IGF-1 decreases collagen degradation in diabetic NOD mesangial cells: implications for diabetic nephropathy.
Nonobese diabetic (NOD) mice develop glomerulosclerosis shortly after the onset of diabetes. We showed that mesangial cells (MCs) from diabetic mice exhibited a stable phenotypic switch, consisting of both increased IGF-1 synthesis and proliferation (Elliot SJ, Striker LJ, Hattori M, Yang CW, He CJ, Peten EP, Striker GE: Mesangial cells from diabetic NOD mice constitutively secrete increased amounts of insulin-like growth factor-I. Endocrinology 133:1783-1788, 1993). Because the extracellular matrix (ECM) accumulation in diabetic glomerulosclerosis may be partly due to decreased degradation, we examined the effect of excess IGF-1 on collagen turnover and the activity of metalloproteinases (MMPs) and tissue inhibitors of metalloproteinase (TIMPs) in diabetic and nondiabetic NOD-MC. Total collagen degradation was reduced by 58 +/- 18% in diabetic NOD-MCs, which correlated with a constitutive decrease in MMP-2 activity and mRNA levels, and nearly undetectable MMP-9 activity and mRNA. TIMP levels were slightly decreased in diabetic NOD-MC. The addition of recombinant IGF-1 to nondiabetic NOD-MC resulted in a decrease in MMP-2 and TIMP activity. Furthermore, treatment of diabetic NOD-MC with a neutralizing antibody against IGF-1 increased the latent form, and restored the active form, of MMP-2. In conclusion, the excessive production of IGF-1 contributes to the altered ECM turnover in diabetic NOD-MC, largely through a reduction of MMP-2 activity. These data suggest that IGF-1 could be a major contributor to the development of diabetic glomerulosclerosis.


Profiling bacterial communities by MinION sequencing of ribosomal operons.
An approach utilizing the long-read capability of the Oxford Nanopore MinION to rapidly sequence bacterial ribosomal operons of complex natural communities was developed. Microbial fingerprinting employs domain-specific forward primers (16S rRNA subunit), reverse primers (23S rRNA subunit), and a high-fidelity Taq polymerase with proofreading capabilities. Amplicons contained both ribosomal subunits for broad-based phylogenetic assignment (~ 3900 bp of sequence), plus the intergenic spacer (ITS) region (~ 300 bp) for potential strain-specific identification. To test the approach, bacterial rRNA operons (~ 4200 bp) were amplified from six DNA samples employing a mixture of farm soil and bioreactor DNA in known concentrations. Each DNA sample mixture was barcoded, sequenced in quadruplicate (n = 24), on two separate 6-h runs using the MinION system (R7.3 flow cell; MAP005 and 006 chemistry). From nearly 90,000 MinION reads, roughly 33,000 forward and reverse sequences were obtained. This yielded over 10,000 2D sequences which were analyzed using a simplified data analysis pipeline based on NCBI Blast and assembly with Geneious software. The method could detect over 1000 operational taxonomic units in the sample sets in a quantitative manner. Global sequence coverage for the various rRNA operons ranged from 1 to 1951x. An iterative assembly scheme was developed to reconstruct those rRNA operons with > 35x coverage from a set of 30 operational taxonomic units (OTUs) among the Proteobacteria, Actinobacteria, Acidobacteria, Firmicutes, and Gemmatimonadetes. Phylogenetic analysis of the 16S rRNA and 23S rRNA genes from each operon demonstrated similar tree topologies with species/strain-level resolution. This sequencing method represents a cost-effective way to profile microbial communities. Because the MinION is small, portable, and runs on a laptop, the possibility of microbiota characterization in the field or on robotic platforms becomes realistic.


Allergenicity of bony and cartilaginous fish - molecular and immunological properties.
Allergy to bony fish is common and probably increasing world-wide. The major heat-stable pan-fish allergen, parvalbumin (PV), has been identified and characterized for numerous fish species. In contrast, there are very few reports of allergic reactions to cartilaginous fish despite widespread consumption. The molecular basis for this seemingly low clinical cross-reactivity between these two fish groups has not been elucidated. PV consists of two distinct protein lineages, α and β. The α-lineage of this protein is predominant in muscle tissue of cartilaginous fish (Chondrichthyes), while β-PV is abundant in muscle tissue of bony fish (Osteichthyes). The low incidence of allergic reactions to ingested rays and sharks is likely due to the lack of molecular similarity, resulting in reduced immunological cross-reactivity between the two PV lineages. Structurally and physiologically, both protein lineages are very similar; however, the amino acid homology is very low with 47-54%. Furthermore, PV from ancient fish species such as the coelacanth demonstrates 62% sequence homology to leopard shark α-PV and 70% to carp β-PV. This indicates the extent of conservation of the PV isoforms lineages across millennia. This review highlights prevalence data on fish allergy and sensitization to fish, and details the molecular diversity of the two protein lineages of the major fish allergen PV among different fish groups, emphasizing the immunological and clinical differences in allergenicity.


Development of outcome indicators for monitoring the quality of public mental health care.
The study attempted to develop a brief and integrated set of reliable and valid outcome measures that could be used by both consumers and providers to assess the quality of public mental health care. A model of outcomes in four domains-consumer satisfaction, functioning, quality of life, and clinical status-was developed from the literature and from the priorities expressed by members of an advisory group of stakeholders. Based largely on extant measures, a consumer survey and a case manager survey were then created to assess these domains. A total of 236 adult consumers of mental health services from six community mental health centers in Washington State were surveyed. The four-item case manager survey to rate consumers' clinical status was completed by 163 of the participants' case managers. Scores and ratings on the survey were analyzed using correlational analysis and principal components analysis to determine whether the data provided empirical support for the four-domain model. Principal components analysis demonstrated support for the four-domain model. Internal consistency of the outcome indicators was adequate, and their concurrent validity was partly supported. The described outcome measures provide a practical, empirically supported structure for monitoring and improving public mental health services.


Targeting imperfect vaccines against drug-resistance determinants: a strategy for countering the rise of drug resistance.
The growing prevalence of antimicrobial resistance in major pathogens is outpacing discovery of new antimicrobial classes. Vaccines mitigate the effect of antimicrobial resistance by reducing the need for treatment, but vaccines for many drug-resistant pathogens remain undiscovered or have limited efficacy, in part because some vaccines selectively favor pathogen strains that escape vaccine-induced immunity. A strain with even a modest advantage in vaccinated hosts can have high fitness in a population with high vaccine coverage, which can offset a strong selection pressure such as antimicrobial use that occurs in a small fraction of hosts. We propose a strategy to target vaccines against drug-resistant pathogens, by using resistance-conferring proteins as antigens in multicomponent vaccines. Resistance determinants may be weakly immunogenic, offering only modest specific protection against resistant strains. Therefore, we assess here how varying the specific efficacy of the vaccine against resistant strains would affect the proportion of drug-resistant vs. -sensitive strains population-wide for three pathogens--Streptococcus pneumoniae, Staphylococcus aureus, and influenza virus--in which drug resistance is a problem. Notably, if such vaccines confer even slightly higher protection (additional efficacy between 1% and 8%) against resistant variants than sensitive ones, they may be an effective tool in controlling the rise of resistant strains, given current levels of use for many antimicrobial agents. We show that the population-wide impact of such vaccines depends on the additional effect on resistant strains and on the overall effect (against all strains). Resistance-conferring accessory gene products or resistant alleles of essential genes could be valuable as components of vaccines even if their specific protective effect is weak.


The Many Faces of CD4+ T Cells: Immunological and Structural Characteristics.
As a major arm of the cellular immune response, CD4+ T cells are important in the control and clearance of infections. Primarily described as helpers, CD4+ T cells play an integral role in the development and activation of B cells and CD8+ T cells. CD4+ T cells are incredibly heterogeneous, and can be divided into six main lineages based on distinct profiles, namely T helper 1, 2, 17 and 22 (Th1, Th2, Th17, Th22), regulatory T cells (Treg) and T follicular helper cells (Tfh). Recent advances in structural biology have allowed for a detailed characterisation of the molecular mechanisms that drive CD4+ T cell recognition. In this review, we discuss the defining features of the main human CD4+ T cell lineages and their role in immunity, as well as their structural characteristics underlying their detection of pathogens.


In-line microfluidic integration of photonic crystal fibres as a highly sensitive refractometer.
Photonic crystal fibres appear to be an ideal platform for the realisation of novel optofluidic devices and sensors due to their waveguide nature and microstructured architecture. In this paper, we present the fabrication and characterisation of an in-line photonic crystal fibre microfluidic refractometer enabled by a C-shaped fibre. The C-shaped fibre spliced in-between the photonic crystal fibre and the single-mode fibre allows simultaneous in-line optical signal delivery and analyte fluid feeding. Through an arc discharge pre-treatment technique, we successfully achieve selective exploitation of only the central two channels of the photonic crystal fibre for microfluidic sensing. After constructing a Sagnac interferometer, a highly sensitive refractometer with a sensitivity of 8699 nm per RIU was achieved experimentally; this agrees very well with the theoretical value of 8675 nm per RIU. As a demonstration for label-free optical sensing application, the refractometer was used to measure the concentration of NaCl solution with a sensitivity of 15.08 nm/(1 wt%) and a detection limit of 2.3 × 10(-3) wt% (23 ppm).


Leukocyte trafficking between stromal compartments: lessons from rheumatoid arthritis.
The trafficking of leukocytes from their site of production in the bone marrow through the circulation and into peripheral tissues is a highly coordinated and tightly regulated process in healthy individuals. Lymphocytes are long-lived cells that visit many lymphoid and peripheral tissues over their lifetime and can even recirculate back to the bone marrow, whereas granulocytes and monocytes are not thought to recirculate so widely. Using rheumatoid arthritis (RA) as an example, this Review explores the migratory journey of leukocytes during the establishment and resolution of disease - from the blood, through the lymphoid tissues and into peripheral sites such as the lungs and the gut before their entry into the synovium. This Review explores our current understanding of differences in the molecular processes that regulate leukocyte trafficking at different phases of disease and in different stromal compartments, which could help to explain the disease heterogeneity seen in patients with RA. Expanding our knowledge of these processes will open new avenues in the clinical management of RA, paving the way for personalized medicine that is founded on the pathological molecular signature of each patient, which varies according to their phase of disease or disease subtype.


