Targeted disruption of the extracellular polymeric network of Pseudomonas aeruginosa biofilms by alginate oligosaccharides.
Acquisition of a mucoid phenotype by Pseudomonas sp. in the lungs of cystic fibrosis (CF) patients, with subsequent over-production of extracellular polymeric substance (EPS), plays an important role in mediating the persistence of multi-drug resistant (MDR) infections. The ability of a low molecular weight (Mn = 3200 g mol-1) alginate oligomer (OligoG CF-5/20) to modify biofilm structure of mucoid Pseudomonas aeruginosa (NH57388A) was studied in vitro using scanning electron microscopy (SEM), confocal laser scanning microscopy (CLSM) with Texas Red (TxRd®)-labelled OligoG and EPS histochemical staining. Structural changes in treated biofilms were quantified using COMSTAT image-analysis software of CLSM z-stack images, and nanoparticle diffusion. Interactions between the oligomers, Ca2+ and DNA were studied using molecular dynamics (MD) simulations, Fourier transform infrared spectroscopy (FTIR) and isothermal titration calorimetry (ITC). Imaging demonstrated that OligoG treatment (≥0.5%) inhibited biofilm formation, revealing a significant reduction in both biomass and biofilm height (P < 0.05). TxRd®-labelled oligomers readily diffused into established (24 h) biofilms. OligoG treatment (≥2%) induced alterations in the EPS of established biofilms; significantly reducing the structural quantities of EPS polysaccharides, and extracellular (e)DNA (P < 0.05) with a corresponding increase in nanoparticle diffusion (P < 0.05) and antibiotic efficacy against established biofilms. ITC demonstrated an absence of rapid complex formation between DNA and OligoG and confirmed the interactions of OligoG with Ca2+ evident in FTIR and MD modelling. The ability of OligoG to diffuse into biofilms, potentiate antibiotic activity, disrupt DNA-Ca2+-DNA bridges and biofilm EPS matrix highlights its potential for the treatment of biofilm-related infections.


Rac2D57N, a dominant inhibitory Rac2 mutant that inhibits p38 kinase signaling and prevents surface ruffling in bone-marrow-derived macrophages.
Rac2 is a Rho GTPase that is expressed in cells of hematopoietic origin, including neutrophils and macrophages. We recently described an immunodeficient patient with severe, recurrent bacterial infections that had a point mutation in one allele of the Rac2 gene, resulting in the substitution of aspartate 57 with asparagine. To ascertain further the effects of Rac2D57N in leukocytes, Rac2D57N was expressed in primary murine bone-marrow-derived macrophages (cells that we show express approximately equal amounts of Rac1 and Rac2). Rac2D57N expression in macrophages inhibited membrane ruffling. Rac2D57N expression inhibited the formation of macropinosomes, demonstrating a functional effect of the loss of surface membrane dynamics. Surprisingly, Rac2D57N induced an elongated, spread morphology but did not affect microtubule networks. Rac2D57N also inhibited lipopolysaccharide-stimulated p38 kinase activation. Examination of guanine nucleotide binding to recombinant Rac2D57N revealed reduced dissociation of GDP and association of GTP. Coimmunoprecipitation studies of Rac2D57N with RhoGDI alpha and Tiam1 demonstrated increased binding of Rac2D57N to these upstream regulators of Rac signaling relative to the wild type. Enhanced binding of Rac2D57N to its upstream regulators would inhibit Rac-dependent effects on actin cytoskeletal dynamics and p38 kinase signaling.


Perimenopausal knowledge of mid-life women in northern Taiwan.
This work has been carried out keeping in view that although knowledge related to perimenopause is an important health issue for women worldwide, little information is known about mid-life Taiwanese women's knowledge of perimenopause. The aim of this paper was to explore the level of perimenopausal knowledge of mid-life women in northern Taiwan, to describe the relationship between demographic factors and women's knowledge, and to identify what information health providers need to offer them. A cross-sectional descriptive survey was conducted. The Perimenopausal Knowledge Questionnaire was used to collect data. The study was carried out in August 2000 to September 2001. A convenience sample was recruited from traditional Chinese medicine clinics, gynecology clinics and communities in northern Taipei. Our findings indicated that the overall mean correct answer weighted Perimenopausal Knowledge score of these mid-life women was 46.31. The rank of weighted scores from highest to lowest was: (1) self-care during perimenopause, (2) the perception of perimenopause, (3) knowledge related to hormone replacement therapy and (4) body changes associated with the declined oestrogen in perimenopause. Only educational level was significantly associated with Perimenopause Knowledge level (P < 0.01). Less than 20% (n = 353) of the women answered the items that measured self-management of symptoms related to perimenopause correctly. Less than 1.4% of the women knew that taking hormone replacement therapy required a physician's prescription and subsequent regular health check-ups. Participants lacked adequate knowledge about perimenopause. RELEVANCE TO CLINICAL PRACTICES: Health providers need to provide factual information about the normal body changes of perimenopause, self-management of perimenopausal disturbances and appropriate use of hormone replacement therapy.


Fabrication, in vitro and in vivo studies of bilayer composite membrane for periodontal guided tissue regeneration.
Development of a guided occlusive biodegradable membrane with controlled morphology in order to restrict the ingrowth of epithelial cells is still a challenge in dental tissue engineering. A bilayer membrane with a non-porous upper layer (polyurethane) and porous lower layer (polycaprolactone and bioactive glass composite) with thermoelastic properties to sustain surgery treatment was developed by lyophilization. Morphology, porosity, and layers attachment were controlled by using the multi-solvent system. In vitro and in vivo biocompatibility, cell attachment, and cell proliferation were analyzed by immunohistochemistry and histology. The cell proliferation rate and cell attachment results showed good biocompatibility of both surfaces, though cell metabolic activity was better on the polycaprolactone-bioactive glass surface. Furthermore, the cells were viable, adhered, and proliferated well on the lower porous bioactive surface, while non-porous polyurethane surface demonstrated low cell attachment, which was deliberately designed and a pre-requisite for guided tissue regeneration/guided bone regeneration membranes. In addition, in vivo studies performed in a rat model for six weeks revealed good compatibility of membranes. Histological analysis (staining with hematoxylin and eosin) indicated no signs of inflammation or accumulation of host immune cells. These results suggested that the fabricated biocompatible bilayer membrane has the potential for use in periodontal tissue regeneration.


Return to contingency: developing a coherent strategy for future R2E/R3 land medical capabilities.
Key to deploying forces in the future will be the provision of a rapidly deployable Deployed Hospital Capability. Developing this capability has been the focus of 34 Field Hospital and 2nd Medical Brigade over the last 18 months and this paper describes a personal account of this development work to date. Future contingent Deployed Hospital Capability must meet the requirements of Defence; that is to be rapidly deployable delivering a hospital standard of care. The excellence seen in clinical delivery on recent operations is intensive; in personnel, equipment, infrastructure and sustainment. The challenge in developing a coherent capability has been in balancing the clinical capability and capacity against strategic load in light of recent advances in battlefield medicine. This paper explores the issues encountered and solutions found to date in reconstituting a Very High Readiness Deployed Hospital Capability.


Beyond-use date determination of buprenorphine buccal solution using a stability-indicating high-performance liquid chromatographic assay.
The objectives of this study included developing and validating a stability-indicating high-performance liquid chromatographic (HPLC) method with ultraviolet (UV) detection for the determination of buprenorphine in a buccal solution for veterinary use, and applying that method to determine the stability of a 3 mg/ml buprenorphine preparation in room temperature and refrigerated storage conditions. This preparation, intended for buccal administration in feline patients, plays an important role in pain management in cats. A stability-indicating HPLC method was developed and validated for system suitability, accuracy, repeatability, intermediate precision, specificity, linearity and robustness based on US Pharmacopeia (USP) General Chapter <1225>. The method was then applied to the study of potency changes over 90 days in a buccal buprenorphine solution stored at two temperatures. All HPLC-UV method data met acceptable criteria for the quantification of buprenorphine in a buccal solution formulation. The buprenorphine concentrations found in each stability sample remained within the 90-110% of label claim throughout the 90 days of study. All stability test bottles of the buprenorphine buccal solution retained their original appearance. For the room temperature bottles, some white particulate matter was noted in the threads of the container bottles starting at day 21. The pH of the preparations during the course of the study was in the range of 3.57-4.06 and 4.01-4.16 for the room temperature and refrigerated samples, respectively. Pharmacists have compounded a concentrated 3 mg/ml buccal solution to use easily in the home care or outpatient setting for treatment of feline pain. Prior to this investigation, pharmacists empirically assigned beyond-use dates to this formulation based on standards in USP General Chapter <795> Pharmaceutical Compounding - Nonsterile Preparations. This study of a 3 mg/ml buprenorphine buccal solution indicates stability through 90 days.


Fermented rice peptides attenuate scopolamine-induced memory impairment in mice by regulating neurotrophic signaling pathways in the hippocampus.
This study investigated the preventive effects of fermented rice peptides (FRPs) against scopolamine-induced memory impairment in mice and their potential mechanisms. Mice were pretreated with FRPs (25 and 100 mg/kg body weight) via intraperitoneal injection for 7 days, followed by intraperitoneal injection of scopolamine. FRP pretreatment suppressed scopolamine-induced cognitive impairment in passive-avoidance test and significantly upregulated levels of brain-derived neurotrophic factor (BDNF) and induced the phosphorylation of cAMP response element binding (CREB) protein and extracellular signal-regulated kinase (ERK) in the hippocampus of scopolamine-treated mice. Additionally, scopolamine-treated mice showed significantly decreased acetylcholine levels and increased acetylcholine-esterase activity in the hippocampus as compared with controls; however, these changes were suppressed by FRP pretreatment. Among the fractions separated by size-exclusion chromatography, the non-glycosylated peptide fraction of FRP suppressed H2O2-induced neuronal damage in SK-N-SH cells via upregulated BDNF levels. Our findings demonstrated that FRP prevented memory impairment, and that the underlying mechanism might involve regulation of the ERK/CREB/BDNF signaling pathway. These results suggest FRP as a potential agent for the prevention of age-related cognitive decline and dementia.


