-wide association data. We also catalogue recent descriptions of novel arteriopathic syndromes that add to the growing list of monogenic connective tissue disease affecting the arterial wall, and further inform our understanding of more common polygenic phenotypes. We also place these cerebrocervical arteriopathies in the context of other systemic nonatherosclerotic, large-vessel vascular disease (e.g. aortic aneurysm and dissection).


[Hygiene status of ambulances and equipment in rescue services].
Focus on hygienic management in ambulances is presented based on findings from hygienic microbiological sampling. In 44 ambulances, the handwashing area, equipment for artificial respiration, insufflation, intubation, aspiration, intravasal catheterisation, blood-pressure measurement, and sterile materials storage were examined hygienically and microbiologically using the following methods: water sampling, imprint technique, and swab technique with subsequent cultivation on blood-, endo-, and Sabouraud agar. The highest contamination was found in the hand-washing area and on insufflation equipment (up to > or = 100,000 cfu/ml), where proof of the potential pathogens Pseudomonas, Acinetobacter, and Alcaligenes spp. was found. The second highest contamination level was found on the sphygmomanometer cuffs, stethoscopes, and respirator masks (e.g., Enterococci and S. aureus were identified). Apparently, the germs chiefly originate from the drinking water at the handwashing station, from the environment and mucous membranes from the skin, and intestines of the ambulance personnel. The predominant hygienic deficiensies were found in the equipment of the handwashing areas, and in the storage of resuscitation equipment and sterile materials. Based on the results of hygienic microbiological analysis, the main focus of hygienic measures--besides hand disinfection--should be on spray disinfection of the sphygmomanometer cuffs and stethoscopes as well as disinfection of ambulance equipment and storage areas for sterile devices, supplemented by dust protection of ready-to-use materials and devices. Stationary dispensers for hand disinfection possessing at least midlength levers must become the standard in ambulances, and conventional handwashing basins with plastic drinking-water containers and pump must be eliminated entirely.


Medicinal plants recommended by the world health organization: DNA barcode identification associated with chemical analyses guarantees their quality.
Medicinal plants are used throughout the world, and the regulations defining their proper use, such as identification of the correct species and verification of the presence, purity and concentration of the required chemical compounds, are widely recognized. Herbal medicines are made from vegetal drugs, the processed products of medicinal species. These processed materials present a number of challenges in terms of botanical identification, and according to the World Health Organization (WHO), the use of incorrect species is a threat to consumer safety. The samples used in this study consisted of the dried leaves, flowers and roots of 257 samples from 8 distinct species approved by the WHO for the production of medicinal herbs and sold in Brazilian markets. Identification of the samples in this study using DNA barcoding (matK, rbcL and ITS2 regions) revealed that the level of substitutions may be as high as 71%. Using qualitative and quantitative chemical analyses, this study identified situations in which the correct species was being sold, but the chemical compounds were not present. Even more troubling, some samples identified as substitutions using DNA barcoding contained the chemical compounds from the correct species at the minimum required concentration. This last situation may lead to the use of unknown species or species whose safety for human consumption remains unknown. This study concludes that DNA barcoding should be used in a complementary manner for species identification with chemical analyses to detect and quantify the required chemical compounds, thus improving the quality of this class of medicines.


The effect of tDCS over the right temporo-parietal junction on pain empathy.
Empathy is a complex psychological phenomenon crucial for social perception and interactions. Several lines of evidence suggest that the right temporo-parietal junction is involved in self-other control mechanisms that play an important role in empathic responses. However, limited direct evidence of the involvement of this region in empathic responses is currently available. In this study, inhibitory transcranial direct current stimulation over this region influenced empathic responses to others' pain. It was found that compared to participants that received anodal or sham transcranial direct current stimulation, participants who received cathodal transcranial direct current stimulation over the right temporo-parietal junction perceived the pain of others as less intense compared to sham stimulation and showed decreased late event related potentials to facial expressions of pain. Furthermore, it was found the stimulation had no significant effect on measures of sensorimotor resonance and physiological responses to pain in others. Our results demonstrate that the right temporo-parietal junction plays a role in empathic responses and that its inhibition can decrease behavioural and cerebral measures related to the cognitive-evaluative component of empathy. It is proposed that the right temporo-parietal junction is a valid stimulation target to study the influence of self-other control in empathic processes and could be useful to study the involvement of this region observed in clinical conditions characterized by altered empathic responses.


