Baulamycins A and B, broad-spectrum antibiotics identified as inhibitors of siderophore biosynthesis in Staphylococcus aureus and Bacillus anthracis.
Siderophores are high-affinity iron chelators produced by microorganisms and frequently contribute to the virulence of human pathogens. Targeted inhibition of the biosynthesis of siderophores staphyloferrin B of Staphylococcus aureus and petrobactin of Bacillus anthracis hold considerable potential as a single or combined treatment for methicillin-resistant S. aureus (MRSA) and anthrax infection, respectively. The biosynthetic pathways for both siderophores involve a nonribosomal peptide synthetase independent siderophore (NIS) synthetase, including SbnE in staphyloferrin B and AsbA in petrobactin. In this study, we developed a biochemical assay specific for NIS synthetases to screen for inhibitors of SbnE and AsbA against a library of marine microbial-derived natural product extracts (NPEs). Analysis of the NPE derived from Streptomyces tempisquensis led to the isolation of the novel antibiotics baulamycins A (BmcA, 6) and B (BmcB, 7). BmcA and BmcB displayed in vitro activity with IC50 values of 4.8 μM and 19 μM against SbnE and 180 μM and 200 μM against AsbA, respectively. Kinetic analysis showed that the compounds function as reversible competitive enzyme inhibitors. Liquid culture studies with S. aureus , B. anthracis , E. coli , and several other bacterial pathogens demonstrated the capacity of these natural products to penetrate bacterial barriers and inhibit growth of both Gram-positive and Gram-negative species. These studies provide proof-of-concept that natural product inhibitors targeting siderophore virulence factors can provide access to novel broad-spectrum antibiotics, which may serve as important leads for the development of potent anti-infective agents.


Infrared drying of sweet potato (Ipomoea batatas L.) slices.
The effect of different infrared power levels (104, 125, 146 and 167 W) on drying kinetics and rehydration ratio of sweet potato slices was investigated. It was observed that the power levels affected the drying time and rehydration ratio. The increase in infrared power level decreased the drying time. The experimental data obtained from drying study were fitted with Newton, Henderson and Pabis and Logarithmic models to evaluate the drying kinetics of the sweet potato slices. The fit quality of the proposed models was evaluated by using the determination of coefficient (R (2) ), mean relative percent error (P), reduced chi-square (χ (2) ) and root means square error (RMSE). Among the three drying models, the Logarithmic model gave a better fit over the other two models. Effective diffusivity varied from 1.31 × 10(-10) to 3.66 × 10(-10) m(2)/s and was significantly influenced by infrared power.


Nonuniform Distribution of High-risk Human Papillomavirus in Squamous Cell Carcinomas of the Oropharynx: Rethinking the Anatomic Boundaries of Oral and Oropharyngeal Carcinoma From an Oncologic HPV Perspective.
The oral cavity and oropharynx have historically been viewed as a single anatomic compartment of the head and neck. The practice of combining the oral cavity and oropharynx has recently been revised, largely owing to the observation that human papillomavirus (HPV)-related carcinogenesis has a strong predilection for the oropharynx but not the oral cavity. The purpose of this study was to determine whether HPV is evenly distributed across squamous cell carcinomas of the oropharynx including those sites that do not harbor tonsillar tissues such as the soft palate. A search of the medical records of the Johns Hopkins Hospital identified 32 primary squamous cell carcinomas of the soft palate (n=31) and posterior pharyngeal wall (n=1). All were evaluated with p16 immunohistochemistry and high-risk HPV in situ hybridization (ISH) (29 by RNA ISH and 3 by DNA ISH). For comparison, we also reviewed the medical records to obtain the HPV status of patients who had undergone HPV testing of primary tonsillar carcinomas over the same time interval as part of their clinical care. High-risk HPV as detected by ISH was present in just 1 (3.1%) of the 32 oropharyngeal squamous cell carcinomas, including 1 of 2 p16-positive carcinomas. The difference in HPV detection rates between tonsillar and nontonsillar sites was significant (1/32, 3.1% vs. 917/997, 92%; P<0.0001). HPV is not frequently detected in squamous cell carcinomas arising from nontonsillar regions of the oropharynx. Indeed, squamous cell carcinomas of the soft palate more closely resemble those arising in the oral cavity than those arising in areas of the oropharynx harboring tonsillar tissue. This finding not only further sharpens our understanding of site-specific targeting by HPV, but may have practical implications regarding HPV testing and even the way the oral vault is oncologically compartmentalized to partition HPV-positive from HPV-negative cancers.


Multiwavelength Raman-fiber-laser-based long-distance remote sensor for simultaneous measurement of strain and temperature.
We propose a simple and flexible multiwavelength Raman-fiber-laser-based long-distance remote-sensing scheme for simultaneous measurement of strain and temperature by use of fiber Bragg gratings. By combining two uniform fiber Bragg gratings with a tunable chirped fiber grating, we readily achieve simultaneous two-channel sensing probes with a high extinction ratio of more than approximately 50 dB over a 50-km distance. When strain and temperature are applied, lasing wavelength separation and shift occur, respectively, since the two uniform fiber Bragg gratings have identical material composition and different cladding diameters. This allows simultaneous measurement of strain and temperature for long-distance sensing applications of more than 50 km.


Intrahepatic cholangiocarcinoma: rising frequency, improved survival, and determinants of outcome after resection.
Despite data suggesting a rising worldwide incidence, intrahepatic cholangiocarcinoma (IHC) remains an uncommon disease. This study analyzes changes in IHC frequency, demographics, and treatment outcome in a consecutive and single institutional cohort. Consecutive patients with confirmed IHC seen and treated over a 16-year period were included. The trend in IHC frequency over the study period was compared with that of hilar cholangiocarcinoma patients (HCCA) seen during the same time. Demographics and patient disposition, histopathologic, treatment, recurrence, and survival data were analyzed; changes in these variables over time were assessed. From December 1990 through July 2006, 594 patients were evaluated (IHC = 270, HCCA = 324). Over the study period, the average annual increase in new IHC patients was 14.2% (P < 0.001). Relative to HCCA, the proportional increase in IHC was nearly 3-fold, and new IHC patients have outnumbered those with HCCA by 2:1 over the last 3 years. Conditions associated with IHC were rarely seen, with only 7 patients having a history of sclerosing cholangitis and/or inflammatory bowel disease and none with hepatolithiasis or biliary parasitic disease; however, heavy tobacco use (27%) and diabetes mellitus (16.4%) were particularly prevalent. The majority of patients were not candidates for resection, most commonly because of advanced hepatic disease. After resection (n = 82), median disease-specific survival was 36 months; recurrence was observed in 62.2% of patients at a median follow-up of 26 months, with the liver remnant involved most frequently (62.7%). Multiple hepatic tumors (P < 0.001), regional nodal involvement (P = 0.012), and large tumor size (P = 0.016) independently predicted poor recurrence-free survival. Most patients (n = 115, 73.7%) with unresectable disease were treated with chemotherapy, either systemic alone (n = 75) or combined with regional hepatic arterial floxuridine (FUDR) (n = 28). Compared with the first 10 years of the study (1990-2000), the last 6 years saw an overall improvement in disease-specific survival for all patients (22 vs. 12 months, P = 0.002), which was particularly notable for patients with unresectable disease (15 vs. 6 months, P = 0.003). At Memorial Sloan-Kettering Cancer Center, IHC incidence has increased dramatically in the last 16 years. Resection offers the best opportunity for long-term survival but is possible in the minority, and patients with large, node-positive or multifocal IHC seem to derive little benefit. Establishing and maintaining control of the intrahepatic disease remains the biggest problem for all IHC patients. The recent increase in survival seems largely because of improved nonoperative therapy for unresectable disease.


Diabetes care in China: meeting the challenge.
In both human and economic terms, diabetes is becoming one of the most serious and costly health conditions worldwide. Economic development, bringing changes from a traditional to a modernized lifestyle, is driving a huge increase in the number of people with obesity-related type 2 diabetes in China. The extraordinary size of the problem is worrying; if current trends continue, diabetes will become a massive health burden in China. In this article, Changyu Pan looks at the status of diabetes care in China and highlights the need for regional and national initiatives to increase awareness amongst the general population of the risk factors for diabetes, and thus prevent a further rapid increase in the prevalence of the condition.


Granulomatous acne rosacea of the eyelids.
We describe the clinicopathologic features of the papular form of granulomatous acne rosacea of the eyelids. This unusual cutaneous disorder is typified by painless eruptions of reddish to yellowish brown, occasionally ulcerative papules symmetrically distributed around the eyelids, lower part of the forehead, nasolabial folds, and upper lip. Histopathologically, the lesions display caseating and noncaseating dermal granulomas that mimic those seen in tuberculosis, tuberculoid leprosy, sarcoidosis, and other diseases. The disorder responds well to systemic tetracycline therapy but may involute spontaneously during a period of months to years. The distinction of this disorder from other dermal granulomatous diseases, such as tuberculosis, leprosy, syphilis, and foreign bodies, is important because of the different therapeutic implications.


Blockage-induced condensation controlled by a local reaction.
We consider the setup of stationary zero range models and discuss the onset of condensation induced by a local blockage on the lattice. We show that the introduction of a local feedback on the hopping rates allows us to control the particle fraction in the condensed phase. This phenomenon results in a current versus blockage parameter curve characterized by two nonanalyticity points.


Clerkship directors' perceptions of the impact of HCFA documentation guidelines.
Chart notes are used to support billing codes under the evaluation and management guidelines of the Health Care Financing Administration (HCFA), in addition to serving as a record of the visit. To better understand the effect of the HCFA documentation guidelines, the authors collected data on how the guidelines affect participation by university- and community-based faculty in clinical education programs. In 2000, the authors sent six copies of their questionnaire to the associate deans of the 125 U.S. medical schools and requested they distribute them to all core clerkship directors. The questionnaire consisted of multiple-choice and short-answer questions regarding documentation of medical visits, participation of community-based faculty, understanding of HCFA documentation guidelines, and effects on education programs. The response rate was about 50%. Most of the 379 clerkship directors who responded (77%) stated they were aware the HCFA documentation guidelines include specifications regarding the role medical students can play and documentation of medical visits, and 64% indicated they were concerned the guidelines would affect their educational programs. Concerns included the loss of student independence and active participation in the patient care environment (37), time constraints and the changing balance between education and service (16), loss of faculty and decreased morale (11), and decreased quality of care for patients (7). Leaders of medical education must work to modify these guidelines to protect the quality of patients' care, while maximizing students' educational opportunity and participation.