Allergic and asthmatic reactions to alcoholic drinks.
Alcoholic drinks are capable of triggering a wide range of allergic and allergic-like responses, including rhinitis, itching, facial swelling, headache, cough and asthma. Limited epidemiological data suggests that many individuals are affected and that sensitivities occur to a variety of drinks, including wine, beer and spirits. In surveys of asthmatics, over 40% reported the triggering of allergic or allergic-like symptoms following alcoholic drink consumption and 30 - 35% reported worsening of their asthma. Sensitivity to ethanol itself can play a role in triggering adverse responses, particularly in Asians, which is due mainly to a reduced capacity to metabolize acetaldehyde. In Caucasians, specific non-alcohol components are the main cause of sensitivities to alcoholic drinks. Allergic sensitivities to specific components of beer, spirits and distilled liquors have been described. Wine is clearly the most commonly reported trigger for adverse responses. Sensitivities to wine appear to be due mainly to pharmacological intolerances to specific components, such as biogenic amines and the sulphite additives. Histamine in wine has been associated with the triggering of a wide spectrum of adverse symptoms, including sneezing, rhinitis, itching, flushing, headache and asthma. The sulphite additives in wine have been associated with triggering asthmatic responses. Clinical studies have confirmed sensitivities to the sulphites in wine in limited numbers of individuals, but the extent to which the sulphites contribute to wine sensitivity overall is not clear. The aetiology of wine-induced asthmatic responses may be complex and may involve several co-factors.


Mammography behavior after receiving a negative BRCA1 mutation test result in the Ashkenazim: a community-based study.
To define the impact of a negative BRCA1 test result on subsequent breast cancer screening behavior in women. Longitudinal study of a community-based sample of Ashkenazi Jews offered testing for the 185delAG BRCA1 mutation in 1996. Of 309 participants, 118 women were mutation negative, of average risk (based on family history of cancer), unaffected with breast cancer, and provided complete data at baseline, and Year 1 and Year 2 follow-up questionnaires. Women age 50 and older had 91.7% compliance with mammography for the year prior to entry (baseline), 88.3% during Year 1, 91.7% during Year 2 (no significant change; P = 0.775). Women under age 50 demonstrated an increase in mammography (49.2% at baseline, 62.7% Year 1, and 67.1% Year 2; P = 0.035). Both groups demonstrated significant decreases in breast cancer worry and perceived risk. Logistic regression analysis on having a mammogram at Year 2 showed that age, physician recommendation, worry, and perceived risk were all significant. Receipt of negative BRCA1 test results in a cohort of Ashkenazi Jewish women did not have a negative impact on mammography behavior 2 years after genetic testing.


Heterogeneity of the genetic risk in age-related macular disease: a population-based familial risk study.
To assess the extent of heterogeneity of the genetic risk of age-related macular disease (AMD) among families. Case-controlled population-based familial aggregation study. Participants comprised 190 first-degree relatives of 65 case probands and 347 relatives of 100 control probands. All probands had been identified from the baseline phase of the Rotterdam Study in The Netherlands. A family score was computed for each family based on the presence and type of macular disease, the expected risk of disease, and the number, extent of kinship, and age of all family members. Presence and stage of AMD as diagnosed on fundus photographs, family score, and logistic regression coefficient. The family score of case families showed a peak of approximately 0 with a skewed tail (14% of families) of higher than expected risks of disease toward a maximum of 2.9. The family score of control families centered on 0, apart from 1 outlier. The risk of AMD increased significantly with higher family scores (beta = 1.34; P<0.001). The heterogeneity of genetic risk among AMD families is considerable, and the proportion of high-risk families is relatively small. The family score method is relevant for genetic counseling as well as for implementation in studies of genetic dissection of AMD.


Reporting of suicide by the New Zealand media.
Rates of suicide in New Zealand are high compared with those of other countries. International evidence suggests that the reporting of suicide may influence rates of suicidal behavior. No research exists, however, on the reporting of suicide by New Zealand media. This study provides the first baseline picture of the reporting of suicide by New Zealand media. The overall objective was to use the findings to inform future development of media guidelines by the Ministry of Health. Newspaper, Internet, television and radio news items on suicide were collected over 12 months. Descriptive statistical data on the nature and extent of the reporting of suicide were generated through content analysis of applicable items. A random sample of 10% was then subjected to a quality analysis to determine whether items aligned with the Ministry of Health's guideline for the reporting of suicide. A total of 3,483 items were extracted, most of which reported on an individual's attempted or completed suicide, while suicide methods were not often mentioned. Few items focused on people overcoming their difficulties or provided information to assist people struggling with suicidal ideation. The reporting of suicide by New Zealand media was extensive and generally of good quality. Better collaboration between the media and mental health professionals is needed, however, to increase information supplied within items on support services. More succinct guidelines and increased journalist awareness of their existence would also contribute to the quality of reporting on suicide.


Parental Child-Rearing Conflicts Through Adolescence: Trajectories and Associations With Child Characteristics and Externalizing Patterns.
Research on longitudinal interparental conflict patterns and offspring development is scarce. The population-based TOPP study (N = 459) was used to investigate (1) child-rearing conflict trajectories through four time points during childhood and adolescence (ages 8-16), and (2) associations between conflict trajectories and child characteristics (i.e., birth order, gender, externalizing patterns from early childhood). Latent profile analysis identified six distinct trajectories. Conflict levels decreased for most respondents over the adolescent offspring period, but offspring's birth order and externalizing problems were related to less typical trajectories and higher levels of conflict. Onset of externalizing problems was of additional importance for the course of parental child-rearing conflicts. The results highlight the perception of the whole family as an interwoven system.


The 'Higgs' amplitude mode at the two-dimensional superfluid/Mott insulator transition.
Spontaneous symmetry breaking plays a key role in our understanding of nature. In relativistic quantum field theory, a broken continuous symmetry leads to the emergence of two types of fundamental excitation: massless Nambu-Goldstone modes and a massive 'Higgs' amplitude mode. An excitation of Higgs type is of crucial importance in the standard model of elementary particle physics, and also appears as a fundamental collective mode in quantum many-body systems. Whether such a mode exists in low-dimensional systems as a resonance-like feature, or whether it becomes overdamped through coupling to Nambu-Goldstone modes, has been a subject of debate. Here we experimentally find and study a Higgs mode in a two-dimensional neutral superfluid close to a quantum phase transition to a Mott insulating phase. We unambiguously identify the mode by observing the expected reduction in frequency of the onset of spectral response when approaching the transition point. In this regime, our system is described by an effective relativistic field theory with a two-component quantum field, which constitutes a minimal model for spontaneous breaking of a continuous symmetry. Additionally, all microscopic parameters of our system are known from first principles and the resolution of our measurement allows us to detect excited states of the many-body system at the level of individual quasiparticles. This allows for an in-depth study of Higgs excitations that also addresses the consequences of the reduced dimensionality and confinement of the system. Our work constitutes a step towards exploring emergent relativistic models with ultracold atomic gases.


Significant differences in the expected versus observed longevity of implantable cardioverter defibrillators (ICDs).
Implantable cardioverter defibrillator (ICD) is a life-saving therapy for patients at risk of ventricular arrhythmias. Due to its high cost, its cost-effectiveness is highly dependent on its longevity, which is currently only based upon the manufacturer's predicted device life span. We sought to assess the ICDs' longevity and the factors influencing it, and to compare the observed (real life) to the expected (manufacturer's prediction) life span at a device level. We retrospectively identified all patients who underwent an ICD implantation in a tertiary care medical center. For each device, an expected longevity was assigned based on the manufacturer/model, pacing percentage, and number of shocks per year. We defined device failure if the observed survival was shorter than 80 % of the expected. Only devices with follow-up time that exceeded the expected longevity were included. Of the 275 devices in the cohort, 79 (29 %) failed. Median device longevity was 5 years and varied markedly between manufacturers (4.3, 4.8, 5.1, and 6.3 years for Biotronik, St. Jude Medical, Boston Scientific, and Medtronic, respectively). There were significant differences among the manufacturers in device failure rates: 48, 17, 22, and 14 % for Biotronik, St. Jude Medical, Boston Scientific, and Medtronic, respectively). In multivariate analysis, manufacturer, earlier year of implantation, congestive heart failure and chronic renal failure significantly predicted device failure. In conclusion, there is a significant device failure rate among ICDs, with variability among manufacturers, impacting both patients and the medical economic systems.


Analysis of the DNA methylome and transcriptome in granulopoiesis reveals timed changes and dynamic enhancer methylation.
In development, epigenetic mechanisms such as DNA methylation have been suggested to provide a cellular memory to maintain multipotency but also stabilize cell fate decisions and direct lineage restriction. In this study, we set out to characterize changes in DNA methylation and gene expression during granulopoiesis using 4 distinct cell populations ranging from the oligopotent common myeloid progenitor stage to terminally differentiated neutrophils. We observed that differentially methylated sites (DMSs) generally show decreased methylation during granulopoiesis. Methylation appears to change at specific differentiation stages and overlap with changes in transcription and activity of key hematopoietic transcription factors. DMSs were preferentially located in areas distal to CpG islands and shores. Also, DMSs were overrepresented in enhancer elements and enriched in enhancers that become active during differentiation. Overall, this study depicts in detail the epigenetic and transcriptional changes that occur during granulopoiesis and supports the role of DNA methylation as a regulatory mechanism in blood cell differentiation.


A combined 3D-SIM/SMLM approach allows centriole proteins to be localized with a precision of ∼4-5 nm.
Centrioles are small barrel-shaped structures that form centrosomes and cilia [1]. Centrioles assemble around a central cartwheel comprising the Sas-6 and Ana2/STIL proteins. The amino termini of nine Sas-6 dimers form a central hub of ∼12 nm radius from which nine dimer spokes radiate, placing the Sas-6 carboxyl termini at the outer edge of the ∼60 nm radius cartwheel [2]. Several centriole proteins are distributed in a toroid around the cartwheel, and super-resolution light microscopy studies have measured the average radii of these ∼100-200 nm radius toroids with a 'precision' - or standard deviation (s.d. or 1σ) - of ±∼10-40 nm. The organization of Ana2/STIL within the cartwheel, however, has not been resolvable. Here, we develop methods to calculate the average toroidal radius of centriolar proteins in the ∼20-60 nm range with a s.d. of just ±∼4-5 nm, revealing that the amino and carboxyl termini of Ana2 are located in the outer cartwheel region.


