DNAJC3 deficiency induces β-cell mitochondrial apoptosis and causes syndromic young-onset diabetes.
DNAJC3, also known as P58IPK, is an Hsp40 family member that interacts with and inhibits PKR-like ER-localized eIF2α kinase (PERK). Dnajc3 deficiency in mice causes pancreatic β-cell loss and diabetes. Loss-of-function mutations in DNAJC3 cause early-onset diabetes and multisystemic neurodegeneration. The aim of our study was to investigate the genetic cause of early-onset syndromic diabetes in two unrelated patients, and elucidate the mechanisms of β-cell failure in this syndrome. Whole exome sequencing was performed and identified variants were confirmed by Sanger sequencing. DNAJC3 was silenced by RNAi in INS-1E cells, primary rat β-cells, human islets, and induced pluripotent stem cell-derived β-cells. β-cell function and apoptosis were assessed, and potential mediators of apoptosis examined. The two patients presented with juvenile-onset diabetes, short stature, hypothyroidism, neurodegeneration, facial dysmorphism, hypoacusis, microcephaly and skeletal bone deformities. They were heterozygous compound and homozygous for novel loss-of-function mutations in DNAJC3. DNAJC3 silencing did not impair insulin content or secretion. Instead, the knockdown induced rat and human β-cell apoptosis and further sensitized cells to endoplasmic reticulum stress, triggering mitochondrial apoptosis via the pro-apoptototic Bcl-2 proteins BIM and PUMA. This report confirms previously described features and expands the clinical spectrum of syndromic DNAJC3 diabetes, one of the five monogenic forms of diabetes pertaining to the PERK pathway of the endoplasmic reticulum stress response. DNAJC3 deficiency may lead to β-cell loss through BIM- and PUMA-dependent activation of the mitochondrial pathway of apoptosis.


The impact of newly produced protein and dietary fiber rich fractions of yellow pea (Pisum sativum L.) on the structure and mechanical properties of pasta-like sheets.
Two fractions from pea (Pisum sativum L.), protein isolate (PPI) and dietary fiber (PF), were newly produced by extraction-fractionation method and characterized in terms of particle size distribution and structural morphology using SEM. The newly produced PPI and PF fractions were processed into pasta-like sheets with varying protein to fiber ratios (100/0, 90/10, 80/20, 70/30 and 50/50, respectively) using high temperature compression molding. We studied protein polymerization, molecular structure and protein-fiber interactions, as well as mechanical performance and cooking characteristics of processed PPI-PF blends. Bi-modal particle size distribution and chemical composition of the PPI and PF fractions influenced significantly the physicochemical properties of the pasta-like sheets. Polymerization was most pronounced for the 100 PPI, 90/10 and 80/20 PPI-PF samples as studied by SE-HPLC, and polymerization decreased with addition of the PF fraction. The mechanical properties, as strength and extensibility, were likewise the highest for the 100 PPI and 90/10 PPI-PF blends, while the E-modulus was similar for all the studied blends (around 38 MPa). The extensibility decreased with the increasing amount of PF in the blend. The highest amounts of β-sheets were found in the pasta-like sheets with high amounts of PPI (100, 90 and 80%), by FT-IR. An increase in PF fraction in the blend, resulted into the high amounts of unordered structures as observed by FT-IR, as well as in an increase in the molecular scattering distances observed by SAXS. The water uptake increased and cooking loss decreased with increased proportions of the PF fraction, and the consistency of 10 min cooked pasta-like sheets were alike al dente texture. The new knowledge obtained in this study on the use of extraction-fractionation method to produce novel PPI and PF fractions for developing innovative high nutritious food can be of a great importance. The obtained knowledge on the pea protein and fiber processing behaviour could greatly contribute to a better control of functional properties of various temperature-processed products from yellow pea.


Chemotherapy with mitomycin C and capecitabine in patients with advanced colorectal cancer pretreated with irinotecan and oxaliplatin.
To assess the activity and tolerability of the combination of mitomycin C and capecitabine in patients with metastatic colorectal cancer after failure of irinotecan and oxaliplatin-containing regimens. We retrospectively reviewed 28 patients with pretreated advanced colorectal cancer who had been treated with mitomycin C, 6 mg/m2 on day 1, and capecitabine, 1900 mg/m2 on days 1-14, every 3 weeks. Tumor assessment was performed every 3 cycles, toxicity assessed at each cycle. Main patient characteristics were median age, 61 years (range, 35-73); male/female ratio, 16/12; single metastatic site involvement, 5/28 (18%); > or =3 metastatic sites, 10/28 (36%). Ninety-six courses of therapy were given (median number, 3; range, 1-9). Twenty-six patients were assessable for response, and all were assessable for toxicity. There was 1 partial response (4%) and 12 had stable disease (43%). Median time to progression was 2 months (range, 1-9) and median overall survival was 6 months (range, 1-29+), with a 1-year overall survival rate of 25%. The regimen was very well tolerated without significant hematological toxicity. Our results are disappointing. Despite the good safety profile, they do not support further investigation or the routine use of this regimen in this setting.


Molecular architecture of bacterial type IV secretion systems.
In Gram-negative bacteria, type IV secretion (T4S) systems form ATP-powered complexes that span the entire cellular envelope and secrete a wide variety of substrates from single proteins to protein-protein and protein-DNA complexes. Recent structural data, namely the electron microscopy structure of the T4S core complex and the atomic-resolution structure of its outer-membrane pore, have profoundly altered our understanding of T4S architecture and mechanisms.


Bringing rights into resilience: revealing complexities of climate risks and social conflict.
Marginalisation and exclusion are expressed in social conflict and are determinative in distributing risk and resilience. This paper builds on recent literature that has adopted a human rights lens to explore how resilience practice can better account for issues of equity and power. Using the illustrative case of Timor-Leste, it presents an analysis of how human rights principles play out in the settings in which rights are given meaning. The approach reveals the reproduction of patterns of conflict and risk, and suggests two key priorities for resilience practice: first, recognising and responding to the deep-rooted narratives and procedures that normalise inequality and marginalisation at different scales; and second, allowing for transformation towards more equitable political and social arrangements as a part of resilience practice. Augmenting resilience with rights-based thinking can situate resilience practice, such that it responds to the complexity of social arrangements, reducing risk and social conflict.


PAHs in background soils from Western Europe: influence of atmospheric deposition and soil organic matter.
The levels and distribution of polynuclear aromatic hydrocarbons (PAHs) were determined in soil samples from background locations in the UK and Norway, to investigate their spatial distribution and the controlling environmental factors. Concentrations ranged between 42 and 11200 microg kg(-1) (geometric mean 640 microg kg(-1)) and 8.6 and 1050 microg kg(-1) (150 microg kg(-1)) dry weight in the UK and Norwegian soil, respectively. Proximity to sources and locations susceptible to high atmospheric depositional inputs resulted in higher concentrations. Statistically significant relationships were observed between PAH and total organic carbon (TOC) in the Norwegian samples. High molecular weight PAHs correlated with black carbon (BC) in UK-woodland soil. These observations support the hypothesis that TOC plays an important role in the retention of PAHs in soil and that PAHs are often combined with BC during combustion emissions. PAHs with 4 and more rings comprised approximately 90% of total PAHs in the UK soil, but only 50% in the Norwegian soil. The mixture of PAHs implied that fractionation occurred during long-range atmospheric transport and deposition. The lighter PAHs with lower K(ow) values more readily reached the most remote sites. The heavier PAHs with higher K(ow) values remained in closer proximity to sources.


Improving power in contrasting linkage-disequilibrium patterns between cases and controls.
Genetic association studies offer an opportunity to find genetic variants underlying complex human diseases. The success of this approach depends on the linkage disequilibrium (LD) between markers and the disease variant(s) in a local region of the genome. Because, in the region with a disease mutation, the LD pattern among markers may differ between cases and controls, in some scenarios, it is useful to compare a measure of this LD, to map disease mutations. For example, using the composite correlation to characterize the LD among markers, Zaykin et al. recently suggested an "LD contrast" test and showed that it has high power under certain haplotype-driven disease models. Furthermore, it is likely that individual variants observed at different positions in a gene act jointly with each other to influence the phenotype, and the LD contrast test is also a useful method to detect such joint action. However, the LD among markers introduced by mutations and their joint action is usually confounded by background LD, which is measured at the population level, especially in a local region with disease mutations. Because the measures of LD that are usually used, such as the composite correlation, represent both effects, they may not be optimal for the purpose of detecting association when high background LD exists. Here, we describe a test that improves the LD contrast test by taking into account the background LD. Because the proposed test is developed in a regression framework, it is very flexible and can be extended to continuous traits and to incorporate covariates. Our simulation results demonstrate the validity and substantially higher power of the proposed method over current methods. Finally, we illustrate our new method by applying it to real data from the International Collaborative Study on Hypertension in Blacks.


Effect of sperm extract injection volume, injection of PLCzeta cRNA, and tissue cell line on efficiency of equine nuclear transfer.
We evaluated the effect of different activation methods on blastocyst development after equine nuclear transfer. All activation treatments were followed by incubation in 2 mM 6-dimethylaminopurine for 4 h. In Experiment 1, reconstructed oocytes were injected with sperm extract for 0.1, 0.2, 0.4, 0.8, or 1.6 sec using a FemtoJet injection device, then treated with ionomycin. The blastocyst rate (9.8%) for 0.1-sec injection was significantly higher than that for 0.2 sec (0%) or 0.8 sec (1.4%). In Experiment 2, injection of murine PLCzeta cRNA before or after ionomycin treatment did not increase blastocyst development (0 and 4.5%) over a control treatment (injection of sperm extract after ionomycin exposure; 5.6%). Transfer of 10 blastocysts produced in Experiments 1 and 2 resulted in five pregnancies, all lost before 70 days of gestation. In Experiment 3, cells from a second biopsy sample from the same horse produced significantly more blastocysts than did the original sample (4/44 vs. 0/58; p < 0.05). Transfer of these four blastocysts produced two viable foals. In Experiment 4, blastocyst development rates did not differ between oocytes in metaphase I or II at the time of nuclear transfer (16.7 and 3.0%, respectively). A healthy foal was produced from a blastocyst originating from a metaphase I oocyte.


