Cultivation of globally distributed soil bacteria from phylogenetic lineages previously only detected in cultivation-independent surveys.
The culturability of microorganisms in a 10 cm core of an Australian pasture soil was investigated using a minimal agar medium with xylan as the growth substrate. Culturability decreased with increasing depth, from a maximum of 19% of the total microscopically countable cells in the 0-2 cm section to 2.4% in the 8-10 cm section. Seventy-one isolates from the core were identified by comparative 16S rRNA gene sequence analysis. Many of these isolates belong to groups of globally distributed soil bacteria, including well-characterized families of the classes Alphaproteobacteria and Betaproteobacteria, and of the subclass Actinobacteridae. Other isolates belong to groups with few or no cultivated representatives: 10 isolates in two subdivisions of the phylum Acidobacteria, five isolates in a new order and nine isolates in a new family of the class Alphaproteobacteria, two isolates in a new order of the class Gammaproteobacteria, three isolates in two new families of the subclass Actinobacteridae, and two isolates in the subclass Rubrobacteridae. These new isolates represent the first laboratory cultures able to be assigned to some of these groups and greatly increase the number of cultivated strains known for others. This demonstrates that a minimal change in cultivation strategy (using a polymeric growth substrate and longer incubation times) can result in the isolation of globally distributed but previously uncultured phylogenetically novel soil bacteria.


Transfection and expression of a gene coding for a human melanoma antigen recognized by autologous cytolytic T lymphocytes.
Human melanoma line MZ2-MEL expresses several antigens recognized by autologous cytolytic T lymphocytes (CTL). As a first step towards the cloning of the gene coding for one of these antigens, we tried to obtain transfectants expressing the antigen. The DNA recipient cell was a variant of MZ2-MEL which had been selected with a CTL clone for the loss of antigen E. It was cotransfected with genomic DNA of the original melanoma line and with selective plasmid pSVtkneo beta. Geneticin-resistant transfectants were obtained at a frequency of 2 x 10(-4). These transfectants were then screened for their ability to stimulate the production of tumor necrosis factor by the anti-E CTL clone. One transfectant expressing antigen E was identified among 70,000 drug-resistant transfectants. Its sensitivity to lysis by the anti-E CTL was equal to that of the original melanoma cell line. When this transfectant was submitted to immunoselection with the anti-E CTL clone, the resulting antigen-loss variants were found to have lost several of the transfected pSVtkneo beta sequences. This indicated that the gene coding for the antigen had been integrated in the vicinity of pSVtkneo beta sequences, as expected for cotransfected DNA.


Lysozyme distribution and conformation in a biodegradable polymer matrix as determined by FTIR techniques.
Lysozyme distribution and conformation in poly(lactic-co-glycolic acid)(PLGA) microspheres was determined using various infrared spectroscopic techniques. Infrared microscopy and confocal laser scanning microscopy indicated that the protein was homogeneously distributed inside the microspheres in small cavities resulting from the water-in-oil emulsification step. Part of the protein was observed at or near the cavity walls, while the rest was located within these cavities. Attenuated total reflectance (ATR) and photoacoustic spectroscopy (PAS) also showed that there is hardly any protein at the surface of the microspheres. Since this microsphere formulation gave a large burst release (ca. 50%), this burst release can not be caused by protein at the surface of the particles. Probably, the protein is rapidly released through pores in the PLGA matrix. Conformational analysis of lysozyme in the PLGA microspheres by KBr pellet transmission suffered from band shape distortion and baseline slope. Despite incomplete subtraction of the PLGA background, a characteristic band of non-covalent aggregates at 1625 cm(-1) was observed in the second derivative spectrum of the protein Amide I region. The other Fourier-transform infrared (FTIR) methods yielded similar results, indicating that the sample preparation procedure did not introduce artifacts. The observed aggregation signal may correspond to the protein adsorbed to the cavity walls inside the microspheres.


A Diagnosis of Biophysical and Socio-Economic Factors Influencing Farmers' Choice to Adopt Organic or Conventional Farming Systems for Cotton Production.
Organic agriculture is one of the most widely known alternative production systems advocated for its benefits to soil, environment, health and economic well-being of farming communities. Rapid increase in the market demand for organic products presents a remarkable opportunity for expansion of organic agriculture. A thorough understanding of the context specific motivations of farmers for adoption of organic farming systems is important so that appropriate policy measures are put in place. With an aim of understanding the social and biophysical motivations of organic and conventional cotton farmers for following their respective farming practices, a detailed farm survey was conducted in Nimar valley of Madhya Pradesh state in central India. The study area was chosen for being an important region for cotton production, where established organic and conventional farms operate under comparable circumstances. We found considerable variation among organic and conventional farmers for their social and biophysical motivations. Organic farmers were motivated by the sustainability of cotton production and growing safer food without pesticides, whereas conventional farmers were sensitive about their reputation in community. Organic farmers with larger holdings were more concerned about closed nutrient cycles and reducing their dependence on external inputs, whereas medium and small holding organic farmers were clearly motivated by the premium price of organic cotton. Higher productivity was the only important motivation for conventional farmers with larger land holdings. We also found considerable yield gaps among different farms, both under conventional and organic management, that need to be addressed through extension and training. Our findings suggest that research and policy measures need to be directed toward strengthening of extension services, local capacity building, enhancing availability of suitable inputs and market access for organic farmers.


Time-dependent expression and activity of cytochrome P450 1s in early life-stages of the zebrafish (Danio rerio).
Zebrafish embryos are being increasingly used as model organisms for the assessment of single substances and complex environmental samples for regulatory purposes. Thus, it is essential to fully understand the xenobiotic metabolism during the different life-stages of early development. The aim of the present study was to determine arylhydrocarbon receptor (AhR)-mediated activity during selected times of early development using qPCR, enzymatic activity through measurement of 7-ethoxyresorufin-O-deethylase (EROD) activity, and protein expression analysis. In the present study, gene expression of cyp1a, cyp1b1, cyp1c1, cyp1c2, and ahr2 as well as EROD activity were investigated up to 120 h postfertilization (hpf) after exposure to either β-naphthoflavone (BNF) or a polycyclic aromatic hydrocarbons (PAH)-contaminated sediment extract from Vering Kanal in Hamburg (VK). Protein expression was measured at 72 hpf after exposure to 20 μg/L BNF. Altered proteins were identified by matrix assisted laser desorption ionization time-of-flight (MALDI-TOF) peptide mass fingerprinting. Distinct patterns of basal messenger RNA (mRNA) expression were found for each of the cyp1 genes, suggesting specific roles during embryonic development. All transcripts were induced by BNF and VK. ahr2 mRNA expression was significantly upregulated after exposure to VK. All cyp1 genes investigated showed a temporal decline in expression at 72 hpf. The significant decline of Hsp 90β protein at 72 hpf after exposure to BNF may suggest an explanation for the decline of cyp1 genes at this time point as Hsp 90β is of major importance for the functioning of the Ah-receptor. EROD activity measured in embryos was significantly induced after 96 hpf of exposure to BNF or VK. Together, these results demonstrate distinct temporal patterns of cyp1 genes and protein activities in zebrafish embryos as well as show a need to investigate further the xenobiotic biotransformation system during early development of zebrafish.


Psychiatric Disorders in Women Seeking Fertility Treatments: A Clinical Investigation in India.
Fertility treatments began in several countries, including India, in the1970s. Despite various advancements in intra uterine insemination (IUI) and in vitro fertilization (IVF), empirical investigations on the psychological endurance and emotional tolerance of Indian women to such treatments are rather scarce. Thus, the aim of this study is to estimate the prevalence of psychiatric disorders in Indian women seeking fertility treatments. It is a cross-sectional study with three hundred women participants undergoing various treatments at the Manipal Assisted Reproductive Centre, Kasturba Medical College, Karnataka, India. Psychiatric disorders were assessed in women using the "ICD- 10 Classification of Mental and Behavioural Disorders" followed by descriptive data analysis. The results show that 78% of women have psychological issues and 45% of them have a diagnosable psychiatric condition. Adjustment Disorders, Anxiety Disorders and Mixed Anxiety and Depression Disorder are established as the top three categories of diagnoses. The findings of this study suggest that women have a high emotional stake in infertility treatments. The data highlights the need for modification of the existing treatment protocol (in Indian clinics) in ways that ensure the emotional wellbeing of patients.


Ongoing ingestive behavior is rapidly suppressed by a preabsorptive, intestinal "bitter taste" cue.
The discovery that cells in the gastrointestinal (GI) tract express the same molecular receptors and intracellular signaling components known to be involved in taste has generated great interest in potential functions of such post-oral "taste" receptors in the control of food intake. To determine whether taste cues in the GI tract are detected and can directly influence behavior, the present study used a microbehavioral analysis of intake, in which rats drank from lickometers that were programmed to simultaneously deliver a brief yoked infusion of a taste stimulus to the intestines. Specifically, in daily 30-min sessions, thirsty rats with indwelling intraduodenal catheters were trained to drink hypotonic (0.12 M) sodium chloride (NaCl) and simultaneously self-infuse a 0.12 M NaCl solution. Once trained, in a subsequent series of intestinal taste probe trials, rats reduced licking during a 6-min infusion period, when a bitter stimulus denatonium benzoate (DB; 10 mM) was added to the NaCl vehicle for infusion, apparently conditioning a mild taste aversion. Presentation of the DB in isomolar lithium chloride (LiCl) for intestinal infusions accelerated the development of the response across trials and strengthened the temporal resolution of the early licking suppression in response to the arrival of the DB in the intestine. In an experiment to evaluate whether CCK is involved as a paracrine signal in transducing the intestinal taste of DB, the CCK-1R antagonist devazepide partially blocked the response to intestinal DB. In contrast to their ability to detect and avoid the bitter taste in the intestine, rats did not modify their licking to saccharin intraduodenal probe infusions. The intestinal taste aversion paradigm developed here provides a sensitive and effective protocol for evaluating which tastants-and concentrations of tastants-in the lumen of the gut can control ingestion.