Next-Generation CDK2/9 Inhibitors and Anaphase Catastrophe in Lung Cancer.
The first generation CDK2/7/9 inhibitor seliciclib (CYC202) causes multipolar anaphase and apoptosis in lung cancer cells with supernumerary centrosomes (known as anaphase catastrophe). We investigated a new and potent CDK2/9 inhibitor, CCT68127 (Cyclacel). CCT68127 was studied in lung cancer cells (three murine and five human) and control murine pulmonary epithelial and human immortalized bronchial epithelial cells. Robotic CCT68127 cell-based proliferation screens were used. Cells undergoing multipolar anaphase and inhibited centrosome clustering were scored. Reverse phase protein arrays (RPPAs) assessed CCT68127 effects on signaling pathways. The function of PEA15, a growth regulator highlighted by RPPAs, was analyzed. Syngeneic murine lung cancer xenografts (n = 4/group) determined CCT68127 effects on tumorigenicity and circulating tumor cell levels. All statistical tests were two-sided. CCT68127 inhibited growth up to 88.5% (SD = 6.4%, P < .003) at 1 μM, induced apoptosis up to 42.6% (SD = 5.5%, P < .001) at 2 μM, and caused G1 or G2/M arrest in lung cancer cells with minimal effects on control cells (growth inhibition at 1 μM: 10.6%, SD = 3.6%, P = .32; apoptosis at 2 μM: 8.2%, SD = 1.0%, P = .22). A robotic screen found that lung cancer cells with KRAS mutation were particularly sensitive to CCT68127 ( P = .02 for IC 50 ). CCT68127 inhibited supernumerary centrosome clustering and caused anaphase catastrophe by 14.1% (SD = 3.6%, P < .009 at 1 μM). CCT68127 reduced PEA15 phosphorylation by 70% (SD = 3.0%, P = .003). The gain of PEA15 expression antagonized and its loss enhanced CCT68127-mediated growth inhibition. CCT68127 reduced lung cancer growth in vivo ( P < .001) and circulating tumor cells ( P = .004). Findings were confirmed with another CDK2/9 inhibitor, CYC065. Next-generation CDK2/9 inhibition elicits marked antineoplastic effects in lung cancer via anaphase catastrophe and reduced PEA15 phosphorylation.


New insight into the role of MMP14 in metabolic balance.
Membrane-anchored matrix metalloproteinase 14 (MMP14) is involved broadly in organ development through both its proteolytic and signal-transducing functions. Knockout of Mmp14 (KO) in mice results in a dramatic reduction of body size and wasting followed by premature death, the mechanism of which is poorly understood. Since the mammary gland develops after birth and is thus dependent for its functional progression on systemic and local cues, we chose it as an organ model for understanding why KO mice fail to thrive. A global analysis of the mammary glands' proteome in the wild type (WT) and KO mice provided insight into an unexpected role of MMP14 in maintaining metabolism and homeostasis. We performed mass spectrometry and quantitative proteomics to determine the protein signatures of mammary glands from 7 to 11 days old WT and KO mice and found that KO rudiments had a significantly higher level of rate-limiting enzymes involved in catabolic pathways. Glycogen and lipid levels in KO rudiments were reduced, and the circulating levels of triglycerides and glucose were lower. Analysis of the ultrastructure of mammary glands imaged by electron microscopy revealed a significant increase in autophagy signatures in KO mice. Finally, Mmp14 silenced mammary epithelial cells displayed enhanced autophagy. Applied to a systemic level, these findings indicate that MMP14 is a crucial regulator of tissue homeostasis. If operative on a systemic level, these findings could explain how Mmp14KO litter fail to thrive due to disorder in metabolism.


Particulate matter exposure aggravates osteoarthritis severity.
Several diseases have been linked to particulate matter (PM) exposure. Outdoor activities, such as road running or jogging, are popular aerobic exercises due to few participatory limitations. Osteoarthritis (OA) is a progressive degenerative joint disease, usually observed at age 40, and not noticed before pain or diagnosis. Although exercise has health benefits, it is unclear whether outdoor jogging in higher PM (standard reference material 1649b, SRM 1649b) concentration environments could affect OA development or severity. Hence, a PM exposure monosodium iodoacetate (MIA)-induced OA animal jogged model was established for investigation. Results showed that high doses of PM (5 mg) significantly increased pro-inflammatory factors such as tumor necrosis factor α (TNF-α), interleukin (IL)-1β, and IL-6, and M1 macrophages in the lung region, also obtained in systemic IL-6 and TNF-α expressions in this MIA-OA rat model. Moreover, levels of osteocalcin, cartilage oligomeric matrix protein (COMP), and N-telopeptides of type I collagen were especially influenced in MIA+PM groups. Morphological and structural changes of the knee joint were detected by micro-computed tomography images (micro-CT) and immunohistochemistry. MIA + PM rats exhibited severe bone density decrease, cartilage wear, and structure damages, accompanied by lower levels of physical activity, than the sham group and groups receiving MIA or PM alone. The findings suggest that the severity of OA could be promoted by PM exposure with a PM concentration effect via systemic inflammatory mechanisms. To the best of our knowledge, this is the first study to provide direct effects of PM exposure on OA severity.


Small-angle light scattering: instrumental design and application to particle sizing.
A small-angle integrated light-scattering (SAILS) instrument was designed with the innovative use of a diffusing plate and a charge-coupled device (CCD) camera. In contrast to previous small-angle light-scattering instruments, SAILS has few optical surfaces, allowing the direct recovery of scattering data. Although this approach bypasses the need for aberration corrections that are due to lenses, geometric corrections still apply and are described in detail. The image on the diffusing plate, when photographed by the CCD camera, yields a digitized two-dimensional array, covering the observed scattering angles from 10 to 20 deg. The size distribution of the scattering particles can be obtained by a discrete inversion of the experimentally obtained intensity versus angle-scattering curve. The mean radii obtained from this inversion of SAILS data are compared with nominal sizes given by the manufacturer, and standard errors are computed. The results indicate that SAILS is an ideal instrument for the study of particulates and, because of its fast readout time, is suitable for studying aggregation phenomena. However, because of the limited Q range of SAILS it is currently not suited for the direct determination of particle diameters smaller than approximately 300 nm.


DMN Operational Synchrony Relates to Self-Consciousness: Evidence from Patients in Vegetative and Minimally Conscious States.
The default mode network (DMN) has been consistently activated across a wide variety of self-related tasks, leading to a proposal of the DMN's role in self-related processing. Indeed, there is limited fMRI evidence that the functional connectivity within the DMN may underlie a phenomenon referred to as self-awareness. At the same time, none of the known studies have explicitly investigated neuronal functional interactions among brain areas that comprise the DMN as a function of self-consciousness loss. To fill this gap, EEG operational synchrony analysis [1, 2] was performed in patients with severe brain injuries in vegetative and minimally conscious states to study the strength of DMN operational synchrony as a function of self-consciousness expression. We demonstrated that the strength of DMN EEG operational synchrony was smallest or even absent in patients in vegetative state, intermediate in patients in minimally conscious state and highest in healthy fully self-conscious subjects. At the same time the process of ecoupling of operations performed by neuronal assemblies that comprise the DMN was highest in patients in vegetative state, intermediate in patients in minimally conscious state and minimal in healthy fully self-conscious subjects. The DMN's frontal EEG operational module had the strongest decrease in operational synchrony strength as a function of selfconsciousness loss, when compared with the DMN's posterior modules. Based on these results it is suggested that the strength of DMN functional connectivity could mediate the strength of self-consciousness expression. The observed alterations similarly occurred across EEG alpha, beta1 and beta2 frequency oscillations. Presented results suggest that the EEG operational synchrony within DMN may provide an objective and accurate measure for the assessment of signs of self-(un)consciousness in these challenging patient populations. This method therefore, may complement the current diagnostic procedures for patients with severe brain injuries and, hence, the planning of a rational rehabilitation intervention.


The use of eye-tracking to explore social difficulties in cognitively able students with autism spectrum disorder: A pilot investigation.
Individuals with autism spectrum disorder do not just 'grow out of' their early difficulties in understanding the social world. Even for those who are cognitively able, autism-related difficulties continue into adulthood. Atypicalities attending to and interpreting communicative signals from others can provide barriers to success in education, employment and relationships. In the current study, we use eye-tracking during real social interaction to explore attention to social cues (e.g. face, eyes, mouth) and links to social awareness in a group of cognitively able University students with autism spectrum disorder and typically developing students from the same University. During the interaction, students with autism spectrum disorder showed less eye fixation and more mouth fixation than typically developing students. Importantly, while 63% of typically developing participants reported thinking they were deceived about the true nature of the interaction, only 9% of autism spectrum disorder participants picked up this subtle social signal. We argue that understanding how these social attentional and social awareness difficulties manifest during adulthood is important given the growing number of adults with autism spectrum disorder who are attending higher level education. These adults may be particularly susceptible to drop-out due to demands of coping in situations where social awareness is so important.


Measuring Behavior in the Home Cage: Study Design, Applications, Challenges, and Perspectives.
The reproducibility crisis (or replication crisis) in biomedical research is a particularly existential and under-addressed issue in the field of behavioral neuroscience, where, in spite of efforts to standardize testing and assay protocols, several known and unknown sources of confounding environmental factors add to variance. Human interference is a major contributor to variability both within and across laboratories, as well as novelty-induced anxiety. Attempts to reduce human interference and to measure more "natural" behaviors in subjects has led to the development of automated home-cage monitoring systems. These systems enable prolonged and longitudinal recordings, and provide large continuous measures of spontaneous behavior that can be analyzed across multiple time scales. In this review, a diverse team of neuroscientists and product developers share their experiences using such an automated monitoring system that combines Noldus PhenoTyper® home-cages and the video-based tracking software, EthoVision® XT, to extract digital biomarkers of motor, emotional, social and cognitive behavior. After presenting our working definition of a "home-cage", we compare home-cage testing with more conventional out-of-cage tests (e.g., the open field) and outline the various advantages of the former, including opportunities for within-subject analyses and assessments of circadian and ultradian activity. Next, we address technical issues pertaining to the acquisition of behavioral data, such as the fine-tuning of the tracking software and the potential for integration with biotelemetry and optogenetics. Finally, we provide guidance on which behavioral measures to emphasize, how to filter, segment, and analyze behavior, and how to use analysis scripts. We summarize how the PhenoTyper has applications to study neuropharmacology as well as animal models of neurodegenerative and neuropsychiatric illness. Looking forward, we examine current challenges and the impact of new developments. Examples include the automated recognition of specific behaviors, unambiguous tracking of individuals in a social context, the development of more animal-centered measures of behavior and ways of dealing with large datasets. Together, we advocate that by embracing standardized home-cage monitoring platforms like the PhenoTyper, we are poised to directly assess issues pertaining to reproducibility, and more importantly, measure features of rodent behavior under more ethologically relevant scenarios.