Spinal dural resection for oncological purposes: a systematic analysis of risks and outcomes in patients with malignant spinal tumors.
Oncological outcomes for many malignant primary spinal tumors and isolated spinal metastases have been shown to correlate with extent of resection. For tumors with dural involvement, some authors have described spinal dural resection at the time of tumor resection in the interest of improving oncological outcomes. The complication profile associated with resection of the spinal dura for oncological purposes, however, and the relative influence of resecting tumor-involved dura on progression-free survival are not well defined. The authors performed a systematic review of the literature and identified cases in which the spinal dura was resected for oncological purposes in the interest of better understanding the associated risks and outcomes of this technique. Electronic databases (PubMed/MEDLINE, Scopus) were systematically searched to identify studies that reported clinical and/or oncological outcomes of patients with malignant spinal neoplasms undergoing resection of tumor-involved dura at the time of surgical intervention. Ten articles describing 15 patients were included in the analysis. The most common tumor histologies were chordoma (3/15, 20%), giant cell tumor (3/15, 20%), epithelioid sarcoma (2/15, 13.3%), osteosarcoma (2/15, 13.3%), and metastasis (2/15, 13.3%). Procedure-related complications were reported in 40% of patients. A trend was seen toward an increased complication rate in redo (66.7%) versus index (16.7%) operations, but this trend did not reach statistical significance (p = 0.24). New, unexpected postoperative neurological deficits were seen in 3 patients (of 14 reporting, 21.4%). A single patient experienced a profound, unexpected neurological deterioration (paraparesis/paraplegia) after surgery, which reportedly improved considerably at latest follow-up. Tumor recurrence was seen in 3 cases (of 12 reporting, 25%) at a mean of 28.34 ± 21.1 months postoperatively. The overall mean radiographic follow-up period was 49.6 ± 36.5 months. Resection of the spinal dura for oncological purposes is rarely performed, although a limited number of reports and small series have demonstrated that it is feasible. Spinal dural resection is primarily performed in patients with isolated, primary spinal neoplasms with an intent to cure. The risk associated with spinal dura resection is nontrivial and the complication profile is significant. The influence of dural resection on oncological outcomes is not well defined, and further study is needed before definitive conclusions may be drawn regarding the oncological benefit of dural resection for any particular patient or pathology.


Rapid increase in mast cell numbers in canine central and peripheral airways.
In preliminary studies of antigen-induced airway inflammation, we noted an apparent increase in peribronchiolar mast cell number. Experiments were thus undertaken to investigate the nature of this migration of mast cells into the central and peripheral airway epithelium and to determine its time course. The tracheae and small airways of 10 anesthetized mongrel dogs were exposed via a bronchoscope to Ascaris suum antigen (Ag), fMet-Leu-Phe (fMLP), ovalbumin (OVA), and isotonic saline (SAL). In the central airways, all stimuli provoked a significant increase (P less than 0.05) in mast cell numbers at the base of the airway epithelium within 3 h. In the peripheral airways, only Ag aerosol stimulated a significant mast cell increase compared with unexposed tissue. In a second series of experiments, the trachea of seven dogs were exposed to 0.026, 0.26, and 2.6 micrograms of Ag. The tissue was collected at 1, 3, 6, and 10 h after exposure. In these experiments, there was a significant mast cell increase seen within 1 h but it was not dose dependent. By 6-10 h after exposure, mast cell counts were not significantly different from the unexposed condition, which is consistent with the idea that some of the cells either degranulated or migrated into the airway lumen. We conclude that mast cell migration is an acute response that can be demonstrated within 1 h of stimulation with Ag. The observation that nonimmunological stimuli may, in some cases, also stimulate mast cell movement affords the possibility that this process represents a generalized response to airway irritation.


The relative contribution of DNA methylation and genetic variants on protein biomarkers for human diseases.
Associations between epigenetic alterations and disease status have been identified for many diseases. However, there is no strong evidence that epigenetic alterations are directly causal for disease pathogenesis. In this study, we combined SNP and DNA methylation data with measurements of protein biomarkers for cancer, inflammation or cardiovascular disease, to investigate the relative contribution of genetic and epigenetic variation on biomarker levels. A total of 121 protein biomarkers were measured and analyzed in relation to DNA methylation at 470,000 genomic positions and to over 10 million SNPs. We performed epigenome-wide association study (EWAS) and genome-wide association study (GWAS) analyses, and integrated biomarker, DNA methylation and SNP data using between 698 and 1033 samples depending on data availability for the different analyses. We identified 124 and 45 loci (Bonferroni adjusted P < 0.05) with effect sizes up to 0.22 standard units' change per 1% change in DNA methylation levels and up to four standard units' change per copy of the effective allele in the EWAS and GWAS respectively. Most GWAS loci were cis-regulatory whereas most EWAS loci were located in trans. Eleven EWAS loci were associated with multiple biomarkers, including one in NLRC5 associated with CXCL11, CXCL9, IL-12, and IL-18 levels. All EWAS signals that overlapped with a GWAS locus were driven by underlying genetic variants and three EWAS signals were confounded by smoking. While some cis-regulatory SNPs for biomarkers appeared to have an effect also on DNA methylation levels, cis-regulatory SNPs for DNA methylation were not observed to affect biomarker levels. We present associations between protein biomarker and DNA methylation levels at numerous loci in the genome. The associations are likely to reflect the underlying pattern of genetic variants, specific environmental exposures, or represent secondary effects to the pathogenesis of disease.


Frequency and outcomes of submandibular gland obstruction following resection of squamous cell carcinomas in the vicinity of the submandibular duct.
The aim of this study was to report rates, associated factors and outcomes of submandibular gland obstruction following surgery for squamous cell carcinoma in the anterior floor of the mouth (FOM) or ventral tongue without a neck dissection where resection has involved (or has been in very close proximity to) the submandibular duct. A retrospective case note review was carried out for the period January 2007 to December 2011. Only patients who received primary local surgical resection for squamous cell carcinoma of the anterior FOM or ventral tongue were included. Fifty-two patients were analysed. Fifteen (29%) developed submandibular gland symptoms following surgery. Symptoms resolved spontaneously for 14 (93%) and 1 patient required the submandibular duct to be repositioned. Comparatively, 2 patients (13%) developed symptoms when the duct was repositioned during primary surgery. No patients had their submandibular gland removed. Resection of small tumours associated with the FOM and ventral tongue in proximity to the submandibular duct is associated with obstructive symptoms in about a quarter of patients; this is reduced by half when redirecting the duct. Symptoms are self-limiting and self-resolving. A prospective clinical trial comparing duct repositioning with subcapsular gland excision would help clarify potential benefits and best treatment modalities.


Characterization of sphenoid sinuses for Sudanese population using computed tomography.
The purpose of this study is to determine the anatomical features of the sphenoid sinus using computerized tomography (CT). 100 Sudanese subjects were investigated for CT sinuses; Characterization of the sphenoid sinus and seven horizontal and vertical measurements were evaluated. Onodi cell was found in 13 subjects, 10 of them were sellar and 3 were pre-sellar. Pneumatization was of the sellar type in 85 %, presellar was 15 %, and no subject was chonchal. The mean length of vertical lines from the center of sphenoid ostium to the roof and bottom were 10.6 ± 3.1 mm, 11.1 ± 3.7 mm respectively. When the sphenoid ostium was located superior to the lowest point of the sella, the line from the center of the sphenoid sinus ostium to the posterior wall of the sinus was 15.2 ± 4.2 mm and when was located inferior, the line was 26.3 ± 5.2 mm on average. The mean length from the lowest point of the sella to the anterior wall of sphenoid sinus was 16.8 ± 3.6 mm. The line from anterior wall to posterior wall of sphenoid sinus lining skull base was 10.9±3.2mm mm. The maximum depth was 25.2 ± 6.9 mm and the maximum width was 18.4 ± 5.9mm. The differences in the sphenoid sinus character take place between males and females. The study provides essential anatomical information for Sudanese subjects and its impact in the clinical surgical practice.


The politics of cognition: liberalism and the evolutionary origins of Victorian education.
In recent years the historical relationship between scientific experts and the state has received increasing scrutiny. Such experts played important roles in the creation and regulation of environmental organizations and functioned as agents dispatched by politicians or bureaucrats to assess health-related problems and concerns raised by the public or the judiciary. But when it came to making public policy, scientists played another role that has received less attention. In addition to acting as advisers and assessors, some scientists were democratically elected members of local and national legislatures. In this essay I draw attention to this phenomenon by examining how liberal politicians and intellectuals used Darwinian cognitive science to conceptualize the education of children in Victorian Britain.


Predictive Modeling of Covid-19 Data in the US: Adaptive Phase-Space Approach.
There are currently intensified efforts by the scientific community world-wide to analyze the dynamics of the Covid-19 pandemic in order to predict key epidemiological effects and assist the proper planning for its clinical management, as well as guide sociopolitical decision-making regarding proper mitigation measures. Most efforts follow variants of the established SIR methodological framework that divides a population into "Susceptible", "Infectious" and "Recovered/Removed" fractions and defines their dynamic inter-relationships with first-order differential equations. This paper proposes a novel approach based on data-guided detection and concatenation of infection waves - each of them described by a Riccati equation with adaptively estimated parameters. This approach was applied to Covid-19 daily time-series data of US confirmed cases, resulting in the decomposition of the epidemic time-course into five "Riccati modules" representing major infection waves to date (June 18th). Four waves have passed the time-point of peak infection rate, with the fifth expected to peak on July 20th. The obtained parameter estimates indicate gradual reduction of infectivity rate, although the latest wave is expected to be the largest. This analysis suggests that, if no new waves of infection emerge, the Covid-19 epidemic will be controlled in the US (<5000 new daily cases) by September 26th, and the maximum of confirmed cases will reach 4,160,000. Importantly, this approach can be used to detect (via rigorous statistical methods) the emergence of possible new waves of infections in the future. Analysis of data from individual states or countries may quantify the distinct effects of different mitigation measures.


Online post LPN to BN students' views of transitioning to a new nursing role.
Vocationally prepared nurses, most of whom are women, experience unique challenges when they enter an online university. And yet, educational research examining their perceptions about transitioning into a new professional role is limited. This qualitative descriptive study investigated beginning Post LPN to BN students' views of role differences among nurses and the process of transitioning between those roles. The project was framed from a constructivist theoretical perspective and applied repertory grid methodology from the field of personal construct psychology. Ten students from a Canadian online university were interviewed. Interview transcripts were analyzed for themes. Findings revealed that students appreciated recognition for their previous accomplishments. They valued affirmation of the unique challenges they faced. And, they associated gains from their new university education with some loss of their hands on bedside nursing role.


