Quantification of ligand-regulated nuclear receptor corepressor and coactivator binding, key interactions determining ligand potency and efficacy for the thyroid hormone receptor.
The potency and efficacy of ligands for nuclear receptors (NR) result both from the affinity of the ligand for the receptor and from the affinity that various coregulatory proteins have for ligand-receptor complexes; the latter interaction, however, is rarely quantified. To understand the molecular basis for ligand potency and efficacy, we developed dual time-resolved fluorescence resonance energy transfer (tr-FRET) assays and quantified binding of both ligand and coactivator or corepressor to the thyroid hormone receptor (TR). Promoter-bound TR exerts dual transcriptional regulatory functions, recruiting corepressor proteins and repressing transcription in the absence of thyroid hormones (THs) and shedding corepressors in favor of coactivators upon binding agonists, activating transcription. Our tr-FRET assays involve a TRE sequence labeled with terbium (fluorescence donor), TRbeta.RXRalpha heterodimer, and fluorescein-labeled NR interaction domains of coactivator SRC3 or corepressor NCoR (fluorescence acceptors). Through coregulator titrations, we could determine the affinity of SRC3 or NCoR for TRE-bound TR.RXR heterodimers, unliganded or saturated with different THs. Alternatively, through ligand titrations, we could determine the relative potencies of different THs. The order of TR agonist potencies is as follows: GC-1 approximately T 3 approximately TRIAC approximately T 4 >> rT 3 (for both coactivator recruitment and corepressor dissociation); the affinities of SRC3 binding to TR-ligand complexes followed a similar trend. This highlights the fact that the low activity of rT 3 is derived both from its low affinity for TR and from the low affinity of SRC for the TR-rT 3 complex. The TR antagonist NH-3 failed to induce SRC3 recruitment but did effect NCoR dissociation. These assays provide quantitative information about the affinity of two key interactions that are determinants of NR ligand potency and efficacy.


Something in the way we move: Motion dynamics, not perceived sex, influence head movements in conversation.
During conversation, women tend to nod their heads more frequently and more vigorously than men. An individual speaking with a woman tends to nod his or her head more than when speaking with a man. Is this due to social expectation or due to coupled motion dynamics between the speakers? We present a novel methodology that allows us to randomly assign apparent identity during free conversation in a video-conference, thereby dissociating apparent sex from motion dynamics. The method uses motion-tracked synthesized avatars that are accepted by naive participants as being live video. We find that 1) motion dynamics affect head movements but that apparent sex does not; 2) judgments of sex are driven almost entirely by appearance; and 3) ratings of masculinity and femininity rely on a combination of both appearance and dynamics. Together, these findings are consistent with the hypothesis of separate perceptual streams for appearance and biological motion. In addition, our results are consistent with a view that head movements in conversation form a low level perception and action system that can operate independently from top-down social expectations.


Combined microspectrophotometry and automated quantitative autoradiography applied to the analysis of the plant cell cycle.
Two methods, which relate grain number to cell cycle phase in Feulgen-stained autoradiographic preparations, have been developed and compared. Both methods automate grain number estimations, one by taking integrated absorbance measurements at different wavelengths, the other by measuring absorption at a single wavelength before and after chemical removal of silver grains. With tritium-labelled tobacco mosaic virus as a probe, a quantitative analysis has been made of the binding of virus particles to plant protoplasts in different compartments of the DNA replication and partition cycle. The preliminary results indicate that the quantity of virus bound by protoplasts is related to their cell cycle phase. Whilst in this case, the methods have been used with plant cells, both techniques are equally applicable to animal cells.


TMEM106B regulates progranulin levels and the penetrance of FTLD in GRN mutation carriers.
To determine whether TMEM106B single nucleotide polymorphisms (SNPs) are associated with frontotemporal lobar degeneration (FTLD) in patients with and without mutations in progranulin (GRN) and to determine whether TMEM106B modulates GRN expression. We performed a case-control study of 3 SNPs in TMEM106B in 482 patients with clinical and 80 patients with pathologic FTLD-TAR DNA-binding protein 43 without GRN mutations, 78 patients with FTLD with GRN mutations, and 822 controls. Association analysis of TMEM106B with GRN plasma levels was performed in 1,013 controls and TMEM106B and GRN mRNA expression levels were correlated in peripheral blood samples from 33 patients with FTLD and 150 controls. In our complete FTLD patient cohort, nominal significance was identified for 2 TMEM106B SNPs (top SNP rs1990622, p(allelic) = 0.036). However, the most significant association with risk of FTLD was observed in the subgroup of GRN mutation carriers compared to controls (corrected p(allelic) = 0.0009), where there was a highly significant decrease in the frequency of homozygote carriers of the minor alleles of all TMEM106B SNPs (top SNP rs1990622, CC genotype frequency 2.6% vs 19.1%, corrected p(recessive) = 0.009). We further identified a significant association of TMEM106B SNPs with plasma GRN levels in controls (top SNP rs1990622, corrected p = 0.002) and in peripheral blood samples a highly significant correlation was observed between TMEM106B and GRN mRNA expression in patients with FTLD (r = -0.63, p = 7.7 × 10(-5)) and controls (r = -0.49, p = 2.2 × 10(-10)). In our study, TMEM106B SNPs significantly reduced the disease penetrance in patients with GRN mutations, potentially by modulating GRN levels. These findings hold promise for the development of future protective therapies for FTLD.


Electrophoretic analysis of nucleic acids isolated from scrapie-infected hamster brain.
The purpose of this study was to investigate previous reports of a scrapie-specific 1.2 kb single-stranded DNA observed in alkaline agarose electrophoresis gels. Protocols were developed to be as consistent as possible with those used previously. Partial subcellular fractionation was applied to the brains of hamsters clinically affected by the 263K strain of scrapie. Nucleic acids were then isolated, and compared electrophoretically to nucleic acids isolated from equivalent fractions, made from the brains of hamsters inoculated with normal brain. Several modifications to the protocols were suggested. Results obtained by using these modifications were compared to results obtained using the original protocols. Scrapie-specific DNA was not observed in a total of eleven consecutive experiments. Thus, these results do not support the hypothesis that the agents of transmissible spongiform encephalopathy contain a single-stranded 1.2 kb DNA species as previously described.


A carnivorous plant fed by its ant symbiont: a unique multi-faceted nutritional mutualism.
Scarcity of essential nutrients has led plants to evolve alternative nutritional strategies, such as myrmecotrophy (ant-waste-derived nutrition) and carnivory (invertebrate predation). The carnivorous plant Nepenthes bicalcarata grows in the Bornean peatswamp forests and is believed to have a mutualistic relationship with its symbiotic ant Camponotus schmitzi. However, the benefits provided by the ant have not been quantified. We tested the hypothesis of a nutritional mutualism, using foliar isotopic and reflectance analyses and by comparing fitness-related traits between ant-inhabited and uninhabited plants. Plants inhabited by C. schmitzi produced more leaves of greater area and nitrogen content than unoccupied plants. The ants were estimated to provide a 200% increase in foliar nitrogen to adult plants. Inhabited plants also produced more and larger pitchers containing higher prey biomass. C. schmitzi-occupied pitchers differed qualitatively in containing C. schmitzi wastes and captured large ants and flying insects. Pitcher abortion rates were lower in inhabited plants partly because of herbivore deterrence as herbivory-aborted buds decreased with ant occupation rate. Lower abortion was also attributed to ant nutritional service. The ants had higher δ(15)N values than any tested prey, and foliar δ(15)N increased with ant occupation rate, confirming their predatory behaviour and demonstrating their direct contribution to the plant-recycled N. We estimated that N. bicalcarata derives on average 42% of its foliar N from C. schmitzi wastes, (76% in highly-occupied plants). According to the Structure Independent Pigment Index, plants without C. schmitzi were nutrient stressed compared to both occupied plants, and pitcher-lacking plants. This attests to the physiological cost of pitcher production and poor nutrient assimilation in the absence of the symbiont. Hence C. schmitzi contributes crucially to the nutrition of N. bicalcarata, via protection of assimilatory organs, enhancement of prey capture, and myrmecotrophy. This combination of carnivory and myrmecotrophy represents an outstanding strategy of nutrient sequestration.


Fast adaptation in vestibular hair cells requires myosin-1c activity.
In sensory hair cells of the inner ear, mechanical amplification of small stimuli requires fast adaptation, the rapid closing of mechanically activated transduction channels. In frog and mouse vestibular hair cells, we found that the rate of fast adaptation depends on both channel opening and stimulus size and that it is modeled well as a release of a mechanical element in series with the transduction apparatus. To determine whether myosin-1c molecules of the adaptation motor are responsible for the release, we introduced the Y61G mutation into the Myo1c locus and generated mice homozygous for this sensitized allele. Measuring transduction and adaptation in the presence of NMB-ADP, an allele-specific inhibitor, we found that the inhibitor not only blocked slow adaptation, as demonstrated previously in transgenic mice, but also inhibited fast adaptation. These results suggest that mechanical activity of myosin-1c is required for fast adaptation in vestibular hair cells.


Synthesis of secondary metabolites by Cladosporium resinae (NRL-6437) under different growth media and chemical inducers and their pharmaceutical activity.
The role of different growth media and chemical enhancer on synthesis of secondary metabolites Cladosporium resinae (NRL-6437) was investigated for their in vitro biological activities. Cladosporium resinae (NRL-6437) were grown in various nutrient media (Czapeak-dox Broth (CB), Czapeak Yeast-extract Broth (CYB), Yeast Extract Sucrose (YES), Potato Dextrose Broth (PDB) and Czapeak-dox (supplemented with glucose and starch) Broth (CGSB) for the production of metabolites. Two chemical epigenetic modifiers (suberoyl-anilide hydroxamic acid (SAHA) and 5-azacytidine (5-AZA) were also used for the expression of silent genes for secondary metabolite production. Our results indicated that among different media, Czapeak yeast extract broth produced more secondary metabolites. Application of 15mM of both modifiers was effective for the expressions of silent genes resulting in an increased metabolites production. Secondary metabolites extracted in ethyl acetate and fractionized in n-Hexane were also tested for their biological activity. The secondary metabolites revealed varying degrees of growth inhibitions of the tested organisms. Similarly, these metabolites were also active against brine shrimps and Lemna.


