The mouse bioassay for the detection of estrogenic activity in rodent diets: II. Comparative estrogenic activity of purified, certified and standard open and closed formula rodent diets.
A major source of exogenous estrogenic substances, which may affect laboratory animals, comes from the diet. To test the possibility that commercially available rodent diets may significantly influence uterine weights and uterine:body weight (U:BW) ratios, estrogen bioassays were performed using female CD-1 mice weaned at 15 days of age and assigned randomly to a variety of commercial test diets or to a control diet (Purina #5002) containing 0 or 6 ppb added diethylstilbestrol (DES) for comparison. Mice were housed five per cage and given deionized water and feed ad libitum. Uterine:BW ratios from 15 mice per diet were determined after 3, 5 and 7 days of feeding. Mice fed The American Institute of Nutrition purified diet (AIN-76A) or the Purina #5015 natural ingredient breeder diet had significantly (P less than 0.05) increased U:BW ratios at 3, 5 and 7 days post weaning when compared to the control diet without added DES. This increase in U:BW ratios was similar to the U:BW ratios observed in a natural ingredient maintenance diet (Purina #5002), containing 6 ppb of DES. These results show that significant differences exist in the level of substances which can cause increase in uterine weight in some commercial diets. The diet may be important when performing or comparing certain types of studies, especially those relating to estrogenic substances. A standardized diet with minimal estrogenic activity may be desirable for such studies. It is unclear from the present studies what substances might be responsible for the uterine growth promoting activity in the diets examined.


Prevalence estimates of asthma or COPD from a health interview survey and from general practitioner registration: what's the difference?
The aim of this study was to compare prevalence estimates of asthma or chronic obstructive pulmonary disease (COPD) derived from self-report in a health interview survey and from general practitioners' (GPs') medical records, and to explain any differences. the presence of asthma or COPD was measured by self-report in a random sample of 104 general practices in the Netherlands (n = 19 685) participating in the second Dutch National Survey of General Practice (DNSGP-2). This was compared with the presence of GP-diagnosed asthma or COPD in the same population as recorded using the International Classification of Primary Care by their GPs during a 12-month period. Gender, age, health insurance, ethnic background, educational level, tobacco exposure, and other symptoms and conditions were evaluated as explanatory variables using logistic models. The prevalence of self-reported asthma or COPD (9.7%) was almost twice as high as the prevalence based on GP information (5.2%). The medical records of patients who reported having asthma or COPD, without having a diagnosis in their medical records, usually included other respiratory conditions. Patients reporting no asthma or COPD but whose medical records carried a diagnosis of asthma or COPD, were relatively older (P < 0.01) and tended to be exposed to smoking in their home (P < 0.05). Two methods for estimating prevalence of asthma or COPD yielded different results: compared with GP medical records, self-reported prevalence shows an overestimation in people who suffer from other respiratory conditions and an underestimation in elderly persons living in a smoky environment.


Celiac disease in Arabs.
A systematic study of celiac disease in a defined population of Arab children has not been previously reported. We therefore performed a prospective study to determine the incidence and clinical presentation of celiac disease in Jordanian children. A total of 34 (12 boys and 22 girls) cases were diagnosed over a period of 36 months. Nine cases were clustered in three families. The incidence was calculated to be 1:2,800 live births. The mean age at presentation was 4.6 years (range 0.3-13), but it was 8.4 years at the time of diagnosis. Diarrhea was the main initial symptom in 15 (44%) patients. Other signs and symptoms included recurrent abdominal pain in five patients (14%), short stature in four (12%), abdominal distension in three (9%), constipation in three (9%), pallor and recurrent mouth ulcers in two patients each. At the time of diagnosis, 24 (71%) had hypochromic microcytic anemia, and nine (26%) had rickets. The relatively high incidence of celiac disease may be related to the large wheat consumption in this population (135 kg/head/year), and the late onset could be related to the widespread prevalence of prolonged breast-feeding in Jordan (12 months) and late introduction of cereals (6.2 months) in children with late presentation. The delay in diagnosis might be due to the presence of other diseases clinically resembling celiac disease in our community.