Doses of primaquine administered to children with Plasmodium vivax according to an age-based dose regimen.
Primaquine is still the first-line drug to eliminate hypnozoites of Plasmodium vivax. The therapeutic efficacy is related to the total dose administered. In several endemic areas, the drug is administered for children in an age-based regimen, which can lead to inadequate exposure, increasing the rates of recurrence of the infection. The present study aims to describe the mg/kg total dose of primaquine administered to children for treatment for vivax malaria when an age-based regimen is used and to measure the plasma concentrations of primaquine and carboxyprimaquine. A total of 85 children were included in the study. The total dose of primaquine administered based on mg/kg had a median value of 3.22 mg/kg. The percentage of patients with a total dose below the required dose of 3.5 mg/kg was 55.75%. The median primaquine maximum concentration was 94 ng/ml. For carboxy-primaquine, the median maximum concentration was 375 ng/ml. The results suggest that age-based dosing regimens likely lead to substantial under-dosing of primaquine, which is evident in the youngest children and is reflected in decreased levels of primaquine and carboxy-primaquine in plasma samples 13.


Fruit and vegetable intake in a sample of 11-year-old children in 9 European countries: The Pro Children Cross-sectional Survey.
An adequate fruit and vegetable intake provides essential nutrients and nutritive compounds and is considered an important part of a healthy lifestyle. No simple instrument has been available for the assessment of fruit and vegetable intake as well as its determinants in school-aged children applicable in different European countries. Within the Pro Children Project, such an instrument has been developed. This paper describes the cross-sectional survey in 11-year-olds in 9 countries. The cross-sectional survey used nationally, and in 2 countries regionally, representative samples of schools and classes. The questionnaires, including a precoded 24-hour recall component and a food frequency part, were completed in the classroom. Data were treated using common syntax files for portion sizes and for merging of vegetable types into four subgroups. The results show that the fruit and vegetable intake in amounts and choice were highly diverse in the 9 participating countries. Vegetable intake was in general lower than fruit intake, boys consumed less fruit and vegetables than girls did. The highest total intake according to the 24-hour recall was found in Austria and Portugal, the lowest in Spain and Iceland. The fruit and vegetable intake in 11-year-old children was in all countries far from reaching population goals and food-based dietary guidelines on national and international levels.


Genetic and environmental exposures constrain epigenetic drift over the human life course.
Epigenetic mechanisms such as DNA methylation (DNAm) are essential for regulation of gene expression. DNAm is dynamic, influenced by both environmental and genetic factors. Epigenetic drift is the divergence of the epigenome as a function of age due to stochastic changes in methylation. Here we show that epigenetic drift may be constrained at many CpGs across the human genome by DNA sequence variation and by lifetime environmental exposures. We estimate repeatability of DNAm at 234,811 autosomal CpGs in whole blood using longitudinal data (2-3 repeated measurements) on 478 older people from two Scottish birth cohorts--the Lothian Birth Cohorts of 1921 and 1936. Median age was 79 yr and 70 yr, and the follow-up period was ∼10 yr and ∼6 yr, respectively. We compare this to methylation heritability estimated in the Brisbane Systems Genomics Study, a cross-sectional study of 117 families (offspring median age 13 yr; parent median age 46 yr). CpG repeatability in older people was highly correlated (0.68) with heritability estimated in younger people. Highly heritable sites had strong underlying cis-genetic effects. Thirty-seven and 1687 autosomal CpGs were associated with smoking and sex, respectively. Both sets were strongly enriched for high repeatability. Sex-associated CpGs were also strongly enriched for high heritability. Our results show that a large number of CpGs across the genome, as a result of environmental and/or genetic constraints, have stable DNAm variation over the human lifetime. Moreover, at a number of CpGs, most variation in the population is due to genetic factors, despite some sites being highly modifiable by the environment.


Suppression of osteoclastogenesis by N,N-dimethyl-D-erythro-sphingosine: a sphingosine kinase inhibition-independent action.
N,N-Dimethyl-D-erythro-sphingosine (DMS) competitively inhibits sphingosine kinase (SPHK) and has been widely used to assess the role of SPHK during cellular events, including motility, proliferation, and differentiation. In the present study, the effect of DMS on the differentiation of bone marrow macrophages (BMMs) to osteoclasts was investigated. When the osteoclast precursor cells were treated with DMS, the receptor activator of nuclear factor kappaB ligand (RANKL)-induced osteoclastogenesis was completely blocked. We were surprised to find, however, that knock-down of SPHK by small interfering RNA (siRNA) in BMMs did not reduce osteoclastogenesis. Furthermore, both overexpression of SPHK and exogenous addition of sphingosine-1-phosphate, the product of SPHK activity, failed to overcome the antiosteoclastogenic effect of DMS. These results suggest that DMS inhibited osteoclastogenesis independently of SPHK. Subsequent characterization of the DMS-mediated suppression of osteoclastogenesis revealed that DMS did not affect RANKL-induced activation of JNK, p38, NF-kappaB, and Ca2+ oscillation. On the other hand, DMS strongly inhibited two separate signaling pathways, the RANKL-induced activation of ERK and Akt, which eventually converged on the transcription factors c-Fos and NFATc1. There was significant increase in the osteoclast formation in the presence of DMS when BMMs were overexpressed with c-Fos, suggesting that c-Fos was a critical downstream target of DMS for the inhibition of osteoclastogenesis. Taken together, our data demonstrate that DMS has an antiosteoclastogenic function independently of its SPHK inhibitory activity. Considering previously reported anticancer properties of DMS, our study may also propose that DMS is an ideal drug candidate for bone metastases, for which osteoclastic bone-resorption is crucial.


ASP-G: an ASP-based method for finding attractors in genetic regulatory networks.
Boolean network models are suitable to simulate GRNs in the absence of detailed kinetic information. However, reducing the biological reality implies making assumptions on how genes interact (interaction rules) and how their state is updated during the simulation (update scheme). The exact choice of the assumptions largely determines the outcome of the simulations. In most cases, however, the biologically correct assumptions are unknown. An ideal simulation thus implies testing different rules and schemes to determine those that best capture an observed biological phenomenon. This is not trivial because most current methods to simulate Boolean network models of GRNs and to compute their attractors impose specific assumptions that cannot be easily altered, as they are built into the system. To allow for a more flexible simulation framework, we developed ASP-G. We show the correctness of ASP-G in simulating Boolean network models and obtaining attractors under different assumptions by successfully recapitulating the detection of attractors of previously published studies. We also provide an example of how performing simulation of network models under different settings help determine the assumptions under which a certain conclusion holds. The main added value of ASP-G is in its modularity and declarativity, making it more flexible and less error-prone than traditional approaches. The declarative nature of ASP-G comes at the expense of being slower than the more dedicated systems but still achieves a good efficiency with respect to computational time. The source code of ASP-G is available at http://bioinformatics.intec.ugent.be/kmarchal/Supplementary_Information_Musthofa_2014/asp-g.zip.


Cuffless Single-Site Photoplethysmography for Blood Pressure Monitoring.
One in three adults worldwide has hypertension, which is associated with significant morbidity and mortality. Consequently, there is a global demand for continuous and non-invasive blood pressure (BP) measurements that are convenient, easy to use, and more accurate than the currently available methods for detecting hypertension. This could easily be achieved through the integration of single-site photoplethysmography (PPG) readings into wearable devices, although improved reliability and an understanding of BP estimation accuracy are essential. This review paper focuses on understanding the features of PPG associated with BP and examines the development of this technology over the 2010-2019 period in terms of validation, sample size, diversity of subjects, and datasets used. Challenges and opportunities to move single-site PPG forward are also discussed.


WNT4 acts downstream of BMP2 and functions via β-catenin signaling pathway to regulate human endometrial stromal cell differentiation.
Differentiation of endometrial stromal cells into decidual cells is a prerequisite for successful embryo implantation. Our previous studies in the mouse have shown that bone morphogenetic protein 2 (BMP2), a morphogen belonging to the TGFβ superfamily, is essential for this differentiation process. BMP2 is markedly induced in human primary endometrial stromal cells (HESCs) as they undergo differentiation in response to steroid hormones and cAMP. The present study was undertaken to identify the BMP2-mediated molecular pathways in primary cultures of HESCs during decidualization. Using gene expression profiling, we identified wingless-related murine mammary tumor virus integration site 4 (WNT4) as a target of BMP2 regulation during decidualization. Attenuation of WNT4 expression in HESCs by small interfering RNA administration greatly reduced BMP2-induced stromal differentiation. Additionally, adenovirus-mediated overexpression of WNT4 in HESCs markedly advanced the differentiation program, indicating that it is a key regulator of decidualization. The stimulatory effect of WNT4 was accompanied by the accumulation of active β-catenin in the nuclei of decidualizing stromal cells, indicating the involvement of the canonical WNT signaling pathway. Functional inhibition of WNT4/β-catenin pathway by Dickkopf-1, an inhibitor of the canonical WNT signaling, or small interfering RNA-mediated silencing of β-catenin expression, greatly reduced the BMP2- and WNT4-induced decidualization. Gene expression profiling revealed that Forkhead box protein O1, a forkhead family transcription factor and previously reported regulator of HESC differentiation, is a common downstream mediator of both BMP2 and WNT4 signaling. Taken together, these studies uncovered a linear pathway involving BMP2, WNT4/β-catenin, and Forkhead box protein O1 that operates in human endometrium to critically control decidualization.