Senior student goal setting during subinternship: a mixed-methods study.
New developments in Subinternship curricula attempt to establish self-directed learning skills that will translate into the 1st year of residency. However, we know little about how well senior medical students' self-directed learning goals match with what is expected of them in residency. We designed a mixed-methods study to examine the goals set by senior students at Georgia Health Sciences University on Pediatric Subinternship and to determine how those goals relate to the 6 Accreditation Council for Graduate Medical Education (ACGME) competencies. We used an iterative process to categorize self-directed learning goals on Pediatric Subinternships (n=188 goals generated by 57 senior students) by (a) the 6 ACGME competencies and (b) general or specific goals. We used tests of association and multivariate modeling to compare goal categories by clinical site and time of year. There were 56.3% of goals addressing patient care. Professionalism and systems-based practice goals were rare. Of the goals, 72% were general, but goals written by students at the newborn nursery and neonatal intensive care unit sites were significantly more likely to be specific than goals written by students on the general inpatient subinternship. The overwhelming majority of senior medical student goals on a Pediatric Subinternship are general and related to patient care. Students may need assistance with developing more specific goals in all 6 competencies. Our findings suggest that understanding and incorporating students' learning goals may be important for enhancing the potential effectiveness of transition-to-residence curricula.


Preliminary randomized double-blind placebo-controlled trial of tryptophan combined with fluoxetine to treat major depressive disorder: antidepressant and hypnotic effects.
Because the initial phase of treatment of depression with a selective serotonin reuptake inhibitor is often complicated by a delayed onset of action of the antidepressant or severe insomnia or both, we investigated whether tryptophan, an amino acid with both antidepressant-augmenting and hypnotic effects, would benefit patients with depression at the beginning of treatment with fluoxetine. Randomized, double-blind, placebo-controlled trial. Thirty individuals with major depressive disorder. Treatment over 8 weeks with 20 mg of fluoxetine per day and either tryptophan (2 to 4 g per day) or placebo. Mood was assessed using the 29-item Hamilton Depression Rating Scale (HDRS-29) and the Beck Depression Inventory (BDI). Laboratory sleep studies were done at baseline and after 4 and 8 weeks of treatment using standard procedures. During the first week of treatment, there was a significantly greater decrease in HDRS-29 depression scores, and a similar trend in BDI scores, in the tryptophan/fluoxetine group than in the placebo/fluoxetine group. No significant differences were noted at later time points. With respect to sleep measures, there was a significant group-by-time interaction for slow-wave sleep at week 4. Further analysis revealed a significant decrease in slow-wave sleep after 4 weeks of treatment in the placebo/fluoxetine group, but not in the tryptophan/fluoxetine group. No cases of serotonin syndrome occurred, and the combination was well tolerated, although the 4 g per day dosage of tryptophan produced daytime drowsiness. Combining 20 mg of fluoxetine with 2 g of tryptophan daily at the outset of treatment for major depressive disorder appears to be a safe protocol that may have both a rapid antidepressant effect and a protective effect on slow-wave sleep. Further large-scale studies are needed to confirm these initial findings.


Developing an ethics curriculum for a family practice residency.
To develop a curriculum in ethics in family practice by studying which ethical issues physicians believe to be important based on frequency of encounter, difficulty in managing the problem, and helpfulness of discussion; to examine whether there are any important differences between faculty and residents and between genders; and to determine the preferred format(s) for teaching these issues. Between November 1991 and June 1992 a total of 475 questionnaires were mailed to all the family practice residents (first- and second-year), graduates in their first two years of practice, and the physician faculty of the Department of Family Medicine at the McGill University Faculty of Medicine. Overall, 319 usable questionnaires were returned, for a final response rate of 67%. The questionnaire asked respondents to evaluate 14 ethical dilemmas in order to determine the importance of teaching specific ethical issues as well as to determine the preferred format for teaching. Chi-square tests, analyses of variance, and Student's t-tests were used to test the significance of differences in responses. No consistent pattern of interrelationship was found among frequency of encounter and difficulty and helpfulness of discussion for most items. Overall, there was little difference in how faculty and residents, men and women, perceived the importance of these ethical issues. Women reported encountering ethical issues less frequently than men [F (14,285) = 1.82, p < .04], while at the same time finding them somewhat more difficult and more deserving of discussion. Small-group, case-oriented discussion appears to have been the favored teaching format regardless of the ethical dilemma. It is difficult to narrow down the content to be included in a curriculum in ethics in family practice. Frequency of encounter, difficulty in management, and helpfulness of discussion can all be argued to be important factors for consideration; they should all be considered separately or in combination for each teaching situation if time restrictions force a choice between topics.


Behavioral and genetic evidence for a novel animal model of Attention-Deficit/Hyperactivity Disorder Predominantly Inattentive Subtype.
According to DSM-IV there are three subtypes of Attention-Deficit/Hyperactivity Disorder, namely: ADHD predominantly inattentive type (ADHD-PI), ADHD predominantly Hyperactive-Impulsive Type (ADHD-HI), and ADHD combined type (ADHD-C). These subtypes may represent distinct neurobehavioral disorders of childhood onset with separate etiologies. The diagnosis of ADHD is behaviorally based; therefore, investigations into its possible etiologies should be based in behavior. Animal models of ADHD demonstrate construct validity when they accurately reproduce elements of the etiology, biochemistry, symptoms, and treatment of the disorder. Spontaneously hypertensive rats (SHR) fulfill many of the validation criteria and compare well with clinical cases of ADHD-C. The present study describes a novel rat model of the predominantly inattentive subtype (ADHD-PI). ADHD-like behavior was tested with a visual discrimination task measuring overactivity, impulsiveness and inattentiveness. Several strains with varied genetic background were needed to determine what constitutes a normal comparison. Five groups of rats were used: SHR/NCrl spontaneously hypertensive and WKY/NCrl Wistar/Kyoto rats from Charles River; SD/NTac Sprague Dawley and WH/HanTac Wistar rats from Taconic Europe; and WKY/NHsd Wistar/Kyoto rats from Harlan. DNA was analyzed to determine background differences in the strains by PCR genotyping of eight highly polymorphic microsatellite markers and 2625 single nucleotide polymorphisms (SNPs). Compared to appropriate comparison strains (WKY/NHsd and SD/NTac rats), SHR/NCrl showed ADHD-C-like behavior: striking overactivity and poor sustained attention. Compared to WKY/NHsd rats, WKY/NCrl rats showed inattention, but no overactivity or impulsiveness. WH/HanTac rats deviated significantly from the other control groups by being more active and less attentive than the WKY/NHsd and SD/NTac rats. We also found substantial genomic differences between the WKY/NCrl and WKY/NHsd rats for eight short tandem repeat loci and 2625 SNPs. About 33.5 percent of the genome differs between the two WKY rat substrains, with large stretches of divergence on each chromosome. These data provide solid behavioral and genetic evidence that the WKY/NCrl and WKY/NHsd rats should be considered as separate substrains. Moreover, the behavioral features of the WKY/NCrl rat indicate that it should be a useful model for ADHD-PI, the primarily inattentive subtype of ADHD. The SD/NTac and the WH/HanTac rats show significant genetic and/or behavioral differences from WKY/NHsd rats and appear not to be appropriate controls in studies using the SHR/NCrl. The present results support the conclusion that SHR/NCrl is the best validated animal model of ADHD-C. The overactivity, impulsiveness and deficient sustained attention of the SHR/NCrl strain are independent behaviors. Thus, overactivity does not account for this strain's impulsiveness and deficient sustained attention. Finally, the present study shows that great care has to be exercised to select the model and comparison groups.


The E3 ubiquitin ligase HECTD3 regulates ubiquitination and degradation of Tara.
Tara was identified as an interacting partner of guanine nucleotide exchange factor Trio and TRF1. Tara is proposed to be involved in many important fundamental cellular processes, ranging from actin remodeling, directed cell movement, to cell cycle regulation. Yet, its exact roles required further elucidation. Here, we identify a novel Tara-binding protein HECTD3, a putative member of HECT E3 ubiquitin ligases. HECTD3 directly binds Tara in vitro and forms a complex with Tara in vivo. Overexpression of HECTD3 enhances the ubiquitination of Tara in vivo and promotes the turnover of Tara, whereas depletion of HECTD3 by small interfering RNA decreases Tara degradation. Furthermore, depletion of HECTD3 leads to multipolar spindle formation. All these findings suggest that HECTD3 may facilitate cell cycle progression via regulating ubiquitination and degradation of Tara.


Over-parameterised, uncertain 'mathematical marionettes' - how can we best use catchment water quality models? An example of an 80-year catchment-scale nutrient balance.
The Integrated Catchment Model of Nitrogen (INCA-N) was applied to the River Lambourn, a Chalk river-system in southern England. The model's abilities to simulate the long-term trend and seasonal patterns in observed stream water nitrate concentrations from 1920 to 2003 were tested. This is the first time a semi-distributed, daily time-step model has been applied to simulate such a long time period and then used to calculate detailed catchment nutrient budgets which span the conversion of pasture to arable during the late 1930s and 1940s. Thus, this work goes beyond source apportionment and looks to demonstrate how such simulations can be used to assess the state of the catchment and develop an understanding of system behaviour. The mass-balance results from 1921, 1922, 1991, 2001 and 2002 are presented and those for 1991 are compared to other modelled and literature values of loads associated with nitrogen soil processes and export. The variations highlighted the problem of comparing modelled fluxes with point measurements but proved useful for identifying the most poorly understood inputs and processes thereby providing an assessment of input data and model structural uncertainty. The modelled terrestrial and instream mass-balances also highlight the importance of the hydrological conditions in pollutant transport. Between 1922 and 2002, increased inputs of nitrogen from fertiliser, livestock and deposition have altered the nitrogen balance with a shift from possible reduction in soil fertility but little environmental impact in 1922, to a situation of nitrogen accumulation in the soil, groundwater and instream biota in 2002. In 1922 and 2002 it was estimated that approximately 2 and 18 kg N ha(-1) yr(-1) respectively were exported from the land to the stream. The utility of the approach and further considerations for the best use of models are discussed.


Energetic constraints on the diet of terrestrial carnivores.
Species in the mammalian order Carnivora exhibit a huge diversity of life histories with body sizes spanning more than three orders of magnitude. Despite this diversity, most terrestrial carnivores can be classified as either feeding on invertebrates and small vertebrates or on large vertebrates. Small carnivores feed predominantly on invertebrates probably because they are a superabundant resource (sometimes 90% of animal biomass); however, intake rates of invertebrate feeders are low, about one tenth of those of vertebrate feeders. Although small carnivores can subsist on this diet because of low absolute energy requirements, invertebrate feeding appears to be unsustainable for larger carnivores. Here we show, by reviewing the most common live prey in carnivore diets, that there is a striking transition from feeding on small prey (less than half of predator mass) to large prey (near predator mass), occurring at predator masses of 21.5-25 kg. We test the hypothesis that this dichotomy is the consequence of mass-related energetic requirements and we determine the predicted maximum mass that an invertebrate diet can sustain. Using a simple energetic model and known invertebrate intake rates, we predict a maximum sustainable mass of 21.5 kg, which matches the point where predators shift from small to large prey.


