Hypoxic-ischemic encephalopathy: pathophysiology and implications for therapy.
Hypoxic-ischemic encephalopathy continues to be a major problem in perinatal medicine because of the high mortality rate and neurological and intellectual impairment in the surviving infants. In addition to the acute necrotic damage that occurs in the neurons initially, reperfusion injury and secondary neuronal damage continue for 6 to 72 hours after the initial insult. Secondary cellular energy failure, membrane breakdown, cellular influx of calcium ions, elaboration of cytokines, and oxidation of excitory amino acids all contribute to enhanced apoptosis of the neurons. Newer forms of therapy including the use of oxygen-free radical inhibitors and mild to moderate cerebral or systemic hypothermia for 72 hours following the asphyxial period are promising adjuncts as follow-up approaches to the affected newborns.


Hyaluronic Acid Modified Hollow Prussian Blue Nanoparticles Loading 10-hydroxycamptothecin for Targeting Thermochemotherapy of Cancer.
This paper reported the fabrication of a multifunctional nanoplatform by modifying hollow Prussian blue nanoparticles with hyaluronic acid grafting polyethylene glycol, followed by loading 10-hydroxycamptothecin for tumor-targeted thermochemotherapy. It was found that the surface modification of hollow Prussian blue nanoparticles with hyaluronic acid grafting polyethylene endowed a great colloidal stability, long blood circulation time and the capability for targeting Hela cells over-expressing the CD44 receptor. The obtained nanoagent exhibited efficient photothermal effect and a light triggered and stepwise release behavior of 10-hydroxycamptothecin due to the strong optical absorption in the near-infrared region. The investigations on the body weight change, histological injury and blood biochemical indexes showed that such nanoagent had excellent biocompatibility for medical application. Both in vitro and in vivo experiments proved that the combination of chemotherapy and photothermal therapy through the agent of hyaluronic acid modified Prussian blue nanoparticles loading 10-hydroxycamptothecin could significantly improve the therapeutic efficacy compared with either therapy alone because of a good synergetic effect.


Bioavailability of tocotrienols: evidence in human studies.
As a minor component of vitamin E, tocotrienols were evident in exhibiting biological activities such as neuroprotection, radio-protection, anti-cancer, anti-inflammatory and lipid lowering properties which are not shared by tocopherols. However, available data on the therapeutic window of tocotrienols remains controversial. It is important to understand the absorption and bioavailability mechanisms before conducting in-depth investigations into the therapeutic efficacy of tocotrienols in humans. In this review, we updated current evidence on the bioavailability of tocotrienols from human studies. Available data from five studies suggested that tocotrienols may reach its target destination through an alternative pathway despite its low affinity for α-tocopherol transfer protein. This was evident when studies reported considerable amount of tocotrienols detected in HDL particles and adipose tissues after oral consumption. Besides, plasma concentrations of tocotrienols were shown to be higher when administered with food while self-emulsifying preparation of tocotrienols was shown to enhance the absorption of tocotrienols. Nevertheless, mixed results were observed based on the outcome from 24 clinical studies, focusing on the dosages, study populations and formulations used. This may be due to the variation of compositions and dosages of tocotrienols used, suggesting a need to understand the formulation of tocotrienols in the study design. Essentially, implementation of a control diet such as AHA Step 1 diet may influence the study outcomes, especially in hypercholesterolemic subjects when lipid profile might be modified due to synergistic interaction between tocotrienols and control diet. We also found that the bioavailability of tocotrienols were inconsistent in different target populations, from healthy subjects to smokers and diseased patients. In this review, the effect of dosage, composition and formulation of tocotrienols as well as study populations on the bioavailability of tocotrienols will be discussed.


Molecular Insights into the Inhibitory Role of α-Crystallin against γD-Crystallin Aggregation.
Cataracts, a major cause of global blindness, contribute significantly to the overall prevalence of blindness. The opacification of the lens, resulting in cataract formation, primarily occurs due to the aggregation of crystallin proteins within the eye lens. Despite the high concentration of these crystallins, they remarkably maintain the lens transparency and refractive index. α-Crystallins (α-crys), acting as chaperones, play a crucial role in preventing crystallin aggregation, although the exact molecular mechanism remains uncertain. In this study, we employed a combination of molecular docking, all-atom molecular dynamics simulations, and advanced free energy calculations to investigate the interaction between γD-crystallin (γD-crys), a major structural protein of the eye lens, and α-crystallin proteins. Our findings demonstrate that α-crys exhibits an enhanced affinity for the NTD2 and CTD4 regions of γD-crys. The NTD2 and CTD4 regions form the interface between the N-terminal domain (NTD) and the C-terminal domain (CTD) of the γD-crys protein. By binding to the interface region between the NTD and CTD of the protein, α-crys effectively inhibits the formation of domain-swapped aggregates and mitigates protein aggregation. Analysis of the Markov state models using molecular dynamics trajectories confirms that minimum free energy conformations correspond to the binding of the α-crystallin domain (ACD) of α-crys to NTD2 and CTD4 that form the interdomain interface.


Serum creatine phosphokinase as prognostic indicator in the management of electrical burn.
To determine the mean difference of serum creatine phosphokinase according to the extent of tissue damage and prognosis of the patients suffering from electrical burn injuries. A case series study. Department of Burns, Liaquat University Hospital, Hyderabad, from September 2005 to August 2006. During the study period, 38 patients with electrical burn injuries were enrolled. Victims of electrical burns with evidence of myocardial injury were excluded. Following admission, serum creatine phosphokinase was measured serially on 10 consecutive occasions. The data was later analyzed statistically using SPSS-10.0. Of the 38 patients, the mean age of the victims was 28 years, with males dominating the study population (82%). A statistically significant association was found between the level of serum creatine phosphokinase and likelihood of death (p=0.000). It was also found that serial monitoring of this enzyme can be used as prognostic indicator in the management of electrical burns injury. The level of creatine phosphokinase increased with the degree of tissue injuries in patients with electrical burns. This prognostic value is of great importance in the local setup, where sophisticated investigations to detect extent of injuries are not available.


Hydrogen- and oxygen from water.
The limitations of thermochemical energy storage devices are the limitations of Carnot devices. Entropy production entailed in product separation further limits the efficiency of thermochemical processes. Thus, high upper temperatures and few reaction steps are desirable. In this article, the one-step effusional separation of water into hydrogen and oxygen is considered. Membrane materials, design, and fabrication techniques are suggested. A parametric analysis of the process suggests that the idea is a tantalizing possibility.


Strategy to Identify Paramedic Transported Sepsis Cases in an Emergency Department Administrative Database.
Background: To evaluate a new strategy for identifying sepsis in Emergency Department (ED) patients that combines administrative diagnosis codes with clinical information from the point of first contact. Methods: This study linked clinical data from adult patients transported by a provincial Emergency Medical Services (EMS) system to ED and inpatient administrative databases. Sepsis cases were identified by searching ED databases for diagnosis codes consistent with infection and organ dysfunction. Organ dysfunction was further assessed using a partial Sequential Organ Failure Assessment (SOFA) score derived from EMS clinical information. Reliability was evaluated by comparing patients' ED diagnosis codes (ICD-10CA) to their inpatient diagnosis codes; criterion validity by comparing cases classified by the new strategy to an existing inpatient administrative algorithm; and construct validity by assessing for clinical characteristics typically associated with sepsis (e.g., mortality). Results: A total of 43,297 patients were included. ED infection codes were more reliable for classifying patients with infection than using ED sepsis codes alone (proportion of agreement with inpatient codes 79% vs. 74%; p-value < 0.001). The novel strategy requiring the presence of an infection code and either an organ dysfunction code or 2 or more SOFA points from EMS clinical information identified 1,379 more ED patients as having sepsis than the inpatient algorithm. These patients had high mortality supporting construct validity. Conclusions: Incorporation of a broader range of diagnostic codes and linking to an electronic database to obtain initial clinical information for the assessment of organ dysfunction improves reliability, criterion, and construct validity for identifying sepsis in ED patients.


Coronary revascularisation outcomes in patients with cancer.
Cancer and coronary artery disease (CAD) overlap in traditional risk factors as well as molecular mechanisms underpinning the development of these two disease states. Patients with cancer are at increased risk of developing CAD, representing a high-risk population that are increasingly undergoing coronary revascularisation. Over 1 in 10 patients with CAD that require revascularisation with either percutaneous coronary intervention or coronary artery bypass grafting have either a history of cancer or active cancer. These patients are typically older, have more comorbidities and have more extensive CAD compared with patients without cancer. Haematological abnormalities with competing risks of thrombosis and bleeding pose further unique challenges during and after revascularisation. Management of patients with concurrent cancer and CAD requiring revascularisation is challenging as these patients carry a higher risk of morbidity and mortality compared with those without cancer, often driven by the underlying cancer and associated comorbidities. However, due to variability by different types and stages of cancer, revascularisation outcomes are specific to cancer characteristics such as the timing of onset, cancer subtype and site, stage, presence of metastases, and cancer-related therapies received. Recent studies have provided insights into defining revascularisation outcomes, procedural considerations and best practices in managing patients with cancer. Nevertheless, many gaps remain that require further studies to inform clinical best practices in this population.


Biotransformation of nitrosamines and precursor secondary amines under methanogenic conditions.
The biotransformation potential of six nitrosamines and their precursor secondary amines by a mixed methanogenic culture was investigated. Among the six nitrosamines tested, N-nitrosodimethylamine (NDMA), N-nitrosomethylethylamine (NMEA), and N-nitrosopyrrolidine (NPYR) were almost completely degraded but only when degradable electron donors were available. On the contrary, N-nitrosodiethylamine (NDEA), N-nitrosodipropylamine (NDPA), and N-nitrosodibutylamine (NDBA) were not degraded. Three precursor secondary amines, corresponding to the three biodegradable nitrosamines, were also completely utilized even with very low levels of available electron donors. The secondary amine precursors of the three, nonbiodegradable nitrosamines were also recalcitrant. A bioassay conducted to elucidate the biotransformation pathway of NDMA in the mixed methanogenic culture using H(2) as the electron donor showed that NDMA was utilized as an electron acceptor and transformed to dimethylamine (DMA), which in turn was degraded to ammonia and methane. The H(2) threshold concentration for NDMA bioreduction ranged between 0.0017 and 0.031 atm. Such a high H(2) threshold concentration suggests that in mixed methanogenic cultures, NDMA reducers are weak competitors to other, H(2)-consuming microbial species, such as homoacetogens and methanogens. Thus, complete removal of nitrosamines in anaerobic digestion systems, where the H(2) partial pressure is typically below 10(-4) atm, is difficult to achieve.


