Comparison of phenobarbital with bromide as a first-choice antiepileptic drug for treatment of epilepsy in dogs.
To compare efficacy and safety of treatment with phenobarbital or bromide as the first-choice antiepileptic drug (AED) in dogs. Double-blinded, randomized, parallel, clinical trial. 46 AED-naïve dogs with naturally occurring epilepsy. Study inclusion was based on age, history, findings on physical and neurologic examinations, and clinicopathologic test results. For either phenobarbital treatment (21 dogs) or bromide treatment (25), a 7-day loading dose period was initiated along with a maintenance dose, which was adjusted on the basis of monthly monitoring. Efficacy and safety outcomes were compared between times (baseline and study end [generally 6 months]) and between drugs. Phenobarbital treatment resulted in eradication of seizures (17/20 [85%]) significantly more often than did bromide (12/23 [52%]); phenobarbital treatment also resulted in a greater percentage decrease in seizure duration (88 ± 34%), compared with bromide (49 ± 75%). Seizure activity worsened in 3 bromide-treated dogs only. In dogs with seizure eradication, mean ± SD serum phenobarbital concentration was 25 ± 6 μg/mL (phenobarbital dosage, 4.1 ± 1.1 mg/kg [1.9 ± 0.5 mg/lb], p.o., q 12 h) and mean serum bromide concentration was 1.8 ± 0.6 mg/mL (bromide dosage, 31 ± 11 mg/kg [14 ± 5 mg/lb], p.o., q 12 h). Ataxia, lethargy, and polydipsia were greater at 1 month for phenobarbital-treated dogs; vomiting was greater for bromide-treated dogs at 1 month and study end. Both phenobarbital and bromide were reasonable first-choice AEDs for dogs, but phenobarbital was more effective and better tolerated during the first 6 months of treatment.


The prevalence of CDKN2A germ-line mutations and relative risk for cutaneous malignant melanoma: an international population-based study.
Germ-line mutations of CDKN2A have been identified as strong risk factors for melanoma in studies of multiple-case families. However, an assessment of their relative risk for melanoma in the general population has been difficult because they occur infrequently. We addressed this issue using a novel population-based case-control study design in which "cases" have incident second- or higher-order melanomas [multiple primary melanoma (MPM)] and "controls" have incident first primary melanoma [single primary melanoma (SPM)]. Participants were ascertained from nine geographic regions in Australia, Canada, Italy, and United States. In the 1,189 MPM cases and 2,424 SPM controls who were eligible and available for analysis, the relative risk of a subsequent melanoma among patients with functional mutations who have an existing diagnosis of melanoma, after adjustments for age, sex, center, and known phenotypic risk factors, is estimated to be 4.3 (95% confidence interval, 2.3-7.7). The odds ratio varied significantly depending on the type of mutation involved. The results suggest that the relative risk of mutation carriers in the population may be lower than currently believed and that different mutations on the CDKN2A gene may confer substantially different risks of melanoma.


A simple iterative model accurately captures complex trapline formation by bumblebees across spatial scales and flower arrangements.
Pollinating bees develop foraging circuits (traplines) to visit multiple flowers in a manner that minimizes overall travel distance, a task analogous to the travelling salesman problem. We report on an in-depth exploration of an iterative improvement heuristic model of bumblebee traplining previously found to accurately replicate the establishment of stable routes by bees between flowers distributed over several hectares. The critical test for a model is its predictive power for empirical data for which the model has not been specifically developed, and here the model is shown to be consistent with observations from different research groups made at several spatial scales and using multiple configurations of flowers. We refine the model to account for the spatial search strategy of bees exploring their environment, and test several previously unexplored predictions. We find that the model predicts accurately 1) the increasing propensity of bees to optimize their foraging routes with increasing spatial scale; 2) that bees cannot establish stable optimal traplines for all spatial configurations of rewarding flowers; 3) the observed trade-off between travel distance and prioritization of high-reward sites (with a slight modification of the model); 4) the temporal pattern with which bees acquire approximate solutions to travelling salesman-like problems over several dozen foraging bouts; 5) the instability of visitation schedules in some spatial configurations of flowers; 6) the observation that in some flower arrays, bees' visitation schedules are highly individually different; 7) the searching behaviour that leads to efficient location of flowers and routes between them. Our model constitutes a robust theoretical platform to generate novel hypotheses and refine our understanding about how small-brained insects develop a representation of space and use it to navigate in complex and dynamic environments.


Enhancement of tissue plasminogen activator-catalyzed plasminogen activation by Escherichia coli S fimbriae associated with neonatal septicaemia and meningitis.
The effect of Escherichia coli strains isolated from blood and cerebrospinal fluid of septic infants on plasminogen activation was studied. These strains typically carry a filamentous surface protein, S fimbria, that has formerly been shown to bind to endothelial cells and interact with plasminogen. The bacteria effectively promoted plasminogen activation by tissue plasminogen activator (t-PA) which was inhibited by epsilon-aminocaproic acid. A recombinant strain expressing S fimbriae accelerated t-PA-catalyzed plasminogen activation to a similar extent as did the wild-type strains whereas the nonfimbriate recipient strain had no effect. After incubation with t-PA and plasminogen, the S-fimbriate strain displayed bacterium-bound plasmin activity whereas the nonfimbriate strain did not. Bacterium-associated plasmin generation was also observed with a strain expressing mutagenized S fimbriae that lack the cell-binding subunit SfaS but not with a strain lacking the major subunit SfaA. Both t-PA and plasminogen bound to purified S fimbriae in a lysine-dependent manner and purified S fimbriae accelerated t-PA-catalyzed plasminogen activation. The results indicate that E. coli S fimbriae form a complex with t-PA and plasminogen which enhances the rate of plasminogen activation and generates bacterium-bound plasmin. This may promote bacterial invasion and persistence in tissues and contribute to the systemic activation of fibrinolysis in septicaemia.


A model to measure insulin effects on glucose transport and phosphorylation in muscle: a three-tracer study.
We studied five healthy subjects with perfused forearm and euglycemic clamp techniques in combination with a three-tracer (D-[12C]mannitol, not transportable; 3-O-[14C]methyl-D-glucose, transportable but not metabolizable; D-[3-3H]glucose, transportable and metabolizable) intra-arterial pulse injection to assess transmembrane transport and intracellular phosphorylation of glucose in vivo in human muscle. The washout curves of the three tracers were analyzed with a multicompartmental model. A priori identifiability analysis of the tracer model shows that the rate constants of glucose transport into and out of the cells and of glucose phosphorylation are uniquely identifiable. Tracer model parameters were estimated by a nonlinear least-squares parameter estimation technique. We then solved for the tracee model and estimated bidirectional transmembrane transport glucose fluxes, glucose intracellular phosphorylation, extracellular and intracellular volumes of glucose distribution, and extracellular and intracellular glucose concentrations. Physiological hyperinsulinemia (473 +/- 22 pM) caused 2.7-fold (63.1 +/- 7.2 vs. 23.4 +/- 6.1 mumol.min-1.kg-1, P < 0.01) and 5.1-fold (42.5 +/- 5.8 vs. 8.4 +/- 2.2 mumol.min-1.kg-1, P < 0.01) increases in transmembrane influx and intracellular phosphorylation of glucose, respectively. Extracellular distribution volume and concentration of glucose were unchanged, whereas intracellular distribution volume of glucose was increased (approximately 2-fold) and intracellular glucose concentration was almost halved by hyperinsulinemia. In summary, 1) a multicompartment model of three-tracer kinetic data can quantify transmembrane glucose fluxes and intracellular glucose phosphorylation in human muscle; and 2) physiological hyperinsulinemia stimulates both transport and phosphorylation of glucose and, in doing so, amplifies the role of glucose transport as a rate-determining step of muscle glucose uptake.


Influence of QuEChERS modifications on recovery and matrix effect during the multi-residue pesticide analysis in soil by GC/MS/MS and GC/ECD/NPD.
A QuEChERS extraction followed by GC/MS/MS and GC-μECD/NPD for 216 pesticide and metabolites determination in soil simultaneously were developed and compared. Volume of water, volume and polarity of solvent, and cleanup sorbents (C18, GCB, PSA) were optimized. The QuEChERS with and without purification step were applied to estimate effectiveness of the method. The recovery and matrix effect (ME) were critical parameters within each tested procedure. The optimal method without cleanup was validated. Accuracy (expressed as recovery), precision (expressed as RSD), linearity, LOQ, and uncertainty were determined. The recoveries at the three spiking levels using matrix-matched standards ranged between 65 and 116% with RSD ≤17 and 60-112% with RSD ≤18% for MS/MS and μEC/NP, respectively. The LOQ ranged from 0.005-0.01 mg/kg for MS/MS to 0.05 mg/kg for μEC/NP. The ME for most of pesticides resulted in enhancement of the signal and depended on the analyte and detection system: MS/MS showed ME from -25 to 74%, while μEC/NP from -45 to 96%. A principal component analysis was performed to explain the relationships between physicochemical parameters and ME of 216 pesticides. The QuEChERS protocol without the cleanup step is a promising option to make the method less expensive and faster. This methodology was applied in routine analysis of 263 soil samples in which p,p' DDT was the most frequently detected (23.5% of samples) and pendimethalin with the highest concentration (1.63 mg/kg).


Discovery of a bacterial 5-methylcytosine deaminase.
5-Methylcytosine is found in all domains of life, but the bacterial cytosine deaminase from Escherichia coli (CodA) will not accept 5-methylcytosine as a substrate. Since significant amounts of 5-methylcytosine are produced in both prokaryotes and eukaryotes, this compound must eventually be catabolized and the fragments recycled by enzymes that have yet to be identified. We therefore initiated a comprehensive phylogenetic screen for enzymes that may be capable of deaminating 5-methylcytosine to thymine. From a systematic analysis of sequence homologues of CodA from thousands of bacterial species, we identified putative cytosine deaminases where a "discriminating" residue in the active site, corresponding to Asp-314 in CodA from E. coli, was no longer conserved. Representative examples from Klebsiella pneumoniae (locus tag: Kpn00632), Rhodobacter sphaeroides (locus tag: Rsp0341), and Corynebacterium glutamicum (locus tag: NCgl0075) were demonstrated to efficiently deaminate 5-methylcytosine to thymine with values of kcat/Km of 1.4 × 10(5), 2.9 × 10(4), and 1.1 × 10(3) M(-1) s(-1), respectively. These three enzymes also catalyze the deamination of 5-fluorocytosine to 5-fluorouracil with values of kcat/Km of 1.2 × 10(5), 6.8 × 10(4), and 2.0 × 10(2) M(-1) s(-1), respectively. The three-dimensional structure of Kpn00632 was determined by X-ray diffraction methods with 5-methylcytosine (PDB id: 4R85 ), 5-fluorocytosine (PDB id: 4R88 ), and phosphonocytosine (PDB id: 4R7W ) bound in the active site. When thymine auxotrophs of E. coli express these enzymes, they are capable of growth in media lacking thymine when supplemented with 5-methylcytosine. Expression of these enzymes in E. coli is toxic in the presence of 5-fluorocytosine, due to the efficient transformation to 5-fluorouracil.


