Flavobacterium psychrophilum, invasion into and shedding by rainbow trout Oncorhynchus mykiss.
The infection route of Flavobacterium psychrophilum into rainbow trout Oncorhynchus mykiss was studied using bath and cohabitation challenges as well as oral challenge with live feed as a vector. Additionally, the number of bacterial cells shed by infected fish into the surrounding water was determined in the cohabitation experiment and in challenge experiments at 3 different water temperatures. The experiments showed that skin and skin mucus abrasion dramatically enhanced the invasion of F. psychrophilum into the affected fish in bath and cohabitation challenges. Disruption of the skin is discussed as an important invasion route for F. psychrophilum into the fish. The shedding rate of F. psychrophilum by infected fish was associated with water temperature and the mortality of the infected fish. High numbers of F. psychrophilum cells were released into the water by dead rainbow trout during a long time period compared to the numbers of cells shed by live fish. The results emphasise the importance of removing dead and moribund fish from rearing tanks in order to diminish the infection pressure against uninfected fish in commercial fish farms. In immunohistochemical examinations of organs and tissues of orally infected fish, F. psychrophilum cells were detected in only 1 fish out of 31 studied. Mortality of the orally challenged fish was not observed in the experiment.


Efficacy and Safety of Serotonin Receptor Ligands in the Treatment of Irritable Bowel Syndrome: A Review.
Irritable bowel syndrome (IBS) is a chronic, recurrent bowel disorder with an unknown etiology, which is most likely multifactorial. Increased mucosal permeability, visceral hypersensitivity and activation status of intestinal mucosal immune cells cause changes in gastrointestinal (GI) motility, secretion and sensation observed in the course of IBS. Permanent, cumbersome symptoms, such as diarrhea, constipation and abdominal pain greatly lower the quality of life of IBS patients. On this basis, according to the Rome IV criteria, different forms of IBS can be distinguished. This article focuses on the role of serotonin system in the pathophysiology of IBS as a potential therapeutic target. We shortly describe several molecules, associated with serotonin receptors, mainly 5-HT3 receptor antagonists and 5-HT4 receptor agonists, that are used in the treatment of motility disorders and visceral pain in IBS patients. We summarize the findings obtained in the clinical trials and elaborate on the safety of the serotonin ligands. Although the majority of serotonin receptor ligands relieve global symptoms, there are also some adverse effects, which can be dangerous for patients. We postulate that currently, among all serotonin-targeting compounds, ramosetron is the best treatment option for IBS-D patients, due to its exceptional efficacy in both genders as well as good tolerability. Whereas, tegaserod is highly recommended for IBS-C sufferers. Nevertheless, numerous studies on the new serotonin receptor ligands are conducted to ensure the delivery of novel compounds with improved efficacy and safety profiles.


Effects of PAF-acether and structural analogues on platelet activation and bronchoconstriction in guinea-pigs.
PAF-acether (platelet-activating factor) (1-O-alkyl-2-acetyl-sn-glycerol-3-phosphorylcholine) induces platelet-dependent bronchoconstriction in the guinea-pig which correlates with its in vivo thrombocytopenic effect. We investigated the influence of modifications of the polar head group in position 3 of the glycerol skeleton of PAF-acether on guinea-pig platelet activation and bronchoconstriction. PAF-acether itself induced concentration-dependent platelet activation (EC50 for aggregation = 0.41 nM and EC20 for secretion of ATP = 0.56 nM). The 3-phosphoryl-N-methyl-morpholino ethanol analogue was slightly more active than PAF-acether and the 3-phosphoryl-N-methyl-piperidinium ethanol, 3-phopshoryl-(N-methyl-piperidino-3') methanol and 3-phosphoryl-(N-methyl-hydroxy-4') piperidine analogues were equieffective to PAF-acether in activating platelets. The 3-phosphoryl-piperidino ethanol analogue was 8 times less active than PAF-acether; the 3-phosphoryl-morpholino ethanol analogue and the 1-O-octadecyl-2-O-acetyl-3-O-[trimethyl-ammonio)-propyl) glycerol were inactive up to 1 microM. Our data show that the choline head group is not a compulsory requirement for activity. When injected i.v. to the propranolol-treated guinea-pigs, the platelet-activating analogues also induced bronchoconstriction. Two PAF-acether antagonists, compounds 48740 RP and BN 52021, inhibited PAF-acether-induced platelet activation when added to PRP at the final concentration of 0.1 mM (aggregation inhibited by 91 +/- 4 and by 94 +/- 3% respect.; secretion inhibited by 80 +/- 12 and 79 +/- 10% respectively, mean +/- S.E.M., n = 4). Both antagonists also suppressed platelet activation and in vivo bronchoconstriction, thrombocytopenia, leukopenia and hypotension induced by PAF-acether and the various analogues. Our results indicate that PAF-acether and the analogues studied trigger platelet activation and the consequent bronchoconstriction through mechanisms which share sensitivity to same antagonists.


Discovery of the First Selective, Nonpeptidic Orexin 2 Receptor Agonists.
In this issue, Nagase and colleagues report the discovery of the first selective nonpeptidic orexin 2 receptor (OX2R) agonists. The discovery of these OX2R selective agonists opens up new avenues for therapies related to the activation of the orexin system, especially with respect to the treatment of sleep disorders such as narcolepsy.


CD16+NK-92 and anti-CD123 monoclonal antibody prolongs survival in primary human acute myeloid leukemia xenografted mice.
Patients with acute myeloid leukemia (AML) often relapse after initial therapy because of persistence of leukemic stem cells that frequently express the IL-3 receptor alpha chain CD123. Natural killer (NK) cell-based therapeutic strategies for AML show promise and we explore the NK cell lines, NK-92 and CD16+ NK-92, as a treatment for AML. NK-92 has been tested in phase I clinical trials with minimal toxicity; irradiation prior to infusion prevents risk of engraftment. The CD16 negative NK-92 parental line was genetically modified to express the high affinity Fc gamma receptor, enabling antibody-dependent cell-mediated cytotoxicity, which we utilized in combination with an anti-CD123 antibody to target leukemic stem cells. NK-92 was preferentially cytotoxic against leukemic stem and progenitor cells compared with bulk leukemia in in vitro assays, while CD16+ NK-92 in combination with an anti-CD123 mAb mediated antibody-dependent cell-mediated cytotoxicity against CD123+ leukemic targets. Furthermore, NK-92 infusions (with or without prior irradiation) improved survival in a primary AML xenograft model. Mice xenografted with primary human AML cells had a superior survival when treated with irradiated CD16+NK-92 cells and an anti-CD123 monoclonal antibody (7G3) versus treatment with irradiated CD16+NK-92 cells combined with an isotype control antibody. In this proof-of-principle study, we show for the first time that a CD16+NK-92 cell line combined with an antibody that targets a leukemic stem cell antigen can lead to improved survival in a relevant pre-clinical model of AML.


Immunohistochemical and molecular analysis of human melanomas for expression of the human cancer-testis antigens NY-ESO-1 and LAGE-1.
NY-ESO-1 and LAGE-1 are homologous cancer-testis antigens, which are expressed in many different cancers. It is essential to type tumors accurately to assess patient suitability for clinical trials which target these. This study evaluates typing strategies used to distinguish these two homologous but distinct antigens and to characterize and quantitate expression of each in clinical samples. We typed 120 malignant melanomas for the expression of NY-ESO-1 and LAGE-1 with immunohistochemistry, reverse transcription-PCR (RT-PCR), and quantitative real-time (qRT-PCR), which was also used to explore the relationship between NY-ESO-1 and LAGE expression. The two monoclonal antibodies ES121 and E978 had very similar immunohistochemistry reactivities. Both were specific for NY-ESO-1 because neither bound to homologous LAGE-1 peptides despite 84% overall amino acid homology. Of 120 melanomas tested by immunohistochemistry, NY-ESO-1 was expressed in >50% of cells in 23 melanomas (19%), between 11 and 50% cells in 15 (12.5%), <11% cells in 16 (13.5%), and negative in 66 (55%). Although specific for both antigens, the PCR methods did not provide this information about microheterogeneity. Polymorphisms in the LAGE-1 gene resulted in false negative LAGE-1 typing by qRT-PCR by inhibiting binding of oligonucleotide primers, thereby showing the exquisite specificity of qRT-PCR as a typing method. For NY-ESO-1 typing, immunohistochemistry compared favorably with the RT-PCR, with the added advantage of being able to characterize heterogeneity of antigen expression. Because neither mAb bound LAGE and because there was no coordinate expression LAGE and NY-ESO-1, separate typing for each is required.


Obesity as a Risk Factor for Prostatic Enlargement: A Retrospective Cohort Study in Korea.
We aimed to evaluate obesity, a risk factor of metabolic syndrome, and its association with prostatic enlargement in a retrospective cohort in Korea. Baseline data were obtained from the Korean Genome and Epidemiology Study on Atherosclerosis Risk of Rural Areas in the Korean General Population (KoGES-ARIRANG). Between March 2015 and November 2015, 2,127 male participants of KoGES-ARIRANG were invited to the Korean Prostate Health Council Screening Program, and 602 participants underwent urological examination, including serum prostate specific antigen measurement and transrectal ultrasonography, and completed the International Prostate Symptom Score questionnaire. The data for 571 participants were analyzed, after excluding 31 men who had a history of prostatic disease or testosterone replacement, or had undergone a prior prostatic surgery or procedure. Among components of metabolic syndrome, waist circumference had a statistically significant linear correlation with incremental increases in prostate volume (B=0.181, P=0.004). Abdominal obesity as determined by anthropometric measures including body mass index (odds ratio [OR], 1.205; 95% confidence interval [CI], 1.088-1.336), waist circumference (OR, 1.073; 95% CI, 1.032-1.115), body fat (OR, 1.126; 95% CI, 1.056-1.202), and visceral fat composition (OR, 1.667; 95% CI, 1.246-2.232) was significantly associated with the presence of high-volume benign prostatic hyperplasia (BPH) (prostate volume≥ 40 mL). Furthermore, the highest quartile of serum leptin (OR, 3.541; 95% CI, 1.103-11.365) and adiponectin levels (OR, 0.315; 95% CI, 0.102-0.971) were significantly correlated with high-volume BPH compared to the lowest quartile of levels. Abdominal obesity and serum leptin level are positively associated with prostate growth, whereas serum adiponectin level is inversely associated with the presence of prostatic enlargement.