Disposition of triprolidine in the male beagle dog.
Three male beagle dogs were given 2.5 mg/kg doses of [14C]triprolidine HCl monohydrate (2.09 mg/kg of the free base) by intravenous and oral routes, in a nonrandomized cross-over experiment. After either route of administration, approximately 75% of the dose was excreted in the urine, and the remainder was excreted in the feces. Triprolidine was extensively metabolized, with less than 1% of the parent drug recovered in the excreta after either route of administration. Three metabolites were isolated from excreta and identified, including the major metabolite (metabolite 1, 219C69), in which the toluene ring methyl group was oxidized to a carboxylic acid, a metabolite (metabolite 2) in which the pyrrolidine ring was opened with oxidation of the terminal carbon to a carboxylic acid (a gamma-aminobutyric acid), and a metabolite (metabolite 3) that was a pyrrolidinone derivative of 219C69. Other metabolites in urine and feces were present in amounts too small for quantitation or identification. Route of administration had little effect on the metabolic pattern of triprolidine. Thus, after oral administration of triprolidine, a mean of 49.1% of the dose was excreted as 219C69, 12.0% as metabolite 2, 3.4% as metabolite 3, and 0.6% as triprolidine, while after intravenous administration, a mean of 50.8% of the dose was excreted as 219C69, 11.1% as metabolite 2, 4.2% as metabolite 3, and 0.8% as triprolidine. Plasma contained triprolidine, 219C69, and metabolite 2, as well as other apparent metabolites that were present at levels too low for quantitation. Mean pharmacokinetic parameters calculated for triprolidine after intravenous dosing were: CL = 24.4 ml/min/kg, Vdss = 5.8 liters/kg, and Vc = 1.6 liters/kg.(ABSTRACT TRUNCATED AT 250 WORDS)


Interaction of testosterone-based compounds with dodecyl sulphate monolayers at the air-water interface.
A series of atomistic molecular dynamics simulations were performed for investigating the interactions between three different testosterone-based compounds (testosterone (T), testosterone propionate (TP) and testosterone enanthate (TE)) and sodium dodecyl sulphate (SDS) and ammonium dodecyl sulphate (ADS) monolayers, which vary only in the sodium or ammonium counterions used to neutralise the sulphate headgroup. These simulations were used to investigate how the structural and interfacial properties of the monolayer were affected by changing the number of drug molecules present per monolayer, and the chemical nature of the surfactant counterions and the testosterone-based compounds. Our results show that the structure of the interfacial water layer is affected by the change of the counterion but not the chemistry of the drug molecules. As a result of the difference in their chemical structure, the T, TP and TE drug molecules prefer different locations and orientations within the monolayers. Finally, we observed that the hydration of the drug molecules encapsulated within the ADS monolayers is significantly less than when they are encapsulated within the SDS monolayers. Understanding the role that the counterion and the chemistry of the drug molecules play in these systems provides us with a detailed description of the interactions that cause ADS micelles to encapsulate significantly less drug molecules than SDS micelles, which we have recently observed experimentally.


A proposed definition of remission from chronic pain, based on retrospective evaluation of 24-month outcomes with spinal cord stimulation.
Objective: In the treatment of chronic diseases, remission is commonly used as a meaningful treatment goal, synonymous with the absence of significant clinical signs and symptoms of a disease, but not representing a cure. The objective of this paper is to propose a definition for remission for use as an outcome to evaluate the long-term efficacy of therapies for chronic pain. Methods: Data from a randomized clinical trial (NCT01609972) testing the efficacy of spinal cord stimulation in low back and leg pain subjects was used to evaluate the association between pain and functional outcomes and identify the cut-off value to predict remission. Available data over 24-month assessment period included visual analog score (VAS), disability (Oswestry Disability Index [ODI]), patient and clinician global impression of change (PGIC and CGIC), and patient satisfaction. Cluster analysis, Pearson's correlation coefficients, sensitivity, and specificity analyses were used to evaluate its utility in predicting higher patient functionality and satisfaction. Results: Though the term remission has been used in the chronic pain field, a consistent definition has not been previously established. Based on the analysis of the clinical data, we propose that a sustained (≥6 months) pain score of ≤3.0 cm out of 10 cm on VAS be defined as remission. Applying this definition to the clinical trial data: subjects in remission at 24 months versus non-remitters were significantly more likely to be in the highest functional category of minimally disabled according to the ODI (31.5 vs. 8.2%, respectively, p = 0.001), and be 'very satisfied' (75.7 vs 22.6%, respectively, p < 0.001). Conclusions: The validity of the proposed definition of remission is supported by the persistence of remission in this study group, and its correspondence with patient satisfaction, and reduced disability. Further evaluation of the definition using clinical data from other long-term studies is needed.


Protective effects of ethanolic extract of Delphinium denudatum in a rat model of Parkinson's disease.
Parkinson's disease (PD) is one of the major neurodegenerative disorders, and oxidative stress has been implicated in playing an important role in the pathogenesis of the disease. In the present study, we investigated if Delphinium denudatum extract can slow down the neuronal injury in 6-hydroxydopamine (6-OHDA) rat model of Parkinsonism. Rats were treated with 200, 400 and 600 mg/kg body weight (b.w.) of D. denudatum extract for 3 weeks. On day 22, 2 microL of 6-OHDA (10 microg in 0.1% ascorbic acid-saline) or vehicle was infused into the right striatum of the animals. Three weeks after the 6-OHDA injections, the rats were killed for estimation of lipid peroxidation (LPO), reduced glutathione (GSH) content, superoxide dismutase (SOD) and catalase (CAT) activities, catecholamines, dopaminergic D2 receptor binding and tyrosine hydroxylase (TH) expression. Increased LPO and significant depletion of reduced GSH content in the substantia nigra resulting from the lesion were appreciably prevented with Delphinium treatment. Delphinium extract also dose-dependently attenuated the activities of SOD and CAT in striatum, which had been reduced significantly by lesioning. A significant decrease in the level of dopamine (DA) and its metabolites and an increase in the number of dopaminergic D2 receptors in striatum were observed after 6-OHDA injection, both parameters were significantly recovered with treatment of the extract. Finally, all these results were confirmed by an increase in expression of TH in the ipsilateral striatum of the lesioned groups following treatment with Delphinium extract. Thus, the study indicates that D. denudatum extract may be helpful in checking neuronal injury in Parkinsonism.


The impact of socioeconomic status on foodborne illness in high-income countries: a systematic review.
Foodborne illness is a major cause of morbidity and loss of productivity in developed nations. Although low socioeconomic status (SES) is generally associated with negative health outcomes, its impact on foodborne illness is poorly understood. We conducted a systematic review to examine the association between SES and laboratory-confirmed illness caused by eight important foodborne pathogens. We completed this systematic review using PubMed for all papers published between 1 January 1980 and 1 January 2013 that measured the association between foodborne illness and SES in highly developed countries and identified 16 studies covering four pathogens. The effect of SES varied across pathogens: the majority of identified studies for Campylobacter, salmonellosis, and E. coli infection showed an association between high SES and illness. The single study of listeriosis showed illness was associated with low SES. A reporting bias by SES could not be excluded. SES should be considered when targeting consumer-level public health interventions for foodborne pathogens.


The catechol-O-methyltransferase (COMT) Val158Met polymorphism moderates the effect of antenatal stress on childhood behavioural problems: longitudinal evidence across multiple ages.
The functional polymorphism Val158Met in the catechol-O-methyltransferase (COMT) gene was analysed to determine its association with maternal stress and childhood total difficulties. Data were collected at birth from a group of infants who were born small for gestational age and a group who were born at an appropriate size for gestational age and had been enrolled in the Auckland Birthweight Collaborative Study. Children were followed up at the ages of 1 year, 3 years 6 months, 7 years, and 11 years. At the age of 11 years, DNA samples were collected from 546 children (270 females, 276 males): 227 children born small for gestational age and 319 children born at an appropriate size for gestational age. The main independent variable was perceived maternal stress at birth and at 7 and 11 years of age, assessed using the total difficulties scale of the Strength and Difficulties Questionnaire. IQ was assessed at the age of 7 years. Met/Met homozygotes were at a significantly increased risk of behavioural and emotional problems at the ages of 7 (p=0.002) and 11 years (p=0.003), relative to either heterozygous or homozygous carriers of the Val158Met polymorphism, but only when they were exposed to maternal stress in utero. Met/Met homozygotes had, on average, IQ scores that were four points higher than those of Val/Val homozygotes (p=0.010). These findings emphasize the potential long-term consequences of prenatal stress for genetically susceptible individuals during neurodevelopment in utero. Our findings add to the general understanding of the aetiology and developmental nature of childhood emotional and behavioural problems.


Acute occupational pesticide-related illness in the US, 1998-1999: surveillance findings from the SENSOR-pesticides program.
Concern about the adverse public health and environmental effects of pesticide use is persistent. Recognizing the importance of surveillance for acute occupational pesticide-related illness, we report on surveillance for this condition across multiple states. Survey data collected between 1998 and 1999 were obtained from the seven states that conduct acute occupational pesticide-related illness surveillance as part of the Sentinel Event Notification System for Occupational Risks (SENSOR) program. Data were collected by these state programs in a standardized manner and analyzed. Acute occupational pesticide-related illness incidence rates for those employed in agriculture and those employed in non-agricultural industries were also calculated. Between 1998 and 1999, a total of 1,009 individuals with acute occupational pesticide-related illness were identified by states participating in the SENSOR-pesticides program. The mean age was 36 years, and incidence rates peaked among 20-24 year-old workers. The overall incidence rate was 1.17 per 100,000 full time equivalents (FTEs). The incidence rate among those employed in agriculture was higher (18.2/100,000 FTEs) compared to those employed in non-agricultural industries (0.53/100,000 FTEs). Most of the illnesses were of low severity (69.7%). Severity was moderate in 29.6% of the cases, and high in four cases (0.4%). Three fatalities were identified. Insecticides were responsible for 49% of all illnesses. Surveillance is an important tool to assess acute pesticide-related illness, and to identify associated risk factors. Our findings suggest that these illnesses continue to be an important occupational health problem, especially in agriculture. As such, greater efforts are needed to prevent acute occupational pesticide-related illness.


Paternal history of mental illness associated with posttraumatic stress disorder among veterans.
This study examined the association between parent and family reported history of non-PTSD mental illness (MI), PTSD specifically, and substance use problems, and participant clinical diagnosis of PTSD. Participants were drawn from the US Department of Veterans Affairs Mid-Atlantic Mental Illness Research, Education and Clinical Center (MIRECC) Post-Deployment Mental Health (PDMH) study (n = 3191), an ongoing multi-site cohort study of US Afghanistan and Iraq conflict era veterans. Participants who recalled a father history of PTSD had a 26-percentage point higher likelihood of meeting criteria for PTSD; while participants reporting any family history of PTSD had a 15-percentage point higher probability of endorsing symptoms consistent with PTSD. Mother history of substance use problems was associated with Veteran current PTSD, but results were sensitive to model specification. Current PTSD was not associated with family/parent history of non-PTSD mental illness, mother history of PTSD, or family/father history of substance use problems. Family history of PTSD may increase PTSD risk among veterans exposed to trauma, particularly when a father history is reported. Knowledge of family history could improve clinical decision-making for trauma-exposed individuals and allow for more effective targeting of programs and clinical services.