Surface Orientation and Binding Strength Modulate Shape of FtsZ on Lipid Surfaces.
We have used a simple model system to test the prediction that surface attachment strength of filaments presenting a torsion would affect their shape and properties. FtsZ from E. coli containing one cysteine in position 2 was covalently attached to a lipid bilayer containing maleimide lipids either in their head group (to simulate tight attachment) or at the end of a polyethylene glycol molecule attached to the head group (to simulate loose binding). We found that filaments tightly attached grew straight, growing from both ends, until they formed a two-dimensional lattice. Further monomer additions to their sides generated a dense layer of oriented filaments that fully covered the lipid membrane. After this point the surface became unstable and the bilayer detached from the surface. Filaments with a loose binding were initially curved and later evolved into straight thicker bundles that destabilized the membrane after reaching a certain surface density. Previously described theoretical models of FtsZ filament assembly on surfaces that include lateral interactions, spontaneous curvature, torsion, anchoring to the membrane, relative geometry of the surface and the filament 'living-polymer' condition in the presence of guanosine triphosphate (GTP) can offer some clues about the driving forces inducing these filament rearrangements.


STIM1 R304W in mice causes subgingival hair growth and an increased fraction of trabecular bone.
Calcium signaling plays a central role in bone development and homeostasis. Store operated calcium entry (SOCE) is an important calcium influx pathway mediated by calcium release activated calcium (CRAC) channels in the plasma membrane. Stromal interaction molecule 1 (STIM1) is an endoplasmic reticulum calcium sensing protein important for SOCE. We generated a mouse model expressing the STIM1 R304W mutation, causing Stormorken syndrome in humans. Stim1R304W/R304W mice showed perinatal lethality, and the only three animals that survived into adulthood presented with reduced growth, low body weight, and thoracic kyphosis. Radiographs revealed a reduced number of ribs in the Stim1R304W/R304W mice. Microcomputed tomography data revealed decreased cortical bone thickness and increased trabecular bone volume fraction in Stim1R304W/R304W mice, which had thinner and more compact bone compared to wild type mice. The Stim1R304W/+ mice showed an intermediate phenotype. Histological analyses showed that the Stim1R304W/R304W mice had abnormal bone architecture, with markedly increased number of trabeculae and reduced bone marrow cavity. Homozygous mice showed STIM1 positive osteocytes and osteoblasts. These findings highlight the critical role of the gain-of-function (GoF) STIM1 R304W protein in skeletal development and homeostasis in mice. Furthermore, the novel feature of bilateral subgingival hair growth on the lower incisors in the Stim1R304W/R304W mice and 25 % of the heterozygous mice indicate that the GoF STIM1 R304W protein also induces an abnormal epithelial cell fate.


Habitat provision is a major driver of native bird communities in restored urban forests.
Urbanization, and the drastic loss of habitat it entails, poses a major threat to global avian biodiversity. Ecological restoration of urban forests is therefore increasingly vital for native bird conservation, but control of invasive predators may also be needed to sustain native bird populations in cities where species invasions have been particularly severe. We evaluated restoration success by investigating changes in native bird communities along a restoration chronosequence of 25 restored urban forests representing 72 years of forest development, which we compared to two target reference systems and a control system. We hypothesized that total species richness and relative abundance of native forest birds would increase with the age of restoration planting. We further hypothesized that relative abundance of rats, possums and cats would negatively impact native birds, while amount of native forest in the surrounding landscape would have a positive effect. We used structural equation modelling (SEM) to investigate the relative influence of forest structure (complexity index, tree height, canopy openness, basal area, species richness and density), landscape attributes (patch area, perimeter length, landscape composition within three buffer zones, distance to the nearest road and water source) and invasive mammalian predator indices of relative abundance on total species richness and relative abundance of native forest birds. Species richness increased with age of restoration planting, with community composition progressing towards that found in target reference systems. SEM revealed that years restored was a direct driver of bird species richness but an indirect driver of abundance, which was directly driven by canopy openness. Contrary to our predictions, invasive mammals had no significant effect on native bird species richness or abundance. Our results demonstrate that provision and improvement of habitat quantity and quality through restoration is the vital first step to re-establishing native forest bird communities in cities.


NeuroChip, an updated version of the NeuroX genotyping platform to rapidly screen for variants associated with neurological diseases.
Genetics has proven to be a powerful approach in neurodegenerative diseases research, resulting in the identification of numerous causal and risk variants. Previously, we introduced the NeuroX Illumina genotyping array, a fast and efficient genotyping platform designed for the investigation of genetic variation in neurodegenerative diseases. Here, we present its updated version, named NeuroChip. The NeuroChip is a low-cost, custom-designed array containing a tagging variant backbone of about 306,670 variants complemented with a manually curated custom content comprised of 179,467 variants implicated in diverse neurological diseases, including Alzheimer's disease, Parkinson's disease, Lewy body dementia, amyotrophic lateral sclerosis, frontotemporal dementia, progressive supranuclear palsy, corticobasal degeneration, and multiple system atrophy. The tagging backbone was chosen because of the low cost and good genome-wide resolution; the custom content can be combined with other backbones, like population or drug development arrays. Using the NeuroChip, we can accurately identify rare variants and impute over 5.3 million common SNPs from the latest release of the Haplotype Reference Consortium. In summary, we describe the design and usage of the NeuroChip array and show its capability for detecting rare pathogenic variants in numerous neurodegenerative diseases. The NeuroChip has a more comprehensive and improved content, which makes it a reliable, high-throughput, cost-effective screening tool for genetic research and molecular diagnostics in neurodegenerative diseases.


The effectiveness of early lens extraction with intraocular lens implantation for the treatment of primary angle-closure glaucoma (EAGLE): study protocol for a randomized controlled trial.
Glaucoma is the leading cause of irreversible blindness. Although primary open-angle glaucoma is more common, primary angle-closure glaucoma (PACG) is more likely to result in irreversible blindness. By 2020, 5·3 million people worldwide will be blind because of PACG. The current standard care for PACG is a stepped approach of a combination of laser iridotomy surgery (to open the drainage angle) and medical treatment (to reduce intraocular pressure). If these treatments fail, glaucoma surgery (eg, trabeculectomy) is indicated. It has been proposed that, because the lens of the eye plays a major role in the mechanisms leading to PACG, early clear lens extraction will improve glaucoma control by opening the drainage angle. This procedure might reduce the need for drugs and glaucoma surgery, maintain good visual acuity, and improve quality of life compared with standard care.EAGLE aims to evaluate whether early lens extraction improves patient-reported, clinical outcomes, and cost-effectiveness, compared with standard care. EAGLE is a multicentre pragmatic randomized trial. All people presenting to the recruitment centres in the UK and east Asia with newly diagnosed PACG and who are at least 50 years old are eligible.The primary outcomes are EQ-5D, intraocular pressure, and incremental cost per quality adjusted life year (QALY) gained. Other outcomes are: vision and glaucoma-specific patient-reported outcomes, visual acuity, visual field, angle closure, number of medications, additional surgery (e.g., trabeculectomy), costs to the health services and patients, and adverse events.A single main analysis will be done at the end of the trial, after three years of follow-up. The analysis will be based on all participants as randomized (intention to treat). 400 participants (200 in each group) will be recruited, to have 90% power at 5% significance level to detect a difference in EQ-5D score between the two groups of 0·05, and a mean difference in intraocular pressure of 1·75 mm Hg. The study will have 80% power to detect a difference of 15% in the glaucoma surgery rate. ISRCTN44464607.


Hepatocellular carcinoma in Egyptians with and without a history of hepatitis B virus infection: association with hepatitis C virus (HCV) infection but not with (HCV) RNA level.
The aim of this study was to analyze the association of hepatocellular carcinoma (HCC) with hepatitis C virus (HCV) in Egypt, using hepatitis B virus (HBV) and hepatitis E virus (HEV) as virus controls. In addition, the association of HCC with HCV RNA levels among persons seropositive for HCV was analyzed. We compared 131 patients with proven HCC, 247 with bladder cancer, and 466 healthy hospital employees. Age, sex, and place of residence were recorded to study confounding factors. Among the healthy controls, 16% were seropositive for HCV, 21% for HBV, and 31% for HEV. When healthy controls were age-matched with HCC patients, the latter were significantly (P < 0.001) more often HCV seropositive (67%) than were the controls (30%). The seropositivity for HBV and HEV did not differ significantly in frequency between the two groups. The seropositivity for HCV was also significantly (P < 0.001) more often found in HCC patients (76%) than in BC patients (47%), with seroprevalences for HBV and HEV not differing significantly in these age-matched groups. In HBV-negative HCC and bladder cancer patients, seroprevalence for HCV was significantly (P = 0.002) higher in HCC patients (68%) than in bladder cancer patients (36%). This difference was even more pronounced (P < 0.001) in HBV-positive HCC and bladder cancer patients (78% versus 52%, respectively). Of HCV-seropositive individuals, 49% were HCV RNA positive by branched DNA assay, and of these, 96% were infected by HCV genotype 4. No correlation between HCV RNA load and seropositivity of HBV or age or disease state was found. Infection with HCV and HCV-HBV double infection, but not HBV or HEV infection alone, is strongly correlated with HCC in Egypt.


Comparing Different Sequential Mediational Interpretations of Beck's Cognitive Model of Depression in Adolescents.
Depression is a developmental phenomenon with significantly increasing rates during adolescence. As Beck's cognitive model of depression has been commonly accepted to explain the development and maintenance of depression, it is crucial to understand how and when cognitive vulnerabilities predicted in this model begin to interact. Three sequential interpretations of this model were compared. The causal mediational interpretation identifies dysfunctional attitudes as most distal to depressive symptoms, followed by cognitive errors, cognitive triad, and negative automatic thoughts, with each construct successively more proximal to depressive symptoms. In the symptom model the causal chain is reversed, with depressive symptoms as the most distal construct, followed by negative automatic thoughts, the cognitive triad, cognitive errors, and then dysfunctional attitudes. The bidirectional model merges both interpretations in which the activation of cognitive constructs causes the development of depressive symptoms which in turn trigger and reinforce already existing dysfunctional attitudes. Further, while Beck's model of depression proposes full mediation, empirical studies identified repeatedly partial mediations. Thus, the causal meditational, the symptoms, and the bidirectional model were each tested as full and partial mediation models. Finally, sex differences in the associations between variables were studied. In the 3-wave longitudinal study, 518 high school students (62.7 % female, average age: 15.09 years) completed questionnaires measuring all mentioned elements of Beck's model. The bidirectional model with partial mediation fits the data best. Cognitive errors emerged as the main mediator in the bidirectional model with partial mediation and significant sex differences in the strengths of associations were identified. The findings demonstrate the relevance of adolescence as developmental period during which the examined associations develop into the network they form in adulthood. Further, psychological interventions focusing on cognitive errors promise to be most effective.