Geographic imbalances of physician supply: an international comparison.
This study was undertaken to compare geographic imbalances of physician manpower in developed and developing countries and to evaluate the success of policies designed to alleviate the imbalances. Data were drawn from a mail survey of countries conducted by the World Health Organization, from national statistical summaries, and from published information on national policies and programs. Information was available from 26 developing countries and 15 developed countries. Among developing countries, the degree of urbanization and rural economic development appeared to be the most important determinants of the level of geographic imbalance. Among developed countries, those with very high overall physician/population ratios tended to have the least degree of imbalance, but this effect was not consistent. At the national level in most countries, the effect of policies or programs specifically designed to redistribute physician manpower appeared to be limited.


Calphostin C triggers calcium-dependent apoptosis in human acute lymphoblastic leukemia cells.
Recent studies have demonstrated that the naturally occurring perylenequinone antibiotic calphostin C is a potent inhibitor of protein kinase C and can induce apoptosis in some tumor cell lines by an as yet unknown mechanism. Here we demonstrate that calphostin C induces dose-dependent apoptosis in DT40 chicken lymphoma B-cells, and targeted disruption of lyn, syk, btk, PLCgamma2, or IP3R genes does not prevent or attenuate its cytotoxicity. In our study, calphostin C also induced rapid apoptosis in human acute lymphoblastic leukemia (ALL) cell lines ALL-1 (BCR-ABL+ pre-pre-B ALL), RS4;11 (MLL-AF4+ pro-B ALL), NALM-6 (pre-B ALL), DAUDI (Burkitt's/B-cell ALL), MOLT-3 (T-ALL), and JURKAT (T-ALL), whereas other potent PKC inhibitors did not. In biochemical studies, calphostin C was discovered to induce rapid calcium mobilization from intracellular stores of ALL cell lines, and its cytotoxicity against ALL cell lines was well correlated with the magnitude of this calcium signal. Calphostin C-induced apoptosis was markedly suppressed by BAPTA/AM, a cell-permeable Ca2+ chelator as well as NiCl2, an inhibitor of Ca2+/Mg2+-dependent endonucleases. Inhibition of the Ca2+/calmodulin-dependent phosphatase calcineurin with perfluoreperazine dimadeate (a calmodulin antagonist) or cyclosporin A (a specific inhibitor of calcineurin) also reduced the magnitude of calphostin C-induced apoptosis in ALL cell lines. Calphostin C was capable of inducing calcium mobilization and apoptosis in freshly obtained primary leukemic cells from children with ALL. Taken together, our results provide unprecedented evidence that calphostin C triggers a Ca2+-dependent apoptotic signal in human ALL cells.


Evaluations by staff, residents, and community seniors of patronizing speech in the nursing home: impact of passive, assertive, or humorous responses.
Two studies tested the impact of alternative communication in accommodation strategies. Nursing home staff and residents (and community-residing seniors in Study 2) rated nurse-resident conversational scenarios in which a resident responded passively, directly assertively, or humorously (indirectly assertively) to a patronizing nurse. The nurse then either maintained a patronizing manner or accommodated with a more respectful speech style. Even though all groups devalued the nurse who maintained a patronizing speech style, nursing home residents predictably showed the most acceptance. The directly assertive response by the resident elicited more devaluation of the nonaccommodating nurse than did either passive or humorous responses, but also the least favorable ratings of the resident. Ratings of the humorous response in Study 2 suggested that humor could be a good compromise response style for allowing the receiver of patronizing speech to express opposition to a request, yet still maintain an appearance of competence and politeness.