[Detection of mecA in strains with oxacillin and cefoxitin disk tests for detection of methicillin-resistant Staphylococcus].
Cinical and Laboratory Standards Institute (CLSI) recommends the use of cefoxitin disks instead of long-used oxacillin disks for screening methicillin-resistant isolates of staphylococci. The frequency of discrepant results and accuracy of the tests were evaluated by detecting mecA gene. A total of 3,123 Stapylococci isolates from patients in Severance Hospital were tested during September 2005 to August 2006 by the CLSI-recommended test using both cefoxitin and oxacillin disks. The mecA gene was detected by PCR and the oxacillin minimum inhibitory concentration (MIC) was determined by using agar dilution method for the isolates with discrepant tests. Among 1,915 S. aureus islolates tested, one isolate was resistant to oxacillin disk but susceptible to cefoxitin disk; the isolate did not have mecA gene. Another isolate susceptible to oxacillin but resistant to cefoxitin had mecA gene. Among 1,208 coagulase-negative staphylococcal isolates, 15 isolates were resistant to oxacillin disk but susceptible to cefoxitin disk; the isolates did not have mecA genes. Two isolates susceptible to oxacillin disk but resistant to cefoxitin disk had mecA genes. Among the 16 Staphylococcus isolates that did not have mecA gene, 15 isolates had the oxacillin MICs of </= 2 microg/mL and were considered as methicillin-susceptible, while 1 isolate with the MIC of 4 microg/mL was considered as methicillin-resistant. Overall, 1.9% of staphylococcal isolates showed discrepant results when the screening tests were performed by using oxacillin and cefoxitin disks. None of the isolates resistant to oxacillin disk but susceptible to cefoxitin disk had mecA gene. In conclusion, the cefoxitin disk test is more reliable than oxacillin disk test in screening methicillin-resistant staphylococcal isolates.


Estradiol: micrograms or milligrams.
Estrogen is used to induce puberty in peripubertal girls with hypogonadism. Although both synthetic and natural forms are available, along with different routes of administration, in the UK oral ethinyl estradiol and the low-dose oral contraceptive pill are commonly used as hormone replacement therapy for practical reasons. We present five peripubertal girls (aged 12.5-14.9 years) with hypogonadism (two with primary hypogonadism due to Turner syndrome and three with central (secondary) hypogonadism as part of multiple pituitary hormone deficiency) who for a variety of reasons have received milligram doses of estradiol (E2) in error for between 6 weeks and 6 months, instead of the expected microgram doses of ethinyl estradiol. Although there are no direct comparisons in peripubertal girls between synthetic and natural estrogens, all girls had vaginal bleeding whilst receiving the milligram doses and have ended up with reduced final heights, below the 9th centile in 1 and below the 2nd centile in 4. Whilst reduction in final height may be part of the underlying condition (especially in Turner syndrome) the two girls with height predictions performed prior to receiving the estrogen overdose have not achieved their predicted height. Estrogen is one of the few drugs which is available in both milligram and microgram formulations. Clinicians need to be alert to the possibility of patients receiving the wrong formulation and dosage in error. Girls with primary and secondary gonadal failure require assistance with pubertal induction.Although several different formulations and route of administration are available, for practical reasons, the majority of girls in the UK receive oral ethinyl estradiol.Estrogen preparations are available in both milligram and microgram formulations, with potential for receiving the wrong dose.Girls receiving milligram rather than microgram preparations all had vaginal bleeding and a short final height.


The lamin B receptor of the nuclear envelope inner membrane: a polytopic protein with eight potential transmembrane domains.
The lamin B receptor is a previously identified integral membrane protein in the nuclear envelope of turkey erythrocytes that associates with the nuclear intermediate filament protein lamin B (Worman, H. J., J. Yuan, G. Blobel, and S. D. Georgatos. 1988. Proc. Natl. Acad. Sci. USA. 85:8531-8534). In the present report, we use cell fractionation and antibodies against the lamin B receptor to localize it to an 8-M urea-extracted membrane fraction of chicken liver nuclei, supporting an inner nuclear membrane localization. We deduced the amino acid sequence of the chicken lamin B receptor from overlapping clones obtained by screening cDNA libraries with a probe generated by the polymerase chain reaction with primers based on the partial protein sequence of the isolated protein. The mature lamin B receptor has a calculated molecular mass of 73,375 D and eight segments of hydrophobic amino acids that could function as transmembrane domains as determined by hydropathy analysis. Preceding the first putative transmembrane segment is a highly charged 204-residue-long amino terminal region that contains two consensus sites for phosphorylation by protein kinase A. Since the lamin B receptor has been shown to be phosphorylated by protein kinase A in vitro and in vivo and this phosphorylation affects lamin B binding (Applebaum, J., G. Blobel, and S. D. Georgatos. 1990. J. Biol. Chem. 265:4181-4185), it is likely that this amino terminal region faces the nucleoplasm. The amino terminal region also contains three DNA-binding motifs that are found in gene regulatory proteins and histones, suggesting that the lamin B receptor may additionally play a role in gene regulation and/or chromatin organization.


The Scientist Citizen and the Citizen Scientist: Blurring the Lines.
The scientific enterprise satisfies the innate human urge to understand the world; these efforts have led to both improvements and dangers to society. The storied history of relationships between scientists and citizens suggests that the lines between these 2 sectors of society are often blurred. Here we discuss these relationships on the context of animal welfare. We briefly outline the history of animal welfare in research, and the entry of citizens into the discussion, leading to the Animal Welfare Act of 1966. The commitment of scientists to society, in this context, is the act of whistleblowing in research. As medical and life sciences technologies continue to expand at breathtaking rates, the landscape that both scientists and citizens must navigate increases in complexity. We discuss the responsibility of both the scientist and the citizen, as members of the voting public, in the face of the challenges of the future.


The rice genome encodes two vacuolar invertases with fructan exohydrolase activity but lacks the related fructan biosynthesis genes of the Pooideae.
* Fructans are believed to contribute to cold and drought tolerance in several plant families (Poaceae, Asparagaceae and Asteraceae), but it is not clear why the ability to accumulate these polymers is found in some genera (e.g. Triticum) but not in others (e.g. Oryza). * As fructan biosynthesis enzymes (FBEs) evolved from vacuolar invertases (VINs), we searched the rice genome sequence for genes related to both FBE and VIN genes of wheat and other members of the Pooideae. We compared them at the levels of exon-intron structure, protein sequence, and the enzymatic properties of recombinant proteins after expression in the yeast Pichia pastoris. * We found that rice possesses two VIN genes (OsVIN1 and OsVIN2) and no FBE genes. FBE genes appear to have arisen in the Pooideae by a series of gene duplications from an ancestor of wheat TaVIN3. Recombinant TaVIN2, OsVIN1 and OsVIN2 behaved as invertases with no FBE activity, but possessed high fructan exohydrolase activity, especially OsVIN1. * The engineering of fructan accumulation into rice for greater stress tolerance could founder on endogenous exohydrolases, but the fact that OsVIN1 transcripts are absent from peduncles of well watered and drought-stressed plants removes one potential obstacle to this endeavour.


Neurofibromatosis type 1 gene as a mutational target in a mismatch repair-deficient cell type.
DNA mismatch repair (MMR) is the process by which incorrectly paired DNA nucleotides are recognized and repaired. A germline mutation in one of the genes involved in the process may be responsible for a dominantly inherited cancer syndrome, hereditary nonpolyposis colon cancer. Cancer progression in predisposed individuals results from the somatic inactivation of the normal copy of the MMR gene, leading to a mutator phenotype affecting preferentially repeat sequences (microsatellite instability, MSI). Recently, we identified children with a constitutional deficiency of MMR activity attributable to a mutation in the h MLH1 gene. These children exhibited a constitutional genetic instability associated with clinical features of de novo neurofibromatosis type 1 (NF1) and early onset of extracolonic cancer. Based on these observations, we hypothesized that somatic NF1 gene mutation was a frequent and possibly early event in MMR-deficient cells. To test this hypothesis, we screened for NF1 mutations in cancer cells. Genetic alterations were identified in five out of ten tumor cell lines with MSI, whereas five MMR-proficient tumor cell lines expressed a wild-type NF1 gene. Somatic NF1 mutations were also detected in two primary tumors exhibiting an MSI phenotype. Finally, a 35-bp deletion in the murine Nf1 coding region was identified in mlh1-/- mouse embryonic fibroblasts. These observations demonstrate that the NF1 gene is a mutational target of MMR deficiency and suggest that its inactivation is an important step of the malignant progression of MMR-deficient cells.


Video game telemetry as a critical tool in the study of complex skill learning.
Cognitive science has long shown interest in expertise, in part because prediction and control of expert development would have immense practical value. Most studies in this area investigate expertise by comparing experts with novices. The reliance on contrastive samples in studies of human expertise only yields deep insight into development where differences are important throughout skill acquisition. This reliance may be pernicious where the predictive importance of variables is not constant across levels of expertise. Before the development of sophisticated machine learning tools for data mining larger samples, and indeed, before such samples were available, it was difficult to test the implicit assumption of static variable importance in expertise development. To investigate if this reliance may have imposed critical restrictions on the understanding of complex skill development, we adopted an alternative method, the online acquisition of telemetry data from a common daily activity for many: video gaming. Using measures of cognitive-motor, attentional, and perceptual processing extracted from game data from 3360 Real-Time Strategy players at 7 different levels of expertise, we identified 12 variables relevant to expertise. We show that the static variable importance assumption is false--the predictive importance of these variables shifted as the levels of expertise increased--and, at least in our dataset, that a contrastive approach would have been misleading. The finding that variable importance is not static across levels of expertise suggests that large, diverse datasets of sustained cognitive-motor performance are crucial for an understanding of expertise in real-world contexts. We also identify plausible cognitive markers of expertise.


Animal colour vision--behavioural tests and physiological concepts.
Over a century ago workers such as J. Lubbock and K. von Frisch developed behavioural criteria for establishing that non-human animals see colour. Many animals in most phyla have since then been shown to have colour vision. Colour is used for specific behaviours, such as phototaxis and object recognition, while other behaviours such as motion detection are colour blind. Having established the existence of colour vision, research focussed on the question of how many spectral types of photoreceptors are involved. Recently, data on photoreceptor spectral sensitivities have been combined with behavioural experiments and physiological models to study systematically the next logical question: 'what neural interactions underlie colour vision?' This review gives an overview of the methods used to study animal colour vision, and discusses how quantitative modelling can suggest how photoreceptor signals are combined and compared to allow for the discrimination of biologically relevant stimuli.