Synthetic aperture superresolution by speckle pattern projection.
We propose a method for increasing the resolution of an aperture limited optical system by illuminating the input with a speckle pattern. The high resolution of the projected speckle pattern demodulates the high frequencies of the sample and permits its passage through the system aperture. A decoding provides the superresolved image. The speckle pattern can be generated in a simple manner in contrast with other structured light superresolution methods. The method is demonstrated in microscopy test images.


Factor structure of the Children's Perception of Interparental Conflict Scale for studies of youths with externalizing behavior problems.
The assessment of children's perception of marital conflict is an important area of research related to family relations, children's cognitions about self, and the development of psychopathology. The leading instrument in this domain is the Children's Perception of Interparental Conflict Scale (CPIC; J. H. Grych, M. Seid, & F. D. Fincham, 1992). It has 48 items organized into 9 conceptual designed subscales, with reports of 3-factor higher order structure to the 9 subscales. However, the 3-factor solution does not capture all 9 subscales well. Further, items have never been subjected to factor analysis to evaluate the best fitting factor structure at the item level. Doing so is particularly important when bringing the scale into new populations such as children with attention-deficit/hyperactivity disorder (ADHD) or disruptive behavior disorders. In the present study, 2 samples of children (total N = 1,190; ages 6-18 years) completed the CPIC. An exploratory factor analysis in Sample 1 (from a clinical study of ADHD and non-ADHD youths) yielded 4 interpretable factors. A confirmatory factor analysis in Sample 2 (a population sample of twins) confirmed the generalizability of the solution with an acceptable fit, although 1 item was dropped. The final solution used 38 of the 48 items. The 4-factor solution captured a Conflict Properties factor, two appraisal factors labeled Threat to Self and Self-Blame, and a Triangulation/Stability factor that included elements of appraisal and content. The authors concluded that the item-based 4-factor solution to the CPIC is preferable to the 9-factor or 3-factor formulation.


Antiviral T-cell-independent type 2 antibody responses induced in vivo in the absence of T and NK cells.
Polyomavirus (PyV) infection induces protective T-cell-independent (TI) IgM and IgG responses in T-cell-deficient (TCR beta x delta-/-) mice. In this study, we show that PyV is a TI -2 antigen: B cells with a mutated Bruton's tyrosine kinase (Xid mutants) do not respond to PyV with antibody secretion in the absence of T cells. We also demonstrate that NK-cell-mediated "help" is not absolutely required for the induction of the TI-2 antibodies to PyV; thus for the first time, we provide evidence for protective IgM and IgG responses against a viral infection induced in mice lacking T and NK cells (CD3Etg). Comparison of the antibody responses observed in T- and NK-cell-deficient mice with those of mice lacking only T cells, however, suggests that NK cells may promote isotype switching to IgG2a. This effect is probably mediated by IFN gamma secretion. In support of this idea, studies on the antibody responses of PyV-infected SCID mice that had been reconstituted with IFN gamma R-/- B cells or wild-type B cells demonstrated the IFN gamma dependence of PyV-specific TI IgG2a secretion and provided evidence that IFN gamma acting directly on B cells plays an important role in TI pathways of isotype switching to IgG2a in vivo.


Detection of antimicrobial residues in poultry meat and slaughter cattle in Nigeria.
The European four plate test (FPT) was used to establish whether and where antimicrobial residues accumulate in the tissue of commercial poultry and slaughter cattle in Nigeria. Bacillus subtilis and Staphylococcus aureus sensitive to different antimicrobials seeded on nutrient agar media at different pH levels were employed. Inhibitory residues were detected in the muscles, liver, and/or kidney of experimental local chickens, commercial broilers and layers and 2 of 24 slaughter cattle. The public health hazards of antimicrobial residues in developing countries are discussed.


Iron-induced cardiac damage: role of apoptosis and deferasirox intervention.
Excess cardiac iron levels are associated with cardiac damage and can result in increased morbidity and mortality. Here, we hypothesize that elevations in tissue iron can activate caspase-dependent signaling, which leads to increased cardiac apoptosis and fibrosis, and that these alterations can be attenuated by iron chelation. Using an iron-overloaded gerbil model, we show that increased cardiac iron is associated with reduced activation of Akt (Ser473 and Thr308), diminished phosphorylation of the proapoptotic regulator Bad (Ser136), and an increased Bax/Bcl-2 ratio. These iron-overload-induced alterations in Akt/Bad phosphorylation and Bax/Bcl-2 ratio were coupled with increased activation of the downstream caspase-9 (40/38- and 17-kDa fragments) and apoptosis executioner caspase-3 (19- and 17-kDa fragments), which were accompanied by evidence of elevated cytoskeletal α-fodrin cleavage (150- and 120-kDa fragments), discontinuity of myocardial membrane dystrophin immunoreactivity, increases in the number of terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL)-positive cells (nucleic DNA fragmentation), and cardiac fibrosis. We demonstrate that the administration of deferasirox, a tridentate iron chelator, is associated with diminished tissue iron deposition, attenuated activation of caspases, reduced α-fodrin cleavage, improved membrane integrity, decreased TUNEL reactivity, and attenuated cardiac fibrosis. These results suggest that the activation of caspase-dependent signaling may play a role in the development of iron-induced cardiac apoptosis and fibrosis, and deferasirox, via a reduction in cardiac tissue iron levels, may be useful for decreasing the extent of iron-induced cardiac damage.


Optimum lymphadenectomy for esophageal cancer.
Using Worldwide Esophageal Cancer Collaboration data, we sought to (1) characterize the relationship between survival and extent of lymphadenectomy, and (2) from this, define optimum lymphadenectomy. What constitutes optimum lymphadenectomy to maximize survival is controversial because of variable goals, analytic methodology, and generalizability of the underpinning data. A total of 4627 patients who had esophagectomy alone for esophageal cancer were identified from the Worldwide Esophageal Cancer Collaboration database. Patient-specific risk-adjusted survival was estimated using random survival forests. Risk-adjusted 5-year survival was averaged for each number of lymph nodes resected and its relation to cancer characteristics explored. Optimum number of nodes that should be resected to maximize 5-year survival was determined by random forest multivariable regression. For pN0M0 moderately and poorly differentiated cancers, and all node-positive (pN+) cancers, 5-year survival improved with increasing extent of lymphadenectomy. In pN0M0 cancers, no optimum lymphadenectomy was defined for pTis; optimum lymphadenectomy was 10 to 12 nodes for pT1, 15 to 22 for pT2, and 31 to 42 for pT3/T4, depending on histopathologic cell type. In pN+M0 cancers and 1 to 6 nodes positive, optimum lymphadenectomy was 10 for pT1, 15 for pT2, and 29 to 50 for pT3/T4. Greater extent of lymphadenectomy was associated with increased survival for all patients with esophageal cancer except at the extremes (TisN0M0 and >or=7 regional lymph nodes positive for cancer) and well-differentiated pN0M0 cancer. Maximum 5-year survival is modulated by T classification: resecting 10 nodes for pT1, 20 for pT2, and >or=30 for pT3/T4 is recommended.


Reconstruction of the immunogenic peptide RNase(43-56) by identification and transfer of the critical residues into an unrelated peptide backbone.
The involvement of each of the amino acid residues of the I-Ak-restricted T cell determinant RNase(43-56) was examined in detail using a series of peptides containing single amino acid substitutions. Four positions were identified as being essential for the formation of the determinant, Phe-46, Val-47, His-48, and Leu-51. When these four residues were substituted into the backbone of the unrelated peptide HA(130-144), a nonstimulatory peptide was obtained. The inclusion of an additional residue, Val-54, resulted in a chimeric peptide, RN/HA2, which was nearly as active as the native molecule. The peptide RN/HA2 was able to prime in vivo for RNase reactivity, confirming that these five residues contained all of the specificity of the RNase(43-56) determinant. The role of three of these critical residues was examined using both a functional competition assay and an in vivo priming assay. It was ascertained that the Phe-46 was directly involved in contacting the TCR, while the His-48 and Leu-51 were either involved in binding to the I-Ak molecule or in determining the conformation of the peptide. Thus, by critically evaluating the contribution of each of the amino acid residues in a T cell determinant, we were able to generate a chimeric peptide only containing 5 of 15 residues from the RNase(43-56) sequence that was functionally identical to the native RNase(43-56) molecule both in vitro and in vivo.


Heat Treatments and Critical Quenching Rates in Additively Manufactured Al-Si-Mg Alloys.
Laser powder-bed fusion (LPBF) has significantly gained in importance and has become one of the major fabrication techniques within metal additive manufacturing. The fast cooling rates achieved in LPBF due to a relatively small melt pool on a much larger component or substrate, acting as heat sink, result in fine-grained microstructures and high oversaturation of alloying elements in the α-aluminum. Al-Si-Mg alloys thus can be effectively precipitation hardened. Moreover, the solidified material undergoes an intrinsic heat treatment, whilst the layers above are irradiated and the elevated temperature in the built chamber starts the clustering process of alloying elements directly after a scan track is fabricated. These silicon-magnesium clusters were observed with atom probe tomography in as-built samples. Similar beneficial clustering behavior at higher temperatures is known from the direct-aging approach in cast samples, whereby the artificial aging is performed immediately after solution annealing and quenching. Transferring this approach to LPBF samples as a possible post-heat treatment revealed that even after direct aging, the outstanding hardness of the as-built condition could, at best, be met, but for most instances it was significantly lower. Our investigations showed that LPBF Al-Si-Mg exhibited a high dependency on the quenching rate, which is significantly more pronounced than in cast reference samples, requiring two to three times higher quenching rate after solution annealing to yield similar hardness results. This suggests that due to the finer microstructure and the shorter diffusion path in Al-Si-Mg fabricated by LPBF, it is more challenging to achieve a metastable oversaturation necessary for precipitation hardening. This may be especially problematic in larger components.