Reductions in cortisol associated with primary care brief mindfulness program for veterans with PTSD.
Patients with posttraumatic stress disorder (PTSD) have significant medical morbidity, which may be mediated by hypothalamic pituitary axis (HPA) dysfunction and reflected in cortisol output. Many veterans with PTSD are hesitant to engage in trauma-focused exposure treatments; therefore briefer, non-exposure-based treatments are needed; one such promising approach is an abbreviated Primary Care brief Mindfulness Program (PCbMP). This study investigated the relationship between dose-response to participation in a veterans PCbMP program and diurnal cortisol. Cortisol reflects HPA function and PTSD is associated with HPA dysregulation. Veterans with PTSD were identified in PC and randomly assigned to treatment as usual (TAU, n=21) or participation in brief 4-week Mindfulness Based Stress Reduction program (n=19). Veterans (n=40) (mean age, 48±16 y; 90% men) with PTSD referred through their VA PC provider and randomly assigned to PCbMP or TAU. As an objective indicator of HPA function, salivary diurnal cortisol was measured from samples collected across 2 consecutive days at baseline and follow-up. Analyses revealed that significant changes in cortisol were associated with PCbMP treatment engagement and dosing (number of mindfulness program sessions completed). Veterans completing 4 mindfulness-based meditation sessions significantly reduced their cortisol awakening response (P≤0.05); and had significant changes in cortisol area under the curve increase compared with TAU participants (P≤0.05). Results indicate that PCbMP has a beneficial physiological impact on veterans with PTSD with a minimum of 4 weeks of practice.


Systemic and splanchnic haemodynamic effects of pentifylline in rats with portal hypertension.
1. The systemic and splanchnic haemodynamic effects of pentifylline (40 mg/kg body weight intravenously) were assessed in rats with portal hypertension associated either with CCl4-induced cirrhosis (n = 13) or portal vein ligation (n = 13). 2. Heparinized catheters were placed into the portal vein, inferior vena cava, aorta and left ventricle with exits from the neck. Haemodynamic studies were performed 4 h after consciousness was regained. Cardiac output and regional blood flows were measured using radiolabelled microspheres and the reference sample method in seven rats in each group; portal-systemic shunting was measured using microsphere injection in the ileo-colic vein in six rats in each group. 3. Forty-five minutes after injection, pentifylline had no effect on mean arterial pressure, cardiac output, peripheral resistance, portal venous flow, hepatic artery flow or portal-systemic shunting in either group of rats with portal hypertension. The drug lowered portal pressure (-18%) in cirrhotic rats, but not in portal-vein-ligated rats. 4. These data demonstrate that pentifylline lowers portal pressure in cirrhotic rats without affecting portal venous flow and portal-systemic shunting; this effect is possibly mediated by changes in intrahepatic resistance related to the effects of pentifylline on blood viscosity and/or on intrahepatic vasomotor tone.


Role of inter-trial phase coherence in atypical auditory evoked potentials to speech and nonspeech stimuli in children with autism.
This autism study investigated how inter-trial phase coherence (ITPC) drives abnormalities in auditory evoked potential (AEP) responses for speech and nonspeech stimuli. Auditory P1-N2 responses and ITPCs in the theta band (4-7 Hz) for pure tones and words were assessed with EEG data from 15 school-age children with autism and 16 age-matched typically developing (TD) controls. The autism group showed enhanced P1 and reduced N2 for both speech and nonspeech stimuli in comparison with the TD group. Group differences were also found with enhanced theta ITPC for P1 followed by ITPC reduction for N2 in the autism group. The ITPC values were significant predictors of P1 and N2 amplitudes in both groups. Abnormal trial-to-trial phase synchrony plays an important role in AEP atypicalities in children with autism. ITPC-driven enhancement as well as attenuation in different AEP components may coexist, depending on the stage of information processing. It is necessary to examine the time course of auditory evoked potentials and the corresponding inter-trial coherence of neural oscillatory activities to better understand hyper- and hypo- sensitive responses in autism, which has important implications for sensory based treatment.


The impact of MTHFR 677C → T risk knowledge on changes in folate intake: findings from the Food4Me study.
It is hypothesised that individuals with knowledge of their genetic risk are more likely to make health-promoting dietary and lifestyle changes. The present study aims to test this hypothesis using data from the Food4Me study. This was a 6-month Internet-based randomised controlled trial conducted across seven centres in Europe where individuals received either general healthy eating advice or varying levels of personalised nutrition advice. Participants who received genotype-based personalised advice were informed whether they had the risk (CT/TT) (n = 178) or non-risk (CC) (n = 141) alleles of the methylenetetrahydrofolate reductase (MTHFR) gene in relation to cardiovascular health and the importance of a sufficient intake of folate. General linear model analysis was used to assess changes in folate intake between the MTHFR risk, MTHFR non-risk and control groups from baseline to month 6 of the intervention. There were no differences between the groups for age, gender or BMI. However, there was a significant difference in country distribution between the groups (p = 0.010). Baseline folate intakes were 412 ± 172, 391 ± 190 and 410 ± 186 μg per 10 MJ for the risk, non-risk and control groups, respectively. There were no significant differences between the three groups in terms of changes in folate intakes from baseline to month 6. Similarly, there were no changes in reported intake of food groups high in folate. These results suggest that knowledge of MTHFR 677C → T genotype did not improve folate intake in participants with the risk variant compared with those with the non-risk variant. ClinicalTrials.gov NCT01530139.


[Non-alcoholic fatty liver disease--new view].
Non-alcoholic fatty liver disease (NAFLD) covers a wide spectrum of liver pathology--from steatosis alone, through the necroinflammatory disorder of non-alcoholic steatohepatitis (NASH) to cirrhosis and liver cancer. NAFLD/NASH is mostly related with visceral adiposity, obesity, type 2 diabetes melitus (DM t.2) and metabolic syndrome. Pathogenetic concepts of NAFLD include overnutrition and underactivity, insulin resistance (IR) and genetic factor. The prevalence of NAFLD has been estimated to be 17-33% in some countries, NASH may be present in about 1/3 of such cases, while 20-25% of NASH cases could progress to cirrhosis. NAFLD is now recognized as one of the most frequent reason of liver tests elevation without clinical symptoms. Insulin resistance is considering as having a central role in NAFLD pathogenesis. In hepatocytes, IR is related to hyperglycaemia and hyperinsulinaemia, formation of advanced glycation end-products, increased free fatty acids and their metabolites, oxidative stress and altered profiles of adipocytokines. Early stages of fatty liver are clinically silent and include elevation of ALT and GGTP, hyperechogenic liver in USG and/or hepatomegaly. Among clinical symptoms, abdominal discomfort is relatively common as well as chronic fatigue. NAFLD/NASH is not a benign disease, progressive liver biopsy have shown histological progression of fibrosis in 32%, the estimated rate of cirrhosis development is 20% and a liver--related death is 12% over 10 years. No treatment has scientifically proved to ameliorate NAFLD or to avoid its progression. The various therapeutic alternatives are aimed at interfering with the risk factors involved in the pathogenesis of the disorder in order to prevent the progression to end-stage liver disease. The most important therapeutic measure is increasing insulin sensitivity by an attempt to change a lifestyle mostly by dieting and physical activity in order to loose weight. The most used agent is metformin, the others are under controlled trials or their effectiveness is low. NASH is not a common indication for liver transplantation because of the older age distribution of patients and high prevalence of comorbidity, related to metabolic syndrome. Recurence of NASH in the grafted liver is also a relatively frequent complication.


Retention of the Native Epigenome in Purified Mammalian Chromatin.
A protocol is presented for the isolation of native mammalian chromatin as fibers of 25-250 nucleosomes under conditions that preserve the natural epigenetic signature. The material is composed almost exclusively of histones and DNA and conforms to the structure expected by electron microscopy. All sequences probed for were retained, indicating that the material is representative of the majority of the genome. DNA methylation marks and histone marks resembled the patterns observed in vivo. Importantly, nucleosome positions also remained largely unchanged, except on CpG islands, where nucleosomes were found to be unstable. The technical challenges of reconstituting biochemical reactions with native mammalian chromatin are discussed.


Risk Factors for Developing Hyponatremia During Terlipressin Treatment: A Retrospective Analyses in Variceal Bleeding.
The aim of this study was to evaluate the risk factors and clinical significance of terlipressin-induced hyponatremia. Patients receiving terlipressin treatment frequently develop hyponatremia. However, its clinical significance and risk factors are not fully elucidated. Records of patients treated with terlipressin for variceal bleeding were analyzed. Hyponatremia was defined as a decrease in serum sodium (Na) level of >5 mEq/L from the baseline level; severe hyponatremia as a decrease in serum Na level of >10 mEq/L from the baseline level; and rapid severe hyponatremia as a decrease in serum Na level of >10 mEq/L within 3 days of treatment. The study involved 151 patients (mean age, 55.1±11.8 y) with male predominance (80.8%). Baseline serum Na and creatinine levels were 137.2±6.1 mEq/L and 0.9±0.4 mg/dL, respectively. Patients were treated with terlipressin for 4.5±1.9 days. Changes in serum Na levels from baseline were 0.4±4.1, -1.1±4.8, -4.0±7.0, -6.5±9.1, and -6.1±11.2 mEq/L, whereas the frequencies of hyponatremia and severe hyponatremia were 13.6%, 30.4%, 50.8%, 63.5%, and 66.9% and 0%, 8.8%, 23.3%, 33.0%, and 38.8% on days 1, 2, 3, 4, and 5 of treatment, respectively. Younger age, lower Child-Pugh score, higher serum Na level, and longer duration of terlipressin treatment were independent risk factors. Rapid severe hyponatremia developed in 29 patients (19.2%); lower body mass index was an additional risk factor in this group. Mortality was not associated with hyponatremia. Terlipressin-induced hyponatremia occurred frequently, especially in young patients with good liver function and higher Na level. Caution is required when administering terlipressin to patients with low body mass index.