MIDAS2: Metagenomic Intra-species Diversity Analysis System.
The Metagenomic Intra-Species Diversity Analysis System (MIDAS) is a scalable metagenomic pipeline that identifies single nucleotide variants (SNVs) and gene copy number variants in microbial populations. Here, we present MIDAS2, which addresses the computational challenges presented by increasingly large reference genome databases, while adding functionality for building custom databases and leveraging paired-end reads to improve SNV accuracy. This fast and scalable reengineering of the MIDAS pipeline enables thousands of metagenomic samples to be efficiently genotyped. The source code is available at https://github.com/czbiohub/MIDAS2. Supplementary data are available at Bioinformatics online.


Age-Related Social Cognitive Performance in Individuals With Psychotic Disorders and Their First-Degree Relatives.
Social cognitive impairment is a recognized feature of psychotic disorders. However, potential age-related differences in social cognitive impairment have rarely been studied. Data came from 905 individuals with a psychotic disorder, 966 unaffected siblings, and 544 never-psychotic controls aged 18-55 who participated in the Genetic Risk and Outcome of Psychosis (GROUP) study. Multilevel linear models were fitted to study group main effects and the interaction between group and age on emotion perception and processing (EPP; degraded facial affect recognition) and theory of mind (ToM; hinting task) performance. Age-related differences in the association between socio-demographic and clinical factors, and EPP and ToM were also explored. Across groups, EPP performance was associated with age (β = -0.02, z = -7.60, 95% CI: -0.02, -0.01, P < .001), with older participants performing worse than younger ones. A significant group-by-age interaction on ToM (X2(2) = 13.15, P = .001) indicated that older patients performed better than younger ones, while no age-related difference in performance was apparent among siblings and controls. In patients, the association between negative symptoms and ToM was stronger for younger than older patients (z = 2.16, P = .03). The findings point to different age-related performance patterns on tests of 2 key social cognitive domains. ToM performance was better in older individuals, although this effect was only observed for patients. EPP was less accurate in older compared with younger individuals. These findings have implications with respect to when social cognitive training should be offered to patients.


Drinking alcohol during pregnancy: evidence from Canadian Community Health Survey 2007/2008.
Drinking alcohol during pregnancy may cause many health problems for the child, one of which is fetal alcohol spectrum disorder (FASD). Since FASD is incurable, actions meant to prevent the occurrence of the disability by targeting drinking women become more important. Epidemiological data on drinking among pregnant women, including prevalence and determinants/risk factors, is essential for designing and evaluating prevention programs. To estimate the prevalence of drinking alcohol during pregnancy and examine the determinants of this behaviour. Using the 2007/8 Canadian Community Health Survey (CCHS) data, we estimated the weighted prevalence of women who drank alcohol during their last pregnancy by provinces. We used a weighted logistic regression to examine associations between drinking patterns, substance abuse behaviours, health-related and socio-demographic characteristics of the women, and the outcome variable. There were two main findings of this study. One was that the 2007/8 prevalence of drinking alcohol during pregnancy in ON, BC, and Canada was estimated at 5.4%, 7.2%, and 5.8%, respectively. The other was that the use of general practitioners (GP) or family physicians (FP) associated with a decreased risk of drinking alcohol during pregnancy. The results suggest that interventions that involve GP or FP and that increase the use of GP or FP by pregnant women can be effective in reducing drinking alcohol during pregnancy.


Knockdown of the Sonic Hedgehog (SHH) Gene Inhibits Proliferation of Hep3B and SMMC-7721 Hepatocellular Carcinoma Cells via the PI3K/Akt/PCK1 Signaling Pathway.
BACKGROUND The PCK1 gene encodes phosphoenolpyruvate carboxykinase (PEPCK), which has been shown have a role in metabolic events in hepatocellular carcinoma (HCC). This study aimed to investigate the role of the SHH gene and its encoded protein, sonic hedgehog (SHH), in two human hepatocellular carcinoma (HCC) cell lines. MATERIAL AND METHODS The human HCC cell lines Hep3B and SMMC-7721 were cultured. Cells were transfected with plasmids carrying specific SHH gene short-hairpin RNA (shRNA) and negative control (NC) shRNA. The effects of knockdown of expression levels of the SHH gene were studied on cell survival, cell apoptosis, the cell cycle, gluconeogenesis, and the expression of PCK1. Anchorage-independent growth, a characteristic of transformed cells, was detected by the colony formation assay. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot were performed 24 h after transfection. RESULTS Knockdown of expression levels of the SHH gene reduced cell proliferation and growth of HCC cells and induced cell apoptosis and G1 cell cycle arrest in both HCC cell lines. Knockdown of the SHH gene decreased the levels of glycolysis products and increased the production of glucose and reduced the phosphorylation of PI3K and Akt but induced the expression of PCK1. CONCLUSIONS Knockdown of the SHH gene reduced cell survival of HCC cells by increasing apoptosis, reducing cell proliferation, inducing G1 cell cycle arrest, and restoring gluconeogenesis, and was associated with the inhibition of the PI3K/Akt axis and induced the expression of PCK1.


Right ventricular infarction and tissue Doppler imaging – insights from acute inferior myocardial infarction after primary coronary intervention –.
Tissue Doppler imaging (TDI) data for acute inferior myocardial infarction (MI) patients who have received primary percutaneous coronary intervention (PCI) are sparse. One hundred and sixty-five patients received primary PCI for acute inferior MI were enrolled. Right ventricular infarction (RVI) was defined as a culprit lesion proximal to the right ventricular branch of right coronary artery (RCA). Echocardiograms and TDI were obtained within 6 h after primary PCI. The prevalence of multi-vessel disease in the RCA-P culprit group (50%) was higher than that in other groups (39% of RCA-D culprit, 43% of left circumflex artery (LCX) culprit). The myocardial performance index (MPI) of the lateral tricuspid annulus provides discriminatory power for identifying RVI, whereas systolic velocity (Sm) of the lateral tricuspid annulus does not. Lateral mitral annular MPI divided by the lateral tricuspid annular MPI is a reliable index for identifying a culprit lesion (>1.06 predicts culprit over LCX; <0.96 predicts culprit over RCA-P and RVI). Kaplan-Meier survival curves revealed that late cardiovascular events were more likely in RVI patients. However, multivariate Cox proportional hazards analysis revealed that the most important factor in hard events and all cardiovascular events was multivessel disease. TDI is useful for identifying RVI and culprit lesions in inferior MI patients received primary PCI. RVI itself isn't associated with 1-year hard events and all cardiovascular events.


Mental Health Consequences of Childhood Physical Abuse in Chinese Populations: A Meta-Analysis.
Childhood physical abuse (CPA) can lead to adverse mental health outcomes in adulthood, but its potential impact on Chinese populations is still unclear. This meta-analysis is the first to examine the association between CPA and mental health outcomes in Chinese populations. Studies published before December 31, 2014 were identified from Embase, CINAHL, Scopus, PsycINFO, PubMed, and Chinese National Knowledge Infrastructure databases. Studies with data on the association between CPA and mental health outcomes from Chinese subjects were included. Twenty-four studies were initially identified but two were excluded because of poor quality. Two reviewers independently extracted data to generate summary effect sizes using a random-effects meta-analytic model. A priori subgroup and sensitivity analyses were performed to evaluate heterogeneity and bias in these studies. Our meta-analysis of 22 studies found a significant positive association between CPA and overall mental health outcomes among all Chinese subjects (pooled effect size: odds ratio [OR] = 2.16, 95% confidence interval [CI] = [1.87, 2.49]) and among community samples (pooled effect size: OR = 2.06, 95% CI [1.71, 2.48]). Based on the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition, Text Revision, diagnostic criteria, CPA was more strongly associated with Axis II (OR = 2.62, 95% CI [2.13, 3.22]) than Axis I disorders (OR = 1.85, 95% CI [1.58, 2.17]). The detrimental effects of CPA on mental health outcomes in Chinese populations were comparable to, if not more than, the West. Contrary to the Chinese belief that physical punishment is a safe way to discipline children, our findings highlight the potential harm to mental health and the need to change this parenting practice.


Class III β-tubulin expression as a predictor of docetaxel-resistance in metastatic castration-resistant prostate cancer.
About half of the patients treated with docetaxel in the setting of metastatic castration-resistant prostate cancer (CRPC) are non-responders. Therefore, a marker of response would be beneficial for clinical decision-making. We evaluated class III β-tubulin (βIII-tubulin) expression as a predictor of resistance in this setting, which previously has been correlated with lack of response to taxanes in other cancers. Patients with CRPC were included if they were treated with at least 3 cycles of docetaxel between 1990 and 2011. βIII-tubulin expression was assessed by immunostaining, which was performed in tissue samples obtained either via biopsy or prostatectomy at the time of diagnosis. Rates of prostate-specific antigen (PSA) response and overall survival (OS) following docetaxel treatment were compared between patients with high (2+ or 3+ staining) vs. low (0 or 1+ staining) βIII-tubulin expression. Of 73 patients, 26 (35%) had a high expression of βIII-tubulin. A PSA decline of 10% or greater occurred in 65% of patients with a high βIII-tubulin expression vs. 89% with a low βIII-tubulin expression (p = 0.0267). The median OS for patients with a high βIII-tubulin expression was 17.4 (95% CI 8.7-21.0) months vs. 19.8 (95% CI 16.6-23.6) months for patients with a low expression (p = 0.039). Our results show that a high βIII-tubulin expression is a negative prognostic factor in metastatic CRPC patients treated with docetaxel.