Prediction and identification of Arabidopsis thaliana microRNAs and their mRNA targets.
A class of eukaryotic non-coding RNAs termed microRNAs (miRNAs) interact with target mRNAs by sequence complementarity to regulate their expression. The low abundance of some miRNAs and their time- and tissue-specific expression patterns make experimental miRNA identification difficult. We present here a computational method for genome-wide prediction of Arabidopsis thaliana microRNAs and their target mRNAs. This method uses characteristic features of known plant miRNAs as criteria to search for miRNAs conserved between Arabidopsis and Oryza sativa. Extensive sequence complementarity between miRNAs and their target mRNAs is used to predict miRNA-regulated Arabidopsis transcripts. Our prediction covered 63% of known Arabidopsis miRNAs and identified 83 new miRNAs. Evidence for the expression of 25 predicted miRNAs came from northern blots, their presence in the Arabidopsis Small RNA Project database, and massively parallel signature sequencing (MPSS) data. Putative targets functionally conserved between Arabidopsis and O. sativa were identified for most newly identified miRNAs. Independent microarray data showed that the expression levels of some mRNA targets anti-correlated with the accumulation pattern of their corresponding regulatory miRNAs. The cleavage of three target mRNAs by miRNA binding was validated in 5' RACE experiments. We identified new plant miRNAs conserved between Arabidopsis and O. sativa and report a wide range of transcripts as potential miRNA targets. Because MPSS data are generated from polyadenylated RNA molecules, our results suggest that at least some miRNA precursors are polyadenylated at certain stages. The broad range of putative miRNA targets indicates that miRNAs participate in the regulation of a variety of biological processes.


Femtosecond Cr:LiSAF and Cr:LiCAF lasers pumped by tapered diode lasers.
We report compact, low-cost and efficient Cr:Colquiriite lasers that are pumped by high brightness tapered laser diodes. The tapered laser diodes provided 1 to 1.2 W of output power around 675 nm, at an electrical-to-optical conversion efficiency of about 30%. Using a single tapered diode laser as the pump source, we have demonstrated output powers of 500 mW and 410 mW together with slope efficiencies of 47% and 41% from continuous wave (cw) Cr:LiSAF and Cr:LiCAF lasers, respectively. In cw mode-locked operation, sub-100-fs pulse trains with average power between 200 mW and 250 mW were obtained at repetition rates around 100 MHz. Upon pumping the Cr:Colquiriite lasers with two tapered laser diodes (one from each side of the crystal), we have observed scaling of cw powers to 850 mW in Cr:LiSAF and to 650 mW in Cr:LiCAF. From the double side pumped Cr:LiCAF laser, we have also obtained ~220 fs long pulses with 5.4 nJ of pulse energy at 77 MHz repetition rate. These are the highest energy levels reported from Cr:Colquiriite so far at these repetition rates. Our findings indicate that tapered diodes in the red spectral region are likely to become the standard pump source for Cr:Colquiriite lasers in the near future. Moreover, the simplified pumping scheme might facilitate efficient commercialization of Cr:Colquiriite systems, bearing the potential to significantly boost applications of cw and femtosecond lasers in this spectral region (750-1000 nm).


Are there early signs that predict development of temporomandibular joint disease?
Temporomandibular joint disorders (TMJD) involve orofacial pain and functional limitations that may limit important daily activities such as chewing and speaking. This observational case-control study attempted to identify factors associated with TMJD development, particularly inflammation. The study participants were patients treated at Karolinska University Hospital, Stockholm, Sweden. The cases were patients who received a diagnosis of TMJD, chronic closed lock, or painful clicking and were treated surgically during the period from 2007 through 2011. The control group was randomly selected from among patients who had undergone tooth extraction and was matched by age and sex. A total of 146 cases and 151 controls were included in the analyses. The response rate was 55.3% for the case group and 21.8% for the control group. The male:female ratio for patients with TMJD was 1:4.4. TMJD was significantly associated with pneumonia (odds ratio [OR], 2.1), asthma (OR, 2.1), allergies (OR, 1.8), headache (OR, 3.1), general joint hypermobility (OR, 3.8), orofacial trauma (OR, 3.9), rheumatism (OR, 2.5), and orthodontic treatment (OR, 2.4) (P < 0.05 for all outcomes). In conclusion, autoimmune diseases and inflammatory conditions are associated with increased risk of TMJD. Moreover, certain lung disorders may predict subsequent development of TMJD.


Distal anterior inferior cerebellar artery aneurysm masquerading as a cerebellopontine angle tumor: case report and review of literature.
We present the case of a distal anterior inferior cerebellar artery (AICA) aneurysm masquerading as a cerebellopontine angle tumor in a 60-year-old right-handed man with previously undiagnosed polyarteritis nodosa (PAN). The patient presented with a 2-month history of progressive right-sided hearing loss, intermittent severe headache, and sudden onset of complete facial paralysis 3 weeks before admission. Magnetic resonance imaging, including post-gadolinium images, showed a 1.2-cm heterogeneously enhancing mass that slightly enlarged the right internal auditory canal. A right suboccipital craniotomy was performed, and a partially thrombosed fusiform AICA aneurysm was discovered just anterior to the VII/VIII nerve complex. The aneurysm was trapped and opened, and a thrombectomy was performed. Postoperatively, the patient experienced abdominal pain; liver function tests were abnormal. Investigation revealed a small retroperitoneal hemorrhage and aneurysms of the celiac axis and gastroduodenal arteries. Further investigation revealed an increased erythrocyte sedimentation rate, and a diagnosis of PAN was made. PAN is a well-identified factor in the genesis of peripheral vascular aneurysms. Aneurysms involving the hepatic, renal, coronary, pancreatic, and tibial arteries have been described. PAN is an extremely rare cause of intracranial aneurysm. Patients who present with aneurysms in unusual locations (e.g., distal AICA) should be investigated for vasculopathy and collagen vascular disorders.


Involving the general public in priority setting: experiences from Australia.
The discussion over whether community preferences have a legitimate role to play in priority setting has been highly polarised. Skeptics warn of the risk of establishing a 'dictatorship of the uninformed', while advocates proclaim the legitimacy of the participatory process. The one group who appears not to be consulted in this debate is the citizens themselves. In this study, a convenience sample of 373 citizens attending two medical clinics in central Sydney were surveyed about whether the general public has a legitimate role to play in informing priority setting in health care. Respondents were presented with three different levels of priority setting: across health care programmes, across medical procedures, and at a global level. To assist respondents in understanding the choices and trade-offs involved, they were given information about current levels of funding and the cost-effectiveness of each alternative. Respondents were asked whether they felt the preferences of the general public should be used to inform priority setting at each level. Of particular interest was the question of whether their willingness to use public preferences depended on the level of priority setting. Respondents were also asked about who else's preferences should be used to inform priority setting at each level. The results suggest that the public overwhelmingly want their preferences to inform priority-setting decisions in health care. This was seen to be particularly important in informing decisions about how to prioritise across broad health care programmes and about the criteria to be used to allocate funds across different population groups. In contrast, the preferences of medical professionals and health service managers were rated most highly in relation to the prioritisation of different treatments and medical procedures. In most cases, however, respondents did not advocate the use of one particular group's preferences. Even when the preferences of the general public were considered most important, it was felt that any decision-making process needed to be informed by the preferences of a range of groups. The preferences of politicians were viewed as least important to processes of priority setting in health care.


A contour-guided deformable image registration algorithm for adaptive radiotherapy.
In adaptive radiotherapy, deformable image registration is often conducted between the planning CT and treatment CT (or cone beam CT) to generate a deformation vector field (DVF) for dose accumulation and contour propagation. The auto-propagated contours on the treatment CT may contain relatively large errors, especially in low-contrast regions. A clinician's inspection and editing of the propagated contours are frequently needed. The edited contours are able to meet the clinical requirement for adaptive therapy; however, the DVF is still inaccurate and inconsistent with the edited contours. The purpose of this work is to develop a contour-guided deformable image registration (CG-DIR) algorithm to improve the accuracy and consistency of the DVF for adaptive radiotherapy. Incorporation of the edited contours into the registration algorithm is realized by regularizing the objective function of the original demons algorithm with a term of intensity matching between the delineated structures set pairs. The CG-DIR algorithm is implemented on computer graphics processing units (GPUs) by following the original GPU-based demons algorithm computation framework (Gu et al 2010 Phys Med Biol. 55 207-219). The performance of CG-DIR is evaluated on five clinical head-and-neck and one pelvic cancer patient data. It is found that compared with the original demons, CG-DIR improves the accuracy and consistency of the DVF, while retaining similar high computational efficiency.


Charged surface area of maurocalcine determines its interaction with the skeletal ryanodine receptor.
The 33 amino acid scorpion toxin maurocalcine (MCa) has been shown to modify the gating of the skeletal-type ryanodine receptor (RyR1). Here we explored the effects of MCa and its mutants ([Ala(8)]MCa, [Ala(19)]MCa, [Ala(20)]MCa, [Ala(22)]MCa, [Ala(23)]MCa, and [Ala(24)]MCa) on RyR1 incorporated into artificial lipid bilayers and on elementary calcium release events (ECRE) in rat and frog skeletal muscle fibers. The peptides induced long-lasting subconductance states (LLSS) on RyR1 that lasted for several seconds. However, their average length and frequency were decreased if the mutation was placed farther away in the 3D structure from the critical (24)Arg residue. The effect was strongly dependent on the direction of the current through the channel. If the direction was similar to that followed by calcium during release, the peptides were 8- to 10-fold less effective. In fibers long-lasting calcium release events were observed after the addition of the peptides. The average length of these events correlated well with the duration of LLSS. These data suggest that the effect of the peptide is governed by the large charged surface formed by residues Lys(20), Lys(22), Arg(23), Arg(24), and Lys(8). Our observations also indicate that the results from bilayer experiments mimic the in situ effects of MCa on RyR1.


Supplementing Conventional Treatment with Pycnogenol® May Improve Hepatitis C Virus-Associated Type 2 Diabetes: A Mini Review.
Hepatitis C virus (HCV) infection and type 2 diabetes mellitus (T2DM) present a significant health burden, with increasing complications and mortality rates worldwide. Pycnogenol® (PYC), a natural product, possesses antidiabetic and antiviral properties that may improve HCV-associated T2DM. In this review, we present previously published data on the effectiveness of PYC against HCV replication and T2DM. We believe that supplementing conventional treatment with PYC may improve the current HCV therapy, attenuate HCV-associated T2DM, and reduce the risk of complications such as cirrhosis or hepatocellular carcinoma and cardiovascular disease.