Development of a common priority list of pharmaceuticals relevant for the water cycle.
Pharmaceutically active compounds (PhACs), including prescription drugs, over-the-counter medications, drugs used in hospitals and veterinary drugs, have been found throughout the water cycle. A desk study was initiated by the Global Water Research Coalition to consolidate a uniform selection of such compounds in order to judge risks of PhACs for the water cycle. By identifying major existing prioritization efforts and evaluating the criteria they use, this study yields a representative and qualitative profile ('umbrella view') of priority pharmaceuticals based on an extensive set of criteria. This can then be used for further studies on analytical methods, occurrence, treatability and potential risks associated with exposure to PhACs in water supply, identifying compounds most likely to be encountered and that may have significant impact on human health. For practical reasons, the present study excludes veterinary drugs. The pragmatic approach adopted provides an efficient tool to manage risks related to pharmaceuticals and provides assistance for selecting compounds for future studies.


Tumor localization and image registration of F-18 FDG coincidence detection scans with computed tomographic scans.
The aim of this study was to determine the feasibility of registering routine clinical F-18 fluorodeoxyglucose (FDG) coincidence detection (CD) scans with computed tomographic (CT) scans for radiation treatment planning and case management. F-18 FDG CD and chest CT scans, performed in 10 randomly selected patients with confirmed or possible adenocarcinoma of the lung, were evaluated. The quality of the matches was verified by comparisons of the center-to-center distance between a region of interest (ROI) manually drawn on the CT slice and warped onto the CD slice with an ROI drawn manually directly on the CD slice. In addition, the overlap between the two ROIs was calculated. All 10 F-18 FDG CD and CT scans were registered with good superimposition of soft tissue density on increased radionuclide activity. The center-to-center distance between the ROIs ranged from 0.29 mm to 8.08 mm, with an average center-to-center distance of 3.89 mm +/- 2.42 mm (0.69 pixels +/- 0.34 pixels). The ROI overlap ranged from 77% to 99%, with an average of 90% +/- 5.6%. Although the use of F-18 FDG CD shows great promise for the identification of tumors, it shares the same drawbacks as those associated with radiolabeled monoclonal antibody SPECT and ligand-based positron emission tomographic scans in that anatomic markers are limited. This study shows that image registration is feasible and may improve the clinical relevance of CD images.


Identification of genes related to floral organ development in pak choi by expression profiling.
Pak choi is a highly nutritious vegetable that is widely grown in China, Southeast Asia, and other parts of the world. Because it reproduces by seed, it is very important to understand the mechanism of floral organ development. Therefore, using the Chinese cabbage genome as a reference, this study analyzed the expression profiles of shoot apex genes at flower bud differentiation stages 1 and 5, in order to identify genes related to floral organ development. The results showed that the proportion of mapped genes was high, with 84.25 and 83.80% of clean reads from the two sample saligned to the reference genome, respectively. A total of 525 differentially expressed genes (DEGs) were identified, 224 of which were upregulated and 301 were downregulated. The expression levels of genes homologous to Chinese cabbage flowering genes were also analyzed at stages 1 and 5; the expression levels of Bra012997 (ap1), Bra000393 (SOC1), and Bra004928 (SOC1) were significantly upregulated at stage 5, suggesting that these three genes positively regulate floral development in pak choi. DEGs involved in floral organ development were analyzed with homologous genes from Arabidopsis thaliana; the homologous genes Bra029281 (AGL42), Bra026577 (ARPN), Bra022954 (SPL3), Bra029293 (ARF2), Bra007978 (AtRLP12), Bra033221 (SPL8), Bra008037 (LOX4), Bra001598 (IAA19), Bra003892 (PATL1), Bra038778 (AT4G21323), Bra025315 (KLCR2), and Bra013906 (DTX35) are directly related to floral organ development in Arabidopsis, suggesting that these genes have corresponding functions during flower organ development in pak choi, and could be candidates for further genetic research. These results provide a foundation for research on the molecular mechanism of flower organ development in pak choi and other Brassica rapa vegetables.


Stiffness and damping in postural control increase with age.
Upright balance is believed to be maintained through active and passive mechanisms, both of which have been shown to be impacted by aging. A compensatory balance response often observed in older adults is increased co-contraction, which is generally assumed to enhance stability by increasing joint stiffness. We investigated the effect of aging on standing balance by fitting body sway data to a previously developed postural control model that includes active and passive stiffness and damping parameters. Ten young (24 +/- 3 years) and seven older (75 +/- 5 years) adults were exposed during eyes-closed stance to perturbations consisting of lateral pseudorandom floor tilts. A least-square fit of the measured body sway data to the postural control model found significantly larger active stiffness and damping model parameters in the older adults. These differences remained significant even after normalizing to account for different body sizes between the young and older adult groups. An age effect was also found for the normalized passive stiffness, but not for the normalized passive damping parameter. This concurrent increase in active stiffness and damping was shown to be more stabilizing than an increase in stiffness alone, as assessed by oscillations in the postural control model impulse response.


Serotonin 5-HTTLPR Genotype Modulates Reactive Visual Scanning of Social and Non-social Affective Stimuli in Young Children.
Previous studies have documented the 5-HTTLPR polymorphisms as genetic variants that are involved in serotonin availability and also associated with emotion regulation and facial emotion processing. In particular, neuroimaging and behavioral studies of healthy populations have produced evidence to suggest that carriers of the Short allele exhibit heightened neurophysiological and behavioral reactivity when processing aversive stimuli, particularly in brain regions involved in fear. However, an additional distinction has emerged in the field, which highlights particular types of fearful information, i.e., aversive information which involves a social component versus non-social aversive stimuli. Although processing of each of these stimulus types (social and non-social) is believed to involve a subcortical neural system which includes the amygdala, evidence also suggests that the amygdala itself may be particularly responsive to socially significant environmental information, potentially due to the critical relevance of social information for humans. Examining individual differences in neurotransmitter systems which operate within this subcortical network, and in particular the serotonin system, may be critically informative for furthering our understanding of the neurobiological mechanisms underlying responses to emotional and affective stimuli. In the present study we examine visual scanning patterns in response to both aversive and positive images of a social or non-social nature in relation to 5-HTTLPR genotypes, in 49 children aged 4-7 years. Results indicate that children with at least one Short 5-HTTLPR allele spent less time fixating the threat-related non-social stimuli, compared with participants with two copies of the Long allele. Interestingly, a separate set of analyses suggests that carriers of two copies of the short 5-HTTLPR allele also spent less time fixating both the negative and positive non-social stimuli. Together, these findings support the hypothesis that genetically mediated differences in serotonin availability mediate behavioral responses to different types of emotional stimuli in young children.


Bromelain Inhibits SARS-CoV-2 Infection in VeroE6 Cells.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). The initial interaction between Transmembrane Serine Protease 2 (TMPRSS2) primed SARS-CoV-2 spike (S) protein and host cell receptor angiotensin-converting enzyme 2 (ACE-2) is a pre-requisite step for this novel coronavirus pathogenesis. Here, we expressed a GFP-tagged SARS-CoV-2 S-Ectodomain in Tni insect cells. That contained sialic acid-enriched N- and O-glycans. Surface resonance plasmon (SPR) and Luminex assay showed that the purified S-Ectodomain binding to human ACE-2 and immunoreactivity with COVID-19 positive samples. We demonstrate that bromelain (isolated from pineapple stem and used as a dietary supplement) treatment diminishes the expression of ACE-2 and TMPRSS2 in VeroE6 cells and dramatically lowers the expression of S-Ectodomain. Importantly, bromelain treatment reduced the interaction between S-Ectodomain and VeroE6 cells. Most importantly, bromelain treatment significantly diminished the SARS-CoV-2 infection in VeroE6 cells. Altogether, our results suggest that bromelain or bromelain rich pineapple stem may be used as an antiviral against COVID-19. Bromelain inhibits / cleaves the expression of ACE-2 and TMPRSS2Bromelain cleaves / degrades SARS-CoV-2 spike proteinBromelain inhibits S-Ectodomain binding and SARS-CoV-2 infection.


Fast and accurate genotype imputation in genome-wide association studies through pre-phasing.
The 1000 Genomes Project and disease-specific sequencing efforts are producing large collections of haplotypes that can be used as reference panels for genotype imputation in genome-wide association studies (GWAS). However, imputing from large reference panels with existing methods imposes a high computational burden. We introduce a strategy called 'pre-phasing' that maintains the accuracy of leading methods while reducing computational costs. We first statistically estimate the haplotypes for each individual within the GWAS sample (pre-phasing) and then impute missing genotypes into these estimated haplotypes. This reduces the computational cost because (i) the GWAS samples must be phased only once, whereas standard methods would implicitly repeat phasing with each reference panel update, and (ii) it is much faster to match a phased GWAS haplotype to one reference haplotype than to match two unphased GWAS genotypes to a pair of reference haplotypes. We implemented our approach in the MaCH and IMPUTE2 frameworks, and we tested it on data sets from the Wellcome Trust Case Control Consortium 2 (WTCCC2), the Genetic Association Information Network (GAIN), the Women's Health Initiative (WHI) and the 1000 Genomes Project. This strategy will be particularly valuable for repeated imputation as reference panels evolve.


Quantitative trait loci mapping of genome regions controlling permethrin resistance in the mosquito Aedes aegypti.
The mosquito Aedes aegypti is the principal vector of dengue and yellow fever flaviviruses. Permethrin is an insecticide used to suppress Ae. aegypti adult populations but metabolic and target site resistance to pyrethroids has evolved in many locations worldwide. Quantitative trait loci (QTL) controlling permethrin survival in Ae. aegypti were mapped in an F(3) advanced intercross line. Parents came from a collection of mosquitoes from Isla Mujeres, México, that had been selected for permethrin resistance for two generations and a reference permethrin-susceptible strain originally from New Orleans. Following a 1-hr permethrin exposure, 439 F(3) adult mosquitoes were phenotyped as knockdown resistant, knocked down/recovered, or dead. For QTL mapping, single nucleotide polymorphisms (SNPs) were identified at 22 loci with potential antixenobiotic activity including genes encoding cytochrome P450s (CYP), esterases (EST), or glutathione transferases (GST) and at 12 previously mapped loci. Seven antixenobiotic genes mapped to chromosome I, six to chromosome II, and nine to chromosome III. Two QTL of major effect were detected on chromosome III. One corresponds with a SNP previously associated with permethrin resistance in the para sodium channel gene and the second with the CCEunk7o esterase marker. Additional QTL but of relatively minor effect were also found. These included two sex-linked QTL on chromosome I affecting knockdown and recovery and a QTL affecting survival and recovery. On chromosome II, one QTL affecting survival and a second affecting recovery were detected. The patterns confirm that mutations in the para gene cause target-site insensitivity and are the major source of permethrin resistance but that other genes dispersed throughout the genome contribute to recovery and survival of mosquitoes following permethrin exposure.