Thermodynamic prediction of protein neutrality.
We present a simple theory that uses thermodynamic parameters to predict the probability that a protein retains the wild-type structure after one or more random amino acid substitutions. Our theory predicts that for large numbers of substitutions the probability that a protein retains its structure will decline exponentially with the number of substitutions, with the severity of this decline determined by properties of the structure. Our theory also predicts that a protein can gain extra robustness to the first few substitutions by increasing its thermodynamic stability. We validate our theory with simulations on lattice protein models and by showing that it quantitatively predicts previously published experimental measurements on subtilisin and our own measurements on variants of TEM1 beta-lactamase. Our work unifies observations about the clustering of functional proteins in sequence space, and provides a basis for interpreting the response of proteins to substitutions in protein engineering applications.


Thermodynamic modelling predicts energetic bottleneck for seabirds wintering in the northwest Atlantic.
Studying the energetics of marine top predators such as seabirds is essential to understand processes underlying adult winter survival and its impact on population dynamics. Winter survival is believed to be the single most important life-history trait in long-lived species but its determinants are largely unknown. Seabirds are inaccessible during this season, so conventional metabolic studies are extremely challenging and new approaches are needed. This paper describes and uses a state-of-the-art mechanistic model, Niche Mapper, to predict energy expenditure and food requirements of the two main seabird species wintering in the northwest Atlantic. We found that energy demand increased throughout the winter phase in both species. Across this period, mean estimated daily energy requirements were 1306 kJ day(-1) for Brünnich's guillemots (Uria lomvia) and 430 kJ day(-1) for little auks (Alle alle) wintering off Greenland and Newfoundland. Mean estimated daily food requirements were 547 g wet food day(-1) for Brünnich's guillemots, and 289 g wet food day(-1) for little auks. For both species and both wintering sites, our model predicts a sharp increase in energy expenditure between November and December, primarily driven by climatic factors such as air temperature and wind speed. These findings strongly suggest the existence of an energetic bottleneck for North Atlantic seabirds towards the end of the year, a challenging energetic phase which might explain recurrent events of winter mass-mortality, so called 'seabird winter wrecks'. Our study therefore emphasizes the relevance of thermodynamics/biophysical modelling for investigating the energy balance of wintering marine top predators and its interplay with survival and population dynamics in the context of global change.


A qualitative study of factors influencing antimicrobial prescribing by non-consultant hospital doctors.
To determine the factors that influence non-consultant hospital doctors (NCHDs) in their decision to prescribe antimicrobial agents. A qualitative study using semi-structured interviews centred on a life grid tracking the medical career of 22 NCHDs employed by a university teaching hospital in the west of Ireland. Early in their careers NCHD prescribing is based primarily on the immediate influence of more senior colleagues. Recollection of formalized undergraduate teaching and hospital guidelines are a very minor influence. As their career progressed and they exercise greater autonomy, personal experience becomes the major influence on prescribing decisions. Hospital guidelines are a minor influence. Participants consider that undergraduate teaching needs to be more practical and taught in a way that is easier to apply to on-ward situations and that hospital prescribing guidelines need to be presented in a 'user-friendly' format and adherence to the guidelines needs to be promoted. The key influences on antimicrobial prescribing by NCHDs are informal. New approaches are required to ensure that formal training and hospital guidelines on antimicrobial prescribing are more influential in shaping antimicrobial prescribing practice.


Identification, subcellular localization, and developmental studies of oleosins in the anther of Brassica napus.
mRNAs encoding putative oleosins have been detected in the tapetum of developing anthers in Brassica and Arabidopsis, but the authentic proteins have not been previously documented. Antibodies against a synthetic 15-residue polypeptide that represents a portion of the putative tapetum oleosins encoded by two cloned Brassica napus genes were raised. Using these antibodies for immunoblotting after SDS-PAGE of the sporophytic extracts of B. napus developing anthers, two oleosins of approximately 48 and 45 kDa were detected. These two oleosins were judged to be the putative oleosins encoded by cloned Brassica genes because of their identical N-terminal sequences. The two oleosins were present in the anthers only during the developmental stage when the tapetum cells were packed with organelles. A fraction of lowdensity organelles was isolated from the developing anthers by flotation centrifugation. The fraction contained plastoglobule-filled plastids and lipid-containing particles. The structures of these two isolated organelles were similar to those in situ in the tapetum cells. Of subcellular fractions of the anther homogenate, the two oleosins were present exclusively in the low-density organelle fraction. They were absent in the surface fractions of the developing microspores and the mature pollen, although fragmented oleosin molecules were earlier reported to be present on the pollen. By immunocytochemistry, immunogold particles were found largely on the periphery of the plastoglobuli inside the plastids in the tapetum cells. The antibodies also detected oleosins on the surface of storage oil bodies inside the maturing microspores. Apparently, the gametophytic microspore oil-body oleosins share common epitopes at the generally non-conserved C-terminal domain with the sporophytic tapetum oleosins.


Runaway Youth: Caring for the Nation's Largest Segment of Missing Children.
The largest segment of missing children in the United States includes runaways, children who run away from home, and thrownaways, children who are told to leave or stay away from home by a household adult. Although estimates vary, as many as 1 in 20 youth run away from home annually. These unaccompanied youth have unique health needs, including high rates of trauma, mental illness, substance use, pregnancy, and sexually transmitted infections. While away, youth who run away are at high risk for additional trauma, victimization, and violence. Runaway and thrownaway youth have high unmet health care needs and limited access to care. Several populations are at particular high risk for runaway episodes, including victims of abuse and neglect; lesbian, gay, bisexual, transgender, and questioning youth; and youth in protective custody. Pediatricians and other health care professionals have a critical role to play in supporting runaway youth, addressing their unique health needs, fostering positive relationships within their families and with other supportive adults, and connecting them with available community resources. This report provides clinical guidance for pediatricians and other health care professionals regarding (1) the identification of adolescents who are at risk for running away or being thrown away and (2) the management of the unique medical, mental health, and social needs of these youth. In partnership with national, state, and local resources, pediatricians can significantly reduce risk and improve long-term outcomes for runaway youth.


Differential effects of yfgL mutation on Escherichia coli outer membrane proteins and lipopolysaccharide.
YfgL together with NlpB, YfiO, and YaeT form a protein complex to facilitate the insertion of proteins into the outer membrane of Escherichia coli. Without YfgL, the levels of OmpA, OmpF, and LamB are significantly reduced, while OmpC levels are slightly reduced. In contrast, the level of TolC significantly increases in a yfgL mutant. When cells are depleted of YaeT or YfiO, levels of all outer membrane proteins examined, including OmpC and TolC, are severely reduced. Thus, while the assembly pathways of various nonlipoprotein outer membrane proteins may vary through the step involving YfgL, all assembly pathways in Escherichia coli converge at the step involving the YaeT/YfiO complex. The negative effect of yfgL mutation on outer membrane proteins may in part be due to elevated sigma E activity, which has been shown to downregulate the synthesis of various outer membrane proteins while upregulating the synthesis of periplasmic chaperones, foldases, and lipopolysaccharide. The data presented here suggest that the yfgL effect on outer membrane proteins also stems from a defective assembly apparatus, leading to aberrant outer membrane protein assembly, except for TolC, which assembles independent of YfgL. Consistent with this view, the simultaneous absence of YfgL and the major periplasmic protease DegP confers a synthetic lethal phenotype, presumably due to the toxic accumulation of unfolded outer membrane proteins. The results support the hypothesis that TolC and major outer membrane proteins compete for the YaeT/YfiO complex, since mutations that adversely affect synthesis or assembly of major outer membrane proteins lead to elevated TolC levels.


The moral economy of contemporary working-class adolescence: managing symbolic capital in a French public 'Adolescent Centre'.
Working-class adolescents of French urban peripheries are key figures in a new social debate that reactivates the nineteenth century spectre of 'dangerous' classes to be controlled. Since the 1990s, French social counselling has privileged two modalities of response: taking account of suffering and government by listening and speech. We hypothesize that the contemporary moral economy allows for social interactions that go beyond social control and institutional domination. This is partly because professionals engaged in this moral undertaking may keep a critical distance, and partly because the concerned populations aren't necessarily devoid of resources to advance their interests or incapable of resistance. The concept of moral economy, coupled with the ethnographic method, is heuristic for fully comprehending the complexity of these issues and their stakes. Our fieldwork was centred on a French Adolescent Centre in an impoverished commune in Paris's periphery, from January 2010 through March 2011. These institutions were established in the early 2000s to respond to adolescent 'suffering' by crossing social work and psychiatry. Adolescents, parents, and other institutions (especially schools) solicit the professionally diverse staff for assistance, which in turn may take on cases and/or make referrals to other support institutions. By paying attention to all the scenes upon which the story of a counselled adolescent evolves, and bearing more general social evolutions in mind by applying the concept of moral economy, we can consider the multiplicity of seemingly contradictory processes as a whole. We see the destabilization of parents and their loss of symbolic capital, partly due to the norms of contemporary parenthood and partly due to the stigmatization of working-class adolescence. But we also discern possibilities for expressing sentiments of injustice and humiliation, for increasing symbolic capital, and in some cases a reappropriation of the system, particularly in trajectories marked by a will for social ascension.