Genetic investigation of novel hantaviruses causing fatal HPS in Brazil.
Although hantavirus pulmonary syndrome (HPS) was discovered in North America in 1993, more recent investigations have shown that the disease is a much larger problem in South America, where a greater number of cases and HPS-associated viruses have now been detected. Here we describe the genetic investigation of three fatal HPS cases from Brazil, including a 1995 case in Castelo dos Sonhos (CAS) in the state of Mato Grosso and two 1996 cases in the counties of Araraquara (ARA) and Franca (FRA), in the state of São Paulo. Reverse transcription-polymerase chain reaction (RT-PCR) products representing fragments of the hantavirus N, G1, and G2 coding regions were amplified from patient acute-phase serum samples, and the nucleotide (nt) sequences (394, 259, and 139 nt, respectively) revealed high deduced amino acid sequence identity between ARA and FRA viruses (99.2%, 96.5%, and 100%, respectively). However, amino acid differences of up to 14.0% were observed when ARA and FRA virus sequences were compared with those of the geographically more distant CAS virus. Analysis of a 643-nt N coding region and a 1734-nt predominantly G2-encoding region of ARA and CAS virus genomes confirmed that these Brazilian viruses were distinct and monophyletic with previously characterized Argentinean hantaviruses, and suggested that Laguna Negra (LN) virus from Paraguay was ancestral to both the Brazilian and Argentinean viruses. The phylogenetic tree based on the N coding fragment also placed LN in a separate clade with Rio Mamore virus from Bolivia. At the amino acid level, ARA and CAS viruses appeared more closely related to the Argentinean viruses than they were to each other. Similarly, analysis of the diagnostic 139-nt G2 fragment showed that the Juquitiba virus detected in a 1993 fatal HPS case close to São Paulo city, Brazil was closer to Argentinean viruses than to ARA or CAS viruses. These data indicate that at least three different hantavirus genetic lineages are associated with Brazilian HPS cases.


Static hyperspectral imaging polarimeter for full linear Stokes parameters.
A compact, static hyperspectral imaging linear polarimeter (HILP) based on a Savart interferometer (SI) is conceptually described. It improves the existing SI by replacing front polarizer with two Wollaston prisms, and can simultaneously acquire four interferograms corresponding to four linearly polarized lights on a single CCD. The spectral dependence of linear Stokes parameters can be recovered with Fourier transformation. Since there is no rotating or moving parts, the system is relatively robust. The interference model of the HILP is proved. The performance of the system is demonstrated through a numerical simulation, and the methods for compensating the imperfection of the polarization elements are described.


How a DNA polymerase clamp loader opens a sliding clamp.
Processive chromosomal replication relies on sliding DNA clamps, which are loaded onto DNA by pentameric clamp loader complexes belonging to the AAA+ family of adenosine triphosphatases (ATPases). We present structures for the ATP-bound state of the clamp loader complex from bacteriophage T4, bound to an open clamp and primer-template DNA. The clamp loader traps a spiral conformation of the open clamp so that both the loader and the clamp match the helical symmetry of DNA. One structure reveals that ATP has been hydrolyzed in one subunit and suggests that clamp closure and ejection of the loader involves disruption of the ATP-dependent match in symmetry. The structures explain how synergy among the loader, the clamp, and DNA can trigger ATP hydrolysis and release of the closed clamp on DNA.


Helicopter-based in-water resuscitation with chest compressions: a pilot study.
Drowning is a relevant worldwide cause of severe disability and death. The delay of ventilations and chest compressions is a crucial problem in drowning victims. Hence, a novel helicopter-based ALS rescue concept with in-water ventilation and chest compressions was evaluated. Cardio pulmonary resuscitation (CPR) and vascular access were performed in a self-inflating Heliboat platform in an indoor wave pool using the Fastrach intubating laryngeal mask, the Oxylator resuscitator, Lund University Cardiopulmonary Assist System (LUCAS) chest compression device and EZ-IO intraosseous power drill. The time requirement and physical exertion on a Visual Analogue Scale (VAS) were compared between a procedure without waves and with moderate swell. Measurement of the elapsed time of the various stages of the procedure did not reveal significant differences between calm water and swell: Ventilation was initiated after 02:48 versus 03:02 and chest compression after 04:20 versus 04:18 min; the intraosseous cannulisation was completed after 05:59 versus 06:30 min after a simulated jump off the helicopter. The attachment of the LUCAS to the mannequin and the intraosseous cannulisation was rated significantly more demanding on the VAS during swell conditions. CPR appears to be possible when performed in a rescue platform with special equipment. The novel helicopter-based strategy appears to enable the rescuers to initiate CPR in an appropriate length of time and with an acceptable amount of physical exertion for the divers. The time for the helicopter to reach the patient will have to be very short to minimise neurological damage in the drowning victim.


The burden and determinants of neck pain in workers: results of the Bone and Joint Decade 2000-2010 Task Force on Neck Pain and Its Associated Disorders.
Systematic review and best evidence synthesis. To describe the prevalence and incidence of neck pain and disability in workers; to identify risk factors for neck pain in workers; to propose an etiological diagram; and to make recommendations for future research. Previous reviews of the etiology of neck pain in workers relied on cross-sectional evidence. Recently published cohorts and randomized trials warrant a re-analysis of this body of research. We systematically searched Medline for literature published from 1980-2006. Retrieved articles were reviewed for relevance. Relevant articles were critically appraised. Articles judged to have adequate internal validity were included in our best evidence synthesis. One hundred and nine papers on the burden and determinants of neck pain in workers were scientifically admissible. The annual prevalence of neck pain varied from 27.1% in Norway to 47.8% in Québec, Canada. Each year, between 11% and 14.1% of workers were limited in their activities because of neck pain. Risk factors associated with neck pain in workers include age, previous musculoskeletal pain, high quantitative job demands, low social support at work, job insecurity, low physical capacity, poor computer workstation design and work posture, sedentary work position, repetitive work and precision work. We found preliminary evidence that gender, occupation, headaches, emotional problems, smoking, poor job satisfaction, awkward work postures, poor physical work environment, and workers' ethnicity may be associated with neck pain. There is evidence that interventions aimed at modifying workstations and worker posture are not effective in reducing the incidence of neck pain in workers. Neck disorders are a significant source of pain and activity limitations in workers. Most neck pain results from complex relationships between individual and workplace risk factors. No prevention strategies have been shown to reduce the incidence of neck pain in workers.


Health aspects of cannabis.
Marijuana seems firmly established as another social drug in Western countries, regardless of its current legal status. Patterns of use vary widely. As with other social drugs, the pattern of use is critical in determining adverse effects on health. Perhaps the major area of concern about marijuana use is among the very young. Using any drug on a regular basis that alters reality may be detrimental to the psychosocial maturation of young persons. Chronic use of marijuana may stunt the emotional growth of youngsters. Evidence for an amotivational syndrome is largely based on clinical reports; whether marijuana use is a cause or effect is uncertain. A marijuana psychosis, long rumored, has been difficult to prove. No one doubts that marijuana use may aggravate existing psychoses or other severe emotional disorders. Brain damage has not been proved. Physical dependence is rarely encountered in the usual patterns of social use, despite some degree of tolerance that may develop. The endocrine effects of the drug might be expected to delay puberty in prepubertal boys, but actual instances have been rare. As with any material that is smoked, chronic smoking of marijuana will produce bronchitis; emphysema or lung cancer have not yet been documented. Cardiovascular effects of the drug are harmful to those with preexisting heart disease; fortunately the number of users with such conditions is minimal. Fears that the drug might accumulate in the body to the point of toxicity have been groundless. The potential deleterious effects of marijuana use on driving ability seem to be self-evident; proof of such impairment has been more difficult. The drug is probably harmful when taken during pregnancy, but the risk is uncertain. One would be prudent to avoid marijuana during pregnancy, just as one would do with most other drugs not essential to life or well-being. No clinical consequences have been noted from the effects of the drug on immune response, chromosomes, or cell metabolites. Contamination of marijuana by spraying with defoliants has created the clearest danger to health; such attempts to control production should be abandoned. Therapeutic uses for marijuana, THC, or cannabinoid homologs are being actively explored. Only the synthetic homolog, nabilone, has been approved for use to control nausea and vomiting associated with cancer chemotherapy.(ABSTRACT TRUNCATED AT 400 WORDS)


Biochemical and Molecular Characterization of Carotenogenic Flavobacterial Isolates from Marine Waters.
Carotenoids are known to possess immense nutraceutical properties and microorganisms are continuously being explored as natural source for production of carotenoids. In this study, pigmented bacteria belonging to Flavobacteriaceae family were isolated using kanamycin-containing marine agar and identified using the molecular techniques and their phenotypic characteristics were studied along with their potential to produce carotenoids. Analysis of random amplification of polymorphic DNA (RAPD) banding patterns and the fragment size of the bands indicated that the 10 isolates fall under two major groups. Based on 16S rRNA gene sequence analysis the isolates were identified as Vitellibacter sp. (3 isolates), Formosa sp. (2 isolates) and Arenibacter sp. (5 isolates). Phenotypically, the isolates showed slight variation from the reported species of these three genera of Flavobacteriaceae. Only the isolates belonging to Vitellibacter and Formosa produced flexirubin, a typical yellow orange pigment produced by most of the organisms of the family Flavobacteriaceae. Vitellibacter sp. and Formosa sp. were found to produce higher amount of carotenoids compared to Arenibacter sp. and zeaxanthin was found to be the major carotenoid produced by these two species. The study indicated that Vitellibacter sp. and Formosa sp. can be exploited for production of carotenoids, particularly zeaxanthin.


Genetic and biochemical studies in yeast reveal that the cotton fibre-specific GhCER6 gene functions in fatty acid elongation.
3-ketoacyl-CoA synthase catalyses the initial condensation reaction during fatty acid elongation using malonyl-CoA and long-chain acyl-CoA as substrates. Previously, it was reported that several genes encoding putative cotton 3-ketoacyl-CoA synthases were significantly up-regulated during early cotton fibre development. In this study, GhCER6 cDNA that contains an open reading frame of 1479 bp, encoding a protein of 492 amino acid residues homologous to the Arabidopsis condensing enzyme CER6, was isolated and cloned. In situ hybridization results demonstrated that GhCER6 mRNA was detected only in the elongating wild-type cotton fibre cells. When GhCER6 was transformed to the Saccharomyces cerevisiae elo3 deletion mutation strain that was deficient in the production of 26-carbon fatty acids and displayed a very slow-growth phenotype, the mutant cells were found to divide similarly compared with those of the wild-type cells. Further, heterologous expression of GhCER6 restored the viability of the S. cerevisiae haploid elo2 and elo3 double-deletion strain. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis showed that GhCER6 was enzymatically active since the yeast elo2 and elo3 double-deletion mutant expressing the cotton gene produced very-long-chain fatty acids that are essential for cell growth. The results suggest that GhCER6 encodes a functional 3-ketoacyl-CoA synthase.