Abnormal erythrocyte and renal frusemide-sensitive sodium transport in idiopathic calcium nephrolithiasis.
1. Anomalous transmembrane anion transport has been observed in erythrocytes of patients with idiopathic calcium nephrolithiasis. 2. To verify whether cation transport is also abnormal, we investigated the frusemide-sensitive Na+ efflux from Na(+)-loaded erythrocytes and the natriuretic response to acute intravenous frusemide administration in calcium oxalate renal stone formers. 3. Frusemide administration induced a statistically significant smaller increase in the fractional excretion of Na+ in patients than in control subjects. Abnormal kinetic properties of erythrocyte Na(+)-K(+)-2Cl- cotransport were observed in approximately 60% of stone formers. The Km for Na+ of Na(+)-K(+)-2Cl- co-transport correlated with urinary Ca2+ excretion. 4. The abnormal kinetic properties of Na(+)-K(+)-2Cl- co-transport may be relevant for stone formation, hampering renal Ca2+ reabsorption in the distal nephron and determining critical physicochemical conditions for calcium/oxalate crystallization.


An epigenetic regulatory element of the Nodal gene in the mouse and human genomes.
Nodal signaling plays critical roles during embryonic development. The Nodal gene is not expressed in adult tissues but is frequently activated in cancer cells, contributing to progression toward malignancy. Although several regulatory elements of the Nodal gene have been identified, the epigenetic mechanisms by which Nodal expression is regulated over the long term remain unclear. We found a region exhibiting dynamic changes in DNA methylation at approximately -3.0 kb to -0.4 kb upstream from the transcriptional start site (TSS) that we termed the epigenetic regulatory element (ERE). The ERE was unmethylated in mouse embryonic stem cells (mESCs) but became increasingly methylated in differentiated cells and tissues, concomitant with the downregulation of Nodal mRNA expression. In vitro reporter assays identified an Oct3/4 binding motif within the ERE, indicating that the ERE is responsible for the activation of Nodal in mESCs. Furthermore, the ERE was a target of differentiation-associated Polycomb silencing, and the chromatin condensed when mESCs differentiated to embryoid bodies (EBs). Pharmacological inhibition of PRC2 led to the reactivation of Nodal expression in EBs and mouse embryonic fibroblasts (MEFs). The ERE was also targeted by PRC2 in normal human cells. In NODAL-expressing human cancer cells, accumulation of EZH2 and trimethylation of H3K27 at the ERE were diminished. In conclusion, Nodal is epigenetically controlled through the ERE in the mouse embryo and human cells.


Determinants of the interindividual variability in response to antiplatelet drugs.
The aim of this review article is to discuss the main determinants of the interindividual variability in response to antiplatelet agents. The main sources of pharmacokinetic and pharmacodynamic variability are reviewed, with particular emphasis on aspirin and clopidogrel. The term 'resistance' is uninformative of the mechanism(s) underlying interindividual variability in response to these antiplatelet agents, and is potentially misleading. Increased awareness of the distinct factors potentially interfering with the desired antiplatelet effects of aspirin or clopidogrel, particularly avoidable drug interactions, may ultimately result in better patient management than requesting unnecessary costly tests of platelet function. Similarly, new studies addressing the interindividual variability in response to these antiplatelet agents should rely upon mechanism-based biochemical end-points rather than platelet aggregation measurements. As with any drug used to prevent atherothrombosis, treatment 'failure' can occur with aspirin or clopidogrel perhaps not surprisingly, given the multifactorial nature of atherothrombosis. There is no scientific basis for changing antiplatelet therapy in the face of a treatment 'failure', as we cannot be sure whether a second vascular event occurring in the same patient will reflect the same pathophysiological event that led to the first. Moreover, we have no controlled evidence that changing therapy is a more effective strategy than maintaining an evidence-based therapy.


Chemical characterization, antioxidant, anti-inflammatory and cytotoxic properties of bee venom collected in Northeast Portugal.
Bee venom (BV) or apitoxin is a complex mixture of substances with reported biological activity. In the present work, five bee venom samples obtained from Apis mellifera iberiensis from the Northeast Portugal (two different apiaries) were chemically characterized and evaluated for their antioxidant, anti-inflammatory and cytotoxic properties. The LC/DAD/ESI-MS(n) analysis of the samples showed that melittin was the most abundant compound, followed by phospholipase A2 and apamin. All the samples revealed antioxidant and anti-inflammatory activity but without a direct relation with any of the individual chemical components identified. The results highlight that there are specific concentrations (present in BV5) in which these compounds are more active. The BV samples showed similar cytotoxicity for all the tested tumour cell lines (MCF-7, NCI-H460, HeLa and HepG2), being MCF-7 and HeLa the most susceptible ones. Nevertheless, the studied samples seem to be suitable to treat breast, hepatocellular and cervical carcinoma because at the active concentrations, the samples were not toxic for non-tumour cells (PLP2). Regarding the non-small cell lung carcinoma, BV should be used under the toxic concentration for non-tumour cells. Overall, the present study corroborates the enormous bioactive potential of BV being the first report on samples from Portugal.


Effect of fibroblast donor cell age and cell cycle on development of bovine nuclear transfer embryos in vitro.
The effects of cell cycle stage and the age of the cell donor animal on in vitro development of bovine nuclear transfer embryos were investigated. Cultures of primary bovine fibroblasts were established from animals of various ages, and the in vitro life span of these cell lines was analyzed. Fibroblasts from both fetuses and calves had similar in vitro life spans of approximately 30 population doublings (PDs) compared with 20 PDs in fibroblasts obtained from adult animals. When fibroblasts from both fetuses and adult animals were cultured as a population, the percentage of cells in G1 increased linearly with time, whereas the percentage of S-phase cells decreased proportionately. Furthermore, the percentage of cells in G1 at a given time was higher in adult fibroblasts than in fetal fibroblasts. To study the individual cells from a population, a shake-off method was developed to isolate cells in G1 stage of the cell cycle and evaluate the cell cycle characteristics of both fetal and adult fibroblasts from either 25% or 100% confluent cultures. Irrespective of the age, the mean cell cycle length in isolated cells was shorter (9.6-15.5 h) than that observed for cells cultured as a population. Likewise, the length of the G1 stage in these isolated cells, as indicated by 5-bromo-deoxyuridine labeling, lasted only about 2-3 h. There were no differences in either the number of cells in blastocysts or the percentage of blastocysts between the embryos reconstructed with G1 cells from 25% or 100% confluent cultures of fetal or adult cell lines. This study suggests that there are substantial differences in cell cycle characteristics in cells derived from animals of different ages or cultured at different levels of confluence. However, these factors had no effect on in vitro development of nuclear transfer embryos.


Mitogenic activities of amino acid substitution mutants of staphylococcal enterotoxin B in human and mouse lymphocyte cultures.
Site-directed mutagenesis has been used to introduce amino acid substitutions at specific residues of the staphylococcal enterotoxin B (SEB) gene cloned from Staphylococcus aureus 10-275. The mitogenic activities of these derivatives were determined in two assay systems: (i) mouse spleen cells and (ii) a mixture of human peripheral blood mononuclear cells and lymphocytes. Substitution of either His-12, His-32, His-121, His-166, Lys-152, or Gly-205 did not significantly alter the mitogenic activity from that of the wild-type toxin in either proliferation assay. Substitution of either residue Asn-23, Phe-44, or Cys-93 reduced the mitogenicity of SEB by a degree that depended upon the assay system used. Similar to the results reported by others measuring toxin activation of mouse lymphoid cells, we found that substitutions of these three residues of SEB caused at least 800-fold reductions of mitogenic activity from that of the wild-type toxin. When tested for toxicity in vivo in D-galactosamine-treated mice, the reduced activities of these mutant toxins, however, were not as pronounced. In contrast, when tested in the human cell mitogenicity assay, these mutant toxins were active. Small alterations in activity (two- to fivefold reduction) were observable only at low concentrations. Our findings reveal the importance of using human lymphocytes in addition to the traditional mouse spleen cell assay when assessing biological activities of staphylococcal enterotoxins.


MUC1 enhances invasiveness of pancreatic cancer cells by inducing epithelial to mesenchymal transition.
Increased motility and invasiveness of pancreatic cancer cells are associated with epithelial to mesenchymal transition (EMT). Snai1 and Slug are zinc-finger transcription factors that trigger this process by repressing E-cadherin and enhancing vimentin and N-cadherin protein expression. However, the mechanisms that regulate this activation in pancreatic tumors remain elusive. MUC1, a transmembrane mucin glycoprotein, is associated with the most invasive forms of pancreatic ductal adenocarcinomas (PDA). In this study, we show that over expression of MUC1 in pancreatic cancer cells triggers the molecular process of EMT, which translates to increased invasiveness and metastasis. EMT was significantly reduced when MUC1 was genetically deleted in a mouse model of PDA or when all seven tyrosines in the cytoplasmic tail of MUC1 were mutated to phenylalanine (mutated MUC1 CT). Using proteomics, RT-PCR and western blotting, we revealed a significant increase in vimentin, Slug and Snail expression with repression of E-Cadherin in MUC1-expressing cells compared with cells expressing the mutated MUC1 CT. In the cells that carried the mutated MUC1 CT, MUC1 failed to co-immunoprecipitate with β-catenin and translocate to the nucleus, thereby blocking transcription of the genes associated with EMT and metastasis. Thus, functional tyrosines are critical in stimulating the interactions between MUC1 and β-catenin and their nuclear translocation to initiate the process of EMT. This study signifies the oncogenic role of MUC1 CT and is the first to identify a direct role of the MUC1 in initiating EMT during pancreatic cancer. The data may have implications in future design of MUC1-targeted therapies for pancreatic cancer.


Observations on the effect of chimeric anti-CD4 monoclonal antibody in patients with mycosis fungoides.
Chimeric (murine/human) anti-CD4 monoclonal antibody was infused into seven patients with mycosis fungoides. Successive patients received doses of 10, 20, 40, and 80 mg of antibody twice a week for 3 consecutive weeks. All patients had some clinical improvement, but responses were of relatively short duration. Serum levels of chimeric antibody varied as a function of dose. At the 80-mg dose level, antibody was readily observed in biopsied skin lesions. Although there was coating by antibody of most CD4 positive cells in the blood, there was no significant depletion of CD4 positive cells. Low-level antibody responses against the mouse Ig variable region and human Ig allotypic constant region determinants were observed in several patients, but none were of clinical significance. All but two patients made primary antibody and T-cell proliferative responses to a simultaneously administered foreign protein test antigen. However, there was marked suppression of the mixed lymphocyte reaction. We conclude that at the dose levels studied, a chimeric anti-CD4 monoclonal antibody (1) had some clinical efficacy against mycosis fungoides; (2) was well tolerated; (3) had a low level of immunogenicity; (4) had immediate immunosuppressive effects; and (5) did not induce tolerance to a co-injected antigen.