Integrase inhibitors in late pregnancy and rapid HIV viral load reduction.
Minimizing time to HIV viral suppression is critical in pregnancy. Integrase strand transfer inhibitors (INSTIs), like raltegravir, are known to rapidly suppress plasma HIV RNA in nonpregnant adults. There are limited data in pregnant women. We describe time to clinically relevant reduction in HIV RNA in pregnant women using INSTI-containing and non-INSTI-containing antiretroviral therapy (ART) options. We conducted a retrospective cohort study of pregnant HIV-infected women in the United States from 2009 through 2015. We included women who initiated ART, intensified their regimen, or switched to a new regimen due to detectable viremia (HIV RNA >40 copies/mL) at ≥20 weeks gestation. Among women with a baseline HIV RNA permitting 1-log reduction, we estimated time to 1-log RNA reduction using the Kaplan-Meier estimator comparing women starting/adding an INSTI in their regimen vs other ART. To compare groups with similar follow-up time, we also conducted a subgroup analysis limited to women with ≤14 days between baseline and follow-up RNA data. This study describes 101 HIV-infected pregnant women from 11 US clinics. In all, 75% (76/101) of women were not taking ART at baseline; 24 were taking non-INSTI containing ART, and 1 received zidovudine monotherapy. In all, 39% (39/101) of women started an INSTI-containing regimen or added an INSTI to their ART regimen. Among 90 women with a baseline HIV RNA permitting 1-log reduction, the median time to 1-log RNA reduction was 8 days (interquartile range [IQR], 7-14) in the INSTI group vs 35 days (IQR, 20-53) in the non-INSTI ART group (P < .01). In a subgroup of 39 women with first and last RNA measurements ≤14 days apart, median time to 1-log reduction was 7 days (IQR, 6-10) in the INSTI group vs 11 days (IQR, 10-14) in the non-INSTI group (P < .01). ART that includes INSTIs appears to induce more rapid viral suppression than other ART regimens in pregnancy. Inclusion of an INSTI may play a role in optimal reduction of HIV RNA for HIV-infected pregnant women presenting late to care or failing initial therapy. Larger studies are urgently needed to assess the safety and effectiveness of this approach.


Two New Species of Sucking Lice (Phthiraptera: Anoplura: Hoplopleuridae and Polyplacidae) from Grant's Rock Mouse, Micaelamys granti, in South Africa.
Two new species of sucking lice (Phthiraptera: Anoplura), Hoplopleura granti n. sp. (Hoplopleuridae) and Polyplax megacephalus n. sp. (Polyplacidae), are described from Grant's rock mouse, Micaelamys granti (Wroughton), from Northern Cape Province, South Africa. Adults of both sexes are illustrated via line drawings and differential interference contrast microscopy images, and are compared with previously described related species that parasitize related hosts: Hoplopleura patersoni Johnson from Aethomys chrysophilus (de Winton) (red veld rat), Aethomys kaiseri (Noack) (Kaiser's aethomys), and Micaelamys namaquensis (A. Smith) (Namaqua rock mouse); Hoplopleura aethomydis Kleynhans from M. namaquensis; Polyplax praomydis Bedford from A. chrysophilus and M. namaquensis; and Polyplax solivaga Johnson from A. chrysophilus. It is not known if these new species of lice are vectors of any pathogens.


Identification of relevant single-nucleotide polymorphisms in Pneumocystis jirovecii: relationship with clinical data.
Pneumocystis jirovecii is a poorly understood pathogen that causes opportunistic pneumonia (Pneumocystis pneumonia (PcP)) in patients with AIDS. The present study was aimed at correlating genetic differences in P. jirovecii isolates and clinical patient data. A description of genetic diversity in P. jirovecii isolates from human immunodeficiency virus-positive patients, based on the identification of multiple single-nucleotide polymorphisms (SNPs) at five distinct loci encoding mitochondrial large-subunit rRNA (mtLSU rRNA), cytochrome b (CYB), superoxide dismutase (SOD), dihydrofolate reductase (DHFR), and dihydropteroate synthase (DHPS), was achieved using PCR with DNA sequencing and restriction fragment length polymorphism analysis. The statistical analysis revealed several interesting correlations among the four most relevant SNPs (mt85, SOD110, SOD215, and DHFR312) and specific clinical parameters: mt85C was associated with undiagnosed or atypical PcP episodes and favourable follow-up; SOD215C was associated with favourable follow-up; and DHFR312T was associated with PcP cases presenting moderate to high parasite burdens. The genotypes mt85C/SOD215C and SOD110T/SOD215C were found to be associated with less virulent P. jirovecii infections, whereas the genotype SOD110T/SOD215T was found to be related to more virulent PcP episodes. The present work demonstrated that potential P. jirovecii haplotypes may be related to the clinical data and outcome of PcP.