Thyroid storm following radioactive iodine (RAI) therapy for pediatric graves disease.
Female, 11 FINAL DIAGNOSIS: Thyroid storm Symptoms: Diarrhea • tachycardia • tachypnea • tremor • wheezing - Clinical Procedure: - Specialty: - Rare disease. A growing number of pediatric endocrinologists treat Graves disease with radioactive iodine (RAI) therapy due to the typically definitive nature of I-131 therapy. Given the published benefits and perceived low risks of RAI when compared to surgery or long-term anti-thyroid medication, the trend towards therapy with RAI is likely to continue. Nevertheless, RAI is not without significant risk. An 11-year-old girl with newly diagnosed Graves disease received RAI for definitive treatment of her hyperthyroidism. Within 24 hours of receiving I-131, she developed increasing sleepiness and eventually became unresponsive. Upon arrival at the emergency department she had a tonic-clonic seizure and was diagnosed with thyroid storm. Despite best efforts to manage her hyperthyroidism, she suffered a stroke of the left cerebral hemisphere that left her with persistent neurological deficits. Although thyroid storm after thyroid ablation is rare, the significant morbidity and potential mortality of pediatric thyroid storm warrant further studies to determine if children with markedly elevated thyroid hormone concentrations at diagnosis should receive prolonged pretreatment with anti-thyroid drugs. While such an approach may reduce the efficacy of I-131 ablation, it can also reduce and hopefully eliminate the risk of post-ablative thyroid storm.


Tonic tensor tympani syndrome in tinnitus and hyperacusis patients: a multi-clinic prevalence study.
Tonic tensor tympani syndrome (TTTS) is an involuntary, anxiety-based condition where the reflex threshold for tensor tympani muscle activity is reduced, causing a frequent spasm. This can trigger aural symptoms from tympanic membrane tension, middle ear ventilation alterations and trigeminal nerve irritability. TTTS is considered to cause the distinctive symptoms of acoustic shock (AS), which can develop after exposure to an unexpected loud sound perceived as highly threatening. Hyperacusis is a dominant AS symptom. Aural pain/blockage without underlying pathology has been noted in tinnitus and hyperacusis patients, without wide acknowledgment. This multiclinic study investigated the prevalence of TTTS symptoms and AS in tinnitus and hyperacusis patients. This study included consecutive patients with tinnitus and/or hyperacusis seen in multiple clinics. Symptoms consistent with TTTS (pain/numbness/burning in and around the ear; aural "blockage"; mild vertigo/nausea; "muffled" hearing; tympanic flutter; headache); onset or exacerbation from exposure to loud/intolerable sounds; tinnitus/hyperacusis severity. All patients were medically cleared of underlying pathology, which could cause these symptoms. 60.0% of the total sample (345 patients), 40.6% of tinnitus only patients, 81.1% of hyperacusis patients had ≥ 1 symptoms (P < 0.001). 68% of severe tinnitus patients, 91.3% of severe hyperacusis patients had ≥ 1 symptoms (P < 0.001). 19.7% (68/345) of patients in the total sample had AS. 83.8% of AS patients had hyperacusis, 41.2% of non-AS patients had hyperacusis (P < 0.001). The high prevalence of TTTS symptoms suggests they readily develop in tinnitus patients, more particularly with hyperacusis. Along with AS, they should be routinely investigated in history-taking.


The direction of membrane lipid flow in locomoting polymorphonuclear leukocytes.
The objective of this study was to determine the direction of membrane lipid flow in locomoting cells. The plasma membrane of human polymorphonuclear leukocytes was stained with a fluorescent lipid analog dihexadecanoyl indocarbocyanine. A line was photobleached on the cell surface perpendicular to the direction of cell motion. Low-light-level fluorescence microscopy and digital image-processing techniques were used to analyze a series of images taken at short intervals after photobleaching. The bleached line remained visible for about 5 seconds before being erased by diffusional recovery. Examination of fluorescence intensity profiles allowed a comparison to be made between the velocities of line and cell movement. Results indicate that the bleached line moves forward with the same velocity as the cell during locomotion, refuting the retrograde lipid flow model of locomotion. Instead, the plasma membrane lipid appears to move forward according to either the unit movement of membrane or the tank track model of locomotion.


Calibration and use of the polar organic chemical integrative sampler--a critical review.
The implementation of strict environmental quality standards for polar organic priority pollutants poses a challenge for monitoring programs. The polar organic chemical integrative sampler (POCIS) may help to address the challenge of measuring low and fluctuating trace concentrations of such organic contaminants, offering significant advantages over traditional sampling. In the present review, the authors evaluate POCIS calibration methods and factors affecting sampling rates together with reported environmental applications. Over 300 compounds have been shown to accumulate in POCIS, including pesticides, pharmaceuticals, hormones, and industrial chemicals. Polar organic chemical integrative sampler extracts have been used for both chemical and biological analyses. Several different calibration methods have been described, which makes it difficult to directly compare sampling rates. In addition, despite the fact that some attempts to correlate sampling rates with the properties of target compounds such as log K(OW) have been met with varying success, an overall model that can predict uptake is lacking. Furthermore, temperature, water flow rates, salinity, pH, and fouling have all been shown to affect uptake; however, there is currently no robust method available for adjusting for these differences. Overall, POCIS has been applied to a wide range of sampling environments and scenarios and has been proven to be a useful screening tool. However, based on the existing literature, a more mechanistic approach is required to increase understanding and thus improve the quantitative nature of the measurements.


Gradients of Rac1 Nanoclusters Support Spatial Patterns of Rac1 Signaling.
Rac1 is a small RhoGTPase switch that orchestrates actin branching in space and time and protrusion/retraction cycles of the lamellipodia at the cell front during mesenchymal migration. Biosensor imaging has revealed a graded concentration of active GTP-loaded Rac1 in protruding regions of the cell. Here, using single-molecule imaging and super-resolution microscopy, we show an additional supramolecular organization of Rac1. We find that Rac1 partitions and is immobilized into nanoclusters of 50-100 molecules each. These nanoclusters assemble because of the interaction of the polybasic tail of Rac1 with the phosphoinositide lipids PIP2 and PIP3. The additional interactions with GEFs and possibly GAPs, downstream effectors, and other partners are responsible for an enrichment of Rac1 nanoclusters in protruding regions of the cell. Our results show that subcellular patterns of Rac1 activity are supported by gradients of signaling nanodomains of heterogeneous molecular composition, which presumably act as discrete signaling platforms.


Glycolipid markers of astrocytomas and oligodendrogliomas.
Neutral glycolipids and gangliosides were analyzed in 149 astrocytomas (A), 46 oligodendrogliomas (O), and 21 oligoastrocytomas (OA) to determine if specific glycolipids correlate with histologic diagnosis and grade. Positivity for asialoGM1 (GA1) and negativity for paragloboside by immuno-TLC correlated with histological diagnosis of O and OA, whereas the reverse pattern correlated with A. High levels (over 5 microg hexose per mg dry weight) of CMH generally correlated with an O component, but the association was not as strong as for either GA1 presence or paragloboside absence. Pilocytic astrocytomas and pleomorphic xanthoastrocytomas had high proportions (> 15%) of globoside, low ratios (< 0.5) of GD1a: GD1b, and identifiable ceramide trihexoside (CTH). Three gangliosides of the 1b pathway were progressively lost with increasing grade of A, but a similar correlation with grade was not seen in O or OA. A high proportion of cases expressing sialosylparagloboside (3'LM1; 6'LM1) were grade 4 A. Glycolipids are synthesized by glycosyltransferases that add specific sugars to the nascent oligosaccharide. Correlation of specific glycolipids with histological diagnoses and grades indicate that these tumor types express specific patterns of glycosyltransferases, several of which have been cloned. It is possible that critical genes coding for these enzymes are deleted, overexpressed, or mutated in certain tumor types and grades, thus leading to the patterns of glycolipids that we found to be associated with these tumors.


Thrombin rapidly induces protein kinase D phosphorylation, and protein kinase C delta mediates the activation.
Thrombin plays a critical role in hemostasis, thrombosis, and inflammation. However, the responsible intracellular signaling pathways triggered by thrombin are still not well defined. We report here that thrombin rapidly and transiently induces activation of protein kinase D (PKD) in aortic smooth muscle cells. Our data demonstrate that protein kinase C (PKC) inhibitors completely block thrombin-induced PKD activation, suggesting that thrombin induces PKD activation via a PKC-dependent pathway. Furthermore, our results show that thrombin rapidly induces PKC delta phosphorylation and that the PKC delta-specific inhibitor rottlerin blocks thrombin-induced PKD activation, suggesting that PKC delta mediates the thrombin-induced PKD activation. Using dominant negative approaches, we demonstrated that expression of a dominant negative PKC delta inhibits the phosphorylation and activation of PKD induced by thrombin, whereas neither PKC epsilon nor PKC zeta affects thrombin-induced PKD activation. In addition, our results of co-immunoprecipitation assays showed that PKD forms a complex with PKC delta in smooth muscle cells. Taken together, the findings of the present study demonstrate that thrombin induces activation of PKD and reveal a novel role of PKC delta in mediating thrombin-induced PKD activation in vascular smooth muscle cells.


