Severe anaemia in children living in a malaria endemic area of Kenya.
Severe anaemia is an important cause of morbidity and mortality in African children, but the causes, particularly falciparum malaria, are difficult to determine. We assessed the contribution of falciparum malaria to anaemia in Kenyan children by clinical examination and measurement of parasitaemia and haemoglobin (Hb) concentration in 559 children in the community and in 2412 children admitted to Kilifi district hospital during a 2-year period. We also attempted to characterize severe malarial anaemia by examining the causes and pathophysiology of anaemia in 101 children admitted with Hb concentration < or = 50 g/l during a 1-year period. Plasmodium falciparum infection was associated with reduced Hb concentration in children in the community and in those admitted to hospital irrespective of diagnosis. Falciparum malaria was the primary cause in 46 cases (46%) of severe anaemia admitted to hospital. There was no difference in the frequency of haemolysis or dyserythropoiesis in the children with malarial anaemia and those with anaemia from other causes, such as iron deficiency or sickle cell disease. The mortality rate in the children with severe malarial anaemia was 8.6% compared with 3.6% in children with severe anaemia due to other causes. Falciparum malaria does not present with a characteristic clinical or haematological picture, but is a major cause of the morbidity and mortality in children with severe anaemia who live on the Kenyan coast, a malaria endemic area.


A tobacco-free world: a call to action to phase out the sale of tobacco products by 2040.
The time has come for the world to acknowledge the unacceptability of the damage being done by the tobacco industry and work towards a world essentially free from the sale (legal and illegal) of tobacco products. A tobacco-free world by 2040, where less than 5% of the world's adult population use tobacco, is socially desirable, technically feasible, and could become politically practical. Three possible ways forward exist: so-called business-as-usual, with most countries steadily implementing the WHO Framework Convention on Tobacco Control (FCTC) provisions; accelerated implementation of the FCTC by all countries; and a so-called turbo-charged approach that complements FCTC actions with strengthened UN leadership, full engagement of all sectors, and increased investment in tobacco control. Only the turbo-charged approach will achieve a tobacco-free world by 2040 where tobacco is out of sight, out of mind, and out of fashion--yet not prohibited. The first and most urgent priority is the inclusion of an ambitious tobacco target in the post-2015 sustainable development health goal. The second priority is accelerated implementation of the FCTC policies in all countries, with full engagement from all sectors including the private sector--from workplaces to pharmacies--and with increased national and global investment. The third priority is an amendment of the FCTC to include an ambitious global tobacco reduction goal. The fourth priority is a UN high-level meeting on tobacco use to galvanise global action towards the 2040 tobacco-free world goal on the basis of new strategies, new resources, and new players. Decisive and strategic action on this bold vision will prevent hundreds of millions of unnecessary deaths during the remainder of this century and safeguard future generations from the ravages of tobacco use.


Control of cystic echinococcosis/hydatidosis: 1863-2002.
Echinococcosis/hydatidosis, caused by Echinococcus granulosus, is a chronic and debilitating zoonotic larval cestode infection in humans, which is principally transmitted between dogs and domestic livestock, particularly sheep. Human hydatid disease occurs in almost all pastoral communities and rangeland areas of the underdeveloped and developed world. Control programmes against hydatidosis have been implemented in several endemic countries, states, provinces, districts or regions to reduce or eliminate cystic echinococcosis (CE) as a public health problem. This review assesses the impact of 13 of the hydatid control programmes implemented, since the first was introduced in Iceland in 1863. Five island-based control programmes (Iceland, New Zealand, Tasmania, Falklands and Cyprus) resulted, over various intervention periods (from <15 to >50 years), in successful control of transmission as evidenced by major reduction in incidence rates of human CE, and prevalence levels in sheep and dogs. By 2002, two countries, Iceland and New Zealand, and one island-state, Tasmania, had already declared that hydatid disease had been eliminated from their territories. Other hydatid programmes implemented in South America (Argentina, Chile, Uruguay), in Europe (mid-Wales, Sardinia) and in East Africa (northwest Kenya), showed varying degrees of success, but some were considered as having failed. Reasons for the eventual success of certain hydatid control programmes and the problems encountered in others are analysed and discussed, and recommendations for likely optimal approaches considered. The application of new control tools, including use of a hydatid vaccine, are also considered.


Bone Material Properties in Osteogenesis Imperfecta.
Osteogenesis imperfecta entrains changes at every level in bone tissue, from the disorganization of the collagen molecules and mineral platelets within and between collagen fibrils to the macroarchitecture of the whole skeleton. Investigations using an array of sophisticated instruments at multiple scale levels have now determined many aspects of the effect of the disease on the material properties of bone tissue. The brittle nature of bone in osteogenesis imperfecta reflects both increased bone mineralization density-the quantity of mineral in relation to the quantity of matrix within a specific bone volume-and altered matrix-matrix and matrix mineral interactions. Contributions to fracture resistance at multiple scale lengths are discussed, comparing normal and brittle bone. Integrating the available information provides both a better understanding of the effect of current approaches to treatment-largely improved architecture and possibly some macroscale toughening-and indicates potential opportunities for alternative strategies that can influence fracture resistance at longer-length scales.


Driver Mutations in Uveal Melanoma: Associations With Gene Expression Profile and Patient Outcomes.
Frequent mutations have been described in the following 5 genes in uveal melanoma (UM): BAP1, EIF1AX, GNA11, GNAQ, and SF3B1. Understanding the prognostic significance of these mutations could facilitate their use in precision medicine. To determine the associations between driver mutations, gene expression profile (GEP) classification, clinicopathologic features, and patient outcomes in UM. Retrospective study of patients with UM treated by enucleation by a single ocular oncologist between November 1, 1998, and July 31, 2014. Clinicopathologic features, patient outcomes, GEP classification (class 1 or class 2), and mutation status were recorded. The study cohort comprised 81 participants. Their mean age was 61.5 years, and 37% (30 of 81) were female. The GEP classification was class 1 in 35 of 81 (43%), class 2 in 42 of 81 (52%), and unknown in 4 of 81 (5%). BAP1 mutations were identified in 29 of 64 (45%), GNAQ mutations in 36 of 81 (44%), GNA11 mutations in 36 of 81 (44%), SF3B1 mutations in 19 of 81 (24%), and EIF1AX mutations in 14 of 81 (17%). Sixteen of the mutations in BAP1 and 6 of the mutations in EIF1AX were previously unreported in UM. GNAQ and GNA11 mutations were mutually exclusive. BAP1, SF3B1, and EIF1AX mutations were almost mutually exclusive with each other. Using multiple regression analysis, BAP1 mutations were associated with class 2 GEP and older patient. EIF1AX mutations were associated with class 1 GEP and the absence of ciliary body involvement. SF3B1 mutations were associated with younger patient age. GNAQ mutations were associated with the absence of ciliary body involvement and greater largest basal diameter. GNA11 mutations were not associated with any of the analyzed features. Using Cox proportional hazards modeling, class 2 GEP was the prognostic factor most strongly associated with metastasis (relative risk, 9.4; 95% CI, 3.1-28.5) and melanoma-specific mortality (relative risk, 15.7; 95% CI, 3.6-69.1) (P < .001 for both). After excluding GEP class, the presence of BAP1 mutations was the factor most strongly associated with metastasis (relative risk, 10.6; 95% CI, 3.4-33.5) and melanoma-specific mortality (relative risk, 9.0; 95% CI, 2.8-29.2) (P < .001 for both). BAP1, SF3B1, and EIF1AX mutations occur during UM tumor progression in an almost mutually exclusive manner and are associated with different levels of metastatic risk. These mutations may have value as prognostic markers in UM.


Is lGnRH-III the most potent GnRH analog containing only natural amino acids that specifically inhibits the growth of human breast cancer cells?
Analogs of the decapeptide, gonadotropin-releasing hormone (GnRH), used in the treatment of hormone-dependent tumors, contain numerous unnatural amino acids, giving rise to many adverse effects. lGnRH-III, a natural isoform of GnRH isolated from the sea lamprey, is a weak agonist of GnRH in the pituitary, but inhibits the growth of human cancer cells in micromolar concentrations. As lGnRH-III is not a natural ligand in humans, it is possible that a more potent peptide, also containing only natural amino acids, can be synthesized. A positional scanning peptide library, focused on the variable region of the GnRH family of peptides, residues 5-8, was synthesized. The synthesized peptides were analyzed in competitive binding experiments and six new analogs were designed on the basis of the results. Their biological activities were evaluated in cell growth experiments. The only natural sequence selected was chicken GnRH-II. The synthetic library did not yield a more potent peptide than lGnRH-III.


A novel KIR3DS1*0130107 allele isolated by sequencing from an Asian donor.
This novel KIR3DS1 allele officially named as KIR3DS1*0130107 was isolated from DNA samples from Asia using high-resolution sequenced-based techniques. KIR3DS1*0130107 differs from the first member of the KIR3DS1*013 subgroup (KIR3DS1*0130101) by a single mutation at position 8922A>G (intron 5), just nine nucleotides away from the start of exon 6.


Angiopoietin-2 is required for postnatal angiogenesis and lymphatic patterning, and only the latter role is rescued by Angiopoietin-1.
VEGF and Angiopoietin-1 requisitely collaborate during blood vessel development. While Angiopoietin-1 obligately activates its Tie2 receptor, Angiopoietin-2 can activate Tie2 on some cells, while it blocks Tie2 activation on others. Our analysis of mice lacking Angiopoietin-2 reveals that Angiopoietin-2 is dispensable for embryonic vascular development but is requisite for subsequent angiogenic remodeling. Unexpectedly, mice lacking Angiopoietin-2 also exhibit major lymphatic vessel defects. Genetic rescue with Angiopoietin-1 corrects the lymphatic, but not the angiogenesis, defects, suggesting that Angiopoietin-2 acts as a Tie2 agonist in the former setting, but as an antagonist in the latter setting. Our studies define a vascular growth factor whose primary role is in postnatal angiogenic remodeling and also demonstrate that members of the VEGF and Angiopoietin families collaborate during development of the lymphatic vasculature.