IL-17A impairs host tolerance during airway chronic infection by Pseudomonas aeruginosa.
Resistance and tolerance mechanisms participate to the interplay between host and pathogens. IL-17-mediated response has been shown to be crucial for host resistance to respiratory infections, whereas its role in host tolerance during chronic airway colonization is still unclear. Here, we investigated whether IL-17-mediated response modulates mechanisms of host tolerance during airways chronic infection by P. aeruginosa. First, we found that IL-17A levels were sustained in mice at both early and advanced stages of P. aeruginosa chronic infection and confirmed these observations in human respiratory samples from cystic fibrosis patients infected by P. aeruginosa. Using IL-17a(-/-) or IL-17ra(-/-) mice, we found that the deficiency of IL-17A/IL-17RA axis was associated with: i) increased incidence of chronic infection and bacterial burden, indicating its role in the host resistance to P. aeruginosa; ii) reduced cytokine levels (KC), tissue innate immune cells and markers of tissue damage (pro-MMP-9, elastin degradation, TGF-β1), proving alteration of host tolerance. Blockade of IL-17A activity by a monoclonal antibody, started when chronic infection is established, did not alter host resistance but increased tolerance. In conclusion, this study identifies IL-17-mediated response as a negative regulator of host tolerance during P. aeruginosa chronic airway infection.


Intravaginal immunization using the recombinant HIV-1 clade-C trimeric envelope glycoprotein CN54gp140 formulated within lyophilized solid dosage forms.
Vaccine-mediated prevention of primary HIV-1 infection at the heterosexual mucosal portal of entry may be facilitated by highly optimised formulations or drug delivery devices for intravaginal (i.vag) immunization. Previously we described hydroxyethylcellulose (HEC)-based rheologically structured gel vehicles (RSVs) for vaginal immunization of an HIV-1 vaccine candidate, a soluble recombinant trimeric HIV-1 clade-C envelope glycoprotein designated CN54gp140. Here we investigated the efficacy of lyophilized solid dosage formulations (LSDFs) for prolonging antigen stability and as i.vag delivery modalities. LSDFs were designed and developed that upon i.vag administration they would reconstitute with the imbibing of vaginal fluid to mucoadhesive, site-retentive semi-solids. Mice were immunized with lyophilized equivalents of (i) RSVs, (ii) modified versions of the RSVs more suited to lyophilization (sodium carboxymethyl cellulose (NaCMC)-based gels) and (iii) Carbopol(®) gel, all containing CN54gp140. NaCMC-based LSDFs provided significantly enhanced antigen stability compared to aqueous-based RSVs. Rheological analysis indicated the NaCMC-based LSDFs would offer enhanced vaginal retention in woman compared to more conventional vaginal gel formulations. All LSDFs were well tolerated in the mouse model. Following i.vag administration, all LSDFs boosted systemic CN54gp140-specific antibody responses in sub-cutaneously primed mice. Induction of CN54gp140-specific antibody responses in the female genital tract was evident. Of all the LSDFs the fastest releasing which was lyophilized Carbopol(®) gel elicited immune responses comparable to buffer instillation of antigen suggesting that rather than slower sustained release, initial high burst release from the LSDFs may suffice. The boosting of specific immune responses upon i.vag administration indicates that LSDFs are viable mucosal vaccine delivery modalities promoting antigen stability and facilitating intimate exposure of CN54gp140 to the mucosal-associated lymphoid tissue of the female genital tract.