Evaluation of a multi-atlas based method for segmentation of cardiac CTA data: a large-scale, multicenter, and multivendor study.
Computed tomography angiography (CTA) is increasingly used for the diagnosis of coronary artery disease (CAD). However, CTA is not commonly used for the assessment of ventricular and atrial function, although functional information extracted from CTA data is expected to improve the diagnostic value of the examination. In clinical practice, the extraction of ventricular and atrial functional information, such as stroke volume and ejection fraction, requires accurate delineation of cardiac chambers. In this paper, we investigated the accuracy and robustness of cardiac chamber delineation using a multiatlas based segmentation method on multicenter and multivendor CTA data. A fully automatic multiatlas based method for segmenting the whole heart (i.e., the outer surface of the pericardium) and cardiac chambers from CTA data is presented and evaluated. In the segmentation approach, eight atlas images are registered to a new patient's CTA scan. The eight corresponding manually labeled images are then propagated and combined using a per voxel majority voting procedure, to obtain a cardiac segmentation. The method was evaluated on a multicenter/multivendor database, consisting of (1) a set of 1380 Siemens scans from 795 patients and (2) a set of 60 multivendor scans (Siemens, Philips, and GE) from different patients, acquired in six different institutions worldwide. A leave-one-out 3D quantitative validation was carried out on the eight atlas images; we obtained a mean surface-to-surface error of 0.94 +/- 1.12 mm and an average Dice coefficient of 0.93 was achieved. A 2D quantitative evaluation was performed on the 60 multivendor data sets. Here, we observed a mean surface-to-surface error of 1.26 +/- 1.25 mm and an average Dice coefficient of 0.91 was achieved. In addition to this quantitative evaluation, a large-scale 2D and 3D qualitative evaluation was performed on 1380 and 140 images, respectively. Experts evaluated that 49% of the 1380 images were very accurately segmented (below 1 mm error) and that 29% were accurately segmented (error between 1 and 3 mm), which demonstrates the robustness of the presented method. A fully automatic method for whole heart and cardiac chamber segmentation was presented and evaluated using multicenter/multivendor CTA data. The accuracy and robustness of the method were demonstrated by successfully applying the method to 1420 multicenter/ multivendor data sets.


Importance of hydrogen bonding for efficiency and specificity of the human mitochondrial DNA polymerase.
To assess the contribution to discrimination afforded by base pair hydrogen bonding during DNA replication by the human mitochondrial DNA polymerase, we examined nucleoside mimics lacking hydrogen bond forming capability but retaining the overall steric shape of the natural nucleotide. We employed oligonucleotide templates containing either a deoxyadenosine shape mimic (dQ) or a deoxythymidine shape mimic (dF). Additionally, the nucleoside triphosphate analogs difluorotoluene deoxynucleoside triphosphate, 9-methyl-1-H-imidazo[(4,5)-b]pyridine deoxyribose triphosphate, and 4-methylbenzimidazole deoxyribose triphosphate (dZTP; another dATP shape mimic) were assayed. We used pre-steady state methods to determine the kinetic parameters governing nucleotide incorporation, k(pol) and K(d). In general, the loss of hydrogen bonding potential led to 2-3 kcal/mol reduction in ground state binding free energy, whereas effects on the maximum rate of polymerization were quite variable, ranging from negligible (dATP:dF) to nearly 4 kcal/mol (dZTP:dT). Although we observed only a 46-fold reduction in discrimination when dF was present in the template, there was a complete elimination of discrimination when dQ was present in the template. Our data with dF indicate that hydrogen bonding contributes 2.2 kcal/mol toward the efficiency of incorporation, whereas data with dQ (which may overestimate the effect due to poor steric mimicry) suggest a contribution of up to 6.8 kcal/mol. Taken together, the data suggest that sterics are necessary but not sufficient to achieve optimal efficiency and fidelity for DNA polymerase. Base pair hydrogen bonding contributes at least a third of the energy underlying nucleoside incorporation efficiency and specificity.


Maintaining methylation activities during salt stress. The involvement of adenosine kinase.
Synthesis of the compatible osmolyte Gly betaine is increased in salt-stressed spinach (Spinacia oleracea). Gly betaine arises by oxidation of choline from phosphocholine. Phosphocholine is synthesized in the cytosol by three successive S-adenosyl-Met-dependent N-methylations of phosphoethanolamine. With each transmethylation, a molecule of S-adenosylhomo-Cys (SAH) is produced, a potent inhibitor of S-adenosyl-Met-dependent methyltransferases. We examined two enzymes involved in SAH metabolism: SAH hydrolase (SAHH) catabolizes SAH to adenosine plus homo-Cys and adenosine kinase (ADK) converts adenosine to adenosine monophosphate. In vitro SAHH and ADK activities increased incrementally in extracts from leaves of spinach plants subjected to successively higher levels of salt stress and these changes reflected increased levels of SAHH and ADK protein and transcripts. Another Gly betaine accumulator, sugar beet (Beta vulgaris), also showed salt-responsive increases in SAHH and ADK activities and protein whereas tobacco (Nicotiana tabacum) and canola (Brassica napus), which do not accumulate Gly betaine, did not show comparable changes in these enzymes. In spinach, subcellular localization positions SAHH and ADK in the cytosol with the phospho-base N-methyltransferase activities. Because SAHH activity is inhibited by its products, we propose that ADK is not a stress-responsive enzyme per se, but plays a pivotal role in sustaining transmethylation reactions in general by serving as a coarse metabolic control to reduce the cellular concentration of free adenosine. In support of this model, we grew Arabidopsis under a short-day photoperiod that promotes secondary cell wall development and found both ADK activity and transcript levels to increase severalfold.


Human Sperm Quality and Metal Toxicants: Protective Effects of some Flavonoids on Male Reproductive Function.
Metals can cause male infertility through affection of spermatogenesis and sperm quality. Strong evidences confirm that male infertility in metal-exposed humans is mediated via various mechanisms such as production of reactive oxygen species (ROS). Flavonoids have antioxidant and metal chelating properties which make them suitable candidates for neutralizing adverse effects of metals on semen quality. In the current study, we have evaluated the effects of five types of flavonoids (rutin, naringin, kaempferol, quercetin, and catechin) on recovery of sperm motility and prevention of membrane oxidative damage from aluminum chloride (AlCl3), cadmium chloride (CdCl2), and lead chloride (PbCl4). In this experimental study, motility and lipid peroxidation of metalexposed sperm was investigated in the presence of different concentrations of five kinds of flavonoids. Malondialdehyde (MDA) production was assessed as a lipid peroxidation marker. Aluminum chloride (AlCl3), cadmium chloride (CdCl2), and lead chloride (PbCl4) diminished sperm motility. Treatment of metal-exposed sperm with rutin, naringin, and kaempferol attenuated the negative effects of the metals on sperm motility. Quercetin and catechin decreased the motility of metal-exposed sperm. Based on the MDA production results, only AlCl3 significantly induced lipid peroxidation. Treatment with rutin, naringin, and kaempferol significantly decreased MDA production.


Endothelial HIF-2 mediates protection and recovery from ischemic kidney injury.
The hypoxia-inducible transcription factors HIF-1 and HIF-2 mediate key cellular adaptions to hypoxia and contribute to renal homeostasis and pathophysiology; however, little is known about the cell type-specific functions of HIF-1 and HIF-2 in response to ischemic kidney injury. Here, we used a genetic approach to specifically dissect the roles of endothelial HIF-1 and HIF-2 in murine models of hypoxic kidney injury induced by ischemia reperfusion or ureteral obstruction. In both models, inactivation of endothelial HIF increased injury-associated renal inflammation and fibrosis. Specifically, inactivation of endothelial HIF-2α, but not endothelial HIF-1α, resulted in increased expression of renal injury markers and inflammatory cell infiltration in the postischemic kidney, which was reversed by blockade of vascular cell adhesion molecule-1 (VCAM1) and very late antigen-4 (VLA4) using monoclonal antibodies. In contrast, pharmacologic or genetic activation of HIF via HIF prolyl-hydroxylase inhibition protected wild-type animals from ischemic kidney injury and inflammation; however, these same protective effects were not observed in HIF prolyl-hydroxylase inhibitor-treated animals lacking endothelial HIF-2. Taken together, our data indicate that endothelial HIF-2 protects from hypoxia-induced renal damage and represents a potential therapeutic target for renoprotection and prevention of fibrosis following acute ischemic injury.


Grasping the Müller-Lyer illusion: the contributions of vision for perception in action.
The present study examines the contributions of vision for perception processes in action. To this end, the influence of allocentric information on different action components (i.e., the selection of an appropriate mode of action, the pre-planning and online control of movement kinematics) is assessed. Participants (n=10) were presented with a shaft of various lengths (i.e., 13-20 cm) that was embedded in a Müller-Lyer figure. Picking up the shaft would, dependent on its length, either require a one- or a two-handed grasp. In different conditions participants were instructed to give a verbal judgement on the size of the shaft (VSJ); to make a manual estimation of the shaft's length (MLE); to indicate verbally whether they would grasp the shaft with one- or two hands (VAE); to actually grasp the shaft (G). We found that the Müller-Lyer figure affected the choice between using a one- or two-handed grasp, both when the participants actually grasped (G) the object and when they made a verbal estimation (VAE). The illusionary bias was of a similar magnitude as the one found in the verbal (VSJ) and manual perception task (MLE). The illusion had only a minor influence on the movement kinematics, and appears to be restricted to participants in which the grasping condition was immediately preceded by the VSJ-condition. We conclude that vision for perception contributes to the selection of an action mode, and that its contributions beyond that stage are dependent on the particular (experimental) circumstances.


The Black cells phenotype is caused by a point mutation in the Drosophila pro-phenoloxidase 1 gene that triggers melanization and hematopoietic defects.
Melanization contributes to arthropod-specific innate immunity through deposition of melanin at wound sites or around parasites, with concomitant release of microbicidal reactive oxygen species. Melanization requires sequential activation of proteolytic enzymes in the hemolymph, including the final enzyme pro-phenoloxidase. Black cells (Bc) is a mutation causing spontaneous melanization of Drosophila crystal cells, a hemocyte cell type producing phenoloxidases. Bc individuals exhibit circulating black spots but fail to melanize upon injury. Although Bc is widely used as a loss-of-function mutant of phenoloxidases, the mutation causing Bc remained unknown. Here, we identified a single point mutation in the pro-phenoloxidase 1 (PPO1) gene of Bc flies causing an Alanine to Valine change in the C-terminal domain of PPO1, predicted to affect the conformation of the N-terminal pro-domain cleavage site at a distance and causing uncontrolled catalytic activity. Genomic insertion of a PPO1(A480V) transgene phenocopies Black cells, proving that A480V is indeed the causal mutation of the historical Bc phenotype.


Prediction of retinopathy at 20 years after onset in younger-onset type 1 diabetes using mean metabolic memory-free HbA1c values: the importance of using HbA1c data of total, not partial, diabetes duration.
Metabolic memory, in which past hyperglycemia could affect future retinopathy, is a potential issue in studying the relationship between glycemia and retinopathy. We examined retrospectively if mean "metabolic memory-free" glycosylated hemoglobin A1C (HbA1c) values covering total diabetes duration could predict retinopathy in younger-onset type 1 diabetes mellitus (T1DM). Inclusion criteria were T1DM onset before age 30 years, first visit to our center between 1988 and 1990 soon after onset, continuous HbA1c data for 20 years, and a 20-year follow-up retinopathy examination. Retinopathy predictive capabilities of HbA1c variables were examined. Of 15 subjects, 5 were retinopathy-positive and 10 were retinopathy-negative at the 20-year follow-up. Mean metabolic memory-free HbA1c values for the 20 years showed a substantial capacity to predict retinopathy at 20 years. The longer the period without HbA1c data following onset in simulation, the less accurate the prediction. HbA1c values may predict retinopathy better if metabolic memory-free data are used.