Whole body hyperthermia improves obesity-induced insulin resistance in diabetic mice.
In this study, we examined the efficacy of whole body hyperthermia (WBH) on obesity-induced insulin resistance in diabetic mice. Male db/db mice were treated with WBH 3 times per week for 12 weeks. The rectal temperature of mice reached 38.0 degrees C 5 min after heating, and was kept at 38.0 degrees C for 30 min. At the end of each week, tail snip glucose levels were determined under fasting conditions. The GLUT-4 gene expression of muscle tissue was analyzed by real-time PCR. (1) WBH-treated db/db mice showed a significant decrease in fasting blood glucose level as compared with untreated db/db mice (p < 0.01). (2) Plasma insulin levels in untreated db/db mice at the age of 10 weeks were significantly increased compared with those of db/+ mice (p < 0.0001). On the other hand, the reduction (31%) in insulin levels in WBH-treated mice indicated improved insulin sensitivity. (3) The ability of WBH to increase insulin sensitivity was further established in glucose tolerance tests and insulin tolerance tests. (4) Urine albumin of db/db mice significantly increased compared with those of db/+ mice at 18 weeks of age (p < 0.001). This increase in urinary albumin was significantly inhibited by WBH (p < 0.01). (5) WBH up-regulated the expression of GLUT4 mRNA in skeletal muscle. Although the mechanisms have not yet been completely investigated, WBH may provide a new therapeutic or preventive modality against obesity-related diseases such as T2DM and metabolic or insulin resistance syndrome.


Storage of ultrashort optical pulses in a resonantly absorbing Bragg reflector.
A practical method of slowing and stopping an incident ultrashort light pulse with a resonantly absorbing Bragg reflector is demonstrated numerically. It is shown that an incident laser pulse with suitable pulse area evolves from a given pulse waveform into a stable, spatially-localized oscillating or standing gap soliton. We show that multiple gap solitons can be simultaneously spatially localized, resulting in efficient optical energy conversion and storage in the resonantly absorbing Bragg structure as atomically coherent states.


The UK10K project identifies rare variants in health and disease.
The contribution of rare and low-frequency variants to human traits is largely unexplored. Here we describe insights from sequencing whole genomes (low read depth, 7×) or exomes (high read depth, 80×) of nearly 10,000 individuals from population-based and disease collections. In extensively phenotyped cohorts we characterize over 24 million novel sequence variants, generate a highly accurate imputation reference panel and identify novel alleles associated with levels of triglycerides (APOB), adiponectin (ADIPOQ) and low-density lipoprotein cholesterol (LDLR and RGAG1) from single-marker and rare variant aggregation tests. We describe population structure and functional annotation of rare and low-frequency variants, use the data to estimate the benefits of sequencing for association studies, and summarize lessons from disease-specific collections. Finally, we make available an extensive resource, including individual-level genetic and phenotypic data and web-based tools to facilitate the exploration of association results.


Advances in ITP--therapy and quality of life--a patient survey.
Current guidelines recommend glucocorticoids and splenectomy as standard 1(st) and 2(nd) line treatments for chronic immune thrombocytopenia (ITP). We sought to find out how German ITP-patients are treated with respect to these guidelines. Members of a patient support association ≥18 years with a self-reported history of chronic ITP>12 months were surveyed with a web-based questionnaire. 122 questionnaires were evaluated. 70% of patients had chronic ITP for more than 5 years and 20% an average platelet count of ≤30·10(9)/L. 41% of the patients reported haematomas or petechiae more than once or twice and up to 12 times or more per year and 17% oropharyngeal and nasal bleeds. 11% had been admitted to hospital during the last 12 months. 88% had received or currently receive glucocorticoids, 27% were splenectomised. IVIG had been given to 55%, rituximab to 22%, anti-D to 12%, ciclosporin to 7%, while complementary and alternative medical treatments had been used by 36%. 50 women responded to questions concerning pregnancy. 14 (28%) had been advised not to become pregnant. 23 reported pregnancies and 10 (44%) required treatment for their ITP during pregnancy. Glucocorticoids are the most common therapy for chronic ITP but complementary and alternative treatments already come second and less than ⅓ of patients are splenectomised. This and the frequent use of complementary medicines suggests patients' dissatisfaction with conventional approaches. Many patients receive off-label therapies. There is a major need for adequate counselling and care for pregnant ITP-patients.


Evolutionary and Modern Image Content Differentially Influence the Processing of Emotional Pictures.
From an evolutionary perspective, environmental threats relevant for survival constantly challenged human beings. Current research suggests the evolution of a fear processing module in the brain to cope with these threats. Recently, humans increasingly encountered modern threats (e.g., guns or car accidents) in addition to evolutionary threats (e.g., snakes or predators) which presumably required an adaptation of perception and behavior. However, the neural processes underlying the perception of these different threats remain to be elucidated. We investigated the effect of image content (i.e., evolutionary vs. modern threats) on the activation of neural networks of emotion processing. During functional magnetic resonance imaging (fMRI) 41 participants watched affective pictures displaying evolutionary-threatening, modern-threatening, evolutionary-neutral and modern-neutral content. Evolutionary-threatening stimuli evoked stronger activations than modern-threatening stimuli in left inferior frontal gyrus and thalamus, right middle frontal gyrus and parietal regions as well as bilaterally in parietal regions, fusiform gyrus and bilateral amygdala. We observed the opposite effect, i.e., higher activity for modern-threatening than for evolutionary-threatening stimuli, bilaterally in the posterior cingulate and the parahippocampal gyrus. We found no differences in subjective arousal ratings between the two threatening conditions. On the valence scale though, subjects rated modern-threatening pictures significantly more negative than evolutionary-threatening pictures, indicating a higher level of perceived threat. The majority of previous studies show a positive relationship between arousal rating and amygdala activity. However, comparing fMRI results with behavioral findings we provide evidence that neural activity in fear processing areas is not only driven by arousal or valence, but presumably also by the evolutionary content of the stimulus. This has also fundamental methodological implications, in the sense to suggest a more elaborate classification of stimulus content to improve the validity of experimental designs.


Metapangenomics of the oral microbiome provides insights into habitat adaptation and cultivar diversity.
The increasing availability of microbial genomes and environmental shotgun metagenomes provides unprecedented access to the genomic differences within related bacteria. The human oral microbiome with its diverse habitats and abundant, relatively well-characterized microbial inhabitants presents an opportunity to investigate bacterial population structures at an ecosystem scale. Here, we employ a metapangenomic approach that combines public genomes with Human Microbiome Project (HMP) metagenomes to study the diversity of microbial residents of three oral habitats: tongue dorsum, buccal mucosa, and supragingival plaque. For two exemplar taxa, Haemophilus parainfluenzae and the genus Rothia, metapangenomes reveal distinct genomic groups based on shared genome content. H. parainfluenzae genomes separate into three distinct subgroups with differential abundance between oral habitats. Functional enrichment analyses identify an operon encoding oxaloacetate decarboxylase as diagnostic for the tongue-abundant subgroup. For the genus Rothia, grouping by shared genome content recapitulates species-level taxonomy and habitat preferences. However, while most R. mucilaginosa are restricted to the tongue as expected, two genomes represent a cryptic population of R. mucilaginosa in many buccal mucosa samples. For both H. parainfluenzae and the genus Rothia, we identify not only limitations in the ability of cultivated organisms to represent populations in their native environment, but also specifically which cultivar gene sequences are absent or ubiquitous. Our findings provide insights into population structure and biogeography in the mouth and form specific hypotheses about habitat adaptation. These results illustrate the power of combining metagenomes and pangenomes to investigate the ecology and evolution of bacteria across analytical scales.


Variation in patterns of metal accumulation in thallus parts of Lessonia trabeculata (Laminariales; Phaeophyceae): implications for biomonitoring.
Seaweeds are well known to concentrate metals from seawater and have been employed as monitors of metal pollution in coastal waters and estuaries. However, research showing that various intrinsic and extrinsic factors can influence metal accumulation, raises doubts about the basis for using seaweeds in biomonitoring programmes. The thallus of brown seaweeds of the order Laminariales (kelps) is morphologically complex but there is limited information about the variation in metal accumulation between the different parts, which might result in erroneous conclusions being drawn if not accounted for in the biomonitoring protocol. To assess patterns of individual metals in the differentiated parts of the thallus (blade, stipe, holdfast), concentrations of a wide range of essential and non-essential metals (Fe, Cr, Cu, Zn, Mn, Pb, Cd, Ni and Al) were measured in the kelp Lessonia trabeculata. Seaweeds were collected from three sampling stations located at 5, 30 and 60 m from an illegal sewage outfall close to Ventanas, Chile and from a pristine location at Faro Curaumilla. For the majority of metals the highest concentrations in bottom sediment and seaweed samples were found at the site closest to the outfall, with concentrations decreasing with distance from the outfall and at control stations; the exception was Cd, concentrations of which were higher at control stations. The patterns of metal concentrations in different thallus parts were metal specific and independent of sampling station. These results and the available literature suggest that biomonitoring of metals using seaweeds must take account of differences in the accumulation of metals in thallus parts of complex seaweeds.


Vinculin regulates osteoclast function.
Osteoclastic bone resorption depends upon the cell's ability to organize its cytoskeleton. Because vinculin (VCL) is an actin-binding protein, we asked whether it participates in skeletal degradation. Thus, we mated VCL(fl/fl) mice with those expressing cathepsin K-Cre (CtsK-VCL) to delete the gene in mature osteoclasts or lysozyme M-Cre (LysM-VCL) to target all osteoclast lineage cells. VCL-deficient osteoclasts differentiate normally but, reflecting cytoskeletal disorganization, form small actin rings and fail to effectively resorb bone. In keeping with inhibited resorptive function, CtsK-VCL and LysM-VCL mice exhibit a doubling of bone mass. Despite cytoskeletal disorganization, the capacity of VCL(-/-) osteoclastic cells to normally phosphorylate c-Src in response to αvβ3 integrin ligand is intact. Thus, integrin-activated signals are unrelated to the means by which VCL organizes the osteoclast cytoskeleton. WT VCL completely rescues actin ring formation and bone resorption, as does VCL(P878A), which is incapable of interacting with Arp2/3. As expected, deletion of the VCL tail domain (VCL(1-880)), which binds actin, does not normalize VCL(-/-) osteoclasts. The same is true regarding VCL(I997A), which also prevents VCL/actin binding, and VCL(A50I) and VCL(811-1066), both of which arrest talin association. Thus, VCL binding talin, but not Arp2/3, is critical for osteoclast function, and its selective inhibition retards physiological bone loss.