"Satanism is witchcraft's younger sibling": Changing perceptions of natural and supernatural anaemia causality in Malawian children.
In countries of sub-Saharan Africa, many children are admitted to hospital with severe forms of anaemia. The late hospital admissions of anaemic children contribute significantly to child morbidity and mortality in these countries. This qualitative study explores local health beliefs and traditional treatment practices that may hinder timely seeking of hospital care for anaemic children. In January of 2019, nine focus group discussions were conducted with 90 participants in rural communities of Malawi. The participants represented four groups of caregivers; mothers, fathers, grandmothers and grandfathers of children under the age of five. The Malawian medical landscape is comprised of formal and informal therapeutic alternatives-and this myriad of modalities is likely to complicate the healthcare choices of caregivers. When dealing with child illness, many participants reported how they would follow a step-by-step, 'multi-try' therapeutic pathway where a combination of biomedical and traditional treatment options were sought at varying time points depending on the perceived cause and severity of symptoms. The participants linked anaemia to naturalistic (malaria, poor nutrition and the local illnesses kakozi and kapamba), societal (the local illness msempho) and supernatural or personalistic (witchcraft and Satanism) causes. Most participants agreed that anaemia due to malaria and poor nutrition should be treated at hospital. As for local illnesses, many grandparents suggested herbal treatment offered by traditional healers, while the majority of parents would opt for hospital care. However, participants across all age groups claimed that anaemia caused by witchcraft and Satanism could only be dealt with by traditional healers or prayer, respectively. The multiple theories of anaemia causality combined with extensive use of and trust in traditional and complementary medicine may explain the frequent delay in admittance of anaemic children to hospital.


Persistence of effects of a brief intervention on parental restrictions of teen driving privileges.
The purpose of this study was to determine the extent to which effects of exposure to a brief intervention designed to increase parental restrictions on teen driving privileges persisted over time. A total of 658 parents and their 16 year old adolescents were recruited from a local motor vehicle administration (MVA) site as adolescents successfully tested for provisional licenses. At the MVA, parents completed written surveys about expected teen driving during the first month of provisional licensure. Afterwards, on weeks assigned as intervention, parents watched a video and were given the video and a driving agreement to take home. Both parents and teens completed follow up telephone interviews about communication, amounts, and limits on teen driving at one month (579 dyads), four months (529 dyads), and nine months (528 dyads). The results indicated that both intervention parents and teens were much more likely to report using a driving agreement at each follow up during the nine month period. Significant treatment group differences persisted for communication about driving, but effects related to limits on teen driving that were evident at one month declined over time. Reports for passenger, road, and overall limits remained significant at four months; fewer were present at nine months. There were no differences in amounts of teen driving at four or nine months. It is possible to reach parents through brief interventions at the MVA and successfully promote increases in initial parental restrictions on teen driving with modest persistence for at least four months.


Health-promotion theories in nutritional interventions for community-dwelling older adults: a systematic review.
To identify theories applied in interventions promoting healthy nutrition among community-dwelling older adults and determine the efficacy of theories in changing knowledge, attitudes, and behaviors. The PubMed, PsycINFO, Embase, and ERIC databases were searched for English articles from January 1990 to December 2015. Mono or multicomponent randomized controlled trial studies were included, whereas research on nutritional interventions related to acute or chronic diseases were excluded. Eight articles satisfied the inclusion criteria. Only three articles referred explicitly to health promotion theories. Nutritional programs varied in terms of contents, outcomes, lengths of interventions and follow-up. Pooling the results and identifying the most effective theories were therefore impossible. Although researchers and practitioners recognize the significance of theoretical models in guiding the health-promoting interventions, referring to a theoretical model for such interventions is still relatively recent.


The anti-inflammatory effect of exercise: its role in diabetes and cardiovascular disease control.
Chronic low-grade systemic inflammation is a feature of chronic diseases such as cardiovascular disease and type 2 diabetes. Regular exercise offers protection against all-cause mortality, primarily by protection against atherosclerosis and insulin resistance and there is evidence that physical training is effective as a treatment in patients with chronic heart diseases and type 2 diabetes. Regular exercise induces anti-inflammatory actions. During exercise, IL-6 (interleukin-6) is produced by muscle fibres. IL-6 stimulates the appearance in the circulation of other anti-inflammatory cytokines such as IL-1ra (interleukin-1 receptor antagonist) and IL-10 (interleukin-10) and inhibits the production of the pro-inflammatory cytokine TNF-alpha (tumour necrosis factor-alpha). In addition, IL-6 enhances lipid turnover, stimulating lipolysis as well as fat oxidation. It is suggested that regular exercise induces suppression of TNF-alpha and thereby offers protection against TNF-alpha-induced insulin resistance. Recently, IL-6 was introduced as the first myokine, defined as a cytokine, that is produced and released by contracting skeletal muscle fibres, exerting its effects in other organs of the body. Myokines may be involved in mediating the beneficial health effects against chronic diseases associated with low-grade inflammation such as diabetes and cardiovascular diseases.


Sperm DNA integrity in cancer patients before and after cytotoxic treatment.
We assessed sperm DNA fragmentation index (DFI) in cancer patients before and after treatment to evaluate if sperm DNA integrity is compromised by cancer itself or its treatment. In a prospective study, DFI was assessed in 127 patients diagnosed with testicular germ cell tumours (TGCT), Hodgkin's lymphoma (HL), non-Hodgkin's lymphoma (NHL) and various malignancies. The severity of cancer and tumour markers at diagnosis was recorded. Follow-up DFI after treatment was available in 52 patients who were mostly less severely affected. In patients diagnosed with TGCT, HL and various malignancies, pretreatment DFI levels were not significantly different from that of proven fertile controls, but in patients with NHL an increased DFI was found. An overall significant decrease in post-treatment DFI (13.2% range 5.0-70.5) compared with pretreatment values (17.1% range 5.1-66.6) was found (P = 0.040). In TGCT patients, post-treatment DFI was significantly higher in patients who were treated with radiotherapy (16.9% range 11.5-39.9) compared with that in patients treated with chemotherapy (CT) alone (10.9% range 5.5-39.9) (P = 0.037). In HL patients, the type of treatment or number of CT cycles was not associated with DFI. Overall, post-treatment DFI in cancer patients was not significantly different from that of proven fertile controls. In this study, the presence of cancer does not seem to negatively affect the sperm DNA integrity in TGCT and HL patients; only NHL patients showed increased DFI at the time of diagnosis compared with healthy controls. Our results confirm previous reports that DFI decreases significantly following various anti-cancer treatments. In contrast, radiotherapy in TGCT patients is associated with an increase in DFI compared with CT treatment alone.


Is it possible to predict the average surface hydrophobicity of a protein using only its amino acid composition?
Hydrophobicity is one of the most important physicochemical properties of proteins. Moreover, it plays a fundamental role in hydrophobic interaction chromatography, a separation technique that, at present time, is used in most industrial processes for protein purification as well as in laboratory scale applications. Although there are many ways of assessing the hydrophobicity value of a protein, recently, it has been shown that the average surface hydrophobicity (ASH) is an important tool in the area of protein separation and purification particularly in protein chromatography. The ASH is calculated based on the hydrophobic characteristics of each class of amino acid present on the protein surface. The hydrophobic characteristics of the amino acids are determined by a scale of aminoacidic hydrophobicity. In this work, the scales of Cowan-Whittaker and Berggren were studied. However, to calculate the ASH, it is necessary to have the three-dimensional protein structure. Frequently this data does not exist, and the only information available is the amino acid sequence. In these cases it would be desirable to estimate the ASH based only on properties extracted from the protein sequence. It was found that it is possible to predict the ASH from a protein to an acceptable level for many practical applications (correlation coefficient > 0.8) using only the aminoacidic composition. Two predictive tools were built: one based on a simple linear model and the other on a neural network. Both tools were constructed starting from the analysis of a set of 1982 non-redundant proteins. The linear model was able to predict the ASH for an independent subset with a correlation coefficient of 0.769 for the case of Cowan-Whittaker and 0.803 for the case of Berggren. On the other hand, the neural model improved the results shown by the linear model obtaining correlation coefficients of 0.831 and 0.836, respectively. The neural model was somewhat more robust than the linear model particularly as it gave similar correlation coefficients for both hydrophobicity scales tested, moreover, the observed variabilities did not overcome 6.1% of the mean square error. Finally, we tested our models in a set of nine proteins with known retention time in hydrophobic interaction chromatography. We found that both models can predict this retention time with correlation coefficients only slightly inferior (11.5% and 5.5% for the linear and the neural network models, respectively) than models that use the information about the three-dimensional structure of proteins.


Ca(2+)-dependent K(+) channels and Na(+)-K(+)-ATPase mediate H(2)O(2)- and superoxide-induced relaxations in canine trachealis.
We examined the ionic mechanisms underlying the responses of canine trachealis to superoxide (generated in vitro by using xanthine oxidase or added exogenously) and peroxide (generated spontaneously in vitro by the dismutation of superoxide or added exogenously). Although neither had any effect on resting tone, both triggered relaxations in carbachol-precontracted tissues. These relaxations were eliminated by catalase but were much less sensitive to the hydroxyl radical scavenger dimethylthiourea, indicating they were mediated primarily by peroxide. These relaxations were decreased in magnitude and/or slowed by nifedipine (10(-6) M), ouabain (10(-6) M), or tetraethylammonium (25 mM), but not by 4-aminopyridine (5 mM), and were small or absent in tissues precontracted with 30 mM KCl. Finally, peroxide triggered membrane hyperpolarization and elevated cytosolic concentration of Ca(2+) (primarily via release from the internal store). Thus peroxide-mediated relaxations seem to involve Ca(2+) release, opening of Ca(2+)-dependent K(+) channels, hyperpolarization, closure of Ca(2+) channels, and relaxation. In addition, some other free radical (hydroxyl radical?) may activate the Na(+)-K(+) pump, also hyperpolarizing the membrane and causing relaxation.