Human Gene Mutation Database (HGMD): 2003 update.
The Human Gene Mutation Database (HGMD) constitutes a comprehensive core collection of data on germ-line mutations in nuclear genes underlying or associated with human inherited disease (www.hgmd.org). Data catalogued includes: single base-pair substitutions in coding, regulatory and splicing-relevant regions; micro-deletions and micro-insertions; indels; triplet repeat expansions as well as gross deletions; insertions; duplications; and complex rearrangements. Each mutation is entered into HGMD only once in order to avoid confusion between recurrent and identical-by-descent lesions. By March 2003, the database contained in excess of 39,415 different lesions detected in 1,516 different nuclear genes, with new entries currently accumulating at a rate exceeding 5,000 per annum. Since its inception, HGMD has been expanded to include cDNA reference sequences for more than 87% of listed genes, splice junction sequences, disease-associated and functional polymorphisms, as well as links to data present in publicly available online locus-specific mutation databases. Although HGMD has recently entered into a licensing agreement with Celera Genomics (Rockville, MD), mutation data will continue to be made freely available via the Internet.


Lasers and Nonsurgical Periodontal Therapy.
The purpose of Linking Research to Clinical Practice is to present evidence based information to clinical dental hygienists so that they can make informed decisions regarding patient treatment and recommendations. Each issue will feature a different topic area of importance to clinical dental hygienists with A BOTTOM LINE to translate the research findings into clinical application.


International service trade and its implications for human resources for health: a case study of Thailand.
This study aims at analysing the impact of international service trade on the health care system, particularly in terms of human resources for health (HRH), using Thailand as a case study. Information was gathered through a literature review and interviews of relevant experts, as well as a brainstorming session.It was found that international service trade has greatly affected the Thai health care system and its HRH. From 1965 to 1975 there was massive emigration of physicians from Thailand in response to increasing demand in the United States of America. The country lost about 1,500 physicians, 20% of its total number, during that period.External migration of health professionals occurred without relation to agreements on trade in services. It was also found that free trade in service sectors other than health could seriously affect the health care system and HRH. Free trade in financial services with free flow of low-interest foreign loans, which started in 1993 in Thailand, resulted in the mushrooming of urban private hospitals between 1994 and 1997. This was followed by intensive internal migration of health professionals from rural public to urban private hospitals.After the economic crisis in 1997, with the resulting downturn of the private health sector, reverse brain drain was evident. At the same time, foreign investors started to invest in the bankrupt private hospitals. Since 2001, the return of economic growth and the influx of foreign patients have started another round of internal brain drain.


Comparison of mammography and Pap test use from the 1987 and 1992 National Health Interview Surveys: are we closing the gaps?
Using data from the National Health Interview Survey Cancer Control Supplements, we examined the trends in mammogram and Pap smear test screening between 1987 and 1992 and the demographic characteristics associated with the use of those screening tests in 1992. In 1992, 67% of women 40 years of age and older reported ever having had a mammogram compared with 36% in 1987. A mammogram within the past year was reported by 29% of women in 1992, an increase from 14% in 1987. Among women 18 and older, 91% had ever had a Pap smear test in 1992, and 43% had one within the past year, an increase from 89% and 38%, respectively, in 1987. These changes represented significant increases in the use of both mammograms and Pap smear tests between 1987 and 1992 (P < .05). Race was not significantly associated with underutilization of mammograms, but income showed a significant relationship, with a declining likelihood of mammogram use as income decreased (odds ratio [OR] = 0.4, 95% confidence interval [CI] = 0.3, 0.6 for the income less than poverty level). Pap smear tests were less likely to be reported by older women ( > or = 65), widows, and never-married women, African-American women were more likely than Caucasian women to have had a Pap smear test, and women of other races were the least likely of any race/ethnic group to have ever had one (OR = 0.2, 95% CI = 0.1, 0.4). Women with less than 12 years of education had about a 40% decreased risk of having had a recent mammogram or Pap smear test. Women without a usual source of medical care also underutilized both screening procedures. Significant gains in the use of mammograms were found for all groups between 1987 and 1992. However, for the Pap smear test, some groups reported no differences during the five-year period. Although race and ethnic differences in the rates of screening use have been reduced somewhat, socioeconomically disadvantaged women and those with poor access to health care are still important target populations for increasing screening, particularly mammograms.


The effect of statin therapy on the formation of arteriovenous fistula stenoses and the rate of reoccurrence of previously treated stenoses.
Statins reduce inflammation in end-stage renal disease patients and improve endothelial function beyond cholesterol lowering. Despite this, statins do not improve the maturation rate, primary patency rate, and the cumulative survival of arteriovenous fistulas (AVFs). It is unknown if statins decrease the number of stenoses developing in AVFs or prolong the intervals between angioplasties needed to treat recurring stenoses. We conducted a retrospective chart review of our 265 active dialysis patients. The statin group was significantly more likely to be diabetic (64% vs. 43.6%) and treated with aspirin (64% vs. 40%) when compared to those not treated with statins (P=0.04 and 0.01). The mean time to first intervention (primary patency) was 16.5 months in statin users and 15.8 months in the nonstatin group (P=0.49) with standard deviations of ± 18.5 and 16.6 months, respectively. Statin use was not associated with a significant decrease in the number of stenoses diagnosed (P=0.28). The mean time between recurrent stenoses' angioplasties was 8.9 months in statin users and 7.3 months in the nonstatin patients (P=0.25). Aspirin users were more likely to have a decreased primary patency (rate ratio=1.65, P=0.03) compared with nonaspirin users. Patients who were prescribed aspirin developed 1.6 (P 0.01) times more stenoses than those not treated with aspirin. We report for the first time that statin therapy does not decrease the number of stenotic lesions developing in the AVF or prolong the interval between procedures required to treat recurrent stenoses.


Severe sepsis exacerbates cell-mediated immunity in the lung due to an altered dendritic cell cytokine profile.
Severe sepsis leads to long-term alterations in the immune response of surviving individuals. We have modeled this alteration in host immunity by studying the survivors of severe experimental sepsis (murine cecal ligation and puncture), which were subsequently challenged with lung granuloma-inducing Schistosoma mansoni eggs. This granulomatous response is a well-studied cell-mediated immune reaction characterized by elevated levels of type-2 cytokines. Pulmonary granulomas induced by S. mansoni eggs in cecal ligation and puncture survivors were significantly larger and contained more eosinophils than granulomas in sham-operated mice. Significantly lower interleukin (IL)-12p40 mRNA and IL-12p70 protein levels were observed in the lungs of postseptic mice with developing granulomas, compared with controls. Postseptic mice had significantly fewer dendritic cells in the lungs during the granulomatous response. Isolated lung dendritic cells from postseptic mice at days 8 and 16 after S. mansoni egg challenge exhibited defective IL-12 synthesis but enhanced IL-10 synthesis after Toll-like receptor agonist challenge. Pulmonary transfection with an IL-12-expressing adenovirus in postseptic mice reversed the skewing of the pulmonary cytokine profile and normalized the lung granulomatous response. Our data indicate that severe sepsis shifts the pulmonary cytokine environment, presumably via effects on pulmonary dendritic cells, which in turn alters the lung cell-mediated immune response.


Risk factors for inflammatory breast cancer and other invasive breast cancers.
We investigated risk factors for inflammatory breast cancer (IBC), a rare, aggressive, and poorly understood breast cancer that is characterized by diffuse breast skin erythema and edema. We included 617 IBC case subjects in a nested case-control study from the Breast Cancer Surveillance Consortium database (1994-2009). We also included 1151 noninflammatory, locally advanced, invasive breast cancers with chest wall/breast skin involvement (LABC), 7600 noninflammatory invasive case subjects without chest wall/breast skin involvement (BC), and 93 654 control subjects matched to case subjects on age and year at diagnosis and mammography registry. We present estimates of rate ratios (RRs) and 95% confidence intervals (CI) from conditional logistic regression analyses for each case group vs control subjects based on multiply imputed datasets. First-degree family history of breast cancer and high mammographic breast density increased risk of IBC, LABC, and BC. High body mass index (BMI) increased IBC risk irrespective of menopausal status and estrogen receptor (ER) expression; rate ratios for BMI 30 and greater vs BMI less than 25 were 3.90 (95% CI = 1.50 to 10.14) in premenopausal women and 3.70 (95% CI = 1.98 to 6.94) in peri/postmenopausal women not currently using hormones. BMI 30 and greater slightly increased risk of ER-positive BC (RR = 1.40; 95% CI = 1.11 to 1.76). Statistically significant reductions in risk of ER-negative IBC with older age at first birth and of ER-positive IBC with higher education were not seen for LABC and BC of the same ER status. Different associations with BMI, age at first birth, and education between IBC and/or LABC and BC suggest a distinct etiology for IBC.


The GRADE Working Group clarifies the construct of certainty of evidence.
To clarify the grading of recommendations assessment, development and evaluation (GRADE) definition of certainty of evidence and suggest possible approaches to rating certainty of the evidence for systematic reviews, health technology assessments, and guidelines. This work was carried out by a project group within the GRADE Working Group, through brainstorming and iterative refinement of ideas, using input from workshops, presentations, and discussions at GRADE Working Group meetings to produce this document, which constitutes official GRADE guidance. Certainty of evidence is best considered as the certainty that a true effect lies on one side of a specified threshold or within a chosen range. We define possible approaches for choosing threshold or range. For guidelines, what we call a fully contextualized approach requires simultaneously considering all critical outcomes and their relative value. Less-contextualized approaches, more appropriate for systematic reviews and health technology assessments, include using specified ranges of magnitude of effect, for example, ranges of what we might consider no effect, trivial, small, moderate, or large effects. It is desirable for systematic review authors, guideline panelists, and health technology assessors to specify the threshold or ranges they are using when rating the certainty in evidence.