Combined clotrimazole irrigation and depot therapy for canine nasal aspergillosis.
To evaluate the effect of short duration 1 per cent clotrimazole flush when combined with 1 per cent clotrimazole cream instilled into the frontal sinuses for the treatment of nasal aspergillosis in 14 dogs. Fourteen dogs with clinical, radiological, serological and rhinoscopic findings consistent with nasal aspergillosis were treated by frontal sinus trephination and a short, five-minute flushing of 1 per cent topical clotrimazole solution followed by a 1 per cent clotrimazole cream instilled as a depot agent. Twelve of the 14 dogs (86 per cent) responded well to treatment and either had no clinical signs after treatment or had signs consistent with mild rhinitis during a minimum follow-up period of six months. Only one dog required multiple treatments. Treatment was well tolerated by all patients, with minimal complications. This treatment compares favourably to previously published data using one-hour topical clotrimazole or enilconazole flushing treatment protocols. The treatment technique significantly reduced treatment time under anaesthesia.


The effects and risks associated to mephedrone and methylone in humans: A review of the preliminary evidences.
New psychoactive substances have drastically modified the world drug scene. An increasingly popular class comprises synthetic or substituted cathinones (legal highs, research chemicals, bath salts). Among the most common psychoactive constituents of bath salts are mephedrone and methylone. Recent reports on the abuse of novel synthetic cathinone derivatives call attention to the serious physical and psychological risks resulting from their consumption, thereby emphasizing the growing use of these drugs might constitute an important public health issue. In this paper, we will review the available data regarding the use and effects of mephedrone and methylone in humans in order to highlight their impact on public health. To reach this objective, a literature search was performed on two representative databases (Pubmed, Google Scholar), the Erowid Center website (a US non-profit educational organization that provides information about psychoactive plants and chemicals), and various governmental websites. The terms used for the database search were "mephedrone", "methylone", "new psychoactive substances", "synthetic cathinones", "substituted cathinones", "substance abuse", "substance use disorder", "adverse effects", "fatalities". The literature search was limited to years 2005-2015 and led to the identification of 71 potentially relevant articles. To date, the actual prevalence rates of their use remains difficult to estimate. Important health-related issues have emerged in relation to the somatic, psychiatric, and addictive consequences of their use. The potential chronic health effects of their prolonged use remain to date unknown (e.g., reproductive toxicity, genotoxicity and carcinogenic potential). Treatment for patients with prolonged exposure to synthetic cathinones should ideally include a drug management plan coupled with psychotherapy taking place in a structured program of care.


IVC filters-Trends in placement and indications, a study of 2 populations.
Inferior vena cava filter (IVCF) placement appears to be expanding over time despite absence of clear directing evidence.Two populations were studied. The first population included patients who received an IVCF between January 2005 and August 2013 at our community hospital center. Demographic information, indications for placement, and retrieval rate was recorded among other variables. The second population comprised of patients receiving an IVCF from 2005 to 2012 according to the Nationwide Inpatient Sample (NIS) using ICD-9CM coding. Patients were divided into 2 groups based on the year of admission for comparison, that is, first group from 2005 to 2008 and the second from 2009 to 2012. In addition, we analyzed annual trends in filter placement, acute venothromboembolic events (VTE) and several underlying comorbidities within this population.At our center, 802 IVCFs were placed (55.2% retrievable); 34% for absolute, 61% for relative, and 5% for prophylactic indications. Major bleeding (27.5%), minor self-limited bleeding (13.7%), and fall history (11.2%) were the commonest indications. Periprocedural complication rate was 0.7%, and filter retrieval rate was 7%. The NIS population (811,487 filters) saw a decline in IVCF placement after year 2009, following an initial uptrend (Ptrend < 0.01). IVCF use among patients with neither acute VTE nor bleeding among prior VTE saw a 3-fold absolute reduction from 2005 to 2012 (33,075-11,655; Ptrend < 0.01). Patients from 2009 to 2012 were more likely to be male and had higher rates of acute VTE, thrombolytic use, cancer, bleeding, hypotension, acute cardiorespiratory failure, shock, prior falls, blood product transfusion, hospital mortality including higher Charlson comorbidity scores. The patients were younger, had shorter length of stay, and were less likely to be associated with strokes including hemorrhagic or require ventilator support. Prior falls (adjusted odds ratio-aOR 2.8), thrombolytic use (aOR 1.76), and shock (aOR 1.45) were most predictive of IVCF placement between 2009 and 2012 on regression analysis.Recent trends suggest that a higher proportion of patients receive temporary IVCF, for predominantly relative indications. Nationally, the number of filters being placed is decreasing, especially among those who did not experience acute VTE or bleeding events. Prior falls, thrombolytic therapy, and shock were most predictive of IVCF placement in latter half of the study period.


Enzyme immunoassays for total and allergen specific IgE in population studies.
Extensive IgE serology in occupational or environmental health studies is often hampered by a lack of technical facilities and finance. The use in population studies of relatively simple and inexpensive enzyme immunoassays (EIAs) was therefore evaluated for the assessment of total serum immunoglobulin E (IgE), and of specific IgE reactions with various common (house dust mites, grass and birch pollen, and cat) or occupational (fungal alpha-amylase and rat urinary protein) allergens. Total IgE was measured with a sandwich EIA, calibrated with commercially available IgE standards. Reproducibility was studied by testing pooled normal human serum samples in each of a large series of test plates. A panel of 156 children's serum samples with known IgE values was used to compare the assay with other total IgE assays. A previously developed EIA for anti-yeast IgE was adapted for the measurement of IgE reacting with various common and occupational allergens. Binding of IgE to microwells coated with commercially available allergen extracts, or allergen preparations from our own laboratory, was measured with a monoclonal anti-human IgE antibody and subsequent incubations with biotinylated rabbit anti-mouse Ig and avidin-peroxidase. Panels of serum samples from school children (n = 116), bakery workers (n = 126), and laboratory animal workers (n = 52) were used to study sensitivity and specificity, with reference to skin prick tests as the standard, and to compare the EIAs with commercially available test kits. The detection limit of the EIA for total IgE was 0.5-1 kU/l for undiluted serum samples, and the coefficient of variation between assays was less than 15% at serum concentrations between 1 and 150 kU/l. Results obtained with the panel of 156 children's serum samples were strongly correlated (r2 = 0.86) with IgE concentrations measured previously by radioimmunoassay. The results of the EIA for various occupational allergens correlated very well, both qualitatively and quantitatively, with the results of commercial test kits. Sensitivity and specificity of the EIA results as a predictor of skin prick test reactivity towards common allergens (house dust mite, grass pollen, birch pollen, and cat) were remarkably high (> 80%-90%) in the series of 116 children's serum samples. In a population of bakery workers the specificity of the EIAs was also very high (> 90%). The sensitivity was notably lower (30%-70%) in this adult population, which is, however, in agreement with results reported for conventional IgE tests. As the costs were estimated to be at least five to 10-fold lower than those of commercial test kits, the EIAs for total and specific IgE may be very useful tools in epidemiological studies of atopic respiratory or other disorders.


Mutations in the cardiac myosin binding protein-C gene on chromosome 11 cause familial hypertrophic cardiomyopathy.
Familial hypertrophic cardiomyopathy (FHC) is an autosomal dominant disorder manifesting as cardiac hypertrophy with myocyte disarray and an increased risk of sudden death. Mutations in five different loci cause FHC and 3 disease genes have been identified: beta cardiac myosin heavy chain, alpha tropomyosin and cardiac troponin T. Because these genes encode contractile proteins, other FHC loci are predicted also to encode sarcomere components. Two further FHC loci have been mapped to chromosomes 11p13-q13 (CMH4, ref. 6) and 7q3 (ref. 7). The gene encoding the cardiac isoform of myosin binding protein-C (cardiac MyBP-C) has recently been assigned to chromosome 11p11.2 and proposed as a candidate FHC gene. Cardiac MyBP-C is arrayed transversely in sarcomere A-bands and binds myosin heavy chain in thick filaments and titin in elastic filaments. Phosphorylation of MyBP-C appears to modulate contraction. We report that cardiac MyBP-C is genetically linked to CMH4 and demonstrate a splice donor mutation in one family with FHC and a duplication mutation in a second. Both mutations are predicted to disrupt the high affinity, C-terminal, myosin-binding domain of cardiac MyBP-C. These findings define cardiac MyBP-C mutations as the cause of FHC on chromosome 11p and reaffirm that FHC is a disease of the sarcomere.


Patched and smoothened mRNA expression in human astrocytic tumors inversely correlates with histological malignancy.
Patched (Ptc) is a transmembrane receptor for sonic hedgehog (Shh) and functionally associated with another transmembrane protein, smoothened (Smo). Ptc is a tumor suppressor gene whereas Smo serves as a proto-oncogene of neuroectodermal tumors. Their downstream molecules, Gli1, Gli2, and Gli3, are oncogenes of glioblastomas. We have analyzed mRNA expression of Ptc, Smo, and Gli family members in human astrocytic tumors. The mRNA expression was quantified by real-time polymerase chain reactions in 40 tumors (diffuse astrocytomas; 6 cases: anaplastic astrocytomas; 12 cases: glioblastomas; 22 cases) and four cell lines derived from astrocytic tumors. The MIB-1 proliferating cell indices (PCIs) of these tumors were analyzed by immunohistochemistry. In comparison with the World Health Organization (WHO) classification, the amount of Ptc and Smo mRNAs decreased in proportion to the progression of histological maliganancy, and similar results were obtained with astrocytic tumor-derived cell lines. However, there was no remarkable correlation between the mRNA expression level of each gene and the MIB-1 PCIs. The mRNA expression level of Gli1 was variable and highly elevated in two cases. No remarkable features were found clinically or histologically in these two cases. In summary, our results indicate that Ptc and Smo mRNA levels have an inverse correlation with histological malignancy and suggest that these gene products are implicated in the suppression of astrocytic tumors. In contrast, there was no significant correlation between the mRNA levels of the Gli family members and histological malignancy, suggesting that Gli proteins are not associated with the progression of astrocytic tumors.