Effect of a Cationic Surfactant on Droplet Wetting on Superhydrophobic Surfaces.
We experimentally and theoretically examine the influence of a double-chain cationic surfactant, didodecyldimethylammonium bromide (DDAB), on the wetting states and contact angles on superhydrophobic (SH) surfaces made of hydrophobic microcylinders. We use two types of micropatterns of different surface roughness, r, and packing fraction, ϕ, and vary nine dimensionless surfactant concentrations (CS), normalized by the critical micelle concentration (CMC), in the experiments. At low CS, some of the surfactant-laden droplets are in a gas-trapping, Cassie-Baxter (CB) state on the high-roughness microstructures. In contrast, some droplets are in a complete-wetting Wenzel (W) state on the low-roughness microtextures. We found that the contact angle of CB drops can be well predicted using a thermodynamic model considering surfactant adsorption at the liquid-vapor (LV) and solid-liquid (SL) interfaces. At high CS, however, all of the DDAB drops wet in a Wenzel mode. Based on a Gibbsian thermodynamic analysis, we find that for the two types of superhydrophobic surfaces used, the Wenzel state has the lowest thermodynamic energy and thus is more favorable theoretically. The CB state, however, is metastable at low CS due to a thermodynamic energy barrier. The metastable CB wetting state becomes more stable on the SH microtextures with greater ϕ and r, in agreement with our experimental observations. Finally, we generalize this surface-energy analysis to provide useful designs of surface parameters for a DDAB-laden surfactant droplet on the SH surface with a stable and robust CB state.


Regional Variations in Documentation of Sexual Trauma Concepts in Electronic Medical Records in the United States Veterans Health Administration.
Background: Experiences of sexual trauma are associated with adverse patient and health system outcomes, but are not systematically documented in electronic health records (EHR). Objective: To describe variations in how sexual trauma is documented in the Veterans Health Adminstration's EHR. Methods: Sexual trauma concepts were extracted from from 362,559 clinical notes using a natural language processing pipeline. Results: We observed variations in the presence of sexual trauma in notes across five United States regions: Pacific, Continental, Midwest, North Atlantic, Southeast. We also observed variations in the types of notes used to document sexual trauma (e.g., mental health, primary care) and sources of sexual trauma (e.g., adult, childhood, military) mentioned in the EHR. Our findings illustrate potential differences in cultural norms related to patient disclosure of sensitive information, and provider documentation. Standardized protocol for eliciting and documenting sexual trauma histories are needed to ensure Veteran access to high quality, trauma-informed care.


Experimental autoimmune dacryoadenitis. I. Lacrimal gland disease in the rat.
Experimental autoimmune dacryoadenitis was produced in Lewis rats by immunization with a single intradermal administration of a 3M KCl extract of exorbital lacrimal gland in CFA, when enhanced by simultaneous i.v. injection of killed Bordetella pertussis. No significant lacrimal lesions were observed in control animals immunized with the extracts of Harderian or salivary glands. Gel filtration of the 3M KCl extract on Sephacryl S-300 column yielded three protein fractions. Only fraction III (MW = 10-55K) induced marked dacryoadenitis following a single injection of 2.0 mg protein in CFA plus pertussis. The infiltrates in the exorbital lacrimal lesions were first apparent around the ducts and associated vasculature. From this area, the infiltrates appeared to spread to the acini drained by these ducts, ultimately involving as much as 30-50% of the gland. The affected glands most commonly showed a diffuse nongranulomatous infiltrate of small lymphocytes, macrophages, and plasma cells; this was focal in nature, involving acinar atrophy and breakdown, and replaced the normal architecture in extreme cases. The Harderian and salivary glands were uninvolved in these animals, suggesting a restricted specificity of this response. Lewis rats immunized with exorbital lacrimal gland fractions I or II in CFA plus pertussis showed only minimal lesions, similar to controls receiving CFA and pertussis without antigen. These findings suggest that an autoantigen exists in the lacrimal gland of the rat that is capable of inducing a specific lymphoproliferative dacryoadenitis.


Understanding Local-Field Correction Factors in the Framework of the Onsager-Böttcher Model.
The determination of the appropriate local-field factor for quantifying the response of a molecule to an external electric field is of major importance in optical spectroscopy. Although numerous studies have dealt with the evolution of the optical properties of emitters as a function of their environment, the choice of the model used to quantify local fields is still ambiguous, and sometimes even arbitrary. In this paper, we review the Onsager-Böttcher model, which introduces the polarizability of the probe molecule as the determinant parameter for the local field factor, and we establish a simple conceptual framework encompassing all commonly used models. Finally, a discussion of published experimental research illustrates the potential of the measurement of local electric fields in dense dielectric media, as well as the subtleties involved in their interpretation.