Search for new phenomena in final states with an energetic jet and large missing transverse momentum in pp collisions at [Formula: see text]TeV with the ATLAS detector.
Results of a search for new phenomena in final states with an energetic jet and large missing transverse momentum are reported. The search uses 20.3 fb[Formula: see text] of [Formula: see text] TeV data collected in 2012 with the ATLAS detector at the LHC. Events are required to have at least one jet with [Formula: see text] GeV and no leptons. Nine signal regions are considered with increasing missing transverse momentum requirements between [Formula: see text] GeV and [Formula: see text] GeV. Good agreement is observed between the number of events in data and Standard Model expectations. The results are translated into exclusion limits on models with either large extra spatial dimensions, pair production of weakly interacting dark matter candidates, or production of very light gravitinos in a gauge-mediated supersymmetric model. In addition, limits on the production of an invisibly decaying Higgs-like boson leading to similar topologies in the final state are presented.


Synthesis and evaluation of analogues of the glycinocin family of calcium-dependent antibiotics.
The glycinocins are a class of calcium-dependent, acidic cyclolipopeptide antibiotics that are structurally related to the clinically approved antibiotic daptomycin. In this article, we describe the synthesis of a small library of glycinocin analogues that differ by variation in the exocyclic fatty acyl substituent. The glycinocin analogues were screened against a panel of Gram-positive bacteria (as well as Gram-negative P. aeruginosa). These analogues exhibited similar calcium-dependent activity to the parent natural products against Gram-positive bacteria but showed no activity against P. aeruginosa. The length of the fatty acid was shown to be important for optimal biological activity, while the hybridisation at the α,β position and branching within the fatty acyl chain had only subtle effects on activity.


Parental height differences predict the need for an emergency caesarean section.
More than 30% of all pregnancies in the UK require some form of assistance at delivery, with one of the more severe forms of assistance being an emergency Caesarean section (ECS). Previously it has been shown that the likelihood of a delivery via ECS is positively associated with the birth weight and size of the newborn and negatively with maternal height. Paternal height affects skeletal growth and mass of the fetus, and thus might also affect pregnancy outcomes. We hypothesized that the effect of newborn birth weight on the risk of ECS would decrease with increasing maternal height. Similarly, we predicted that there would be an increase in ECS risk as a function of paternal height, but that this effect would be relative to maternal height (i.e., parental height differences). We used data from the Millennium Cohort Study: a large-scale survey (N = 18,819 births) with data on babies born and their parents from the United Kingdom surveyed 9 to 12-months after birth. We found that in primiparous women, both maternal height and parental height differences interacted with birth weight and predicted the likelihood of an ECS. When carrying a heavy newborn, the risk of ECS was more than doubled for short women (46.3%) compared to tall women (21.7%), in agreement with earlier findings. For women of average height carrying a heavy newborn while having a relatively short compared to tall partner reduced the risk by 6.7%. In conclusion, the size of the baby, the height of the mother and parental height differences affect the likelihood of an ECS in primiparous women.


Ethnobotanical authentication and identification of Khrog-sman (Lower Elevation Medicinal Plants) of Bhutan.
The Bhutanese form of g.so-ba-rig-pa medicine, which is a scholarly medical system, belongs to a larger system of medicinal corpus that spreads from Mongolia to India. It uses medicinal plants as a bulk ingredient but only 'Higher Elevation Medicinal Plants' have been botanically identified so far. Our study reports the botanical identification of 'Lower Elevation Medicinal Plants' and their ethnomedical uses. To botanically identify the 'Lower Elevation Medicinal Plants' used in Bhutanese traditional medicine. A five stage process was conducted which consisted of: (1) a survey of specialized ancient ethnomedical literatures (Pharmacopoeias and formularies); (2) freely listing the 'Lower Elevation Medicinal Plants' reported in the ancient Bhutanese medical texts and translating their ethnomedical uses in equivalent terms of English; (3) making field visits, collecting herbarium specimens and photographs, and spot identification of plants; (4) double blind testing of Bhutanese traditional medicine practitioners for authentication and standardization of Bhutanese g.so-ba-rig-pa names; (5) organising workshops for open forum discussions on medicinal plants involving Traditional Physicians and other professional participants of the relevant areas. We identified the botanical names of 113 'Lower Elevation Medicinal Plants' belonging to 68 families and 104 genera. Out of 113 medicinal plant species identified, 92 species are currently used in Bhutan and the remaining 21 species were not used in the current formulation, but described in the Bhutanese traditional medical texts. The identification of these 21 species was achieved both ethnomedically and botanically for the first time. Out of the 28 plant species that are currently imported from India, we found for the first time, even to the knowledge of Traditional Physicians, that 16 of them are actually growing abundantly in Bhutan. Among the plant parts collected, seeds were the most prominent followed by fruits and then roots. Our study identified 113 'Lower Elevation Medicinal Plants' out of which 92 of them are used daily in formulating 102 multi-ingredient prescription medicines in Bhutan.


Nicotinic acetylcholine receptor efficacy and pharmacological properties of 3-(substituted phenyl)-2β-substituted tropanes.
There is a need for different and better aids to tobacco product use cessation. Useful smoking cessation aids, bupropion (2) and varenicline (3), share some chemical features with 3-phenyltropanes (4), which have promise in cocaine dependence therapy. Here we report studies to generate and characterize pharmacodynamic features of 3-phenyltropane analogues. These studies extend our work on the multiple molecular target model for aids to smoking cessation. We identified several new 3-phenyltropane analogues that are superior to 2 in inhibition of dopamine, norepinephrine, and sometimes serotonin reuptake. All of these ligands also act as inhibitors of nicotinic acetylcholine receptor (nAChR) function with a selectivity profile that favors, like 2, inhibition of α3β4*-nAChR. Many of these ligands also block acute effects of nicotine-induced antinociception, locomotor activity, and hypothermia. Importantly, all except one of the analogues tested have better potencies in inhibition of nicotine conditioned place preference than 2. We have identified new compounds that have utility as research tools and possible promise for treatment of nicotine dependence.


NMDA receptor activation-responsive phosphoproteins in the developing optic tectum.
A front phosphorylation assay followed by two-dimensional gel electrophoresis was used to detect proteins in the tadpole optic tectum, the phosphorylation state of which is regulated by NMDA receptor activation. Five proteins with isoelectric points between 4 and 7 displayed marked increases in their phosphorylation state in response to application of 10 microM glutamate and 50 microM NMDA. This response was inhibited by 60 microM 2-amino-5-phosphopentanoic acid. These proteins are termed NMDA receptor activation-responsive phosphoproteins (NARPPs). Two NARPPs were identified as both in vitro and in vivo substrates for protein kinase C. Of these two NARPPs, one was located in the postsynaptic density (NARPP-50), and one was located in the nuclear fraction (NARPP-21). Phosphorylation of NARPP-21 was also induced by application of the metabotropic glutamate receptor agonist trans-(+/-)-1-amino-1,3-cyclopentanedicarboxylic acid (trans-ACPD) (100 microM). Phosphorylation of all NARPPs was eliminated by dantrolene, which inhibits release of calcium from intracellular stores. In adult tecta, only NARPP-21 and -50 were phosphorylation. Thus the phosphorylation state of most NARPPs is regulated differently when synaptic plasticity is low. Further characterization of NARPPs should lead to identification of second messenger systems involved in NMDA receptor signaling and developmental synaptic plasticity.


Identification using 1H NMR spectroscopy of slowly exchanging amide hydrogens of hen lysozyme in solution.
Resonances of over 20 of the most slowly exchanging amide hydrogens have been identified and assigned in the 1H NMR spectrum of hen lysozyme. This was achieved by combining information about spin-spin coupling patterns with nuclear Overhauser enhancement measurements. A computer-based search program was used to permit the assumptions and constraints in this procedure to be closely defined and to reveal possible ambiguities. In addition, experimental values of coupling constants were compared with values calculated on the basis of the torsion angles found in the crystal structure. The close correlation between these gave further confidence in the assignment procedures and provided information concerning the nature of fluctuations about the average solution structure. The very slowly exchanging amide hydrogens are largely buried in alpha-helical and beta-sheet regions of the protein structure.


The Association of Modifiable Breast Cancer Risk Factors and Somatic Genomic Alterations in Breast Tumors: The Cancer Genome Atlas Network.
The link between modifiable breast cancer risk factors and tumor genomic alterations remains largely unexplored. We evaluated the association of prediagnostic body mass index (BMI), cigarette smoking, and alcohol consumption with somatic copy number variation (SCNV), total somatic mutation burden (TSMB), seven single base substitution (SBS) signatures (SBS1, SBS2, SBS3, SBS5, SBS13, SBS29, and SBS30), and nine driver mutations (CDH1, GATA3, KMT2C, MAP2K4, MAP3K1, NCOR1, PIK3CA, RUNX1, and TP53) in a subset of The Cancer Genome Atlas (TCGA). Clinical and genomic data were retrieved from the TCGA database. Risk factor information was collected from four TCGA sites (n = 219 women), including BMI (1 year before diagnosis), cigarette smoking (smokers/nonsmokers), and alcohol consumption (current drinkers/nondrinkers). Multivariable regression analyses were conducted in all tumors and stratified according to estrogen receptor (ER) status. Increasing BMI was associated with increasing SCNV in all women (P = 0.039) and among women with ER- tumors (P = 0.031). Smokers had higher SCNV and TSMB versus nonsmokers (P < 0.05 all women). Alcohol drinkers had higher SCNV versus nondrinkers (P < 0.05 all women and among women with ER+ tumors). SBS3 (defective homologous recombination-based repair) was exclusively found in alcohol drinkers with ER- disease. GATA3 mutation was more likely to occur in women with higher BMI. No association was significant after multiple testing correction. This study provides preliminary evidence that BMI, cigarette smoking, and alcohol consumption can influence breast tumor biology, in particular, DNA alterations. This study demonstrates a link between modifiable breast cancer risk factors and tumor genomic alterations.


Effects of exposure to dust in swine confinement buildings--a working group report.
Pulmonary and other symptoms among workers in swine confinement buildings were evaluated by an international working group. In several studies in five different countries a total of about 2000 workers has been studied in clinical and epidemiologic investigations. Symptoms indicative of acute and chronic airway inflammation were widespread, as were systemic reactions of organic dust toxic syndrome. The base-line, and across workshift, pulmonary function changes were moderate. There was no evidence that antigen-antibody reactions are important in the pathogenesis. Longitudinal studies are recommended to establish the relationship between acute and chronic symptoms and end stage disease.