The pansigmoidoscope: one year's experience in a gastrointestinal diagnostic unit.
Since May 1976, the Olympus pansigmoidoscope has been available for routine use at the University of Oregon Health Sciences center. Two hundred sixty-five examinations were performed over the next year. The average distance examined was 49 cm. Time per examination ranged from 3 to 15 minutes, with an average of 8 minutes. Preparation consisted of one or two tap water enemas, except in known inflammatory bowel disease where no preparation was given. No patient received sedation and there were no complications. Small biopsy (2.8 mm), large biopsy (4.0 mm), "hot biopsy" and polypectomy were performed when indicated. The procedure was most helpful for the following indications: 1) differential diagnosis and follow-up of inflammatory bowel disease, 2) hematochezia, 3) evaluation of abnormal barium enema, 4) left-sided polypectomy, 5) diarrhea with normal barium enema, and 6) guaiac-positive stools. It was of no value in patients with abdominal pain with normal barium enema. Comparing the frequency of examinations this year with last year we found a 50% decrease in use of the rigid (25 cm) sigmoidoscope (538 to 270 exams) and a 98% decrease in use of the MB2 (100 cm) colonoscope (80 to 2 exams).


Prevalence and intensity of rhinoconjunctivitis in subjects with occupational asthma.
Subjects with occupational asthma may also report symptoms of rhinoconjunctivitis. The aims of this study were: 1) to assess the prevalence of rhinoconjunctivitis in association with occupational asthma, and the severity of rhinoconjunctivitis according to the type of agent (high (HMW) and low (LMW) molecular weight agents) causing occupational asthma; and 2) to evaluate the timing of occurrence of symptoms of rhinoconjunctivitis in relation to those of occupational asthma. A questionnaire on symptoms of rhinoconjunctivitis and its timing in relation to the development of chest symptoms was prospectively addressed to 143 subjects consecutively referred to an occupational asthma clinic. Objective testing through specific inhalation challenges confirmed the diagnosis of occupational asthma in 40 subjects. Symptoms of rhinitis were reported at some time by 37 of the 40 subjects (92%), and of conjunctivitis by 29 of the 40 subjects (72%). The prevalence of symptoms was not different for HMW and LMW agents, although rhinitis was more intense for HMW (19 out of 24 subjects with three or more of the following symptoms: runny nose, itchy nose, nasal blockage, and sneezing) than for LMW (5 out of 14 subjects) (p<0.01). There were significantly fewer subjects with occupational asthma due to LMW agents, with rhinitis appearing before asthma (p=0.03). Figures for conjunctivitis showed a similar trend, but did not reach statistical significance. In conclusion, symptoms of rhinoconjunctivitis are often associated with occupational asthma. Rhinitis is less pronounced in the case of low molecular weight agents, but more often appears before occupational asthma in the case of high molecular weight agents.


Dominant Role of Entropy in Stabilizing Sugar Isomerization Transition States within Hydrophobic Zeolite Pores.
Lewis acid sites in zeolites catalyze aqueous-phase sugar isomerization at higher turnover rates when confined within hydrophobic rather than within hydrophilic micropores; however, relative contributions of competitive water adsorption at active sites and preferential stabilization of isomerization transition states have remained unclear. Here, we employ a suite of experimental and theoretical techniques to elucidate the effects of coadsorbed water on glucose isomerization reaction coordinate free energy landscapes. Transmission IR spectra provide evidence that water forms extended hydrogen-bonding networks within hydrophilic but not hydrophobic micropores of Beta zeolites. Aqueous-phase glucose isomerization turnover rates measured on Ti-Beta zeolites transition from first-order to zero-order dependence on glucose thermodynamic activity, as Lewis acidic Ti sites transition from water-covered to glucose-covered, consistent with intermediates identified from modulation excitation spectroscopy during in situ attenuated total reflectance IR experiments. First-order and zero-order isomerization rate constants are systematically higher (by 3-12×, 368-383 K) when Ti sites are confined within hydrophobic micropores. Apparent activation enthalpies and entropies reveal that glucose and water competitive adsorption at Ti sites depend weakly on confining environment polarity, while Gibbs free energies of hydride-shift isomerization transition states are lower when confined within hydrophobic micropores. DFT calculations suggest that interactions between intraporous water and isomerization transition states increase effective transition state sizes through second-shell solvation spheres, reducing primary solvation sphere flexibility. These findings clarify the effects of hydrophobic pockets on the stability of coadsorbed water and isomerization transition states and suggest design strategies that modify micropore polarity to influence turnover rates in liquid water.


Changes in language services use by US pediatricians.
Access to appropriate language services is critical for ensuring patient safety and reducing the impact of language barriers. This study compared language services use by US pediatricians in 2004 and 2010 and examined variation in use in 2010 by pediatrician, practice, and state characteristics. We used data from 2 national surveys of pediatricians (2004: n = 698; 2010: n = 683). Analysis was limited to postresidency pediatricians with patients with limited English proficiency (LEP). Pediatricians reported use of ≥ 1 communication methods with LEP patients: bilingual family member, staff, physician, formal interpreter (professional, telephone), and primary-language written materials. Bivariate analyses examined 2004 to 2010 changes in methods used, and 2010 use by characteristics of pediatricians (age, sex, ethnicity), practices (type, location, patient demographics), and states (LEP population, Latino population growth, Medicaid/Children's Health Insurance Program language services reimbursement). Multivariate logistic regression was performed to determine adjusted odds of use of each method. Most pediatricians reported using family members to communicate with LEP patients and families, but there was a decrease from 2004 to 2010 (69.6%, 57.1%, P < .01). A higher percentage of pediatricians reported formal interpreter use (professional and/or telephone) in 2010 (55.8%) than in 2004 (49.7%, P < .05); the increase was primarily attributable to increased telephone interpreter use (28.2%, 37.8%, P < .01). Pediatricians in states with reimbursement had twice the odds of formal interpreter use versus those in nonreimbursing states (odds ratio 2.34; 95% confidence interval 1.24-4.40). US pediatricians' use of appropriate language services has only modestly improved since 2004. Expanding language services reimbursement may increase formal interpreter use.


Functional role of endothelial adhesion molecules in the early stages of brain metastasis.
Cellular adhesion molecules (CAMs), which are normally associated with leukocyte trafficking, have also been shown to play an essential role in tumor metastasis to non-CNS sites. However, the role played by CAMs in brain metastasis is largely unexplored. It is known that leukocyte recruitment to the brain is very atypical and that mechanisms of disease in peripheral tissues cannot be extrapolated to the brain. Here, we have established the spatiotemporal expression of 12 key CAMs in the initial phases of tumor seeding in 2 different models of brain metastasis. BALB/c or SCID mice were injected intracardially (10(5) cells/100 μL phosphate-buffered saline with either 4T1-GFP or MDA231BR-GFP cells, respectively (n = 4-6/group), and expression of the CAMs was determined by immunohistochemistry and immunofluorescence colocalisation. Endothelial expression of E-selectin, VCAM-1, ALCAM, ICAM-1, VLA-4, and β4 integrin was markedly increased early in tumor seeding. At the same time, the natural ligands to these adhesion molecules were highly expressed on the metastatic tumor cells both in vitro and in vivo. Two of these ligands showed particularly high tumor cell expression (ALCAM and VLA-4), and consequently their functional role in tumor seeding was determined. Antibody neutralization of either ALCAM or VLA-4 significantly reduced tumor seeding within the brain (>60% decrease in tumor number/mm(2) brain; P < .05-0.01). These findings suggest that ALCAM/ALCAM and VLA-4/VCAM-1 interactions play an important functional role in the early stages of metastasis seeding in the brain. Moreover, this work identifies a specific subset of ligand-receptor interactions that may yield new therapeutic and diagnostic targets for brain metastasis.


Rates of diagnostic transition and cognitive change at 18-month follow-up among 1,112 participants in the Australian Imaging, Biomarkers and Lifestyle Flagship Study of Ageing (AIBL).
The Australian Imaging, Biomarkers and Lifestyle (AIBL) Flagship Study of Ageing is a prospective study of 1,112 individuals (211 with Alzheimer's disease (AD), 133 with mild cognitive impairment (MCI), and 768 healthy controls (HCs)). Here we report diagnostic and cognitive findings at the first (18-month) follow-up of the cohort. The first aim was to compute rates of transition from HC to MCI, and MCI to AD. The second aim was to characterize the cognitive profiles of individuals who transitioned to a more severe disease stage compared with those who did not. Eighteen months after baseline, participants underwent comprehensive cognitive testing and diagnostic review, provided an 80 ml blood sample, and completed health and lifestyle questionnaires. A subgroup also underwent amyloid PET and MRI neuroimaging. The diagnostic status of 89.9% of the cohorts was determined (972 were reassessed, 28 had died, and 112 did not return for reassessment). The 18-month cohort comprised 692 HCs, 82 MCI cases, 197 AD patients, and one Parkinson's disease dementia case. The transition rate from HC to MCI was 2.5%, and cognitive decline in HCs who transitioned to MCI was greatest in memory and naming domains compared to HCs who remained stable. The transition rate from MCI to AD was 30.5%. There was a high retention rate after 18 months. Rates of transition from healthy aging to MCI, and MCI to AD, were consistent with established estimates. Follow-up of this cohort over longer periods will elucidate robust predictors of future cognitive decline.


Coalescent-based species delimitation approach uncovers high cryptic diversity in the cosmopolitan lichen-forming fungal genus Protoparmelia (Lecanorales, Ascomycota).
Species recognition in lichen-forming fungi has been a challenge because of unsettled species concepts, few taxonomically relevant traits, and limitations of traditionally used morphological and chemical characters for identifying closely related species. Here we analyze species diversity in the cosmopolitan genus Protoparmelia s.l. The ~25 described species in this group occur across diverse habitats from the boreal-arctic/alpine to the tropics, but their relationship to each other remains unexplored. In this study, we inferred the phylogeny of 18 species currently assigned to this genus based on 160 specimens and six markers: mtSSU, nuLSU, ITS, RPB1, MCM7, and TSR1. We assessed the circumscription of species-level lineages in Protoparmelia s. str. using two coalescent-based species delimitation methods--BP&P and spedeSTEM. Our results suggest the presence of a tropical and an extra-tropical lineage, and eleven previously unrecognized distinct species-level lineages in Protoparmelia s. str. Several cryptic lineages were discovered as compared to phenotype-based species delimitation. Many of the putative species are supported by geographic evidence.