Opt-out testing for hepatitis B and C infections in adults attending the emergency department of a large London teaching hospital.
The National Health Service (NHS) in England commissioned opt-out testing in London Emergency Departments (ED) in April 2022 to allow early identification and management of hepatitis B (HBV) and hepatitis C virus (HCV) infection in patients unaware of their infection status. All adults over the age of 16 undergoing blood tests in the ED at the Royal Free Hospital were tested for HBV surface antigen and anti-HCV IgG unless they opted out. Data was collected between the 12th of April and 22nd of August 2022. Of 11,215 patients tested for HCV, 164 patients were found to be anti-HCV IgG positive, giving a seroprevalence rate of 1.46 %. 52 of the anti-HCV IgG positive patients did not have any previous HCV serology result. 23 of the anti-HCV IgG positive patients were also HCV RNA positive giving an RNA seroprevalence of 0.21 %, and 17 of those were new diagnoses of HCV viraemia. For HBV testing, 82 (0.73 %) out of 11,192 patients tested were found to be HBsAg positive, including one patient who presented acutely with a positive HBV core IgM. 39 of the HBsAg positive patients were previously unknown to us; of these, 9 had an HBV viral load of more than 2000 IU/mL, including 3 patients with positive HBV e antigen and one patient with hepatitis D virus co-infection. Opt-out screening of HBV and HCV in ED is effective at identifying patients with previously undiagnosed viral hepatitis infection and providing an opportunity to engage them in specialist care.


Accelerated cytotoxicity mechanism screening using drug metabolising enzyme modulators.
By assessing how drug/new chemical entity (NCE) cytotoxicity is affected when their metabolic pathways are inhibited or activated, the metabolic pathways that activate versus detoxify drugs/NCEs can be identified. Reactive metabolites contributing to cytotoxicity can also be identified. In the following, the drug metabolizing enzyme inhibitors and activators used in vitro with freshly isolated rat hepatocytes for the accelerated cytotoxicity mechanism screening (ACMS) of drugs/NCEs (a technique used in our laboratory) are reviewed and, this technique is useful for determining in vivo rat hepatotoxicity mechanisms. The enzyme inhibitors/activators have been chosen on the basis of their selectivity, modulator effectiveness, and their lack of toxicity. The use of these inhibitors/activators with human hepatocytes or subcellular fractions for assessing human hepatotoxicity mechanisms is also reviewed.


Relationships between inhalable and total hexavalent chromium exposures in steel passivation, welding and electroplating operations of Ontario.
The main aim of the study was to assess the relationship between inhalable hexavalent chromium and "total" hexavalent chromium. Air sampling was conducted at steel passivation operation of a steel manufacturer at a stainless steel welding operation and at two hard chrome electroplaters. Air samples were taken side-by-side for "total" dust using closed-face 37-mm diameter cassette samplers and for inhalable dust using Institute of Occupational Medicine inhalable samplers. A total of 40 pairs of total and inhalable dust samples were collected and later analyzed. The range of "total" dust and inhalable dust concentrations in μg/m3 measured were 30-410 and 0.02 to 740 respectively for steel passivation; 260 to 1520 and 477 to 6970 for welding; and 0.01 to 1500 and 204 to 2130 for electroplaters. The range of "total" dust hexavalent chromium and inhalable dust hexavalent chromium concentrations in μg/m3 were 0.02-89 and 0.02 to 150 respectively for steel making; 4.1 to 4.9 and 2.2 to 6.9 for welding and 0.01 to 9.3 and 0.01 to 21 for electroplaters. A linear relationship between inhalable hexavalent chromium and "total" hexavalent chromium was found with a slope of 1.4 (CI:1.3, 1.5) and 0 offset. A ratio of 1.4 can thus be used as a conversion factor to convert previous data of hexavalent chromium taken on "total" dust basis to inhalable hexavalent chromium concentrations.


Capturing of the monoterpene olefin limonene produced in Saccharomyces cerevisiae.
Monoterpene olefins such as limonene are plant compounds with applications as flavouring and fragrance agents, as solvents and potentially also in polymer and fuel chemistry. We engineered baker's yeast Saccharomyces cerevisiae to express a (-)-limonene synthase from Perilla frutescens and a (+)-limonene synthase from Citrus limon. Both proteins were expressed either with their native plastid targeting signal or in a truncated form in which the plastidial sorting signal was removed. The yeast host strain for expression was AE9 K197G, which expresses a mutant Erg20 enzyme. This enzyme catalyses the formation of geranyl diphosphate, which is the precursor for monoterpenes. Several methods were tested to capture limonene produced by the yeast. Extraction from the culture medium by pentane, or by the addition of CaCl2 followed by solid-phase micro-extraction, did not lead to detectable limonene, indicating that limonene is rapidly lost from the culture medium. Volatile terpenes such as limonene may also be trapped in a dodecane phase added to the medium during fermentation. This method resulted in recovery of 0.028 mg/l (+)-limonene and 0.060 mg/l (-)-limonene in strains using the truncated Citrus and Perilla synthases, respectively. Trapping the headspace during culture of the limonene synthase-expressing strains resulted in higher titres, at 0.12 mg/l (+)-limonene and 0.49 mg/l (-)-limonene. These results show that the volatile properties of the olefins produced require specific methods for efficient recovery of these molecules from biotechnological production systems.


Feasibility of high-intensity interval training and moderate-intensity continuous training in adults with inactive or mildly active Crohn's disease: study protocol for a randomised controlled trial.
Structured exercise training has been proposed as a useful adjunctive therapy for Crohn's disease by improving immune function and psychological health, reducing fatigue and promoting gains in muscle and bone strength. However, the evidence for exercise in Crohn's disease is sparse, with only a handful of small prospective trials [1, 2], with methodological limitations, including the use of non-randomised and non-controlled study designs and small sample sizes. Here, we describe the protocol for a study that aims to assess the feasibility and acceptability of two common types of exercise training-high-intensity interval training (HIIT) and moderate-intensity continuous training (MICT)-in adults with inactive or mildly active Crohn's disease (CD). This is a randomised, controlled, assessor-blinded, feasibility trial with three parallel groups. Forty-five adults with inactive or mildly active Crohn's disease will be randomly assigned 1:1:1 to HIIT, MICT or usual care control. Participants in the HIIT and MICT groups will be invited to undertake three sessions of supervised exercise each week for 12 consecutive weeks. HIIT sessions will consist of ten 1-min intervals of cycling exercise at 90% of peak power output separated by 1 min of active recovery. MICT sessions will involve 30 min of continuous cycling at 35% of peak power output. Participants will be assessed before randomisation and 13 and 26 weeks after randomisation. Feasibility outcomes include rates of recruitment, retention and adherence. Interviews with participants will explore the acceptability of the exercise programmes and study procedures. Clinical/health outcomes include cardiorespiratory fitness, body mass index, resting blood pressure, markers of disease activity (faecal calprotectin and Crohn's Disease Activity Index) and activated T cell cytokine profiles. Study questionnaires include the Inflammatory Bowel Disease Quality of Life Questionnaire, EQ-5D-5L, IBD Fatigue Scale, Hospital and Anxiety Depression Scale, and International Physical Activity Questionnaire. This study will provide useful information on the feasibility and acceptability of supervised exercise training in adults with inactive and mildly active Crohn's disease and will inform the design of a subsequent, adequately powered, multi-centre trial. The trial is registered with the International Standard Randomised Controlled Trial Register (ISRCTN13021107). Date registration assigned was 02/12/2015.


Isolation, characterization, and comparison of recombinant DNAs derived from genomes of human hepatitis B virus and woodchuck hepatitis virus.
The human hepatitis B virus (HBV) and the woodchuck hepatitis virus (WHV) are closely related by several criteria and belong to the same class of DNA viruses. The DNA genomes from these viruses are difficult to obtain in quantities required for biochemical analysis. We have, therefore, cloned these two DNAs in the vector lambda gtWES and subcloned into the kanamycin resistance plasmid pA01. Comparison of the recombinant DNAs with authentic viral DNAs by specific hybridization, size, and restriction enzyme analysis suggests that the recombinants contain the complete genome of each virus. The nominal size of the cloned HBV genome was 3150 base pairs, compared to 3200 base pairs for the cloned WHV genome. The small amount of nucleic acid homology previously reported between the HBV and WHV DNAs could be demonstrated between the cloned viral DNAs.


Serological, hematologic, and PCR studies of cattle in an area of Switzerland in which tick-borne fever (caused by Ehrlichia phagocytophila) is endemic.
The purpose of this study was to examine the seasonal variations in seroprevalence to Ehrlichia phagocytophila in cattle pastured during the summer months in an area where tick-borne fever is endemic. The study was performed during a 1-year period from April 1996 to March 1997 and involved 34 cows, 22 pregnant heifers, and 14 calves. Blood samples, collected from all 70 cattle once a month, were used to determine serum immunoglobulin G titers by indirect immunofluorescence. In addition, blood smears were examined for Ehrlichia organisms, and PCR amplification was performed for the molecular detection of E. phagocytophila. Prior to the pasture period, the seroprevalence was 16%. Two weeks after the start of pasturing, it was 43%, after which it progressively increased and reached a maximum of 63% in September. The seroprevalence progressively decreased after the end of pasturing to a low of 23%. The variation in antibody titers was similar to that of seroprevalence. E. phagocytophila organisms were detected in blood smears of 7 animals and by nested PCR in 12. Only four cows, which were on the pastures of endemicity for the first time, had clinical signs of ehrlichiosis. This study demonstrated marked seasonal variations in seroprevalence and in serum titers of antibody to E. phagocytophila in cattle. The incidence of clinical signs of ehrlichiosis was increased in cattle grazing on the pastures of endemicity for the first time.