Simultaneous measurement of 19 components in serum-containing animal cell culture media by fourier transform near-infrared spectroscopy.
Animal cell cultures generate maximal amounts of desired products when maintained in a controlled environment with low and constant concentrations of nutrients and wastes. Traditionally this has involved slow addition of glucose and glutamine; however, recent studies have indicated that a number of low concentration amino acids are required to prevent initiation of apoptosis. Therefore, optimal control of animal cell cultures will likely require measurement of a large number of chemical components. We present here the evaluation of a near-infrared spectroscopic (NIRS) monitoring scheme to quantify 19 cellular nutrients and wastes in culture medium with and without serum. The components include glucose, lactate, ammonia, pyruvate, glutamine, and 14 other amino acids. Spectroscopic calibrations were generated for a synthetic version of a standard culture medium (DMEM) in which the concentrations of 17 DMEM components and ammonia and lactate were varied in a random fashion. This randomization provides a stringent evaluation of the measurement scheme. Reasonably accurate measurements of these 19 components could be accomplished in the absence or presence of 10% horse serum by volume with percent errors ranging from 3% to 37%. Analytes with concentrations as low as 0.3 mM could be reliably quantified. The presence of serum, when properly included in the calibration, has little effect on measurement error. These results provide an important step toward application of NIRS for monitoring the large number of varying components of animal cell cultivations.


Prevalence and associated factors of job-related depression in laboratory technicians in Hospital Universiti Sains Malaysia (HUSM) and Ministry of Health Malaysia (MOH) Hospitals in Kelantan.
Karasek's job strain model postulates that workers chronically exposed to adverse psychosocial work environment (high strain job)--high psychological job demands and low job control, including poor social support, increased physical demand and hazardous work condition--will eventually develop both physical illnesses such as cardiovascular diseases and psychological disorders such as depression. In order to determine the prevalence and associated factors of job-related depression, a cross-sectional study was conducted among laboratory technicians in Hospital Universiti Sains Malaysia (HUSM) and Kementerian Kesihatan Malaysia (KKM) Hospitals in Kelantan between September 2001 and February 2002. One hundred and two laboratory technicians in HUSM and 79 laboratory technicians in 7 KKM Hospitals were selected; 84/102 (82.4%) in HUSM and 71/79 (89.9%) in KKM Hospitals responded. Data was collected using self-administered questionnaires of the validated Malay version of the Job Content Questionnaire (JCQ) originally developed by Robert Karasek. Results indicated that the prevalence of high job strain in laboratory technicians in HUSM and KKM hospitals was 33.3% (28/84) and 26.8% (19/71), respectively (p>0.05). Significantly higher proportion (59.5%) (50/84) of laboratory technicians in HUSM compared to those in KKM Hospitals (39.4%) (28/71) (p = 0.016) experienced job-related depression. Significant associated factors of job-related depression were low social support (HUSM: adjusted OR 4.7, 95% CI 1.2-18.8; KKM: adjusted OR 14.8, 95%CI 2.4-89.3), high psychological demand (HUSM: adjusted OR 3.0, 95%CI 1.0-8.8), and low decision authority (KKM: adjusted OR 9.7, 95%CI 1.0-91.1). We conclude that strengthening the social support network (supervisors' and coworkers' support), reducing psychological job demand, and increasing decision-making authority of laboratory technicians may go a long way towards reducing job-related depression.