Mitogenic effects of human recombinant insulin on B-cell precursor acute lymphoblastic leukemia cells.
Insulin and the insulin-like growth factors (IGF-I, IGF-II) constitute a family of peptides capable of stimulating diverse cellular responses, including cell proliferation. In order to determine the effects of these peptides on malignant cells, we analyzed the expression and function of insulin, IGF-I, and IGF-II receptors on B-cell precursor acute lymphoblastic leukemia (BCP ALL) cell lines, utilizing competitive binding, affinity crosslinking, and cell proliferation assays. The BCP ALL cells bound to each peptide with mean specific binding for 125I-insulin, 125I-IGF-I, and 125I-IGF-II of 19.6%, 7.1%, and 4.3% of radioligand added, respectively. Competitive binding to intact cells demonstrated that 125I-IGF-I was displaced by IGF-I = IGF-II >> insulin, 125I-IGF-II was displaced by IGF-II > insulin = IGF-I, and 125I-insulin was displaced by insulin >> IGF-II > IGF-I. These data were remarkable for the potency of IGF-II displacement of 125I-IGF-I and 125I-insulin. Affinity crosslinking of radioligands to SUP-B2 cell membranes demonstrated the high affinity insulin and IGF-I (type 1 IGF) receptors. IGF binding proteins were also present in BCP ALL cell membrane preparations. In the cell proliferation studies, insulin stimulated a 50-130% increase in leukemic cell growth with a half-maximal concentration of 0.1-3.0 ng/ml in three BCP ALL cell lines. The proliferative response to insulin was blocked by the addition of an insulin receptor antibody. However, no response was observed with IGF-I, and IGF-II was only weakly mitogenic with a proliferative response noted at 100 ng/ml. Thus, while BCP ALL cells possess receptors for insulin and IGF-I, only the insulin receptor mediated a proliferative response.


Decreased mitochondrial respiration in aneurysmal aortas of Fibulin-4 mutant mice is linked to PGC1A regulation.
Thoracic aortic aneurysms are a life-threatening condition often diagnosed too late. To discover novel robust biomarkers, we aimed to better understand the molecular mechanisms underlying aneurysm formation. In Fibulin-4R/R mice, the extracellular matrix protein Fibulin-4 is 4-fold reduced, resulting in progressive ascending aneurysm formation and early death around 3 months of age. We performed proteomics and genomics studies on Fibulin-4R/R mouse aortas. Intriguingly, we observed alterations in mitochondrial protein composition in Fibulin-4R/R aortas. Consistently, functional studies in Fibulin-4R/R vascular smooth muscle cells (VSMCs) revealed lower oxygen consumption rates, but increased acidification rates. Yet, mitochondria in Fibulin-4R/R VSMCs showed no aberrant cytoplasmic localization. We found similar reduced mitochondrial respiration in Tgfbr-1M318R/+ VSMCs, a mouse model for Loeys-Dietz syndrome (LDS). Interestingly, also human fibroblasts from Marfan (FBN1) and LDS (TGFBR2 and SMAD3) patients showed lower oxygen consumption. While individual mitochondrial Complexes I-V activities were unaltered in Fibulin-4R/R heart and muscle, these tissues showed similar decreased oxygen consumption. Furthermore, aortas of aneurysmal Fibulin-4R/R mice displayed increased reactive oxygen species (ROS) levels. Consistent with these findings, gene expression analyses revealed dysregulation of metabolic pathways. Accordingly, blood ketone levels of Fibulin-4R/R mice were reduced and liver fatty acids were decreased, while liver glycogen was increased, indicating dysregulated metabolism at the organismal level. As predicted by gene expression analysis, the activity of PGC1α, a key regulator between mitochondrial function and organismal metabolism, was downregulated in Fibulin-4R/R VSMCs. Increased TGFβ reduced PGC1α levels, indicating involvement of TGFβ signalling in PGC1α regulation. Activation of PGC1α restored the decreased oxygen consumption in Fibulin-4R/R VSMCs and improved their reduced growth potential, emphasizing the importance of this key regulator. Our data indicate altered mitochondrial function and metabolic dysregulation, leading to increased ROS levels and altered energy production, as a novel mechanism, which may contribute to thoracic aortic aneurysm formation.


Weekly versus monthly testosterone administration on fast and slow skeletal muscle fibers in older adult males.
In older adults, loss of mobility due to sarcopenia is exacerbated in men with low serum T. T replacement therapy is known to increase muscle mass and strength, but the effect of weekly (WK) vs monthly (MO) administration on specific fiber types is unknown. To determine the efficacy of WK vs MO T replacement on the size and functional capacity of individual fast and slow skeletal muscle fiber types. Subjects were randomized into a 5-month, double-blind, placebo-controlled trial. All subjects (ages, 61-71 y) were community-dwelling men who had T levels < 500 ng/dL. Subjects were dosed weekly for 5 months, receiving continuous T (WK, n = 5; 100 mg T enanthate, im injection), monthly cycled T (MO, n = 7; alternating months of T and placebo), or placebo (n = 7). Muscle biopsies of the vastus lateralis were obtained before and after treatment. Main outcomes for individual slow and fast fibers included fiber diameter, peak force (P0), rate of tension development, maximal shortening velocity, peak power, and Ca(2+) sensitivity. Both treatments increased fiber diameter and peak power, with WK treatment 5-fold more effective than MO in increasing type I fiber P0. WK effects on fiber diameter and force were 1.5-fold higher in slow fibers compared to fast fibers. In fast type II fibers, diameter and P0 increased similarly between treatments. The increased power was entirely due to increased fiber size and force. In conclusion, T replacement effects were fiber-type dependent, restricted to increases in cell size, P0, and peak power, and dependent on the paradigm selected (WK vs MO).


Quasispecies of bovine enteric and respiratory coronaviruses based on complete genome sequences and genetic changes after tissue culture adaptation.
The genetic diversity of 2 pairs (AH65 and AH187) of wild type bovine coronaviruses (BCoV) sequenced directly from nasal (respiratory) and rectal (enteric) swabs of two feedlot calves with respiratory and enteric symptoms [Hasoksuz, M., Sreevatsan, S., Cho, K.O., Hoet, A.E., Saif, L.J., 2002b. Molecular analysis of the S1 subunit of the spike glycoprotein of respiratory and enteric bovine coronavirus isolates. Virus Res. 84 (1-2), 101-109.]. was analyzed. Sequence analysis of the complete genomes revealed differences at 123 and 149 nucleotides (nt) throughout the entire genome between the respiratory and enteric strains for samples AH65 and AH187, respectively, indicating the presence of intra-host BCoV quasispecies. In addition, significant numbers of sequence ambiguities were found in the genomes of some BCoV-R and BCoV-E strains, suggesting intra-isolate quasispecies. The tissue culture (TC) passaged counterparts of AH65 respiratory BCoV (AH65-R-TC) and enteric BCoV (AH65-E-TC) were also sequenced after 14 and 15 passages and 1 plaque purification in human rectal tumor cells (HRT-18), respectively. Compared to the parental wild type strains, tissue culture passage generated 104 nt changes in the AH65-E-TC isolate but only 8 nt changes in the AH65-R-TC isolate. Particularly noteworthy, the majority of nucleotide changes in the AH65-E-TC isolate occurred at the identical positions as the mutations occurring in the AH65-R strain from the same animal. These data suggest that BCoV evolves through quasispecies development, and that enteric BCoV isolates are more prone to genetic changes and may mutate to resemble respiratory BCoV strains after tissue culture passage.


Mutant prenyltransferase-like mitochondrial protein (PLMP) and mitochondrial abnormalities in kd/kd mice.
Mice that are homozygous for the kidney disease (kd) mutation are apparently healthy for the first 8 weeks of life, but spontaneously develop a severe form of interstitial nephritis that progresses to end-stage renal disease (ESRD) by 4 to 8 months of age. By testing for linkage to microsatellite markers, we previously localized the kd gene to a YAC/BAC contig. The sequence of the entire critical region was examined, and candidate genes were identified. These candidate genes were sequenced in both mutant (kd/kd) mice and normal controls. The phenotype was further characterized by immunohistochemistry and electron microscopy. Transgenic mice were constructed that carried the wild-type allele of the prime candidate gene, and this transgene was transferred to a kd/kd background by breeding. We have obtained evidence that kd is a mutant allele of a novel gene for a prenyltransferase-like mitochondrial protein (PLMP). This gene is alternatively spliced, with the larger gene product having one domain that resembles transprenyltransferase and another that is similar to geranylgeranyl pyrophosphate synthase. The smaller gene product includes only the first domain. An antiserum to PLMP localizes to mitochondria, and ultrastructural defects are present in the mitochondria of renal tubular epithelial cells, and to a lesser extent, hepatocytes and heart cells from kd/kd mice. In a line of kd/kd mice that carried the wild-type PLMP allele as a transgene, only 1 out of 13 animals expressed the disease by 120 days of age. The kd allele codes for a novel protein that localizes to the mitochondria, and the kd/kd mouse has dysmorphic mitochondria in the renal tubular epithelial cells. This mouse is therefore a unique animal model for studying mechanisms that lead to tubulointerstitial nephritis.


Single Synonymous Mutations in KRAS Cause Transformed Phenotypes in NIH3T3 Cells.
Synonymous mutations in the KRAS gene are clustered at G12, G13, and G60 in human cancers. We constructed 9 stable NIH3T3 cell lines expressing KRAS, each with one of these synonymous mutations. Compared to the negative control cell line expressing the wild type human KRAS gene, all the synonymous mutant lines expressed more KRAS protein, grew more rapidly and to higher densities, and were more invasive in multiple assays. Three of the cell lines showed dramatic loss of contact inhibition, were more refractile under phase contrast, and their refractility was greatly reduced by treatment with trametinib. Codon usage at these glycines is highly conserved in KRAS compared to HRAS, indicating selective pressure. These transformed phenotypes suggest that synonymous mutations found in driver genes such as KRAS may play a role in human cancers.


A Chiral Nitrogen Ligand for Enantioselective, Iridium-Catalyzed Silylation of Aromatic C-H Bonds.
Iridium catalysts containing dative nitrogen ligands are highly active for the borylation and silylation of C-H bonds, but chiral analogs of these catalysts for enantioselective silylation reactions have not been developed. We report a new chiral pyridinyloxazoline ligand for enantioselective, intramolecular silylation of symmetrical diarylmethoxy diethylsilanes. Regioselective and enantioselective silylation of unsymmetrical substrates was also achieved in the presence of this newly developed system. Preliminary mechanistic studies imply that C-H bond cleavage is irreversible, but not the rate-determining step.