Re-envisioning paediatric nurse training in a re-engineered health care system.
South African's infant and child mortality rates remain high and at the current rate of decline will not meet the Millennium Development Goals of a two thirds decrease by 2015. At the latest available count, there were fewer than 1500 qualified paediatric nurses on the National South African Nursing Council register, with only about 100 nurses graduating with this qualification from South African nursing schools annually. It is not clear, however, if current paediatric nurse training programmes adequately equip nurses to make a real impact on reducing the under-5 mortality rate. In their 2011 interim report, the Ministerial Committee on Morbidity and Mortality in Children under 5 years recommended strengthening paediatric nurses' training as a strategy to reduce the under-5 mortality rate. In response to the Committee on Morbidity and Mortality in Children recommendation, a colloquium was convened as a national forum for schools of nursing, departments of health, health care facilities, clinicians and regulatory bodies to advance children's nursing in South Africa. The goals of the colloquium were to thoroughly investigate the situation in South Africa's paediatric nurse training, plot ways to strengthen and expand postgraduate paediatric programmes to meet priority child health needs, and to build relationships between the various schools and stakeholders. Outcomes included the clarification and strengthening of a 'stakeholder grid' in nurse training, recognition of the need for more active teaching and learning strategies in curricula linked to national child health priorities, as well as the need to develop and support clinical nursing practice in facilities.


Identification and functional assay of an extracellular calcium-sensing receptor in Necturus gastric mucosa.
In mammals and amphibians, increases in extracellular Ca2+ can activate bicarbonate secretion and other protective functions of gastric mucosa. We hypothesized that the recently cloned extracellular Ca(2+)-sensing receptor (CaR) is functioning in the gastric mucosa. In Necturus maculosus gastric mucosa, reverse transcription-polymerase chain reaction using primers based on previously cloned CaR sequences amplified a 326-bp DNA fragment that had 84% nucleotide sequence identity with the rat kidney CaR. Immunohistochemical localization of the CaR using specific anti-CaR antiserum revealed its presence on the basal aspect of gastric epithelial cells. In microelectrode studies of Necturus antral mucosa, exposure to elevated Ca2+ (4.8 mM) and the CaR agonists NPS-467 and neomycin sulfate resulted in significant hyperpolarizations of basal membrane electrical potentials and increases in apical-to-basal membrane resistance ratios. Circuit analysis revealed that these changes reflected specific decreases in basolateral membrane resistance. Inhibition of prostaglandin synthesis using indomethacin significantly attenuated these effects. We conclude that the CaR is present and functioning in Necturus gastric antrum.


Facial suture synostosis of newborn Fgfr1(P250R/+) and Fgfr2(S252W/+) mouse models of Pfeiffer and Apert syndromes.
Apert and Pfeiffer syndromes are hereditary forms of craniosynostosis characterized by midfacial hypoplasia and malformations of the limbs and skull. A serious consequence of midfacial hypoplasia in these syndromes is respiratory compromise due to airway obstruction. In this study, we have evaluated Fgfr1(P250R/+) and Fgfr2(S252W/+) mouse models of these human conditions to study the pathogenesis of midfacial hypoplasia. Our histologic and micro-CT evaluation revealed premature synostosis of the premaxillary-maxillary, nasal-frontal, and maxillary-palatine sutures of the face and dysplasia of the premaxilla, maxilla, and palatine bones. These midfacial abnormalities were detected in the absence of premature ossification of the cranial base at postnatal day 0. Our results indicate that midfacial hypoplasia is not secondary to premature cranial base ossification but rather primary synostosis of facial sutures. Birth Defects Research (Part A), 2011.


Assessing pulmonary circulation in severe bronchopulmonary dysplasia using functional echocardiography.
Pulmonary hypertension (PH) is common in infants with severe bronchopulmonary dysplasia (BPD) and increases the risk of death. The objectives of this preliminary study were to compare responses of pulmonary circulation parameters to 100% oxygen (O2 ) and inhaled nitric oxide (iNO) in infants with BPD and PH using echocardiography. Responses between fetal growth restriction (FGR) and appropriate for gestational age infants were compared. Ten infants <28 weeks GA at birth were assessed at ≥36 weeks corrected gestation. Baseline echocardiography1 was performed which was repeated (echocardiography2) after 30 minutes of O2 . After a gap of 2-3 hours, iNO was administered for 15 minutes and echocardiography3 was performed, followed by iNO weaning. The gestation and birthweight of the cohort were 25.9 ± 1.6 weeks and 612 ± 175 g. Assessments were performed at 38.7 ± 1.4 weeks corrected gestational age. Baseline time to peak velocity: right ventricular ejection time (TPV/RVETc) increased from 0.24 ± 0.02 to 0.27 ± 0.02 (O2 , p = .01) and 0.31 ± 0.03 (iNO, p < .001), indicating a decrease in pulmonary vascular resistance [PVR]. Baseline tricuspid annular plane systolic excursion (TAPSE) increased from 8.1 ± 0.6 mm to 9.3 ± 0.7 mm (O2 , p = .01) and 10.5 ± 1.1 mm (iNO, p = .0004), indicating improved ventricular systolic performance. Percentage change for all parameters was greater with iNO. Significant correlations between cardiac performance and PVR were noted. FGR infants noted higher baseline PVR (TPV/RVETc, 0.21 ± 0.02 vs. 0.25 ± 0.01, p = .002), lower ventricular performance (TAPSE, 7 ± 1.2 mm vs. 8.6 ± 6 mm, p = .003), and lower percentage change with O2 and iNO. A reactive component of pulmonary circulation provides real-time physiological information, which could rationalize treatment decisions.


Dissociative protonation sites: reactive centers in protonated molecules leading to fragmentation in mass spectrometry.
It is often found in mass spectrometry that when a molecule is protonated at the thermodynamically most favorable site, no fragmentation occurs, but a major reaction is observed when the proton migrates to a different position. For benzophenones, acetophenones, and dibenzyl ether, which are all preferentially protonated at the oxygen, deacylation or dealkylation was observed in the collision-induced dissociation of the protonated molecules. For para-monosubstituted benzophenones, electron-withdrawing substituents favor the formation of RC6H4CO+ (R = substituent), whereas electron-releasing groups favor the competing reaction leading to C6H5CO+. The ln[(RC6H4CO+)/(C6H5CO+)] values are well-correlated with the sigmap+ substituent constants. In the fragmentation of protonated acetophenones, deacetylation proceeds to give an intermediate proton-bound dimeric complex of ketene and benzene. The distribution of the product ions was found to depend on the proton affinities of ketene and substituted benzenes, and the kinetic method was applied in identifying the reaction intermediate. Protonated dibenzyl ether loses formaldehyde upon dealkylation, via an ion-neutral complex of the benzyloxymethyl cation and neutral benzene. These gas-phase retro-Friedel-Crafts reactions occurred as a result of the attack of the proton at the carbon atom to which the carbonyl or the methylene group is attached on the aromatic ring, which is described as the dissociative protonation site.


Monitoring the He+ ion channel formation by high-order harmonic generation.
The macroscopic build-up of the high-order harmonic signal depends on the free electron density in the generation medium. The free electrons affect the harmonic yield and spectral shape through modifying the refractive index and the phase matching conditions. These dependences allow studying the He(+) ion channel formation in a He gas jet. The evolution of an ion channel created by an ultrashort laser pulse via optical field ionization was monitored using the harmonic signal generated by a collinear propagating second laser pulse. From the measured high harmonic signal as function of the delay we are able to gain information about the free electron density. Under our experimental condition, the ion channel has been fully formed 300 fs after the first laser pulse, resulting in an enhancement of harmonic yield of the second laser pulse by two orders of magnitude.


Generation and characterization of transgenic mice expressing tamoxifen-inducible cre-fusion protein specifically in mouse liver.
To establish transgenic mice expressing tamoxifen-inducible Cre-ERt recombinase specifically in the liver and to provide an efficient animal model for studying gene function in the liver and creating various mouse models mimicking human diseases. Alb-Cre-ERt transgenic mice were produced by microinjecting the construct with Cre-ERt fusion gene of DNA fragments into fertilized eggs derived from inbred C57BL/6 strain. Transgenic mice were identified by using PCR and Southern blotting. Expression of Cre-ERt fusion gene was analyzed in the liver, kidney, brain and lung from F1 generation transgenic mice at 8 weeks of age by reverse transcription (RT)-PCR. Four hundred and fourteen fertilized eggs of C57 BL/6 mice were microinjected with recombinant Alb-Cre-ERt DNA fragments, and 312 survival eggs injected were transferred to the oviducts of 12 pseudopregnant recipient mice, 6 of 12 recipient mice became pregnant and gave birth to 44 offsprings. Of the 44 offsprings, two males and one female carried the hybrid Cre-ERt fusion gene. Three mice were determined as founders, and were back crossed to set up F1 generations with other inbred C57BL/6 mice. Transmission of Cre-ERt fusion gene in F1 offspring followed Mendelian rules. The expression of Cre-ERt mRNA was detected only in the liver of F1 offspring from two of three founder mice. Transgenic mice expressing tamoxifen-inducible Cre-ERt recombinase under control of the liver-specific promoter are preliminary established.


Mobile phones as fomites for potential pathogens in hospitals: microbiome analysis reveals hidden contaminants.
Smartphones used in clinical settings harbour potentially pathogenic bacteria, and this may pose an infection risk. Previous studies have relied on culture-based methods. To characterize the quantity and diversity of microbial contamination of hospital staff smartphones using culture-dependent and culture-independent methods; to determine the prevalence of antibiotic-resistant potential pathogens; to compare microbial communities of hospital staff and control group phones. Smartphones of 250 hospital staff and 191 control group participants were swabbed. The antibiotic resistance profile of Staphylococcus aureus and Enterococcus isolates was determined. Swabs were pooled into groups according to the hospital area staff worked in, and DNA was extracted. The microbial community of the phone was characterized using an Illumina MiSeq metabarcoding pipeline. Almost all (99.2%) of hospital staff smartphones were contaminated with potential pathogens, and bacterial colony forming units (CFUs) were significantly higher on hospital phones than in the control group. Meticillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus (VRE) were only detected on hospital mobile phones. Metabarcoding revealed a far greater abundance of Gram-negative contaminants, and much greater diversity, than culture-based methods. Bacillus spp. were significantly more abundant in the hospital group. This study reinforces the need to consider infection-control policies to mitigate the potential risks associated with the increased use of smartphones in clinical environments, and highlights the limitations of culture-based methods for environmental swabbing.