Gene essentiality and synthetic lethality in haploid human cells.
Although the genes essential for life have been identified in less complex model organisms, their elucidation in human cells has been hindered by technical barriers. We used extensive mutagenesis in haploid human cells to identify approximately 2000 genes required for optimal fitness under culture conditions. To study the principles of genetic interactions in human cells, we created a synthetic lethality network focused on the secretory pathway based exclusively on mutations. This revealed a genetic cross-talk governing Golgi homeostasis, an additional subunit of the human oligosaccharyltransferase complex, and a phosphatidylinositol 4-kinase β adaptor hijacked by viruses. The synthetic lethality map parallels observations made in yeast and projects a route forward to reveal genetic networks in diverse aspects of human cell biology.


Alcohols as alkylating agents in heteroarene C-H functionalization.
Redox processes and radical intermediates are found in many biochemical processes, including deoxyribonucleotide synthesis and oxidative DNA damage. One of the core principles underlying DNA biosynthesis is the radical-mediated elimination of H2O to deoxygenate ribonucleotides, an example of 'spin-centre shift', during which an alcohol C-O bond is cleaved, resulting in a carbon-centred radical intermediate. Although spin-centre shift is a well-understood biochemical process, it is underused by the synthetic organic chemistry community. We wondered whether it would be possible to take advantage of this naturally occurring process to accomplish mild, non-traditional alkylation reactions using alcohols as radical precursors. Because conventional radical-based alkylation methods require the use of stoichiometric oxidants, increased temperatures or peroxides, a mild protocol using simple and abundant alkylating agents would have considerable use in the synthesis of diversely functionalized pharmacophores. Here we describe the development of a dual catalytic alkylation of heteroarenes, using alcohols as mild alkylating reagents. This method represents the first, to our knowledge, broadly applicable use of unactivated alcohols as latent alkylating reagents, achieved via the successful merger of photoredox and hydrogen atom transfer catalysis. The value of this multi-catalytic protocol has been demonstrated through the late-stage functionalization of the medicinal agents, fasudil and milrinone.


The primary mechanism of attenuation of bacillus Calmette-Guerin is a loss of secreted lytic function required for invasion of lung interstitial tissue.
Tuberculosis remains a leading cause of death worldwide, despite the availability of effective chemotherapy and a vaccine. Bacillus Calmette-Guérin (BCG), the tuberculosis vaccine, is an attenuated mutant of Mycobacterium bovis that was isolated after serial subcultures, yet the functional basis for this attenuation has never been elucidated. A single region (RD1), which is absent in all BCG substrains, was deleted from virulent M. bovis and Mycobacterium tuberculosis strains, and the resulting DeltaRD1 mutants were significantly attenuated for virulence in both immunocompromised and immunocompetent mice. The M. tuberculosis DeltaRD1 mutants were also shown to protect mice against aerosol challenge, in a similar manner to BCG. Interestingly, the DeltaRD1 mutants failed to cause cytolysis of pneumocytes, a phenotype that had been previously used to distinguish virulent M. tuberculosis from BCG. A specific transposon mutation, which disrupts the Rv3874 Rv3875 (cfp-10 esat-6) operon of RD1, also caused loss of the cytolytic phenotype in both pneumocytes and macrophages. This mutation resulted in the attenuation of virulence in mice, as the result of reduced tissue invasiveness. Moreover, specific deletion of each transcriptional unit of RD1 revealed that three independent transcriptional units are required for virulence, two of which are involved in the secretion of ESAT-6 (6-kDa early secretory antigenic target). We conclude that the primary attenuating mechanism of bacillus Calmette-Guérin is the loss of cytolytic activity mediated by secreted ESAT-6, which results in reduced tissue invasiveness.


Fluoride Exposure Induces Inhibition of Sodium/Iodide Symporter (NIS) Contributing to Impaired Iodine Absorption and Iodine Deficiency: Molecular Mechanisms of Inhibition and Implications for Public Health.
The sodium iodide symporter (NIS) is the plasma membrane glycoprotein that mediates active iodide transport in the thyroid and other tissues, such as the salivary, gastric mucosa, rectal mucosa, bronchial mucosa, placenta and mammary glands. In the thyroid, NIS mediates the uptake and accumulation of iodine and its activity is crucial for the development of the central nervous system and disease prevention. Since the discovery of NIS in 1996, research has further shown that NIS functionality and iodine transport is dependent on the activity of the sodium potassium activated adenosine 5'-triphosphatase pump (Na+, K+-ATPase). In this article, I review the molecular mechanisms by which F inhibits NIS expression and functionality which in turn contributes to impaired iodide absorption, diminished iodide-concentrating ability and iodine deficiency disorders. I discuss how NIS expression and activity is inhibited by thyroglobulin (Tg), tumour necrosis factor alpha (TNF-α), transforming growth factor beta 1 (TGF-β1), interleukin 6 (IL-6) and Interleukin 1 beta (IL-1β), interferon-γ (IFN-γ), insulin like growth factor 1 (IGF-1) and phosphoinositide 3-kinase (PI3K) and how fluoride upregulates expression and activity of these biomarkers. I further describe the crucial role of prolactin and megalin in regulation of NIS expression and iodine homeostasis and the effect of fluoride in down regulating prolactin and megalin expression. Among many other issues, I discuss the potential conflict between public health policies such as water fluoridation and its contribution to iodine deficiency, neurodevelopmental and pathological disorders. Further studies are warranted to examine these associations.


Chromatin as a key consumer in the metabolite economy.
In eukaryotes, chromatin remodeling and post-translational modifications (PTMs) shape the local chromatin landscape to establish permissive and repressive regions within the genome, orchestrating transcription, replication, and DNA repair in concert with other epigenetic mechanisms. Though cellular nutrient signaling encompasses a huge number of pathways, recent attention has turned to the hypothesis that the metabolic state of the cell is communicated to the genome through the type and concentration of metabolites in the nucleus that are cofactors for chromatin-modifying enzymes. Importantly, both epigenetic and metabolic dysregulation are hallmarks of a range of diseases, and this metabolism-chromatin axis may yield a well of new therapeutic targets. In this Perspective, we highlight emerging themes in the inter-regulation of the genome and metabolism via chromatin, including nonenzymatic histone modifications arising from chemically reactive metabolites, the expansion of PTM diversity from cofactor-promiscuous chromatin-modifying enzymes, and evidence for the existence and importance of subnucleocytoplasmic metabolite pools.


Diagnostic accuracy of urine dipstick testing in screening for microalbuminuria in type 2 diabetes: a cohort study in primary care.
Clinical guidelines recommend annual screening for microalbuminuria in diabetes. Detection of microalbuminuria is important because it is associated with increased morbidity and mortality. Dipstick tests for microalbuminuria may be convenient, but their accuracy is uncertain. To assess the utility of urine dipstick testing for microalbuminuria in type 2 diabetes. In a 6-week cohort study in four general practices in Oxfordshire, UK, first-pass urine samples were obtained at two weekly intervals from patients with type 2 diabetes and tested in the practice using Micral-Test and Microalbustix urine dipsticks. Parallel samples were sent for laboratory albumin-creatinine ratio (ACR) assay. Results of single dipstick tests and sequences of dipstick and laboratory tests were compared with a clinical testing strategy based on current guidelines to assess the accuracy and estimate costs of testing. The prevalence of microalbuminuria was 12.5% (n = 88). Mean (standard deviation) age was 68 (10) years, 56 (57%) were men. Median (interquartile range) diabetes duration was 6.2 (2.0-10.0) years. The sensitivity and specificity, respectively, of a single Micral-Test were 91.7% and 44.0% and of a Microalbustix test 33.3% and 92.0%. Testing strategies involving dipstick and laboratory ACR measurements or dipstick tests had similar accuracy. The costs of using dipstick tests were overall lower than laboratory ACR-based testing. Dipstick testing in this study did not reliably identify diabetes patients with microalbuminuria. Although dipstick testing would decrease testing costs, it could either fail to diagnose most patients with microalbuminuria or increase the numbers of patients retested depending on the dipstick used.


Cisplatin in combination with Phenethyl Isothiocyanate (PEITC), a potential new therapeutic strategy for malignant pleural mesothelioma.
Malignant pleural mesothelioma (MPM) is a very aggressive form of cancer with a poor diagnosis and prognosis. The first line treatment for MPM is a combination of cisplatin and Pemetrexed, which displayed limited efficacy and severe side effects. The naturally occurring compound phenethyl isothiocyanate (PEITC) previously showed interesting anti-tumor properties on several cancer cell lines. We thus aim at evaluating PEITC used alone or in combination with cisplatin in order to improve MPM treatment. Nine MPM cell lines and primary mesothelial cells (PMC), co-cultured or not with M2 macrophages present in MPM microenvironment, were used to assess PEITC and cisplatin anti-tumor properties. Compounds were used alone or in combination. Both PEITC and cisplatin were cytotoxic on MPM cells in a dose dependent manner. We herein showed that PEITC-induced cytotoxicity was due to the generation of reactive oxygen species. Moreover, we showed that cisplatin-PEITC combination allowed the potentialization of both compounds' cytotoxic effects and prevented the emergence of resistant MPM cells. Interestingly, PMC were not sensitive to the combination. Finally, we showed that M2 macrophages did not alter the anti-tumor properties of the combination. Cisplatin-PEITC combination thus represents a promising strategy to induce a selective toxicity towards malignant cells.


Increased H3K9 methylation and impaired expression of Protocadherins are associated with the cognitive dysfunctions of the Kleefstra syndrome.
Kleefstra syndrome, a disease with intellectual disability, autism spectrum disorders and other developmental defects is caused in humans by haploinsufficiency of EHMT1. Although EHMT1 and its paralog EHMT2 were shown to be histone methyltransferases responsible for deposition of the di-methylated H3K9 (H3K9me2), the exact nature of epigenetic dysfunctions in Kleefstra syndrome remains unknown. Here, we found that the epigenome of Ehmt1+/- adult mouse brain displays a marked increase of H3K9me2/3 which correlates with impaired expression of protocadherins, master regulators of neuronal diversity. Increased H3K9me3 was present already at birth, indicating that aberrant methylation patterns are established during embryogenesis. Interestingly, we found that Ehmt2+/- mice do not present neither the marked increase of H3K9me2/3 nor the cognitive deficits found in Ehmt1+/- mice, indicating an evolutionary diversification of functions. Our finding of increased H3K9me3 in Ehmt1+/- mice is the first one supporting the notion that EHMT1 can quench the deposition of tri-methylation by other Histone methyltransferases, ultimately leading to impaired neurocognitive functioning. Our insights into the epigenetic pathophysiology of Kleefstra syndrome may offer guidance for future developments of therapeutic strategies for this disease.