Role of proline peptide bond isomerization in unfolding and refolding of ribonuclease.
The isomerization of the proline peptide bond between tyrosine-92 and proline-93 in bovine pancreatic ribonuclease A has been investigated in the unfolded protein as well as during the slow refolding process. This bond is in the cis state in the native protein. By comparison of various homologous ribonucleases we show that isomerization of proline-93 is associated with a change in fluorescence of tyrosine-92. This provides a spectroscopic probe to monitor this process in the disordered chain after unfolding as well as its reversal in the course of slow refolding. In unfolded ribonuclease incorrect trans isomers of proline-93 are found in both slow-folding species. trans----cis reversal of isomerization of this proline peptide bond during refolding shows kinetics that are identical with the time course of formation of native protein. Isomerization of proline-93 is slower than the formation of a native-like folded intermediate that accumulates on the major slow refolding pathway. Models to explain these results are discussed.


Opioid maintenance treatment: trajectories in and out of treatment.
Although efficacy studies of opioid maintenance treatment (OMT) have shown evidence of treatment benefits, there is still need for studies on its effectiveness in natural clinical processes. This study investigates the development in health, substance use and social conditions of those who applied for OMT, including those denied access or discharged. First, persons assessed for admittance in 2005-2011 (n = 127) were categorized into four trajectory groups based on whether they were admitted or denied (n = 19), discharged (n = 31), readmitted (n = 21) or had been undergoing OMT without interruption (n = 56). Second, 99 of these, the analytical sample, were interviewed at follow-up using (a) the Addiction Severity Index (ASI) for seven problem-areas and housing, and (b) self-rated change in 11 problem areas. The ASI was compared to baseline interviews after 55 months (mean). Third, outcomes within groups was studied in relation to alternative interventions. Within the analytical sample, those denied OMT showed no improvements at group level, those discharged had some improvements, more if readmitted than if not and those with uninterrupted OMT showed the most comprehensive improvements. Those outside OMT, denied and discharged, had considerable mortality risks related to ongoing drug use, especially in lack of well-planned alternative interventions. Improvements strongly relate to access to OMT. This study underscores that access to OMT improves the situation in all areas investigated and decreases the risk for drug-related death. It underscores the importance of two major risk situations, i.e. being denied OMT and being discharged.


Leukocyte subpopulations elicited by a nontumorigenic variant of B16 melanoma: their role in direct rejection of the melanoma and in prevention of tumorigenesis in Winn assays.
The mechanisms by which various leukocyte subpopulations elicited by an immunogenic, nontumorigenic subclone (C3471) of B16 melanoma caused rejection of the tumorigenic parental melanoma (B559), were investigated. Leukocytes from C3471-immune mice were co-injected with B559 tumor cells in Winn assays into normal syngeneic recipients. Tumor formation by B559 cells was prevented when C3471-immune (a) unfractionated peritoneal leukocytes, or (b) glass-adherent peritoneal cells (90% macrophages), or (c) nylon wool purified nonadherent cells (95% Thy-1.2+) were used in the Winn assays. If the C3471-immunized mice were treated with antithymocyte serum before harvest of their peritoneal cells, none of these leukocyte populations were effective in the Winn assay. However, macrophages from these immunologically compromised donors regained their tumoricidal activity after incubation in vitro with T lymphocytes from untreated C3471-immune donors; similarly, C3471-immune lymphocytes rendered normal resident peritoneal macrophages tumoricidal in Winn assays. When C3471-immunized mice were irradiated or treated with antithymocyte serum before direct challenge with B559 cells, melanomas developed, thus providing additional evidence for the need for intact T cell function to establish immunity against the melanoma. Furthermore, when Winn assay recipients were treated with antithymocyte serum, neither C3471-immune macrophages nor T cells were able to prevent tumor formation. These findings indicate that antithymocyte serum-sensitive (Thy-1.2+) lymphocytes are necessary both for the generation of tumoricidal leukocytes in C3471-immunized mice, and for the rejection of B559 melanoma by demonstrably tumoricidal macrophages in Winn assay recipients. In addition, long-lasting immunity developed in 50% of the normal mice that had received both C3471-immune peritoneal cells and B559 tumor cells, as manifested by their capacity to reject a second challenge with B559 cells 40-60 d later.


Influence of Rainfall on E. Coli Concentrations in Clams: Results of Collaboration Between Competent Health Authority and Producers' Association in the Province of Fermo (Italy).
The Area Vasta di Fermo (the competent health authority of the Marche Region), in collaboration with the local producers' Association conducted a series of studies on the presence of E. coli as an indicator of faecal contamination in the environment, in shellfish and in fishery waters, and on the factors that can affect their presence. These studies, carried out from 2008 to 2011, included an assessment of the currents along the coast, of the precipitations, and data from the monitoring of E. coli on shellfish harvested in the collection areas were examined. The results showed that in most cases, small concentrations of microorganisms in shellfish corresponded to little or no precipitations, while an increase in the levels of E. coli was preceded by more or less abundant rainfalls. The conclusions suggest that it is advisable to carry out a more detailed risk analysis which should take into account the above-mentioned factors. Furthermore, monitoring alone based on the determination of indicator organisms, especially when carried out as a single analysis or with a small number of E. coli determinations, does not provide a satisfactory indication of safety. The regional surveillance plans should be applied timely and rigorously, together with onsite investigations aimed at identifying changes that can affect the presence of E. coli in shellfish. Food business operators themselves could implement good manufacturing practices to verify whether the microbiological parameters are within the prescribed limits after rainfalls, especially if heavy.


Symmetric diffusion equations, barodiffusion, and cross-diffusion in concentrated liquid mixtures.
In models of diffusion in multicomponent mixtures, the current practice is to derive equations for an isobaric system. The equations are nonsymmetric in relation to the components of the mixture, and the concentration of solvent is assumed to be governed by the conservation of mass instead of its own corresponding diffusion equation. For concentrated mixtures, the solvent component is selected arbitrarily, which makes interpretation of the experimental data dependent on the choice of the interpreter. In this work, we derive a symmetric system of equations, made possible by the introduction of a spontaneously produced secondary pressure gradient. The effect of that pressure gradient is barodiffusion (barophoresis), defined by the force expressed as the secondary pressure gradient multiplied by the molecular volume. The model also considers the cross-diffusion (diffusiophoresis) that results from the hydrodynamic stresses associated with the local concentration-induced pressure gradient in liquid layers surrounding individual molecules. The resulting system of diffusion equations, which contains the secondary pressure gradient and component concentrations, is applied to a binary (nonionic) mixture of benzene and 1,2-dichloroethane. The steady-state system is placed in a uniform force field, and the effect of the secondary pressure gradient on the field-induced migration is discussed. Fluctuation dynamics in a system with no external force field is also discussed. The numerical results predict the establishment of lower concentration gradients compared to standard theory. Also, the predicted concentration dependence in the effective diffusion coefficient measured by dynamic light scattering is different compared to standard theory. Finally, experiments are proposed to further evaluate differences between the new model and the standard approach.


The relationship of depressive symptoms to parenting competence and social support in inner-city mothers of young children.
Despite the high prevalence of maternal depression and its negative consequences for children, many pediatricians fail to identify this problem. Our goal was to determine whether simple questions about parenting competence and the adequacy of maternal social support might be useful to providers in determining which inner-city mothers are likely to be depressed. We surveyed a convenience sample of 279 English-speaking mothers of children 6 months to 3 years old prior to a routine visit at an urban, hospital-based general pediatrics clinic. The mothers self-completed the Psychiatric Symptom Index (PSI) and the Parenting Stress Index Sense of Competence subscale, and rated the adequacy of their social support, and provided health and sociodemographic data by face-to-face interview. 41% of mothers had "high" PSI symptom levels and 22% had scores above a criterion that suggests major depressive disorder. In addition, 15% experienced high parenting stress (low competence) and 42% reported little or no social support. High distress was unrelated to a variety of sociodemographic risk factors, but significantly associated with a poor sense of parenting competence (Adj. OR = 3.3, 95% CI = 1.5, 7.0) and inadequate perceived social support (Adj. OR = 2.3, 95% CI = 1.2, 4.4), as well as with having health-related activity limitations (Adj. OR = 3.2, 95% CI = 1.1, 9.0). Negative ratings of parenting competence, low perceived social support, and presence of health-related activity restrictions can be useful markers of likely depression among inner-city mothers of young children. These factors are often assessed during routine pediatric visits and may be helpful to pediatricians in identifying mothers needing further evaluation or treatment by mental health specialists.


The DEAD-box RNA helicase Vad1 regulates multiple virulence-associated genes in Cryptococcus neoformans.
The study of fungal regulatory networks is essential to the understanding of how these pathogens respond to host environmental signals with effective virulence-associated traits. In this study, a virulence-associated DEAD-box RNA helicase-encoding gene (VAD1) was isolated from a mutant defective in the virulence factor laccase. A Deltavad1 mutant exhibited a profound reduction in virulence in a mouse model that was restored after reconstitution with WT VAD1. Loss of VAD1 resulted in upregulation of NOT1, a gene encoding a global repressor of transcription. NOT1 was found to act as an intermediary transcriptional repressor of laccase. Vad1 was located within macromolecular complexes that formed cytoplasmic granular bodies in mature cells and during infection of mouse brain. In addition, VAD1 was shown by in situ hybridization to be expressed in the brain of an AIDS patient coinfected with C. neoformans. To understand the role of VAD1 in virulence, a functional genomics approach was used to identify 3 additional virulence determinants dependent on VAD1: PCK1, TUF1, and MPF3, involved in gluconeogenesis, mitochondrial protein synthesis, and cell wall integrity, respectively. These data show that fungal virulence-associated genes are coordinately regulated and that an analysis of such transcriptomes allows for the identification of important new genes involved in the normal growth and virulence of fungal pathogens.


Fresnel filtering in lasing emission from scarred modes of wave-chaotic optical resonators.
We study lasing emission from asymmetric resonant cavity GaN microlasers. By comparing far-field intensity patterns with images of the microlaser we find that the lasing modes are concentrated on three-bounce unstable periodic ray orbits; i.e., the modes are scarred. The high-intensity emission directions of these scarred modes are completely different from those predicted by applying Snell's law to the ray orbit. This effect is due to the process of "Fresnel filtering" which occurs when a beam of finite angular spread is incident at the critical angle for total internal reflection.