Effects of steroid pulse therapy on immunoglobulin-resistant Kawasaki disease.
The use of intravenous immunoglobulin (IVIG) is well established as an initial therapy for Kawasaki disease (KD), but treatment for IVIG-resistant KD remains uncertain To analyse the effects of intravenous methylprednisolone (IVMP) pulse therapy compared with additional IVIG in IVIG-resistant patients. IVMP was administered to patients with KD who had persistent or recurrent fever after a single dose of IVIG, at Juntendo University Hospital and affiliated medical institutions between May 2003 and March 2006. The effectiveness of the treatment and the incidence of coronary lesions in patients who received IVMP and those who received additional IVIG were retrospectively analysed and compared by chart review. 411 patients with KD were treated with a single dose of IVIG. Of the 63 IVIG-resistant patients, 44 were then given IVMP and 19 were given additional IVIG. Treatment was successful in 34 (77%) of the patients who received IVMP and 12 (63%) who received additional IVIG. Five of the 10 patients who did not respond to IVMP and two of the seven who did not respond to additional IVIG developed coronary artery aneurysms. Although fever initially resolved faster in the IVMP-resistant group, there was a delay in fever recurrence, which ultimately delayed the final resolution of fever. The findings suggest that IVMP is an effective additional treatment for IVIG-resistant KD. However, there was a tendency for fever to recur later in IVMP-resistant patients, which could potentially delay the therapeutic decision-making process.


Growth inhibition of human tumor cells in athymic mice by anti-epidermal growth factor receptor monoclonal antibodies.
Monoclonal antibodies (MoAbs) were raised against epidermal growth factor (EGF) receptors on a human epidermoid carcinoma cell line, A431. Administration of anti-EGF receptor MoAbs inhibited tumor formation in athymic mice by A431 cells and by another epidermal carcinoma cell line, T222. When one of the same MoAbs was used in therapy against Li-7 (a human hepatoma) and HeLa cells (a cervical carcinoma), tumor growth was not affected. The number of EGF receptors on A431 cells was about 100-fold higher than on T222, Li-7, and HeLa cells, suggesting that the number of EGF receptors may not be an important determinant in suppressing tumor growth. Three anti-EGF receptor MoAbs were used in the present studies. MoAbs 528 (immunoglobulin G2a) and 225 (immunoglobulin G1) are capable of competing with EGF for receptor binding and inhibit proliferation of A431 cells in culture. The other MoAb, 455 (immunoglobulin G1), is incapable of blocking the binding of EGF to its receptors and has no effect on the proliferation of cultured A431 cells. All three MoAbs inhibited A431 tumor growth in athymic mice, indicating that the antibody isotype and the site of binding on the EGF receptor are not the determinants of antiproliferative activity in vivo. The observation that MoAb against the receptor for EGF is cytostatic rather than cytocidal in vitro against A431 cells, yet completely prevents tumor growth in vivo, suggests that some host animal responses also may be involved in the antitumor effect. MoAbs against growth factor receptors could provide useful immunotherapeutic agents.


Overcoming challenges to sustainable immunization financing: early experiences from GAVI graduating countries.
Over the 5-year period ending in 2018, 16 countries with a combined birth cohort of over 6 million infants requiring life-saving immunizations are scheduled to transition (graduate) from outside financial and technical support for a number of their essential vaccines. This support has been provided over the past decade by the GAVI Alliance. Will these 16 countries be able to continue to sustain these vaccination efforts? To address this issue, GAVI and its partners are supporting transition planning, entailing country assessments of readiness to graduate and intensive dialogue with national officials to ensure a smooth transition process. This approach was piloted in Bhutan, Republic of Congo, Georgia, Moldova and Mongolia in 2012. The pilot showed that graduating countries are highly heterogeneous in their capacity to assume responsibility for their immunization programmes. Although all possess certain strengths, each country displayed weaknesses in some of the following areas: budgeting for vaccine purchase, national procurement practices, performance of national regulatory agencies, and technical capacity for vaccine planning and advocacy. The 2012 pilot experience further demonstrated the value of transition planning processes and tools. As a result, GAVI has decided to continue with transition planning in 2013 and beyond. As the graduation process advances, GAVI and graduating countries should continue to contribute to global collective thinking about how developing countries can successfully end their dependence on donor aid and achieve self-sufficiency.


Anticipating the impact of the COVID-19 pandemic on TB patients and TB control programmes.
The COVID-19 pandemic has currently overtaken every other health issue throughout the world. There are numerous ways in which this will impact existing public health issues. Here we reflect on the interactions between COVID-19 and tuberculosis (TB), which still ranks as the leading cause of death from a single infectious disease globally. There may be grave consequences for existing and undiagnosed TB patients globally, particularly in low and middle income countries (LMICs) where TB is endemic and health services poorly equipped. TB control programmes will be strained due to diversion of resources, and an inevitable loss of health system focus, such that some activities cannot or will not be prioritised. This is likely to lead to a reduction in quality of TB care and worse outcomes. Further, TB patients often have underlying co-morbidities and lung damage that may make them prone to more severe COVID-19. The symptoms of TB and COVID-19 can be similar, with for example cough and fever. Not only can this create diagnostic confusion, but it could worsen the stigmatization of TB patients especially in LMICs, given the fear of COVID-19. Children with TB are a vulnerable group especially likely to suffer as part of the "collateral damage". There will be a confounding of symptoms and epidemiological data through co-infection, as happens already with TB-HIV, and this will require unpicking. Lessons for COVID-19 could be learned from the vast experience of running global TB control programmes, while the astonishingly rapid and relatively well co-ordinated response to COVID-19 demonstrates how existing programmes could be significantly improved.


Alteration of brain glycogen turnover in the conscious rat after 5h of prolonged wakefulness.
Although glycogen (Glyc) is the main carbohydrate storage component, the role of Glyc in the brain during prolonged wakefulness is not clear. The aim of this study was to determine brain Glyc concentration ([]) and turnover time (tau) in euglycemic conscious and undisturbed rats, compared to rats maintained awake for 5h. To measure the metabolism of [1-(13)C]-labeled Glc into Glyc, 23 rats received a [1-(13)C]-labeled Glc solution as drink (10% weight per volume in tap water) ad libitum as their sole source of exogenous carbon for a "labeling period" of either 5h (n=13), 24h (n=5) or 48 h (n=5). Six of the rats labeled for 5h were continuously maintained awake by acoustic, tactile and olfactory stimuli during the labeling period, which resulted in slightly elevated corticosterone levels. Brain [Glyc] measured biochemically after focused microwave fixation in the rats maintained awake (3.9+/-0.2 micromol/g, n=6) was not significantly different from that of the control group (4.0+/-0.1 micromol/g, n=7; t-test, P>0.5). To account for potential variations in plasma Glc isotopic enrichment (IE), Glyc IE was normalized by N-acetyl-aspartate (NAA) IE. A simple mathematical model was developed to derive brain Glyc turnover time as 5.3h with a fit error of 3.2h and NAA turnover time as 15.6h with a fit error of 6.5h, in the control rats. A faster tau(Glyc) (2.9h with a fit error of 1.2h) was estimated in the rats maintained awake for 5h. In conclusion, 5h of prolonged wakefulness mainly activates glycogen metabolism, but has minimal effect on brain [Glyc].


Bounds on entanglement dimensions and quantum graph parameters via noncommutative polynomial optimization.
In this paper we study optimization problems related to bipartite quantum correlations using techniques from tracial noncommutative polynomial optimization. First we consider the problem of finding the minimal entanglement dimension of such correlations. We construct a hierarchy of semidefinite programming lower bounds and show convergence to a new parameter: the minimal average entanglement dimension, which measures the amount of entanglement needed to reproduce a quantum correlation when access to shared randomness is free. Then we study optimization problems over synchronous quantum correlations arising from quantum graph parameters. We introduce semidefinite programming hierarchies and unify existing bounds on quantum chromatic and quantum stability numbers by placing them in the framework of tracial polynomial optimization.


Activation of HIV transcription with short-course vorinostat in HIV-infected patients on suppressive antiretroviral therapy.
Human immunodeficiency virus (HIV) persistence in latently infected resting memory CD4+ T-cells is the major barrier to HIV cure. Cellular histone deacetylases (HDACs) are important in maintaining HIV latency and histone deacetylase inhibitors (HDACi) may reverse latency by activating HIV transcription from latently infected CD4+ T-cells. We performed a single arm, open label, proof-of-concept study in which vorinostat, a pan-HDACi, was administered 400 mg orally once daily for 14 days to 20 HIV-infected individuals on suppressive antiretroviral therapy (ART). The primary endpoint was change in cell associated unspliced (CA-US) HIV RNA in total CD4+ T-cells from blood at day 14. The study is registered at ClinicalTrials.gov (NCT01365065). Vorinostat was safe and well tolerated and there were no dose modifications or study drug discontinuations. CA-US HIV RNA in blood increased significantly in 18/20 patients (90%) with a median fold change from baseline to peak value of 7.4 (IQR 3.4, 9.1). CA-US RNA was significantly elevated 8 hours post drug and remained elevated 70 days after last dose. Significant early changes in expression of genes associated with chromatin remodeling and activation of HIV transcription correlated with the magnitude of increased CA-US HIV RNA. There were no statistically significant changes in plasma HIV RNA, concentration of HIV DNA, integrated DNA, inducible virus in CD4+ T-cells or markers of T-cell activation. Vorinostat induced a significant and sustained increase in HIV transcription from latency in the majority of HIV-infected patients. However, additional interventions will be needed to efficiently induce virus production and ultimately eliminate latently infected cells. ClinicalTrials.gov NCT01365065.


Variation in the nasal cavity of baboon hybrids with implications for late Pleistocene hominins.
Hybridization is increasingly proving to be an important force shaping human evolution. Comparisons of both ancient and modern genomes have provided support for a complex evolutionary scenario over the past million years, with evidence for multiple incidents of gene exchange. However, to date, genetic evidence is still limited in its ability to pinpoint the precise time and place of ancient admixture. For that we must rely on evidence of admixture from the skeleton. The research presented here builds on previous work on the crania of baboon hybrids, focusing on the nasal cavity of olive baboons, yellow baboons, and first generation (F1) hybrids. The nasal cavity is a particularly important anatomical region for study, given the clear differentiation of this feature in Neanderthals relative to their contemporaries, and therefore it is a feature that will likely differ in a distinctive manner in hybrids of these taxa. Metric data consist of 45 linear, area, and volume measurements taken from CT scans of known-pedigree baboon crania. Results indicate that there is clear evidence for differences among the nasal cavities of the parental taxa and their F1 hybrids, including a greater degree of sexual dimorphism in the hybrids. There is also some evidence for transgressive phenotypes in individual F1 animals. The greatest amount of shape variation occurs in the anterior bony cavity, the choana, and the mid-nasopharynx. Extrapolating our results to the fossil record, we would expect F1 hybrid fossils to have larger nasal cavities, on average, than either parental taxon, with overall nasal cavity shape showing the most profound changes in regions that are distinct between the parental taxa (e.g., anterior nasal cavity). We also expect size and shape differences to be more pronounced in male F1 hybrids than in females. Because of pronounced anterior nasal cavity differences between Neanderthals and their contemporaries, we suggest that this model might be effective for examining the fossil record of late Pleistocene contact.