Relationship between attention-deficit hyperactivity disorder symptoms and problem gambling: A mediation analysis of influential factors among 7,403 individuals from the UK.
Our goal was to examine the association between attention-deficit hyperactivity disorder (ADHD) symptoms and gambling problems, and to identify potential mediating factors of this association. This study used cross-sectional, community-based data from 7,403 people aged ≥16 years who participated in the Adult Psychiatric Morbidity Survey 2007. ADHD symptoms were assessed using the Adult ADHD Self-Report Scale (ASRS) Screener. Problem gambling was assessed using a questionnaire based on the 10 DSM-IV diagnostic criteria for pathological gambling. Respondents were classified as having no problem, at-risk, or problem gambling. Logistic regression and mediation analyses were conducted to analyze the association between ADHD symptoms (i.e., ASRS score ≥14) and problem gambling and the role of several variables in this association. The prevalence of at-risk (5.3% vs. 2.4%) and problem gambling (2.4% vs. 0.6%) was higher in individuals with ADHD symptoms than in those without ADHD symptoms. ADHD symptoms were significantly associated with both at-risk (OR = 2.15; 95% CI = 1.22-3.79) and problem gambling (OR = 3.57; 95% CI = 1.53-8.31) when adjusted for age, sex, and ethnicity. Common mental disorders (CMDs; i.e., depression and anxiety disorders) (mediated percentage = 22.4%), borderline personality disorder (BPD) traits (22.1%), stressful life events (13.2%), stress at work or home (12.6%), alcohol dependence (11.8%), and impulsivity (11.2%) were significant mediators in the ADHD-gambling association. Overall, ADHD symptoms were positively associated with problem gambling. CMDs, BPD traits, and stressful life events were important mediators in this relationship.


Recognising Bias in Studies of Diagnostic Tests Part 1: Patient Selection.
In this two-part series on sources of bias in studies of diagnostic test performance, we outline common errors and optimal conditions during three study phases: patient selection, interpretation of the index test and disease verification by a gold standard. Here in part 1, biases associated with suboptimal participant selection are discussed through the lens of partial verification bias and spectrum bias, both of which increase the proportion of participants who are the 'sickest of the sick' or the 'wellest of the well.' Especially through retrospective methodology, partial verification introduces bias by including patients who are test positive by a gold standard, since patients with a positive index test are more likely to go on to further gold standard testing. Spectrum bias is frequently introduced through case-control design, dropping of indeterminate results or convenience sampling. After reading part 1, the informed clinician should be better able to judge the quality of a diagnostic test study, its inherent limitations and whether its results could be generalisable to their practice. Part 2 will describe how interpretation of the index test and disease verification by a gold standard can contribute to diagnostic test bias.


[Present situation of hepatitis B in Chile].
Hepatitis B virus infection generates carriers and 8% will evolve to a chronic phase. To perform a compilation of studies on hepatitis B in Chile and other sources of information to estimate the impact of this disease in our country. Published and unpublished evidence about the infection, in the general population and risk groups in our country, was compiled and reviewed critically. Informal interviews with experts, revision of the mandatory notification book of the Ministry of Health and collection of data from laboratories that study hepatitis B virus, were also carried out. The seroprevalence of chronic carriers in blood donors is nearly O.3%. Among risk groups such as health care personnel, the figure is O.7%, among homosexuals 29%, among HIV positive patients 30%, among sexual workers 2% and among children with chronic hemodialysis, 9%. Prevalence rate according to notified cases in 2004 was 1.8 x 100,000 inhabitants. Detection of viral hepatitis B surface antigen in laboratories occurs in 0.2% of donors and 1.396 of non donors. The seroprevalence of hepatitis B virus, the lack of notification, and the introduction of hepatitis B vaccine to our Regular Program of Immunizations, are arguments to develop in Chile a hepatitis B and C surveillance system.


On the potential of using nanocellulose for consolidation of painting canvases.
Nanocellulose has been recently proposed as a novel consolidant for historical papers. Its use for painting canvas consolidation, however, remains unexplored. Here, we show for the first time how different nanocelluloses, namely mechanically isolated cellulose nanofibrils (CNF), carboxymethylated cellulose nanofibrils (CCNF) and cellulose nanocrystals (CNC), act as a bio-based alternative to synthetic resins and other conventional canvas consolidants. Importantly, we demonstrate that compared to some traditional consolidants, all tested nanocelluloses provided reinforcement in the adequate elongation regime. CCNF showed the best consolidation per added weight; however, it had to be handled at very low solids content compared to other nanocelluloses, exposing canvases to larger water volumes. CNC reinforced the least per added weight but could be used in more concentrated suspensions, giving the strongest consolidation after an equivalent number of coatings. CNF performed between CNC and CCNF. All nanocelluloses showed better consolidation than lining with synthetic adhesive (Beva 371) and linen canvas in the elongation region of interest.