'Sweet talk': framing the merits of a sugar tax in Australia.
Although rising rates of obesity are recognized as a major concern for Australian public health, debate on what (if any) responsive action should be undertaken is conceptually and normatively complex. It is shaped by diverse values and interests; different representations of the problem; and many options for action by government, the private sector or individuals. This paper presents research documenting arguments for and against implementing a sugar tax in Australia. It is based on semi-structured interviews with representatives from industry (n = 4), public health (n = 4), policy think tanks (n = 2); and document and media analyses. The research design was informed by framing and agenda setting theory with results reported under four main themes: framing economic impacts, framing equity, framing obesity and framing the 'nanny state' versus individual liberty argument. We found that proposals for a sugar tax as part of policy responses to the issue of overweight and obesity in Australia are framed very differently by actors who either support or oppose it. A conclusion is that policy makers and public health advocates involved in policy debates on a sugar tax need to understand the role of problem and 'solution' framing, and develop positions based on protecting the public interest as a basic ethical responsibility of governments and public agencies.


Prescribing selective serotonin reuptake inhibitors in older age.
Apart from commercial reasons, two motivations have led to the introduction of SSRIs to replace the first and second generation antidepressants already available. One was the search for a more rational treatment, based on specific mechanisms, the other the development of effective treatments with fewer side effects, particularly for older patients, who have a greater sensitivity to cardio-vascular and central nervous system effects. The first has been frustrated up to a point, in that SSRIs and other single mechanism drugs do not appear to be more effective than the earliest relatively non-specific antidepressants. The second has been fulfilled, in that SSRIs generally are better tolerated in older patients and in overdose. However, there is a spectrum of other side effects that are particularly relevant in older age and that need attention when treating depression in this particular patient group.


Partial inhibition of the ubiquitin-proteasome system ameliorates cardiac dysfunction following ischemia-reperfusion in the presence of high glucose.
Acute hyperglycemia co-presenting with myocardial infarction (in diabetic and non-diabetic individuals) is often associated with a poor prognosis. Although acute hyperglycemia induces oxidative stress that can lead to dysregulation of the ubiquitin-proteasome system (UPS), it is unclear whether increased/decreased UPS is detrimental with ischemia-reperfusion under such conditions. As our earlier data implicated the UPS in cardiac damage, we hypothesized that its inhibition results in cardioprotection with ischemia-reperfusion performed under conditions that simulate acute hyperglycemia. Ex vivo rat heart perfusions were performed with Krebs-Henseleit buffer containing 33 mM glucose vs. controls (11 mM glucose) for 60 min stabilization, followed by 20 min global ischemia and 60 min reperfusion ± 5 µM lactacystin and 10 µM MG-132, respectively. The UPS inhibitors were added during the first 20 min of the reperfusion phase and various cardiac functional parameters evaluated. In parallel experiments, infarct sizes were assessed following 20 min regional ischemia and 120 min reperfusion ± each of the respective UPS inhibitors (added during reperfusion). Heart tissues were collected and analyzed for markers of oxidative stress, UPS activation, inflammation and autophagy. The proteasome inhibitor doses and treatment duration here employed resulted in partial UPS inhibition during the reperfusion phase. Both lactacystin and MG-132 administration resulted in cardioprotection in our experimental system, with MG-132 showing a greater effect. The proteasome inhibitors also enhanced cardiac superoxide dismutase protein levels (SOD1, SOD2), attenuated pro-inflammatory effects and caused an upregulation of autophagic markers. This study established that partial proteasome inhibition elicits cardioprotection in hearts exposed to ischemia-reperfusion with acute simulated hyperglycemia. These data reveal that protease inhibition triggered three major protective effects, i.e. (a) enhancing myocardial anti-oxidant defenses, (b) attenuating inflammation, and (c) increasing the autophagic response. Thus the UPS emerges as a unique therapeutic target for the treatment of ischemic heart disease under such conditions.


Ethanol withdrawal and hyperalgesia.
Hyperalgesia has been observed during ethanol withdrawal, comparable to the hyperalgesia observed during withdrawal from opioids. To determine the extent of this phenomenon and its potential mechanisms, both behavioral and in vitro studies are examined, and the roles of GABA(A), glutamate and other receptors in mediating the acute and chronic antinociceptive effects of ethanol are reviewed. Hyperalgesia during ethanol withdrawal is a robust phenomenon that has been observed in various strains of mice and rats, with different methods of exposure to ethanol, and with a variety of nociceptive assays. GABA receptors play an important role in mediating the antinociceptive effects of ethanol, but too little research has examined the role of glutamate receptors to make any conclusion about their importance. Adenosine receptors, calcium channels, and protein kinase C appear to play central roles in mediating tolerance to antinociceptive effects of ethanol and mediating the hyperalgesia seen during withdrawal. Although some key pathways have been identified, further mechanistic work is necessary to fully characterize the mechanisms for the development of hyperalgesia following chronic exposure to ethanol. An understanding of how the hyperalgesia may fit in with other manifestations of ethanol withdrawal may be an important variable in determining treatment outcome. Clinical research is essential to determine the significance of the hyperalgesia to the severity of withdrawal and to relapse.


Residential greenness, air pollution and psychological well-being among urban residents in Guangzhou, China.
China's rapid urbanization has led to an increasing level of exposure to air pollution and a decreasing level of exposure to vegetation among urban populations. Both trends may pose threats to psychological well-being. Previous studies on the interrelationships among greenness, air pollution and psychological well-being rely on exposure measures from remote sensing data, which may fail to accurately capture how people perceive vegetation on the ground. To address this research gap, this study aimed to explore relationships among neighbourhood greenness, air pollution exposure and psychological well-being, using survey data on 1029 adults residing in 35 neighbourhoods in Guangzhou, China. We used the Normalized Difference Vegetation Index (NDVI) and streetscape greenery (SVG) to assess greenery exposure at the neighbourhood level, and we distinguished between trees (SVG-tree) and grasses (SVG-grass) when generating streetscape greenery exposure metrics. We used two objective (PM2.5 and NO2 concentrations) measures and one subjective (perceived air pollution) measure to quantify air pollution exposure. We quantified psychological well-being using the World Health Organization Well-Being Index (WHO-5). Results from multilevel structural equation models (SEM) showed that, for parallel mediation models, while the association between SVG-grass and psychological well-being was completely mediated by perceived air pollution and NO2, the relationship between SVG-tree and psychological well-being was completely mediated by ambient PM2.5, NO2 and perceived air pollution. None of three air pollution indicators mediated the association between psychological well-being and NDVI. For serial mediation models, measures of air pollution did not mediate the relationship between NDVI and psychological well-being. While the linkage between SVG-grass and psychological well-being scores was partially mediated by NO2-perceived air pollution, SVG-tree was partially mediated by both ambient PM2.5-perceived air pollution and NO2-perceived air pollution. Our results suggest that street trees may be more related to lower air pollution levels and better mental health than grasses are.


EEL ELECTROPLAQUES: SPIKE ELECTROGENESIS WITHOUT POTASSIUM ACTIVATION.
Measurements with the voltage clamp technique demonstrate that only an early conductance increase occurs during spike electrogenesis of eel electroplaques. The delayed increase which is characteristic of spike electrogenesis in many other cells is absent. Instead, the membrane resistance increases two- to threefold above its resting value. The brief initial increase in conductance is due to sodium activation followed by rapid sodium inactivation. The influx of sodium causes an inward current of up to 80 ma/cm2. The inward current is abolished by eliminating the sodium from the normal medium (substitution of choline chloride for NaCl); by blocking sodium activation with tetrodotoxin; or by causing sodium inactivation through enrichment of the potassium in the medium. The delayed increase in membrane resistance is not affected by eliminating sodium influx, nor by substituting various impermeable anions for the chlorine of the normal medium. Thus, the increase in resistance signifies the occurrence of potassium inactivation which is unmasked by the absence of potassium activation.


Psychometric Characteristics of the Mutuality Scale in Stroke Patients and Caregivers.
The Mutuality Scale (MS) is composed of four theoretically derived factors (love, shared pleasurable activities, shared values, and reciprocity), but this structure has never been confirmed. Also, research involving the patient's perspective on the MS is limited. In this study, we tested the factorial structure of the MS and its reliability in stroke patients and caregivers. Cross-sectional, with a follow-up after 15 days for test-retest reliability. A total of 248 stroke patients and 163 stroke caregivers completed the MS. Stroke patients and their caregivers were enrolled in 10 rehabilitation hospitals across Italy. MS factorial structure was analyzed using confirmatory factor analysis; internal consistency reliability was evaluated with Cronbach's α and model-based internal consistency index; test-retest reliability was evaluated with intraclass correlation coefficient. Confirmatory factor analysis supported the four-factor structure of MS in its patient and caregiver version (CFI = 0.94; RMSEA = 0.06, for both). Cronbach's αs and model-based internal consistency index were >0.90 and intraclass correlations ranged between 0.66 and 0.93 in MS patient and caregiver version. This study tested the theoretical dimensions of the MS in stroke patients and their caregivers. From a scientific and clinical point of view, an assessment of stroke patient and caregiver mutuality would allow dyadic approaches to data analysis and care that account for the nonindependence between the stroke patient and the caregiver.