Empowering pharmacists in asthma management through interactive SMS (EmPhAsIS): study protocol for a randomized controlled trial.
Medication regimens for asthma are particularly vulnerable to adherence problems because of the requirement for long-term use and periods of symptom remission experienced by patients. Pharmacists are suited to impact medication adherence given their training, skills, and frequent contact with patients. The Empowering pharmacists in asthma management through interactive SMS (EmPhAsIS) trial involves an intervention leveraging mobile health (mHealth) technology to support community pharmacy practice with the hypothesis of improved medication adherence in asthma. This study is a pragmatic pharmacy-based, cluster, randomized controlled trial with 12 months of intervention delivery and follow-up. Pharmacies (the clusters) will be randomized at a 1:1 ratio to provide intervention or usual care. The EmPhAsIS intervention consists of patient asthma education, short message service (SMS)-based monthly assessment of adherence, and follow-up of non-adherent individuals by community pharmacists. There are no inclusion or exclusion criteria for pharmacies. Patients are eligible if they: are 14 years of age or older, fill a prescription for inhaled corticosteroid (either monotherapy or in a combination inhaler with long-acting beta-agonists), have been diagnosed with asthma, possess a mobile phone with SMS capabilities, and have no communication difficulties such as inability to communicate in English, or significant impairment in vision, hearing, or speech. The primary outcome is adherence to inhaled corticosteroids ascertained by the medication possession ratio, the ratio of the days of medication supplied to days in a given time interval. This study will also evaluate secondary outcomes including: asthma control, asthma-related quality of life, asthma-related hospital admissions, and use of reliever medications during the follow-up period. A nested economic evaluation using a probabilistic decision-analytic model will be used to perform a cost-effectiveness analysis from the societal perspective of the intervention compared with usual care over a 10-year time horizon. Considering the prevalence of asthma, the extent of the non-adherence problem in this disease, and the availability of effective treatments, there is a tremendous potential to reduce the burden of asthma through improving adherence. This is the first study of an intervention based on mobile communication technology involving community pharmacists in asthma management. ClinicalTrials.gov identifier: NCT02170883; date of registration: 19 June 2014.


Expression of multidrug resistance 1 and glutathione-S-transferase-Pi protein in nasopharyngeal carcinoma.
Radiotherapy is the modality of choice for the treatment of nasopharyngeal carcinoma (NPC). However, systemic chemotherapy has recently been found to play an increasing role in the treatment of advanced or metastatic disease. The status of drug resistance gene expression that has crucial impact on chemotherapy has not been fully addressed for patients with NPC. In this study, we examined the expression of multidrug resistance 1 (MDR-1) and glutathione-S-transferase-Pi (GST-Pi) in primary, recurrent, and metastatic NPC using results of immunohistochemical examinations. The results were correlated with the expression of Epstein-Barr virus (EBV) latent protein, latent membrane protein 1 (LMP1), and clinicopathologic features, including stage, histopathologic types, and survival rates. MDR-1 protein expression was detected in 18 (12.6%) of 143 patients with primary NPC, 14 (32.6%) of 43 with recurrent NPC, and O (0%) of 20 with metastatic NPC, whereas 83 (58%) of 143 patients with primary NPC, 30 (69.8%) of 43 with recurrent NPC, and 13 (65%) of 20 with metastatic NPC expressed GST-Pi. EBV-LMP1 was expressed in 59 (41.3%) of 143 patients with primary NPC, 23 (53.5%) of 43 with recurrent NPC, and 9 (45%) of 20 with metastatic NPC. Simultaneous expression of MDR1 and GST-Pi was observed in 13 (72.2%) of 18 patients with primary NPC and 12 (85.7%) of 14 with recurrent NPC. The expression of LMP1 was detected in only 6 of the 13 patients with primary NPC and 6 of the 12 with recurrent NPC. We concluded that the expression of GST-Pi was more frequent in NPC tumor tissues than the expression of MDR-1. The expression of MDR-1 correlated with clinicopathologic features of primary NPC, including the histopathologic types and survival rates, but not with disease stage. The expression of GST-Pi did not correlate with clinicopathologic features. The expression of MDR-1 and GST-Pi did not correlate with expression of EBV-LMP1 for patients with NPC.


The alpha isoform of protein kinase C is involved in signaling the response of desmosomes to wounding in cultured epithelial cells.
Initiation of reepithelialization upon wounding is still poorly understood. To enhance this understanding, we focus here on changes in the adhesive state of desmosomes of cultured Madin-Darby canine kidney cells in response to wounding of confluent cell sheets. Previous results show that desmosomal adhesion in Madin-Darby canine kidney cells changes from a calcium-dependent state to calcium independence in confluent cell sheets. We show that this change, which requires culture confluence to develop, is rapidly reversed upon wounding of confluent cell sheets. Moreover, the change to calcium dependence in wound edge cells is propagated to cells hundreds of micrometers away from the wound edge. Rapid transition from calcium independence to calcium dependence also occurs when cells are treated with phorbol esters that activate PKC. PKC inhibitors, including the conventional isoform inhibitor Gö6976, cause rapid transition from calcium dependence to calcium independence, even in subconfluent cells. The cellular location of the alpha isoform of PKC correlates with the calcium dependence of desmosomes. Upon monolayer wounding, PKCalpha translocates rapidly to the cell periphery, becomes Triton X-100 insoluble, and also becomes concentrated in lamellipodia. The PKCalpha translocation upon wounding precedes both the increase in PKC activity in the membrane fraction and the reversion of desmosomes to calcium dependence. Specific depletion of PKCalpha with an antisense oligonucleotide increases the number of cells with calcium-independent desmosomes. These results show that PKCalpha participates in a novel signaling pathway that modulates desmosomal adhesion in response to wounding.


Community-university partnerships in community-based research.
Community-engaged research (CEnR) is a complex, collaborative process that presents many challenges and requires investment of time and commitment by both community and university research partners. This paper describes the experience of a group of university and community members developing a set of guidelines for the ethical conduct of CEnR projects. The paper outlines the process of guideline development and lessons learned from this collaborative effort, which was based upon approaches and methods of community-based participatory research (CBPR). The guidelines are included and may serve as a framework to be individualized by other partnerships. Our experience shows that the very process of review, revision, and engagement is extremely helpful in creating a framework that works for the specific communities and for establishing working relationships among the partners so that all stakeholders feel ownership and investment in the framework and the collaborative research efforts.


Thought disorder and frontotemporal volumes in pediatric epilepsy.
The aim of this study was to determine if volumes of frontotemporal regions associated with language were related to thought disorder in 42 children, aged 5-16 years, with cryptogenic epilepsy, all of whom had complex partial seizures (CPS). The children with CPS and 41 age- and gender-matched healthy children underwent brain MRI scans at 1.5 T. Tissue was segmented, and total brain, frontal lobe, and temporal lobe volumes were computed. Thought disorder measures, IQ, and seizure information were collected for each patient. The subjects with CPS had more thought disorder, smaller total gray matter and orbital frontal gray matter volumes, as well as larger temporal lobe white matter volumes than the control group. In the CPS group, thought disorder was significantly related to smaller orbital frontal and inferior frontal gray matter volumes, increased Heschl's gyrus gray matter volumes, and smaller superior temporal gyrus white matter volumes. However, significantly larger orbital frontal gyrus, superior temporal gyrus, and temporal lobe gray matter volumes and decreased Heschl's gyrus white matter volumes were associated with thought disorder in the control group. These findings suggest that thought disorder might represent a developmental disability involving frontotemporal regions associated with language in pediatric CPS.


Inverse association of pulse pressure augmentation during exercise with heart failure and death.
Resting pulse pressure (PP) is a risk factor for heart failure (HF); however, whether PP augmentation during exercise, a parameter easily obtained from routine treadmill stress testing, is associated with incident HF is unknown. Thus, we aimed to study the relationship between a novel parameter, the pulse pressure stress index (P2SI), and adverse outcomes among adults undergoing clinical exercise stress testing in the Henry Ford Exercise Testing Project. The P2SI was calculated as PP at peak exercise divided by resting PP and was analysed continuously and categorically using quartiles. Cox models examined the association between P2SI and adjusted HR (aHR) of incident HF, myocardial infarction (MI) or death. Receiver operating curve (ROC) analyses tested the optimal prognostic cut-point for P2SI. Among 55 524 participants without prior MI or HF, mean (SD) age was 53 (13) years, 51% were men and 29% black. A total of 2516 HF, 1606 MI and 6224 mortality outcomes occurred. Quartile 3 P2SI (2.0-2.4) was chosen as the reference category based on ROC analyses. There was a graded inverse association of low P2SI with excess HF (aHR of 1.3 (95% CI 1.1 to 1.5) for quartile 2 and 1.5 (95% CI 1.2 to 1.8) for quartile 1, p for trend<0.001) and mortality (aHR of 1.1 (95% CI 1.01 to 1.2) for quartile 2 and 1.3 (95% CI 1.2 to 1.5) for quartile 1, p for trend<0.001). There was no association between P2SI and MI after adjustment. P2SI added significant prognostic information to more established stress testing parameters such as peak systolic blood pressure, per cent maximal predicted heart rate achieved and metabolic equivalents of task achieved. Poor augmentation of PP with exercise, specifically a P2SI below 2, is a novel and readily quantifiable exercise-based risk feature for HF and death.


N-cadherin in the maintenance of human corneal limbal epithelial progenitor cells in vitro.
To demonstrate the role of N-cadherin (N-cad) in maintaining the progenitor status of primary human limbal epithelial cells in vitro. Immunohistochemistry and immunoelectron microscopy against N-cad was performed in human limbal tissue. The expression of N-cad, cytokeratin (K) 12, and K14 was also observed in primary cultured limbal epithelial cell colonies in 3T3 feeder cells, which also express N-cad. Laser microdissection of individual colonies was performed to separate central cells from peripheral cells to compare secondary colony formation. Finally, an artificial small interfering RNA (siRNA) expression vector was used to downregulate N-cad in 3T3 cells (N-cad(low) 3T3) to observe changes in colony formation and cultivated epithelial sheet phenotype. N-cad was expressed in clusters of basal limbal epithelial cells. Limbal epithelial cell colonies cocultured with N-cad(+) 3T3 feeder cells showed N-cad expression along the edge of each colony. When individual colonies were divided into peripheral and central sections by laser microdissection, peripheral cells had a significantly higher secondary colony-forming efficiency than did central cells. Furthermore, colonies using N-cad(low) 3T3 cells were significantly smaller than mock-transfected cells. Then a duplex-feeder model with two layers of either 3T3 or N-cad(low) 3T3 cells was used to produce stratified epithelial sheets. Only N-cad(+) 3T3 cells produced epithelial sheets with basal K15/ suprabasal K12 expression observed in limbal tissue. N-cad plays a pivotal role in the maintenance of the progenitor phenotype in cultured limbal epithelial cells.