Ejaculate quality and constraints in relation to sperm competition levels among eutherian mammals.
The outcome of sperm competition is influenced by the relative quantity and quality of sperm among competing ejaculates. Whereas it is well established that individual ejaculate traits evolve rapidly under postcopulatory sexual selection, little is known about other factors that might influence the evolution of ejaculates. For example, the metabolic rate is likely to affect the sperm production rate and the cellular activity or metabolism of sperm, and it has recently been suggested to constrain the evolution of sperm length in large but not small mammals. I thus examined in eutherian mammals how ejaculate quality traits vary with one another and with testis mass, body size, and metabolism. I found all ejaculate traits to covary positively with one another and to increase with relative testis mass. When controlling for testis mass, small-bodied species showed superior sperm quality (but not sperm number). Furthermore, sperm motility and viability were positively associated with the mass-corrected metabolic rate, but the percentage of morphologically normal and acrosome-intact sperm were not. These results indicate that body size and the energy budget may also influence the evolution of ejaculate quality, although these influences appear to vary among traits.


Phylogenetic relationships of Middle American cichlids (Cichlidae, Heroini) based on combined evidence from nuclear genes, mtDNA, and morphology.
Heroine cichlids are the second largest and very diverse tribe of Neotropical cichlids, and the only cichlid group that inhabits Mesoamerica. The taxonomy of heroines is complex because monophyly of most genera has never been demonstrated, and many species groups are without applicable generic names after their removal from the catch-all genus Cichlasoma (sensu Regan, 1905). Hence, a robust phylogeny for the group is largely wanting. A rather complete heroine phylogeny based on cytb sequence data is available [Concheiro Pérez, G.A., Rícan O., Ortí G., Bermingham, E., Doadrio, I., Zardoya, R. 2007. Phylogeny and biogeography of 91 species of heroine cichlids (Teleostei: Cichlidae) based on sequences of the cytochrome b gene. Mol. Phylogenet. Evol. 43, 91-110], and in the present study, we have added and analyzed independent data sets (nuclear and morphological) to further confirm and strengthen the cytb-phylogenetic hypothesis. We have analyzed a combined cytb-nuclear (RAG1 and two S7 introns) data set of 48 species representing main heroine lineages to achieve further resolution of heroine higher taxonomic levels and a combined cytb-morphological data set of 92 species to stabilize generic taxonomy. The recovered phylogenies supported the circumamazonian--CAM--Heroini (sensu Concheiro Peréz et al., 2007) as a monophyletic group, that could be divided into six main clades: (1) australoheroines (the southernmost heroine genus Australoheros), (2) nandopsines (the Antillean genus Nandopsis), (3) caquetaines (including the north western Amazonian genera Caquetaia and Heroina), (4) astatheroines (including Astatheros, Herotilapia and Rocio), (5) amphilophines (including Amphilophus and related genera), and (6) herichthyines (including Herichthyis and related genera). Nuclear and mitochondrial data partitions arrived at highly congruent topologies. Suprageneric relationships were influenced mainly by the nuclear signal, as well as the most basal phylogenetic position of Australoheros within CAM heroines. The new phylogeny of the tribe Heroini provides robust framework to stabilize the taxonomy of the group and for future comparative studies on these morphologically and ecologically diverse freshwater fishes. Morphology was mostly informative at the genus level and aid in determining the monophyly and composition of heroine genera. Upon acceptance of all putative genera, as recovered in this study, the Heroini would be with 35 genera the most genus-rich clade of Neotropical cichlids.


Holographic particle-image velocimetry in the first International Microgravity Laboratory aboard the Space Shuttle Discovery.
In January 1992 the Space Shuttle Discovery carried the first International Microgravity Laboratory into Earth orbit for eight days. One of the many experiments carried out during the orbit was a combined study of triglycine sulfate crystal growth from solution and fluid-particle-dynamics studies in microgravity. Optical diagnostics included holocameras to provide concentration measurements and three-dimensional particle tracking. More than 1000 holograms that were recorded in space have been analyzed since the flight, providing a wide range of interesting conclusions about microgravity, crystal growth, and particle dynamics. This paper focuses on the results of holographic particle-image velocimetry experiments and provides an excellent example, along with new techniques, for exploiting holography for particle and flow diagnostics.


Diet quality of individuals with rheumatoid arthritis using the Healthy Eating Index (HEI)-2010.
Rheumatoid arthritis (RA) afflicts approximately 1.5 million American adults and is a major cause of disability. As disease severity worsens, individuals with RA may experience functional decline that can impact dietary intake. The objective of this study is to assess the diet quality of individuals with RA using the Healthy Eating Index (HEI)-2010 and examine associations between diet quality and disease activity and functional status. This cross-sectional study assessed diet quality and disease activity and functional status in adults with RA. Participants completed seven-day weighed food records, which were scored using the HEI-2010. Participants had a fasting blood draw and completed the Multidimensional Health Assessment Questionnaire to determine disease activity and functional status. The mean age of individuals with RA ( N = 84) was 53 ± 14 years, and 86.9% were female. The mean HEI-2010 total score was 58.7 ± 15.9, with 7.1% of participants scoring "good", 58.3% "fair", and 34.5% "poor". Most participants did not adhere to recommended intakes of total fruit, total vegetables, whole grains, fatty acids, refined grains, sodium, and empty calories. An unadjusted multiple linear regression model found duration of morning stiffness and C-reactive protein concentration to be significant variables to inversely predict HEI-2010 total score. The diet quality of many individuals with RA needs improvement and may be related to functional disability associated with RA. Healthcare providers should encourage individuals with RA to meet dietary guidelines and maintain a healthy diet. Moreover, healthcare providers should be aware of the potential impacts of functional disability on diet quality in individuals with RA.


The importance of imported infections in maintaining hepatitis B in The Netherlands.
In The Netherlands, in May 1999 an enhanced surveillance of hepatitis B was begun to collect detailed information of patients with acute hepatitis B virus (HBV) infection. The objective was to gain insight in transmission routes and source of infection of new HBV cases. Through public health services, patients were interviewed on risk factors. It appeared that the majority (59%) acquired the infection through sexual contact; 52% of these by homosexual and 48% by heterosexual contact. In 60% of the heterosexual cases, the source of infection was a partner originating from a hepatitis B-endemic region. Sexual transmission is the most common route of transmission of acute hepatitis B in The Netherlands and introduction of infections from abroad plays a key role in the current epidemiology of HBV. As well as prevention programmes targeted at sexual high-risk groups, prevention efforts should focus more on the heterosexual transmission from HBV carriers.


Decay-Associated Fourier Spectroscopy: Visible to Shortwave Infrared Time-Resolved Photoluminescence Spectra.
We describe and implement an interferometric approach to decay-associated photoluminescence spectroscopy, which we term decay-associated Fourier spectroscopy (DAFS). In DAFS, the emitted photon stream from a substrate passes through a variable path length Mach-Zehnder interferometer prior to detection and timing. The interferometer encodes spectral information in the intensity measured at each detector enabling simultaneous spectral and temporal resolution. We detail several advantages of DAFS, including wavelength-range insensitivity, drift-noise cancellation, and optical mode retention. DAFS allows us to direct the photon stream into an optical fiber, enabling the implementation of superconducting nanowire single photon detectors for energy-resolved spectroscopy in the shortwave infrared spectral window (λ = 1-2 μm). We demonstrate the broad applicability of DAFS, in both the visible and shortwave infrared, using two Förster resonance energy transfer pairs: a pair operating with conventional visible wavelengths and a pair showing concurrent acquisition in the visible and the shortwave infrared regime.


Rhythmic modulation of θ oscillations supports encoding of spatial and behavioral information in the rat hippocampus.
Oscillatory patterns of activity in various frequency ranges are ubiquitously expressed in cortical circuits. While recent studies in humans emphasized rhythmic modulations of neuronal oscillations ("second-order" rhythms), their potential involvement in information coding remains an open question. Here, we show that a rhythmic (~0.7 Hz) modulation of hippocampal theta power, unraveled by second-order spectral analysis, supports encoding of spatial and behavioral information. The phase preference of neuronal discharge within this slow rhythm significantly increases the amount of information carried by action potentials in various motor/cognitive behaviors by (1) distinguishing between the spikes fired within versus outside the place field of hippocampal place cells, (2) disambiguating place firing of neurons having multiple place fields, and (3) predicting between alternative future spatial trajectories. This finding demonstrates the relevance of second-order spectral components of brain rhythms for decoding neuronal information.


Mitochondrial DNA disease and developmental implications for reproductive strategies.
Mitochondrial diseases are potentially severe, incurable diseases resulting from dysfunctional mitochondria. Several important mitochondrial diseases are caused by mutations in mitochondrial DNA (mtDNA), the genetic material contained within mitochondria, which is maternally inherited. Classical and modern therapeutic approaches exist to address the inheritance of mtDNA disease, but are potentially complicated by the fact that cellular mtDNA populations evolve according to poorly-understood dynamics during development and organismal lifetimes. We review these therapeutic approaches and models of mtDNA dynamics during development, and discuss the implications of recent results from these models for modern mtDNA therapies. We particularly highlight mtDNA segregation-differences in proliferative rates between different mtDNA haplotypes-as a potential and underexplored issue in such therapies. However, straightforward strategies exist to combat this and other potential therapeutic problems. In particular, we describe haplotype matching as an approach with the power to potentially ameliorate any expected issues from mtDNA incompatibility.


Outcome and resource use associated with myomectomy.
To evaluate the outcomes and cost of myomectomy through retrospective claims data analysis. The study was performed using a retrospective database of private insurance claims from 1995 to 1997. Records were selected for analysis based on the presence of ICD-9-CM procedure and/or CPT-4 codes associated with myomectomy. In addition, diagnosis of uterine leiomyoma and related symptoms for these patients were confirmed through ICD-9-CM diagnosis codes. Inpatient, outpatient, and physician costs were estimated. All cost data were converted into 1997 dollars. A total of 4394 women, between the ages of 14 and 70, were available for analysis. Of these, 3305 were classified by type of myomectomy procedure, and complete data were available on 820 at 1 year and 236 at 2 years. Abdominal myomectomies were the most common procedures, followed by hysteroscopic and laparoscopic myomectomies. Conversion to a more invasive procedure occurred in 5.4% of the patients. The rate of additional surgeries was 8.3% in 6 months, 10.6% in 1 year, and 16.5% in 2 years. Overall cost increased from an initial 6,737 dollars per patient to 7,575 dollars in 1 year and to 8,001 dollars in 2 years. The repeat procedures required after the initial myomectomy add significantly to total cost and highlight the importance of assessing post-procedure health care use. This comprehensive analysis facilitates the systematic evaluation of myomectomy with current and emerging alternative treatments for uterine leiomyomas.