Isoliquiritigenin inhibits the proliferation, migration and metastasis of Hep3B cells via suppressing cyclin D1 and PI3K/AKT pathway.
The overall survival rate of patients with hepatocellular carcinoma (HCC) has remained unchanged over the last several decades. Therefore, novel drugs and therapies are required for HCC treatment. Isoliquiritigenin (ISL), a natural flavonoid predominantly isolated from the traditional Chinese medicine Glycyrrhizae Radix (Licorice), has a high anticancer potential and broad application value in various cancers. Here, we aimed to investigate the anticancer role of ISL in the HCC cell line Hep3B. Functional analysis revealed that ISL inhibited the proliferation of Hep3B cells by causing G1/S cell cycle arrest in vitro. Meanwhile, the inhibitory effect of ISL on proliferation was also observed in vivo. Further analysis revealed that ISL could suppress the migration and metastasis of Hep3B cells in vitro and in vivo. Mechanistic analysis revealed that ISL inhibited cyclin D1 and up-regulated the proteins P21, P27 that negatively regulate the cell cycle. Furthermore, ISL induced apoptosis while inhibiting cell cycle transition. In addition, phosphatidylinositol 3'-kinase/protein kinase B (PI3K/AKT) signal pathway was suppressed by ISL treatment, and the epithelial marker E-cadherin was up-regulated when the mesenchymal markers Vimentin and N-cadherin were down-regulated. In brief, our findings suggest that ISL could be a promising agent for preventing HCC tumorigenesis and metastasis.


Antibiotic resistance in Salmonella isolates from imported chicken carcasses in Bhutan and from pig carcasses in Vietnam.
The antibiotic resistance in Salmonella isolates from 400 imported chicken carcasses in Bhutan and from 178 pig carcasses in Vietnam were analyzed on a random basis against 14 antimicrobial agents. Among the poultry samples tested, 13% were positive for Salmonella. Salmonella Enteritidis dominated with a prevalence of 80.7%, and 40 of the 42 isolates harbored two or more resistance determinants. For the 178 pigs investigated, 49.4% of the swabs and 34.8% of the lymph nodes were Salmonella positive. The most prevalent serotypes in lymph nodes were Salmonella Derby (50.0%) and Salmonella Typhimurium (27.4%). From the Salmonella isolates from pigs, only 6% were sensitive to the antimicrobial agents tested. The high resistance level of Salmonella isolates from pigs and chicken carcasses to different classes of antimicrobials should be emphasized and encourage a prudent use of these agents in animal farming, especially in pig production.


Aptamer-based immunosensor on the ZnO nanorods networks.
This paper presents the fabrication and characteristics of a new aptamer-based electrochemical immunosensor on the patterned zinc oxide nanorod networks (ZNNs) for detecting thrombin. Aptamers are single-stranded RNA or DNA sequence that binds to target materials with high specificity and affinity. An antibody-antigen-aptamer sandwich structure was employed to this immunosensor for detecting thrombin. First, hydrothermally grown ZNNs were patterned on the patterned 0.02 cm2 Au/Ti electrodes on a glass substrate by lift-off process. The high isoelectric point (IEP, approximately 9.5) of nanostructured ZnO makes it suitable for immobilizing proteins with low IEP. Then 5 microL of the 500 nM antibody was immobilized on the ZNNs electrode. 5 micro/L of the mixture of 1 microM aptamer labeled by ferrocene (Fc) and thrombin was dropped on the electrode for antibody-antigen binding. The peak oxidation currents of the immunosensors at various thrombin concentrations were measured by using cyclic voltammetry. The peak oxidation current was observed at 340 mV versus Ag/AgCl electrode, and the peak oxidation current increased linearly from 62.26 nA to 354.13 nA with the logarithmic concentration of thrombin in the range from 100 pM to 250 nM. Fabrication of an aptamer-based immunosensor for thrombin detection is a new attempt and the characteristics of the fabricated immunosensors showed that the fabricated aptamer-baded immunosensor worked electrochemically well and had a low detection limit (approximately 91.04 pM) and good selectivity.


Identification of Proteins and Genes Expressed by Methylophaga thiooxydans During Growth on Dimethylsulfide and Their Presence in Other Members of the Genus.
Dimethylsulfide is a volatile organic sulfur compound that provides the largest input of biogenic sulfur from the oceans to the atmosphere, and thence back to land, constituting an important link in the global sulfur cycle. Microorganisms degrading DMS affect fluxes of DMS in the environment, but the underlying metabolic pathways are still poorly understood. Methylophaga thiooxydans is a marine methylotrophic bacterium capable of growth on DMS as sole source of carbon and energy. Using proteomics and transcriptomics we identified genes expressed during growth on dimethylsulfide and methanol to refine our knowledge of the metabolic pathways that are involved in DMS and methanol degradation in this strain. Amongst the most highly expressed genes on DMS were the two methanethiol oxidases driving the oxidation of this reactive and toxic intermediate of DMS metabolism. Growth on DMS also increased expression of the enzymes of the tetrahydrofolate linked pathway of formaldehyde oxidation, in addition to the tetrahydromethanopterin linked pathway. Key enzymes of the inorganic sulfur oxidation pathway included flavocytochrome c sulfide dehydrogenase, sulfide quinone oxidoreductase, and persulfide dioxygenases. A sulP permease was also expressed during growth on DMS. Proteomics and transcriptomics also identified a number of highly expressed proteins and gene products whose function is currently not understood. As the identity of some enzymes of organic and inorganic sulfur metabolism previously detected in Methylophaga has not been characterized at the genetic level yet, highly expressed uncharacterized genes provide new targets for further biochemical and genetic analysis. A pan-genome analysis of six available Methylophaga genomes showed that only two of the six investigated strains, M. thiooxydans and M. sulfidovorans have the gene encoding methanethiol oxidase, suggesting that growth on methylated sulfur compounds of M. aminisulfidivorans is likely to involve different enzymes and metabolic intermediates. Hence, the pathways of DMS-utilization and subsequent C1 and sulfur oxidation are not conserved across Methylophaga isolates that degrade methylated sulfur compounds.


Aboriginal status is a prognostic factor for mortality among antiretroviral naïve HIV-positive individuals first initiating HAART.
Although the impact of Aboriginal status on HIV incidence, HIV disease progression, and access to treatment has been investigated previously, little is known about the relationship between Aboriginal ethnicity and outcomes associated with highly active antiretroviral therapy (HAART). We undertook the present analysis to determine if Aboriginal and non-Aboriginal persons respond differently to HAART by measuring HIV plasma viral load response, CD4 cell response and time to all-cause mortality. A population-based analysis of a cohort of antiretroviral therapy naïve HIV-positive Aboriginal men and women 18 years or older in British Columbia, Canada. Participants were antiretroviral therapy naïve, initiated triple combination therapy between August 1, 1996 and September 30, 1999. Participants had to complete a baseline questionnaire as well as have at least two follow-up CD4 and HIV plasma viral load measures. The primary endpoints were CD4 and HIV plasma viral load response and all cause mortality. Cox proportional hazards models were used to determine the association between Aboriginal status and CD4 cell response, HIV plasma viral load response and all-cause mortality while controlling for several confounder variables. A total of 622 participants met the study criteria. Aboriginal status was significantly associated with no AIDS diagnosis at baseline (p = 0.0296), having protease inhibitor in the first therapy (p = 0.0209), lower baseline HIV plasma viral load (p < 0.001), less experienced HIV physicians (P = 0.0133), history of IDU (p < 0.001), not completing high school (p = 0.0046), and an income of less than $10,000 per year (p = 0.0115). Cox proportional hazards models controlling for clinical characteristics found that Aboriginal status had an increased hazard of mortality (HR = 3.12, 95% CI: 1.77-5.48) but did not with HIV plasma viral load response (HR = 1.15, 95% CI: 0.89-1.48) or CD4 cell response (HR = 0.95, 95% CI: 0.73-1.23). Our study demonstrates that HIV-infected Aboriginal persons accessing HAART had similar HIV treatment response as non-Aboriginal persons but have a shorter survival. This study highlights the need for continued research on medical interventions and behavioural changes among HIV-infected Aboriginal and other marginalized populations.


Adipose tissue energy metabolism: altered gene expression profile of mice subjected to long-term caloric restriction.
We investigated the influences of short-term and lifespan-prolonging long-term caloric restriction (LCR) on gene expression in white adipose tissue (WAT). Over 11,000 genes were examined using high-density oligonucleotide microarrays in four groups of 10- to 11-month-old male C57Bl6 mice that were either fasted for 18 h before death (F), subjected to short-term caloric restriction for 23 days (SCR), or LCR for 9 months and compared with nonfasted control (CO) mice. Only a few transcripts of F and SCR were differentially expressed compared with CO mice. In contrast, 345 transcripts of 6,266 genes found to be expressed in WAT were altered significantly by LCR. The expression of several genes encoding proteins involved in energy metabolism was increased by LCR. Further, many of the shifts in gene expression after LCR are known to occur during adipocyte differentiation. Selected LCR-associated alterations of gene expression were supported by quantitative reverse transcriptase-polymerase chain reaction, histology, and histochemical examinations. Our data provide new insights on the metabolic state associated with aging retardation by LCR.


Automatic segmentation of choroidal thickness in optical coherence tomography.
The assessment of choroidal thickness from optical coherence tomography (OCT) images of the human choroid is an important clinical and research task, since it provides valuable information regarding the eye's normal anatomy and physiology, and changes associated with various eye diseases and the development of refractive error. Due to the time consuming and subjective nature of manual image analysis, there is a need for the development of reliable objective automated methods of image segmentation to derive choroidal thickness measures. However, the detection of the two boundaries which delineate the choroid is a complicated and challenging task, in particular the detection of the outer choroidal boundary, due to a number of issues including: (i) the vascular ocular tissue is non-uniform and rich in non-homogeneous features, and (ii) the boundary can have a low contrast. In this paper, an automatic segmentation technique based on graph-search theory is presented to segment the inner choroidal boundary (ICB) and the outer choroidal boundary (OCB) to obtain the choroid thickness profile from OCT images. Before the segmentation, the B-scan is pre-processed to enhance the two boundaries of interest and to minimize the artifacts produced by surrounding features. The algorithm to detect the ICB is based on a simple edge filter and a directional weighted map penalty, while the algorithm to detect the OCB is based on OCT image enhancement and a dual brightness probability gradient. The method was tested on a large data set of images from a pediatric (1083 B-scans) and an adult (90 B-scans) population, which were previously manually segmented by an experienced observer. The results demonstrate the proposed method provides robust detection of the boundaries of interest and is a useful tool to extract clinical data.