Rapid feline immunodeficiency virus provirus quantitation by polymerase chain reaction using the TaqMan fluorogenic real-time detection system.
An improved quantitative polymerase chain reaction (qPCR) method based on a combination of real-time detection and the 5'-3' nuclease activity of the Taq DNA polymerase was developed to quantify the provirus load of feline immunodeficiency virus (FIV), a lentivirus of veterinary importance and an animal model for AIDS research. Two fluorogenic probes were designed to detect FIV provirus in genomic DNA of peripheral lymphocytes and tissues infected with different FIV subtypes. The most sensitive assay can detect one copy of FIV provirus. The assay showed excellent precision within-run and between-runs. Comparison of the TaqMan system with a conventional seminested PCR assay revealed a comparable detection limit and good correlation. Furthermore the design of the two probes allowed the detection of various FIV isolates of clade A and B.


A longitudinal study of back pain and radiological changes in the lumbar spines of middle aged women. II. Radiographic findings.
The natural history of radiological changes in the lumbar spine was evaluated in two groups of middle aged Dutch women selected from the general population. One group (n = 236) had recurrent back pain and the other (n = 241) had never experienced back pain. At the beginning of the study disc degeneration was more common in the group with back pain. Osteoporotic vertebral fractures were equally common in both groups. Nine years later both groups showed an increase in prevalence of disc degeneration and osteoporotic fractures. The strongest predictor for change in disc degeneration was the presence of degeneration at the beginning of the study. The development of disc degeneration for the first time was related to body mass index.


High frequency of recurrent viremia after hepatitis B e antigen seroconversion and consolidation therapy.
The primary treatment endpoint for hepatitis B e antigen (HBeAg)-positive chronic hepatitis B is HBeAg seroconversion; however, data on the durability of response are inconsistent. Our goal was to investigate the rate of recurrent viremia after HBeAg seroconversion and subsequent discontinuation of therapy. We retrospectively studied 88 consecutive Asian American patients who achieved HBeAg seroconversion [loss of HBeAg and development of antibody to HBeAg (anti-HBe)] among 458 HBeAg-positive patients who received oral antiviral therapy at 3 US clinics between March 1998 and November 2010. Recurrent viremia was defined as reappearance of detectable serum hepatitis B virus DNA (>100 IU/mL) on 2 consecutive laboratory tests from previously undetectable levels. Antiviral medications used at the time of HBeAg seroconversion included: lamivudine (23%), adefovir (34%), entecavir (36%), tenofovir (4%), and combination therapy (3%). Antiviral therapy was continued after HBeAg seroconversion in 49 patients (group I) and discontinued in the other 39 patients after consolidation therapy [median=12 months (range, 1 to 55 mo)] (group II). No patients in group I experienced recurrent viremia, whereas 90% in group II did. Elevated alanine aminotransferase also occurred in 38% of group II patients [median peak alanine aminotransferase 249 IU/mL (range, 93 to 1070 IU/mL)]. Despite consolidation therapy, almost all patients who discontinued therapy after achieving HBeAg seroconversion and complete viral suppression experienced recurrent viremia, and close to half also experienced biochemical flares. HBeAg seroconversion does not seem to be a durable treatment endpoint for many patients, and they should be monitored carefully for virologic relapse and biochemical flares if antiviral therapy is withdrawn.


Mortality of United Kingdom acrylonitrile workers--an extended and updated study.
The mortality experience of 2763 men employed between 1950 and 1978 for at least 1 year at 6 factories involved in the polymerization of acrylonitrile and the spinning of acrylic fiber was followed to the end of 1991. Overall, cancer deaths did not exceed the expected numbers. There were, however, excess cancer deaths among the workers in the jobs more highly exposed to acrylonitrile. The excesses did not reach conventional levels of statistical significance apart from an excess of lung cancer among workers under 45 years of age. Detailed analyses provided no consistent support for a causal association between acrylonitrile exposure and carcinogenesis. The limitations of the study, including a lack of information on smoking habits and very limited estimates of acrylonitrile exposure, need to be borne in mind.


The Effect of Noseband Tightening on Horses' Behavior, Eye Temperature, and Cardiac Responses.
Restrictive nosebands are common in equestrian sport. This is concerning, as recent evidence suggests that very tight nosebands can cause a physiological stress response, and may compromise welfare. The objective of the current study was to investigate relationships that noseband tightness has with oral behavior and with physiological changes that indicate a stress response, such as increases in eye temperature (measured with infrared thermography) and heart rate and decreases in heart rate variability (HRV). Horses (n = 12) wearing a double bridle and crank noseband, as is common in dressage at elite levels, were randomly assigned to four treatments: unfastened noseband (UN), conventional area under noseband (CAUN) with two fingers of space available under the noseband, half conventional area under noseband (HCAUN) with one finger of space under the noseband, and no area under the noseband (NAUN). During the tightest treatment (NAUN), horse heart rate increased (P = 0.003), HRV decreased (P < 0.001), and eye temperature increased (P = 0.011) compared with baseline readings, indicating a physiological stress response. The behavioral results suggest some effects from bits alone but the chief findings are the physiological readings that reflect responses to the nosebands at their tightest. Chewing decreased during the HCAUN (P < 0.001) and NAUN (P < 0.001) treatments. Yawning rates were negligible in all treatments. Similarly, licking was eliminated by the NAUN treatment. Following the removal of the noseband and double bridle during the recovery session, yawning (P = 0.015), swallowing (P = 0.003), and licking (P < 0.001) significantly increased compared with baseline, indicating a post-inhibitory rebound response. This suggests a rise in motivation to perform these behaviors and implies that their inhibition may place horses in a state of deprivation. It is evident that a very tight noseband can cause physiological stress responses and inhibit the expression of oral behaviors.


IL-21 and CD40L synergistically promote plasma cell differentiation through upregulation of Blimp-1 in human B cells.
After undergoing Ig somatic hypermutation and Ag selection, germinal center (GC) B cells terminally differentiate into either memory or plasma cells (PCs). It is known that the CD40L and IL-21/STAT3 signaling pathways play critical roles in this process, yet it is unclear how the B cell transcription program interprets and integrates these two types of T cell-derived signals. In this study, we characterized the role of STAT3 in the GC-associated PC differentiation using purified human tonsillar GC B cells and a GC B cell-like cell line. When primary GC B cells were cultured under PC differentiation condition, STAT3 inhibition by AG490 prevented the transition from GC centrocytes to preplasmablast, suggesting that STAT3 is required for the initiation of PC development. In a GC B cell-like human B cell line, although IL-21 alone can induce low-level Blimp-1 expression, maximum Blimp-1 upregulation and optimal PC differentiation required both IL-21 and CD40L. CD40L, although having no effect on Blimp-1 as a single agent, greatly augmented the amplitude and duration of IL-21-triggered Jak-STAT3 signaling. In the human PRDM1 locus, CD40L treatment enhanced the ability of STAT3 to upregulate Blimp-1 by removing BCL6, a potent inhibitor of Blimp-1 expression, from a shared BCL6/STAT3 site in intron 3. Thus, IL-21 and CD40L collaborate through at least two distinct mechanisms to synergistically promote Blimp-1 activation and PC differentiation.


Perceived adverse effects of antiretroviral therapy.
Adverse effects from antiretroviral therapy (ARV) for HIV are associated with medication nonadherence. The purposes of this study were to explore group differences in the reporting of adverse effects, identify individual adverse effects that are linked to nonadherence, and to explore the role of coping in the relationship between adverse effects and adherence. Cross-sectional interviews of 2,765 HIV-positive adults on ARV therapies in four U.S. cities were performed using a computerized assessment of self-reported adverse effects, coping self-efficacy, and adherence. There were no gender differences in the rate or severity of adverse effects reported. Latino respondents reported more adverse effects than either White or African Americans. Those taking a protease inhibitor (PI) reported a higher rate and greater severity of adverse effects. Older participants reported fewer adverse effects despite being more likely to be on a regimen containing a PI. Respondents with less than 90% adherence reported greater numbers and severity of adverse effects overall. In multivariate analyses, nausea, skin problems, vomiting, and memory adverse effects were independently related to less than 90% adherence over the prior three days. Coping moderated the relationship between nausea and adherence such that individuals who reported lower coping self-efficacy and experienced nausea were at increased risk for nonadherence, regardless of the length of time on the current ARV regimen. Women and men are similar in their overall reports of adverse effects, and Latinos report more adverse effects to ARVs than White or African American patients. Specific adverse effects (skin problems, memory problems, vomiting, and nausea) are more likely than others to be associated with missing ARV medications. Increasing adaptive coping self-efficacy among patients experiencing nausea may be a particularly effective strategy in increasing medication adherence.


Estimation of extracellular volume from regularized multi-shell diffusion MRI.
Diffusion MRI measures micron scale displacement of water molecules, providing unique insight into microstructural tissue architecture. However, current practical image resolution is in the millimeter scale, and thus diffusivities from many tissue compartments are averaged in each voxel, reducing the sensitivity and specificity of the measurement to subtle pathologies. Recent studies have pointed out that eliminating the contribution of extracellular water increases the sensitivity of the diffusion measures to tissue architecture. Moreover, in brain imaging, estimation of the extracellular volume appears to indicate pathological processes such as atrophy, edema and neuroinflammation. Here we study the free-water method, which assumes a bi-tensor model. We add low b-value shells to a regular DTI acquisition and present methods to improve the estimation of the model parameters using the extra information. In addition, we define a Laplace-Beltrami regularization operator that further stabilizes the multi-shell estimation.