Severe hypoxemia associated with liver disease: Mayo Clinic experience and the experimental use of almitrine bismesylate.
Severe hypoxemia associated with chronic liver disease is an uncommon disorder most likely due to an intrapulmonary vascular abnormality that has characteristics of both ventilation-perfusion mismatching and diffusion limitation. Anatomically, the intrapulmonary vascular abnormalities can occasionally be detected by angiography. Physiologically, the gas exchange abnormalities can be substantiated by contrast-enhanced two-dimensional echocardiography. Although orthodeoxia and platypnea have frequently been found in these patients, echocardiographic data suggest that vascular abnormalities can exist in the absence of orthodeoxia. We describe 11 patients who had severe hypoxemia and chronic liver disease and review their pulmonary angiographic, contrast echocardiographic, and arterial blood gas findings. Among five of these patients who were given almitrine bismesylate, an experimental medication thought to alter ventilation-perfusion relationships in patients with chronic obstructive pulmonary disease, one had improved oxygenation. We recommend that patients with hypoxemia associated with chronic liver disease have detailed studies to rule out reversible forms of hypoxemia and that those with severe hypoxemia undergo testing to determine the existence of intrapulmonary vascular abnormalities, especially if liver transplantation is considered.


Illuminating the dark side of indoor oxidants.
The chemistry of oxidants and their precursors (oxidants*) plays a central role in outdoor environments but its importance in indoor air remains poorly understood. Ozone (O3) chemistry is important in some indoor environments and, until recently, ozone was thought to be the dominant oxidant indoors. There is now evidence that formation of the hydroxyl radical by photolysis of nitrous acid (HONO) and formaldehyde (HCHO) may be important indoors. In the past few years, high time-resolution measurements of oxidants* indoors have become more common and the importance of event-based release of oxidants* during activities such as cleaning has been proposed. Here we review the current understanding of oxidants* indoors, including drivers of the formation and loss of oxidants*, levels of oxidants* in indoor environments, and important directions for future research.


Thiol protein groups correlate with cognitive impairment in patients with recurrent depressive disorder.
Depressive disorders are multifactorial diseases in which cognitive impairments are one of typical features. Thiol protein groups (TPGs) are elements of chemical structure of compounds having antioxidative properties (glutathione peroxidase, metallothioneins) and their oxidation reflects the lost of compensatory capacity of antioxidative mechanisms. The purpose of this study was to determine the level of TPGs in patients with recurrent depressive disorder (rDD) and to define relationship between plasma levels of TPGs and the cognitive performance. The study comprised 76 subjects: patients with rDD (n=43) and healthy subjects (comparison group, CG - n=33). Cognitive function assessment was based on the following 4 tests: the Trail Making Test (TMT), the Stroop Test, Verbal Fluency Test (VFT) and Auditory Verbal Learning Test (AVLT). The level of TPGs was higher in patients with rDD. In rDD group, the elevated concentration of TPGs in plasma was associated with a decrease in efficiency of declarative-memory, working memory and verbal fluency. The higher was the concentration of plasma TPGs, the greater was the severity of depressive symptoms measured by 21-item Hamilton Depression Rating Scale (HDRS), before and after pharmacotherapy. In CG group, the elevated TPGs levels were associated with worse cognitive test performance (AVLT and VFT tests). 1) Our study confirms previous results showing increased TPGs level in depression. 2) Our data suggest relation between increased plasma TPGs level in depression and cognitive impairment. 3) The elevated levels of plasma TPGs are related to impairment of the short-term and delayed declarative memory, verbal fluency and working memory.


Siderophile Elements in Tracing Planetary Formation and Evolution.
The siderophile, or iron-loving elements have many applications in the Earth and planetary sciences. In primitive meteorites, differences in the relative abundances of these elements are likely due to both nebular and parent body processes. In addition, some siderophile elements are also characterised by isotopically distinctive nucleosynthetic signatures. Thus, the relative abundances and isotopic compositions of these elements can be used to trace the genetics of primary planetary building blocks. Although these elements are largely concentrated in the metallic cores of differentiated planetary bodies, their absolute and relative abundances, as well as their isotopic compositions can also reveal important information regarding conditions of core formation and the chemical evolution of the silicate portions of the planetary bodies. The lithophile-siderophile nature of the radiogenic 182Hf-182W system allow it to be used to place chronologic constraints on planetary core formation. The differing incompatibilities of the two elements in silicate systems further mean that the system can also be used to study early differentiation processes and subsequent efficiency of mixing in the silicate portions of differentiated bodies, including Earth. The abundances of siderophile elements in the terrestrial mantle are used to assess primary and secondary melting processes, and resulting metasomatic interactions. In addition, the Re-Os isotope system can, in some instances, be used to place chronologic constraints on when these processes occurred. The abundances of siderophile elements, and 187Os/188Os and 186Os/188Os ratios in the mantle sources of ocean island basalts can be used to place constraints on the age of recycled materials, and in some instances, the types of recycled materials present in these mantle domains.


Detailed DNA methylation profiles of the E-cadherin promoter in the NCI-60 cancer cells.
E-cadherin (E-cad) is a transmembrane adhesion glycoprotein, the expression of which is often reduced in invasive or metastatic tumors. To assess E-cad's distribution among different types of cancer cells, we used bisulfite-sequencing for detailed, base-by-base measurement of CpG methylation in E-cad's promoter region in the NCI-60 cell lines. The mean methylation levels of the cell lines were distributed bimodally, with values pushed toward either the high or low end of the methylation scale. The 38 epithelial cell lines showed substantially lower (28%) mean methylation levels compared with the nonepithelial cell lines (58%). The CpG site at -143 with respect to the transcriptional start was commonly methylated at intermediate levels, even in cell lines with low overall DNA methylation. We also profiled the NCI-60 cell lines using Affymetrix U133 microarrays and found E-cad expression to be correlated with E-cad methylation at highly statistically significant levels. Above a threshold of approximately 20% to 30% mean methylation, the expression of E-cad was effectively silenced. Overall, this study provides a type of detailed analysis of methylation that can also be applied to other cancer-related genes. As has been shown in recent years, DNA methylation status can serve as a biomarker for use in choosing therapy.


Systemic inflammation, nutritional status and tumor immune microenvironment determine outcome of resected non-small cell lung cancer.
Hypothesizing that nutritional status, systemic inflammation and tumoral immune microenvironment play a role as determinants of lung cancer evolution, the purpose of this study was to assess their respective impact on long-term survival in resected non-small cell lung cancers (NSCLC). Clinical, pathological and laboratory data of 303 patients surgically treated for NSCLC were retrospectively analyzed. C-reactive protein (CRP) and prealbumin levels were recorded, and tumoral infiltration by CD8+ lymphocytes and mature dendritic cells was assessed. We observed that factors related to nutritional status, systemic inflammation and tumoral immune microenvironment were correlated; significant correlations were also found between these factors and other relevant clinical-pathological parameters. With respect to outcome, at univariate analysis we found statistically significant associations between survival and the following variables: Karnofsky index, American Society of Anesthesiologists (ASA) class, CRP levels, prealbumin concentrations, extent of resection, pathologic stage, pT and pN parameters, presence of vascular emboli, and tumoral infiltration by either CD8+ lymphocytes or mature dendritic cells and, among adenocarcinoma type, tumor grade (all p<0.05). In multivariate analysis, prealbumin levels (Relative Risk (RR): 0.34 [0.16-0.73], p = 0.0056), CD8+ cell count in tumor tissue (RR = 0.37 [0.16-0.83], p = 0.0162), and disease stage (RR 1.73 [1.03-2.89]; 2.99[1.07-8.37], p = 0.0374- stage I vs II vs III-IV) were independent prognostic markers. When taken together, parameters related to systemic inflammation, nutrition and tumoral immune microenvironment allowed robust prognostic discrimination; indeed patients with undetectable CRP, high (>285 mg/L) prealbumin levels and high (>96/mm2) CD8+ cell count had a 5-year survival rate of 80% [60.9-91.1] as compared to 18% [7.9-35.6] in patients with an opposite pattern of values. When stages I-II were considered alone, the prognostic significance of these factors was even more pronounced. Our data show that nutrition, systemic inflammation and tumoral immune contexture are prognostic determinants that, taken together, may predict outcome.


Predictive and prognostic value of de novo MET expression in patients with advanced non-small-cell lung cancer.
Cellular-mesenchymal-epithelial transition (MET) protein has recently been identified as a novel target that shows promise for the treatment of non-small-cell lung cancer (NSCLC). However, the relationship between de novo MET expression and patient outcomes remains unclear. We reviewed the data of patients who had been diagnosed with NSCLC between December 2013 and October 2014. All the patients were evaluated for MET expression status. MET-positive was defined as having an H-score ≥ 60 by immunohistochemistry analysis. MET expression was analyzed in 158 patients who were negative for the common driver genes, including EGFR, ALK, KRAS and ROS1. A chi-squared test was used to assess the clinicopathological parameters. Multivariate analyses were performed using the Cox proportional hazards model. MET data were available for analysis in 158 advanced NSCLC patients. Of these, based on the MET H-score criteria, the IHC-positive rate was 48.1% (76/158). There were more patients with adenocarcinoma in the MET-positive group compared with the MET-negative group (P=0.01). Nine patients with lymphoepithelioma-like carcinoma had no MET expression. There was no significant difference in overall survival (OS) between MET-positive and -negative patients. There was also no significant difference in the efficacy (Z=-0.44, P=0.66) or progression free survival (PFS) of first-line chemotherapy between the MET-positive and -negative patients (mPFS 6.8 months [95% CI: 5.4-8.2] vs. 5.9 months [5.5-6.3], P=0.92). In the cell model, the MET-positive cell showed no difference in chemotherapy but with a increase in percentage of growth inhibition upon treatment with MET inhibitor INC280 compared to MET-negative cell. Our data showed that de novo MET expression was not rare. It was not a predictive or prognostic factor for stage IV NSCLC patients, but this should be confirmed in larger population cohort.