Tumour compartment transcriptomics demonstrates the activation of inflammatory and odontogenic programmes in human adamantinomatous craniopharyngioma and identifies the MAPK/ERK pathway as a novel therapeutic target.
Adamantinomatous craniopharyngiomas (ACPs) are clinically challenging tumours, the majority of which have activating mutations in CTNNB1. They are histologically complex, showing cystic and solid components, the latter comprised of different morphological cell types (e.g. β-catenin-accumulating cluster cells and palisading epithelium), surrounded by a florid glial reaction with immune cells. Here, we have carried out RNA sequencing on 18 ACP samples and integrated these data with an existing ACP transcriptomic dataset. No studies so far have examined the patterns of gene expression within the different cellular compartments of the tumour. To achieve this goal, we have combined laser capture microdissection with computational analyses to reveal groups of genes that are associated with either epithelial tumour cells (clusters and palisading epithelium), glial tissue or immune infiltrate. We use these human ACP molecular signatures and RNA-Seq data from two ACP mouse models to reveal that cell clusters are molecularly analogous to the enamel knot, a critical signalling centre controlling normal tooth morphogenesis. Supporting this finding, we show that human cluster cells express high levels of several members of the FGF, TGFB and BMP families of secreted factors, which signal to neighbouring cells as evidenced by immunostaining against the phosphorylated proteins pERK1/2, pSMAD3 and pSMAD1/5/9 in both human and mouse ACP. We reveal that inhibiting the MAPK/ERK pathway with trametinib, a clinically approved MEK inhibitor, results in reduced proliferation and increased apoptosis in explant cultures of human and mouse ACP. Finally, we analyse a prominent molecular signature in the glial reactive tissue to characterise the inflammatory microenvironment and uncover the activation of inflammasomes in human ACP. We validate these results by immunostaining against immune cell markers, cytokine ELISA and proteome analysis in both solid tumour and cystic fluid from ACP patients. Our data support a new molecular paradigm for understanding ACP tumorigenesis as an aberrant mimic of natural tooth development and opens new therapeutic opportunities by revealing the activation of the MAPK/ERK and inflammasome pathways in human ACP.


The complete mitochondrial genome of Wonwhang (Pyrus pyrifolia).
This is a de novo assembly and annotation of a complete mitochondrial genome from Pyrus pyrifolia in the family Rosaceae. The complete mitochondrial genome of P. pyrifolia was assembled from PacBio RSII P6-C4 sequencing reads. The circular genome was 458,873 bp in length, containing 39 protein-coding genes, 23 tRNA genes and three rRNA genes. The nucleotide composition was A (27.5%), T (27.3%), G (22.6%) and C (22.6%) with GC content of 45.2%. Most of protein-coding genes use the canonical start codon ATG, whereas nad1, cox1, matR and rps4 use ACG, mttB uses ATT, rpl16 and rps19 uses GTG. The stop codon is also common in all mitochondrial genes. The phylogenetic analysis showed that P. pyrifolia was clustered with the Malus of Rosaceae family. Maximum-likelihood analysis suggests a clear relationship of Rosids and Asterids, which support the traditional classification.


Associations Between Parental Perceptions of the Neighborhood Environment and Childhood Physical Activity: Results from ISCOLE-Kenya.
A physical activity transition to declining activity levels, even among children, now poses a serious public health concern because of its contribution to a rising prevalence of noncommunicable diseases. Childhood physical activity levels are associated with parental perceptions of the neighborhood; however, these relationships have not been explored in sub-Saharan Africa (SSA). The objective was to investigate relationships between parental perceptions of the neighborhood and physical activity indicators among Kenyan children. Data were collected from children 9 to 11 years old in Nairobi as part of the International Study of Childhood Obesity, Lifestyle and Environment. Child physical activity was assessed by accelerometry, and information on obtaining sufficient physical activity, active transport, and parental perceptions of the neighborhood collected using questionnaires. Of 563 participating children, 45.7%, 12.6%, and 11.4% used active school transportation, met physical activity guidelines, and were sufficiently active, respectively. Parental perception of positive neighborhood social cohesion, positive environs and connectivity, and negative child safety concerns, were associated with child physical activity outcomes. Aspects of parental perceptions of the neighborhood were associated with child physical activity outcomes and should be further explored to appropriately inform policy and practice in curbing declining physical activity levels among children in SSA.


Recent contributions of glaciers and ice caps to sea level rise.
Glaciers and ice caps (GICs) are important contributors to present-day global mean sea level rise. Most previous global mass balance estimates for GICs rely on extrapolation of sparse mass balance measurements representing only a small fraction of the GIC area, leaving their overall contribution to sea level rise unclear. Here we show that GICs, excluding the Greenland and Antarctic peripheral GICs, lost mass at a rate of 148 ± 30 Gt yr(-1) from January 2003 to December 2010, contributing 0.41 ± 0.08 mm yr(-1) to sea level rise. Our results are based on a global, simultaneous inversion of monthly GRACE-derived satellite gravity fields, from which we calculate the mass change over all ice-covered regions greater in area than 100 km(2). The GIC rate for 2003-2010 is about 30 per cent smaller than the previous mass balance estimate that most closely matches our study period. The high mountains of Asia, in particular, show a mass loss of only 4 ± 20 Gt yr(-1) for 2003-2010, compared with 47-55 Gt yr(-1) in previously published estimates. For completeness, we also estimate that the Greenland and Antarctic ice sheets, including their peripheral GICs, contributed 1.06 ± 0.19 mm yr(-1) to sea level rise over the same time period. The total contribution to sea level rise from all ice-covered regions is thus 1.48 ± 0.26 mm (-1), which agrees well with independent estimates of sea level rise originating from land ice loss and other terrestrial sources.


Dinitroazetidines are a novel class of anticancer agents and hypoxia-activated radiation sensitizers developed from highly energetic materials.
In an effort to develop cancer therapies that maximize cytotoxicity, while minimizing unwanted side effects, we studied a series of novel compounds based on the highly energetic heterocyclic scaffold, dinitroazetidine. In this study, we report the preclinical validation of 1-bromoacetyl-3,3-dinitroazetidine (ABDNAZ), a representative lead compound currently in a phase I clinical trial in patients with cancer. In tumor cell culture, ABDNAZ generated reactive free radicals in a concentration- and time-dependent manner, modulating intracellular redox status and triggering apoptosis. When administered to mice as a single agent, ABDNAZ exhibited greater cytotoxicity than cisplatin or tirapazamine under hypoxic conditions. However, compared with cisplatin, ABDNAZ was better tolerated at submaximal doses, yielding significant tumor growth inhibition in the absence of systemic toxicity. Similarly, when combined with radiation, ABDNAZ accentuated antitumor efficacy along with the therapeutic index. Toxicity studies indicated that ABDNAZ was not myelosuppressive and no dose-limiting toxicity was apparent following daily administration for 14 days. Taken together, our findings offer preclinical proof-of-concept for ABDNAZ as a promising new anticancer agent with a favorable toxicity profile, either as a chemotherapeutic agent or a radiosensitizer.


Weak value amplified optical activity measurements.
We present a new form of optical activity measurement based on a modified weak value amplification scheme. It has recently been shown experimentally that the left- and right-circular polarization components refract with slightly different angles of refraction at a chiral interface causing a linearly polarized light beam to split into two. By introducing a polarization modulation that does not give rise to a change in the optical rotation it is possible to differentiate between the two circular polarization components even after post-selection with a linear polarizer. We show that such a modified weak value amplification measurement permits the sign of the splitting and thus the handedness of the optically active medium to be determined. Angular beam separations of Δθ ∼ 1 nanoradian, which corresponds to a circular birefringence of Δn ∼ 1 × 10(-9), could be measured with a relative error of less than 1%.


Traffic-related air pollution correlates with adult-onset asthma among never-smokers.
Traffic-related pollution is associated with the onset of asthma in children. Its effect on adult-onset asthma is poorly investigated. The SAPALDIA cohort study was used to investigate associations between the 11-year change (1991-2002) in home outdoor traffic-related particulate matter up to 10 microm in diameter (TPM(10)) and the incidence of asthma. Never-smokers without asthma at baseline aged 18-60 years in 1991 were eligible for inclusion in the study. Subjects reporting doctor-diagnosed asthma at follow-up were considered incident cases. TPM(10) at baseline and follow-up was predicted and interpolated to subjects' place of residence by dispersion models using emission and meteorological data. Cox proportional hazard models for time to asthma onset were adjusted (age, gender, baseline atopy, body mass index, bronchial reactivity, maternal allergies). Of 2725 never-smokers, 41 reported asthma onset in 2002. Home outdoor TPM(10) concentrations improved during the interval (mean -0.6; range -9 to +7.2; IQR 0.6 microg/m(3)). The incidence of asthma was associated with a change in TPM(10). The hazard ratio (1.30; 95% CI 1.05 to 1.61) per 1 microg/m(3) change in TPM(10) (IQR) was not sensitive to further adjustments (education, workplace exposure, passive smoking, parental asthma or allergies, random area effects, lung function or co-pollutants such as regional, secondary, total PM(10) or proximity to busy roads). The data suggest a role for traffic-related pollution in adult-onset asthma. Space, time and source-specific individual assignment of exposure to traffic-related pollution is a key strength of SAPALDIA. It may explain why findings were statistically significant despite the limited number of new cases. As traffic-related pollution prevails, the finding may be of substantial public health relevance.


Similarity in percent body fat between white and Vietnamese women: implication for a universal definition of obesity.
It has been widely assumed that for a given BMI, Asians have higher percent body fat (PBF) than whites, and that the BMI threshold for defining obesity in Asians should be lower than the threshold for whites. This study sought to test this assumption by comparing the PBF between US white and Vietnamese women. The study was designed as a comparative cross-sectional investigation. In the first study, 210 Vietnamese women ages between 50 and 85 were randomly selected from various districts in Ho Chi Minh City (Vietnam). In the second study, 419 women of the same age range were randomly selected from the Rancho Bernardo Study (San Diego, CA). In both studies, lean mass (LM) and fat mass (FM) were measured by dual-energy X-ray absorptiometry (DXA) (QDR 4500; Hologic). PBF was derived as FM over body weight. Compared with Vietnamese women, white women had much more FM (24.8 +/- 8.1 kg vs. 18.8 +/- 4.9 kg; P < 0.0001) and greater PBF (36.4 +/- 6.5% vs. 35.0 +/- 6.2%; P = 0.012). However, there was no significant difference in PBF between the two groups after matching for BMI (35.1 +/- 6.2% vs. 35.0 +/- 5.7%; P = 0.87) or for age and BMI (35.6 +/- 5.1% vs. 35.8 +/- 5.9%; P = 0.79). Using the criteria of BMI >or=30, 19% of US white women and 5% of Vietnamese women were classified as obese. Approximately 54% of US white women and 53% of Vietnamese women had their PBF >35% (P = 0.80). Although white women had greater BMI, body weight, and FM than Vietnamese women, their PBF was virtually identical. Further research is required to derive a more appropriate BMI threshold for defining obesity for Asian women.