Resistance to Phytophthora citrophthora and P. parasitica and Nursery Characteristics of Several Citrus Rootstocks.
Studies were conducted to compare existing and potential citrus rootstocks with respect to resistance to root rot and gummosis caused by Phytophthora citrophthora and P. parasitica in greenhouse and growth chamber experiments and horticultural performance under simulated nursery conditions. Depending upon rootstock and experiment, mean root weights resulting from inoculation with P. citrophthora were 27 to 96% lower than the comparable controls. In similar experiments with the same rootstocks, inoculation with P. parasitica resulted in root weights that were 38 to 95% less than weights of the noninoculated controls. During 1994 or 1995, mean root weight reduction compared with noninoculated plants among Citrus macrophylla, rough lemon, C. volkameriana, and Sunki mandarin × Flying Dragon trifoliate (62-109-19) attributable to P. citrophthora and mean root weight reduction among C. macrophylla, C. volkameriana, rough lemon, Sacaton citrumelo, Sunki mandarin × Flying Dragon trifoliate (62-109-19), African shaddock × Rubidoux trifoliate, and Shekwasha mandarin × English trifoliate attributable to P. parasitica were significantly less than those recorded for all other tested rootstocks. Rootstocks that sustained a low percentage of root weight reduction generally experienced a low percentage of shoot weight reduction and survived longer as well. In evaluation of resistance to gummosis, depending on rootstock and experiment, the mean length of stem lesions caused by P. citrophthora on rootstocks ranged from 0.2 to 25.0 mm, whereas values for P. parasitica ranged from 0.2 to 18.5 mm. Stem lesions smaller than 5 mm in length were recorded for 21 and 14 of 36 different rootstocks inoculated with P. citrophthora and P. parasitica, respectively. On the other hand, P. citrophthora and P. parasitica caused stem lesions of at least 10 mm in length on 8 and 16 citrus rootstocks, respectively. Desirable nursery characteristics, including vigorous growth, minimal branching, and high leaf chlorophyll content, were demonstrated most prominently by Gomiri rough lemon, C. volkameriana, and Benton citrange, and to a lesser degree by some other rootstocks. Possible factors that could account for inconsistent classification of some citrus rootstocks as susceptible or resistant to Phytophthora root rot and gummosis are discussed.


Using functional neuroimaging combined with a think-aloud protocol to explore clinical reasoning expertise in internal medicine.
Clinical reasoning is essential to medical practice, but because it entails internal mental processes, it is difficult to assess. Functional magnetic resonance imaging (fMRI) and think-aloud protocols may improve understanding of clinical reasoning as these methods can more directly assess these processes. The objective of our study was to use a combination of fMRI and think-aloud procedures to examine fMRI correlates of a leading theoretical model in clinical reasoning based on experimental findings to date: analytic (i.e., actively comparing and contrasting diagnostic entities) and nonanalytic (i.e., pattern recognition) reasoning. We hypothesized that there would be functional neuroimaging differences between analytic and nonanalytic reasoning theory. 17 board-certified experts in internal medicine answered and reflected on validated U.S. Medical Licensing Exam and American Board of Internal Medicine multiple-choice questions (easy and difficult) during an fMRI scan. This procedure was followed by completion of a formal think-aloud procedure. fMRI findings provide some support for the presence of analytic and nonanalytic reasoning systems. Statistically significant activation of prefrontal cortex distinguished answering incorrectly versus correctly (p < 0.01), whereas activation of precuneus and midtemporal gyrus distinguished not guessing from guessing (p < 0.01). We found limited fMRI evidence to support analytic and nonanalytic reasoning theory, as our results indicate functional differences with correct vs. incorrect answers and guessing vs. not guessing. However, our findings did not suggest one consistent fMRI activation pattern of internal medicine expertise. This model of employing fMRI correlates offers opportunities to enhance our understanding of theory, as well as improve our teaching and assessment of clinical reasoning, a key outcome of medical education.


Genomics of Dementia: APOE- and CYP2D6-Related Pharmacogenetics.
Dementia is a major problem of health in developed societies. Alzheimer's disease (AD), vascular dementia, and mixed dementia account for over 90% of the most prevalent forms of dementia. Both genetic and environmental factors are determinant for the phenotypic expression of dementia. AD is a complex disorder in which many different gene clusters may be involved. Most genes screened to date belong to different proteomic and metabolomic pathways potentially affecting AD pathogenesis. The ε4 variant of the APOE gene seems to be a major risk factor for both degenerative and vascular dementia. Metabolic factors, cerebrovascular disorders, and epigenetic phenomena also contribute to neurodegeneration. Five categories of genes are mainly involved in pharmacogenomics: genes associated with disease pathogenesis, genes associated with the mechanism of action of a particular drug, genes associated with phase I and phase II metabolic reactions, genes associated with transporters, and pleiotropic genes and/or genes associated with concomitant pathologies. The APOE and CYP2D6 genes have been extensively studied in AD. The therapeutic response to conventional drugs in patients with AD is genotype specific, with CYP2D6-PMs, CYP2D6-UMs, and APOE-4/4 carriers acting as the worst responders. APOE and CYP2D6 may cooperate, as pleiotropic genes, in the metabolism of drugs and hepatic function. The introduction of pharmacogenetic procedures into AD pharmacological treatment may help to optimize therapeutics.


Fate of the mammalian cranial neural crest during tooth and mandibular morphogenesis.
Neural crest cells are multipotential stem cells that contribute extensively to vertebrate development and give rise to various cell and tissue types. Determination of the fate of mammalian neural crest has been inhibited by the lack of appropriate markers. Here, we make use of a two-component genetic system for indelibly marking the progeny of the cranial neural crest during tooth and mandible development. In the first mouse line, Cre recombinase is expressed under the control of the Wnt1 promoter as a transgene. Significantly, Wnt1 transgene expression is limited to the migrating neural crest cells that are derived from the dorsal CNS. The second mouse line, the ROSA26 conditional reporter (R26R), serves as a substrate for the Cre-mediated recombination. Using this two-component genetic system, we have systematically followed the migration and differentiation of the cranial neural crest (CNC) cells from E9.5 to 6 weeks after birth. Our results demonstrate, for the first time, that CNC cells contribute to the formation of condensed dental mesenchyme, dental papilla, odontoblasts, dentine matrix, pulp, cementum, periodontal ligaments, chondrocytes in Meckel's cartilage, mandible, the articulating disc of temporomandibular joint and branchial arch nerve ganglia. More importantly, there is a dynamic distribution of CNC- and non-CNC-derived cells during tooth and mandibular morphogenesis. These results are a first step towards a comprehensive understanding of neural crest cell migration and differentiation during mammalian craniofacial development. Furthermore, this transgenic model also provides a new tool for cell lineage analysis and genetic manipulation of neural-crest-derived components in normal and abnormal embryogenesis.


The anti-inflammatory potential of diet and nonalcoholic fatty liver disease: the ATTICA study.
Nonalcoholic fatty liver disease (NAFLD) is correlated with low-grade inflammation and dietary habits. Until today, there have been limited epidemiologic data assessing the role of diet's inflammatory potential on NAFLD. The aim was to evaluate the relationship between an anti-inflammatory diet, as reflected by the Dietary Anti-Inflammation Index (D-AII), and NAFLD among cardiovascular disease (CVD)-free adults. ATTICA is a prospective, population-based study that recruited 3042 adults without pre-existing CVD from the Greek population (Whites; age ⩾18 years; 1514 men and 1528 women). D-AII was calculated using a standard procedure. The baseline study captured various sociodemographic, lifestyle and clinical characteristics as well as hepatic markers. These were used to calculate four NAFLD assessment indices: triglyceride-glucose (TyG) index, fatty liver index (FLI), hepatic steatosis index (HSI), and NAFLD Fatty Liver Score (NAFLD-FLS). Specific cutoffs were applied to capture NAFLD. D-AII showed a significant inverse association with NAFLD, applying the four indices with NAFLD cutoffs [odds ratio (OR) with 95% confidence interval (CI); TyG (0.95, 0.93-0.98); HSI (0.89, 0.86-0.92); FLI (0.88, 0.85-0.91); NAFLD-FLS (0.89, 0.86-0.92)], after adjusting for various confounders. Participants in the highest D-AII tertile had lower odds of having NAFLD, compared with those in the lowest D-AII tertile [(OR, 95% CI); TyG (0.33, 0.24-0.47); HSI (0.13, 0.08-0.23); FLI (0.05, 0.02-0.11); NAFLD-FLS (0.13, 0.07-0.23)]. Anti-inflammatory nutrition was related to lower odds of NAFLD among daily alcohol drinkers and individuals with metabolic syndrome. Anti-inflammatory diet is an important predictor of NAFLD among adults without pre-existing CVD. Adherence to a high anti-inflammatory diet seems to contribute to NAFLD prevention.


Perspective: medical education research and the institutional review board: reexamining the process.
Medical school and residency training curricula across the country have undergone extensive revisions and, much like clinical quality improvement (QI) initiatives, require assessments of new programs. Because sharing knowledge is a hallmark of academic medicine, program evaluation may come under the purview of the institutional review board (IRB); however, the distinction between QI and research is often unclear. And yet a medical education (ME) inquiry can be designed according to either paradigm. The purpose of this article is to bring IRBs and ME researchers closer to a shared understanding of key concepts underlying human participation in research and QI activities, and to consensus on the application of these concepts. The current QI discourse provides a useful framework for making this distinction; the authors identify key theoretical principles and practical considerations derived from this work that are relevant to ME and training, such as the application of the regulatory definition of human subject research to ME inquiries. For ME inquiries defined as human subject research, and therefore subject to IRB review, this article explores the application of the human research regulations to ME research. It concludes with practical suggestions for institutions, IRBs, and ME researchers, which range from formal procedures for making the QI versus research distinction, to instruction in study design and development and the human subject regulatory implications. The intent is to promote a discussion that will result in greater consensus and a more consistent application of the regulatory framework.


Identification of an Immunomodulating Agent from Mycobacterium leprae.
A search for an immunomodulating agent from mycobacteria was carried out using Mycobacterium leprae. The antigenicity of each fraction of the bacterial membrane, which contains the most antigenic components of M. leprae, was assessed by using sera from paucibacillary leprosy. N-terminal sequencing of the serum-reactive protein and functional assessment of the membrane fractions using monocyte-derived dendritic cells (DCs) identified major membrane protein II (MMP-II) as one of the efficient T-cell-activating candidates. Purified MMP-II stimulated DCs from healthy individuals to produce interleukin-12 p70 and up-regulated the surface expression of major histocompatibility complex class I and II, CD86, and CD83 molecules. Also, there was an increase in the percentage of CD83(+) cells in the DC population. Furthermore, MMP-II-pulsed DCs expressed their derivatives on their surfaces. Using Toll-like receptor 2 (TLR-2)-dependent receptor constructs, we found that TLR-2 signaling was involved in DC maturation induced by MMP-II. Taken together, MMP-II can be recognized as an immunomodulating protein in terms of activation of antigen-presenting cells and innate immunity.