[Development of an instrument for the assessment of chronic care models].
To present the context, aim and process of designing the Instrument for the Assessment of Chronic Care Models (Instrumento de Evaluación de Modelos de Atención ante la Cronicidad [IEMAC]), which was developed to make the conceptual framework of the chronic care model operational in the Spanish national health system. The IEMAC was developed by a series of national experts with distinct profiles of expertise using qualitative research techniques. A matrix was built with the dimensions selected as basic for the new model. In each dimension, actions were identified and categorized, creating a taxonomy of components and interventions. The clarity and appropriateness of each intervention, and the degree of evidence to support it, were assessed. The resulting questionnaire was validated by other experts from diverse disciplines and settings. Finally, the IEMAC 1.0 was piloted at macro, meso and micro levels. The IEMAC is a tool to be self-administered by health organizations at macro, meso and micro levels. This instrument is composed of six dimensions, 27 components and 80 interventions, whose implementation is assessed with the aid of a scale that combines deployment, systematic evaluation, and orientation improvement. The IEMAC uses a systemic, population-based approach and integrates promotion, prevention, and coordination with social services. The IEMAC contains a set of interventions that can be used as a road map by decision makers, managers and clinicians interested in building a state-of-the-art chronic care model. At the same time, the IEMAC allows healthcare organizations to identify their baseline score and the progress achieved after improvement interventions.


Characterization of the in vitro responses of equine cecal longitudinal smooth muscle to endothelin-1.
To characterize the in vitro response of equine cecal longitudinal smooth muscle (CLSM) to endothelin (ET)-1 and assess the role of ETA and ETB receptors in those ET-1-induced responses. 36 horses without gastrointestinal tract disease. To determine cumulative concentration-response relationships, CLSM strips were suspended in tissue baths containing graded concentrations of ET-1 (10(-9) to 10(-6)M) with or without BQ-123 (ETA receptor antagonist); with or without IRL-1038 (ETB receptor antagonist); or with both antagonists at concentrations of 10(-9), 10(-7), and 10(-5)M. To determine the percentage change in baseline tension of CLSM, the areas under the curve during the 3-minute periods before and after addition of each dose were compared. Also, the effects of ET-1 and a combination of selective ETA and ETB receptor antagonists on electrically evoked contractions were studied. ET-1 caused sustained increases in CLSM tension in a concentration-dependent manner. Contractile responses to ET-1 were not significantly inhibited by either BQ-123 or IRL-1038 alone at any concentration; however, responses were significantly inhibited by exposure to the antagonists together at a concentration of 10(-5)M. Electrical field stimulation did not change the spontaneous contractile activity of CLSM and did not significantly alter the tissue response to ET-1, BQ-123, or IRL-1038. Results indicated that ET-1 has a contractile effect on equine CLSM that is mediated via ETA and ETB receptors. In vitro spontaneous contractions of equine CLSM apparently originate in the smooth muscle and not the enteric nervous system.


Practice Preferences for Glaucoma Surgery: A Survey of the American Glaucoma Society.
The purpose of this study is to assess surgical practice patterns among the American Glaucoma Society (AGS) membership. An anonymous online survey evaluating the use of glaucoma surgeries in various clinical settings was redistributed to AGS members. Survey responses were compared with prior results from 1996, 2002, and 2008 to determine shifts in surgical practice patterns. Questions were added to assess the preferred approach to primary incisional glaucoma surgery and phacoemulsification combined with glaucoma surgery. A total of 252 of 1091 (23%) subscribers to the AGS-net participated in the survey. Percentage use (mean±SD) of trabeculectomy with mitomycin C (MMC), glaucoma drainage device (GDD), and minimally invasive glaucoma surgery (MIGS) as an initial surgery in patients with primary open angle glaucoma was 59%±30%, 23%±23%, and 14%±20%, respectively. Phacoemulsification cataract extraction alone was the preferred surgical approach in 44%±32% of patients with primary open angle glaucoma and visually significant cataract, and phacoemulsification cataract extraction was combined with trabeculectomy with MMC in 24%±23%, with MIGS in 22%±27%, and with GDD in 9%±14%. Although trabeculectomy was selected most frequently to surgically manage glaucoma in 8 of 8 clinical settings in 1996, GDD was preferred in 7 of 8 clinical settings in 2016. The use of GDD has increased and that of trabeculectomy has concurrently decreased over the past 2 decades. Trabeculectomy with MMC is the most popular primary incisional surgery when performed alone or in combination with phacoemulsification cataract extraction. Surgeons frequently manage coexistent cataract and glaucoma with cataract extraction alone, rather than as a combined cataract and glaucoma procedure.


Comparative analysis of bone stresses and strains in the Intoss dental implant with and without a flexible internal post.
The clinical success of any dental implant is dependent upon the maintenance of good-quality bone supporting it. Previous studies have shown high values of strain around the neck of an implant under lateral loading. These high values may lead to fatigue damage and resorption in lower strength cancellous bone. In this study, the finite element method has been used to study the bone strain distribution around the following implants: (a) an Intoss dental implant, referred to as the 'standard' implant; (b) a comparative Branemark implant and (c) a modified Intoss implant with a central flexible post, referred to as the 'modified' implant. Three different bone distributions have been investigated under axial and lateral loading: (a) implant surrounded by cortical bone; (b) implant tip supported by cortical bone with a thin layer of cancellous bone along the length and top of the implant; (c) implant tip and top supported by cortical bone with a thin layer of cancellous bone along the remaining length. For the standard implant, similar maximum equivalent strain values were predicted for the bone surrounding a comparable length Branemark-type implant. Modification of the standard implant design to include a flexible central post resulted in a decrease in the maximum von Mises stresses and equivalent strains in the cancellous bone. It is postulated that this will reduce the likelihood of bone fatigue failure and subsequent resorption in this bone. Thus the proposed design change is predicted to be highly beneficial in terms of bone load transfer.


Sub-micrometre imaging of metal surface corrosion by scanning Kelvin nanoprobe.
The detection of the initial changes in the surface microstructure and local chemical properties connected to the corrosion process is possible using a high-resolution scanning Kelvin nanoprobe. This technique provides the simultaneous imaging of the topographical features and potential distribution across a surface at the sub-micrometer level. Nanoprobe measurements performed on samples of Al, Cu, Fe, Ni, Ag and Pt before and after exposure to different corroding solutions reveal the significant changes that take place during the first stages of the corrosion process. The similarities of the images obtained subsequent to corrosion are reflective of surface fractal behavior.


Pantropical climate interactions.
The El Niño-Southern Oscillation (ENSO), which originates in the Pacific, is the strongest and most well-known mode of tropical climate variability. Its reach is global, and it can force climate variations of the tropical Atlantic and Indian Oceans by perturbing the global atmospheric circulation. Less appreciated is how the tropical Atlantic and Indian Oceans affect the Pacific. Especially noteworthy is the multidecadal Atlantic warming that began in the late 1990s, because recent research suggests that it has influenced Indo-Pacific climate, the character of the ENSO cycle, and the hiatus in global surface warming. Discovery of these pantropical interactions provides a pathway forward for improving predictions of climate variability in the current climate and for refining projections of future climate under different anthropogenic forcing scenarios.


FT-IR Hyperspectral Imaging and Artificial Neural Network Analysis for Identification of Pathogenic Bacteria.
Identification of microorganisms by Fourier transform infrared (FT-IR) spectroscopy is known as a promising alternative to conventional identification techniques in clinical, food, and environmental microbiology. In this study we demonstrate the application of FT-IR hyperspectral imaging for rapid, objective, and cost-effective diagnosis of pathogenic bacteria. The proposed method involves a relatively short cultivation step under standardized conditions, transfer of the microbial material onto suitable IR windows by a replica method, FT-IR hyperspectral imaging measurements, and image segmentation by machine learning classifiers, a hierarchy of specifically optimized artificial neural networks (ANN). For cultivation, aliquots of the initial microbial cell suspension were diluted to guarantee single-colony growth on solid agar plates. After a short incubation period when microbial microcolonies achieved diameters between 50 and 300 μm, microcolony imprints were produced by using a specifically developed stamping device which allowed spatially accurate transfer of the microcolonies' upper cell layers onto IR-transparent CaF2 windows. Dry microcolony imprints were subsequently characterized using a mid-IR microspectroscopic imaging system equipped with a focal plane array (FPA) detector. Spectral data analysis involved preprocessing, quality tests, and the application of supervised modular ANN classifiers for hyperspectral image segmentation. The resulting easily interpretable segmentation maps suggest a taxonomic resolution below the species level.


Vertical jump performance after 90 days bed rest with and without flywheel resistive exercise, including a 180 days follow-up.
Muscle atrophy and neuromuscular de-conditioning occur in response to space flight and bed-rest. In this study, we investigated the efficacy of flywheel training to conserve jumping power and height during 90 days bed rest. Twenty-four young healthy men underwent strict bed-rest (-6 degrees head down tilt) for 90 days. Eight participants were assigned to a flywheel group (FW) and 16 to a control group (Ctrl). The ground reaction force was measured during vertical jump tests twice during baseline data collection, and on day 4, 7, 14, 90 and 180 of recovery. In half of the participants, jump tests were also performed within minutes after re-ambulation and on four more occasions during the first 2 days of recovery. Jump height was reduced from 40.6 cm (SD 6.1 cm) during the first baseline measurement to 27.6 cm (SD 5.6 cm) on day 4 of recovery in Ctrl, but only from 38.6 cm (SD 3.9 cm) to 34.4 cm (SD 6.5 cm) in FW (P < 0.001). At the same time, peak power was reduced from 47.4 W/kg (SD 8.0 W/kg) to 34.5 W/kg in Ctrl, but only from 46.2 W/kg (6.0 W/kg) to 42.2 W/kg SD 4.6 W/kg) in FW (P < 0.001). Jump height and peak power were completely recovered after 163 and 140 days in Ctrl, respectively, and after 72 and 18 days in FW (regression analysis). In conclusion, flywheel exercise could effectively offset neuromuscular de-conditioning during bed-rest, and led to full recovery at an earlier stage. These findings nourish the hope that adequate training paradigms can fully sustain neuromuscular function under microgravity conditions.