Function of the homeodomain protein GHF1 in pituitary cell proliferation.
Mutations that cause pituitary dwarfism in the mouse reside in the gene encoding the transcription factor growth hormone factor 1 (GHF1 or pit1). These dwarf mice (dw and dwJ) are deficient in growth hormone (GH) and prolactin (PRL) synthesis and exhibit pituitary hypoplasia, suggesting a stem cell defect. With antisense oligonucleotide technology, a cell culture model of this genetic defect was developed. Specific inhibition of GHF1 synthesis by complementary oligonucleotides led to a marked decrease in GH and PRL expression and to a marked decrease in proliferation of somatotrophic cell lines. These results provide direct evidence that the homeodomain protein GHF1 is required not only for the establishment and maintenance of the differentiated phenotype but for cell proliferation as well.


Mass Spectrometric Analysis of TRPM6 and TRPM7 Phosphorylation Reveals Regulatory Mechanisms of the Channel-Kinases.
TRPM7 and TRPM6 were the first identified bifunctional channels to contain their own kinase domains, but how these channel-kinases are regulated is poorly understood. Previous studies identified numerous phosphorylation sites on TRPM7, but very little is known about TRPM6 phosphorylation or sites on TRPM7 transphosphorylated by TRPM6. Our mass spectrometric analysis of homomeric and heteromeric TRPM7 and TRPM6 channels identified phosphorylation sites on both proteins, as well as several prominent sites on TRPM7 that are commonly modified through autophosphorylation and transphosphorylation by TRPM6. We conducted a series of amino acid substitution analyses and identified S1777, in TRPM7's catalytic domain, and S1565, in TRPM7's exchange domain that mediates kinase dimerization, as potential regulatory sites. The phosphomimetic S1777D substitution disrupted catalytic activity, most likely by causing an electrostatic perturbation at the active site. The S1565D phosphomimetic substitution also inactivated the kinase but did so without interfering with kinase dimerization. Molecular modeling indicates that phosphorylation of S1565 is predicted to structurally affect TRPM7's functionally conserved N/D loop, which is thought to influence the access of substrate to the active site pocket. We propose that phosphorylation of S1565 within the exchange domain functions as a regulatory switch to control TRPM7 catalytic activity.


Informal and formal caregivers' involvement in nursing home care activities: impact of integrated care.
This paper reports a study to investigate the relationships between informal and formal care, changing relationships over time, impact of integrated care, and theoretical and methodological lessons that can be drawn from research on this topic. The ratio of informal to formal care provided in nursing homes varies. In many countries the predominance of traditional, formal care is being challenged by the emergence of integrated care models in which formal and informal caregivers co-operate and co-ordinate their activities to deliver tailor-made care. This move towards integrated care is likely to result in changes in the degree and nature of involvement of formal and informal caregivers in care activities. However, very little research has investigated this. Previous research is restricted to one point in time, as opposed to focussing on the changing relationships between formal and informal caregivers over time. The quasi-experimental design encompassed a traditional care comparison setting and an experimental setting with an integrated care intervention. At three measurement points, one before the intervention in May/June 2000 and two at 6 and 14 months after the intervention, informal and formal caregivers recorded their care activities. Informal care relationships were determined by calculating contribution rates and informal/formal care ratios for 14 activities. Integrated care did not bring about the expected major changes. There was a limited amount of substitution of formal care with informal care. There were limited changes in the extent of dual specialization, in which informal and formal caregivers perform separate activities. There was little supplementation of formal care with informal care. Furthermore, relationships changed over time, resulting in a complex pattern of linkages at the level of separate activities. Informal caregivers contribute to many activities. The impact of integrated care on the degree and nature of involvement, however, was moderate. A possible explanation is the contextual situation for nursing home care. These findings point to the need for further research into the (changing) relationship between formal and informal carers in residential care activities and the evolution of this relationship over time.


Plasma lipid profiles discriminate bacterial from viral infection in febrile children.
Fever is the most common reason that children present to Emergency Departments. Clinical signs and symptoms suggestive of bacterial infection are often non-specific, and there is no definitive test for the accurate diagnosis of infection. The 'omics' approaches to identifying biomarkers from the host-response to bacterial infection are promising. In this study, lipidomic analysis was carried out with plasma samples obtained from febrile children with confirmed bacterial infection (n = 20) and confirmed viral infection (n = 20). We show for the first time that bacterial and viral infection produces distinct profile in the host lipidome. Some species of glycerophosphoinositol, sphingomyelin, lysophosphatidylcholine and cholesterol sulfate were higher in the confirmed virus infected group, while some species of fatty acids, glycerophosphocholine, glycerophosphoserine, lactosylceramide and bilirubin were lower in the confirmed virus infected group when compared with confirmed bacterial infected group. A combination of three lipids achieved an area under the receiver operating characteristic (ROC) curve of 0.911 (95% CI 0.81 to 0.98). This pilot study demonstrates the potential of metabolic biomarkers to assist clinicians in distinguishing bacterial from viral infection in febrile children, to facilitate effective clinical management and to the limit inappropriate use of antibiotics.


Exercise in the Prevention and Treatment of Breast Cancer: What Clinicians Need to Tell Their Patients.
In 2016, nearly a quarter-million women were diagnosed with invasive breast cancer, the second deadliest cancer type. Lifestyle behaviors are well-established risk factors for both the development of, and negative outcomes from, breast cancer. New findings demonstrate that engaging in even minimal amounts of exercise is protective against breast cancer. Further, numerous studies have identified the role of exercise in the treatment of breast cancer and improvement in treatment side effects, quality of life, and overall survival. Unfortunately, few patients are physically active due to multiple barriers. Physicians may be helpful in engaging patients in being more active. Further research is necessary to identify programs and approaches to help patients both with, and at-risk for, breast cancer to engage in exercise.


Intrinsic ferromagnetism and quantum transport transition in individual Fe-doped Bi2Se3 topological insulator nanowires.
Time-reversal symmetry is broken by magnetic doping in topological insulators (TIs). An energy gap at the Dirac point opens and thus, generates numerous surface carriers. TI nanostructures are an ideal platform to investigate exotic surface transport behavior due to their large surface-to-volume ratio, which enhances the contribution of the TI surface states. However, magnetic doping into TI nanostructures has been challenging, and induced magnetic behavior has remained elusive. Herein, we have synthesized Fe-doped Bi2Se3 nanowires using a facile chemical vapor deposition with a doping concentration of ∼1 at%. The combined structural characterizations illustrate the homogeneous distribution of the Fe dopants. Cryogenic magnetic force microscopy gives direct evidence of the spontaneous magnetization with a Curie temperature of ∼40 K in a single nanowire. The transport measurements show a quantum transition from weak anti-localization to weak localization behavior. All the evidence indicates the existence of intrinsic ferromagnetism and gapped topological surface states in the TI nanowires, paving a way for future memory and magnetoelectric nanodevice applications.


Maximally recommended doses of angiotensin-converting enzyme (ACE) inhibitors do not completely prevent ACE-mediated formation of angiotensin II in chronic heart failure.
The added benefits of angiotensin II type I receptor (AT(1)) blockers (ARBs) to ACE inhibition suggests that recommended doses of ACE inhibitors provide only partial inhibition of ACE in chronic heart failure (CHF). Accordingly, the level of ACE inhibition was assessed by the pressor response to angiotensin (Ang) I in patients who had been treated with recommended doses of ACE inhibitors. Forty-two patients with CHF receiving 40 mg/d of a long-acting ACE inhibitor or 150 mg of captopril were studied. Radial artery systolic pressure (RASP, mm Hg) was monitored noninvasively. The pressor response to ascending doses of Ang I was evaluated in all patients before and after administration of the ARB valsartan. The pressor response to Ang I before and after valsartan was also reevaluated in 11 patients after the dose of ACE inhibitor was doubled for 1 week. RASP increased linearly with significantly ascending doses of Ang I despite treatment with ACE inhibitors. The pressor response to Ang I was blunted significantly by valsartan. Ang I-induced increase in RASP did not correlate with duration of ACE inhibitor therapy. After the dose of ACE inhibitors was doubled, the pressor response to Ang I was no longer different from that noted after valsartan. Recommended doses of ACE inhibitors do not fully inhibit ACE in CHF. The level of ACE inhibition achieved is not related to duration of ACE inhibitor therapy. Greater ACE inhibition is also achieved at twice the recommended doses of ACE inhibitors.


Intermethod variation in serum carcinoembryonic antigen (CEA) measurement. fresh serum pools and control materials compared.
This study was undertaken to evaluate the feasibility of using commercial control materials in a regional external quality assessment scheme (EQAS) for serum carcinoembryonic antigen (CEA) measurement. We have assessed the commutability of 12 commercial control materials using five automated immunochemical systems. We compared the intermethod behavior of the materials with that of 12-14 patient serum pools. In a total of 48 comparisons (12 materials x 4 pairs of analytical systems), seven instances of non-commutability were apparent, as shown by normalized residuals falling outside the +/-3 interval. The use of non-commutable materials generates two negative effects. In EQAS, the interlaboratory variation recorded is not representative of the variation expected in the assay of patient sera; in interlaboratory harmonization programs, recalibration with non-commutable materials increases, instead of decreasing, the interlaboratory variation. Both these effects were shown to occur in CEA measurement with the tested materials and systems. The materials planned to be used in our EQAS turned out to be commutable: this gave us the guarantee of measuring realistic interlaboratory variation values, although the check for commutability should be extended to all the analytical systems used by the participants in the scheme.