Regional variation of Guillain-Barré syndrome.
Guillain-Barré syndrome is a heterogeneous disorder regarding the clinical presentation, electrophysiological subtype and outcome. Previous single country reports indicate that Guillain-Barré syndrome may differ among regions, but no systematic comparative studies have been conducted. Comparative studies are required to identify factors determining disease susceptibility, variation and prognosis, and to improve diagnostic criteria. The International Guillain-Barré Syndrome Outcome Study is a prospective, observational cohort study including all patients within the diagnostic spectrum, aiming to describe the heterogeneity of Guillain-Barré syndrome worldwide. The current study was based on the first 1000 inclusions with a follow-up of at least 1 year and confirmed the variation in clinical presentation, course and outcome between patients. The full clinical spectrum of Guillain-Barré syndrome was observed in patients from all countries participating in the International Guillain-Barré Syndrome Outcome Study, but the frequency of variants differed between regions. We compared three regions based on geography, income and previous reports of Guillain-Barré syndrome subtypes: 'Europe/Americas', 'Asia' (without Bangladesh), and 'Bangladesh'. We excluded 75 (8%) patients because of alternative diagnoses, protocol violations, or missing data. The predominant clinical variant was sensorimotor in Europe/Americas (n = 387/562, 69%) and Asia (n = 27/63, 43%), and pure motor in Bangladesh (n = 74/107, 69%). Miller Fisher syndrome and Miller Fisher-Guillain-Barré overlap syndrome were more common in Asia (n = 14/63, 22%) than in the other two regions (Europe/Americas: n = 64/562, 11%; Bangladesh: n = 1/107, 1%) (P < 0.001). The predominant electrophysiological subtype was demyelinating in all regions (Europe/Americas: n = 312/573, 55%; Asia: n = 29/65, 45%; Bangladesh: n = 38/94, 40%). The axonal subtype occurred more often in Bangladesh (n = 34/94, 36%) than in Europe/Americas (n = 33/573, 6%) and other Asian countries (n = 4/65, 6%) (P < 0.001). In all regions, patients with the axonal subtype were younger, had fewer sensory deficits, and showed a trend towards poorer recovery compared to patients with the demyelinating subtype. The proportion of patients able to walk unaided after 1 year varied between Asia (n = 31/34, 91%), Europe/Americas (n = 334/404, 83%) and Bangladesh (n = 67/97, 69%) (P = 0.003). A similar variation was seen for mortality, being higher in Bangladesh (n = 19/114, 17%) than in Europe/Americas (n = 23/486, 5%) and Asia (n = 1/45, 2%) (P < 0.001). This study showed that factors related to geography have a major influence on clinical phenotype, disease severity, electrophysiological subtype, and outcome of Guillain-Barré syndrome.


Targeting TEAD/YAP-transcription-dependent necrosis, TRIAD, ameliorates Huntington's disease pathology.
Neuronal cell death in neurodegenerative diseases is not fully understood. Here we report that mutant huntingtin (Htt), a causative gene product of Huntington's diseases (HD) selectively induces a new form of necrotic cell death, in which endoplasmic reticulum (ER) enlarges and cell body asymmetrically balloons and finally ruptures. Pharmacological and genetic analyses revealed that the necrotic cell death is distinct from the RIP1/3 pathway-dependent necroptosis, but mediated by a functional deficiency of TEAD/YAP-dependent transcription. In addition, we revealed that a cell cycle regulator, Plk1, switches the balance between TEAD/YAP-dependent necrosis and p73/YAP-dependent apoptosis by shifting the interaction partner of YAP from TEAD to p73 through YAP phosphorylation at Thr77. In vivo ER imaging with two-photon microscopy detects similar ER enlargement, and viral vector-mediated delivery of YAP as well as chemical inhibitors of the Hippo pathway such as S1P recover the ER instability and necrosis in HD model mice. Intriguingly S1P completely stops the decline of motor function of HD model mice even after the onset of symptom. Collectively, we suggest approaches targeting the signalling pathway of TEAD/YAP-transcription-dependent necrosis (TRIAD) could lead to a therapeutic development against HD.


Somatic GPR101 Duplication Causing X-Linked Acrogigantism (XLAG)-Diagnosis and Management.
Recent reports have proposed that sporadic or familial germline Xq26.3 microduplications involving the GPR101 gene are associated with early-onset X-linked acrogigantism (XLAG) with a female preponderance. A 4-year-old boy presented with rapid growth over the previous 2 years. He complained of sporadic headaches and had coarse facial features. His height Z-score was +4.89, and weight Z-score was +5.57. Laboratory testing revealed elevated serum prolactin (185 μg/L; normal, <18 μg/L), IGF-1 (745 μg/L; normal, 64-369 μg/L), and fasting GH > 35.0 μg/L. Magnetic resonance imaging demonstrated a homogenous bulky pituitary gland (18 × 15 × 13 mm) without obvious adenoma. A pituitary biopsy showed hyperplastic pituitary tissue with enlarged cords of GH and prolactin cells. Germline PRKAR1A, MEN1, AIP, DICER1, CDKN1B, and somatic GNAS mutations were negative. Medical management was challenging until institution of continuous sc infusion of short-acting octreotide combined with sc pegvisomant and oral cabergoline. The patient remains well controlled with minimal side effects 7 years after presentation. His phenotype suggested XLAG, but his peripheral leukocyte-, saliva-, and buccal cell-derived DNA tested negative for microduplication in Xq26.3 or GPR101. However, DNA isolated from the pituitary tissue and forearm skin showed duplicated dosage of GPR101, suggesting that he is mosaic for this genetic abnormality. Our patient is the first to be described with somatic microduplication leading to typical XLAG phenotype. This patient demonstrates that a negative test for Xq26.3 microduplication or GPR101 duplication on peripheral blood DNA does not exclude the diagnosis of XLAG because it can result from a mosaic mutation affecting the pituitary.


Islands and streams: clusters and gene flow in wild barley populations from the Levant.
The domestication of plants frequently results in a high level of genetic differentiation between domesticated plants and their wild progenitors. This process is counteracted by gene flow between wild and domesticated plants because they are usually able to inter-mate and to exchange genes. We investigated the extent of gene flow between wild barley Hordeum spontaneum and cultivated barley Hordeum vulgare, and its effect on population structure in wild barley by analysing a collection of 896 wild barley accessions (Barley1K) from Israel and all available Israeli H. vulgare accessions from the Israeli gene bank. We compared the performance of simple sequence repeats (SSR) and single nucleotide polymorphisms (SNP) marker data genotyped over a core collection in estimating population parameters. Estimates of gene flow rates with SSR markers indicated a high level of introgression from cultivated barley into wild barley. After removing accessions from the wild barley sample that were recently admixed with cultivated barley, the inference of population structure improved significantly. Both SSR and SNP markers showed that the genetic population structure of wild barley in Israel corresponds to the three major ecogeographic regions: the coast, the Mediterranean north and the deserts in the Jordan valley and the South. Gene flow rates were estimated to be higher from north to south than in the opposite direction. As has been observed in other crop species, there is a significant exchange of alleles between the wild species and domesticated varieties that needs to be accounted for in the population genetic analysis of domestication.


Principles of Ice-Free Cryopreservation by Vitrification.
Vitrification is an alternative to cryopreservation by freezing that enables hydrated living cells to be cooled to cryogenic temperatures in the absence of ice. Vitrification simplifies and frequently improves cryopreservation because it eliminates mechanical injury from ice, eliminates the need to find optimal cooling and warming rates, eliminates the importance of differing optimal cooling and warming rates for cells in mixed cell type populations, eliminates the need to find a frequently imperfect compromise between solution effects injury and intracellular ice formation, and can enable chilling injury to be "outrun" by using rapid cooling without a risk of intracellular ice formation. On the other hand, vitrification requires much higher concentrations of cryoprotectants than cryopreservation by freezing, which introduces greater risks of both osmotic damage and cryoprotectant toxicity. Fortunately, a large number of remedies for the latter problem have been discovered over the past 35 years, and osmotic damage can in most cases be eliminated or adequately controlled by paying careful attention to cryoprotectant introduction and washout techniques. Vitrification therefore has the potential to enable the superior and convenient cryopreservation of a wide range of biological systems (including molecules, cells, tissues, organs, and even some whole organisms), and it is also increasingly recognized as a successful strategy for surviving harsh environmental conditions in nature. But the potential of vitrification is sometimes limited by an insufficient understanding of the complex physical and biological principles involved, and therefore a better understanding may not only help to improve present outcomes but may also point the way to new strategies that may be yet more successful in the future. This chapter accordingly describes the basic principles of vitrification and indicates the broad potential biological relevance of this alternative method of cryopreservation.


A Pr3+ doping strategy for simultaneously optimizing the size and near infrared persistent luminescence of ZGGO:Cr3+ nanoparticles for potential bio-imaging.
Spinel-phase Zn2Ga2.98-xGe0.75O8:Cr0.020,Prx (ZGGO:Cr3+,Pr3+) near infrared (NIR) persistent luminescence nanoparticles (PLNPs) with different amounts of Pr3+ dopant were prepared by a hydrothermal method in combination with a subsequent annealing in a vacuum. For these nanoparticles, the averaged particle size decreases from 64 to 37 nm with increasing Pr3+ doping concentration from 0 to 0.025 and Cr3+ and Pr3+ ions are uniformly doped into the interior and surface of a single nanoparticle. It can be found that Pr3+ doping leads to the appearance of more anti-site pairs () around distorted octahedral Cr3+ ions and enhanced NIR emissions around 697 nm, which originate from the 2E(2G) → 4A2(4F) and 4T2(4F) → 4A2(4F) transitions of the interior and surface Cr3+ ions in the nanoparticles. In particular, for the interior Cr3+ ions in the Pr3+ doped nanoparticles, the enhanced NIR luminescence can be attributed to the suppressed energy transfer of the excited electrons from the 4T2(4F) level to the trap level related to anti-site pairs () around the distorted octahedral Cr3+ ions. Our results suggest that Pr3+ doped ZGGO:Cr3+ PLNPs have potential applications for bio-imaging.