Dimerization of soluble disulfide trap single-chain major histocompatibility complex class I molecules dependent on peptide binding affinity.
Stable presentation of peptide epitope by major histocompatibility complex (MHC) class I molecules is a prerequisite for the efficient expansion of CD8(+) T cells. The construction of single-chain MHC class I molecules in which the peptide, β(2)-microglobulin, and MHC heavy chain are all joined together via flexible linkers increases peptide-MHC stability. We have expressed two T cell epitopes that may be useful in leukemia treatment as single-chain MHC class I molecules, aiming to develop a system for the expansion of antigen-specific CD8(+) T cells in vitro. Disulfide trap versions of these single-chain MHC molecules were also created to improve anchoring of the peptides in the MHC molecule. Unexpectedly, we observed that soluble disulfide trap single-chain molecules expressed in eukaryotic cells were prone to homodimerization, depending on the binding affinity of the peptide epitope. The dimers were remarkably stable and efficiently recognized by conformation-specific antibodies, suggesting that they consisted of largely correctly folded molecules. However, dimerization was not observed when the disulfide trap molecules were expressed as full-length, transmembrane-anchored molecules. Our results further emphasize the importance of peptide binding affinity for the efficient folding of MHC class I molecules.


TGF-β-induced epigenetic deregulation of SOCS3 facilitates STAT3 signaling to promote fibrosis.
Fibroblasts are key effector cells in tissue remodeling. They remain persistently activated in fibrotic diseases, resulting in progressive deposition of extracellular matrix. Although fibroblast activation may be initiated by external factors, prolonged activation can induce an "autonomous," self-maintaining profibrotic phenotype in fibroblasts. Accumulating evidence suggests that epigenetic alterations play a central role in establishing this persistently activated pathologic phenotype of fibroblasts. We demonstrated that in fibrotic skin of patients with systemic sclerosis (SSc), a prototypical idiopathic fibrotic disease, TGF-β induced the expression of DNA methyltransferase 3A (DNMT3A) and DNMT1 in fibroblasts in a SMAD-dependent manner to silence the expression of suppressor of cytokine signaling 3 (SOCS3) by promoter hypermethylation. Downregulation of SOCS3 facilitated activation of STAT3 to promote fibroblast-to-myofibroblast transition, collagen release, and fibrosis in vitro and in vivo. Reestablishment of the epigenetic control of STAT3 signaling by genetic or pharmacological inactivation of DNMT3A reversed the activated phenotype of SSc fibroblasts in tissue culture, inhibited TGF-β-dependent fibroblast activation, and ameliorated experimental fibrosis in murine models. These findings identify a pathway of epigenetic imprinting of fibroblasts in fibrotic disease with translational implications for the development of targeted therapies in fibrotic diseases.


Down's syndrome suppression of tumour growth and the role of the calcineurin inhibitor DSCR1.
The incidence of many cancer types is significantly reduced in individuals with Down's syndrome, and it is thought that this broad cancer protection is conferred by the increased expression of one or more of the 231 supernumerary genes on the extra copy of chromosome 21. One such gene is Down's syndrome candidate region-1 (DSCR1, also known as RCAN1), which encodes a protein that suppresses vascular endothelial growth factor (VEGF)-mediated angiogenic signalling by the calcineurin pathway. Here we show that DSCR1 is increased in Down's syndrome tissues and in a mouse model of Down's syndrome. Furthermore, we show that the modest increase in expression afforded by a single extra transgenic copy of Dscr1 is sufficient to confer significant suppression of tumour growth in mice, and that such resistance is a consequence of a deficit in tumour angiogenesis arising from suppression of the calcineurin pathway. We also provide evidence that attenuation of calcineurin activity by DSCR1, together with another chromosome 21 gene Dyrk1a, may be sufficient to markedly diminish angiogenesis. These data provide a mechanism for the reduced cancer incidence in Down's syndrome and identify the calcineurin signalling pathway, and its regulators DSCR1 and DYRK1A, as potential therapeutic targets in cancers arising in all individuals.


Surveillance for Health Care Access and Health Services Use, Adults Aged 18-64 Years - Behavioral Risk Factor Surveillance System, United States, 2014.
As a result of the 2010 Patient Protection and Affordable Care Act, millions of U.S. adults attained health insurance coverage. However, millions of adults remain uninsured or underinsured. Compared with adults without barriers to health care, adults who lack health insurance coverage, have coverage gaps, or skip or delay care because of limited personal finances might face increased risk for poor physical and mental health and premature mortality. 2014. The Behavioral Risk Factor Surveillance System (BRFSS) is an ongoing, state-based, landline- and cellular-telephone survey of noninstitutionalized adults aged ≥18 years residing in the United States. Data are collected from states, the District of Columbia, and participating U.S. territories on health risk behaviors, chronic health conditions, health care access, and use of clinical preventive services (CPS). An optional Health Care Access module was included in the 2014 BRFSS. This report summarizes 2014 BRFSS data from all 50 states and the District of Columbia on health care access and use of selected CPS recommended by the U.S. Preventive Services Task Force or the Advisory Committee on Immunization Practices among working-aged adults (aged 18-64 years), by state, state Medicaid expansion status, expanded geographic region, and federal poverty level (FPL). This report also provides analysis of primary type of health insurance coverage at the time of interview, continuity of health insurance coverage during the preceding 12 months, and other health care access measures (i.e., unmet health care need because of cost, unmet prescription need because of cost, medical debt [medical bills being paid off over time], number of health care visits during the preceding year, and satisfaction with received health care) from 43 states that included questions from the optional BRFSS Health Care Access module. In 2014, health insurance coverage and other health care access measures varied substantially by state, state Medicaid expansion status, expanded geographic region (i.e., states categorized geographically into nine regions), and FPL category. The following proportions refer to the range of estimated prevalence for health insurance and other health care access measures by examined geographical unit (unless otherwise specified), as reported by respondents. Among adults with health insurance coverage, the range was 70.8%-94.5% for states, 78.8%-94.5% for Medicaid expansion states, 70.8%-89.1% for nonexpansion states, 73.3%-91.0% for expanded geographic regions, and 64.2%-95.8% for FPL categories. Among adults who had a usual source of health care, the range was 57.2%-86.6% for states, 57.2%-86.6% for Medicaid expansion states, 61.8%-83.9% for nonexpansion states, 64.4%-83.6% for expanded geographic regions, and 61.0%-81.6% for FPL categories. Among adults who received a routine checkup, the range was 52.1%-75.5% for states, 56.0%-75.5% for Medicaid expansion states, 52.1%-71.1% for nonexpansion states, 56.8%-70.2% for expanded geographic regions, and 59.9%-69.2% for FPL categories. Among adults who had unmet health care need because of cost, the range was 8.0%-23.1% for states, 8.0%-21.9% for Medicaid expansion states, 11.9%-23.1% for nonexpansion states, 11.6%-20.3% for expanded geographic regions, and 5.3%-32.9% for FPL categories. Estimated prevalence of cancer screenings, influenza vaccination, and having ever been tested for human immunodeficiency virus also varied by state, state Medicaid expansion status, expanded geographic region, and FPL category. The prevalence of insurance coverage varied by approximately 25 percentage points among racial/ethnic groups (range: 63.9% among Hispanics to 88.4% among non-Hispanic Asians) and by approximately 32 percentage points by FPL category (range: 64.2% among adults with household income <100% of FPL to 95.8% among adults with household income >400% of FPL). The prevalence of unmet health care need because of cost varied by nearly 14 percentage points among racial/ethnic groups (range: 11.3% among non-Hispanic Asians to 25.0% among Hispanics), by approximately 17 percentage points among adults with and without disabilities (30.8% versus 13.7%), and by approximately 28 percentage points by FPL category (range: 5.3% among adults with household income >400% of FPL to 32.9% among adults with household income <100% of FPL). Among the 43 states that included questions from the optional module, a majority of adults reported private health insurance coverage (63.4%), followed by public health plan coverage (19.4%) and no primary source of insurance (17.1%). Financial barriers to health care (unmet health care need because of cost, unmet prescribed medication need because of cost, and medical bills being paid off over time [medical debt]) were typically lower among adults in Medicaid expansion states than those in nonexpansion states regardless of source of insurance. Approximately 75.6% of adults reported being continuously insured during the preceding 12 months, 12.9% reported a gap in coverage, and 11.5% reported being uninsured during the preceding 12 months. The largest proportion of adults reported ≥3 visits to a health care professional during the preceding 12 months (47.3%), followed by 1-2 visits (37.1%), and no health care visits (15.6%). Adults in expansion and nonexpansion states reported similar levels of satisfaction with received health care by primary source of health insurance coverage and by continuity of health insurance coverage during the preceding 12 months. This report presents for the first time estimates of population-based health care access and use of CPS among adults aged 18-64 years. The findings in this report indicate substantial variations in health insurance coverage; other health care access measures; and use of CPS by state, state Medicaid expansion status, expanded geographic region, and FPL category. In 2014, health insurance coverage, having a usual source of care, having a routine checkup, and not experiencing unmet health care need because of cost were higher among adults living below the poverty level (i.e., household income <100% of FPL) in states that expanded Medicaid than in states that did not. Similarly, estimates of breast and cervical cancer screening and influenza vaccination were higher among adults living below the poverty level in states that expanded Medicaid than in states that did not. These disparities might be due to larger differences to begin with, decreased disparities in Medicaid expansion states versus nonexpansion states, or increased disparities in nonexpansion states. BRFSS data from 2014 can be used as a baseline by which to assess and monitor changes that might occur after 2014 resulting from programs and policies designed to increase access to health care, reduce health disparities, and improve the health of the adult population. Post-2014 changes in health care access, such as source of health insurance coverage, attainment and continuity of coverage, financial barriers, preventive care services, and health outcomes, can be monitored using these baseline estimates.