Caerulein-induced acute pancreatitis in mice that constitutively overexpress Reg/PAP genes.
The cystic fibrosis (CF) mouse pancreas has constitutively elevated expression of the Reg/PAP cell stress genes (60-fold greater Reg3alpha, and 10-fold greater PAP/Reg3beta and Reg3gamma). These genes are suggested to be involved in protection or recovery from pancreatic injury. To test this idea the supramaximal caerulein model was used to induce acute pancreatitis in wild type and CF mice. Serum amylase, pancreatic water content (as a measure of edema), pancreatic myeloperoxidase activity, and Reg/PAP expression were quantified. In both wild type and CF mice caerulein induced similar elevations in serum amylase (maximal at 12 h), pancreatic edema (maximal at 7 h), and pancreatic myeloperoxidase activity (MPO, a marker of neutrophil infiltration; maximal at 7 h). By immunohistochemistry, Reg3alpha was strongly expressed in the untreated CF pancreas but not in wild type. During pancreatitis, Reg3alpha was intensely expressed in foci of inflamed tissue in both wild type and CF. These data demonstrate that the severity of caerulein-induced pancreatitis is not ameliorated in the CF mouse even though the Reg/PAP stress genes are already highly upregulated. While Reg/PAP may be protective they may also have a negative effect during pancreatitis due to their anti-apoptotic activity, which has been shown to increase the severity of pancreatitis.


Arguments against toxic effects of chemotherapy on liver injury and regeneration in an experimental model of partial hepatectomy.
New chemotherapy regimens are increasingly used in metastatic colorectal cancer to the liver before surgery. Some clinical observations have suggested that chemotherapy may affect liver regeneration. The aim of this study was to evaluate liver damage and liver regeneration after chemotherapy treatment in a model of partial hepatectomy. C57BL/6 mice were repeatedly treated with intraperitoneal injections of either saline or different chemotherapy regimens including the drugs 5-fluorouracyl (5-FU), irinotecan, oxaliplatin, gemcitabine and combined treatments with 5-FU/irinotecan, 5-FU/oxaliplatin. A 70% partial hepatectomy was performed 1 week after the last injection. Ki-67 and PCNA immunohistochemistry were performed to assess liver regeneration, serum liver enzymes and histology analysis to evaluate injury. A variety of chemotherapeutic agents used at maximum tolerated doses compatible with survival affected body weight and blood cell levels. However, these regimens did not affect liver injury before and after hepatectomy nor did they impair liver regeneration. Liver histology showed no steatosis, fibrosis or inflammation before hepatectomy. We therefore tested whether chemotherapy in presence of diet-induced steatosis may trigger injury. Even under these conditions, we did not observe histological signs of inflammation or sinusoidal injury. Liver injury and liver regeneration are not impaired after neoadjuvant chemotherapy with 5-FU, irinotecan, oxaliplatin and gemcitabine in non-tumoural liver parenchyma. In addition, combined treatments disclose no adverse effects on liver regeneration. Chemotherapy alone induces no histological alterations even in the presence of steatosis.


Expression of heparan sulphate N-deacetylase/N-sulphotransferase by vascular smooth muscle cells.
Heparan sulphate is an important mediator in determining vascular smooth muscle cell (SMC) phenotype. The sulphation pattern of the heparan sulphate chains is critical to their function. We have examined the initial step in the biosynthesis of the sulphated domains mediated by the enzyme heparan sulphate N-deacetylase/N-sulphotransferase (NDST). Rabbit aortic SMC in primary culture exhibited NDST enzyme activity and expressed NDST-1 in their Golgi apparatus, with maximal expression in SMC 2 days after dispersal in primary culture confirmed by Western blot analysis. Endothelial cells, macrophages and fibroblasts expressed NDST-1 but had generally less intense staining than SMC, although SMC expression decreased with culture. The uninjured rat aorta also showed widespread expression of NDST-1. After balloon de-endothelialisation, NDST-1 could not be detected in SMC of the neointima in the early stages of neointimal formation, but was re-expressed at later time points (after 12 weeks). In human coronary arteries, SMC of the media and the diffuse intimal thickening expressed NDST-1, while SMC in the atherosclerotic plaque were negative for NDST-1. We conclude that SMC may regulate their heparan sulphate sulphation at the level of expression of the enzyme heparan sulphate NDST in a manner related to their phenotypic state.


Clinical algorithms for the identification of sick newborns in community-based settings.
Clinical algorithms can be powerful tools for the identification of sick newborns at risk of neonatal mortality. Several studies have evaluated clinical signs for newborns aged 0-60 days to identify severe illness; however, few studies have focused specifically on the most vulnerable time period for neonatal death, the first week of life. Therefore, we reviewed the studies that evaluated clinical signs in newborns 0-60 days, focusing on infants 0 to <7 days. Based on a comparison of relevant studies, we then identified the common, important clinical signs shown to be useful for the identification of at-risk newborns by health workers in community-based and low-resource settings. We concluded that further work is urgently needed to develop a clinical algorithm for widespread validation in various community-based settings, which focuses specifically on newborns <7 days at risk of early neonatal mortality.


Isolation and characterization of specialized phi80 transducing phages carrying regions of the Salmonella typhimurium trp operon.
We have isolated a series of nondefective phi80 specialized transducing phage which carry segments of the Salmonella typhimurium trp operon. These phage were obtained from a lysogenic derivative of a merozygote constructed by transferring an S. typhimurium trp episome into an Escherichia coli strain which lacks the normal phi80 attachment site. The deoxyribonucleic acid (DNA) from one such phage was purified and employed in DNA-ribonucleic acid (RNA) hybridization studies. The results obtained show that, under our hybridization conditions, heterologous hybridization is less efficient than homologous hybridization. It was also observed that not all S. typhimurium trp messenger RNA can readily anneal to E. coli trp operon DNA. Heterologous hybrids consisting of S. typhimurium trp messenger RNA and E. coli trp operon DNA were estimated to have a dissociation constant 10-fold larger than that of homologous hybrids.


[Metabonomic profiling of plasma metabolites in Wistar rats to study the effect of aging by means of GC/TOFMS-based techniques].
The global metabolite profiles of endogenous compounds of Wistar rats from 12 to 20 weeks old were investigated to take deep insight into and get better understanding of the pathogenesis of development and aging. Plasma from Wistar rats at 12, 14, 16, 18, and 20 weeks old were analyzed using GC/TOFMS. Multivariate data analysis was then used to process the metabonomic data which indicated excellent separation between different weeks and showed that the metabolic profiles of the samples changed with age, enabling age-related metabolic trajectories to be visualized. Decreased concentrations of citric acid, cis-aconitic acid, 9-(z)-hexadecenoic acid along with increased levels of hexanedioic acid, alpha-tocopherol, 3-indole propionic acid, etc contributed to the separation. Several major metabolic pathways were identified to be involved in metabolic regulation. This suggests that GC/TOFMS-based metabonomics is a powerful alternative approach to identifying potential biomarkers and investigating the physiological developments of aging and it is important to employ suitable age-match control group in metabonomic study of physiological monitoring, drug safety assessment, and disease diagnosis, etc.


Transient stimulation of distinct subpopulations of striatal neurons mimics changes in action value.
In changing environments, animals must adaptively select actions to achieve their goals. In tasks involving goal-directed action selection, striatal neural activity has been shown to represent the value of competing actions. Striatal representations of action value could potentially bias responses toward actions of higher value. However, no study to date has demonstrated the direct effect of distinct striatal pathways in goal-directed action selection. We found that transient optogenetic stimulation of dorsal striatal dopamine D1 and D2 receptor-expressing neurons during decision-making in mice introduced opposing biases in the distribution of choices. The effect of stimulation on choice was dependent on recent reward history and mimicked an additive change in the action value. Although stimulation before and during movement initiation produced a robust bias in choice behavior, this bias was substantially diminished when stimulation was delayed after response initiation. Together, our data suggest that striatal activity is involved in goal-directed action selection.


Formation of prostaglandins E2 and D2 via the isoprostane pathway: a mechanism for the generation of bioactive prostaglandins independent of cyclooxygenase.
It has heretofore been assumed that the cyclooxygenases (COXs) are solely responsible for peostaglandin (PG) synthesis in vivo. An important structural feature of PGH2 formed by COX is the trans-configuration of side chains relative to the prostane ring. Previously, we reported that a series of PG-like compounds termed isoprostanes (IsoPs) are formed in vivo in humans from the free radical-catalyzed peroxidation of arachidonate independent of COX. A major difference between these compounds and PGs is that IsoPs are formed from endoperoxide intermediates, the vast majority of which contain side chains that are cis relative to the prostane ring. In addition, unlike the formation of eicosanoids from COX, IsoPs are formed as racemic mixtures because they are generated nonenzymatically. IsoPs containing E- and D-type prostane rings (E2/D2-IsoPs) are one class of IsoPs formed, and we have reported previously that one of the major IsoPs generated is 15-E2t-IsoP (8-iso-PGE2). Unlike PGE2, 15-E2t-IsoP is significantly more unstable in buffered solutions in vitro and undergoes epimerization to PGE2. Analogously, the D-ring IsoP (15-D2c-IsoP) would be predicted to rearrange to PGD2. We now report that compounds identical in all respects to PGE2 and PGD2 and their respective enantiomers are generated in vivo via the IsoP pathway, presumably by epimerization of racemic 15-E2t-IsoP and 15-D2c-IsoP, respectively. Racemic PGE2 and PGD2 were present esterified in phospholipids derived from liver tissue from rats exposed to oxidant stress at levels of 24 +/- 16 and 37 +/- 12 ng/g of tissue, respectively. In addition, racemic PGs, particularly PGD2, were present unesterified in urine from normal animals and humans and represented up to 10% of the total PG detected. Levels of racemic PGD2 increased 35-fold after treatment of rats with carbon tetrachloride to induce oxidant stress. In this setting, PGD2 and its enantiomer generated by the IsoP pathway represented approximately 30% of the total PGD2 present in urine. These findings strongly support the contention that a second pathway exists for the formation of bioactive PGs in vivo that is independent of COX.