Tumor suppressor cylindromatosis acts as a negative regulator for Streptococcus pneumoniae-induced NFAT signaling.
Gram-positive bacterium Streptococcus pneumoniae is an important human pathogen that colonizes the upper respiratory tract and is also the major cause of morbidity and mortality worldwide. S. pneumoniae causes invasive diseases such as pneumonia, meningitis, and otitis media. Despite the importance of pneumococcal diseases, little is known about the molecular mechanisms by which S. pneumoniae-induced inflammation is regulated, especially the negative regulatory mechanisms. Here we show that S. pneumoniae activates nuclear factor of activated T cells (NFAT) signaling pathway and the subsequent up-regulation of inflammatory mediators via a key pneumococcal virulence factor, pneumolysin. We also demonstrate that S. pneumoniae activates NFAT transcription factor independently of Toll-like receptors 2 and 4. Moreover, S. pneumoniae induces NFAT activation via both Ca(2+)-calcineurin and transforming growth factor-beta-activated kinase 1 (TAK1)-mitogen-activated protein kinase kinase (MKK) 3/6-p38alpha/beta-dependent signaling pathways. Interestingly, we found for the first time that tumor suppressor cylindromatosis (CYLD) acts as a negative regulator for S. pneumoniae-induced NFAT signaling pathway via a deubiquitination-dependent mechanism. Finally, we showed that CYLD interacts with and deubiquitinates TAK1 to negatively regulate the activation of the downstream MKK3/6-p38alpha/beta pathway. Our studies thus bring new insights into the molecular pathogenesis of S. pneumoniae infections through the NFAT-dependent mechanism and further identify CYLD as a negative regulator for NFAT signaling, thereby opening up new therapeutic targets for these diseases.


Statistical Considerations When Assessing Short Latency Stretch Reflexes in the Human Soleus Muscle.
The aim was to investigate trial-by-trial response characteristics in the short-latency stretch reflex (SSR). Fourteen dorsiflexion stretches were applied to the ankle joint with a pre-contracted soleus muscle on two days. The magnitude and variability of trial-by-trial responses of the SSR were assessed. The SSR was log-normally distributed and variance heterogeneous between subjects. For some subjects, the magnitude and variance differed between days and stretches. As velocity increased, variance heterogeneity tended to decrease and response magnitude increased. The current study demonstrates the need to assess trial-by-trial response characteristics and not averaged curves. Moreover, it provides an analysis of SSR characteristics accounting for log-normally distributed and variance heterogeneous trial-by-trial responses.


Growth Characteristics and Thermodynamics of Syntrophic Acetate Oxidizers.
Syntrophic acetate oxidation (SAO) plays a pivotal role in biogas production processes when aceticlastic methanogens are inhibited. Despite the importance of SAO, the metabolic interactions and syntrophic growth of the organisms involved are still poorly understood. Therefore, we studied growth parameters and interactions within constructed defined cocultures comprising the methanogen Methanoculleus bourgensis and one, or several, of the syntrophic acetate oxidizers Syntrophaceticus schinkii, [ Clostridium] ultunense, and Tepidanaerobacter acetatoxydans and a novel, uncharacterized bacterium. Cultivation experiments in a design-of-experiment approach revealed positive effects on methane production rate of increased ammonium levels (up to 0.2 M), temperature (up to 45 °C), and acetate concentrations (0.15-0.30 M). Molecular analyses and thermodynamic calculations demonstrated close interlinkages between the microorganisms, with available energies of -10 kJ/mol for acetate oxidation and -20 kJ/mol for hydrogenotrophic methanogenesis. The estimated generation time varied between 3 and 20 days for all syntrophic microorganisms involved, and the acetate minimum threshold level was 0.40-0.45 mM. The rate of methanogenesis depended on the SAO bacteria present in the culture. These data are beneficial for interpretation of SAO prevalence and competiveness against aceticlastic methanogens in anaerobic environments.


Hearing aid effectiveness after aural rehabilitation: individual versus group trial results.
We designed this noninferiority randomized trial to determine (1) if group hearing aid fitting visits and group hearing aid follow-up visits were at least as effective as individual visits in terms of hearing-related function (measured with the Inner Effectiveness of Auditory Rehabilitation [EAR] scale) and adherence, and (2) if group visits were less costly over the 6 mo post-fitting period. For 644 participants randomized to receive their hearing aid visits in an individual or group format at the Department of Veterans Affairs Puget Sound Health Care System, 6 mo outcomes showed no significant differences in Inner EAR scores between individual and group fitting (p = 0.53) or follow-up (p = 0.11), or in the number of hours per day hearing aids were worn between individual and group fitting (p = 0.90) or follow-up (p = 0.24). We found significantly higher cost for individual compared with group fitting (p < 0.001) and follow-up (p < 0.001). We conclude that outcomes for group hearing aid fitting and hearing aid follow-up visits were not inferior to individual hearing aid fitting and follow-up visits, with group fitting and follow-up visits yielding cost savings.


Normal targeting of a tagged Kv1.5 channel acutely transfected into fresh adult cardiac myocytes by a biolistic method.
The transfection of cardiac myocytes is difficult, and so most of the data regarding the regulation of trafficking and targeting of cardiac ion channels have been obtained using heterologous expression systems. Here we apply the fast biolistic transfection procedure to adult cardiomyocytes to show that biolistically introduced exogenous voltage-gated potassium channel, Kv1.5, is functional and, like endogenous Kv1.5, localizes to the intercalated disc, where it is expressed at the surface of that structure. Transfection efficiency averages 28.2 +/- 5.7% of surviving myocytes at 24 h postbombardment. Ventricular myocytes transfected with a tagged Kv1.5 exhibit an increased sustained current component that is approximately 40% sensitive to 100 microM 4-aminopyridine and which is absent in myocytes transfected with a fluorescent protein-encoding construct alone. Kv1.5 deletion mutations known to reduce the surface expression of the channel in heterologous cells similarly reduce the surface expression in transfected ventricular myocytes, although targeting to the intercalated disc per se is generally unaffected by both NH(2)- and COOH-terminal deletion mutants. Expressed current levels in wild-type Kv1.5, Kv1.5DeltaSH3(1), Kv1.5DeltaN209, and Kv1.5DeltaN135 mutants were well correlated with apparent surface expression of the channel at the intercalated disc. Our results conclusively demonstrate functionality of channels present at the intercalated disc in native myocytes and identify determinants of trafficking and surface targeting in intact cells. Clearly, biolistic transfection of adult cardiac myocytes will be a valuable method to study the regulation of surface expression of channels in their native environment.


Rapid and selective detection of fatty acylated proteins using omega-alkynyl-fatty acids and click chemistry.
Progress in understanding the biology of protein fatty acylation has been impeded by the lack of rapid direct detection and identification methods. We first report that a synthetic omega-alkynyl-palmitate analog can be readily and specifically incorporated into GAPDH or mitochondrial 3-hydroxyl-3-methylglutaryl-CoA synthase in vitro and reacted with an azido-biotin probe or the fluorogenic probe 3-azido-7-hydroxycoumarin using click chemistry for rapid detection by Western blotting or flat bed fluorescence scanning. The acylated cysteine residues were confirmed by MS. Second, omega-alkynyl-palmitate is preferentially incorporated into transiently expressed H- or N-Ras proteins (but not nonpalmitoylated K-Ras), compared with omega-alkynyl-myristate or omega-alkynyl-stearate, via an alkali sensitive thioester bond. Third, omega-alkynyl-myristate is specifically incorporated into endogenous co- and posttranslationally myristoylated proteins. The competitive inhibitors 2-bromopalmitate and 2-hydroxymyristate prevented incorporation of omega-alkynyl-palmitate and omega-alkynyl-myristate into palmitoylated and myristoylated proteins, respectively. Labeling cells with omega-alkynyl-palmitate does not affect membrane association of N-Ras. Furthermore, the palmitoylation of endogenous proteins including H- and N-Ras could be easily detected using omega-alkynyl-palmitate as label in cultured HeLa, Jurkat, and COS-7 cells, and, promisingly, in mice. The omega-alkynyl-myristate and -palmitate analogs used with click chemistry and azido-probes will be invaluable to study protein acylation in vitro, in cells, and in vivo.


Light scattering induced by opposite microdomains in LiNbO3:Fe:Hf crystals.
We report on the light scattering phenomenon in annealed multidomain LiNbO3:Fe:Hf crystals. The scattering sources are found to be some fog-like "defects", which cause the polarization-dependent scattering of the light, and can be removed completely by the illumination of visible light. Based on these results and the etch patterns, these "defects" are suggested to be refractive index fluctuations induced by the space charges accumulated at the boundary of opposite microdomains. The influence of quick heating-up on the "defects" is also studied and the results firmly support our suggestion about the nature of the "defects". At last, the temporal curves of the transmitted intensity during the light scattering are explained. The mechanism for the opposite microdomain formation is also explained from the view of crystal growth.


HLA class I sensitization in islet transplant recipients: report from the Collaborative Islet Transplant Registry.
Pancreatic islet transplantation is a promising treatment option for patients severely affected with type 1 diabetes. This report from CITR presents pre- and posttransplant human leukocyte antigen (HLA) class I sensitization rates in islet-alone transplantation. Data came from 303 recipients transplanted with islet-alone between January 1999 and December 2008. HLA class I sensitization was determined by the presence of anti-HLA class I antibodies. Panel-reactive antibodies (PRA) from prior to islet infusion and at 6 months, and yearly posttransplant was correlated to measures of islet graft failure. The cumulative number of mismatched HLA alleles increased with each additional islet infusion from a median of 3 for one infusion to 9 for three infusions. Pretransplant PRA was not predictive of islet graft failure. However, development of PRA >20% posttransplant was associated with 3.6-fold (p < 0.001) increased hazard ratio for graft failure. Patients with complete graft loss who had discontinued immunosuppression had significantly higher rate of PRA ≥ 20% compared to those with functioning grafts who remained on immunosuppression. Exposure to repeat HLA class I mismatch at second or third islet infusions resulted in less frequent development of de novo HLA class I antibodies when compared to increased class I mismatch. The development of HLA class I antibodies while on immunosuppression is associated with subsequent islet graft failure. The risk of sensitization may be reduced by minimizing the number of islet donors used per recipient, and in the absence of donor-specific anti-HLA antibodies, repeating HLA class I mismatches with subsequent islet infusions.