Spontaneous haemorrhagic perforation of gallbladder in acute cholecystitis as a complication of antiplatelet, immunosuppressant and corticosteroid therapy.
An older lady presented 1 week after being discharged from hospital with acute cholecystitis. She suffered a sudden onset lower abdominal pain and was in hypovolaemic shock upon arrival. It was noted that she had been on antiplatelet therapy after suffering a recent myocardial infarction, an immunosuppressor and steroids for rheumatoid arthritis. Her admission bloods revealed a platelet count of 83 with normal clotting factors. After resuscitation, a CT scan confirmed fluid in the abdomen possibly arising from the right subhepatic space. During laparotomy, bleeding was noted from a perforated and ischaemic-looking gallbladder, with an intact cystic artery and duct and no biliary calculi evident. The gallbladder was removed and the patient was transferred to intensive therapy unit. She recovered well within the subsequent 8 days and was discharged. Her histology described 'haemorrhage within the gallbladder wall along with oedema, fibrosis and patchy inflammation and no signs of malignancy or gangrene'.


Impact of introducing near patient testing for standard investigations in general practice.
To assess the clinical and economic impact of surgery based near patient testing in general practice for six commonly used biochemical and bacteriological tests. After four months' monitoring, equipment for two bacteriological and four biochemical tests was introduced without cost into 12 practices using a crossover design. Structured request forms were used to monitor laboratory investigations. 12 general practices in west midlands and south west Thames with list sizes above 9000. Investigation rates per 1000 consultations. Changes from baseline rates. Reasons for requesting investigations and provisional diagnoses. Cost per test and sensitivity of costs to rate of use. Investigation rates for the six tests rose by 16.5% (from 78.6/1000 consultations to 91.6/1000) when equipment was available in the surgery and reverted to baseline rates when it was withdrawn. The average weekly number of tests when equipment was available ranged from 0.5 to 10.5 (mean 9.0). Cholesterol tests were used as an addition to laboratory testing, usually for screening. Midstream urine analysis was often done in the surgery instead of in the laboratory, although 30% of samples were tested by both methods. Doctors' reasons for investigation and conditions tested were largely unaffected by availability of surgery tests. Costs for surgery tests were higher for all tests except midstream urine. Availability of surgery based testing increased the number of tests performed. It was cost effective only for midstream urine analysis.


Active regulator of SIRT1 is required for cancer cell survival but not for SIRT1 activity.
The NAD(+)-dependent deacetylase SIRT1 is involved in diverse cellular processes, and has also been linked with multiple disease states. Among these, SIRT1 expression negatively correlates with cancer survival in both laboratory and clinical studies. Active regulator of SIRT1 (AROS) was the first reported post-transcriptional regulator of SIRT1 activity, enhancing SIRT1-mediated deacetylation and downregulation of the SIRT1 target p53. However, little is known regarding the role of AROS in regulation of SIRT1 during disease. Here, we report the cellular and molecular effects of RNAi-mediated AROS suppression, comparing this with the role of SIRT1 in a panel of human cell lines of both cancerous and non-cancerous origins. Unexpectedly, AROS is found to vary in its modulation of p53 acetylation according to cell context. AROS suppresses p53 acetylation only following the application of cell damaging stress, whereas SIRT1 suppresses p53 under all conditions analysed. This supplements the original characterization of AROS but indicates that SIRT1 activity can persist following suppression of AROS. We also demonstrate that knockdown of AROS induces apoptosis in three cancer cell lines, independent of p53 activation. Importantly, AROS is not required for the viability of three non-cancer cell lines indicating a putative role for AROS in specifically promoting cancer cell survival.


Low-affinity B cells transport viral particles from the lung to the spleen to initiate antibody responses.
The lung is an important entry site for pathogens; its exposure to antigens results in systemic as well as local IgA and IgG antibodies. Here we show that intranasal administration of virus-like particles (VLPs) results in splenic B-cell responses with strong local germinal-center formation. Surprisingly, VLPs were not transported from the lung to the spleen in a free form but by B cells. The interaction between VLPs and B cells was initiated in the lung and occurred independently of complement receptor 2 and Fcγ receptors, but was dependent upon B-cell receptors. Thus, B cells passing through the lungs bind VLPs via their B-cell receptors and deliver them to local B cells within the splenic B-cell follicle. This process is fundamentally different from delivery of blood or lymph borne particulate antigens, which are transported into B cell follicles by binding to complement receptors on B cells.


Regulation of gene expression in gastric epithelial cell populations of fetal, neonatal, and adult transgenic mice.
Little is known about lineage relationships and differentiation programs of various epithelial cells present in mouse gastric units. We have previously used rat liver fatty acid binding protein/human growth hormone (L-FABP/hGH) transgenes to define epithelial cell lineages relationships in the small intestine of fetal and adult mice and to examine regulation of their terminal differentiation programs along the crypt-to-villus and duodenal-to-ileal axes. We have now used these transgenes to explore similar issues in the stomach. Immunocytochemical studies of fetal and adult transgenic L-FABP/hGH animals and their normal littermates revealed that the intact endogenous mouse L-FABP gene (Fabpl) is not expressed in gastric epithelium. Nucleotides-596 to +21 of the rat L-FABP gene direct "inappropriate" expression of hGH in the gastric epithelium as early as fetal day 15. From 1 to 13 mo, L-FABP-596 to +21/hGH expression occurs only in surface mucous cells of zymogenic and mucous gastric units; the reporter is not detectable in the enteroendocrine, parietal and chief cell populations of zymogenic glands. Electron microscopic immunocytochemistry revealed that hGH is directed to apical secretory granules in surface and pit mucous cells expressing the transgene. hGH levels vary widely among surface mucous cells both within single pits and between gastric units in a given animal. The heterogeneity noted in reporter expression suggests that there are marked differences in the regulatory environments of individual cells of a single type within a given gastric unit. This raises the possibility that cell differentiation programs in the stomach may not be as tightly coupled to cellular translocation as in the small intestine. Finally, the lack of expression of L-FABP-596 to +21/hGH in gastrin- and serotonin-immunoreactive cells of the stomach contrasts with its efficient expression in comparable cell types located in the duodenum; providing a model system for examining differential regulation of gene expression in terminally differentiated cell types represented in both gastric and intestinal epithelium.


Current pattern of primary aldosteronism diagnosis: Delayed and complicated
Primary aldosteronism is the most common specifically treatable and potentially curable cause of hypertension. The aim of this study was to analyse the referral pattern and disease characteristics of patients with hypertension and suspected primary aldosteronism. We prospectively collected clinical data from patients who attended the Endocrine Hypertension Service (EHS) at Monash Health from May 2016 to May 2017. Of 87 patients, 69.2% had indications for primary aldosteronism screening in the primary care setting, although only 3.7% were actually screened. At the time of referral to the EHS, 61% had hypertension for over 10 years and 42% of the patients with primary aldosteronism had associated end-organ damage. Targeted treatment of primary aldosteronism led to biochemical and clinical improvement in all patients. The diagnosis of primary aldosteronism is currently delayed and associated with significant end-organ damage. Increased awareness of primary aldosteronism and its screening indications in primary and tertiary care is needed for earlier diagnosis and improved outcomes.


A Gateway(®) -compatible bacterial adenylate cyclase-based two-hybrid system.
The bacterial adenylate cyclase two-hybrid (BACTH) system has been widely used to characterize protein-protein interactions in the prokaryotic world. This system relies on the interaction-mediated reconstitution of adenylate cyclase activity in Escherichia coli by bringing together two complementary fragments of the catalytic domain of the adenylate cyclase toxin of Bordetella pertussis. A limiting factor in performing large-scale two-hybrid interaction screens with full-length open reading frames (ORFs) is the need to clone each ORF individually into the plasmids used to express the hybrid proteins. The Gateway(®) (GW) cloning system (Life Technologies, Grand Island, NY, USA) partially circumvents this limitation, and we describe here modifications to the BACTH system for compatibility with this recombineering technology. We validated and tested the functionality of the BACTH Gateway (BACTHGW ) system using several models of protein-protein interactions, focusing particularly on those involved in bacterial cell division. We further modified the BACTH plasmids to incorporate a transmembrane (TM) segment downstream of the cyclase fragments to permit analysis of extracytoplasmic protein interactions. This approach was also useful to identify putative TM segments and to experimentally validate bioinformatically identified TM domains. The BACTHGW system will prove a useful addition to the study of protein-protein interactions.


[Whole Exome Sequencing in daily practice: the possibilities and impossibilities of this diagnostic test].
Whole Exome Sequencing (WES) is becoming a standard diagnostic tool for patients who potentially suffer from an underlying monogenic condition. This is illustrated by the use of WES panels for specific medical conditions; in such instances, analysis is focused on a subset of genes that can be implicated in the pathogenesis of a condition such as epilepsy, intellectual disability, cardiomyopathy, etc. Physicians requesting WES diagnostics should be informed about the possibilities, limitations and pitfalls of this test. Here, we give a short introduction on the background of WES and also explain its potential in today's practice. In addition, we will discuss the limitations of WES and the WES classification system, which is used to indicate the pathogenic character of identified gene variants, will be further explained. We expect that WES will soon become a standard diagnostic tool for physicians from many disciplines.


Striatal shape abnormalities as novel neurodevelopmental endophenotypes in schizophrenia: a longitudinal study.
There are varying, often conflicting, reports with respect to altered striatal volume and morphometry in the major psychoses due to the influences of antipsychotic medications on striatal volume. Thus, disassociating disease effects from those of medication become exceedingly difficult. For the first time, using a longitudinally studied sample of structural magnetic resonance images from patients with childhood onset schizophrenia (COS; neurobiologically contiguous with the adult onset form of schizophrenia), their nonpsychotic siblings (COSSIBs), and novel shape mapping algorithms that are volume independent, we report the familial contribution of striatal morphology in schizophrenia. The results of our volumetric analyses demonstrate age-related increases in overall striatal volumes specific only to COS. However, both COS and COSSIBs showed overlapping shape differences in the striatal head, which normalized in COSSIBs by late adolescence. These results mirror previous studies from our group, demonstrating cortical thickness deficits in COS and COSSIBs as these deficits normalize in COSSIBs in the same age range as our striatal findings. Finally, there is a single region of nonoverlapping outward displacement in the dorsal aspect of the caudate body, potentially indicative of a response to medication. Striatal shape may be considered complimentary to volume as an endophenotype, and, in some cases may provide information that is not detectable using standard volumetric techniques. Our striatal shape findings demonstrate the striking localization of abnormalities in striatal the head. The neuroanatomical localization of these findings suggest the presence of abnormalities in the striatal-prefrontal circuits in schizophrenia and resilience mechanisms in COSSIBs with age dependent normalization.