The visual story of data storage: From storage properties to user interfaces.
About fifty times more data has been created than there are stars in the observable universe. Current trends in data creation and consumption mean that the devices and storage media we use will require more physical space. Novel data storage media such as DNA are considered a viable alternative. Yet, the introduction of new storage technologies should be accompanied by an evaluation of user requirements. To assess such needs, we designed and conducted a survey to rank different storage properties adapted for visualization. That is, accessibility, capacity, usage, mutability, lifespan, addressability, and typology. Withal, we reported different storage devices over time while ranking them by their properties. Our results indicated a timeline of three distinct periods: magnetic, optical and electronic, and alternative media. Moreover, by investigating user interfaces across different operating systems, we observed a predominant presence of bar charts and tree maps for the usage of a medium and its file directory hierarchy, respectively. Taken together with the results of our survey, this allowed us to create a customized user interface that includes data visualizations that can be toggled for both user groups: Experts and Public.


Machine learning on genome-wide association studies to predict the risk of radiation-associated contralateral breast cancer in the WECARE Study.
The purpose of this study was to identify germline single nucleotide polymorphisms (SNPs) that optimally predict radiation-associated contralateral breast cancer (RCBC) and to provide new biological insights into the carcinogenic process. Fifty-two women with contralateral breast cancer and 153 women with unilateral breast cancer were identified within the Women's Environmental Cancer and Radiation Epidemiology (WECARE) Study who were at increased risk of RCBC because they were ≤ 40 years of age at first diagnosis of breast cancer and received a scatter radiation dose > 1 Gy to the contralateral breast. A previously reported algorithm, preconditioned random forest regression, was applied to predict the risk of developing RCBC. The resulting model produced an area under the curve (AUC) of 0.62 (p = 0.04) on hold-out validation data. The biological analysis identified the cyclic AMP-mediated signaling and Ephrin-A as significant biological correlates, which were previously shown to influence cell survival after radiation in an ATM-dependent manner. The key connected genes and proteins that are identified in this analysis were previously identified as relevant to breast cancer, radiation response, or both. In summary, machine learning/bioinformatics methods applied to genome-wide genotyping data have great potential to reveal plausible biological correlates associated with the risk of RCBC.


Endotoxemia, inflammation, and atrial fibrillation.
There is emerging evidence to support a link between inflammation and atrial fibrillation (AF). Lipopolysaccharide (LPS) infusion is a well established experimental model used to investigate host systemic inflammatory responses. It was hypothesized that LPS challenge, by virtue of its provoked host inflammatory response, might increase the propensity to the development of new-onset AF. A post hoc analysis was performed of prospective data collected for 652 healthy men (mean age 27+/-5 years, all without history of AF) who were challenged with LPS according to a standard experimental protocol. All subjects underwent a detailed health screening before inclusion. After LPS challenge, all subjects underwent continuous cardiac monitoring for a minimum of 8 hours and were reviewed again using rhythm assessments at 24 hours and after 7+/-3 days. Effects of LPS on high-sensitivity C-reactive protein, interleukin-6, tumor necrosis factor-alpha, and neutrophil counts as indexes of an inflammatory response were also assessed. LPS led to overall marked increases in high-sensitivity C-reactive protein, interleukin-6, tumor necrosis factor-alpha, and neutrophil counts (all p<0.0001) and an average temperature increase of 1.1 degrees C. There was no evidence of new-onset AF in the subjects challenged. In conclusion, experimental LPS challenge led to a significant increase in acute inflammatory indexes, but did not increase the propensity to new-onset AF in a young low-risk population.


Processing of adenovirus 2-induced proteins.
Analysis of (35)S-methionine-labeled extracts of adenovirus 2-infected KB cells revealed 22 virus-induced polypeptide components. Most proteins of the virion were easily detected in extracts of whole cells labeled for short periods between 15 and 30 h after infection; however, several virion components were conspicuously absent. Radioactivity appeared in two of these virion components during a chase in nonradioactive medium, and this appearance was paralleled by a decrease in the radioactivity associated with two nonvirion adenovirus-induced proteins, results which imply precursor-product relationships for these components. Comparison of one of the chasable adenovirus-induced components (designated P-VII; mass of 20,000 daltons) and the major core protein (VII; mass of 18,500 daltons) of the virion showed that they have four common methionine-containing tryptic peptides; P-VII has an additional methionine residue which is not found in the major core protein. We propose that at least two of the adenovirus 2 virion components are derived by the cleavage of higher molecular weight precursor polypeptides.


[Radiographic planning and the evaluation of implant treatment].
Radiographic techniques can be used during the pre-, per- and postoperative stages of implant treatment. In the preoperative stage information is needed regarding the quantity and quality of the bone and the relevant anatomical structures of the jaw. Various radiographic techniques are available for this and all have their inherent merits and disadvantages. Occasionally there is a need of a preoperative radiograph i.e. a panoramic or periapical radiograph. In the postoperative stage radiographic techniques are used for the evaluation of bone and implants. In this stage there is a need for detailed and reproducible information regarding the cervical peri-implant bone level, the total height of the jaw and the radiographic density of the peri-implant bone. There is a limited number of radiographic techniques available in this stage.


Minimally invasive in vivo human lung tissue bioimpedance measurements during the bronchoscopy procedure.
Respiratory diseases, which include diseases of the lung, pleura, bronchial tree, trachea, upper respiratory tract and of the respiratory muscles and nerves, are a common and important cause of illness and death among the population. Experimental evidences have shown that tissue lesions have different electrical properties compared with normal tissue. Therefore, lung tissues lesions may be differentiated from lung normal tissue by comparing the tissue passive electrical properties. The manuscript reports a feasibility study for minimally invasive in vivo human lung tissue tetrapolar bioimpedance measurements using a catheter during the bronchoscopy procedure based on multisine Electrical Impedance Spectroscopy (EIS) at 10 kHz - 1 MHz.


[Comprehensive quality indicators for measuring hospital antibiotic use].
As in other countries, the growing resistance to antimicrobial drugs is also taking place in the Netherlands; the primary cause being the total consumption of antibiotics. Given the steady decline in the discovery of new antimicrobials, better use of agents currently available is warranted. Guidelines describing appropriate antimicrobial therapy play an important role; however, such guidelines are not optimally used in daily practice. Quality indicators can be used to assess the quality of antibiotic treatment and evaluate the impact of interventions aimed at improving care. Quality indicators used for evaluating treatment of infections of the respiratory and urinary tracts are developed previously. A comprehensive set of indicators that could be used to assess the quality of hospital antibiotic use for all bacterial infections has not yet been developed. A new project has recently been started in the Netherlands called 'The development of Reliable generic quality Indicators for the optimalisation of ANTibiotic use in the hospital' (RIANT study) for developing such a set of comprehensive indicators.


Acid exposure disrupts mucus secretion and impairs mucociliary transport in neonatal piglet airways.
Tenacious mucus produced by tracheal and bronchial submucosal glands is a defining feature of several airway diseases, including cystic fibrosis (CF). Airway acidification as a driving force of CF airway pathology has been controversial. Here we tested the hypothesis that transient airway acidification produces pathologic mucus and impairs mucociliary transport. We studied pigs challenged with intra-airway acid. Acid had a minimal effect on mucus properties under basal conditions. However, cholinergic stimulation in acid-challenged pigs revealed retention of mucin 5B (MUC5B) in the submucosal glands, decreased concentrations of MUC5B in the lung lavage fluid, and airway obstruction. To more closely mimic a CF-like environment, we also examined mucus secretion and transport following cholinergic stimulation under diminished bicarbonate and chloride transport conditions ex vivo. Under these conditions, airways from acid-challenged pigs displayed extensive mucus films and decreased mucociliary transport. Pretreatment with diminazene aceturate, a small molecule with ability to inhibit acid detection through blockade of the acid-sensing ion channel (ASIC) at the doses provided, did not prevent acid-induced pathologic mucus or transport defects but did mitigate airway obstruction. These findings suggest that transient airway acidification early in life has significant impacts on mucus secretion and transport properties. Furthermore, they highlight diminazene aceturate as an agent that might be beneficial in alleviating airway obstruction.


Prevalence and genetic diversity of Batrachochytrium dendrobatidis in Central African island and continental amphibian communities.
The fungal pathogen Batrachochytrium dendrobatidis (Bd) infects hundreds of amphibian species and is implicated in global amphibian declines. Bd is comprised of several lineages that differ in pathogenicity, thus, identifying which Bd strains are present in a given amphibian community is essential for understanding host-pathogen dynamics. The presence of Bd has been confirmed in Central Africa, yet vast expanses of this region have not yet been surveyed for Bd prevalence, and the genetic diversity of Bd is largely unknown in this part of the world. Using retrospective surveys of museum specimens and contemporary field surveys, we estimated the prevalence of Bd in Central African island and continental amphibian assemblages, and genotyped strains of Bd present in each community. Our sampling of museum specimens included just a few individuals collected in the Gulf of Guinea archipelago prior to 1998, yet one of these individuals was Bd-positive indicating that the pathogen has been on Bioko Island since 1966. We detected Bd across all subsequent sample years in our study and found modest support for a relationship between host life history and Bd prevalence, a positive relationship between prevalence and host community species richness, and no significant relationship between elevation and prevalence. The Global Panzootic Lineage (Bd GPL) was present in all the island and continental amphibian communities we surveyed. Our results are consistent with a long-term and widespread distribution of Bd in amphibian communities of Gabon and the Gulf of Guinea archipelago.


Human disturbance causes the formation of a hybrid swarm between two naturally sympatric fish species.
Most evidence for hybrid swarm formation stemming from anthropogenic habitat disturbance comes from the breakdown of reproductive isolation between incipient species, or introgression between allopatric species following secondary contact. Human impacts on hybridization between divergent species that naturally occur in sympatry have received considerably less attention. Theory predicts that reinforcement should act to preserve reproductive isolation under such circumstances, potentially making reproductive barriers resistant to human habitat alteration. Using 15 microsatellites, we examined hybridization between sympatric populations of alewife (Alosa pseudoharengus) and blueback herring (A. aestivalis) to test whether the frequency of hybridization and pattern of introgression have been impacted by the construction of a dam that isolated formerly anadromous populations of both species in a landlocked freshwater reservoir. The frequency of hybridization and pattern of introgression differed markedly between anadromous and landlocked populations. The rangewide frequency of hybridization among anadromous populations was generally 0-8%, whereas all landlocked individuals were hybrids. Although neutral introgression was observed among anadromous hybrids, directional introgression leading to increased prevalence of alewife genotypes was detected among landlocked hybrids. We demonstrate that habitat alteration can lead to hybrid swarm formation between divergent species that naturally occur sympatrically, and provide empirical evidence that reinforcement does not always sustain reproductive isolation under such circumstances.