Resistance to paclitaxel therapy is related with Bcl-2 expression through an estrogen receptor mediated pathway in breast cancer.
Taxanes are approved for the treatment of breast cancer that has spread to the lymph nodes, following surgery and doxorubicin containing chemotherapy. Taxanes have improved the survival of breast cancer patients, especially in estrogen receptor (ER) negative population in clinical settings. This time we examined the relationship between chemosensitivity to Taxanes and expresson of ERalpha in breast cancer cell lines. In vitro effects of paclitaxel in 4 ER-positive and 3 ER-negative breast cancer cell lines were investigated by MTT assay. We also investigated members of Bcl-2 family by Western blotting and RT-PCR to clarify their role in paclitaxel resistance both in ER-positive and in ER-negative cells. ER-negative cell lines were more sensitive to paclitaxel than ER-positive cells. ER-negative KPL-4 and ZR-75-30 cells, which were sensitive to paclitaxel, became resistant when they were treated with demethylation agent, 5-aza-2'-deoxycytidine. Analysis of proapoptotic (Bax) and antiapoptotic (Bcl-2) molecules suggested that Bcl-2 is likely to have a role in the resistance of ER-positive cells. Bcl-2 expression was increased in a time-dependent manner after treatment of ER-positive cell lines with estrogen (E2). On the other hand, Bcl-2 was not detected in ER-negative cell lines. However, no significant difference was detected for Bax mRNA levels before and after E2 treatment in ER-positive and negative cell lines. Activation of ER gene expression in ER-negative KPL-4 cells by 5-aza-2'-deoxycytidine resulted in up-regulation of Bcl-2 mRNA. To support our data, we examined paclitaxel sensitivity in ER-negative MDA-MB-231 and ER stable transfectant cells S30 and JM6. This experiment also showed ER-negative cells were sensitive to paclitaxel but ER-positive cells were resistant to it. These results suggest that ER influenced chemosensitivity to paclitaxel through regulation of Bcl-2 family and regulation of the pathway may be crucial to increase the efficacy of taxanes in ER-positive breast cancer.


New guidelines for the management of thyroid nodules and differentiated thyroid cancer.
The last 5 years witnessed publication of several multidisciplinary guidelines that address management of benign and malignant thyroid diseases. The primary goal of this review was to excerpt highlights from the most recent 2009 guidelines of the American Thyroid Association that provided recommendations for treating thyroid nodules and differentiated thyroid cancer. There is a systematic algorithm for the evaluation of thyroid nodules based on clinical history, physical examination, measurement of thyroid stimulating hormone (TSH), and neck ultrasound, which has become the preferred radiologic modality for imaging thyroid disease. Radionuclide thyroid scanning is reserved for the evaluation of patients with suppressed TSH levels. Ultrasound-guided fine needle aspiration biopsy (FNAB) is indicated for cytologic assessment of most thyroid nodules >1 cm; additional criteria are specified for FNAB of nodules <1 cm. There is an expanded classification system for reporting thyroid cytologic finding, based on risk of malignancy. Treatment guidelines for thyroid cancer have rapidly evolved. Total thyroidectomy is advocated for initial surgical treatment of differentiated thyroid cancers >1 cm in size. Concurrent therapeutic central and lateral neck dissections remain essential to treat known cervical metastases, but prophylactic central neck dissection is controversial and should be cautiously chosen based on individual patient risk factors. Updates have been made for the use of radioactive iodine ablation and long-term surveillance for thyroid cancer recurrence, where ultrasound imaging is also important. A risk stratification paradigm is proposed to define the chance of recurrence and death from thyroid cancer, and modify treatment plans at various stages of management to the level of risk. Recommendations addressing both benign and malignant thyroid disease topics were graded according to the strength of available and published clinical evidence. Awareness of multidisciplinary recommendations for the treatment of benign and malignant thyroid diseases can enhance the practice of evidence-based medicine and provide practical tools for decision-making relevant to daily clinical encounters.


A Descriptive Study of Lower Limb Torsional Kinematic Profiles in Children With Spastic Diplegia.
Lower limb rotational anomalies in spastic diplegic children with cerebral palsy (CP) are common and difficult to identify through physical examination alone. The identification and treatment of the overall rotational disorders must be considered to restore physiological lever-arms lengths and lever-arms orientation.The aims of the study were to assess the prevalence of lower limb rotational malalignment and to describe the distribution of the different kinematic torsional profiles in children with spastic diplegia. Instrumented gait analysis data from 188 children with spastic diplegia were retrospectively reviewed. None of the patients had undergone surgery previously or received botulinum toxin treatment within 6 months before the review. Kinematic data, collected at the midstance phase, included: pelvic, hip, and ankle rotation and foot progression angle. The prevalence of kinematic rotational deviations was 98.4%. Sixty-one percent of the children walked with an internal foot progression angle and 21% exhibited external alignment. The pelvis was internally rotated in 41% of the cases and externally in another 27%. Hip rotation was internal in 29% and external in 27% of the cases. Ankle rotation was internal in 55% and external in 16% of the cases. Lower limb rotational anomalies involved more than one level in 77% of the limbs. A kinematic compensatory deviation was identified in at least one level in 48% of the limbs. Kinematic rotational anomalies were identified in nearly all the 188 children in the study. The multilevel involvement of lower limb malalignment was not systematically associated with compensatory mechanisms between the levels. Ankle rotational anomalies were the most frequent cause of lower limb torsional deviations followed by pelvic malalignment. Level IV.


CTLA4 blockade in mesothelioma: finally a competing strategy over cytotoxic/target therapy?
No second-line treatment significantly prolongs the survival of malignant mesothelioma patients who have a high unmet medical need. Here, we comment on the therapeutic potential of cytotoxic T-lymphocyte-associated protein (CTLA)4-blockade by the anti-CTLA4 monoclonal antibody (mAb) tremelimumab of refractory malignant mesothelioma patients. We also focus on the critical role of an accurate tumor assessment in the course of treatment with immunomodulating mAb. Finally, treatment with potentially effective, second-generation checkpoint(s) inhibiting mAb, as well future combination strategies in this deadly disease, will be discussed.


Paracellular nutrient absorption is higher in bats than rodents: integrating from intact animals to the molecular level.
Flying vertebrates have been hypothesized to rely heavily on paracellular absorption of nutrients to compensate for having smaller intestines than non-flyers. We tested this hypothesis in an insectivorous bat (Myotis lucifugus) and two insect-eating rodents (Onychomys leucogaster and Peromyscus leucopus). In intact animals, the fractional absorption of orally dosed l-arabinose (Mr 150) was 82% in M. lucifugus, which was more than twice that of the rodents. Absorption of creatinine (Mr 113) was greater than 50% for all species and did not differ between M. lucifugus and the rodents. We also conducted intestinal luminal perfusions on anesthetized animals. Absorption of l-arabinose per nominal surface area in M. lucifugus was nearly double that of the rodents, while absorption of creatinine was not different among species. Using an everted sleeve preparation, we demonstrated that high concentrations of l-arabinose and creatinine did not inhibit their own uptake, validating their use as passive, paracellular probes. Histological measurements indicated that M. lucifugus has more cells, and presumably more tight junctions, per nominal surface area than P. leucopus. This seems unlikely to explain entirely the higher absorption of l-arabinose in M. lucifugus during perfusions, because l-arabinose absorption normalized to the number of enterocytes was still double that of P. leucopus. As an alternative, we investigated tight junction gene expression. M. lucifugus had higher expression of claudin-1 and claudin-15, and lower expression of claudin-2 relative to P. leucopus. Expression of claudin-7 and occludin did not differ among species. Taken together, our results support the hypothesis that bats have evolved higher paracellular nutrient absorption than non-flying animals, and that this phenomenon might be driven by both histological characteristics and differences in tight junction gene expression.


The lifetime history of major depression in women. Reliability of diagnosis and heritability.
In epidemiologic samples, the assessment of lifetime history (LTH) of major depression (MD) is not highly reliable. In female twins, we previously found that LTH of MD, as assessed at a single personal interview, was moderately heritable (approximately 40%). In that analysis, errors of measurement could not be discriminated from true environmental effects. In 1721 female twins from a population-based register, including both members of 742 pairs, LTH of MD, covering approximately the same time period, was obtained twice, once by self-administered questionnaire and once at personal interview. Reliability of LTH of MD was modest (kappa = +.34, tetrachoric r = +.56) and was predicted by the number of depressive symptoms, treatment seeking, number of episodes, and degree of impairment. Deriving an "index of caseness" from these predictors, the estimated heritability of LTH of MD was greater for more restrictive definitions. Incorporating error of measurement into a structural equation model including both occasions of measurement, the estimated heritability of the liability to LTH of MD increased substantially (approximately 70%). More than half of what was considered environmental effects when LTH of MD was analyzed on the basis of one assessment appeared, when two assessments were used, to reflect measurement error. Major depression, as assessed over the lifetime, may be a rather highly heritable disorder of moderate reliability rather than a moderately heritable disorder of high reliability.


Classification of biliary tract cancers established by the Japanese Society of Hepato-Biliary-Pancreatic Surgery: 3(rd) English edition.
The 3(rd) English edition of the Japanese classification of biliary tract cancers was released approximately 10 years after the 5(th) Japanese edition and the 2(nd) English edition. Since the first Japanese edition was published in 1981, the Japanese classification has been in extensive use, particularly among Japanese surgeons and pathologists, because the cancer status and clinical outcomes in surgically resected cases have been the main objects of interest. However, recent advances in the diagnosis, management and research of the disease prompted the revision of the classification that can be used by not only surgeons and pathologists but also by all clinicians and researchers, for the evaluation of current disease status, the determination of current appropriate treatment, and the future development of medical practice for biliary tract cancers. Furthermore, during the past 10 years, globalization has advanced rapidly, and therefore, internationalization of the classification was an important issue to revise the Japanese original staging system, which would facilitate to compare the disease information among institutions worldwide. In order to achieve these objectives, the new Japanese classification of the biliary tract cancers principally adopted the 7(th) edition of staging system developed by the International Union Against Cancer (UICC) and the American Joint Committee on Cancer (AJCC). However, because there are some points pending in these systems, several distinctive points were also included for the purpose of collection of information for the future optimization of the staging system. Free mobile application of the new Japanese classification of the biliary tract cancers is available via http://www.jshbps.jp/en/classification/cbt15.html.


Suicide Risk Assessment in Hospitals: An Expert System-Based Triage Tool.
The November 2010 Joint Commission Sentinel Event Alert on the prevention of suicides in medical/surgical units and the emergency department (ED) mandates screening every patient treated as an outpatient or admitted to the hospital for suicide risk. Our aim was to develop a suicide risk assessment tool to (1) predict the expert psychiatrist's assessment for risk of committing suicide within 72 hours in the hospital, (2) replicate the recommended intervention by the psychiatrist, and (3) demonstrate acceptable levels of participant satisfaction. The 3 phases of tool development took place between October 2012 and February 2014. An expert panel developed key questions for a tablet-based suicide risk questionnaire. We then performed a randomized cross-sectional study comparing the questionnaire to the interview by a psychiatrist, for model derivation. A neural network model was constructed using 255 ED participants. Evaluation was the agreement between the risk/intervention scores using the questionnaire and the risk/intervention scores given by psychiatrists to the same patients. The model was validated using a new population of 124 participants from the ED and 50 participants from medical/surgical units. The suicide risk assessment tool performed at a remarkably high level. For levels of suicide risk (minimal or low, moderate, or high), areas under the curves were all above 0.938. For levels of intervention (routine, specialized, highly specialized, or secure), areas under the curves were all above 0.914. Participants reported that they liked the tool, and it took less than a minute to use. An expert-based neural network model predicted psychiatrists' assessments of risk of suicide in the hospital within 72 hours. It replicated psychiatrist-recommended interventions to mitigate risk in EDs and medical/surgical units.