Contributions of Kv3 channels to neuronal excitability.
Four mammalian Kv3 genes have been identified, each of which generates, by alternative splicing, multiple protein products differing in their C-terminal sequence. Products of the Kv3.1 and Kv3.2 genes express similar delayed-rectifier type currents in heterologous expression systems, while Kv3.3 and Kv3.4 proteins express A-type currents. All Kv3 currents activate relatively fast at voltages more positive than -10 mV, and deactivate very fast. The distribution of Kv3 mRNAs in the rodent CNS was studied by in situ hybridization, and the localization of Kv3.1 and Kv3.2 proteins has been studied by immunohistochemistry. Most Kv3.2 mRNAs (approximately 90%) are present in thalamic-relay neurons throughout the dorsal thalamus. The protein is expressed mainly in the axons and terminals of these neurons. Kv3.2 channels are thought to be important for thalamocortical signal transmission. Kv3.1 and Kv3.2 proteins are coexpressed in some neuronal populations such as in fast-spiking interneurons of the cortex and hippocampus, and neurons in the globus pallidus. Coprecipitation studies suggest that in these cells the two types of protein form heteromeric channels. Kv3 proteins appear to mediate, in native neurons, similar currents to those seen in heterologous expression systems. The activation voltage and fast deactivation rates are believed to allow these channels to help repolarize action potentials fast without affecting the threshold for action potential generation. The fast deactivating current generates a quickly recovering after hyperpolarization, thus maximizing the rate of recovery of Na+ channel inactivation without contributing to an increase in the duration of the refractory period. These properties are believed to contribute to the ability of neurons to fire at high frequencies and to help regulate the fidelity of synaptic transmission. Experimental evidence has now become available showing that Kv3.1-Kv3.2 channels play critical roles in the generation of fast-spiking properties in cortical GABAergic interneurons.


Circadian variation in ictus of aneurysmal subarachnoid hemorrhage.
Temporal patterns in aneurysmal subarachnoid hemorrhage (aSAH) may provide insight into modulation, and therefore, prevention of hemorrhage. We investigated the time of hemorrhage and its relationship to traditional risk factors among patients admitted with aSAH. Admitted patients with aSAH were prospectively followed through outcomes and baseline demographics were abstracted through chart review. The group temporal distribution by hour of onset was summarized with cosinor nonlinear least squares. aSAH onset was gathered into night (2300-0500), morning (0500-1100), afternoon (1100-1700), and evening (0500-2300) daily phases. The odds ratio (OR) with 95% CI was calculated for having an aSAH during the morning, afternoon, and evening hours using night as a reference. Multinomial logit models were fitted using aSAH cases across time blocks to determine their associations with different risk factors. 202 patients had the hour of hemorrhage available, and 49 had phase identifiable [total 251: 38 (15%) night, 98 (39%) morning, 58 (23%) afternoon, 57 (23%) evening]. The peak hours of aSAH were between 0700 and 0800 representing 13% of the sample, with a significant cosinor-fitted phase of 7.33(95% CI 5.30, 9.36). For all aSAH cases, morning onset was significantly more common than night onset (OR = 2.58, 95% CI = 1.77-3.75). Nonsmokers were more likely to have aSAH in the morning than smokers (P = 0.043, OR = 3.10, 95% CI = 1.33-7.23). aSAH occur in a diurnal, morning prevalent pattern regardless of traditional aSAH risk factors. The association of these risk factors with existing onset patterns should be investigated in future studies.


A plausible model correlates prebiotic peptide synthesis with the primordial genetic code.
Aminoacyladenylates (5'-aa-AMPs) are key intermediates in peptide synthesis. Here we report analogs of 5'-aa-AMPs, namely nucleotide amidates (aa-N-NMPs), obtained under Hadean conditions. Significantly, dipeptides were detected from the above reactions and their yields varied with different nucleosides through the formation of different aa-N-NMPs. This model provides both prebiotic peptides and the primordial version of the genetic code through reactions that occurred under potentially prebiotic conditions.


A simultaneous equations model of crash frequency by severity level for freeway sections.
This paper presents a simultaneous equations model of crash frequencies by severity level for freeway sections using five-year crash severity frequency data for 275 multilane freeway segments in the State of Washington. Crash severity is a subject of much interest in the context of freeway safety due to higher speeds of travel on freeways and the desire of transportation professionals to implement measures that could potentially reduce crash severity on such facilities. This paper applies a joint Poisson regression model with multivariate normal heterogeneities using the method of Maximum Simulated Likelihood Estimation (MSLE). MSLE serves as a computationally viable alternative to the Bayesian approach that has been adopted in the literature for estimating multivariate simultaneous equations models of crash frequencies. The empirical results presented in this paper suggest the presence of statistically significant error correlations across crash frequencies by severity level. The significant error correlations point to the presence of common unobserved factors related to driver behavior and roadway, traffic and environmental characteristics that influence crash frequencies of different severity levels. It is found that the joint Poisson regression model can improve the efficiency of most model coefficient estimators by reducing their standard deviations. In addition, the empirical results show that observed factors generally do not have the same impact on crash frequencies at different levels of severity.


Identification of human Asf1 chromatin assembly factors as substrates of Tousled-like kinases.
First described in Arabidopsis thaliana, Tousled-like kinases (Tlks) are highly conserved in both plants and animals. In plants, Tousled kinase is essential for proper flower and leaf development, but no direct functional link to any other plant gene product has yet been established. Likewise, the role of Tlks in animals is unknown. In human cells, two structurally similar Tlks, Tlk1 and Tlk2, were recently shown to be cell cycle-regulated kinases with maximal activities during S phase. Here, we report the identification of two human homologs of the Drosophila chromatin assembly factor Asf1 (anti-silencing function 1) as physiological substrates of Tlks. We show that human Asf1 proteins are phosphorylated by Tlks both in vivo and in vitro. Furthermore, Asf1 proteins are phosphorylated during S phase, when Tlks are maximally active. Conversely, Asf1 proteins are dephosphorylated upon the activation of the DNA replication checkpoint, concomitant with the rapid inactivation of Tlks. These data indicate that Tlk family members regulate chromatin assembly during DNA replication, and they suggest a plausible explanation for the pleiotropic developmental defects of plant tousled mutants.


Abdominal obesity and the risk of all-cause, cardiovascular, and cancer mortality: sixteen years of follow-up in US women.
Accumulating evidence indicates that abdominal adiposity is positively related to cardiovascular disease (CVD) risk and some other diseases independently of overall adiposity. However, the association of premature death resulting from these diseases with abdominal adiposity has not been widely studied, and findings are inconsistent. In a prospective cohort study of 44,636 women in the Nurses' Health Study, associations of abdominal adiposity with all-cause and cause-specific mortality were examined. During 16 years of follow-up, 3507 deaths were identified, including 751 cardiovascular deaths and 1748 cancer deaths. After adjustment for body mass index and potential confounders, the relative risks across the lowest to the highest waist circumference quintiles were 1.00, 1.11, 1.17, 1.31, and 1.79 (95% confidence interval [CI], 1.47 to 1.98) for all-cause mortality; 1.00, 1.04, 1.04, 1.28, and 1.99 (95% CI, 1.44 to 2.73) for CVD mortality; and 1.00, 1.18, 1.20, 1.34, and 1.63 (95% CI, 1.32 to 2.01) for cancer mortality (all P<0.001 for trend). Among normal-weight women (body mass index, 18.5 to < 25 kg/m(2)), abdominal obesity was significantly associated with elevated CVD mortality: Relative risk associated with waist circumference > or = 88 cm was 3.02 (95% CI, 1.31 to 6.99) and for waist-to-hip ratio > 0.88 was 3.45 (95% CI, 2.02 to 6.92). After adjustment for waist circumference, hip circumference was significantly and inversely associated with CVD mortality. Anthropometric measures of abdominal adiposity were strongly and positively associated with all-cause, CVD, and cancer mortality independently of body mass index. Elevated waist circumference was associated with significantly increased CVD mortality even among normal-weight women.


Prior short-term consumption of resistant starch enhances postprandial insulin sensitivity in healthy subjects.
Diets rich in insoluble-fibre are linked to a reduced risk of both diabetes and cardiovascular disease; however, the mechanism of action remains unclear. The aim of this study was to assess whether acute changes in the insoluble-fibre (resistant starch) content of the diet would have effects on postprandial carbohydrate and lipid handling. Ten healthy subjects consumed two identical, low-residue diets on separate occasions for 24 h (33% fat; <2 g dietary fibre). Of the diets one was supplemented with 60 g resistant starch (Novelose 260). On the following morning a fibre-free meal tolerance test (MTT) was carried out (59 g carbohydrate; 21 g fat; 2.1 kJ) and postprandial insulin sensitivity (SI(ORAL)) assessed using a minimal model approach. Prior resistant starch consumption led to lower postprandial plasma glucose (p=0.037) and insulin (p=0.038) with a higher insulin sensitivity(44+/-7.5 vs 26+/-3.5 x 10(-4) dl kg(-1) min(-1) per micro Uml(-1); p=0.028) and C-peptide-to-insulin molar ratio (18.7+/-6.5 vs 9.7+/-0.69; p=0.017). There was no effect of resistant starch consumption on plasma triacylglycerol although non-esterified fatty acid and 3-hydroxybutyrate levels were suppressed 5 h after the meal tolerance test. Prior acute consumption of a high-dose of resistant starch enhanced carbohydrate handling in the postprandial period the following day potentially due to the increased rate of colonic fermentation.