Road safety in drivers with Parkinson disease.
To assess road safety and its predictors in drivers with Parkinson disease (PD). Licensed, active drivers with PD (n = 84; age = 67.3 +/- 7.8, median Hoehn & Yahr stage II) and controls (n = 182; age = 67.6 +/- 7.5) underwent cognitive, visual, and motor tests, and drove a standardized route in urban and rural settings in an instrumented vehicle. Safety errors were judged and documented by a driving expert based on video data review. Drivers with PD committed more total safety errors compared to controls (41.6 +/- 14.6 vs 32.9 +/- 12.3, p < 0.0001); 77.4% of drivers with PD committed more errors than the median total error count of the controls (medians: PD = 39.5, controls = 31.0). Lane violations were the most common error category in both groups. Group differences in some error categories became insignificant after results were adjusted for demographics and familiarity with the local driving environment. The PD group performed worse on tests of motor, cognitive, and visual abilities. Within the PD group, older age and worse performances on tests of visual acuity, contrast sensitivity, attention, visuospatial abilities, visual memory, and general cognition predicted error counts. Measures of visual processing speed and attention and far visual acuity were jointly predictive of error counts in a multivariate model. Overall, drivers with Parkinson disease (PD) had poorer road safety compared to controls, but there was considerable variability among the drivers with PD, and some performed normally. Familiarity with the driving environment was a mitigating factor against unsafe driving in PD. Impairments in visual perception and cognition were associated with road safety errors in drivers with PD.


Sorbicillinoid analogs with cytotoxic and selective anti-Aspergillus activities from Scytalidium album.
As part of an ongoing project to explore filamentous fungi for anticancer and antibiotic leads, 11 compounds were isolated and identified from an organic extract of the fungus Scytalidium album (MSX51631) using bioactivity-directed fractionation against human cancer cell lines. Of these, eight compounds were a series of sorbicillinoid analogs (1-8), of which four were new (scalbucillin A (2), scalbucillin B (3), scalbucillin C (6) and scalbucillin D (8)), two were phthalides (9-10) and one was naphthalenone (11). Compounds (1-11) were tested in the MDA-MB-435 (melanoma) and SW-620 (colon) cancer cell lines. Compound 1 was the most potent with IC50 values of 1.5 and 0.5 μM, followed by compound 5 with IC50 values of 2.3 and 2.5 μM at 72 h. Compound 1 showed a 48-h IC50 value of 3.1 μM when tested against the lymphocytic leukemia cell line OSU-CLL, while the nearly identical compound 5 had almost no activity in this assay. Compounds 1 and 5 showed selective and equipotent activity against Aspergillus niger with minimum IC values of 0.05 and 0.04 μg ml(-1) (0.20 and 0.16 μM), respectively. The in vitro hemolytic activity against sheep erythrocytes of compounds 1 and 5 was investigated and were found to provoke 10% hemolysis at 52.5 and 45.0 μg ml(-1), respectively, indicative of a promising safety factor.


Multivitamins, folate, and green vegetables protect against gene promoter methylation in the aerodigestive tract of smokers.
One promising approach for early detection of lung cancer is by monitoring gene promoter hypermethylation events in sputum. Epidemiologic studies suggest that dietary fruits and vegetables and the micronutrients they contain may reduce risk of lung cancer. In this study, we evaluated whether diet and multivitamin use influenced the prevalence of gene promoter methylation in cells exfoliated from the aerodigestive tract of current and former smokers. Members (N = 1,101) of the Lovelace Smokers Cohort completed the Harvard Food Frequency Questionnaire and provided a sputum sample that was assessed for promoter methylation of eight genes commonly silenced in lung cancer and associated with risk for this disease. Methylation status was categorized as low (fewer than two genes methylated) or high (two or more genes methylated). Logistic regression models were used to identify associations between methylation status and 21 dietary variables hypothesized to affect the acquisition of gene methylation. Significant protection against methylation was observed for leafy green vegetables [odds ratio (OR) = 0.83 per 12 monthly servings; 95% confidence interval (95% CI), 0.74-0.93] and folate (OR, 0.84 per 750 microg/d; 95% CI, 0.72-0.99). Protection against gene methylation was also seen with current use of multivitamins (OR, 0.57; 95% CI, 0.40-0.83). This is the first cohort-based study to identify dietary factors associated with reduced promoter methylation in cells exfoliated from the airway epithelium of smokers. Novel interventions to prevent lung cancer should be developed based on the ability of diet and dietary supplements to affect reprogramming of the epigenome.


Interleukin-10 regulates hepcidin in Plasmodium falciparum malaria.
Acute malarial anemia remains a major public health problem. Hepcidin, the major hormone controlling the availability of iron, is raised during acute and asymptomatic parasitemia. Understanding the role and mechanism of raised hepcidin and so reduced iron availability during infection is critical to establish evidence-based guidelines for management of malaria anemia. Our recent clinical evidence suggests a potential role of IL-10 in the regulation of hepcidin in patients with acute P. falciparum malaria. We have measured secretion of hepcidin by primary macrophages and the hepatoma cell line HepG2 stimulated with IL-10, IL-6 and Plasmodium falciparum-infected erythrocytes. We have observed that IL-10 and IL-6 production increased in primary macrophages when these cells were co-cultured with Plasmodium falciparum-infected erythrocytes. We found that IL-10 induced hepcidin secretion in primary macrophages in a dose-dependent manner but not in HepG2 cells. These effects were mediated through signal transducer and activator of transcription (STAT) 3-phosphorylation and completely abrogated by a specific STAT3 inhibitor. IL-10 can directly regulate hepcidin in primary macrophages but not in HepG2 cells. This effect can be modulated by Plasmodium falciparum. The results are consistent with a role for IL-10 in modulating iron metabolism during acute phase of infection.


Bioinformatics approaches and software for detection of secondary metabolic gene clusters.
The accelerating pace of microbial genomics is sparking a renaissance in the field of natural products research. Researchers can now get a preview of the organism's secondary metabolome by analyzing its genomic sequence. Combined with other -omics data, this approach may provide a cost-effective alternative to industrial high-throughput screening in drug discovery. In the last few years, several computational tools have been developed to facilitate this process by identifying genes involved in secondary metabolite biosynthesis in bacterial and fungal genomes. Here, we review seven software programs that are available for this purpose, with an emphasis on antibiotics & Secondary Metabolite Analysis SHell (antiSMASH) and Secondary Metabolite Unknown Regions Finder (SMURF), the only tools that can comprehensively detect complete secondary metabolite biosynthesis gene clusters. We also discuss five related software packages-CLUster SEquence ANalyzer (CLUSEAN), ClustScan, Structure Based Sequence Analysis of Polyketide Synthases (SBSPKS), NRPSPredictor, and Natural Product searcher (NP.searcher)-that identify secondary metabolite backbone biosynthesis genes. This chapter offers detailed protocols, suggestions, and caveats to assist researchers in using these tools most effectively.


Menopause and the influence of culture: another gap for Indigenous Australian women?
There is great variation in experience of menopause in women around the world. The purpose of this study was to review current understanding of Australian Aboriginal and Torres Strait Islander (Indigenous) women's experiences of menopause. The literature pertaining to the perception, significance and experience of menopause from a number of cultural groups around the world has been included to provide context for why Indigenous women's experience might be important for their health and differ from that reported in other studies of Australian women and menopause. A search of databases including Ovid Medline, Pubmed, Web of Science, AUSThealth, AMED, EMBASE, Global Health and PsychINFO was undertaken from January 2011 to April 2011 using the search terms menopause, Indigenous, Aboriginal, attitudes, and perceptions and repeated in September 2012. Considerable research shows significant variation across cultures in the menopausal experience. Biological, psychological, social and cultural factors are associated with either positive or negative attitudes, perceptions or experiences of menopause in various cultures. Comparative international literature shows that neither biological nor social factors alone are sufficient to explain the variation in experiences of the menopausal transition. However, a strong influence of culture on the menopause experience can be found. The variation in women's experience of menopause indicates that different cultural groups of women may have different understandings and needs during the menopausal transition. While considerable literature exists for Australian women as a whole, there has been little investigation of Australian Indigenous women, with only two research studies related to Indigenous women's experiences of menopause identified. Differences in biocultural experience of menopause around the world suggest the importance of biocultural research. For the Indigenous women of Australia, the relative contribution of culture, social disadvantage and poor general health compared with non-Indigenous women to the experience of menopause is unknown. As such, further research and understanding of the experience of Indigenous women around Australia is needed. This information could assist individuals, families, cultural groups and healthcare providers to enhance management and support for Indigenous Australian women.


Down-regulation of vitamin D receptor in mammospheres: implications for vitamin D resistance in breast cancer and potential for combination therapy.
Vitamin D signaling in mammary cancer stem cells (MCSCs), which are implicated in the initiation and progression of breast cancer, is poorly understood. In this study, we examined vitamin D signaling in mammospheres which are enriched in MCSCs from established breast cancer cell lines. Breast cancer cells positive for aldehyde dehydrogenase (ALDH(+)) had increased ability to form mammospheres compared to ALDH(-) cells. These mammospheres expressed MCSC-specific markers and generated transplantable xenografts in nude mice. Vitamin D receptor (VDR) was significantly down-regulated in mammospheres, as well as in ALDH(+) breast cancer cells. TN aggressive human breast tumors as well as transplantable xenografts obtained from SKBR3 expressed significantly lower levels of VDR but higher levels of CD44 expression. Snail was up-regulated in mammospheres isolated from breast cancer cells. Inhibition of VDR expression by siRNA led to a significant change in key EMT-specific transcription factors and increased the ability of these cells to form mammospheres. On the other hand, over-expression of VDR led to a down-regulation of Snail but increased expression of E-cad and significantly compromised the ability of cells to form mammospheres. Mammospheres were relatively insensitive to treatment with 1,25-dihydroxyvitamin D (1,25D), the active form of vitamin D, compared to more differentiated cancer cells grown in presence of serum. Treatment of H-Ras transformed HMLE(HRas) cells with DETA NONOate, a nitric oxide (NO)-donor led to induction of MAP-kinase phosphatase -1 (MKP-1) and dephosphorylation of ERK1/2 in the mammospheres. Combined treatment of these cells with 1,25D and a low-concentration of DETA NONOate led to a significant decrease in the overall size of mammospheres and reduced tumor volume in nude mice. Our findings therefore, suggest that combination therapy using 1,25D with drugs specifically targeting key survival pathways in MCSCs warrant testing in prospective clinical trial for treatment of aggressive breast cancer.