Serum melatonin circadian profiles in women suffering from cervical cancer.
Although there is an increasing evidence that the pineal gland may play a role in human malignancy, the studies on melatonin concentrations in different types of malignant tumors brought about controversial results. However, changes in melatonin concentrations have been observed in some types of human malignant tumors. Therefore, we decided to study the circadian melatonin rhythm in patients suffering from cervical cancer in different stages of progression and to compare them with those in subjects free from neoplastic disease. A total of 45 women were analyzed in this study. The subjects were divided into two groups. The first group consisted of 31 patients [mean age 52.1 +/- 1.8 yr (mean +/- S.E.M.), range 32-77 yr] with cervical cancer in various stages of the disease. The second group consisted of 14 healthy volunteers [mean age 53.5 +/- 2.0 yr (mean +/- S.E.M.), range 42-63] who served as the control group. Blood samples were collected at 08:00, 12:00, 16:00, 20:00, 22:00, 24:00, 02:00, 04:00, 06:00, and 08:00 hours. Melatonin concentration was measured by immunoenzymatic method. There were significant differences in circadian melatonin profiles as well as in the area under curve among the two studied groups. Melatonin concentrations were significantly lower in cancer patients in comparison with healthy individuals. Taking into consideration stage of the cervical cancer significantly lower melatonin secretion has been found in all subgroups of patients in comparison with that of tumor-free control group. Additionally, nocturnal melatonin concentrations as well as area under curve were significantly lower in advanced stage of cancer (stages 3 and 4) in comparison with patients with preinvasive cancer (stage 0) at 24:00, 02:00, and 04:00 hours and patients with stage 1 disease at 02:00 and 04:00 hours. The results of the present study indicate that the presence of cervical cancer influences melatonin levels in women. Moreover, stage dependence in reduction of melatonin concentrations has been found.


Shape-based cortical surface segmentation for visualization brain mapping.
We describe a knowledge-based approach to cortical surface segmentation that uses learned knowledge of the overall shape and range of variation of the cortex (excluding the detailed gyri and sulci) to guide the search for the grey-CSF boundary in a structural MRI image volume. The shape knowledge is represented by a radial surface model, which is a type of geometric constraint network (GCN) that we hypothesize can represent shape by networks of locally interacting constraints. The shape model is used in a protocol for visualization-based mapping of cortical stimulation mapping (CSM) sites onto the brain surface, prior to integration with other mapping modalities or as input to existing surface analysis and reconfiguration programs. Example results are presented for CSM data related to language organization in the cortex, but the methods should be applicable to other situations where a realistic visualization of the brain surface, as seen at neurosurgery, is desired.


Cognitive and behavioral predictors of sexually transmitted disease risk behavior among sexually active adolescents.
To identify important cognitive and behavioral predictors of sexually transmitted disease (STD) risk behavior among a sexually active adolescent cohort. One-year longitudinal study of health beliefs, sexual behaviors, and STD acquisition among 549 adolescents, 14 to 21 years of age. School- and community-based health clinics in a large metropolitan area. Data from 410 sexually active adolescents completing surveys at baseline and 1-year follow-up. None. Sexually transmitted disease risk behavior--a composite measure of condom use consistency with most recent sexual partner(s), number of vaginal sex partners, and frequency of intercourse with most recent sexual partner(s). For girls (n = 335), a model including baseline STD risk behavior, condom use self-efficacy, oral contraceptive use, communication with sexual partners about STD prevention, and alcohol use in connection with sexual activity explained 21.1% of the variance in STD risk behavior at 1-year follow-up. For boys (n = 75), the strongest predictors of STD risk behavior at follow-up included baseline STD risk behavior, perceived susceptibility to STD, condom use self-efficacy, negative outcome expectations associated with condom use, and perceived barriers to STD prevention. Efforts targeting reduction in STD risk behavior must begin before the onset of somewhat stable patterns of sexual risk behavior. Among adolescents who are sexually active, interventions should include components that increase condom use self-efficacy, build skills to communicate with sexual partners about STD prevention, and address behaviors associated with STD risk behavior including oral contraceptive use.


Apoprotein B of lipoprotein(a) of human plasma.
Lipoprotein(a) was purified by agarose-gel chromatography from human plasma from which lipoproteins of Sf greater than 0 had been removed either by sequential or by density-gradient ultracentrifugation. After delipidation, the apoprotein B of this lipoprotein was analysed by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. It could not be distinguished from the apoprotein B of low-density lipoproteins (rho 1.019-1.063 g/ml). A significant increase in the concentration of apoprotein B in plasma from which the Sf greater than 0 lipoproteins had been removed was observed in six subjects 4 h after a fatty meal.


Patients with polymyositis show changes in muscle protein charges.
Polymyositis (PM) appears with indolent proximal muscle weakness and is an inflammatory disease with breakdown of muscle cells. In our study the protein charge concentrations of the contractile proteins in the A and I bands were determined, applying a microelectrode technique. Patients with PM show a lower protein charge concentration than healthy control subjects which may be caused by the breakdown and removal of the proteins in the contractile filaments. A tool to judge the state of the disease as well as an aid in diagnosis may have been found in this method.


Loss of heterozygosity in sporadic primary cutaneous melanoma.
Difficulties in obtaining clinical samples from primary melanomas have meant that most genetic analyses of melanoma have concentrated on cell lines and metastases. Because the Breslow thickness of the primary tumour is the single best prognostic indicator, it is important to identify genetic abnormalities in primary melanomas and relate these changes to the thickness of the lesion. We have investigated 47 sporadic melanomas, of which 41 were primary lesions, for loss of heterozygosity (LOH) on several chromosomal arms, including areas where genes involved in familial melanoma and other relevant hereditary syndromes map, and where LOH has previously been reported in cell lines, or metastatic lesions. LOH was identified at 66 (18%) of 358 informative loci in primary melanomas, and there was a significant relationship between the overall frequency of LOH and Breslow thickness (P < 0.0005). Loss of chromosome arm 9p was most frequent, occurring in 15 (47%) of 32 informative primary tumours, and was observed in 3 of 11 informative lesions < or = 1.5 mm in depth. LOH on chromosome arms 3p, 6q, 10q, 11q, and 17p was also relatively frequent, with loss of 3p and 10q heterozygosity in lesions < or = 1.5 mm in depth, while LOH on 6q, 11q, and 17p was only detected in more invasive tumours. The results suggest that loss of these chromosome regions are important in sporadic cutaneous melanoma, and are consistent with chromosome arm 9p loss occurring before loss of other chromosome arms.


Control of the light-response in supramolecular metallopolymeric gels by tuning the coordination metal.
Two novel supramolecular metallo-heteropolymers bearing a photo-isomerizable telechelic bis-terpyridine ligand and either Fe(ii) or Co(ii) coordination metal were synthesized. Both polymers induced gelation of organic solvents at a concentration as low as 0.12 wt% yielding thixotropic gels. Judicious choice of the electronic and photophysical properties of both ditopic ligand and metal ion enabled to achieve control over photomechanical response in supramolecular organogels upon UV light irradiation through molecular design.


Finite-size scaling in silver nanowire films: design considerations for practical devices.
We report the first application of finite-size scaling theory to nanostructured percolating networks, using silver nanowire (AgNW) films as a model system for experiment and simulation. AgNWs have been shown to be a prime candidate for replacing Indium Tin Oxide (ITO) in applications such as capacitive touch sensing. While their performance as large area films is well-studied, the production of working devices involves patterning of the films to produce isolated electrode structures, which exhibit finite-size scaling when these features are sufficiently small. We demonstrate a generalised method for understanding this behaviour in practical rod percolation systems, such as AgNW films, and study the effect of systematic variation of the length distribution of the percolating material. We derive a design rule for the minimum viable feature size in a device pattern, relating it to parameters which can be derived from a transmittance-sheet resistance data series for the material in question. This understanding has direct implications for the industrial adoption of silver nanowire electrodes in applications where small features are required including single-layer capacitive touch sensors, LCD and OLED display panels.


A selective inhibitor of dipeptidyl peptidase I impairs generation of CD8+ T cell cytotoxic effector function.
CTL express high levels of dipeptidyl peptidase I (DPPI), a granule thiol protease able to convert the zymogen precursors of granzymes A and B into active proteases. In the present studies, the effects of specific inhibition of DPPI on generation of CTL effector functions were examined. When T cell DPPI activity was inhibited by >95% throughout 5-day MLC, a significant reduction in the generation of CD8+ T cell BLT esterase activity (<30% of control) and cytolytic activity (<10% of control) was observed. DPPI inhibition during the second to fourth days of 5-day MLC also was associated with reduced proliferation of CD8+ T cells, but had no effect on CD4+ T cell proliferation or IL-2 production by either population. CTL generated in the continuous presence of DPPI inhibition also exhibited impaired lysis of anuclear erythrocyte targets and diminished killing of nucleated targets by perforin-independent pathways. In contrast, inhibition of DPPI during only the last 24 h of 5-day MLC was associated only with reduced generation of BLT esterase activity and reduced lysis of nucleated targets by perforin-dependent pathways. Repeated or delayed inhibition of DPPI in MLC containing granzyme B-deficient responder cells also impaired generation of cytotoxic activity. These results indicate that DPPI or other DPPI-like protease activities not only are required for the activation of granzymes, but also play a role in the expansion and differentiation of full CD8+ T cell cytolytic activity.