Seroprevalence and risk factors of Q fever in goats on commercial dairy goat farms in the Netherlands, 2009-2010.
The aim of this study was to estimate the seroprevalence of Coxiella burnetii in dairy goat farms in the Netherlands and to identify risk factors for farm and goat seropositivity before mandatory vaccination started. We approached 334 eligible farms with more than 100 goats for serum sampling and a farm questionnaire. Per farm, median 21 goats were sampled. A farm was considered positive when at least one goat tested ELISA positive. In total, 2,828 goat serum samples from 123 farms were available. Farm prevalence was 43.1% (95%CI: 34.3%-51.8%). Overall goat seroprevalence was 21.4% (95%CI: 19.9%-22.9%) and among the 53 positive farms 46.6% (95%CI: 43.8%-49.3%). Multivariable logistic regression analysis included 96 farms and showed that farm location within 8 kilometres proximity from a bulk milk PCR positive farm, location in a municipality with high cattle density (≥ 100 cattle per square kilometre), controlling nuisance animals through covering airspaces, presence of cats or dogs in the goat stable, straw imported from abroad or unknown origin and a herd size above 800 goats were independent risk factors associated with Q fever on farm level. At animal level almost identical risk factors were found, with use of windbreak curtain and artificial insemination as additional risk factors. In 2009-2010, the seroprevalence in dairy goats in the Netherlands increased on animal and farm level compared to a previous study in 2008. Risk factors suggest spread from relatively closely located bulk milk-infected small ruminant farms, next to introduction and spread from companion animals, imported straw and use of artificial insemination. In-depth studies investigating the role of artificial insemination and bedding material are needed, while simultaneously general biosecurity measures should be updated, such as avoiding companion animals and vermin entering the stables, next to advice on farm stable constructions on how to prevent introduction and minimize airborne transmission from affected dairy goat farms to prevent further spread to the near environment.


Randomized, double-blind, comparative-effectiveness study comparing pulsed radiofrequency to steroid injections for occipital neuralgia or migraine with occipital nerve tenderness.
Occipital neuralgia (ON) is characterized by lancinating pain and tenderness overlying the occipital nerves. Both steroid injections and pulsed radiofrequency (PRF) are used to treat ON, but few clinical trials have evaluated efficacy, and no study has compared treatments. We performed a multicenter, randomized, double-blind, comparative-effectiveness study in 81 participants with ON or migraine with occipital nerve tenderness whose aim was to determine which treatment is superior. Forty-two participants were randomized to receive local anesthetic and saline, and three 120 second cycles of PRF per targeted nerve, and 39 were randomized to receive local anesthetic mixed with deposteroid and 3 rounds of sham PRF. Patients, treating physicians, and evaluators were blinded to interventions. The PRF group experienced a greater reduction in the primary outcome measure, average occipital pain at 6 weeks (mean change from baseline -2.743 ± 2.487 vs -1.377 ± 1.970; P < 0.001), than the steroid group, which persisted through the 6-month follow-up. Comparable benefits favoring PRF were obtained for worst occipital pain through 3 months (mean change from baseline -1.925 ± 3.204 vs -0.541 ± 2.644; P = 0.043), and average overall headache pain through 6 weeks (mean change from baseline -2.738 ± 2.753 vs -1.120 ± 2.1; P = 0.037). Adverse events were similar between groups, and few significant differences were noted for nonpain outcomes. We conclude that although PRF can provide greater pain relief for ON and migraine with occipital nerve tenderness than steroid injections, the superior analgesia may not be accompanied by comparable improvement on other outcome measures.