Critical role of both retinoid nuclear receptors and retinoid-X-receptors in mediating growth inhibition of ovarian cancer cells by all-trans retinoic acid.
Retinoids have been shown to inhibit the growth of a number of human tumor cells, including several ovarian adenocarcinoma cell lines. All-trans retinoic acid (RA) is an effective growth suppressor of CA-OV3 cells but not SK-OV3 cells. Since the effects of RA are known to be mediated via the nuclear receptors (RARs and RXRs), we initially compared levels of the various RARs and RXRs in the CA-OV3 and SK-OV3 cell lines. The RA resistant SK-OV3 cells expressed reduced levels of RAR-alpha and RXR-alpha. Furthermore, induction of RAR-alpha by RA was impaired in the RA resistant SK-OV3 cells as was RARE binding and RARE-dependent transcriptional activity. These results suggested that changes in the amounts and/or activity of RARs and/or RXR-alpha could determine the growth response of ovarian tumor cells to RA. This was confirmed by modulating the levels of RARs and RXR-alpha in the SK-OV3 cells using the LacSwitch inducible expression system. Stably transfected clones of RA resistant SK-OV3 cells exhibited a significant inhibition of growth by RA treatment when RAR-alpha was induced. Overexpression of both RAR-alpha and RXR-alpha resulted in a level of growth inhibition nearly equal to that exhibited by the RA sensitive CA-OV3 cell line. Similar results were obtained when a combination of RXR-alpha and either RAR-beta or RAR-gamma was overexpressed in SK-OV3 cells. Our results show that the nuclear receptors and RXR-alpha play a critical role in mediating growth suppression by RA in ovarian cancer cells.


Changes in genetic diversity and differentiation in Red-cockaded woodpeckers (Dryobates borealis) over the past century.
Red-cockaded woodpeckers (RCW; Dryobates borealis) declined after human activities reduced their fire-maintained pine ecosystem to <3% of its historical range in the southeastern United States and degraded remaining habitat. An estimated 1.6 million RCW cooperative breeding groups declined to about 3,500 groups with no more than 10,000 birds by 1978. Management has increased RCW population abundances since they were at their lowest in the 1990s. However, no range-wide study has been undertaken since then to investigate the impacts of this massive bottleneck or infer the effects of conservation management and recent demographic recoveries. We used mitochondrial DNA sequences (mtDNA) and nine nuclear microsatellite loci to determine if range-wide demographic declines resulted in changes to genetic structure and diversity in RCW by comparing samples collected before 1970 (mtDNA data only), between 1992 and 1995 (mtDNA and microsatellites), and between 2010 and 2014 (mtDNA and microsatellites). We show that genetic diversity has been lost as detected by a reduction in the number of mitochondrial haplotypes. This reduction was apparent in comparisons of pre-1970 mtDNA data with data from the 1992-1995 and 2010-2014 time points, with no change between the latter two time points in mtDNA and microsatellite analyses. The mtDNA data also revealed increases in range-wide genetic differentiation, with a genetically panmictic population present throughout the southeastern United States in the pre-1970s data and subsequent development of genetic structure that has remained unchanged since the 1990s. Genetic structure was also uncovered with the microsatellite data, which like the mtDNA data showed little change between the 1992-1995 and 2010-2014 data sets. Temporal haplotype networks revealed a consistent, star-like phylogeny, suggesting that despite the overall loss of haplotypes, no phylogenetically distinct mtDNA lineages were lost when the population declined. Our results may suggest that management during the last two decades has prevented additional losses of genetic diversity.


Feedback from the heart: Emotional learning and memory is controlled by cardiac cycle, interoceptive accuracy and personality.
Feedback processing is critical to trial-and-error learning. Here, we examined whether interoceptive signals concerning the state of cardiovascular arousal influence the processing of reinforcing feedback during the learning of 'emotional' face-name pairs, with subsequent effects on retrieval. Participants (N=29) engaged in a learning task of face-name pairs (fearful, neutral, happy faces). Correct and incorrect learning decisions were reinforced by auditory feedback, which was delivered either at cardiac systole (on the heartbeat, when baroreceptors signal the contraction of the heart to the brain), or at diastole (between heartbeats during baroreceptor quiescence). We discovered a cardiac influence on feedback processing that enhanced the learning of fearful faces in people with heightened interoceptive ability. Individuals with enhanced accuracy on a heartbeat counting task learned fearful face-name pairs better when feedback was given at systole than at diastole. This effect was not present for neutral and happy faces. At retrieval, we also observed related effects of personality: First, individuals scoring higher for extraversion showed poorer retrieval accuracy. These individuals additionally manifested lower resting heart rate and lower state anxiety, suggesting that attenuated levels of cardiovascular arousal in extraverts underlies poorer performance. Second, higher extraversion scores predicted higher emotional intensity ratings of fearful faces reinforced at systole. Third, individuals scoring higher for neuroticism showed higher retrieval confidence for fearful faces reinforced at diastole. Our results show that cardiac signals shape feedback processing to influence learning of fearful faces, an effect underpinned by personality differences linked to psychophysiological arousal.


Acute plasma volume expansion in the untrained alters the hormonal response to prolonged moderate-intensity exercise.
To investigate the role of an increase in plasma volume (PV), characteristically observed with short-term endurance training, on the endocrine response to prolonged moderate intensity exercise, eight untrained males (VO2 peak = 3.52 +/- 0.12 l x min(-1)) performed 90 min of cycle ergometry at approximately 60% VO2peak both before (CON) and following (PVX) PV expansion. Acute PV expansion, which was accomplished using a solution of Dextran (6%) or Pentispan (10%) (6.7 ml kg(-1)), resulted in a calculated 15.8+/-2.2% increase (p<0.05) in PV. The prolonged exercise resulted in increases (p<0.05) in plasma vasopressin (AVP), plasma rennin activity (PRA), aldosterone (ALD), atrial naturetic peptide (alpha-ANP), and the catecholamines norepinephrine (NE) and epinephrine (EPI). PVX blunted the increases (p<0.05) in AVP, PRA, ALD, NE and EPI, during the exercise itself. The concentration of alpha-ANP was also lower (p<0.05) during exercise following PVX, an effect that could be attributed to the lower resting levels. No differences in osmolality was observed between conditions. These results demonstrate that PVX alters the fluid regulatory hormonal response in untrained subjects to moderate intensity dynamic exercise in a manner similar to that observed following short-term training induced alterations in PV. The specific mechanisms responsible for these alterations remain unclear, but appear to be related directly to the increase in PV.


Mitochondrial complex I structure reveals ordered water molecules for catalysis and proton translocation.
Mitochondrial complex I powers ATP synthesis by oxidative phosphorylation, exploiting the energy from ubiquinone reduction by NADH to drive protons across the energy-transducing inner membrane. Recent cryo-EM analyses of mammalian and yeast complex I have revolutionized structural and mechanistic knowledge and defined structures in different functional states. Here, we describe a 2.7-Å-resolution structure of the 42-subunit complex I from the yeast Yarrowia lipolytica containing 275 structured water molecules. We identify a proton-relay pathway for ubiquinone reduction and water molecules that connect mechanistically crucial elements and constitute proton-translocation pathways through the membrane. By comparison with known structures, we deconvolute structural changes governing the mammalian 'deactive transition' (relevant to ischemia-reperfusion injury) and their effects on the ubiquinone-binding site and a connected cavity in ND1. Our structure thus provides important insights into catalysis by this enigmatic respiratory machine.


BEHRTDAY: Dynamic Mortality Risk Prediction using Time-Variant COVID-19 Patient Specific Trajectories.
Incorporating repeated measurements of vitals and laboratory measurements can improve mortality risk-prediction and identify key risk factors in individualized treatment of COVID-19 hospitalized patients. In this observational study, demographic and laboratory data of all admitted patients to 5 hospitals of Mount Sinai Health System, New York, with COVID-19 positive tests between March 1st and June 8th, 2020, were extracted from electronic medical records and compared between survivors and non-survivors. Next day mortality risk of patients was assessed using a transformer-based model BEHRTDAY fitted to patient time series data of vital signs, blood and other laboratory measurements given the entire patients' hospital stay. The study population includes 3699 COVID-19 positive (57% male, median age: 67) patients. This model had a very high average precision score (0.96) and area under receiver operator curve (0.92) for next-day mortality prediction given entire patients' trajectories, and through masking, it learnt each variable's context.


Biochemical and histopathological effects on the rat testis after exposure to electromagnetic field during fetal period.
Electromagnetic radiation (ER) emitted from cell phones may exert a detrimental influence on human health and may affect the man reproductive system. We aimed to study the biological and morphological effects on the testes of 60-day-old male rats after ER exposure (900 MHz), which was applied continuously throughout embryogenesis. A total of six pregnant Sprague Dawley rats were included in the study. Three pregnant rats (experimental group) were exposed to radiation from a cell phone set to talking mode for 24 hours a day for 20 days, and the other 3 pregnant rats (control group) were not to exposed to radiation. Newborn male rats were included from the experimental group (n=7) and the control group (n=7). At the end of 60 days, the rats' testes were excised, and testis length, width, depth, and weight were measured. Histopathological examinations were compared and serum testosterone (T) levels were assayed biochemically. While serum T level (3.51±0.21 ng/ml) of ER Exposed group was significantly lower than the control group (4.04±0.47 ng/ml, p=0.018), Caspase-3 enzyme activity (2.00±0.88) was significantly higher than the control group control (1.00±0.63, p=0.026). Johnsen score (8.4±0.5) of ER group was fairly lower than the control group (9.4±0.5, p=0.010). Our study demonstrated that ER exposure throughout embryogenesis may cause reductions in serum total T levels and in the size and weight of the testes of male rats, while causing modest increase in apoptosis.


Osmotic upshift transiently inhibits uptake via ABC transporters in gram-negative bacteria.
ATP-binding cassette transporters from several rhizobia and Salmonella enterica serovar Typhimurium, but not secondarily coupled systems, were inhibited by high concentrations (100 to 500 mM) of various osmolytes, an effect reversed by the removal of the osmolyte. ABC systems were also inactivated in isolated pea bacteroids, probably due to the obligatory use of high-osmolarity isolation media. Measurement of nutrient cycling in isolated pea bacteroids is impeded by this effect.