Food loss and waste management in Turkey.
Food waste can be an environmental and economic problem if not managed properly but it can meet various demands of a country if it is considered as a resource. The purpose of this report is to review the existing state of the field in Turkey and identify the potential of food waste as a resource. Food loss and waste (FLW) was examined throughout the food supply chain (FSC) and quantified using the FAO model. Edible FLW was estimated to be approximately 26milliontons/year. The amount of biodegradable waste was estimated based on waste statistics and research conducted on household food waste in Turkey. The total amount of biodegradable waste was found to be approximately 20milliontons/year, where more than 8.6milliontons/year of this waste is FLW from distribution and consumption in the FSC. Options for the end-of-life management of biodegradable wastes are also discussed in this review article.


Central command neurons of the sympathetic nervous system: basis of the fight-or-flight response.
During stress, the activity of the sympathetic nervous system is changed in a global fashion, leading to an increase in cardiovascular function and a release of adrenal catecholamines. This response is thought to be regulated by a common set of brain neurons that provide a dual input to the sympathetic preganglionic neurons regulating cardiac and adrenal medullary functions. By using a double-virus transneuronal labeling technique, the existence of such a set of central autonomic neurons in the hypothalamus and brainstem was demonstrated. These neurons innervate both of the sympathetic outflow systems and likely function in circumstances where parallel sympathetic processing occurs, such as in the fight-or-flight response.


Population structure, fitness surfaces, and linkage in the shifting balance process.
Wright first introduced the idea that random genetic drift and classical mass-action selection might combine in such a way as to allow populations to find the highest peak in complicated adaptive surfaces. His theory assumes large but structured populations, in which mating is spatially local. If gene flow is sufficiently low, and the subpopulations (demes) are small enough, they will be subject to genetic drift. Distant demes drift independently, allowing many independent searches of the adaptive surface to take place. A deme that has shifted to a higher peak can, by emigration, cause the rest of the demes to shift to the higher peak. The probability of this shift depends on the migration rate. Previous studies have concluded that very little migration is necessary to effect the shift in adaptive peaks that characterizes the last phase of Wright's Shifting Balance Process (SBP). Here we present the results of a computer study that investigates the roles of dispersal distance, the degree of epistasis in the fitness surface, and recombination on the shifting balance process. In particular, we measure their effect on the population's mean fitness. We show that over a range of dispersal distances the advantage of the SBP is a monotonically increasing function of the amount of epistasis. Our results show that the extent of dispersal that results in the greatest effect of the SBP in increasing mean fitness depends on the extent of epistasis. Finally, for low levels of epistasis, higher recombination performs better, while for intermediate levels, lower recombination results in a greater advantage of the SBP.


Suppressed tyrosine hydroxylase gene expression in the tuberoinfundibular dopaminergic system during lactation.
Suckling-induced PRL secretion is regulated in part by a reduction in tuberoinfundibular dopamine (TIDA) neuronal activity. We have examined the effects of suckling on TIDA activity in the arcuate nucleus by measuring changes in gene expression of tyrosine hydroxylase (TH), the rate-limiting enzyme in catecholamine synthesis. TH gene expression was assessed by performing in situ hybridization, using a 35S-labeled antisense riboprobe for quantitating TH mRNA and analyzing grain density with the aid of an Optimas Bioscan image analysis system. Lactating rats suckled by eight pups were studied on postpartum day 10, and diestrous day 1 rats were used as controls. The results showed that lactation suppressed TH mRNA content throughout the arcuate nucleus to about 10% of diestrous levels. The dramatic reduction in TH mRNA during lactation was specific to the arcuate nucleus, as TH mRNA levels in the zona incerta were similar during lactation and diestrus. The suckling stimulus was the primary signal responsible for the suppression of TH mRNA in the arcuate nucleus, as removal of the pups for 6 h restored TH mRNA content to diestrous levels. By 24 h after pup removal, TH mRNA had reached almost twice diestrous levels. In view of the dramatic reduction in TH mRNA levels during lactation, we examined whether TH protein in the arcuate nucleus was similarly diminished. TH protein was detected by immunocytochemistry using a monoclonal antibody to TH. Qualitatively, TH staining was heavier in cell bodies, nerve fibers, and median eminence during diestrus. There was a small, but significant, decrease in TH-positive cell numbers during lactation (14% reduction) compared to those on diestrus. These data provide clear evidence that TH expression is suppressed during lactation, as evidenced by the decrease in TH mRNA and TH protein. The reduction in TH expression most likely contributes to the decrease in dopaminergic tone during lactation.


Using isotemporal substitution to predict the effects of changing physical behaviour on older adults' cardio-metabolic profiles.
It has been advocated that older adults should concomitantly spend less time in sedentary behaviour (SB), and engage in sufficient physical activity (PA), to reduce their risk of cardio-metabolic diseases. However, it is not clear what intensity of PA must be done to offset SB engagement. Model how cardio-metabolic profiles could change if older adults replaced an hour per day (hr·day-1) of a physical behaviour intensity with 1 hr·day-1 of another physical behaviour of a different intensity. Older adults (n = 93, 60-89 years old, 55% female) wore a thigh-mounted triaxial accelerometer for seven consecutive free-living days to estimate mean daily hourly engagement in SB, Standing, Light Intensity PA (LIPA), sporadic moderate to vigorous physical activity (sMVPA, bouts <10 continuous minutes), and 10-minute MVPA (10MVPA, bouts ≥10 continuous minutes. Fasting whole blood concentration of plasma glucose, triglyceride, total cholesterol, and glycated haemoglobin (%), along with serum concentration of lipoprotein lipase (LPL), interleukin-6 (IL-6), and procollagen III N-terminal propeptide (PIIINP) were measured. Isotemporal Substitution, with covariate adjustment, suggested that: total cholesterol concentration could theoretically decrease when 1 hr·day-1 of SB is replaced with Standing, when 1 hr.day-1 of LIPA is replaced with Standing, and when 1 hr·day-1 of sMVPA is replaced with Standing. Triglyceride concentration theoretically decreased when 1 hr·day-1 of SB, Standing, LIPA, or sMVPA is replaced with 10MVPA. Triglyceride concentration theoretically increases when 1 hr·day-1 of 10MVPA is replaced with SB, Standing, or LIPA. No associations with time reallocation appears to exist for LPL, HbA1c, IL-6, and PIIINP. The type of physical behaviour being replaced could be crucial for total cholesterol maintenance. Engagement in 10MVPA could be necessary to improve triglyceride concentration.


Rescue of T cell-specific V(D)J recombination in SCID mice by DNA-damaging agents.
Assembly of antigen receptor V (variable), D (diversity), and J (joining) gene segments requires lymphocyte-specific genes and ubiquitous DNA repair activities. Severe combined immunodeficient (SCID) mice are defective in general double-strand (ds) DNA break repair and V(D)J coding joint formation, resulting in arrested lymphocyte development. A single treatment of newborn SCID mice with DNA-damaging agents restored functional, diverse, T cell receptor beta chain coding joints, as well as development and expansion of thymocytes expressing both CD4 and CD8 coreceptors, but did not promote B cell development. Thymic lymphoma developed in all mice treated with DNA-damaging agents, suggesting an interrelation between V(D)J recombination, dsDNA break repair, and lymphomagenesis.


Early Age of Alcohol Initiation and its Association with Suicidal Behaviors.
The relationship between alcohol use and suicidal behaviors is well-accepted, but less is known about the contribution of its early initiation. This study was designed to test the association of early alcohol initiation versus later initiation with suicidal ideation and attempt in an ethnically diverse sample. The Collaborative Psychiatric Epidemiology Surveys (CPES), 2001-2003 (n = 20,013), database was used. A total of 13,867 participants were selected included 56.9% females and 43.1% males. Race and ethnicity were reported as 28.8% non-Hispanic White, 39.1% Black, 20.3% Latino, and 11.9% Asian. Logistic regression analyses tested the associations between early (< =14 years) and later (> =15) age alcohol initiation with suicide ideation and attempts. Alcohol initiation was indexed by self-report of the first time that any alcohol product was consumed. Potential confounders were controlled. Early alcohol initiation was associated with higher odds (AOR = 3.64, 95% CI [2.51, 5.28]) of suicide ideation as compared with adults who had initiated > = age 15 (AOR = 2.11, 95% CI [1.46, 3.04]). Early age initiation was also associated with higher odds (AOR = 3.81, 95% CI [2.02, 7.18]) of lifetime suicide attempt versus later age initiators (AOR = 2.03, 95% CI [1.08, 3.79]). Significant differences were found between early and later age of initiation. Conclusion: Early age of alcohol initiation has profoundly increased odds of suicide ideation or attempt. It is critical that effective prevention programs for children and their caregivers be implemented to prevent or delay alcohol initiation and lessen the risk for future suicidal behaviors.


Cost-effective generation of precise label-free quantitative proteomes in high-throughput by microLC and data-independent acquisition.
Quantitative proteomics is key for basic research, but needs improvements to satisfy an increasing demand for large sample series in diagnostics, academia and industry. A switch from nanoflowrate to microflowrate chromatography can improve throughput and reduce costs. However, concerns about undersampling and coverage have so far hampered its broad application. We used a QTOF mass spectrometer of the penultimate generation (TripleTOF5600), converted a nanoLC system into a microflow platform, and adapted a SWATH regime for large sample series by implementing retention time- and batch correction strategies. From 3 µg to 5 µg of unfractionated tryptic digests that are obtained from proteomics-typical amounts of starting material, microLC-SWATH-MS quantifies up to 4000 human or 1750 yeast proteins in an hour or less. In the acquisition of 750 yeast proteomes, retention times varied between 2% and 5%, and quantified the typical peptide with 5-8% signal variation in replicates, and below 20% in samples acquired over a five-months period. Providing precise quantities without being dependent on the latest hardware, our study demonstrates that the combination of microflow chromatography and data-independent acquisition strategies has the potential to overcome current bottlenecks in academia and industry, enabling the cost-effective generation of precise quantitative proteomes in large scale.