Adipokines and their Relation to Endothelial Dysfunction in Patients with Chronic Kidney Disease.
Chronic Kidney Disease (CKD) patients are at high risk of cardiovascular diseases (CVDs). Reduced nitric oxide (NO) bioavailability is a key element in connecting kidney disease to endothelial dysfunction (ED) and cardiovascular (CV) complications. Further, inflammation is implicated in ED in CKD. Besides these, adipose tissue factors were thought to have a role in inflammation and ED in CKD. It is proposed to evaluate the concentration changes of adipokines, inflammatory and ED markers in CKD patients compared to healthy controls. Further, to assess the associations between adipokines, inflammation and ED in CKD patients. A total of 120 CKD patients were included and classified into 3 groups based on Glomerular filtration rate (GFR). Group I (n=40) patients had a GFR between 60-119 ml/min/1.73m(2) (stage I, II), group II (n=40) had 15-59 ml/min/1.73m(2) (stage III, IV) and group III (n=40) had <15 ml/min/1.73m(2) (stage V). Forty healthy subjects served as controls. Adiponectin, Leptin, Interleukin-10 (IL-10), Interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α) were estimated by ELISA. High sensitivity C-reactive protein (hsCRP) was estimated by immunoturbidimetry and NO by Griess method. Mann-Whitney U test was used to compare the difference in variables between controls and CKD patients. One-way ANOVA Kruskalwallis test was used for comparison of variables between groups in CKD patients. Spearman's rank correlation was used to explore the associations between variables. Simple univariate linear regression analysis was used to predict the value of variable from another variable. A significant increase in leptin, IL-6, TNF-α, IL-6/IL-10 ratio, hsCRP and decrease in adiponectin, IL-10, NO was observed in CKD patients compared to controls (p<0.05). In CKD patients, adiponectin, leptin, IL-6, IL-6/IL-10 ratio, TNF-α were significantly increased and IL-10 levels were decreased from group I to group III (p<0.05). In group III CKD patients IL-6 showed a significant negative correlation with NO (r=-0.557; p=0.005). In linear regression analysis also, IL-6 showed a significant negative association with NO (B±SE=-0.038±0.11; p=0.002) in CKD patients. The present study demonstrates that adipokine levels are altered from initial to final stages of CKD due to renal dysfunction which in association with an exaggerated inflammation may contribute to the ED and CV events.


IGF2 but not H19 shows loss of imprinting in human glioma.
Genomic imprinting is a gamete-specific modification resulting in the allele-specific expression of genes in somatic cells. A loss of imprinting (LOI) has been found in many embryonal and adult tumors, suggesting that it plays a role in tumor development. The incidence of LOI, however, does not seem to be ubiquitous among tumors because neuroblastoma and colorectal cancer revealed no LOI. We examined the involvement of LOI of IGF2 and H19 genes in human gliomas. The two genes were imprinted in normal brain subcortex tissues. In glioma, 8 of 14 informative cases (57%) revealed LOI in IGF2. The frequency did not depend on the tumor grade. For H19, in contrast, all 13 informative cases maintained imprinting. These results suggest that LOI of IGF2 but not H19 plays a role in the development of human glioma.


Aspects of energetic substrate metabolism of in vitro and in vivo bovine embryos.
Although the metabolism of early bovine embryos has not been fully elucidated, several publications have addressed this important issue to improve culture conditions for cattle reproductive biotechnologies, with the ultimate goal of producing in vitro embryos similar in quality to those developing in vivo. Here, we review general aspects of bovine embryo metabolism in vitro and in vivo, and discuss the use of metabolic analysis of embryos produced in vitro to assess viability and predict a viable pregnancy after transference to the female tract.


Impact of Early Postnatal Nutrition on the NMR Urinary Metabolic Profile of Infant.
NMR-based metabolomics was used to compare the metabolic urinary profiles of exclusively breast-fed term infants (n = 11) with those of a double-blinded controlled trial with 49 formula-fed term newborns randomized to receive either an infant formula enriched by functional ingredients (n = 24) or a standard formula (n = 25). Anthropometric measurements and urine samples were taken at enrollment (within the first month of life), at around 60 days of life, and at the end of study period (average age of 130 days). The metabolic profiles were examined in relation to time and diet strategy. A common age-dependent modification of the urine metabolome was observed for the three types of nutrition, mainly characterized by similar temporal trends of choline, betaine, myoinositol, taurine, and citrate. Contrariwise, differences in the metabolic profiles were identified according to the type of diet (human versus formula milk), while no significant difference was observed between the two formulas. These modifications are discussed mainly in terms of the different milk compositions. Despite the low number of enrolled infants (n = 60), these findings pointed out the potential of the metabolomics approach for neonatal nutritional science, in particular to provide important contributions to the optimization of formula milk.