Energy choices that europe faces: a European view of energy.
In conclusion, I feel that the energy challenge, tough as it is, does not pose unsurmountable technological problems, even in Europe. At least in principle, the necessary technology is already there. This article is meant to make that statement plausible. It is not the intention to insist on certain ideas. It is important, however, to have a consistent approach, and this means to obey the timing of the problem. Therefore the most important aspect during the transition phase probably is the buildup of a modern secondary energy system. In the long run it will ibe energy embedding and not the production of energy which will be the principal driving force for the development, because in principle at least there is more than one option to provide almost unlimited amounts of energy. In order to meet the demand for an appropriate embedding of energy, the concept of primary energy parks in the open sea seems to be most promising.


Theoretical analysis of obliquely excited surface plasmon self-interference.
We present the theoretical analysis of surface plasmon polaritons induced by a tightly focused light beam at oblique incidence. Firstly, we propose a geometrical model to explain the evolution of SPPs effect as light deviating from normal incidence, and introduce a concept of critical oblique angle (θ(co)) which is one of the key factors affecting the stability, efficiency and lateral resolution of SPPs. Secondly, the integral expressions for the transmitted SPP field excited by a linearly polarized vortex beam are derived, using angular spectrum representation and rotation matrix trans-formation, for the oblique directions as parallel and perpendicular to polarization plane. An interesting finding is that the system completely goes out of SPP self-interference resonance at an incident angle smaller than θ(co) at parallel obliquity, while larger than θ(co) at perpendicular obliquity.


Altered glycogen metabolism in cultured astrocytes from mice with chronic glutathione deficit; relevance for neuroenergetics in schizophrenia.
Neurodegenerative and psychiatric disorders including Alzheimer's, Parkinson's or Huntington's diseases and schizophrenia have been associated with a deficit in glutathione (GSH). In particular, a polymorphism in the gene of glutamate cysteine ligase modulatory subunit (GCLM) is associated with schizophrenia. GSH is the most important intracellular antioxidant and is necessary for the removal of reactive by-products generated by the utilization of glucose for energy supply. Furthermore, glucose metabolism through the pentose phosphate pathway is a major source of NADPH, the cofactor necessary for the regeneration of reduced glutathione. This study aims at investigating glucose metabolism in cultured astrocytes from GCLM knockout mice, which show decreased GSH levels. No difference in the basal metabolism of glucose was observed between wild-type and knockout cells. In contrast, glycogen levels were lower and its turnover was higher in knockout astrocytes. These changes were accompanied by a decrease in the expression of the genes involved in its synthesis and degradation, including the protein targeting to glycogen. During an oxidative challenge induced by tert-Butylhydroperoxide, wild-type cells increased their glycogen mobilization and glucose uptake. However, knockout astrocytes were unable to mobilize glycogen following the same stress and they could increase their glucose utilization only following a major oxidative insult. Altogether, these results show that glucose metabolism and glycogen utilization are dysregulated in astrocytes showing a chronic deficit in GSH, suggesting that alterations of a fundamental aspect of brain energy metabolism is caused by GSH deficit and may therefore be relevant to metabolic dysfunctions observed in schizophrenia.


The relationship between the lactate turnpoint and the time at VO2max during a constant velocity run to exhaustion.
The present investigation examined the relationship between the running velocity at the lactate turnpoint (vLTP) and the time at which VO2max can be sustained (TVO2max) during a continuous run to exhaustion at the minimal running velocity that elicits VO2max (vVO2max). Seven moderately-trained endurance runners undertook three tests on a treadmill. The first test was to determine vVO2max; the second to determine the time to exhaustion during a constant velocity run at vVO2max (Tlim vVO2max) and TVO2max; and the third to determine the vLTP. Pearson's correlation coefficient was used to determine the association between the vLTP (%vVO2max; i.e. the relative vLTP) and TVO2max, and between other selected physiological variables. Correlations between the relative vLTP and TVO2max, expressed as a percentage of T(lim vVO2max (the relative TVO2max; r=0.82), and between TVO2max and Tlim vVO2max (r=0.89), were significant at the p<0.05 level. All other correlations between selected measured physiological variables were found to be statistically insignificant. The main finding of this present study is that the relative vLTP demonstrated a significant positive correlation with the relative TVO2max. The physiological mechanism by which the lactate turnpoint may influence the relative TVO2max has not been elucidated, but may be due to a combination of decreasing the time to attain VO2max and increasing Tlim vVO2max. The present investigation has demonstrated that the lactate turnpoint may influence the relative time at which VO2max can be sustained during a continuous run to exhaustion at vVO2max, although further research is required to substantiate these findings.


Latitudinal variation in spongivorous fishes and the effectiveness of sponge chemical defenses.
It has been proposed that predation pressure declines with increasing latitude and a positive correlation exists between predation intensity and the investment into chemical defenses. However, little direct evidence supports the idea that tropical species are better defended chemically than their temperate counterparts. Temperate reefs of the South Atlantic Bight (SAB) off Georgia, USA, provide a unique opportunity to study tropical sponges in a temperate environment. We documented sponge species richness and abundance, sponge predator density, and examined the ability of eight sponge species to chemically deter predation by fishes on two reefs in the SAB. We used rarefaction analysis and ANOVA to compare our results for sponge species richness and density, respectively, with similar published studies conducted on reefs of the sub-tropical Atlantic (i.e., Florida Keys). These analyses were combined with similar statistical comparisons for spongivorous fish species richness and density. Results showed that sponge species richness was lower, but sponge density was higher, on the temperate SAB reefs than on the subtropical reefs. Both spongivorous fish diversity and density were lower on the SAB reefs. The greater abundance of sponges and lower density of predators on SAB reefs suggest a lower frequency of predation on sponges on SAB reefs. Of the eight sponge species assayed from the SAB reefs, five possessed chemical extracts that were significantly less deterrent to fish predators than their tropical/subtropical conspecifics. When the results were combined across all sponge species, the chemical deterrence of fish predators was significantly lower for extracts obtained from the temperate sponge community as compared to the tropical/subtropical assemblage. These results support the more general hypothesis that a lower density and diversity of sponge predators occurs at high as compared to low latitudes in the western Atlantic and may contribute to decreased investment in chemical defenses.


Levels of the retinoic acid synthesizing enzyme aldehyde dehydrogenase-1A2 are lower in testicular tissue from men with infertility.
To determine whether decreased testicular levels of enzymes necessary for retinoic acid biosynthesis were associated with male infertility, as retinoic acid is known to be necessary for spermatogenesis. Observational analysis of testicular tissue samples, sperm indices, and serum hormone concentrations. Two infertility centers in Chile. 32 infertile men and 11 control men. Measurement of the three enzymes necessary for retinoic acid biosynthesis, aldehyde dehydrogenase (ALDH) 1A1, 1A2, and 1A3, in testicular tissue by a novel liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) peptide assay. ALDH isozyme levels compared by type of infertility and correlated with testicular germ cell numbers, sperm parameters, and serum and intratesticular hormone concentrations. Men with infertility had statistically significantly reduced levels of ALDH1A2 but not ALDH1A1 or ALDH1A3 in their testicular tissue compared with men with normal spermatogenesis. The ALDH1A2 protein levels were strongly correlated with the number of germ cells found via testicular biopsy. These findings suggest that ALDH1A2 is the enzyme involved in retinoic acid biosynthesis in human germ cells. Further study of the relationship between intratesticular ALDH1A2 and male infertility is warranted to determine whether men with infertility have a reduced ability to synthesize retinoic acid within their germ cells that could impair spermatogenesis.


Clinical management of patients with genetic obesity during COVID-19 pandemic: position paper of the ESE Growth & Genetic Obesity COVID-19 Study Group and Rare Endo-ERN main thematic group on Growth and Obesity.
This article aims to provide guidance on prevention and treatment of COVID-19 in patients with genetic obesity. Key principals of the management of patients with genetic obesity during COVID-19 pandemic for patients that have contracted COVID-19 are to be aware of: possible adrenal insufficiency (e.g., POMC deficiency, PWS); a more severe course in patients with concomitant immunodeficiency (e.g., LEP and LEPR deficiency), although defective leptin signalling could also be protective against the pro-inflammatory phenotype of COVID-19; disease severity being masked by insufficient awareness of symptoms in syndromic obesity patients with intellectual deficit (in particular PWS); to adjust medication dose to increased body size, preferably use dosing in m2; the high risk of malnutrition in patients with Sars-Cov2 infection, even in case of obesity. Key principals of the obesity management during the pandemic are to strive for optimal obesity management and a healthy lifestyle within the possibilities of the regulations to prevent weight (re)gain and to address anxiety within consultations, since prevalence of anxiety for COVID-19 is underestimated.


Effect of sodium hypochlorite contamination on microhardness of dental core build-up materials.
This study aimed to determine the influence of sodium hypochlorite (NaOCl) contamination on the microhardness of build-up composites. Fifty-two samples, from each of three build-up materials (LuxaCore Dual, MultiCore flow and Rebilda DC) were prepared. Half of the samples from each material were stored in physiologic saline (baseline control) while the other halves were stored in NaOCl. After 1 h, the samples were rinsed with tap water, cut axially and measured for Knoop hardness at different depth levels. The results were analysed by ANOVA and unpaired t-tests (p<0.05). Significant differences in microhardness were observed for LuxaCore Dual up to 0.2 mm, Rebilda DC up to 0.3 mm, and for MultiCore flow up to 0.4 mm under the surface level. Contact with sodium hypochlorite on build-up materials causes reduction of the microhardness. The softening is not only limited on the surface, but can also be found in deeper layers of build-up materials.