Priming of hippocampal population bursts by individual perisomatic-targeting interneurons.
Hippocampal population bursts ("sharp wave-ripples") occur during rest and slow-wave sleep and are thought to be important for memory consolidation. The cellular mechanisms involved are incompletely understood. Here we investigated the cellular mechanisms underlying the initiation of sharp waves using a hippocampal slice model. To this end, we used a combination of field recordings with planar multielectrode arrays and whole-cell patch-clamp recordings of individual anatomically identified pyramidal neurons and interneurons. We found that GABA(A) receptor-mediated inhibition is necessary for sharp wave generation. Moreover, the activity of individual perisomatic-targeting interneurons can both suppress, and subsequently enhance, the local generation of sharp waves. Finally, we show that this is achieved by the tight control of local excitation and inhibition by perisomatic-targeting interneurons. These results suggest that perisomatic-targeting interneurons assist in selecting the subset of pyramidal neurons that initiate each hippocampal sharp wave-ripple.


Integrative analysis of Multiple Sclerosis using a systems biology approach.
Multiple sclerosis (MS) is a chronic autoimmune disorder characterized by inflammatory-demyelinating events in the central nervous system. Despite more than 40 years of MS research its aetiology remains unknown. This study aims to identify the most frequently reported and consistently regulated molecules in MS in order to generate molecular interaction networks and thereby leading to the identification of deregulated processes and pathways which could give an insight of the underlying molecular mechanisms of MS. Driven by an integrative systems biology approach, gene-expression profiling datasets were combined and stratified into "Non-treated" and "Treated" groups and additionally compared to other disease patterns. Molecular identifiers from dataset comparisons were matched to our Multiple Sclerosis database (MuScle; www.padb.org/muscle ). From 5079 statistically significant molecules, correlation analysis within groups identified a panel of 16 high-confidence genes unique to the naïve MS phenotype, whereas the "Treated" group reflected a common pattern associated with autoimmune disease. Pathway and gene-ontology clustering identified the Interferon gamma signalling pathway as the most relevant amongst all significant molecules, and viral infections as the most likely cause of all down-stream events observed. This hypothesis-free approach revealed the most significant molecular events amongst different MS phenotypes which can be used for further detailed studies.


Importance of folate-homocysteine homeostasis during early embryonic development.
Although the beneficial effects of maternal folate supplementation in the periconceptional period have been shown to prevent neural tube defects, congenital heart defects and orofacial clefts, the exact protective mechanism of folates remains unknown. Folates affect DNA synthesis, amino acid metabolism and methylation of genes, proteins and lipids via S-adenosylmethionine-mediated one-carbon transfer reactions. Our laboratory has created several mouse knock out models of folate transport using gene targeting to inactivate folate receptor 1 (Folr1), folate receptor 2 (Folr2) and reduced folate carrier 1 (Slc19a1) genes. Gene ablation of both Folr1 and Slc19a1 leads to lethality, but with maternal folate supplementation, nullizygous embryos for both genes present with neural tube defects (NTDs) and congenital heart defects (CHDs). Folr1 nullizygous mice also exhibit orofacial clefts when the dams are provided with low folate supplementation during pregnancy. Finally, women with NTD-affected pregnancies have been reported to have high autoantibody titers against the folate receptor, potentially inhibiting the transport of folate to the developing embryo. This may be an explanation for some of the folate-responsive NTDs and perhaps other congenital malformations. Herein, we propose how homocysteinylation of the folate receptor may contribute to generation of these autoantibodies against the folate receptor.


Awareness of Clenching and Underweight are Risk Factors for Onset of Crowding in Young Adults: A Prospective 3-Year Cohort Study.
Bruxism is a parafunctional activity that can seriously affect quality of life. Although bruxism induces many problems in the oral and maxillofacial area, whether it contributes to the onset of malocclusion remains unclear. The purpose of this prospective cohort study was to investigate the association between the onset of malocclusion and awareness of clenching during the daytime in young adults. Among 1,092 Okayama University students who underwent normal occlusion at baseline, we analysed 238 who had undergone a dental examination and had complete data after 3 years (2013⁻2016). We also performed subgroup analysis to focus on the association between awake bruxism and the onset of crowding (n = 216). Odds ratios (ORs) were calculated using multivariate logistic regression analyses. The incidences of malocclusion and crowding were 53.8% and 44.5%, respectively. In multivariate logistic regression, awareness of clenching was a risk factor for crowding (OR: 3.63; 95% confidence interval [CI]: 1.08⁻12.17). Moreover, underweight (body mass index < 18.5 kg/m²) was related to the onset of malocclusion (OR: 2.34; 95%CI: 1.11⁻4.92) and crowding (OR: 2.52, 95%CI: 1.25⁻5.76). These results suggest that awareness of clenching during the daytime and underweight are risk factors for the onset of crowding in young adults.


Dissociation between humoral and cellular immune responses to lens antigens.
Rabbits immunized against lens extracts from various other (xenogeneic) species reacted well with the immunizing lens extracts by immunological assays measuring either humoral or cellular immune responses. A dissociation was observed, however, between the humoral and cellular immune responses when lenses from rabbit (allogeneic) or from nonimmunizing species were tested. Antibodies from these rabbits cross-reacted with lenses from rabbit or all other tested species, with levels similar to those obtained with the immunizing lens extracts. The antibodies were determined by both serological tests and the Arthus skin reaction. On the other hand, these rabbits failed to cross-react with rabbit or nonimmunizing animal lens extracts by the cell-mediated delayed skin reaction, and their lymphocytes reacted poorly in culture with the nonimmunizing lenses. A similar dissociation between the humoral and cellular responses was found in rabbits immunized against allogeneic lens. Most of these rabbits produced moderate levels of lens antibodies demonstrated by the serological tests and the Arthus skin reaction but failed to exhibit delayed-type hypersensitivity, and their lymphocyte cultures reacted poorly or not at all to the rabbit or other lenses. The results are discussed in view of their relevance to the hypotheses concerning the pathogenesis of phacogenic uveitis.


Feeding habits of the speckled guitarfish Rhinobatos glaucostigma (Elasmobranchii: Rhinobatidae).
The stomachs of 464 speckled guitarfish Rhinobatos glaucostigma were sampled from the south-eastern Gulf of California (GC) to determine diet composition. Numerical indices and prey-specific index of relative importance ((%I)PSIR ) were used to determine the feeding strategy of the species. An analysis of similarity (ANOSIM) was used to determine differences in diet with respect to sex, season (dry or rainy) and maturity stages (immature or mature). The diversity and niche breadth (by sex, season and maturity) and a general trophic level were determined. The overall diet was dominated by shrimps ((%I)PSIR = 43.47), amphipods ((%I)PSIR = 18.89) and crabs ((%I)PSIR = 18.07). ANOSIM demonstrated differences in the diet by maturity and season, but not by sex. Rainy and dry season diets were dominated by shrimps and amphipods, respectively. Immature specimens fed mainly on amphipods, whereas mature fish preferred shrimps and crabs. Rhinobatos glaucostigma showed a narrow niche breadth with an intermediary trophic level (TL = 3.72) and can be considered as a secondary consumer in the soft-bottom demersal community of the south-east GC. Understanding the feeding habits and trophic level of R. glaucostigma is vital to help identify the segments of the population vulnerable to overfishing by artisanal and industrial fisheries, and to aid in conservation and management of this elasmobranch.


Impact of age and gender on the prevalence and prognostic importance of the metabolic syndrome and its components in Europeans. The MORGAM Prospective Cohort Project.
To investigate the influence of age and gender on the prevalence and cardiovascular disease (CVD) risk in Europeans presenting with the Metabolic Syndrome (MetS). Using 36 cohorts from the MORGAM-Project with baseline between 1982-1997, 69094 men and women aged 19-78 years, without known CVD, were included. During 12.2 years of follow-up, 3.7%/2.1% of men/women died due to CVD. The corresponding percentages for fatal and nonfatal coronary heart disease (CHD) and stroke were 8.3/3.8 and 3.1/2.5. The prevalence of MetS, according to modified definitions of the International Diabetes Federation (IDF) and the revised National Cholesterol Education Program-Adult Treatment Panel III (NCEP-ATPIII), increased across age groups for both genders (P<0.0001); with a 5-fold increase in women from ages 19-39 years to 60-78 years (7.4%/7.6% to 35.4%/37.6% for IDF/NCEP-ATPIII) and a 2-fold increase in men (5.3%/10.5% to 11.5%/21.8%). Using multivariate-adjusted Cox regressions, the associations between MetS and all three CVD events were significant (P<0.0001). For IDF/NCEP-ATPIII in men and women, hazard ratio (HR) for CHD was 1.60/1.62 and 1.93/2.03, for CVD mortality 1.73/1.65 and 1.77/2.06, and for stroke 1.51/1.53 and 1.58/1.77. Whereas in men the HRs for CVD events were independent of age (MetS*age, P>0.05), in women the HRs for CHD declined with age (HRs 3.23/3.98 to 1.55/1.56; MetS*age, P=0.01/P=0.001 for IDF/NCEP-ATPIII) while the HRs for stroke tended to increase (HRs 1.31/1.25 to 1.55/1.83; MetS*age, P>0.05). In Europeans, both age and gender influenced the prevalence of MetS and its prognostic significance. The present results emphasise the importance of being critical of MetS in its current form as a marker of CVD especially in women, and advocate for a redefinition of MetS taking into account age especially in women.