Bilateral hegu acupoints have the same effect on the heart rate variability of the healthy subjects.
Background. The specificity of acupuncture points (acupoints) is one of the key concepts in traditional acupuncture theory, but the question of whether there is adequate scientific evidence to prove or disprove specificity has been vigorously debated in recent years. Acupoint laterality is an important aspect of acupoint specificity. Data is particularly scarce regarding the laterality of the same channel, namesake acupoint located on opposite sides of the body. Our previous study results suggest that Neiguan acupoint (PC6) has the laterality. The aim of this study was to investigate whether Hegu (LI4) also has laterality from the perspective of heart rate variability. Methods. A total of twenty-eight healthy female volunteers were recruited for this study and were randomly separated into the group I (n = 14) and the group II (n = 14) according to the register order. In the group I, left LI4 was stimulated in the first epoch and the right LI4 was stimulated in the second epoch. In the group II, right LI4 was stimulated in the first epoch and left LI4 was stimulated in the second epoch. Electrocardiogram was recorded and heart rate variability was analyzed. Results. The results show that there were no significant differences of heart rate variablity between the group I and the group II in the time domain and in the frequency domain. Conclusions. Bilateral Hegu acupoints have the same effect on the heart rate variability of the healthy subjects.


Healthcare reform: it's working in Oregon.
Oregon is beginning to operate a highly controversial program with explicit definitions of coverage. Oregon's Director of the Health Plan, Jean Thorne, has focused on the national and state political attention that Oregon is receiving and knows that "...there are many people who are waiting for us to fail." Thorne has requested that we keep our focus on making major changes to improve the chance that more people in poverty will receive coverage. We continue to believe that the Oregon Health Plan will improve overall healthcare in our state and serve as a blueprint for others to study and improve.


Development and application of stable carbon isotope analysis to the detection of cortisol administration in cattle.
The use of anabolic agents in food-producing animals has been prohibited within the EU since 1988. The control of the illegal use of natural steroid hormones in cattle is still an exciting analytical challenge as no definitive method and nonambiguous analytical criteria are available. We have used gas chromatography/combustion/isotope ratio mass spectrometry (GC/C/IRMS) to demonstrate the administration of cortisol to cattle. The method consisted of an efficient combination between OASIS HLB solid-phase extraction (SPE), oxidation, SiOH SPE and semi-preparative high-performance liquid chromatography (HPLC) for glucocorticoid purification. By comparison of the (13)C/(12)C isotopic ratio of the oxidised product of cortisol, i.e. 5 beta-androstane-3,11,17-trione (5 beta AAT), with an endogenous reference compound (ERC), dehydroepiandrosterone (DHEA), the differentiation of cortisol metabolite origin, either endogenous or exogenous, has been achieved. After treatment of an animal, the delta(13)C(VPDB) values of 5 beta AAT reached -30 to -32 per thousand, whereas the delta(13)C(VPDB) values of DHEA remained at -25 per thousand. A significant difference in the delta(13)C(VPDB) values between DHEA and 5 beta AAT was measurable over a period of 3 days after a single administration of cortisol to the animal.


Altered Bulbar Conjunctival Microcirculation in Response to Contact Lens Wear.
This study was conducted to determine blood flow velocities and corresponding vessel diameters to characterize the response of the bulbar conjunctival microvasculature to contact lens wear. A functional slit-lamp biomicroscope (FSLB), an adapted traditional slitlamp, was used to image the temporal bulbar conjunctiva of 22 healthy subjects before and after 6 hr of contact lens wear. All of the measurable venules on the conjunctiva were processed to yield vessel diameters and blood flow velocities. The average blood flow velocity increased from 0.51±0.20 to 0.65±0.22 mm/sec (P<0.001) after 6 hr of lens wear. The blood flow velocity distribution showed a velocity increase that correlated with the vessel diameter increase from the baseline (r=0.826, P<0.05). This pattern maintained a similar trend after 6 hr of lens wear (r=0.925, P<0.05), and increased velocities were found across all of the vessel diameter ranges (P<0.001). Blood flow velocity increases across all of the vessel diameter ranges in response to contact lens wear. Functional slitlamp biomicroscope is capable of characterizing the bulbar microvascular response to contact lens wear.


Patient Positioning During In-Office Otologic Procedures Impacts Physician Ergonomics.
The purpose of this study was to evaluate the impact of patient positioning on physician ergonomics during in-office otologic procedures. A previous simulation study suggested that placing patients supine during in-office otology procedures is ergonomically favorable. This study aims to substantiate these findings during the routine care of patients in an otolaryngology practice setting. Observational study. Outpatient otology clinic within tertiary care facility. Two neurotology attending physicians. Physicians performed cerumen removal procedures in the office with patients either in the seated position (n = 24) or supine position (n = 24). The rapid upper limb assessment (RULA), a validated instrument that measures body positioning with a focus on the upper arm, was used to measure ergonomic positioning. RULA scores correlate occupational body positioning with a numeric representation of musculoskeletal injury risk ranging from 1 (minimal risk) to 7 (very high risk). Overall median RULA scores were 4.5 (medium risk) with patients in the seated position, and 2 (low risk) with patients in the supine position (p < 0.0001). Similarly, RULA scores were significantly lower with patients in the supine position when each physician was evaluated independently (p < 0.0001, for both). Placing patients in a supine position for cerumen removal results in more favorable ergonomic positioning for the physician, thus reducing risk for work-related musculoskeletal disorders. This suggests that physicians should consider placing patients in the supine position for in-office otologic procedures. Further study is needed to investigate optimal ergonomic positioning for other common otolaryngology procedures.


Neopterin activates transcription factor nuclear factor-kappa B in vascular smooth muscle cells.
We have previously shown that the pteridine compound neopterin stimulates inducible nitric oxide synthase (iNOS) gene expression in vascular smooth muscle cells in vitro. The mechanisms whereby neopterin exhibits these effects remained unclear. The present study demonstrates that neopterin induces the translocation of the transcription factor nuclear factor-kappa B (NF-kappa B) to the nucleus. Pretreatment of cells with the antioxidant pyrrolidine dithiocarbamate completely suppressed the effects of neopterin on NF-kappa B activation, iNOS gene expression, and nitric oxide release. From these data we conclude that neopterin activates the translocation of NF-kappa B subunits to the nucleus by modulating the intracellular redox state. This is one possible explanation for the impact of neopterin on iNOS gene expression.


Landscape heterogeneity drives intra-population niche variation and reproduction in an arctic top predator.
While intra-population variability in resource use is ubiquitous, little is known of how this measure of niche diversity varies in space and its role in population dynamics. Here we examined how heterogeneous breeding environments can structure intra-population niche variation in both resource use and reproductive output. We investigated intra-population niche variation in the Arctic tundra ecosystem, studying peregrine falcon (Falco peregrinus tundrius, White) breeding within a terrestrial-marine gradient near Rankin Inlet, Nunavut, Canada. Using stable isotope analysis, we found that intra-population niches varied at the individual level; we examined within-nest and among-nest variation, though only the latter varied along the terrestrial-marine gradient (i.e., increased among-nest variability among birds nesting within the marine environment, indicating higher degree of specialization). Terrestrial prey species (small herbivores and insectivores) were consumed by virtually all falcons. Falcons nesting within the marine environment made use of marine prey (sea birds), but depended heavily on terrestrial prey (up to 90% of the diet). Using 28-years of peregrine falcon nesting data, we found a positive relationship between the proportion of terrestrial habitat surrounding nest sites and annual nestling production, but no relationship with the likelihood of successfully rearing at least one nestling reaching 25 days old. Annually, successful inland breeders raised 0.47 more young on average compared to offshore breeders, which yields potential fitness consequences for this long-living species. The analyses of niche and reproductive success suggest a potential breeding cost for accessing distant terrestrial prey, perhaps due to additional traveling costs, for those individuals with marine nest site locations. Our study indicates how landscape heterogeneity can generate proximate (niche variation) and ultimate (reproduction) consequences on a population of generalist predator. We also show that within-individual and among-individual variation are not mutually exclusive, but can simultaneously arise and structure intra-population niche variation.


[Has the age at sexual debut changed after the Safe Sex Campaign?].
In 1984, immediately before information about AIDS had reached the general public and the campaign for "Safe sex" had begun in earnest, 286 women (percentage replies 75.3) and 336 men (percentage replies 77.8) who were over 16 years but not yet 21 years were interviewed about sexual knowledge, sexual experience and contraception. In October 1989, the investigation was repeated and 359 women (percentage replies 77.9) and 400 men (percentage replies 76.3) participated. Both of the investigations were carried out as personal interviews by the professional interviewer staff of the Institute for Social Research. The age profiles for the first coitus were found to be practically identical in the two investigations. The median age for the first coitus was calculated to be 16.7 years in 1984 and 16.8 years in 1989 for females and 16.9 years for males in both 1984 and 1989. In both of the investigations, the majority stated that the first coitus was with a person of the same age group. Just under 1/4 of the women and nearly 1/3 of the men stated that the first coitus was also the first for the partner. In the 1989 investigation, significantly more stated that they had employed contraception at the first coitus. The campaign for "Safe sex" had recommended condoms. In the 1989 investigation, 61.0% of the women and 62.0% of the men had employed condoms at the first coitus as compared with 36.5% of the women and 42.1% of the men, five years previously. Parental advice about contraception had also changed as a great many more stated that their parents had advised them to use condoms as a contraceptive measure in the 1989 investigation. Among the young people who had not yet had coitus, 24.4% of the women and 34.7% of the men stated in 1989 that they always had a condom handy when they thought they might need it.