Attributable risk estimates for cataract to prioritize medical and public health action.
Cataract is the most common cause of blindness in the world. The purpose of this study was to estimate the population attributable risk associated with identified risk factors for cortical, nuclear, and posterior subcapsular (PSC) cataract in a representative sample of the Victorian population aged 40 years and older. Cluster, stratified sampling was used and participants were recruited through a household census. At locally established test sites, standardized clinical examinations were performed to assess cataract and personal interviews were conducted to quantify potential risk factors. Multivariate logistic regression was used to determine the independent risk factors associated with the three types of cataract, and the population attributable risk was calculated. A total of 3271 (83% of eligible) of the urban residents and 1473 (92%) rural residents participated. The urban residents ranged in age from 40 to 98 years (mean, 59 years), and 1511 (46%) were men. The rural residents ranged in age from 40 to 103 years (mean, 60 years), and 701 (48%) were men. The overall prevalence of cortical cataract was 12.1% (95% CL 10.5, 13.8), nuclear cataract 12.6% (95% CL 9.61, 15.7), and PSC cataract 4.93% (95% CL 3.68, 6.17). Significant risk factors for cortical cataract included age, female gender, diabetes for greater than 5 years, gout for greater than 20 years, arthritis, myopia, average annual ocular UV-B exposure, and family history of cataract (parents or siblings). Significant risk factors for nuclear cataract included age, female gender, rural residence, age-related maculopathy, diabetes for greater than 5 years, smoker for greater than 30 years, and myopia. The significant risk factors for PSC cataract were age, rural residence, thiazide diuretic use, and myopia. Of the modifiable risk factors, ocular UV-B exposure explains 10% of the cortical cataract in the community, and cigarette smoking accounts for 17% of the nuclear cataract. Because of the near universal exposure to UV-B in the environment, ocular protection has one of the highest modifiable attributable risks for cortical cataract and would therefore be an ideal target for public health intervention. Quit smoking campaigns can be expanded to incorporate information about the excess cataract in the community associated with long-term smoking. Nonmodifiable risk factors such as age, gender, and long-term medication use have implications for the timely referral and treatment for those at higher risk of cataract.


A 40-basepair VNTR polymorphism in the dopamine transporter (DAT1) gene and the rapid response to antidepressant treatment.
Finding predictors of the response to antidepressant therapy is a major goal of molecular psychiatry. The genes encoding the serotonin (SERT) and dopamine (DAT1) transporters are among the possible candidate genes modulating an individual's antidepressant response. In a naturalistic prospective cohort study with a total of 190 fully assessed patients, improvement of depression symptoms during the 3 weeks following initiation of antidepressant therapy was recorded using the 21-item Hamilton Depression Rating Scale (HDRS). The SLC6A3 3' UTR 40-bp variable number of tandem repeats (VNTR) and the SLC6A4 5' 44-bp insertion/deletion polymorphism were analyzed by polymerase chain reaction. There was a significantly smaller number of rapid responders among homozygous carriers of the DAT1 9-repeat allele (9/9) than among heterozygous (9/10) and homozygous (10/10) carriers of the 10-repeat allele (19 versus 37 versus 52%, respectively, P=0.0037). Median decline in HDRS score was 35, 40, and 52% in patients with the 9/9, 9/10, and 10/10 genotypes, respectively (P=0.013). The effect was found in all classes of medications (selective serotonin reuptake inhibitors (SSRIs), tricyclics, mirtazapine, venlafaxine) and statistically significant also within the subgroup of patients having received SSRIs. The serotonin promoter insertion/deletion genotype had no effect in the entire study group, but there was an insignificant trend of better response in the l/l and l/s carriers who received SSRIs or mirtazapine. In conclusion, the dopamine transporter VNTR polymorphism influenced rapid response to antidepressant therapy. Compared with homozygous carriers of the 10-repeat allele, carriers of the 9/10 genotype had an odds ratio (OR) calculated by logistic regression analysis of 1.6 (95% CI 0.8-3.2) and carriers of the 9/9 genotype had an OR of 6.0 (1.5-24.4) for no or poor response. Further studies are required to confirm this clinical association and to elucidate the underlying mechanisms.


Down-regulation of the trypanosomatid signal recognition particle affects the biogenesis of polytopic membrane proteins but not of signal peptide-containing proteins.
Protein translocation across the endoplasmic reticulum is mediated by the signal recognition particle (SRP). In this study, the SRP pathway in trypanosomatids was down-regulated by two approaches: RNA interference (RNAi) silencing of genes encoding SRP proteins in Trypanosoma brucei and overexpression of dominant-negative mutants of 7SL RNA in Leptomonas collosoma. The biogenesis of both signal peptide-containing proteins and polytopic membrane proteins was examined using endogenous and green fluorescent protein-fused proteins. RNAi silencing of SRP54 or SRP68 in T. brucei resulted in reduced levels of polytopic membrane proteins, but no effect on the level of signal peptide-containing proteins was observed. When SRP deficiency was achieved in L. collosoma by overexpression of dominant-negative mutated 7SL RNA, a major effect was observed on polytopic membrane proteins but not on signal peptide-containing proteins. This study included two trypanosomatid species, tested various protein substrates, and induced depletion of the SRP pathway by affecting either the levels of SRP binding proteins or that of SRP RNA. Our results demonstrate that, as in bacteria but in contrast to mammalian cells, the trypanosome SRP is mostly essential for the biogenesis of membrane proteins.


Intelligence in DSM-IV combined type attention-deficit/hyperactivity disorder is not predicted by either dopamine receptor/transporter genes or other previously identified risk alleles for attention-deficit/hyperactivity disorder.
A major goal of genetic studies of attention deficit hyperactivity disorder (ADHD) is to identify individual characteristics that might help segregate the disorder's inherent heterogeneity. [Mill et al. (2006); Arch Ger Psychiatry 63:462-469] recently reported a potentially important association between two dopamine-related risk polymorphisms (DRD4 variable number tandem repeat (VNTR) in exon 3 and DAT1 VNTR in the 3' UTR) and lowered IQ in ADHD. The objective of the current study was to replicate the [Mill et al. (2006); Arch Ger Psychiatry 63:462-469] findings in a clinical sample and to extend the analysis to a large range of alternative SNP markers of putative ADHD risk alleles identified in a recent study [Brookes et al. (2006); Mol Genet 11:934-953]. Participants were 1081 children and adolescents with a research-confirmed combined type ADHD diagnosis and 1300 unaffected siblings who took part in the International Multi-centre ADHD Genetics (IMAGE) project. They were recruited from multiple settings from across Europe: Belgium, Britain, Germany, Ireland, Israel, Netherlands, Spain and Switzerland. The results were that ADHD was associated with reduced IQ. However, there was no association between the two dopamine-related risk polymorphisms and IQ in either the probands or their siblings. Furthermore, other selected genetic markers previously demonstrated to be associated with ADHD in this sample were not associated with IQ. This large scale study with a clinically ascertained and regorously diagnosed sample failed to replicate the association between genetic polymorphisms in the dopamine system and IQ in ADHD. We also observed no association of other SNPs with IQ in ADHD.


Functional cardiomyocytes derived from Isl1 cardiac progenitors via Bmp4 stimulation.
As heart failure due to myocardial infarction remains a leading cause of morbidity worldwide, cell-based cardiac regenerative therapy using cardiac progenitor cells (CPCs) could provide a potential treatment for the repair of injured myocardium. As adult CPCs may have limitations regarding tissue accessibility and proliferative ability, CPCs derived from embryonic stem cells (ESCs) could serve as an unlimited source of cells with high proliferative ability. As one of the CPCs that can be derived from embryonic stem cells, Isl1 expressing cardiac progenitor cells (Isl1-CPCs) may serve as a valuable source of cells for cardiac repair due to their high cardiac differentiation potential and authentic cardiac origin. In order to generate an unlimited number of Isl1-CPCs, we used a previously established an ESC line that allows for isolation of Isl1-CPCs by green fluorescent protein (GFP) expression that is directed by the mef2c gene, specifically expressed in the Isl1 domain of the anterior heart field. To improve the efficiency of cardiac differentiation of Isl1-CPCs, we studied the role of Bmp4 in cardiogenesis of Isl1-CPCs. We show an inductive role of Bmp directly on cardiac progenitors and its enhancement on early cardiac differentiation of CPCs. Upon induction of Bmp4 to Isl1-CPCs during differentiation, the cTnT+ cardiomyocyte population was enhanced 2.8±0.4 fold for Bmp4 treated CPC cultures compared to that detected for vehicle treated cultures. Both Bmp4 treated and untreated cardiomyocytes exhibit proper electrophysiological and calcium signaling properties. In addition, we observed a significant increase in Tbx5 and Tbx20 expression in differentiation cultures treated with Bmp4 compared to the untreated control, suggesting a link between Bmp4 and Tbx genes which may contribute to the enhanced cardiac differentiation in Bmp4 treated cultures. Collectively these findings suggest a cardiomyogenic role for Bmp4 directly on a pure population of Isl1 expressing cardiac progenitors, which could lead to enhancement of cardiac differentiation and engraftment, holding a significant therapeutic value for cardiac repair in the future.


Hepatobiliary Scintigraphy to Evaluate Biliary Complications of Pediatric Liver Transplantation: An Account of an Experience.
Hepatobiliary scintigraphy (HS) is a noninvasive imaging technique whose use in the follow-up of liver transplantation has not been duly documented. The main objective of this study is to describe the experience of using this technique to detect biliary complications in pediatric patients following liver transplantation. A retrospective, observational, and descriptive study involving 86 pediatric patients who had undergone liver transplantation between 2013 and 2018. Of the 86, 31 had undergone at least one HS during their postoperative period. A total of 45 studies were performed on 31 patients (36% of the patients undergoing transplantation during that time period). Patient ages ranged from 5 to 204 months (mean = 50 months). A total of 22 transplants (71%) were from living donors and 9 (29%) were from cadaveric donors. Of the 45 studies, 22 were positive for biliary complications, and all of them had an impact on clinical decision-making. The remaining 23 studies were negative. Of these 23, 19 continued under medical treatment and the other four underwent an additional intervention with positive surgical outcomes in all cases. All scintigraphy studies revealed hepatocellular dysfunction and cholestasis. The HS is a useful, noninvasive, and diagnostic procedure for the early diagnosis of biliary complications that may impact the evolution of disease in liver transplant patients. It allows the treating physician to make a more informed decision regarding expectant management, surgical management, or a less invasive course of action for transplantation complications.