Extrinsic purinergic regulation of neural stem/progenitor cells: implications for CNS development and repair.
There has been tremendous progress in understanding neural stem cell (NSC) biology, with genetic and cell biological methods identifying sequential gene expression and molecular interactions guiding NSC specification into distinct neuronal and glial populations during development. Data has emerged on the possible exploitation of NSC-based strategies to repair adult diseased brain. However, despite increased information on lineage specific transcription factors, cell-cycle regulators and epigenetic factors involved in the fate and plasticity of NSCs, understanding of extracellular cues driving the behavior of embryonic and adult NSCs is still very limited. Knowledge of factors regulating brain development is crucial in understanding the pathogenetic mechanisms of brain dysfunction. Since injury-activated repair mechanisms in adult brain often recapitulate ontogenetic events, the identification of these players will also reveal novel regenerative strategies. Here, we highlight the purinergic system as a key emerging player in the endogenous control of NSCs. Purinergic signalling molecules (ATP, UTP and adenosine) act with growth factors in regulating the synchronized proliferation, migration, differentiation and death of NSCs during brain and spinal cord development. At early stages of development, transient and time-specific release of ATP is critical for initiating eye formation; once anatomical CNS structures are defined, purinergic molecules participate in calcium-dependent neuron-glia communication controlling NSC behaviour. When development is complete, some purinergic mechanisms are silenced, but can be re-activated in adult brain after injury, suggesting a role in regeneration and self-repair. Targeting the purinergic system to develop new strategies for neurodevelopmental disorders and neurodegenerative diseases will be also discussed.


International Research Capacity-Building Programs for Nurses to Study the Drug Phenomenon in Latin America: challenges and perspectives.
The First International Research Capacity-Building Program for Nurses to Study the Drug Phenomenon in the Americas is a result of a partnership between the Inter-American Drug Abuse Control Commission (CICAD) of the Organization of American States (OAS) and the Faculty of Nursing in the University of Alberta, with financial support from the Government of Canada. The program was divided into two parts. The first part of the program was held at the University of Alberta in Edmonton, Alberta, Canada. It involved capacity-building in research methodologies at the Faculty of Nursing, which lead to the preparation of four multi-centric research proposals for drug demand reduction in the home countries of the eleven participants in the program. The second part of the program was related to the implementation of multi-centric research proposals in seven countries in Latin America and in Canada. This program presented expertise in research methodology to members of Latin American Schools of Nursing and introduced Latin American expertise to members of a Canadian Faculty of Nursing. The International Research Capacity-Building Program for Nurses to Study the Drug Phenomenon in the Americas has fostered the kind of inter-cultural respect and mutual appreciation necessary to confront the global health problem of the abuse of both licit and illicit drugs.


Nocturnal variations in lower-leg subcutaneous blood flow in paraplegic men.
1. Lower-leg subcutaneous adipose tissue blood flow rates were measured over 12-20 h under ambulatory conditions by means of the 133Xe-washout technique in nine paraplegic men, all with complete spinal cord lesions at or below the Th 6 level, and in nine age-matched healthy men. Portable CdTe(Cl) detectors and data-storage units were used. 2. The central and local sympathetic vasoconstrictive activity at the lower leg was measured under laboratory conditions by means of the 133Xe-washout technique and a stationary NaI(Tl) detector system. 3. The paraplegic men were found to have intact central and local sympathetic vasoconstrictive activity in their lower legs. Moreover, they all had a nocturnal hyperaemic blood flow phase of the same magnitude and duration as the control subjects. 4. The possibility that the somaesthetic nerves play a role in the hyperaemic response could be excluded, as all the paraplegic men suffered from complete lower-leg somaesthetic denervation. 5. A significant correlation was found between the time of going to bed and the nightly hyperaemic response in the right and left lower legs (P less than 0.01). 6. It is concluded that the present data are in accordance with the concept of a central nervous or humoral elicitation of nocturnal hyperaemia, although local metabolic and other factors might participate as well. Paraplegic men have an intact regulation of the postural and nocturnal changes in peripheral blood flow whether of central sympathetic or humoral origin.


Differential Growth Rates and In Vitro Drug Susceptibility to Currently Used Drugs for Multiple Isolates of Naegleria fowleri.
The free-living amoeba Naegleria fowleri, which typically dwells within warm, freshwater environments, can opportunistically cause primary amoebic meningoencephalitis (PAM), a disease with a mortality rate of >97%. The lack of positive treatment outcomes for PAM has prompted the discovery and development of more effective therapeutics, yet most studies utilize only one or two clinical isolates. The inability to assess possible heterogenic responses to drugs among isolates from various geographical regions hinders progress in the discovery of more effective drugs. Here, we conducted drug efficacy and growth rate determinations for 11 different clinical isolates by applying a previously developed CellTiter-Glo 2.0 screening technique and flow cytometry. We found significant differences in the susceptibilities of these isolates to 7 of 8 drugs tested, all of which make up the cocktail that is recommended to physicians by the U.S. Centers for Disease Control and Prevention. We also discovered significant variances in growth rates among isolates, which draws attention to the differences among the amoeba isolates collected from different patients. Our results demonstrate the need for additional clinical isolates of various genotypes in drug assays and highlight the necessity for more targeted therapeutics with universal efficacy across N. fowleri isolates. Our data establish a needed baseline for drug susceptibility among clinical isolates and provide a segue for future combination therapy studies as well as research related to phenotypic or genetic differences that could shed light on mechanisms of action or predispositions to specific drugs. IMPORTANCE Naegleria fowleri, also known as the brain-eating amoeba, is ubiquitous in warm freshwater and is an opportunistic pathogen that causes primary amoebic meningoencephalitis. Although few cases are described each year, the disease has a case fatality rate of >97%. In most laboratory studies of this organism, only one or two well-adapted lab strains are used; therefore, there is a lack of data to discern if there are major differences in potency of currently used drugs for multiple strains and genotypes of the amoeba. In this study, we found significant differences in the susceptibilities of 11 N. fowleri isolates to 7 of the 8 drugs currently used to treat the disease. The data from this study provide a baseline of drug susceptibility among clinical isolates and suggest that new drugs should be tested on a larger number of isolates in the future.


Qualitative factors in patients who die shortly after emergency department discharge.
Early death after emergency department (ED) discharge may signal opportunities to improve care. Prior studies are limited by incomplete mortality ascertainment and lack of clinically important information in administrative data. The goal in this hypothesis-generating study was to identify patient and process of care themes that may provide possible explanations for early postdischarge mortality. This was a qualitative analysis of medical records of adult patients who visited the ED of any of six hospitals in an integrated health system (Kaiser Permanente Southern California [KPSC]) and died within 7 days of discharge in 2007 and 2008. Nonmembers, visits to non-health plan hospitals, patients receiving or referred to hospice care, and patients with do not attempt resuscitation or do not intubate orders (DNAR/DNI) were excluded. Under the guidance of two qualitative research scientists, a team of three emergency physicians used grounded theory techniques to identify patient clinical presentations and processes of care that serve as potential explanations for poor outcome after discharge. The source population consisted of a total of 290,092 members with 446,120 discharges from six KPSC EDs in 2007 and 2008. A total of 203 deaths occurred within 7 days of ED discharge (0.05%). Sixty-one randomly chosen cases were reviewed. Patient-level themes that emerged included an unexplained persistent acute change in mental status, recent fall, abnormal vital signs, ill-appearing presentation, malfunctioning indwelling device, and presenting symptoms remaining at discharge. Process-of-care factors included a discrepancy in history of present illness, incomplete physical examination, and change of discharge plan by a third party, such as a consulting or admitting physician. In this hypothesis-generating study, qualitative research techniques were used to identify clinical and process-of-care factors in patients who died within days after discharge from an ED. These potential predictors will be formally tested in a future quantitative study.


Multiplexed phase-space imaging for 3D fluorescence microscopy.
Optical phase-space functions describe spatial and angular information simultaneously; examples of optical phase-space functions include light fields in ray optics and Wigner functions in wave optics. Measurement of phase-space enables digital refocusing, aberration removal and 3D reconstruction. High-resolution capture of 4D phase-space datasets is, however, challenging. Previous scanning approaches are slow, light inefficient and do not achieve diffraction-limited resolution. Here, we propose a multiplexed method that solves these problems. We use a spatial light modulator (SLM) in the pupil plane of a microscope in order to sequentially pattern multiplexed coded apertures while capturing images in real space. Then, we reconstruct the 3D fluorescence distribution of our sample by solving an inverse problem via regularized least squares with a proximal accelerated gradient descent solver. We experimentally reconstruct a 101 Megavoxel 3D volume (1010×510×500µm with NA 0.4), demonstrating improved acquisition time, light throughput and resolution compared to scanning aperture methods. Our flexible patterning scheme further allows sparsity in the sample to be exploited for reduced data capture.


RNA-seq Identification of RACGAP1 as a Metastatic Driver in Uterine Carcinosarcoma.
Uterine carcinosarcoma is a rare aggressive malignancy frequently presenting at advanced stage of disease with extrauterine metastases. Median survival is less than 2 years due to high relapse rates after surgery and poor response to chemotherapy or radiotherapy. The goal of this study was to identify novel therapeutic targets. We applied RNA-seq analysis to prospectively collected uterine carcinosarcoma tumor samples from patients undergoing primary surgical resection and for comparison, normal endometrial tissues from postmenopausal women undergoing hysterectomy for benign indications. Functional assays were done in primary carcinosarcoma cell lines developed from patients and in established cell lines, as well as a cell line-derived xenograft model. Validation was done by analysis of an independent cohort of patients with uterine carcinosarcoma from The Cancer Genome Atlas (TCGA). Rac GTPase-activating protein 1 (RACGAP1) was identified to be highly upregulated in uterine carcinosarcoma. Functional assays showed that RACGAP1 mediates motility and invasion via regulation of STAT3 phosphorylation and survivin expression. RACGAP1 depletion or survivin inhibition abrogated motility and invasiveness of carcinosarcoma cells, while RACGAP1 overexpression conferred invasiveness to endometrial adenocarcinoma cells. In the TCGA cohort, RACGAP1 expression correlated with survivin expression and extrauterine spread of disease. The RACGAP1-STAT3-survivin signaling pathway is required for the invasive phenotype of uterine carcinosarcoma and is a newly identified therapeutic target in this lethal disease. Clin Cancer Res; 22(18); 4676-86. ©2016 AACR.