Survival of patients treated with online hemodiafiltration compared to conventional hemodialysis.
Accumulating data from observational studies showed that online hemodiafiltration (OLHDF) might improve survival in chronic hemodialysis (HD) patients. According to this data, the aim of our study was to investigate whether there was a difference in survival of patients treated with OLHDF compared to standard, conventional HD. We included 85 prevalent patients with end-stage renal disease (ESRD) treated with HD as a method of renal replacement therapy (RRT) for more than three months. Patients were previously treated with HD and divided into two groups: in 42 patients new treatment with OLHDF was introduced, and 43 patients were treated with HD. Both groups were followed over a period of 36 months. The study showed significantly better survival of patients treated with OLHDF, compared to the survival of patients treated with HD in the whole study population, as well as in the subgroups of diabetics, of patients who were on RRT with HD for more than five years and of the patients who were older than 65 years. In the nondiabetics, patients who were on RRT for less than five years and in the patients who were younger than 65 years, survival results in the OLHDF group were not significantly better compared to those in the HD group. As in our study, there are accumulating data from observational studies that HDF may improve survival in chronic HD patients, but new, prospective randomized trials are needed to support evidence about this hypothesis.


A STATISTICAL MODEL TO ASSESS (ALLELE-SPECIFIC) ASSOCIATIONS BETWEEN GENE EXPRESSION AND EPIGENETIC FEATURES USING SEQUENCING DATA.
Sequencing techniques have been widely used to assess gene expression (i.e., RNA-seq) or the presence of epigenetic features (e.g., DNase-seq to identify open chromatin regions). In contrast to traditional microarray platforms, sequencing data are typically summarized in the form of discrete counts, and they are able to delineate allele-specific signals, which are not available from microarrays. The presence of epigenetic features are often associated with gene expression, both of which have been shown to be affected by DNA polymorphisms. However, joint models with the flexibility to assess interactions between gene expression, epigenetic features and DNA polymorphisms are currently lacking. In this paper, we develop a statistical model to assess the associations between gene expression and epigenetic features using sequencing data, while explicitly modeling the effects of DNA polymorphisms in either an allele-specific or nonallele-specific manner. We show that in doing so we provide the flexibility to detect associations between gene expression and epigenetic features, as well as conditional associations given DNA polymorphisms. We evaluate the performance of our method using simulations and apply our method to study the association between gene expression and the presence of DNase I Hypersensitive sites (DHSs) in HapMap individuals. Our model can be generalized to exploring the relationships between DNA polymorphisms and any two types of sequencing experiments, a useful feature as the variety of sequencing experiments continue to expand.


Microbial mineralization of ring-substituted anilines through an ortho-cleavage pathway.
Moraxella sp. strain G is able to utilize as sole source of carbon and nitrogen aniline, 4-fluoroaniline, 2-chloroaniline, 3-chloroaniline, 4-chloroaniline (PCA), and 4-bromoaniline but not 4-iodoaniline, 4-methylaniline, 4-methoxyaniline, or 3,4-dichloroaniline. The generation time on PCA was 6 h. The pathway for the degradation of PCA was investigated by analysis of catabolic intermediates and enzyme activities. Mutants of strain G were isolated to enhance the accumulation of specific pathway intermediates. PCA was converted by an aniline oxygenase to 4-chlorocatechol, which in turn was degraded via a modified ortho-cleavage pathway. Synthesis of the aniline oxygenase was inducible by various anilines. This enzyme exhibited a broad substrate specificity. Its specific activity towards substituted anilines seemed to be correlated more with the size than with the electron-withdrawing effect of the substituent and was very low towards anilines having substituents larger than iodine or a methyl group. The initial enzyme of the modified ortho-cleavage pathway, catechol 1,2-dioxygenase, had similar characteristics to those of corresponding enzymes of pathways for the degradation of chlorobenzoic acid and chlorophenol, that is, a broad substrate specificity and high activity towards chlorinated and methylated catechols.


Cyclosporine A versus methotrexate in the treatment of polymyositis and dermatomyositis.
To determine the effectiveness and tolerance of treatment with cyclosporine A (CyA) or methotrexate (MTX) added to corticosteroids in patients with severe, active polymyositis (PM) and dermatomyositis (DM). Thirty-six patients (20 with DM, 16 with PM) were enrolled into the study and randomized in MTX (n = 17) and CyA (n = 19) groups. Muscle endurance and functional test (MEFT), clinical assessment (CA), global patient's assessment (GPA), muscle MRI, serum CK, myoglobin, IL-1Ra, and autoantibody status were used to assess the response to therapy after 1, 3, and 6 months. Significant improvement in MEFT, CA, GPA, and muscle MRI was found in both groups. Patients treated with MTX showed insignificantly better response than patients with CyA. CK levels in the MTX group decreased significantly after 1, 3, and 6 months, whereas a significant reduction in the CyA group was first observed after 6 months. IL-1Ra serum levels significantly dropped in the CyA group after two weeks, whereas in the MTX group the significant decrease was first seen after 3 months of treatment. Good correlation was found between each of the clinical parameters (MEFT, CA, and GPA), none of them showed any correlation with CK or IL-1Ra levels. Administration of MTX or CyA added to corticosteroids was associated with clinical and laboratory improvement. Changes in CK and IL-1Ra levels were not associated with parameters of clinical disease severity measured in this study.


ECG mapping and imaging of cardiac electrical function.
Inverse electrocardiography has been developed for several years. By coupling electrocardiographic mapping and 3D-time anatomical data, the electrical excitation sequence can be imaged completely non-invasively in the human heart. In this study, a bidomain theory based surface heart model activation time imaging approach was applied to single beat data of atrial and ventricular depolarization. For sinus and paced rhythms, the sites of early activation and the areas with late activation were estimated with sufficient accuracy. In particular for focal arrhythmias, this model-based imaging approach might allow the guidance and evaluation of antiarrhythmic interventions, for instance, in case of catheter ablation or drug therapy.


Visible-light-controlled homo- and copolymerization of styrenes by a bichromophoric Ir-Pd catalyst.
Visible-light-controlled polymerization was achieved by a bichromophoric organopalladium catalyst which possesses a naphthyl-substituted cyclometallated Ir(III) light-absorbing moiety. The complex was highly active toward styrene polymerization upon visible-light irradiation, and its photoactivity toward polymerization and dimerization was switchable. On the basis of the switching activity, controlled copolymerization of styrene and vinyl ether was achieved upon photo-irradiation to give the corresponding copolymers.


Bioorthogonal Bond Cleavage Chemistry for On-demand Prodrug Activation: Opportunities and Challenges.
Time- and space-resolved drug delivery is highly demanded for cancer treatment, which, however, can barely be achieved with a traditional prodrug strategy. In recent years, the prodrug strategy based on a bioorthogonal bond cleavage chemistry has emerged with the advantages of high temporospatial resolution over drug activation and homogeneous activation irrespective of individual heterogeneity. In the past five years, tremendous progress has been witnessed in this field with one such bioorthogonal prodrug entering Phase II clinical trials. This Perspective aims to highlight these new advances (2019-2023) and critically discuss their pros and cons. In addition, the remaining challenges and potential strategic directions for future progress will also be included.


[The two-in-one hit model of the short-cut carcinogenesis of colorectal carcinomas in MLH1-associated Lynch syndrome].
In a recently published study a new genetic hypothesis was established that explained the existence of CTNNB1 mutations in Lynch syndrome-associated colorectal carcinomas (MLH1-LS-CRC). This hypothesis states that a mitotic recombination on chromosome 3p simultaneously leads to inactivation of the mismatch repair gene MLH1 and to the activation of CTNNB1. This explains the increased frequency of CTNNB1 mutations in MLH1-LS-CRC compared with other colon carcinomas. To test this hypothesis, various experiments were carried out that show that the first phase of recombination occurs in non-cancerous tissues, which favours the development of CTNNB1 mutations. This mechanism could explain the rapid tumour progression in MLH1-LS-CRC. The results highlight the importance of mitotic recombination in carcinogenesis and provide an insight into the genetic basis of colorectal carcinoma in the context of Lynch syndrome.


Genome-wide association study of bipolar disorder accounting for effect of body mass index identifies a new risk allele in TCF7L2.
Bipolar disorder (BD) is associated with higher body mass index (BMI) and increased metabolic comorbidity. Considering the associated phenotypic traits in genetic studies of complex diseases, either by adjusting for covariates or by investigating interactions between genetic variants and covariates, may help to uncover the missing heritability. However, obesity-related traits have not been incorporated in prior genome-wide analyses of BD as covariates or potential interacting factors. To investigate the genetic factors underlying BD while considering BMI, we conducted genome-wide analyses using data from the Genetic Association Information Network BD study. We analyzed 729,454 genotyped single-nucleotide polymorphism (SNP) markers on 388 European-American BD cases and 1020 healthy controls with available data for maximum BMI. We performed genome-wide association analyses of the genetic effects while accounting for the effect of maximum BMI, and also evaluated SNP-BMI interactions. A joint test of main and interaction effects demonstrated significant evidence of association at the genome-wide level with rs12772424 in an intron of TCF7L2 (P=2.85E-8). This SNP exhibited interaction effects, indicating that the bipolar susceptibility risk of this SNP is dependent on BMI. TCF7L2 codes for the transcription factor TCF/LF, part of the Wnt canonical pathway, and is one of the strongest genetic risk variants for type 2 diabetes (T2D). This is consistent with BD pathophysiology, as the Wnt pathway has crucial implications in neurodevelopment, neurogenesis and neuroplasticity, and is involved in the mechanisms of action of BD and depression treatments. We hypothesize that genetic risk for BD is BMI dependent, possibly related to common genetic risk with T2D.


[Not Available]
How was medicine-taught and studied in the early modern period? How did teachers and students relate to the tradition of medical texts? Historians have addressed these questions mainly through research on printed sources. This article uses student notes as a window on medical training in action in Montpellier. The student was Franqois Boissier de Sauvages; the teacher, Jacques Lazerme. The notes record Lazerme lecturing in the form of commentary on the first 14 Hippocratic Aphorisms and setting quite different emphases from those of earlier commentators: professional ethics and how to handle a case (on aphorism 1), moderation in. bloodletting (on 2), disease classification (on 4-6), similarity of symptoms among different diseases (on 7), treating fever (on 8-io), periodicity of disease (on ii), weather and disease (on iz), "calor innatus" and oscillations in the circulation of the blood, with reference to Isaac Newton (on 14). These commentaries allow a fresh look at medical training between theory and practice.