Human papillomavirus vaccination: recommendations of the Advisory Committee on Immunization Practices (ACIP).
This report summarizes the epidemiology of human papillomavirus (HPV) and associated diseases, describes the licensed HPV vaccines, provides updated data from clinical trials and postlicensure safety studies, and compiles recommendations from CDC's Advisory Committee on Immunization Practices (ACIP) for use of HPV vaccines. Persistent infection with oncogenic HPV types can cause cervical cancer in women as well as other anogenital and oropharyngeal cancers in women and men. HPV also causes genital warts. Two HPV vaccines are licensed in the United States. Both are composed of type-specific HPV L1 protein, the major capsid protein of HPV. Expression of the L1 protein using recombinant DNA technology produces noninfectious virus-like particles (VLPs). Quadrivalent HPV vaccine (HPV4) contains four HPV type-specific VLPs prepared from the L1 proteins of HPV 6, 11, 16, and 18. Bivalent HPV vaccine (HPV2) contains two HPV type-specific VLPs prepared from the L1 proteins of HPV 16 and 18. Both vaccines are administered in a 3-dose series. ACIP recommends routine vaccination with HPV4 or HPV2 for females aged 11 or 12 years and with HPV4 for males aged 11 or 12 years. Vaccination also is recommended for females aged 13 through 26 years and for males aged 13 through 21 years who were not vaccinated previously. Males aged 22 through 26 years may be vaccinated. ACIP recommends vaccination of men who have sex with men and immunocompromised persons (including those with HIV infection) through age 26 years if not previously vaccinated. As a compendium of all current recommendations for use of HPV vaccines, information in this report is intended for use by clinicians, vaccination providers, public health officials, and immunization program personnel as a resource. ACIP recommendations are reviewed periodically and are revised as indicated when new information and data become available.


What do internal medicine residents need to enhance their diabetes care?
To identify areas that should be targeted for improvement in care, we examined internal medicine resident practice patterns and beliefs regarding diabetes in a large urban hospital outpatient clinic. Internal medicine residents were surveyed to assess the frequency at which they performed key diabetes quality of care indicators. Responses were compared with recorded performance derived from chart and laboratory database reviews. Resident attitudes about diabetes were determined using the Diabetes Attitude Survey for Practitioners. Finally, an eight-item scale was used to assess barriers to diabetes care. Both self-described and recorded performance of recommended diabetes services short of national recommendations. For yearly eye examinations and lipid screening, recorded performance levels were similar to trainees' reports. However, documented inquiries about patient self-monitoring of blood glucose, performance of foot examinations, and urine protein screening were lower than trainees' reports. Some 49% of the residents selected a target HbA1c of 6.6-7.5% as an attainable goal, yet half of the patients using oral agents or insulin had HbA1c values > 8.0%. No differences in self-described or recorded performance were found by year of training. Most residents did not perceive themselves to need additional training related to diabetes care, and residents were generally neutral about patient autonomy. Patient nonadherence and time constraints within the clinic were most often cited as barriers to care. The study identifies several areas that require improvement in resident care of diabetes in the ambulatory setting. Because experience during training contributes to future practice patterns, developing a program that teaches trainees how to implement diabetes practice guidelines and methods to achieve optimal glycemic control may be key to future improvements in the quality of diabetes care.


A hump-shaped relationship between isolation and abundance of Notonecta irrorata colonists in aquatic mesocosms.
We examined the relationship between the isolation of experimental aquatic mesocosms and the abundance of an aquatic insect colonist, Notonecta irrorata, over two years. We used a curve-fitting approach to assess whether linear or quadratic models better describe the relationship between isolation and abundance. For two measures of mesocosm isolation, distance to nearest source and distance to the largest source population, there was a significant quadratic relationship between isolation and abundance. Abundance of colonizing N. irrorata was not found to be significantly related to a third measure of isolation, mesocosm connectivity. These results indicate that the relationship between habitat isolation and colonist abundance may not be a monotonic decline across all spatial scales, a finding that contradicts the usual assumption incorporated in measures of habitat connectivity. Our results suggest that under some circumstances individuals that have undertaken dispersal may bypass patches they encounter early in this process and preferentially settle in patches encountered later. This behavioral preference in conjunction with decreased numbers of potential colonists at sites far from the source environment could lead to the "hump-shaped" colonist abundance by habitat isolation relationship we observed in this study. We suggest that simple assumptions about the relationship between habitat isolation and the probability a site is colonized need to be reexamined and alternative possible forms of this relationship tested.


Boosting accuracy of automated classification of fluorescence microscope images for location proteomics.
Detailed knowledge of the subcellular location of each expressed protein is critical to a full understanding of its function. Fluorescence microscopy, in combination with methods for fluorescent tagging, is the most suitable current method for proteome-wide determination of subcellular location. Previous work has shown that neural network classifiers can distinguish all major protein subcellular location patterns in both 2D and 3D fluorescence microscope images. Building on these results, we evaluate here new classifiers and features to improve the recognition of protein subcellular location patterns in both 2D and 3D fluorescence microscope images. We report here a thorough comparison of the performance on this problem of eight different state-of-the-art classification methods, including neural networks, support vector machines with linear, polynomial, radial basis, and exponential radial basis kernel functions, and ensemble methods such as AdaBoost, Bagging, and Mixtures-of-Experts. Ten-fold cross validation was used to evaluate each classifier with various parameters on different Subcellular Location Feature sets representing both 2D and 3D fluorescence microscope images, including new feature sets incorporating features derived from Gabor and Daubechies wavelet transforms. After optimal parameters were chosen for each of the eight classifiers, optimal majority-voting ensemble classifiers were formed for each feature set. Comparison of results for each image for all eight classifiers permits estimation of the lower bound classification error rate for each subcellular pattern, which we interpret to reflect the fraction of cells whose patterns are distorted by mitosis, cell death or acquisition errors. Overall, we obtained statistically significant improvements in classification accuracy over the best previously published results, with the overall error rate being reduced by one-third to one-half and with the average accuracy for single 2D images being higher than 90% for the first time. In particular, the classification accuracy for the easily confused endomembrane compartments (endoplasmic reticulum, Golgi, endosomes, lysosomes) was improved by 5-15%. We achieved further improvements when classification was conducted on image sets rather than on individual cell images. The availability of accurate, fast, automated classification systems for protein location patterns in conjunction with high throughput fluorescence microscope imaging techniques enables a new subfield of proteomics, location proteomics. The accuracy and sensitivity of this approach represents an important alternative to low-resolution assignments by curation or sequence-based prediction.


Synthetic Glycolipids as Molecular Vaccine Adjuvants: Mechanism of Action in Human Cells and In Vivo Activity.
Modern adjuvants for vaccine formulations are immunostimulating agents whose action is based on the activation of pattern recognition receptors (PRRs) by well-defined ligands to boost innate and adaptive immune responses. Monophosphoryl lipid A (MPLA), a detoxified analogue of lipid A, is a clinically approved adjuvant that stimulates toll-like receptor 4 (TLR4). The synthesis of MPLA poses manufacturing and quality assessment challenges. Bridging this gap, we report here the development and preclinical testing of chemically simplified TLR4 agonists that could sustainably be produced in high purity and on a large scale. Underpinned by computational and biological experiments, we show that synthetic monosaccharide-based molecules (FP compounds) bind to the TLR4/MD-2 dimer with submicromolar affinities stabilizing the active receptor conformation. This results in the activation of MyD88- and TRIF-dependent TLR4 signaling and the NLRP3 inflammasome. FP compounds lack in vivo toxicity and exhibit adjuvant activity by stimulating antibody responses with a potency comparable to MPLA.


Fe(III) reduction during pyruvate fermentation by Desulfotomaculum reducens strain MI-1.
Desulfotomaculum reducens MI-1 is a Gram-positive, sulfate-reducing bacterium also capable of reducing several metals, among which is Fe(III). Very limited knowledge is available on the potential mechanism(s) of metal reduction among Gram-positive bacteria, despite their preponderance in the microbial communities that inhabit some inhospitable environments (e.g., thermal or hyperthermal ecosystems, extreme pH or salinity environments, heavy metal or radionuclide contaminated sediments). Here, we show that in the presence of pyruvate, this micro-organism is capable of reducing both soluble Fe(III)-citrate and solid-phase hydrous ferric oxide, although growth is sustained by pyruvate fermentation rather than Fe(III) respiration. Despite the fact that Fe(III) reduction does not support direct energy conservation, D. reducens uses it as a complementary means of discarding excess reducing equivalent after H2 accumulation in the culture headspace renders proton reduction unfavorable. Thus, Fe(III) reduction permits the oxidation of greater amounts of pyruvate than fermentation alone. Fe(III) reduction by D. reducens is mediated by a soluble electron carrier, most likely riboflavin. Additionally, an intracellular electron storage molecule acts as a capacitor and accumulates electrons during pyruvate oxidation for slow release to Fe(III). The reductase responsible for the transfer of electrons from the capacitor to the soluble carrier has not been identified, but data presented here argue against the involvement of c-type cytochromes.


Immobilized rhBMP-2/succinylated type I atelocollagen gene expression of intracellular signaling molecules on ST2 cells.
Recombinant human bone morphogenetic protein-2 (rhBMP-2) chemically-bonded to succinylated type I atelocollagen, a biomaterial carrier with a porous structure, was reported to augment cellular activity of ST2 cells. The Smad protein family has been suggested to play an important role in the intracellular signaling pathway of BMP by its binding to receptors on target cells. However, there has been no study analyzing the downstream genes of the rhBMP-2 induced intracellular signal transduction pathway. The purpose of this study was to examine the effect of immobilized rhBMP-2 on gene expression of intracellular signaling molecules on ST2 cells. Our study showed two expression patterns of downstream genes of rhBMP-2 intracellular signal transduction pathway. In the first pattern, BMPR-IA, Smad 1, and Smad 5 genes showed high basic expression before the addition of rhBMP, and the high level of gene expression continued for long period and decreased in the late stage when rhBMP-2 was immobilized. In the second pattern, Smad 6, Smad 7, and Smad 8 genes showed low basic expression before the addition of rhBMP-2 and a continuous increase from the beginning was followed by a decrease in the late stage when rhBMP-2 was immobilized. Our results also showed that intracellular signaling continued for prolonged period when rhBMP-2 was immobilized to succinylated type I atelocollagen. This study indicated that immobilizing rhBMP-2 is an efficient method to increase bone induction.