Elastic Relaxation of Fluid-Driven Cracks and the Resulting Backflow.
Cracks filled with fluid propagation when the pressurized fluid is injected into the crack. Subsequently, when the fluid inlet is exposed to a lower pressure, the fluid flows backwards (backflow) and the crack closes due to the elastic relaxation of the solid. Here we study the dynamics of the crack closure during the backflow. We find that the crack radius remains constant and the fluid volume in the crack decreases with time in a power-law manner at late times. The balance between the viscous stresses in the fluid and elastic stresses in the fluid and the elastic stresses in the solid yields a scaling law that agrees with the experimental results for different fluid viscosities, Young's moduli of the solid, and initial radii of the cracks. Furthermore, we visualize the time-dependent crack shapes, and the convergence to a universal dimensionless shape demonstrates the self-similarity of the crack shapes during the backflow process.


Dietary intakes and risk of lymphoid and myeloid leukemia in the European Prospective Investigation into Cancer and Nutrition (EPIC).
The etiology of leukemias cannot entirely be explained by known risk factors, including ionizing radiation, benzene exposure, and infection with human T cell leukemia virus. A number of studies suggested that diet influences the risk of adult leukemias. However, results have been largely inconsistent. We examined the potential association between dietary factors and risk of leukemias among participants of the European Prospective Investigation into Cancer and Nutrition study. Among the 477,325 participants with mean follow-up of 11.34 yr (SD = 2.47), 773 leukemias (373 and 342 cases of lymphoid and myeloid leukemia, respectively) were identified. Diet over the previous 12 mo was assessed at baseline using a validated country-specific dietary questionnaire. Cox proportional hazards regression was used to explore the association between dietary factors that have previously been associated with leukemia risk, including red and processed meat, poultry, offal, fish, dairy products, vegetables, fruits, and seeds/nuts, and risk of both lymphoid and myeloid leukemias. No significant associations were observed between dietary measures and total, lymphoid, and myeloid leukemias. Additional subtype analyses showed no dietary association with risk of major subtypes of leukemias. In summary, this study did not support a possible link between selected dietary factors and risk of leukemias.


Effect of soil permeability on virus removal through soil columns.
Laboratory experiments were performed on four different soils, using 100 cm long columns, to determine the extent of virus movement when wastewater percolated through the soils at various hydraulic flow rates. Unchlorinated secondary sewage effluent seeded with either poliovirus type 1 (strain LSc) or echovirus type 1 (isolate V239) was continuously applied to soil columns for 3 to 4 days at constant flow rates. Water samples were extracted daily from ceramic samplers at various depths of the column for the virus assay. The effectiveness of virus removal from wastewater varied greatly among the different soil types but appeared to be largely related to hydraulic flow rates. At a flow rate of 33 cm/day, Anthony sandy loam removed 99% of seeded poliovirus within the first 7 cm of the column. At flow rates of 300 cm/day and above, Rubicon sand gave the poorest removal of viruses; less than 90% of the seeded viruses were removed by passage of effluent through the entire length of the soil column. By linear regression analyses, the rate of virus removal in soil columns was found to be negatively correlated with the flow of the percolating sewage effluent. There was no significant difference in rate of removal between poliovirus and echovirus in soil columns 87 cm long. The rate of virus removal in the upper 17 cm of the soil column was found to be significantly greater than in the lower depths of the soil column. This study suggests that the flow rate of water through the soil may be the most important factor in predicting the potential of virus movement into the groundwater. Furthermore, the length of the soil column is critical in obtaining useful data to predict virus movement into groundwater.


Forest water use and water use efficiency at elevated CO2 : a model-data intercomparison at two contrasting temperate forest FACE sites.
Predicted responses of transpiration to elevated atmospheric CO2 concentration (eCO2 ) are highly variable amongst process-based models. To better understand and constrain this variability amongst models, we conducted an intercomparison of 11 ecosystem models applied to data from two forest free-air CO2 enrichment (FACE) experiments at Duke University and Oak Ridge National Laboratory. We analysed model structures to identify the key underlying assumptions causing differences in model predictions of transpiration and canopy water use efficiency. We then compared the models against data to identify model assumptions that are incorrect or are large sources of uncertainty. We found that model-to-model and model-to-observations differences resulted from four key sets of assumptions, namely (i) the nature of the stomatal response to elevated CO2 (coupling between photosynthesis and stomata was supported by the data); (ii) the roles of the leaf and atmospheric boundary layer (models which assumed multiple conductance terms in series predicted more decoupled fluxes than observed at the broadleaf site); (iii) the treatment of canopy interception (large intermodel variability, 2-15%); and (iv) the impact of soil moisture stress (process uncertainty in how models limit carbon and water fluxes during moisture stress). Overall, model predictions of the CO2 effect on WUE were reasonable (intermodel μ = approximately 28% ± 10%) compared to the observations (μ = approximately 30% ± 13%) at the well-coupled coniferous site (Duke), but poor (intermodel μ = approximately 24% ± 6%; observations μ = approximately 38% ± 7%) at the broadleaf site (Oak Ridge). The study yields a framework for analysing and interpreting model predictions of transpiration responses to eCO2 , and highlights key improvements to these types of models.


Mid-infrared gas sensing using a photonic bandgap fiber.
We demonstrate methane sensing based on Fourier transform infrared spectroscopy using a hollow-core photonic bandgap fiber guiding in the mid-infrared and idler pulses from a femtosecond optical parametric oscillator. Transmission measurements are presented for several fibers, and sensing is demonstrated using a fiber whose bandgap overlaps the methane fundamental absorption lines. The gas filling process of the air core is described, and qualitative methane concentrations measurements to 1000 ppm (parts in 10(6)) are reported. Operation down to 50 ppm based on our current experiment is predicted.


Five-Year Outcomes From the Randomized, Phase III Trials CheckMate 017 and 057: Nivolumab Versus Docetaxel in Previously Treated Non-Small-Cell Lung Cancer.
Immunotherapy has revolutionized the treatment of advanced non-small-cell lung cancer (NSCLC). In two phase III trials (CheckMate 017 and CheckMate 057), nivolumab showed an improvement in overall survival (OS) and favorable safety versus docetaxel in patients with previously treated, advanced squamous and nonsquamous NSCLC, respectively. We report 5-year pooled efficacy and safety from these trials. Patients (N = 854; CheckMate 017/057 pooled) with advanced NSCLC, ECOG PS ≤ 1, and progression during or after first-line platinum-based chemotherapy were randomly assigned 1:1 to nivolumab (3 mg/kg once every 2 weeks) or docetaxel (75 mg/m2 once every 3 weeks) until progression or unacceptable toxicity. The primary end point for both trials was OS; secondary end points included progression-free survival (PFS) and safety. Exploratory landmark analyses were investigated. After the minimum follow-up of 64.2 and 64.5 months for CheckMate 017 and 057, respectively, 50 nivolumab-treated patients and nine docetaxel-treated patients were alive. Five-year pooled OS rates were 13.4% versus 2.6%, respectively; 5-year PFS rates were 8.0% versus 0%, respectively. Nivolumab-treated patients without disease progression at 2 and 3 years had an 82.0% and 93.0% chance of survival, respectively, and a 59.6% and 78.3% chance of remaining progression-free at 5 years, respectively. Treatment-related adverse events (TRAEs) were reported in 8 of 31 (25.8%) nivolumab-treated patients between 3-5 years of follow-up, seven of whom experienced new events; one (3.2%) TRAE was grade 3, and there were no grade 4 TRAEs. At 5 years, nivolumab continued to demonstrate a survival benefit versus docetaxel, exhibiting a five-fold increase in OS rate, with no new safety signals. These data represent the first report of 5-year outcomes from randomized phase III trials of a programmed death-1 inhibitor in previously treated, advanced NSCLC.


Fully orthogonal optical-code multiplex for broadcasting.
A novel method of optical-code multiplex transmission from a central location is proposed. It has the advantages that the receivers can be configured to any channel quickly, the channels have in principle zero cross talk, and the bandwidth-expansion factors are less than for other optical-code-division multiple access arrangements. The proposed method is based on arrays of optoelectronic switching detectors that are at present under development for broadband matrix switching.


Asymmetric temporal integration of layer 4 and layer 2/3 inputs in visual cortex.
Neocortical neurons in vivo receive concurrent synaptic inputs from multiple sources, including feedforward, horizontal, and feedback pathways. Layer 2/3 of the visual cortex receives feedforward input from layer 4 and horizontal input from layer 2/3. Firing of the pyramidal neurons, which carries the output to higher cortical areas, depends critically on the interaction of these pathways. Here we examined synaptic integration of inputs from layer 4 and layer 2/3 in rat visual cortical slices. We found that the integration is sublinear and temporally asymmetric, with larger responses if layer 2/3 input preceded layer 4 input. The sublinearity depended on inhibition, and the asymmetry was largely attributable to the difference between the two inhibitory inputs. Interestingly, the asymmetric integration was specific to pyramidal neurons, and it strongly affected their spiking output. Thus via cortical inhibition, the temporal order of activation of layer 2/3 and layer 4 pathways can exert powerful control of cortical output during visual processing.


Sensitivity to Movement-Evoked Pain and Multi-Site Pain are Associated with Work-Disability Following Whiplash Injury: A Cross-Sectional Study.
Objectives Previous research has shown that sensitivity to movement-evoked pain is associated with higher scores on self-report measures of disability in individuals who have sustained whiplash injuries. However, it remains unclear whether sensitivity to movement-evoked pain is associated with work-disability. The aim of the present study was to examine the relation between sensitivity to movement-evoked pain and occupational status in individuals receiving treatment for whiplash injury. Methods A sample of 105 individuals with whiplash injuries participated in a testing session where different measures of pain (i.e. spontaneous pain, multi-site pain, sensitivity to movement-evoked pain) were collected during the performance of a simulated occupational lifting task. Results Hierarchical logistic regression analysis revealed that the measures of multisite pain and sensitivity to movement-evoked pain made significant independent contributions to the prediction of work-disability. Discussion The findings suggest that including measures of multisite pain and sensitivity to movement evoked pain in assessment protocols has the potential to increase the value of pain assessments for the prediction of occupational disability associated with whiplash injury. Clinical and theoretical implications of the findings are addressed.