Awareness, Care and Treatment In Obesity maNagement to inform Haemophilia Obesity Patient Empowerment (ACTION-TO-HOPE): Results of a survey of US patients with haemophilia and obesity (PwHO) and their partners and caregivers.
The ACTION study identified barriers to initiating and maintaining weight loss in patients with obesity; however, joint-related issues (pain, mobility and bleeding) may affect perceptions of patients with haemophilia and obesity (PwHO). To identify patient and caregiver insights on the unique challenges of PwHO. Following IRB approval, adults who self-identified as PwHO, spouses/partners of adult PwHO, and caregivers of adolescent PwHO (aged 12-17 years) completed an online survey between December 2017 and April 2018. Respondents included 124 adult PwHO, 45 spouses/partners and 42 caregivers. By calculated BMI, most adults were overweight (43%) or had obesity (51%); this differed from self-reported weight category. PwHO goals were improving health conditions (60%), having more energy (54%), reducing risks of weight (46%), and losing any weight (44%). Issues related to joint health were secondary for PwHO but frequently reported by spouses/parents. Most perceived weight loss to be a high priority (66%) and their responsibility (64%) but required a complete lifestyle change (63%). Most anticipated that weight loss would reduce joint pain (62%), bleeding (58%) and factor use (52%) and increase mobility (62%). Weight discussions with healthcare providers (HCPs) were commonly reported (51%). HCP discussions targeted improving health conditions (46%), achieving any weight loss (44%), being more active (73%) and improving eating habits (72%). Most PwHO (65%) perceived obesity as a disease and believe that 10% weight loss would be extremely beneficial (78%). In the past 5 years, 80% discussed being overweight and 68% losing weight; a minority reported being successful (9%) or somewhat successful (38%) with weight loss. More realistic or specific (51%/47%) goals, resources (46%), referrals to weight-loss programmes (41%) or dietitians (38%), meals or recipes (54%/50%), local or national (42%/41%) programmes for PwHO and success stories of PwHO (40%) are needed or would be helpful. PwHO, spouse/partners and caregivers exhibited awareness of general and haemophilia-specific consequences of excess body weight. Most have tried general approaches to improve eating and increase activity with little success and desire more education on weight management and more details on specific actionable recommendations distributed through existing haemophilia channels. These insights will better inform the creation of weight-loss programmes for this community.


Lipoxins, aspirin-triggered epi-lipoxins, lipoxin stable analogues, and the resolution of inflammation: stimulation of macrophage phagocytosis of apoptotic neutrophils in vivo.
Lipoxins (LX) are eicosanoids with antiinflammatory activity in glomerulonephritis (GN) and inflammatory diseases, hypersensitivity, and ischemia reperfusion injury. It has been demonstrated that LXA(4) stimulates non-phlogistic phagocytosis of apoptotic polymorphonuclear neutrophils (PMN) by monocyte-derived macrophages (Mphi) in vitro, suggesting a role for LX as endogenous pro-resolution lipid mediators. It is here reported that LXA(4), LXB(4), the aspirin-triggered LX (ATL) epimer, 15-epi-LXB(4), and a stable synthetic analogue 15(R/S)-methyl-LXA(4) stimulate phagocytosis of exogenously administered excess apoptotic PMN by macrophages (M phi) in vivo in a classic model of acute inflammation, namely thioglycollate-induced peritonitis. Significant enhancement of phagocytosis in vivo was observed with 15-min exposure to LX and with intraperitoneal doses of LXA(4), LXB(4), 15(R/S)-methyl-LXA(4), and 15-epi-LXB(4) of 2.5 to 10 micro g/kg. Non-phlogistic LX-stimulated phagocytosis by M phi was sensitive to inhibition of PKC and PI 3-kinase and associated with increased production of transforming growth factor-beta(1) (TGF-beta(1)). LX-stimulated phagocytosis was not inhibited by phosphatidylserine receptor (PSR) antisera and was abolished by prior exposure of M phi to beta 1,3-glucan, suggesting a novel M phi-PMN recognition mechanism. Interestingly, the recently described peptide agonists of the LXA(4) receptor (MYFINITL and LESIFRSLLFRVM) stimulated phagocytosis through a process associated with increased TGF-beta(1) release. These data provide the first demonstration that LXA(4), LXB(4), ATL, and LX stable analogues rapidly promote M phi phagocytosis of PMN in vivo and support a role for LX as rapidly acting, pro-resolution signals in inflammation. Engagement of the LXR by LX generated during cell-cell interactions in inflammation and by endogenous LXR peptide agonists released from distressed cells may be an important stimulus for clearance of apoptotic cells and may be amenable to pharmacologic mimicry for therapeutic gain.