Association Between Genetically Proxied Inhibition of HMG-CoA Reductase and Epithelial Ovarian Cancer.
Preclinical and epidemiological studies indicate a potential chemopreventive role of statins in epithelial ovarian cancer risk. To evaluate the association of genetically proxied inhibition of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase (ie, genetic variants related to lower function of HMG-CoA reductase, target of statins) with epithelial ovarian cancer among the general population and in BRCA1/2 mutation carriers. Single-nucleotide polymorphisms (SNPs) in HMGCR, NPC1L1, and PCSK9 associated with low-density lipoprotein (LDL) cholesterol in a genome-wide association study (GWAS) meta-analysis (N ≤196 475) were used to proxy therapeutic inhibition of HMG-CoA reductase, Niemann-Pick C1-Like 1 (NPC1L1) and proprotein convertase subtilisin/kexin type 9 (PCSK9), respectively. Summary statistics were obtained for these SNPs from a GWAS meta-analysis of case-control analyses of invasive epithelial ovarian cancer in the Ovarian Cancer Association Consortium (OCAC; N = 63 347) and from a GWAS meta-analysis of retrospective cohort analyses of epithelial ovarian cancer among BRCA1/2 mutation carriers in the Consortium of Investigators of Modifiers of BRCA1/2 (CIMBA; N = 31 448). Across the 2 consortia, participants were enrolled between 1973 and 2014 and followed up through 2015. OCAC participants came from 14 countries and CIMBA participants came from 25 countries. SNPs were combined into multi-allelic models and mendelian randomization estimates representing lifelong inhibition of targets were generated using inverse-variance weighted random-effects models. Primary exposure was genetically proxied inhibition of HMG-CoA reductase and secondary exposures were genetically proxied inhibition of NPC1L1 and PCSK9 and genetically proxied circulating LDL cholesterol levels. Overall and histotype-specific invasive epithelial ovarian cancer (general population) and epithelial ovarian cancer (BRCA1/2 mutation carriers), measured as ovarian cancer odds (general population) and hazard ratio (BRCA1/2 mutation carriers). The OCAC sample included 22 406 women with invasive epithelial ovarian cancer and 40 941 control individuals and the CIMBA sample included 3887 women with epithelial ovarian cancer and 27 561 control individuals. Median ages for the cohorts ranged from 41.5 to 59.0 years and all participants were of European ancestry. In the primary analysis, genetically proxied HMG-CoA reductase inhibition equivalent to a 1-mmol/L (38.7-mg/dL) reduction in LDL cholesterol was associated with lower odds of epithelial ovarian cancer (odds ratio [OR], 0.60 [95% CI, 0.43-0.83]; P = .002). In BRCA1/2 mutation carriers, genetically proxied HMG-CoA reductase inhibition was associated with lower ovarian cancer risk (hazard ratio, 0.69 [95% CI, 0.51-0.93]; P = .01). In secondary analyses, there were no significant associations of genetically proxied inhibition of NPC1L1 (OR, 0.97 [95% CI, 0.53-1.75]; P = .91), PCSK9 (OR, 0.98 [95% CI, 0.85-1.13]; P = .80), or circulating LDL cholesterol (OR, 0.98 [95% CI, 0.91-1.05]; P = .55) with epithelial ovarian cancer. Genetically proxied inhibition of HMG-CoA reductase was significantly associated with lower odds of epithelial ovarian cancer. However, these findings do not indicate risk reduction from medications that inhibit HMG-CoA reductase; further research is needed to understand whether there is a similar association with such medications.


Isolation and genetic analysis of mutations allowing the degradation of furans and thiophenes by Escherichia coli.
Successive mutations of Escherichia coli yielded a strain that was able to degrade a variety of heterocyclic oxygen- and sulfur-containing ring compounds. In particular, this strain could use both furan-2-carboxylic acid and thiophene-2-carboxylic acid as sole carbon and energy sources. Nitrogen-containing heterocyclic compounds were not degraded. This mutant was isolated by selecting first for oxidation of furan derivatives and then for thiophene degradation. Genetic analysis revealed that mutations in three novel genes, thdA (12 min), thdC (92 min), and thdD (98 min), were required for thiophene degradation. In addition, constitutively at both of the previously characterized fadR and atoC loci was required for efficient thiophene breakdown. The pathway of furan and thiophene degradation remains obscure, but the inability of our mutants to degrade 5-nitro- or 5-bromo-substituted furan derivatives suggests that hydroxylation at position 5 may be involved. Thiophene derivatives were toxic when they were present at concentrations of 0.1% or greater; however, addition of trace amounts of phenylalanine plus tyrosine greatly reduced this effect.


Flagellin gene variation between clinical and environmental isolates of Burkholderia pseudomallei contrasts with the invariance among clinical isolates.
The flagellin gene sequence from a clinical isolate of Burkholderia pseudomallei was used to design oligonucleotide primers for PCR/RFLP analysis of flagellin gene variation among clinical and environmental isolates of B. pseudomallei. Genes from four clinical and six environmental isolates were amplified and compared by RFLP. The clinical isolates were indistinguishable, but variation was detected among some of the environmental isolates. Sequence analysis of flagellin gene amplified products demonstrated high levels of conservation amongst the flagellin genes of clinical isolates (>99% similarity), compared to the variation observed between the clinical isolates and one of the environmental isolates (<90% similarity). Genomic comparisons with pulsed-field gel electrophoresis (PFGE) revealed differences between the relationships inferred by flagellin genotyping and PFGE, suggesting that a combination of molecular methods may be useful for the subtyping of B. pseudomallei strains.


The second green revolution? Production of plant-based biodegradable plastics.
Biodegradable plastics are those that can be completely degraded in landfills, composters or sewage treatment plants by the action of naturally occurring micro-organisms. Truly biodegradable plastics leave no toxic, visible or distinguishable residues following degradation. Their biodegradability contrasts sharply with most petroleum-based plastics, which are essentially indestructible in a biological context. Because of the ubiquitous use of petroleum-based plastics, their persistence in the environment and their fossil-fuel derivation, alternatives to these traditional plastics are being explored. Issues surrounding waste management of traditional and biodegradable polymers are discussed in the context of reducing environmental pressures and carbon footprints. The main thrust of the present review addresses the development of plant-based biodegradable polymers. Plants naturally produce numerous polymers, including rubber, starch, cellulose and storage proteins, all of which have been exploited for biodegradable plastic production. Bacterial bioreactors fed with renewable resources from plants--so-called 'white biotechnology'--have also been successful in producing biodegradable polymers. In addition to these methods of exploiting plant materials for biodegradable polymer production, the present review also addresses the advances in synthesizing novel polymers within transgenic plants, especially those in the polyhydroxyalkanoate class. Although there is a stigma associated with transgenic plants, especially food crops, plant-based biodegradable polymers, produced as value-added co-products, or, from marginal land (non-food), crops such as switchgrass (Panicum virgatum L.), have the potential to become viable alternatives to petroleum-based plastics and an environmentally benign and carbon-neutral source of polymers.


Leveraging Human Genetics to Identify Safety Signals Prior to Drug Marketing Approval and Clinical Use.
When a new drug or biologic product enters the market, its full spectrum of side effects is not yet fully understood, as use in the real world often uncovers nuances not suggested within the relatively narrow confines of preapproval preclinical and trial work. We describe a new, phenome-wide association study (PheWAS)- and evidence-based approach for detection of potential adverse drug effects. We leveraged our established platform, which integrates human genetic data with associated phenotypes in electronic health records from 29,722 patients of European ancestry, to identify gene-phenotype associations that may represent known safety issues. We examined PheWAS data and the published literature for 16 genes, each of which encodes a protein targeted by at least one drug or biologic product. Initial data demonstrated that our novel approach (safety ascertainment using PheWAS [SA-PheWAS]) can replicate published safety information across multiple drug classes, with validated findings for 13 of 16 gene-drug class pairs. By connecting and integrating in vivo and in silico data, SA-PheWAS offers an opportunity to supplement current methods for predicting or confirming safety signals associated with therapeutic agents.


[Features of metabolism in subcellular fractions of the motor cortex during restoration of visual nerve impulses].
Experimental data are obtained on cytochromoxidase (CO), acetylcholinesterase (AchE), MAO and ATPase activities in subfractions of synaptosomes and cellular mitochondria from the rabbit motor cortex following long-term visual deprivation. The first stage is characterized by elevated protein content and activation of enzymatic systems except K+; Na+--ATPase and CO. The tendency towards normalizing of protein content and activity of enzymes, particularly AchE and MAO occurred within 4 weeks. These data show that changes of enzymatic activity in subcellular components of the motor cortex following the sensory deprivation, are reversible but the process of neuronal recompensation in the integrative--starting system is complex and demands a longer time than in visual one.


Functional impact and targetability of PI3KCA, GNAS, and PTEN mutations in a spindle cell rhabdomyosarcoma with MYOD1 L122R mutation.
Spindle cell/sclerosing rhabdomyosarcoma (ssRMS) is a rare subtype of rhabdomyosarcoma, commonly harboring a gain-of-function L122R mutation in the muscle-specific master transcription factor MYOD1. MYOD1-mutated ssRMS is almost invariably fatal, and development of novel therapeutic approaches based on the biology of the disease is urgently needed. MYOD1 L122R affects the DNA-binding domain and is believed to confer MYC-like properties to MYOD1, driving oncogenesis. Moreover, the majority of the MYOD1-mutated ssRMS harbor additional alterations activating the PI3K/AKT pathway. It is postulated that the PI3K/AKT pathway cooperates with MYOD1 L122R. To address this biological entity, we established and characterized a new patient-derived ssRMS cell line OHSU-SARC001, harboring MYOD1 L122R as well as alterations in PTEN, PIK3CA, and GNAS We explored the functional impact of these aberrations on oncogenic signaling with gain-of-function experiments in C2C12 murine muscle lineage cells. These data reveal that PIK3CAI459_T462del, the novel PIK3CA variant discovered in this patient specimen, is a constitutively active kinase, albeit to a lesser extent than PI3KCAE545K, a hotspot oncogenic mutation. Furthermore, we examined the effectiveness of molecularly targeted PI3K/AKT/mTOR and RAS/MAPK inhibitors to block oncogenic signaling and suppress the growth of OHSU-SARC001 cells. Dual PI3K/mTOR (LY3023414, bimiralisib) and AKT inhibitors (ipatasertib, afuresertib) induced dose-dependent reductions in cell growth. However, mTOR-selective inhibitors (everolimus, rapamycin) alone did not exert cytotoxic effects. The MEK1/2 inhibitor trametinib did not impact proliferation even at the highest doses tested. Our data suggest that molecularly targeted strategies may be effective in PI3K/AKT/mTOR-activated ssRMS. Taken together, these data highlight the importance of utilizing patient-derived models to assess molecularly targetable treatments and their potential as future treatment options.