The TNF family member 4-1BBL sustains inflammation by interacting with TLR signaling components during late-phase activation.
Activation of Toll-like receptor (TLR)-dependent signaling leads to the expression of genes encoding proinflammatory factors, such as tumor necrosis factor-α (TNF-α), and this proinflammatory gene expression is sustained for the duration of the inflammatory response. TLR4-mediated inflammation, which occurs in two phases, depends on the TNF family member 4-1BB ligand (4-1BBL) to sustain TNF-α production during late-phase signaling. We showed that Toll-interleukin-1 receptor (TIR) domain-containing adaptor protein (TIRAP) and the kinase IRAK2 interacted with 4-1BBL to mediate late-phase TLR4 signaling. Expression of 4-1bbl depended on early TLR4 signaling that also induced Tnf expression, and 4-1BBL translocated to the plasma membrane, where it interacted with TLR4 to mediate late-phase signaling. TLR4-4-1BBL-mediated signaling depended on TIRAP and IRAK2, as well as a complex consisting of the E3 ubiquitin ligase TRAF6 (TNF receptor-associated factor 6), the kinase TAK1 (transforming growth factor-β-activated kinase 1), and the adaptor protein TAB1 (TAK-binding protein 1). Inhibition of this late-phase pathway reduced the extent of TNF-α production by mouse macrophages exposed to the TLR4 ligand lipopolysaccharide (LPS) and ameliorated LPS-induced sepsis in mice. Together, these data suggest that TIRAP and IRAK2 are critical for the sustained inflammatory response that is mediated by late-phase signaling by the TLR-4-1BBL complex.


Depression and smoking characteristics among HIV-positive smokers in Russia: A cross-sectional study.
Globally, persons with HIV infection, depression and substance use disorders have a higher smoking prevalence and smoke more heavily than other populations. These associations have not been explored among Russian smokers with HIV infection and substance use disorders. The purpose of this study was to examine the relationship between the presence of depressive symptoms and smoking outcomes in an HIV-positive cohort of Russian smokers with a history of substance use disorders (alcohol and/or drug use disorders). We performed a cross-sectional secondary data analysis of a cohort of HIV-positive regular smokers with a history of substance use disorders recruited in St. Petersburg, Russia in 2012-2015. The primary outcome was heavy smoking, defined as smoking > 20 cigarettes per day. Nicotine dependence (moderate-very high) was a secondary outcome. The main independent variable was a high level of depressive symptoms in the past 7 days (defined as CES-D > = 24). We used multivariable logistic regression to examine associations between depressive symptoms and the outcomes, controlling for age, sex, education, income, running out of money for housing/food, injection drug use, and alcohol use measured by the AUDIT. Among 309 regular smokers, 79 participants (25.6%) had high levels of depressive symptoms, and 65 participants (21.0%) were heavy smokers. High levels of depressive symptoms were not significantly associated with heavy smoking (adjusted odds ratio [aOR] 1.50, 95% CI 0.78-2.89) or with moderate-very high levels of nicotine dependence (aOR 1.35, 95% CI 0.75-2.41). This study did not detect an association between depressive symptoms and smoking outcomes among HIV-positive regular smokers in Russia.


Experimental and theoretical studies of nanofluid thermal conductivity enhancement: a review.
Nanofluids, i.e., well-dispersed (metallic) nanoparticles at low- volume fractions in liquids, may enhance the mixture's thermal conductivity, knf, over the base-fluid values. Thus, they are potentially useful for advanced cooling of micro-systems. Focusing mainly on dilute suspensions of well-dispersed spherical nanoparticles in water or ethylene glycol, recent experimental observations, associated measurement techniques, and new theories as well as useful correlations have been reviewed.It is evident that key questions still linger concerning the best nanoparticle-and-liquid pairing and conditioning, reliable measurements of achievable knf values, and easy-to-use, physically sound computer models which fully describe the particle dynamics and heat transfer of nanofluids. At present, experimental data and measurement methods are lacking consistency. In fact, debates on whether the anomalous enhancement is real or not endure, as well as discussions on what are repeatable correlations between knf and temperature, nanoparticle size/shape, and aggregation state. Clearly, benchmark experiments are needed, using the same nanofluids subject to different measurement methods. Such outcomes would validate new, minimally intrusive techniques and verify the reproducibility of experimental results. Dynamic knf models, assuming non-interacting metallic nano-spheres, postulate an enhancement above the classical Maxwell theory and thereby provide potentially additional physical insight. Clearly, it will be necessary to consider not only one possible mechanism but combine several mechanisms and compare predictive results to new benchmark experimental data sets.


Augmenting antidepressants with deep transcranial magnetic stimulation (DTMS) in treatment-resistant major depression.
Deep transcranial magnetic stimulation (DTMS) has been shown to be efficacious and relatively safe for major depressive disorder (MDD). However, its clinical utility as an augmenting strategy for treatment-resistant depression (TRD) remains unexplored. In an open label trial, 17 outpatients with severe TRD received 4 weeks of daily high frequency DTMS over the left dorsolateral prefrontal cortex. Depressive and anxious symptoms, suicidality and quality of life (QOL) were measured at baseline (i.e., in the week prior to the start of the DTMS treatment) and at week 5 (i.e., in the week following the end of the DTMS treatment). Primary outcome measures were rates of response and remission at week 5 using an intention-to-treat approach. Response and remission rates at week 5 were 70.6 and 41.2%, respectively. Also, depression, anxiety, and suicidality ratings were significantly improved by week 5 (with Hedges' g estimates ranging from 0.6 to 1.72), as well as four of the five QOL domain scores (i.e., global, psychological, environmental and social). Finally, two patients dropped out of the study at week 1 because of significant scalp discomfort during stimulation. Our study suggests that DTMS, when used as an augmenting strategy for antidepressants in severe TRD, is efficacious, safe and relatively well tolerated. However, controlled studies with larger samples are needed to confirm and expand our preliminary findings.


Impact of COVID-19 confinement on eating behaviours across 16 European countries: The COVIDiet cross-national study.
We aimed to evaluate the changes in eating behaviours of the adult population across 16 European countries due to the COVID-19 confinement and to evaluate whether these changes were somehow related to the severity of the containment measures applied in each country. An anonymous online self-reported questionnaire on socio-demographic characteristics, validated 14-items Mediterranean diet (MedDiet) Adherence Screener (MEDAS) as a reference of a healthy diet, eating and lifestyle behaviours prior to and during the COVID-19 confinement was used to collect data. The study included an adult population residing in 16 European countries at the time of the survey. Aggregated Stringency Index (SI) score, based on data from the Oxford COVID-19 Government Response Tracker, was calculated for each country at the time the questionnaire was distributed (range: 0-100). A total of 36,185 participants completed the questionnaire (77.6% female, 75.2% with high educational level and 42.7% aged between 21 and 35 years). In comparison to pre-confinement, a significantly higher adherence to the MedDiet during the confinement was observed across all countries (overall MEDAS score prior to- and during confinement: 5.23 ± 2.06 vs. 6.15 ± 2.06; p < 0.001), with the largest increase seen in Greece and North Macedonia. The highest adherence to MedDiet during confinement was found in Spain and Portugal (7.18 ± 1.84 and 7.34 ± 1.95, respectively). Stricter contingency restrictions seemed to lead to a significantly higher increase in the adherence to the MedDiet. The findings from this cross-sectional study could be used to inform current diet-related public health guidelines to ensure optimal nutrition is followed among the population, which in turn would help to alleviate the current public health crisis.


Optimized laser-induced breakdown spectroscopy for determination of xenobiotic silver in monosodium glutamate and its verification using ICP-AES.
Laser-induced breakdown spectroscopy (LIBS) was applied as a potential tool for the determination of xenobiotic metal in monosodium glutamate (MSG). In order to achieve a high-sensitivity LIBS system required to determine trace amounts of metallic silver in MSG and to attain the best detection limit, the parameters used in our experiment (impact of focusing laser energy on the intensity of LIBS emission signals, the influence of focusing lens distance on the intensity of LIBS signals, and time responses of the plasma emissions) were optimized. The spectra of MSG were obtained in air using a suitable detector with an optical resolution of 0.06 nm, covering a spectral region from 220 to 720 nm. Along with the detection of xenobiotic silver, other elements such as Ca, Mg, S, and Na were also detected in MSG. To determine the concentration of xenobiotic silver in MSG, the calibration curve was plotted by preparing standard samples having different silver abundances in an MSG matrix. The LIBS results of each sample were cross-verified by analyzing with a standard analytical technique such as inductively coupled plasma-atomic emission spectroscopy (ICP-AES). Both (LIBS and ICP-AES) results were in mutual agreement. The limit of detection of the LIBS setup was found to be 0.57 ppm for silver present in MSG samples.


In vitro assembly of PB2 with a PB1-PA dimer supports a new model of assembly of influenza A virus polymerase subunits into a functional trimeric complex.
Influenza virus RNA-dependent RNA polymerase is a heterotrimeric complex of PB1, PB2, and PA. We show that the individually expressed PB2 subunit can be assembled with the coexpressed PB1-PA dimer in vitro into a transcriptionally active complex. Furthermore, we demonstrate that a model viral RNA promoter can bind to the PB1-PA dimer prior to assembly with PB2. Our results are consistent with a recently proposed model for the sequential assembly of viral RNA polymerase complex in which the PB1-PA dimeric complex and the PB2 monomer are transported into the nucleus separately and then assembled in the nucleus.


PIK3CA mutations in Peruvian patients with HER2-amplified and triple negative non-metastatic breast cancers.
To determine the frequency of PIK3CA mutations in a Peruvian cohort with HER2-amplified and triple negative breast cancers (TNBC). We analyzed two cohorts of 134 primary non-metastatic breast cancer patients from Peru. Cohorts consisted of 51 hormone receptors (+)/HER2-amplified breast tumor patients surgically resected as first treatment included in the ALTTO trial (ALTTO cohort) and 81 TNBC patients with residual disease after neoadjuvant treatment (neoadjuvant cohort). Genomic DNA was extracted from paraffin-embedded tumor samples. Samples from the ALTTO and neoadjuvant cohorts were taken at biopsies and from residual tumors, respectively. PIK3CA mutations were detected by sequencing DNA fragments obtained by PCR amplification of exons and their flanking introns. All of the detected PIK3CA mutations were confirmed in a second independent run of sample testing. PIK3CA mutations were present in 21/134 cases (15.7%). Mutations in exon 9 and 20 were present in 10/134 (7.5%) and 11/134 (8.2%), respectively. No cases had mutations in both exons. Mutations in exon 9 consisted of E545A (seven cases), E545K (two cases) and E545Q (one case); while in exon 20, mutations consisted of H1047R (10 cases) and H1047L (one case). Compared to TNBC patients, HER2-amplified patients were more likely to have PIK3CA mutated (23% vs 9.6%; P=0.034). There were no associations between mutational status of PIK3CA with estrogen receptor status (P=0.731), progesterone receptor status (P=0.921), age (P=0.646), nodal status (P=0.240) or histological grade (P=1.00). No significant associations were found between PIK3CA mutational status and clinicopathological features. We found a similar frequency of PIK3CA mutations to that reported in other series. Although we did not include HR+/HER2 patients, those with HER2-amplified tumors were more likely to present PIK3CA mutations compared to patients with triple negative tumors.