Mouse models of human chromosomal translocations and approaches to cancer therapy.
Cancer arises because of genetic changes in somatic cells, eventually giving rise to overt malignancy. Principle among genetic changes found in tumor cells are chromosomal translocations which give rise to fusion genes or enforced oncogene expression. These mutations are tumor-specific and result in production of tumor-specific mRNAs and proteins and are attractive targets for therapy. Also, in acute leukemias, many of these molecules are transcription regulators which involve cell-type-specific complexes, offering an alternative therapy via interfering with protein-protein interaction. We are studying these various features of tumor cells to evaluate new therapeutic methods. We describe a mouse model of de novo chromosomal translocations using the Cre-loxP system in which interchromosomal recombination occurs between the Mll and Af9 genes. We are also developing other in vivo methods designed, like the Cre-loxP system, to emulate the effects of these chromosomal abnormalities in human tumors. In addition, we describe new technologies to facilitate the intracellular targeting of fusion mRNAs and proteins resulting from such chromosomal translocations. These include a masked antisense RNA method with the ability to discriminate between closely related RNA targets and the selection and use of intracellular antibodies to bind to target proteins in vivo and cause cell death. These approaches should also be adaptable to targeting point mutations or to differentially expressed tumor-associated proteins. We hope to develop therapeutic approaches for use in cancer therapy after testing their efficacy in our mouse models of human cancer.


Side-chain substitutions within angiotensin II reveal different requirements for signaling, internalization, and phosphorylation of type 1A angiotensin receptors.
Binding of the peptide hormone angiotensin II (AngII) to the type 1 (AT(1A)) receptor and the subsequent activation of phospholipase C-mediated signaling, involves specific determinants within the AngII peptide sequence. In contrast, the contribution of such determinants to AT(1A) receptor internalization, phosphorylation and activation of mitogen-activated protein kinase (MAPK) signaling is not known. In this study, the internalization of an enhanced green fluorescent protein-tagged AT(1A) receptor (AT(1A)-EGFP), in response to AngII and a series of substituted analogs, was visualized and quantified using confocal microscopy. AngII-stimulation resulted in a rapid, concentration-dependent internalization of the chimeric receptor, which was prevented by pretreatment with the nonpeptide AT(1) receptor antagonist EXP3174. Remarkably, AT(1A) receptor internalization was unaffected by substitution of AngII side chains, including single and double substitutions of Tyr(4) and Phe(8) that abolish phospholipase C signaling through the receptor. AngII-induced receptor phosphorylation was significantly inhibited by several substitutions at Phe(8) as well as alanine replacement of Asp(1). The activation of MAPK was only significantly inhibited by substitutions at position eight in the peptide and specific substitutions did not equally inhibit inositol phosphate production, receptor phosphorylation and MAPK activation. These results indicate that separate, yet overlapping, contacts made between the AngII peptide and the AT(1A) receptor select/induce distinct receptor conformations that preferentially affect particular receptor outcomes. The requirements for AT(1A) receptor internalization seem to be less stringent than receptor activation and signaling, suggesting an inherent bias toward receptor deactivation.


Polycystin-1 is required for stereocilia structure but not for mechanotransduction in inner ear hair cells.
The polycystic kidney disease-1 (Pkd1) gene encodes a large transmembrane protein (polycystin-1, or PC-1) that is reported to function as a fluid flow sensor in the kidney. As a member of the transient receptor potential family, PC-1 has also been hypothesized to play a role in the elusive mechanoelectrical transduction (MET) channel in inner ear hair cells. Here, we analyze two independent mouse models of PC-1, a knock-in (KI) mutant line and a hair cell-specific inducible Cre-mediated knock-out line. Both models exhibit normal MET channel function at neonatal ages despite hearing loss and ultrastructural abnormalities of sterecilia that remain properly polarized at adult ages. These findings demonstrate that PC-1 plays an essential role in stereocilia structure and maintenance but not directly in MET channel function or planar cell polarity. We also demonstrate that PC-1 is colocalized with F-actin in hair cell stereocilia in vivo, using a hemagglutinin-tagged PC-1 KI mouse model, and in renal epithelial cell microvilli in vitro. These results not only demonstrate a novel role for PC-1 in the cochlea, but also suggest insight into the development of polycystic kidney disease.