Prognostic Value for Incidental Antihypertensive Therapy With β-Blockers in Metastatic Colorectal Cancer.
Previous studies suggested that the incidental use of β-blockers might influence clinical outcome in solid tumors. We assessed the correlation between the incidental use of β-blockers and clinical outcome in colorectal cancer patients treated with first-line chemotherapy alone or in combination with bevacizumab in metastatic colorectal cancer patients. We collected data from 235 metastatic colorectal cancer patients treated with first-line chemotherapy alone (128 patients) or with bevacizumab (107 patients). Patients were stratified for clinical factors such as β-blockers use, age, sex, and site of metastases, previous adjuvant chemotherapy and ECOG performance status. In the chemotherapy alone group patients receiving β-blockers showed an improved overall survival (median OS 41.3 vs 25.7 months, P = 0.03, HR: 2.26, 95% CI: 1.05-3.24). A significant relationship with improved response rate was also evident for B-blocker users (P = 0.044). On the contrary in the β-blockers users group treated with chemotherapy in combination with bevacizumab we observed a trend toward a worse overall survival although nonstatistically significant (median OS 18.5 vs 23.6 months, HR: 0. 89, 95% CI: 0.38-2.03, P = 0.77). Our analysis confirmed a potential prognostic role for the use of β-blockers in colorectal cancer patients treated with chemotherapy. Our findings also suggest a potential worse outcome for patients on β-blockers receiving bevacizumab. Future prospective studies should include the incidental use of β-blockers as stratification factor for clinical outcome.


The Repertoire of Glycan Alterations and Glycoproteins in Human Cancers.
Cancer as the leading cause of death worldwide has many issues that still need to be addressed. Since the alterations on the glycan compositions or/and structures (i.e., glycosylation, sialylation, and fucosylation) are common features of tumorigenesis, glycomics becomes an emerging field examining the structure and function of glycans. In the past, cancer studies heavily relied on genomics and transcriptomics with relatively little exploration of the glycan alterations and glycoprotein biomarkers among individuals and populations. Since glycosylation of proteins increases their structural complexity by several orders of magnitude, glycome studies resulted in highly dynamic biomarkers that can be evaluated for cancer diagnosis, prognosis, and therapy. Glycome not only integrates our genetic background with past and present environmental factors but also offers a promise of more efficient patient stratification compared with genetic variations. Therefore, studying glycans holds great potential for better diagnostic markers as well as developing more efficient treatment strategies in human cancers. While recent developments in glycomics and associated technologies now offer new possibilities to achieve a high-throughput profiling of glycan diversity, we aim to give an overview of the current status of glycan research and the potential applications of the glycans in the scope of the personalized medicine strategies for cancer.


A photostable AIE fluorogen for lysosome-targetable imaging of living cells.
Disruptive variation in intracellular pH and its fluctuations in lysosomes have a close relationship with the more acidic lysosome lumen of cancer cells (pH 4.5-5.5). Traditional lysosome-targeted probes, such as LysoTracker Green DND-26 (LTG) and LysoTracker Red DND-99 (LTR), can fluoresce when the weak base units in the probes are removed after donating protons under the photoinduced electron-transfer (PET) effect. However they can only be used at low concentration to avoid the aggregation-caused quenching (ACQ) effect and are also easily photobleached under continuous excitation irradiation, displaying low photostability. Herein, a tetraphenylethylene (TPE)-based lysosome-targetable fluorescence probe, TPE-CA, was synthesized, which could selectively monitor the pH change in subcellular organelles and exhibited a strong blue emission under an acidic condition with pH = 4. Using crystallographic, NMR and HRMS analyses, the mechanism regarding the pH dependent fluorescent performance of TPE-CA has been illustrated at the molecular level. In addition, experimental results show that TPE-CA is cell-permeable and biocompatible with HeLa, MCF-7 and HLF cells. The punctate fluorescent spots in the co-staining experiment of TPE-CA with LTG and LTR proves that the blue fluorescence spots of TPE-CA are indeed localized in the most acidic lysosome organelles. In particular, TPE-CA also inherits the aggregation-induced emission (AIE) feature of TPE, showing better photostability under continuous UV illumination compared with the commercial dyes (LTG and LTR). These results show that TPE-CA would be beneficial for understanding the acid environment of lysosomes in related cells and organs with potential biological significance.


TriMix and tumor antigen mRNA electroporated dendritic cell vaccination plus ipilimumab: link between T-cell activation and clinical responses in advanced melanoma.
We previously reported that dendritic cell-based mRNA vaccination plus ipilimumab (TriMixDC-MEL IPI) results in an encouraging rate of tumor responses in patients with pretreated advanced melanoma. Here, we report the TriMixDC-MEL IPI-induced T-cell responses detected in the peripheral blood. Monocyte-derived dendritic cells electroporated with mRNA encoding CD70, CD40 ligand, and constitutively active TLR4 (TriMix) as well as the tumor-associated antigens tyrosinase, gp100, MAGE-A3, or MAGE-C2 were administered together with IPI for four cycles. For 18/39 patients, an additional vaccine was administered before the first IPI administration. We evaluated tumor-associated antigen specific T-cell responses in previously collected peripheral blood mononuclear cells, available from 15 patients. Vaccine-induced enzyme-linked immunospot assay responses detected after in vitro T-cell stimulation were shown in 12/15 patients. Immune responses detected in patients with a complete or partial response were significantly stronger and broader, and exhibited a higher degree of multifunctionality compared with responses in patients with stable or progressive disease. CD8+ T-cell responses from patients with an ongoing clinical response, either elicited by TriMixDC-MEL IPI or on subsequent pembrolizumab treatment, exhibited the highest degree of multifunctionality. TriMixDC-MEL IPI treatment results in robust CD8+ T-cell responses in a meaningful portion of stage III or IV melanoma patients, and obviously in patients with a clinical response. The levels of polyfunctional and multiantigen T-cell responses measured in patients with a complete response, particularly in patients evidently cured after 5+ years of follow-up, may provide a benchmark for the level of immune stimulation needed to achieve a durable clinical remission. NCT01302496.


Skin tests and challenge-based drug allergy diagnosis: a retrospective study of patients with confirmed drug allergy.
Objectives. To describe clinical manifestations and performed diagnostic workup, focusing drug challenge tests (DCT), in patients with drug allergy. Methods. Retrospective study including all patients with skin tests (STs) or DCT-based drug allergy diagnosis, between 01/2014 - 06/2018 in a Portuguese allergy unit. Data were collected from electronic and paper-based clinical records. Results. We had 75 drug allergy diagnoses. Most index reactions were mild and major or equal 1 hour after drug intake. 59 (78%) diagnoses were based on DCTs, all based on multistep protocols with major or equal 3 predicted steps. Only 10% of the DCT were positive during up-dosing; timing and severity of the index reaction predicted DCT interruption during up-dosing. Conclusions. Most drug allergy diagnoses were based on multistep DCT. The identified predictors of DCT interruption during up-dosing can support the development of more personalized DCTs protocols.


Promotion of colonization and virulence by cholera toxin is dependent on neutrophils.
The innate immune response to Vibrio cholerae infection is poorly understood, but this knowledge is critical for the design of safe, effective vaccines. Using an adult mouse intestinal infection model, this study examines the contribution of neutrophils to host immunity, as well as the effect of cholera toxin and other secreted factors on this response. Depletion of neutrophils from mice with anti-Ly6G IA8 monoclonal antibody led to similar survival rates of mice infected with low or moderate doses of toxigenic V. cholerae El Tor O1. At a high dose, neutropenic mice showed increased rates of survival compared to neutrophil-replete animals. Expression of cholera toxin was found to be protective to the neutropenic host, and this phenotype can be replicated by the administration of purified toxin. Neutrophils do not effectively clear colonizing bacteria from the small intestine, nor do they alter induction of early immune-modulating signals. In both neutropenic and neutrophil-replete animals, the local response to infection is characterized by expression of interleukin 6 (IL-6), IL-10, and macrophage inflammatory protein 2 alpha (MIP-2). Overall, these data indicate that the innate immune response to toxigenic V. cholerae infection differs dramatically from the host response to nontoxigenic infection or vaccination, where neutrophils are protective to the host. In the absence of neutrophils, cholera toxin induces immunomodulatory effects that increase host survival. In cholera toxin-producing strains, similar to nontoxigenic infection, accessory toxins are critical to virulence, indicating that cholera toxin and the other secreted toxins modulate the host response by different mechanisms, with both contributing to bacterial persistence and virulence.


Candidatus Amarolinea and Candidatus Microthrix Are Mainly Responsible for Filamentous Bulking in Danish Municipal Wastewater Treatment Plants.
Filamentous bulking is a common serious operational problem leading to deteriorated sludge settling that has long been observed in activated sludge biological wastewater treatment systems. A number of bacterial genera found therein possess filamentous morphology, where some have been shown to be implicated in bulking episodes (e.g., Ca. Microthrix), the impact of many others is still not clear. In this study we performed a survey of 17 Danish municipal wastewater treatment plants (WWTPs) with nutrient removal using 16S rRNA amplicon sequencing over a period of 13 years, where all known filamentous bacteria from 30 genera were analyzed. The filamentous community constituted on average 13 ± 6%, and up to 43% of total read abundance with the same genera common to all plants. Ca. Microthrix and several genera belonging to phylum Chloroflexi were among the most abundant filamentous bacteria. The effect of filamentous bacteria on sludge settling properties was analyzed using measurements of the diluted sludge volume index (DSVI). Strong positive correlations with DSVI were observed only for Ca. Microthrix and Ca. Amarolinea, the latter being a novel, recently characterized genus belonging to the phylum Chloroflexi. The bulking potential of other filamentous bacteria was not significant despite their presence in many plants. Low phylogenetic diversity was observed for both Ca. Microthrix and Ca. Amarolinea, making physiological characterization of individual species and potential development of control strategies more feasible. In this study we show that, despite the high diversity of filamentous phylotypes in Danish WWTPs, only few of them were responsible for severe bulking episodes.