Factors associated with satisfaction or regret following male-to-female sex reassignment surgery.
This study examined factors associated with satisfaction or regret following sex reassignment surgery (SRS) in 232 male-to-female transsexuals operated on between 1994 and 2000 by one surgeon using a consistent technique. Participants, all of whom were at least 1-year postoperative, completed a written questionnaire concerning their experiences and attitudes. Participants reported overwhelmingly that they were happy with their SRS results and that SRS had greatly improved the quality of their lives. None reported outright regret and only a few expressed even occasional regret. Dissatisfaction was most strongly associated with unsatisfactory physical and functional results of surgery. Most indicators of transsexual typology, such as age at surgery, previous marriage or parenthood, and sexual orientation, were not significantly associated with subjective outcomes. Compliance with minimum eligibility requirements for SRS specified by the Harry Benjamin International Gender Dysphoria Association was not associated with more favorable subjective outcomes. The physical results of SRS may be more important than preoperative factors such as transsexual typology or compliance with established treatment regimens in predicting postoperative satisfaction or regret.


Digital atlas of the zebra finch (Taeniopygia guttata) brain: a high-resolution photo atlas.
We describe a set of new comprehensive, high-quality, high-resolution digital images of histological sections from the brain of male zebra finches (Taeniopygia guttata) and make them publicly available through an interactive website (http://zebrafinch.brainarchitecture.org/). These images provide a basis for the production of a dimensionally accurate and detailed digital nonstereotaxic atlas. Nissl- and myelin-stained brain sections are provided in the transverse, sagittal, and horizontal planes, with the transverse plane approximating the more traditional Frankfurt plane. In addition, a separate set of brain sections in this same plane is stained for tyrosine hydroxylase, revealing the distribution of catecholaminergic neurons (dopaminergic, noradrenergic, and adrenergic) in the songbird brain. For a subset of sagittal sections we also prepared a corresponding set of drawings, defining and annotating various nuclei, fields, and fiber tracts that are visible under Nissl and myelin staining. This atlas of the zebra finch brain is expected to become an important tool for birdsong research and comparative studies of brain organization and evolution.


Centromere remodeling in Hoolock leuconedys (Hylobatidae) by a new transposable element unique to the gibbons.
Gibbons (Hylobatidae) shared a common ancestor with the other hominoids only 15-18 million years ago. Nevertheless, gibbons show very distinctive features that include heavily rearranged chromosomes. Previous observations indicate that this phenomenon may be linked to the attenuated epigenetic repression of transposable elements (TEs) in gibbon species. Here we describe the massive expansion of a repeat in almost all the centromeres of the eastern hoolock gibbon (Hoolock leuconedys). We discovered that this repeat is a new composite TE originating from the combination of portions of three other elements (L1ME5, AluSz6, and SVA_A) and thus named it LAVA. We determined that this repeat is found in all the gibbons but does not occur in other hominoids. Detailed investigation of 46 different LAVA elements revealed that the majority of them have target site duplications (TSDs) and a poly-A tail, suggesting that they have been retrotransposing in the gibbon genome. Although we did not find a direct correlation between the emergence of LAVA elements and human-gibbon synteny breakpoints, this new composite transposable element is another mark of the great plasticity of the gibbon genome. Moreover, the centromeric expansion of LAVA insertions in the hoolock closely resembles the massive centromeric expansion of the KERV-1 retroelement reported for wallaby (marsupial) interspecific hybrids. The similarity between the two phenomena is consistent with the hypothesis that evolution of the gibbons is characterized by defects in epigenetic repression of TEs, perhaps triggered by interspecific hybridization.


Risk factors for sleep-disordered breathing in pregnancy.
Symptoms of sleep-disordered breathing (SDB) are common among pregnant women, and several studies link SDB symptoms with gestational hypertension and preeclampsia. However, few prospective studies objectively measuring SDB during pregnancy have been performed. We performed a prospective cohort study examining risk factors for third trimester SDB in pregnant women. 105 pregnant women from the Hospital of the University of Pennsylvania obstetrics practices completed first and third trimester overnight polysomnography studies. We examined whether the number of SDB events per hour of sleep increased during pregnancy. We performed unadjusted and multivariable logistic regression analyses to estimate the effects of usual and pregnancy-specific characteristics on development of third trimester obstructive sleep apnoea (OSA). In secondary analyses, we examined the relationship between objectively measured SDB, hypertensive disorders of pregnancy, and other adverse maternal-fetal outcomes. Mean Apnoea-Hypopnoea Index increased from 2.07 (SD 3.01) events/h at baseline (first trimester) to 3.74 (SD 5.97) in the third trimester (p=0.009). 10.5% of women had OSA in the first trimester. By the third trimester, 26.7% of women had OSA. In multivariable analyses, first trimester body mass index (BMI) and maternal age were significantly associated with third trimester OSA. Third trimester OSA is common. Risk factors for third trimester OSA among women without baseline SDB include higher baseline BMI and maternal age.


Recurrent Rearrangements of Human Amylase Genes Create Multiple Independent CNV Series.
The human amylase gene cluster includes the human salivary (AMY1) and pancreatic amylase genes (AMY2A and AMY2B), and is a highly variable and dynamic region of the genome. Copy number variation (CNV) of AMY1 has been implicated in human dietary adaptation, and in population association with obesity, but neither of these findings has been independently replicated. Despite these functional implications, the structural genomic basis of CNV has only been defined in detail very recently. In this work, we use high-resolution analysis of copy number, and analysis of segregation in trios, to define new, independent allelic series of amylase CNVs in sub-Saharan Africans, including a series of higher-order expansions of a unit consisting of one copy each of AMY1, AMY2A, and AMY2B. We use fiber-FISH (fluorescence in situ hybridization) to define unexpected complexity in the accompanying rearrangements. These findings demonstrate recurrent involvement of the amylase gene region in genomic instability, involving at least five independent rearrangements of the pancreatic amylase genes (AMY2A and AMY2B). Structural features shared by fundamentally distinct lineages strongly suggest that the common ancestral state for the human amylase cluster contained more than one, and probably three, copies of AMY1.


A randomized 'blinded' comparison of two doses of antivenom in the treatment of Bothrops envenoming in São Paulo, Brazil.
An earlier study in São Paulo state suggested that the dose for patients with mild or moderate envenoming by Bothrops snakes (mainly Bothrops jararaca) could be effectively decreased to 4 ampoules (40 mL) of Brazilian Brothrops polyspecific antivenom. The present 'blinded' study examined the lowest dose studied in the first trial (equivalent to 4 x 10 mL ampoules) and half that dose of antivenom (equivalent to 2 x 10 mL ampoules) in 2 similar groups of 170 patients who were comparable in all respects before treatment. The majority of patients showed rapid clinical improvement after treatment with either dose regimen and rapid restoration of blood coagulability and cessation of bleeding. There was no apparent difference between the 2 groups of patients in any respect. The study confirmed that, in such patients, the dose of antivenom can be decreased from 4 ampoules to 2 ampoules without reduction of therapeutic efficacy, and it is highly likely that this reduction will result in a decrease of early anaphylactic reactions caused by the antivenom.


Patients' perceptions of transrectal prostate biopsy: a qualitative study.
This study explored men's experiences of transrectal prostate biopsy. Fifty men who had had a prostate biopsy talked about the experience as part of an in-depth interview; 36 were interviewed in 2000 about all aspects of prostate cancer, and 14 in 2005 about their experience of prostate-specific antigen testing, subsequent investigations and treatment. Men were recruited via urologists, general practitioners and support groups. In both studies, we aimed to include men of various ages, from different backgrounds, who lived, and had been investigated and treated, in different parts of the UK. A qualitative interpretive approach was taken, combining thematic analysis with constant comparison. Most men described the procedure as merely 'uncomfortable', but some found it stressful, exhausting and extremely painful. Worries included the fear that cancer cells might pass from a man to his wife during ejaculation, and that a biopsy might spread cancer cells to other parts of the body. Men should be given detailed information before a biopsy, so that they are well aware of what might happen. They should also be given the opportunity to voice their fears, so that they can be reassured, and offered some form of pain relief.


Do soil microbes and abrasion by soil particles influence persistence and loss of physical dormancy in seeds of tropical pioneers?
Germination from the soil seed bank (SSB) is an important determinant of species composition in tropical forest gaps, with seed persistence in the SSB allowing trees to recruit even decades after dispersal. The capacity to form a persistent SSB is often associated with physical dormancy, where seed coats are impermeable at the time of dispersal. Germination literature often speculates, without empirical evidence, that dormancy-break in physically dormant seeds is the result of microbial action and/or abrasion by soil particles. We tested the microbial/soil abrasion hypothesis in four widely distributed neotropical pioneer tree species (Apeiba membranacea, Luehea seemannii, Ochroma pyramidale, and Cochlospermum vitifolium). Seeds were buried in five common gardens in a lowland tropical forest in Panama, and recovered at 1, 3, 6, and 12 months after burial. Seed permeability, microbial infection, seed coat thickness, and germination were measured. Parallel experiments compared the germination fraction of fresh and aged seeds without soil contact, and in seeds as a function of seed permeability. Contrary to the microbial/soil abrasion hypothesis the proportion of permeable seeds, and of seeds infected by cultivable microbes, decreased as a function of burial duration. Furthermore, seeds stored in dark and dry conditions for 2 years showed a higher proportion of seed germination than fresh seeds in identical germination conditions. We determined that permeable seeds of A. membranacea and O. pyramidale had cracks in the chalazal area or lacked the chalazal plug, whereas all surfaces of impermeable seeds were intact. Our results are inconsistent with the microbial/soil abrasion hypothesis of dormancy loss and instead suggest the existence of multiple dormancy phenotypes, where a fraction of each seed cohort is dispersed in a permeable state and germinates immediately, while the impermeable seed fraction accounts for the persistent SSB. Thus, we conclude that fluctuations in the soil temperature in the absence of soil abrasion and microbial infection are sufficient to break physical dormancy on seeds of tropical pioneer trees.