L-ascorbate attenuates the endotoxin-induced production of inflammatory mediators by inhibiting MAPK activation and NF-κB translocation in cortical neurons/glia Cocultures.
In response to acute insults to the central nervous system, such as pathogen invasion or neuronal injuries, glial cells become activated and secrete inflammatory mediators such as nitric oxide (NO), cytokines, and chemokines. This neuroinflammation plays a crucial role in the pathophysiology of chronic neurodegenerative diseases. Endogenous ascorbate levels are significantly decreased among patients with septic encephalopathy. Using the bacterial endotoxin lipopolysaccharide (LPS) to induce neuroinflammation in primary neuron/glia cocultures, we investigated how L-ascorbate (vitamin C; Vit. C) affected neuroinflammation. LPS (100 ng/ml) induced the expression of inducible NO synthase (iNOS) and the production of NO, interleukin (IL)-6, and macrophage inflammatory protein-2 (MIP-2/CXCL2) in a time-dependent manner; however, cotreatment with Vit. C (5 or 10 mM) attenuated the LPS-induced iNOS expression and production of NO, IL-6, and MIP-2 production. The morphological features revealed after immunocytochemical staining confirmed that Vit. C suppressed LPS-induced astrocytic and microglial activation. Because Vit. C can be transported into neurons and glia via the sodium-dependent Vit. C transporter-2, we examined how Vit. C affected LPS-activated intracellular signaling in neuron/glia cocultures. The results indicated the increased activation (caused by phosphorylation) of mitogen-activated protein kinases (MAPKs), such as p38 at 30 min and extracellular signal-regulated kinases (ERKs) at 180 min after LPS treatment. The inhibition of p38 and ERK MAPK suppressed the LPS-induced production of inflammatory mediators. Vit. C also inhibited the LPS-induced activation of p38 and ERK. Combined treatments of Vit. C and the inhibitors of p38 and ERK yielded no additional inhibition compared with using the inhibitors alone, suggesting that Vit. C functions through the same signaling pathway (i.e., MAPK) as these inhibitors. Vit. C also reduced LPS-induced IκB-α degradation and NF-κB translocation. Thus, Vit. C suppressed the LPS-stimulated production of inflammatory mediators in neuron/glia cocultures by inhibiting the MAPK and NF-κB signaling pathways.


[Life satisfaction among elderly population in the city of Botucatu, Southern Brazil].
As a result of overall growing population's life expectancy, it has become increasingly important to ensure not only that the elderly have greater longevity but also happiness and life satisfaction. The objective of the study was to describe factors associated with life satisfaction among elderly people. Three hundred and sixty-five older persons, selected by means of random stratified proportional sampling, were interviewed in 2003. The instrument used was a combination of Flanagan and Nahas questionnaires and WHOQOL-100. There were added questions concerning physical activity extracted from International Physical Activity Questionnaire, questions regarding reported morbidity and emotional assessment, sociodemographic condition and an open question. The level of life satisfaction was measured using a scale from one to seven by means of visual recognition. Hierarchical logistic regression analysis was performed including "life satisfaction" as a dependent variable and those included the final questionnaire, in blocks, as independent variables. Most elderly were generally rather satisfied with life as well as with specific aspects. The level of life satisfaction was associated with: comfort at home (OR=11.82; 95% CI: 3.27; 42.63); appraising leisure as quality of life (OR=3.82; 95% CI: 2.28; 6.39); waking up feeling well in the morning (OR=2.80; 95% CI: 1.47; 5.36); not reporting loneliness (OR=2.68; 95% CI: 1.54; 4.65); having three or more daily meals (OR=2.63; 95% CI: 1.75; 5.90) and not reporting Diabetes Mellitus (OR=2.63; 95% CI: 1.31; 5.27). Most elderly in the study were satisfied with life and their satisfaction was associated with situations related to "being well" and not being diabetic.


Second order Jahn-Teller interactions at unusually high molecular orbital energy separations.
Second order Jahn-Teller (SOJT) effects arise from interactions between filled and empty molecular orbitals of like symmetry. These interactions often lead to structural distortions whose extent is inversely proportional to the energy difference between the interacting orbitals. The main objectives of the work described here are (1) the calculation (using density functional theory methods) of the energies of the valence molecular orbitals in the species EH3 (E = N, P, As or Sb), HEEH (E = C, Si, Ge or Sn), and H2EEH2, (E = C, Si, Ge or Sn) and (2) the correlation of these energies with barriers for planarization or linearization. The calculations suggest an upper limit of about 12 eV energy separation of the interacting levels for SOJT effects to be significant, which is considerably larger than previously thought and implies that SOJT effects may be more common than currently realized.


Can we achieve better recruitment by providing better information? Meta-analysis of 'studies within a trial' (SWATs) of optimised participant information sheets.
The information given to people considering taking part in a trial needs to be easy to understand if those people are to become, and then remain, trial participants. However, there is a tension between providing comprehensive information and providing information that is comprehensible. User-testing is one method of developing better participant information, and there is evidence that user-tested information is better at informing participants about key issues relating to trials. However, it is not clear if user-testing also leads to changes in the rates of recruitment in trials, compared to standard trial information. As part of a programme of research, we embedded 'studies within a trial' (SWATs) across multiple ongoing trials to see if user-tested materials led to better rates of recruitment. Seven 'host' trials included a SWAT evaluation and randomised their participants to receive routine information sheets generated by the research teams, or information sheets optimised through user-testing. We collected data on trial recruitment and analysed the results across these trials using random effects meta-analysis, with the primary outcome defined as the proportion of participants randomised in a host trial following an invitation to take part. Six SWATs (n=27,805) provided data on recruitment. Optimised participant information sheets likely result in little or no difference in recruitment rates (7.2% versus 6.8%, pooled odds ratio = 1.03, 95% CI 0.90 to 1.19, p-value = 0.63, I2 = 0%). Participant information sheets developed through user testing did not improve recruitment rates. The programme of work showed that co-ordinated testing of recruitment strategies using SWATs is feasible and can provide both definitive and timely evidence on the effectiveness of recruitment strategies. Healthlines Depression (ISRCTN14172341) Healthlines CVD (ISRCTN27508731) CASPER (ISRCTN02202951) ISDR (ISRCTN87561257) ECLS (NCT01925625) REFORM (ISRCTN68240461) HeLP Diabetes (ISRCTN02123133).


TRIPOD statement: a preliminary pre-post analysis of reporting and methods of prediction models.
To assess the difference in completeness of reporting and methodological conduct of published prediction models before and after publication of the Transparent Reporting of a multivariable prediction model for Individual Prognosis Or Diagnosis (TRIPOD) statement. In the seven general medicine journals with the highest impact factor, we compared the completeness of the reporting and the quality of the methodology of prediction model studies published between 2012 and 2014 (pre-TRIPOD) with studies published between 2016 and 2017 (post-TRIPOD). For articles published in the post-TRIPOD period, we examined whether there was improved reporting for articles (1) citing the TRIPOD statement, and (2) published in journals that published the TRIPOD statement. A total of 70 articles was included (pre-TRIPOD: 32, post-TRIPOD: 38). No improvement was seen for the overall percentage of reported items after the publication of the TRIPOD statement (pre-TRIPOD 74%, post-TRIPOD 76%, 95% CI of absolute difference: -4% to 7%). For the individual TRIPOD items, an improvement was seen for 16 (44%) items, while 3 (8%) items showed no improvement and 17 (47%) items showed a deterioration. Post-TRIPOD, there was no improved reporting for articles citing the TRIPOD statement, nor for articles published in journals that published the TRIPOD statement. The methodological quality improved in the post-TRIPOD period. More models were externally validated in the same article (absolute difference 8%, post-TRIPOD: 39%), used measures of calibration (21%, post-TRIPOD: 87%) and discrimination (9%, post-TRIPOD: 100%), and used multiple imputation for handling missing data (12%, post-TRIPOD: 50%). Since the publication of the TRIPOD statement, some reporting and methodological aspects have improved. Prediction models are still often poorly developed and validated and many aspects remain poorly reported, hindering optimal clinical application of these models. Long-term effects of the TRIPOD statement publication should be evaluated in future studies.


Oral antineoplastic handling at health care institutions in the United States: survey of nurses and pharmacists.
To characterize and identify trends in current practices for ordering, handling, dispensing, administering, and disposing of oral antineoplastics. An electronic survey was designed and sent to pharmacists and nurses via professional society listservs. One hundred and twenty-three practitioners responded to the survey. Of those responding, 76% described having an official policy regarding the handling of oral antineoplastics. Prescribing was limited to attending physicians or oncologists the majority of the time (42% and 98%, respectively), with 11% accepting telephone orders for oral antineoplastics. Personal protective equipment was required by many of the respondents; 70% required gloves be worn. Patient contact precautions were utilized by 79% of practitioners, of which 81% followed similar precautions for targeted therapies. Compounding was required to be performed in a biological safety cabinet by 88% of respondents, and 83% required decontamination of supplies and equipment after exposure to oral antineoplastics. Although practices varied slightly, the majority of respondents follow available guidance when handling oral antineoplastics.


Alterations in the Emotional Regulation Process in Gambling Addiction: The Role of Anger and Alexithymia.
This study aims at the assessment of alexithymia and anger levels in 100 treatment-seeking pathological gamblers compared with controls, who were matched for age, gender and education. Furthermore a positive correlation between alexithymia, anger and severity of gambling disorder and a relationship between gambling behaviour and anger after controlling for alexithymia, are investigated. Finally the role that gender plays in anger in pathological gamblers was also evaluated. Psychological assessment includes the South Oaks Gambling Screen, State-Trait Anger Expression Inventory-2 and the twenty-item Toronto Alexithymia Scale. Statistical analysis of the results shows a higher level of anger in pathological gamblers than in controls, together with alterations in emotional processing. Severity of gambling behaviour positively correlates with alexithymia scores, state-anger and trait-anger. Moreover, a significant contribution of anger in predicting gambling behaviour was suggested after controlling for alexithymia. In conclusion, anger and alexithymia must be regarded as relevant components of the assessment of pathological gamblers, in order to select the best therapeutical strategies to prevent self-defeating behaviours and to reduce drop-out from treatments.


Giant placental chorioangioma: natural history and pregnancy outcome.
Giant placental chorioangiomas are associated with a high prevalence of pregnancy complications and a poor perinatal outcome. The aim of the study was to evaluate the natural history, intrauterine treatment and outcome of pregnancies complicated by giant placental chorioangioma. This was a retrospective study of 19 cases of giant placental chorioangioma, in which the natural history, intrauterine treatment and outcome of pregnancy were evaluated. Eighteen of the 19 cases were associated with a wide variety of fetal complications, including polyhydramnios, growth restriction, hyperdynamic circulation, cardiomegaly, anemia and non-immune hydrops. Two-thirds of the cases developed complications that required either elective delivery for fetal growth restriction (n = 6) or intervention for cardiovascular effects (n = 7). Fetal therapy, when required, was tailored to the particular complication and resulted in a successful pregnancy in all cases. The presence of large placental chorioangioma is associated with the development of fetal growth restriction and hyperdynamic state in a significant number of cases. This series elucidates some of the putative underlying mechanisms for these complications, and reports on a safe and effective treatment modality, percutaneous ultrasound-guided interstitial laser therapy.