Subcutaneous adipose fatty acid profiles and related rumen bacterial populations of steers fed red clover or grass hay diets containing flax or sunflower-seed.
Steers were fed 70∶30 forage∶concentrate diets for 205 days, with either grass hay (GH) or red clover silage (RC), and either sunflower-seed (SS) or flaxseed (FS), providing 5.4% oil in the diets. Compared to diets containing SS, FS diets had elevated (P<0.05) subcutaneous trans (t)-18:1 isomers, conjugated linoleic acids and n-6 polyunsaturated fatty acid (PUFA). Forage and oilseed type influenced total n-3 PUFA, especially α-linolenic acid (ALA) and total non-conjugated diene biohydrogenation (BH) in subcutaneous fat with proportions being greater (P<0.05) for FS or GH as compared to SS or RC. Of the 25 bacterial genera impacted by diet, 19 correlated with fatty acids (FA) profile. Clostridium were most abundant when levels of conjugated linolenic acids, and n-3 PUFA's were found to be the lowest in subcutaneous fat, suggestive of their role in BH. Anerophaga, Fibrobacter, Guggenheimella, Paludibacter and Pseudozobellia were more abundant in the rumen when the levels of VA in subcutaneous fat were low. This study clearly shows the impact of oilseeds and forage source on the deposition of subcutaneous FA in beef cattle. Significant correlations between rumen bacterial genera and the levels of specific FA in subcutaneous fat maybe indicative of their role in determining the FA profile of adipose tissue. However, despite numerous correlations, the dynamics of rumen bacteria in the BH of unsaturated fatty acid and synthesis of PUFA and FA tissue profiles require further experimentation to determine if these correlations are consistent over a range of diets of differing composition. Present results demonstrate that in order to achieve targeted FA profiles in beef, a multifactorial approach will be required that takes into consideration not only the PUFA profile of the diet, but also the non-oil fraction of the diet, type and level of feed processing, and the role of rumen microbes in the BH of unsaturated fatty acid.


Processes of change in Acceptance and Commitment Therapy and Applied Relaxation for long-standing pain.
The utility of cognitive behavioural (CB) interventions for chronic pain has been supported in numerous studies. This includes Acceptance and Commitment Therapy (ACT), which has gained increased empirical support. Previous research suggests that improvements in pain catastrophizing and psychological inflexibility are related to improvements in treatment outcome in this type of treatment. Although a few studies have evaluated processes of change in CB-interventions, there is a particular need for mediation analyses that use multiple assessments to model change in mediators and outcome over time, and that incorporate the specified timeline between mediator and outcome in the data analytic model. This study used session-to-session assessments to evaluate if psychological inflexibility, catastrophizing, and pain intensity mediated the effects of treatment on pain interference. Analyses were based on data from a previously conducted randomized controlled trial (n = 60) evaluating the efficacy of ACT and Applied Relaxation (AR). A moderated mediation model based on linear mixed models was used to analyse the data. Neither catastrophizing nor pain intensity mediated changes in pain interference for any of the treatments. In contrast, psychological inflexibility mediated effects on outcome in ACT but not in AR. Results add to previous findings illustrating the role of psychological inflexibility as a mediator in ACT.


Clinical and biochemical effects of impeded androgen (oxymetholone) therapy of hereditary angioedema.
Daily therapy and alternate-day therapy with the attenuated androgen oxymetholone were compared in patients with hereditary angioedema (HAE). Fifteen of 16 patients who experienced at least monthly attacks of HAE without treatment were asymptomatic on administration of 5 mg oxymetholene daily. When 13 of the patients who had been maintained asymptomatically on 5 mg oxymetholone daily were advanced to a treatment schedule of 5 mg every other day, seven attacks occurred during a cummulative 50 mo of therapy. The adverse effects that occurred with daily oxymetholone therapy largely subsided when the patients received alternate-day therapy, while a significant mean rise in C4 protein and function occurred only on daily therapy. Statistically significant mean increases in serum levels of C1INH occurred with daily therapy and were maintained with alternate-day therapy. Clinical benefit can be obtained with a treatment program that does not produce a statistically significant rise in C4 protein or function and does not raise C1INH to the lower limit of normal. The finding that alternate-day therapy diminished the side effects of the drug while affording a substantial reduction in the incidence and severity of attacks indicates the feasibility of this therapeutic approach.


Detecting RNA modifications in the epitranscriptome: predict and validate.
RNA modifications are emerging players in the field of post-transcriptional regulation of gene expression, and are attracting a comparable degree of research interest to DNA and histone modifications in the field of epigenetics. We now know of more than 150 RNA modifications and the true potential of a few of these is currently emerging as the consequence of a leap in detection technology, principally associated with high-throughput sequencing. This Review outlines the major developments in this field through a structured discussion of detection principles, lays out advantages and drawbacks of new high-throughput methods and presents conventional biophysical identification of modifications as meaningful ways for validation.


Circuitry linking the Csr and stringent response global regulatory systems.
CsrA protein regulates important cellular processes by binding to target mRNAs and altering their translation and/or stability. In Escherichia coli, CsrA binds to sRNAs, CsrB and CsrC, which sequester CsrA and antagonize its activity. Here, mRNAs for relA, spoT and dksA of the stringent response system were found among 721 different transcripts that copurified with CsrA. Many of the transcripts that copurified with CsrA were previously determined to respond to ppGpp and/or DksA. We examined multiple regulatory interactions between the Csr and stringent response systems. Most importantly, DksA and ppGpp robustly activated csrB/C transcription (10-fold), while they modestly activated csrA expression. We propose that CsrA-mediated regulation is relieved during the stringent response. Gel shift assays confirmed high affinity binding of CsrA to relA mRNA leader and weaker interactions with dksA and spoT. Reporter fusions, qRT-PCR and immunoblotting showed that CsrA repressed relA expression, and (p)ppGpp accumulation during stringent response was enhanced in a csrA mutant. CsrA had modest to negligible effects on dksA and spoT expression. Transcription of dksA was negatively autoregulated via a feedback loop that tended to mask CsrA effects. We propose that the Csr system fine-tunes the stringent response and discuss biological implications of the composite circuitry.


Cancer biomarker detection: recent achievements and challenges.
The early detection of cancer can significantly reduce cancer mortality and saves lives. Thus, a great deal of effort has been devoted to the exploration of new technologies to detect early signs of the disease. Cancer biomarkers cover a broad range of biochemical entities, such as nucleic acids, proteins, sugars, small metabolites, and cytogenetic and cytokinetic parameters, as well as entire tumour cells found in the body fluid. They can be used for risk assessment, diagnosis, prognosis, and for the prediction of treatment efficacy and toxicity and recurrence. In this review, we provide an overview of recent advances in cancer biomarker detection. Several representative examples using different approaches for each biomarker have been reviewed, and all these cases demonstrate that the multidisciplinary technology-based cancer diagnostics are becoming an increasingly relevant alternative to traditional techniques. In addition, we also discuss the unsolved problems and future challenges in the evaluation of cancer biomarkers. Clearly, solving these hurdles requires great effort and collaboration from different communities of chemists, physicists, biologists, clinicians, material-scientists, and engineering and technical researchers. A successful outcome will result in the realization of point-of-care diagnosis and individualized treatment of cancers by non-invasive and convenient tests in the future.


Ontogeny of circadian organization in the rat.
The mammalian circadian system is orchestrated by a master pacemaker in the brain, but many peripheral tissues also contain independent or quasi-independent circadian oscillators. The adaptive significance of clocks in these structures must lie, in large part, in the phase relationships between the constituent oscillators and their micro- and macroenvironments. To examine the relationship between postnatal development, which is dependent on endogenous programs and maternal/environmental influences, and the phase of circadian oscillators, the authors assessed the circadian phase of pineal, liver, lung, adrenal, and thyroid tissues cultured from Period 1-luciferase (Per1-luc ) rat pups of various postnatal ages. The liver, thyroid, and pineal were rhythmic at birth, but the phases of their Per1-luc expression rhythms shifted remarkably during development. To determine if the timing of the phase shift in each tissue could be the result of changing environmental conditions, the behavior of pups and their mothers was monitored. The circadian phase of the liver shifted from the day to night around postnatal day (P) 22 as the pups nursed less during the light and instead ate solid food during the dark. Furthermore, the phase of Per1-luc expression in liver cultures from nursing neonates could be shifted experimentally from the day to the night by allowing pups access to the dam only during the dark. Peak Per1-luc expression also shifted from midday to early night in thyroid cultures at about P20, concurrent with the shift in eating times. The phase of Per1-luc expression in the pineal gland shifted from day to night coincident with its sympathetic innervation at around P5. Per1-luc expression was rhythmic in adrenal cultures and peaked around the time of lights-off throughout development; however, the amplitude of the rhythm increased at P25. Lung cultures were completely arrhythmic until P12 when the pups began to leave the nest. Taken together, the data suggest that the molecular machinery that generates circadian oscillations matures at different rates in different tissues and that the phase of at least some peripheral organs is malleable and may shift as the organ's function changes during development.


Sperm competition experiments between lines of crickets producing different sperm lengths.
Sperm numbers can be important determinants of fertilization success in sperm competition. However, the importance of variation in sperm size is less well understood. Sperm size varies significantly both between and within species and comparative studies have suggested that some of this variance can be explained by sperm competition. In this study we examine whether variation in sperm length has consequences for fertilization precedence using controlled sperm competition experiments in the field cricket Gryllus bimaculatus. This species is an ideal model for such investigations because the mechanism of sperm competition generates complete mixing of different males' spermatozoa in the female (thereby allowing individual sperm to express their own competitive abilities). We successfully bred lines of crickets, the males of which produced short, medium and long sperm types with narrow and non-overlapping distributions. Males of different lines were then sequentially mated with control females in order to create two-male sperm competitions. The paternity outcomes of these competitions were measured after matings using an irradiated male technique (with a full reciprocal design that controls for natural fertility and any irradiation effects on gamete competitiveness) over a 12 day oviposition period. However, having successfully bred diverging sperm length lines and competing males that differed in sperm length, we found no evidence that a male's sperm size explained any of the variation in their relative fertilization success. Males from lines producing longer sperm showed no fertilization advantage over males producing shorter sperm across 97 double matings. There was also no advantage for males producing a sperm length close to the population mean over those competitors whose sperm length had been selectively diverged across 63 matings.