Long-term effect of pregnancy-related factors on the development of endometrial neoplasia: A nationwide retrospective cohort study.
By identifying pregnancy-related risk factors for endometrial neoplasia, women's risk of developing this disease after childbirth can be predicted and high-risk women can be screened for early detection. Study data from women who gave birth in Korea in 2007 were collected from the Korea National Health Insurance (KNHI) claims database between 2007 and 2015. The adjusted hazard ratios (HRs) and 95% confidence intervals (CIs) for the development of endometrial neoplasia were estimated by multivariate Cox proportional hazards models. Data from 386,614 women were collected for this study. By 2015, 3,370 women from the initial cohort had been diagnosed with endometrial neoplasia secondary to delivery. Multivariate Cox proportional hazards regression revealed that preeclampsia (HR 1.55, 95% CI 1.29, 1.86), advanced maternal age (≥ 35; HR 1.52, 95% CI 1.39, 1.66), multifetal pregnancy (HR 1.81, 95% CI 1.46, 2.23), multiparity (HR 1.16, 95% CI 1.08, 1.24), cesarean section (HR 1.15, 95% CI 1.07, 1.23) and delivery of a large-for-gestational-age infant (HR 1.19, 95% CI 1.02, 1.39) were independent risk factors for future endometrial neoplasia. The risk for endometrial neoplasia increased as the number of risk factors increased (risk factors ≥3: HR 2.11, 95% CI 1.86-2.40). This study showed that six pregnancy-related factors-advanced maternal age, multiparity, multifetal pregnancy, cesarean section, delivery of a large-for-gestational-age infant, and preeclampsia-are positively correlated with future development of endometrial neoplasia, including endometrial hyperplasia or cancer. Close observation and surveillance are warranted to enable early diagnosis of endometrial diseases, including endometrial cancer after pregnancy in high-risk women. However, due to unavailability of clinical information, many clinical/epidemiological factors can become confounders. Further research is needed on factors associated with the risk of endometrial neoplasia.


Altered renal expression of angiotensin II receptors, renin receptor, and ACE-2 precede the development of renal fibrosis in aging rats.
The susceptibility to fibrosis and progression of renal disease is mitigated by inhibition of the renin-angiotensin system (RAS). We hypothesized that activation of the intrarenal RAS predisposes to renal fibrosis in aging. Intrarenal expression of angiotensin II type 1 (AT(1)R), type 2 (AT(2)R), and (pro)renin receptors, ACE and ACE-2, as well as pro- and antioxidant enzymes were measured in 3-month-old (young), 14-month-old (middle-aged), and 24-month-old (old) male Sprague-Dawley rats. Old rats manifested glomerulosclerosis and severe tubulointerstitial fibrosis with increased fibronectin and TGF-β expression (7-fold). AT(1)R /AT(2)R ratios were increased in middle-aged (cortical 1.6-fold, medullary 5-fold) and old rats (cortical 2-fold, medullary 4-fold). Similarly, (pro)renin receptor expression was increased in middle-aged (cortical 2-fold, medullary 3-fold) and old (cortical 5-fold, medullary 3-fold) rats. Cortical ACE was increased (+35%) in old rats, whereas ACE-2 was decreased (-50%) in middle-aged and old rats. NADPH oxidase activity was increased (2-fold), whereas antioxidant capacity and expression of the mitochondrial enzyme manganese superoxide dismutase (cortical -40%, medullary -53%) and medullary endothelial nitric oxide synthase (-48%) were decreased in old rats. Age-related intrarenal activation of the RAS preceded the development of severe renal fibrosis, suggesting that it contributes to the increased susceptibility to renal injury observed in the elderly.


From DARPins to LoopDARPins: novel LoopDARPin design allows the selection of low picomolar binders in a single round of ribosome display.
Antibodies are the most versatile binding proteins in nature with six loops creating a flexible continuous interaction surface. However, in some molecular formats, antibodies are aggregation prone. Designed ankyrin repeat proteins (DARPins) were successfully created as alternative design solutions. Nevertheless, their concave shape, rigidity and incompletely randomized binding surface may limit the epitopes that can be targeted by this extremely stable scaffold. Combining conformational diversity and a continuous convex paratope found in many antibodies with the beneficial biophysical properties of DARPins, we created LoopDARPins, a next generation of DARPins with extended epitope binding properties. We employed X-ray structure determination of a LoopDARPin for design validation. Biophysical characterizations show that the introduction of an elongated loop through consensus design does not decrease the stability of the scaffold,consistent with molecular dynamics simulations. Ribosome-display selections against extracellular signal-regulated kinase 2 (ERK2) and four members of the BCL-2 family (BCL-2, BCL-XL, BCL-W and MCL-1) of anti-apoptotic regulators yielded LoopDARPins with affinities in the mid-picomolar to low nanomol arrange against all targets. The BCL-2 family binders block the interaction with their natural interaction partner and will be valuable reagents to test the apoptotic response in functional assays. With the LoopDARPin scaffold, binders for BCL-2 with an affinity of 30 pM were isolated with only a single round of ribosome display,an enrichment that has not been described for any scaffold. Identical stringent one-round selections with conventional DARPins without loop yielded no binders. The LoopDARPin scaffold may become a highly valuable tool for biotechnological high-throughput applications.


MAML1, a human homologue of Drosophila mastermind, is a transcriptional co-activator for NOTCH receptors.
Notch receptors are involved in cell-fate determination in organisms as diverse as flies, frogs and humans. In Drosophila melanogaster , loss-of-function mutations of Notch produce a 'neurogenic' phenotype in which cells destined to become epidermis switch fate and differentiate to neural cells. Upon ligand activation, the intracellular domain of Notch (ICN) translocates to the nucleus, and interacts directly with the DNA-binding protein Suppressor of hairless (Su(H)) in flies, or recombination signal binding protein Jkappa (RBP-Jkappa) in mammals, to activate gene transcription. But the precise mechanisms of Notch-induced gene expression are not completely understood. The gene mastermind has been identified in multiple genetic screens for modifiers of Notch mutations in Drosophila. Here we clone MAML1, a human homologue of the Drosophila gene Mastermind, and show that it encodes a protein of 130 kD localizing to nuclear bodies. MAML1 binds to the ankyrin repeat domain of all four mammalian NOTCH receptors, forms a DNA-binding complex with ICN and RBP-Jkappa, and amplifies NOTCH-induced transcription of HES1. These studies provide a molecular mechanism to explain the genetic links between mastermind and Notch in Drosophila and indicate that MAML1 functions as a transcriptional co-activator for NOTCH signalling.


The genome of the vervet (Chlorocebus aethiops sabaeus).
We describe a genome reference of the African green monkey or vervet (Chlorocebus aethiops). This member of the Old World monkey (OWM) superfamily is uniquely valuable for genetic investigations of simian immunodeficiency virus (SIV), for which it is the most abundant natural host species, and of a wide range of health-related phenotypes assessed in Caribbean vervets (C. a. sabaeus), whose numbers have expanded dramatically since Europeans introduced small numbers of their ancestors from West Africa during the colonial era. We use the reference to characterize the genomic relationship between vervets and other primates, the intra-generic phylogeny of vervet subspecies, and genome-wide structural variations of a pedigreed C. a. sabaeus population. Through comparative analyses with human and rhesus macaque, we characterize at high resolution the unique chromosomal fission events that differentiate the vervets and their close relatives from most other catarrhine primates, in whom karyotype is highly conserved. We also provide a summary of transposable elements and contrast these with the rhesus macaque and human. Analysis of sequenced genomes representing each of the main vervet subspecies supports previously hypothesized relationships between these populations, which range across most of sub-Saharan Africa, while uncovering high levels of genetic diversity within each. Sequence-based analyses of major histocompatibility complex (MHC) polymorphisms reveal extremely low diversity in Caribbean C. a. sabaeus vervets, compared to vervets from putatively ancestral West African regions. In the C. a. sabaeus research population, we discover the first structural variations that are, in some cases, predicted to have a deleterious effect; future studies will determine the phenotypic impact of these variations.


Stall no more at polyproline stretches with the translation elongation factors EF-P and IF-5A.
Synthesis of polyproline proteins leads to translation arrest. To overcome this ribosome stalling effect, bacteria depend on a specialized translation elongation factor P (EF-P), being orthologous and functionally identical to eukaryotic/archaeal elongation factor e/aIF-5A (recently renamed 'EF5'). EF-P binds to the stalled ribosome between the peptidyl-tRNA binding and tRNA-exiting sites, and stimulates peptidyl-transferase activity, thus allowing translation to resume. In their active form, both EF-P and e/aIF-5A are post-translationally modified at a positively charged residue, which protrudes toward the peptidyl-transferase center when bound to the ribosome. While archaeal and eukaryotic IF-5A strictly depend on (deoxy-) hypusination (hypusinylation) of a conserved lysine, bacteria have evolved diverse analogous modification strategies to activate EF-P. In Escherichia coli and Salmonella enterica a lysine is extended by β-lysinylation and subsequently hydroxylated, whereas in Pseudomonas aeruginosa and Shewanella oneidensis an arginine in the equivalent position is rhamnosylated. Inactivation of EF-P, or the corresponding modification systems, reduces not only bacterial fitness, but also impairs virulence. Here, we review the function of EF-P and IF-5A and their unusual posttranslational protein modifications.


TP53INP2 is the new guest at the table of self-eating.
We identified a TP53INP1-related gene, which was named TP53INP2, using a bioinformatic approach. TP53INP1 and TP53INP2 sequences were found in higher eukaryotes but not in lower eukaryotes nor in prokaryotes. To gain insight into the function of the TP53INP2 protein, we carried out a number of protein-protein interaction approaches which showed that TP53INP2 binds to the Atg8-like proteins, as well as with the autophagosome transmembrane protein VMP1. TP53INP2 translocates from the nucleus to autophagosomes after activation of autophagy. We also showed that TP53INP2 expression is necessary for autophagosome development since its siRNA-mediated knockdown strongly decreases sensitivity to induced-autophagy. We suggest that TP53INP2 is a scaffold protein that recruits Atg8-like proteins to the autophagosome membrane by interacting with the transmembrane protein VMP1.


Photoflocculation of TiO2 microgel mixed suspensions.
Photocatalytic TiO 2 was derivatized with an amino silane to give a positively charged colloidal suspension in water. UV irradiation of the cationic titania oxidized the organo silane coating converting the titania to a negatively charged colloid. Irradiation of mixed suspensions of cationic titania and cationic polyvinylamine microgel induced catastrophic flocculation. Electrophoresis and XPS evidence suggests that the UV removed the amine groups from the titania resulting in heteroflocculation of anionic titania with cationic microgel particles.