The role of mercury and cadmium heavy metals in vascular disease, hypertension, coronary heart disease, and myocardial infarction.
Mercury, cadmium, and other heavy metals have a high affinity for sulfhydryl (-SH) groups, inactivating numerous enzymatic reactions, amino acids, and sulfur-containing antioxidants (NAC, ALA, GSH), with subsequent decreased oxidant defense and increased oxidative stress. Both bind to metallothionein and substitute for zinc, copper, and other trace metals reducing the effectiveness of metalloenzymes. Mercury induces mitochondrial dysfunction with reduction in ATP, depletion of glutathione, and increased lipid peroxidation; increased oxidative stress is common. Selenium antagonizes mercury toxicity. The overall vascular effects of mercury include oxidative stress, inflammation, thrombosis, vascular smooth muscle dysfunction, endothelial dysfunction, dyslipidemia, immune dysfunction, and mitochondrial dysfunction. The clinical consequences of mercury toxicity include hypertension, CHD, MI, increased carotid IMT and obstruction, CVA, generalized atherosclerosis, and renal dysfunction with proteinuria. Pathological, biochemical, and functional medicine correlations are significant and logical. Mercury diminishes the protective effect of fish and omega-3 fatty acids. Mercury, cadmium, and other heavy metals inactivate COMT, which increases serum and urinary epinephrine, norepinephrine, and dopamine. This effect will increase blood pressure and may be a clinical clue to heavy metal toxicity. Cadmium concentrates in the kidney, particularly inducing proteinuria and renal dysfunction; it is associated with hypertension, but less so with CHD. Renal cadmium reduces CYP4A11 and PPARs, which may be related to hypertension, sodium retention, glucose intolerance, dyslipidemia, and zinc deficiency. Dietary calcium may mitigate some of the toxicity of cadmium. Heavy metal toxicity, especially mercury and cadmium, should be evaluated in any patient with hypertension, CHD, or other vascular disease. Specific testing for acute and chronic toxicity and total body burden using hair, toenail, urine, serum, etc. with baseline and provoked evaluation should be done.


2015 proceedings of the National Heart, Lung, and Blood Institute's State of the Science in Transfusion Medicine symposium.
On March 25 and 26, 2015, the National Heart, Lung, and Blood Institute sponsored a meeting on the State of the Science in Transfusion Medicine on the National Institutes of Health (NIH) campus in Bethesda, Maryland, which was attended by a diverse group of 330 registrants. The meeting's goal was to identify important research questions that could be answered in the next 5 to 10 years and which would have the potential to transform the clinical practice of transfusion medicine. These questions could be addressed by basic, translational, and/or clinical research studies and were focused on four areas: the three "classical" transfusion products (i.e., red blood cells, platelets, and plasma) and blood donor issues. Before the meeting, four working groups, one for each area, prepared five major questions for discussion along with a list of five to 10 additional questions for consideration. At the meeting itself, all of these questions, and others, were discussed in keynote lectures, small-group breakout sessions, and large-group sessions with open discourse involving all meeting attendees. In addition to the final lists of questions, provided herein, the meeting attendees identified multiple overarching, cross-cutting themes that addressed issues common to all four areas; the latter are also provided. It is anticipated that addressing these scientific priorities, with careful attention to the overarching themes, will inform funding priorities developed by the NIH and provide a solid research platform for transforming the future practice of transfusion medicine.


Reduced ceftazidime and ertapenem susceptibility due to production of OXA-2 in Klebsiella pneumoniae ST258.
OXA-2 is a class D β-lactamase that confers resistance to penicillins, as well as narrow-spectrum cephalosporins. OXA-2 was recently reported to also possess carbapenem-hydrolysing activity. Here, we describe a KPC-2-encoding Klebsiella pneumoniae isolate that demonstrated reduced susceptibility to ceftazidime and ertapenem due to production of OXA-2. To elucidate the role of OXA-2 production in reduced ceftazidime and ertapenem susceptibility in a K. pneumoniae ST258 clinical isolate. MICs were determined by the agar dilution method. WGS was conducted to identify and compare resistance genes between isolates. Expression of KPC-2 was quantified by quantitative RT-PCR and immunoblotting. OXA-2 was expressed in Escherichia coli TOP10, as well as in K. pneumoniae ATCC 13883, to define the relative contribution of OXA-2 in β-lactam resistance. Kinetic studies were conducted using purified OXA-2 enzyme. K. pneumoniae 1761 belonged to ST258 and carried both blaKPC-2 and blaOXA-2. However, expression of blaKPC-2 was substantially reduced due to an IS1294 insertion in the promoter region. K. pneumoniae 1761, K. pneumoniae ATCC 13883 and E. coli TOP10 carrying blaOXA-2-harbouring plasmids showed reduced susceptibility to ertapenem and ceftazidime, but meropenem, imipenem and cefepime were unaffected. blaOXA-2 was carried on a 2910 bp partial class 1 integron containing aacA4-blaOXA-2-qacEΔ1-sul1 on an IncA/C2 plasmid, which was not present in the earlier ST258 isolates possessing blaKPC-2 with intact promoters. Hydrolysis of ertapenem by OXA-2 was confirmed using purified enzyme. Production of OXA-2 was associated with reduced ceftazidime and ertapenem susceptibility in a K. pneumoniae ST258 isolate.


Architecture of cell-cell junctions in situ reveals a mechanism for bacterial biofilm inhibition.
Many bacteria, including the major human pathogen Pseudomonas aeruginosa, are naturally found in multicellular, antibiotic-tolerant biofilm communities, in which cells are embedded in an extracellular matrix of polymeric molecules. Cell-cell interactions within P. aeruginosa biofilms are mediated by CdrA, a large, membrane-associated adhesin present in the extracellular matrix of biofilms, regulated by the cytoplasmic concentration of cyclic diguanylate. Here, using electron cryotomography of focused ion beam-milled specimens, we report the architecture of CdrA molecules in the extracellular matrix of P. aeruginosa biofilms at intact cell-cell junctions. Combining our in situ observations at cell-cell junctions with biochemistry, native mass spectrometry, and cellular imaging, we demonstrate that CdrA forms an extended structure that projects from the outer membrane to tether cells together via polysaccharide binding partners. We go on to show the functional importance of CdrA using custom single-domain antibody (nanobody) binders. Nanobodies targeting the tip of functional cell-surface CdrA molecules could be used to inhibit bacterial biofilm formation or disrupt preexisting biofilms in conjunction with bactericidal antibiotics. These results reveal a functional mechanism for cell-cell interactions within bacterial biofilms and highlight the promise of using inhibitors targeting biofilm cell-cell junctions to prevent or treat problematic, chronic bacterial infections.


Immune cells have sex and so should journal articles.
Males and females have the same immunological cells, proteins, and pathways in place to protect against the development of disease. The kinetics, magnitude, and skewing of the responses mounted against pathogens, allergens, toxins, or self-antigens, however, can differ dramatically between the sexes. Generally, females mount higher innate and adaptive immune responses than males, which can result in faster clearance of pathogens but also contributes to increased susceptibility to inflammatory and autoimmune diseases in females compared with males. Hormonal and genetic factors contribute significantly to sex differences in immune function and disease pathogenesis. In particular, the expression of X-linked genes and microRNA as well as sex steroid hormones signaling through hormone receptors in immune cells can affect responses to immunological stimuli differently in males and females. Despite data illustrating profound differences between the sexes in immune function, sex differences in the pathogenesis of disease are often overlooked in biomedical research. Establishing journal policies that require authors to report the sex of their cells, animals, and subjects will improve our understanding of the pathogenesis of diseases, with the long-term goal of personalizing treatments for immune-mediated diseases differently for males and females in an effort to protect us equally.


Blastocystis: to treat or not to treat...
Parasites in the genus Blastocystis comprise several subtypes (genotypes) and have a worldwide distribution. In some surveys, these are the most common parasites found in human stool specimens. An emerging literature suggests that the pathogenicity of Blastocystis is related to specific subtypes and parasite burden, although even individuals with small numbers of cysts may be symptomatic. Some data suggest an association between infection with Blastocystis and irritable bowel syndrome. However, there are few clinical studies demonstrating a direct relationship between the presence of this parasite and disease, few animal models to explore this relationship, and no consensus as to appropriate treatment. We recommend that asymptomatic individuals with few cysts not be treated. However, those who have gastrointestinal or dermatologic signs and symptoms and many cysts in stool specimens may require treatment. Metronidazole is the drug of choice. Additional studies are required to determine pathogenicity and appropriate therapy.


Multicentre, randomised controlled trial to investigate the effects of parental touch on relieving acute procedural pain in neonates (Petal).
Newborn infants routinely undergo minor painful procedures as part of postnatal care, with infants born sick or premature requiring a greater number of procedures. As pain in early life can have long-term neurodevelopmental consequences and lead to parental anxiety and future avoidance of interventions, effective pain management is essential. Non-pharmacological comfort measures such as breastfeeding, swaddling and sweet solutions are inconsistently implemented and are not always practical or effective in reducing the transmission of noxious input to the brain. Stroking of the skin can activate C-tactile fibres and reduce pain, and therefore could provide a simple and safe parent-led intervention for the management of pain. The trial aim is to determine whether parental touch prior to a painful clinical procedure provides effective pain relief in neonates. This is a multicentre randomised controlled trial. A total of 112 neonates born at 35 weeks' gestation or more requiring a blood test in the first week of life will be recruited and randomised to receive parental stroking either preprocedure or postprocedure. We will record brain activity (EEG), cardiac and respiratory dynamics, oxygen saturation and facial expression to provide proxy pain outcome measures. The primary outcome will be the reduction of noxious-evoked brain activity in response to a heel lance. Secondary outcomes will be a reduction in clinical pain scores (Premature Infant Pain Profile-Revised), postprocedural tachycardia and parental anxiety. The study has been approved by the London-South East Research Ethics Committee (ref: 21/LO/0523). The results will be widely disseminated through peer-reviewed publications, international conferences and via our partner neonatal charities Bliss and Supporting the Sick Newborn And their Parents (SSNAP). If the parental tactile intervention is effective, recommendations will be submitted via the National Health Service clinical guideline adoption process. Commenced September 2021. NCT04901611; 14 135 962.