Direct interaction between uracil-DNA glycosylase and a proliferating cell nuclear antigen homolog in the crenarchaeon Pyrobaculum aerophilum.
Proliferating cell nuclear antigen (PCNA) acts as a sliding clamp on duplex DNA. Its homologs, present in Eukarya and Archaea, are part of protein complexes that are indispensable for DNA replication and DNA repair. In Eukarya, PCNA is known to interact with more than a dozen different proteins, including a human major nuclear uracil-DNA glycosylase (hUNG2) involved in immediate postreplicative repair. In Archaea, only three classes of PCNA-binding proteins have been reported previously: replication factor C (the PCNA clamp loader), family B DNA polymerase, and flap endonuclease. In this study, we report a direct interaction between a uracil-DNA glycosylase (Pa-UDGa) and a PCNA homolog (Pa-PCNA1), both from the hyperthermophilic crenarchaeon Pyrobaculum aerophilum (T(opt) = 100 degrees C). We demonstrate that the Pa-UDGa-Pa-PCNA1 complex is thermostable, and two hydrophobic amino acid residues on Pa-UDGa (Phe(191) and Leu(192)) are shown to be crucial for this interaction. It is interesting to note that although Pa-UDGa has homologs throughout the Archaea and bacteria, it does not share significant sequence similarity with hUNG2. Nevertheless, our results raise the possibility that Pa-UDGa may be a functional analog of hUNG2 for PCNA-dependent postreplicative removal of misincorporated uracil.


Urinary tract infections attributed to diverse ExPEC strains in food animals: evidence and data gaps.
Between 70 and 95% of urinary tract infections (UTI) are caused by strains of Escherichia coli. These strains, often termed Extraintestinal Pathogenic E. coli (ExPEC), possess specific virulence traits allowing them to colonize more inhospitable environments, such as the urogenital tract. Some ExPEC isolates from humans have similar virulence factor profiles to ExPEC isolates from animals, and because of the potential for these strains to cause UTI in people, these infections have been referred to as foodborne UTI, or FUTI. Finding similarities in ExPEC in animals and humans is not necessarily proof of transmission, particularly a unidirectional pathway from animals to humans; similarities in virulence factor profiles should be expected given the specific bacterial requirements for colonizing physiological compartments with similar characteristics in all animals. Many of the most important strains of human ExPEC globally, such as ST131, are highly virulent and clonal implying routes of transmission other than food. Documenting routes of transmission is particularly difficult due to the wide range of potential ExPEC sources, including the human intestinal tract, and non-human reservoirs such as food animals and retail meat products, sewage and other environmental sources, and companion animals. The significant environmental reservoir of ExPEC, including strains such as ST131, could potentially explain much more completely the global dissemination of virulent ExPEC clones and the rapid dissemination of new strains within the community. Taken in its totality, the link between ExPEC in animals and UTI in humans might exist, but studies conducted to date do not enable an estimation of the relative importance of this route of transmission. To reduce the burden of illness associated with ExPEC, the scientific community needs to push forward with ecologically-based, scientifically-sound study designs that can address the plethora of ways in which E. coli can spread.


An empirical comparison of meta-analysis and mega-analysis of individual participant data for identifying gene-environment interactions.
For analysis of the main effects of SNPs, meta-analysis of summary results from individual studies has been shown to provide comparable results as "mega-analysis" that jointly analyzes the pooled participant data from the available studies. This fact revolutionized the genetic analysis of complex traits through large GWAS consortia. Investigations of gene-environment (G×E) interactions are on the rise since they can potentially explain a part of the missing heritability and identify individuals at high risk for disease. However, for analysis of gene-environment interactions, it is not known whether these methods yield comparable results. In this empirical study, we report that the results from both methods were largely consistent for all four tests; the standard 1 degree of freedom (df) test of main effect only, the 1 df test of the main effect (in the presence of interaction effect), the 1 df test of the interaction effect, and the joint 2 df test of main and interaction effects. They provided similar effect size and standard error estimates, leading to comparable P-values. The genomic inflation factors and the number of SNPs with various thresholds were also comparable between the two approaches. Mega-analysis is not always feasible especially in very large and diverse consortia since pooling of raw data may be limited by the terms of the informed consent. Our study illustrates that meta-analysis can be an effective approach also for identifying interactions. To our knowledge, this is the first report investigating meta-versus mega-analyses for interactions.


Recurrent insect outbreaks caused by temperature-driven changes in system stability.
Insects often undergo regular outbreaks in population density but identifying the causal mechanism for such outbreaks in any particular species has proven difficult. Here, we show that outbreak cycles in the tea tortrix Adoxophyes honmai can be explained by temperature-driven changes in system stability. Wavelet analysis of a 51-year time series spanning more than 200 outbreaks reveals a threshold in outbreak amplitude each spring when temperature exceeds 15°C and a secession of outbreaks each fall as temperature decreases. This is in close agreement with our independently parameterized mathematical model that predicts the system crosses a Hopf bifurcation from stability to sustained cycles as temperature increases. These results suggest that temperature can alter system stability and provide an explanation for generation cycles in multivoltine insects.


Capsular polysaccharide from Actinobacillus actinomycetemcomitans inhibits IL-6 and IL-8 production in human gingival fibroblast.
We previously reported that a capsular polysaccharide (CP) from Actinobacillus actinomycetemcomitans Y4 induces bone resorption in a mouse organ culture system and osteoclast formation in mouse bone marrow cultures. However, the effects of A. actinomycetemcomitans Y4 CP on human gingival fibroblasts (HGF) are still unclear. The present study was undertaken to test the hypothesis that A. actinomycetemcomitans Y4 CP alters the production of inflammatory cytokines, such as interleukin-6 (IL-6) and IL-8 by HGF. When HGF were cultured with various concentrations of Y4 CP for 24 h, IL-6 and IL-8 production decreased in a concentration-dependent manner. Y4 CP (100 microg/ml) suppressed the release of IL-6 from 9.09 +/- 0.08 ng/ml to 0.34 +/- 0.21 ng/ml (P < 0.01) and IL-8 production decreased from 3.76 +/- 0.03 ng/ml to 0.09 +/- 0.01 ng/ml (P < 0.01). Y4 CP suppressed 70-80% of the release of IL-6 and IL-8 from HGF stimulated with Y4 lipopolysaccharide (LPS), too. Interestingly, anti-A. actinomycetemcomitans Y4 CP completely inhibited the effect of A. actinomycetemcomitans Y4 CP on IL-6 and IL-8 production from HGF. These results indicate that Y4 CP inhibits the release of IL-6 and IL-8 from HGF, suggesting that A. actinomycetemcomitans Y4 modulates the inflammatory response in periodontitis. Remarkably, this inhibitory effect was reversed by specific anti-A. actinomycetemcomitans Y4 CP suggesting an important relationship between the organism and the humoral host response.


Effects of alcohol septal ablation on coronary microvascular function and myocardial energetics in hypertrophic obstructive cardiomyopathy.
This study investigated the effects of alcohol septal ablation (ASA) on microcirculatory function and myocardial energetics in patients with hypertrophic cardiomyopathy (HCM) and left ventricular outflow tract (LVOT) obstruction. In 15 HCM patients who underwent ASA, echocardiography was performed before and 6 mo after the procedure to assess the LVOT gradient (LVOTG). Additionally, [(15)O]water PET was performed to obtain resting myocardial blood flow (MBF) and coronary vasodilator reserve (CVR). Changes in LV mass (LVM) and volumes were assessed by cardiovascular magnetic resonance imaging. Myocardial oxygen consumption (MVo(2)) was evaluated by [(11)C]acetate PET in a subset of seven patients to calculate myocardial external efficiency (MEE). After ASA, peak LVOTG decreased from 41 ± 32 to 23 ± 19 mmHg (P = 0.04), as well as LVM (215 ± 74 to 169 ± 63 g; P < 0.001). MBF remained unchanged (0.94 ± 0.23 to 0.98 ± 0.15 ml·min(-1)·g(-1); P = 0.45), whereas CVR increased (2.55 ± 1.23 to 3.05 ± 1.24; P = 0.05). Preoperatively, the endo-to-epicardial MBF ratio was lower during hyperemia compared with rest (0.80 ± 0.18 vs. 1.18 ± 0.15; P < 0.001). After ASA, the endo-to-epicardial hyperemic (h)MBF ratio increased to 1.03 ± 0.26 (P = 0.02). ΔCVR was correlated to ΔLVOTG (r = -0.82; P < 0.001) and ΔLVM (r = -0.54; P = 0.04). MEE increased from 15 ± 6 to 20 ± 9% (P = 0.04). Coronary microvascular dysfunction in obstructive HCM is at least in part reversible by relief of LVOT obstruction. After ASA, hMBF and CVR increased predominantly in the subendocardium. The improvement in CVR was closely correlated to the absolute reduction in peak LVOTG, suggesting a pronounced effect of LV loading conditions on microvascular function of the subendocardium. Furthermore, ASA has favorable effects on myocardial energetics.


The effect of the global positioning system on the driving performance of people with mild Alzheimer's disease.
The initial symptoms of Alzheimer's disease (AD) include attention deficits, memory loss and deficiencies in topographic and spatial orientation. People with AD may have way-finding difficulties in driving due to the deterioration of their navigation ability. Although the use of the Global Positioning System (GPS) has been proven to be a useful aid for older people in driving, there is no evidence to suggest that the benefit could extend to drivers with AD. This study aims to investigate the effectiveness of the GPS in assisting drivers with mild AD in finding their destination safely. Twenty-eight drivers with mild to very mild AD, diagnosed by a general practitioner or a psychogeriatrician, completed all clinical and psychometric assessments including the Mini Mental State Examination, Trail Making Test B and Doors and People Test. During the driving assessment, 3 driving trials with different settings (normal, visual-only and audio-only) of the GPS were administered to all participants. The participants were required to follow instructions from the GPS and perform a variety of driving tasks on a driving simulator. The driving performances of participants were assessed by criteria specific to AD drivers. The driving assessment criteria were first combined to form the overall driving performance score: a higher score indicated a better overall driving performance. The other outcome of this study was the success of participants to navigate to a predetermined destination. The driving performance of participants was different among the 3 settings (F = 72.8, p < 0.001) and the pairwise comparison between the 3 settings was significant (p < 0.001). The driving performance score was highest in the audio-only setting (mean: 20.0, SD: 2.2), moderate in the normal setting (mean: 16.7, SD: 2.6) and lowest in the visual-only setting (mean: 14.3, SD: 3.3). When compared with the visual-only setting, drivers using the audio-only setting (OR: 37.2, 95% CI: 9.2-151.2) and normal setting (OR: 37.2, 95% CI: 4.8-286.9) were more likely to successfully find their destination (p < 0.05). This study has found that using single, simple auditory instructions with the absence of the visual output of the GPS could potentially help people with mild AD to improve their driving ability and reach their destination.