Do microplastic loads reflect the population demographics along the southern African coastline?
Plastic pollution is a major anthropogenic contaminant effecting the marine environment and is often associated with high human population densities and industrial activities. The microplastic (63 to 5000μm) burden of beach sediment and surf-zone water was investigated at selected sites along the entire length of the South African coastline. It was predicted that samples collected in areas of high population density, would contain a higher microplastic burden than those along coasts that demonstrate very low population densities. With the exception of water column microplastics within Richard's Bay Harbour (413.3±77.53particles·m-3) and Durban Harbour (1200±133.2particles·m-3), there were no significant spatial differences in microplastic loads. This supports the theory that harbours act as a source of microplastics for the surrounding marine environment. Additionally, the absence of any spatial variation highlights the possible long range distribution of microplastic pollutants by large scale ocean currents.


Tyrosine phosphorylation of Pyk2 is selectively regulated by Fyn during TCR signaling.
The Src family protein tyrosine kinases (PTKs), Lck and Fyn, are coexpressed in T cells and perform crucial functions involved in the initiation of T cell antigen receptor (TCR) signal transduction. However, the mechanisms by which Lck and Fyn regulate TCR signaling are still not completely understood. One important question is whether Lck and Fyn have specific targets or only provide functional redundancy during TCR signaling. We have previously shown that Lck plays a major role in the tyrosine phosphorylation of the TCR-zeta chain and the ZAP-70 PTK. In an effort to identify the targets that are specifically regulated by Fyn, we have studied the tyrosine phosphorylation of Pyk2, a recently discovered new member of the focal adhesion kinase family PTK. We demonstrated that Pyk2 was rapidly tyrosine phosphorylated following TCR stimulation. TCR-induced tyrosine phosphorylation of Pyk2 was selectively dependent on Fyn but not Lck. Moreover, in heterologous COS-7 cells, coexpression of Pyk2 with Fyn but not Lck resulted in substantial increases in Pyk2 tyrosine phosphorylation. The selective regulation of Pyk2 tyrosine phosphorylation by Fyn in vivo correlated with the preferential phosphorylation of Pyk2 by Fyn in vitro. Our results demonstrate that Pyk2 is a specific target regulated by Fyn during TCR signaling.


Further tests of a recombination model in which chi removes the RecD subunit from the RecBCD enzyme of Escherichia coli.
When one of two infecting lambda phage types in a replication-blocked cross is chi + and DNA packaging is divorced from the RecBCD-chi interaction, complementary chi-stimulated recombinants are recovered equally in mass lysates only if the chi + parent is in excess in the infecting parental mixture. Otherwise, the chi 0 recombinant is recovered in excess. This observation implies that, along with the chi 0 chromosome, two chi + parent chromosomes are involved in the formation of each chi + recombinant. The trimolecular nature of chi +-stimulated recombination is manifest in recombination between lambda and a plasmid. When lambda recombines with a plasmid via the RecBCD pathway, the resulting chromosome has an enhanced probability of undergoing lambda x lambda recombination in the interval into which the plasmid was incorporated. These two observations support a model in which DNA is degraded by Exo V from cos, the sequence that determines the end of packaged lambda DNA and acts as point of entry for RecBCD enzyme, to chi, the DNA sequence that stimulates the RecBCD enzyme to effect recombination. The model supposes that chi acts by ejecting the RecD subunit from the RecBCD enzyme with two consequences. (1) ExoV activity is blocked leaving a highly recombinagenic, frayed duplex end near chi, and (2) as the enzyme stripped of the RecD subunit travels beyond chi it is competent to catalyze reciprocal recombination.


Loss of Runx2 in committed osteoblasts impairs postnatal skeletogenesis.
The Runx2 transcription factor is critical for commitment to the osteoblast lineage. However, its role in committed osteoblasts and its functions during postnatal skeletogenesis remain unclear. We established a Runx2-floxed line with insertion of loxP sites around exon 8 of the Runx2 gene. The Runx2 protein lacking the region encoded by exon 8 is imported into the nucleus and binds target DNA but exhibits diminished transcriptional activity. We specifically deleted the Runx2 gene in committed osteoblasts using 2.3-kb col1a-Cre transgenic mice. Surprisingly, the homozygous Runx2 mutant mice were born alive. The Runx2 heterozygous and homozygous null were grossly indistinguishable from wild-type littermates at birth. Runx2 deficiency did not alter proliferative capacity of osteoblasts during embryonic development (E18). Chondrocyte differentiation and cartilage growth in mutants was similar to wild-type mice from birth to 3 months of age. Analysis of the embryonic skeleton revealed poor calcification in homozygous mutants, which was more evident in bones formed by intramembranous ossification. Runx2 mutants showed progressive retardation in postnatal growth and exhibited significantly low bone mass by 1 month of age. Decreased bone formation was associated with decreased gene expression of osteoblast markers and impaired collagen assembly in the extracellular matrix. Consequently, Runx2 mutant bones exhibited decreased stiffness and structural integrity. By 3 months of age, bone acquisition in mutant mice was roughly half that of wild-type littermates. In addition to impaired osteoblast function, mutant mice showed markedly decreased osteoclast number and postnatal bone resorption. Taken together, functional deficiency of Runx2 in osteoblasts does not result in failed embryonic skeletogenesis but disrupts postnatal bone formation.


Genomic and metagenomic analysis of microbes in a soil environment affected by the 2011 Great East Japan Earthquake tsunami.
The Great East Japan Earthquake of 2011 triggered large tsunami waves, which flooded broad areas of land along the Pacific coast of eastern Japan and changed the soil environment drastically. However, the microbial characteristics of tsunami-affected soil at the genomic level remain largely unknown. In this study, we isolated microbes from a soil sample using general low-nutrient and seawater-based media to investigate microbial characteristics in tsunami-affected soil. As expected, a greater proportion of strains isolated from the tsunami-affected soil than the unaffected soil grew in the seawater-based medium. Cultivable strains in both the general low-nutrient and seawater-based media were distributed in the genus Arthrobacter. Most importantly, whole-genome sequencing of four of the isolated Arthrobacter strains revealed independent losses of siderophore-synthesis genes from their genomes. Siderophores are low-molecular-weight, iron-chelating compounds that are secreted for iron uptake; thus, the loss of siderophore-synthesis genes indicates that these strains have adapted to environments with high-iron concentrations. Indeed, chemical analysis confirmed the investigated soil samples to be rich in iron, and culture experiments confirmed weak cultivability of some of these strains in iron-limited media. Furthermore, metagenomic analyses demonstrated over-representation of denitrification-related genes in the tsunami-affected soil sample, as well as the presence of pathogenic and marine-living genera and genes related to salt-tolerance. Collectively, the present results would provide an example of microbial characteristics of soil disturbed by the tsunami, which may give an insight into microbial adaptation to drastic environmental changes. Further analyses on microbial ecology after a tsunami are envisioned to develop a deeper understanding of the recovery processes of terrestrial microbial ecosystems.


Azopolymer photopatterning for directional control of angiogenesis.
Understanding cellular behavior in response to microenvironmental stimuli is central to tissue engineering. An increasing number of reports emphasize the high sensitivity of cells to the physical characteristics of the surrounding milieu and in particular, topographical cues. In this work, we investigated the influence of dynamic topographic signal presentation on sprout formation and the possibility to obtain a space-time control over sprouting directionality without growth factors, in order to investigate the contribution of just topography in the angiogenic process. To test our hypothesis, we employed a 3D angiogenesis assay based on the use of spheroids derived from human umbilical vein endothelial cells (HUVECs). We then modulated the in situ presentation of topographical cues during early-stage angiogenesis through real-time photopatterning of an azobenzene-containing polymer, poly (Disperse Red 1 methacrylate) (pDR1m). Pattern inscription on the polymer surface was made using the focused laser of a confocal microscope. We demonstrate that during early-stage angiogenesis, sprouts followed the pattern direction, while spheroid cores acquired a polarized shape. These findings confirmed that sprout directionality was influenced by the photo-inscribed pattern, probably through contact guidance of leader cells, thus validating the proposed platform as a valuable tool for understanding complex processes involved in cell-topography interactions in multicellular systems. The complex relationship between endothelial cells and the surrounding environment that leads to formation of a newly formed vascular network during tissue repair is currently unknown. We have developed an innovative in vitro platform to study these mechanisms in a space and time controlled fashion simulating what happens during regeneration. In particular, we combine a "smart" surface, namely a polymer film, with a three-dimensional living cell aggregate. The polymer is activated by light through which we can design a path to guide cells toward the formation of a new vessel. Our work lies at the intersection of stimuli-responsive biointerfaces and cell biology and may be particularly inspiring for those interested in designing biomaterial surface related to angiogenesis.


First-pass gastric mucosal metabolism of ethanol is negligible in the rat.
Ethanol metabolism by gastric alcohol dehydrogenase (ADH) is thought to be an important determinant of peripheral ethanol time-concentration curves (AUCs) in rats and humans. We quantitated this metabolism in rats by measuring the gastric absorption of oral ethanol (0.25 g/kg) and the gastric venous-arterial (V-A) difference of ethanol versus ethanol metabolites (acetate, acetaldehyde, and bicarbonate). Over 1 h, approximately 20% of the ethanol was absorbed from the stomach and 70% was emptied into the duodenum. The gastric V-A difference of ethanol metabolites was less than 4% of that of ethanol. Thus, gastric metabolism accounted for less than 1% (less than 4% of 20% absorbed) of the dose. This negligible metabolism was predictable from the low affinity of gastric ADH for ethanol. In contrast, gastric ADH has a high affinity for octanol, and 66% of this compound was metabolized during gastric absorption. Evidence supporting gastric metabolism of ethanol largely derives from the lower AUCs observed after oral than after intravenous administration; however, we observed increasingly higher AUCs with increasingly rapid portal vein infusions of identical ethanol doses. We conclude that gastric metabolism of ethanol is negligible in the rat, and differences in AUCs ascribed to gastric metabolism may reflect differences in ethanol absorption.