Clinical evaluation of the efficacy and safety of a medical device in various forms containing Triticum vulgare for the treatment of venous leg ulcers - a randomized pilot study.
This study was carried out to assess the efficacy and tolerability of the topical application of an aqueous extract of Triticum vulgare (TV) in different vehicles (cream, impregnated gauzes, foam, hydrogel, and dressing gel) for the treatment of venous lower leg ulcers. Fifty patients were randomized to receive one of the five investigational vehicles. Treatment was performed up to complete healing or to a maximum of 29 days. The wound size reduction from baseline was the primary efficacy variable, which was measured by means of a noninvasive laser scanner instrument for wound assessment. In all groups, apart from the foam group, a similar trend toward the reduction of the surface area was observed. The cream showed the greatest effect on the mean reduction of the lesion size. At last visit, six ulcers were healed: two in the cream group, three in the gauze group, and one in the dressing gel group. In the patients treated with the cream, the gauzes, the hydrogel, and the dressing gel, the reduction of lesion size was 40%-50%; the reduction was smaller in the foam group. No impact in terms of age on the healing process was found. The Total Symptoms Score decreased in all groups during the study; a greater efficacy in terms of signs/symptoms was observed in the patients treated with the gauzes. In the dressing gel group, one patient had an infection of the wound after 3 weeks of treatment and 2 of colonization, leading to a systemic antibiotic treatment. The events were judged as nonrelated to the device used. On the basis of the results, it could be argued that the medical device may be useful in the treatment of chronic venous ulcers.


Real-time synthetic aperture sonar imaging using a parallel architecture.
This paper describes a parallel architecture that has been developed to perform real-time synthetic aperture sonar imaging as part of the Acoustical Imaging Development (ACID) project. The project has successfully developed a synthetic aperture sonar system for producing high resolution images of the sea floor and that has been tested during a series of sea trials in May 1993 off the south coast of France. This paper describes the synthetic aperture processing system developed by the University of Newcastle upon Tyne and its use of transputer modules and associated devices in order to obtain real-time imaging performance, the software structure of the processing system and the load balancing techniques that have been developed in order to provide efficient processing. The use of a parallel distributed architecture has also allowed a processing system that can readily be extended to deliver greater computational power in the future. Images produced by the synthetic aperture processor from data collected from around the Toulon coastal region are presented. These images highlight the improvement in azimuth resolution that can be obtained from synthetic aperture processing over conventional sidescan sonars.


Simian immunodeficiency virus SIVagm.sab infection of Caribbean African green monkeys: a new model for the study of SIV pathogenesis in natural hosts.
Caribbean-born African green monkeys (AGMs) were classified as Chlorocebus sabaeus by cytochrome b sequencing. Guided by these phylogenetic analyses, we developed a new model for the study of simian immunodeficiency virus (SIV) infection in natural hosts by inoculating Caribbean AGMs with their species-specific SIVagm.sab. SIVagm.sab replicated efficiently in Caribbean AGM peripheral blood mononuclear cells in vitro. During SIVagm.sab primary infection of six Caribbean AGMs, the virus replicated at high levels, with peak viral loads (VLs) of 10(7) to 10(8) copies/ml occurring by day 8 to 10 postinfection (p.i.). Set-point values of up to 2 x 10(5) copies/ml were reached by day 42 p.i. and maintained throughout follow-up (through day 450 p.i.). CD4(+) T-cell counts in the blood showed a transient depletion at the peak of VL, and then returned to near preinfection values by day 28 p.i. and remained relatively stable during the chronic infection. Preservation of CD4 T cells was also found in lymph nodes (LNs) of chronic SIVagm.sab-infected Caribbean AGMs. No activation of CD4(+) T cells was detected in the periphery in SIV-infected Caribbean AGMs. These virological and immunological profiles from peripheral blood and LNs were identical to those previously reported in African-born AGMs infected with the same viral strain (SIVagm.sab92018). Due to these similarities, we conclude that Caribbean AGMs are a useful alternative to AGMs of African origin as a model for the study of SIV infection in natural African hosts.