The effect of a single dose of multivitamin and mineral combinations with and without guaraná on functional brain activity during a continuous performance task.
Relatively few studies have explored the possibility of acute cognitive effects of multivitamin ingestion. This report explores the acute brain electrophysiological changes associated with multivitamin and mineral supplementation, with and without guaraná, using the steady-state visually evoked potential (SSVEP). Based on the known SSVEP correlates of A-X continuous performance task (CPT) performance, and sensitivity to acute psychopharmacological manipulations, the A-X CPT was adopted as a task paradigm to explore treatment-related neurophysiological changes in attentional processing. Twenty healthy non-smoking adults aged 21-39 years (mean age = 28.35 years, SD = 5.52) took part in this double-blind, placebo-controlled, randomized, balanced crossover design study. The study demonstrated both transient and tonic changes in the SSVEP response during completion of the A-X CPT following multivitamin and mineral treatment both with and without guaraná. Transient changes in SSVEP response in prefrontal regions were observed after a single dose of a multivitamin and mineral preparation indicative of enhanced activity within brain regions engaged by the attentional demands of the task. This pattern of change in frontal regions was correlated with improved behavioural performance after treatment with the multivitamin and mineral combination. Where tonic shifts in SSVEP response were investigated, multivitamin and mineral treatment was associated with a pattern of increased inhibition across posterior regions, with enhanced excitatory processing in prefrontal regions. In contrast, multivitamin and mineral treatment with additional guaraná showed a tonic shift towards greater excitatory processes after a single treatment, consistent with the caffeine content of this treatment. While preliminary in nature, these findings suggest a single multivitamin/mineral dose is sufficient to impact on functional brain activity in task-related brain regions.


Applications of next generation sequencing in molecular ecology of non-model organisms.
As most biologists are probably aware, technological advances in molecular biology during the last few years have opened up possibilities to rapidly generate large-scale sequencing data from non-model organisms at a reasonable cost. In an era when virtually any study organism can 'go genomic', it is worthwhile to review how this may impact molecular ecology. The first studies to put the next generation sequencing (NGS) to the test in ecologically well-characterized species without previous genome information were published in 2007 and the beginning of 2008. Since then several studies have followed in their footsteps, and a large number are undoubtedly under way. This review focuses on how NGS has been, and can be, applied to ecological, population genetic and conservation genetic studies of non-model species, in which there is no (or very limited) genomic resources. Our aim is to draw attention to the various possibilities that are opening up using the new technologies, but we also highlight some of the pitfalls and drawbacks with these methods. We will try to provide a snapshot of the current state of the art for this rapidly advancing and expanding field of research and give some likely directions for future developments.


Linear parameter-varying model for a refuellable zinc-air battery.
Due to the increasing trend of using renewable energy, the development of an energy storage system (ESS) attracts great research interest. A zinc-air battery (ZAB) is a promising ESS due to its high capacity, low cost and high potential to support circular economy principles. However, despite ZABs' technological advancements, a generic dynamic model for a ZAB, which is a key component for effective battery management and monitoring, is still lacking. ZABs show nonlinear behaviour where the steady-state gain is strongly dependent on operating conditions. The present study aims to develop a dynamic model, being capable of predicting the nonlinear dynamic behaviour of a refuellable ZAB, using a linear parameter-varying (LPV) technique. The LPV model is constructed from a family of linear time-invariant models, where the discharge current level is used as a scheduling parameter. The developed LPV model is benchmarked against linear and nonlinear model counterparts. Herein, the LPV model performs remarkably well in capturing the nonlinear behaviour of a ZAB. It significantly outperforms the linear model. Overall, the LPV approach provides a systematic way to construct a robust dynamic model which well represents the nonlinear behaviour of a ZAB.


Carbamazepine versus valproate monotherapy for epilepsy.
Carbamazepine and valproate are drugs of first choice for epilepsy. Despite the lack of hard evidence from individual randomized controlled trials, there is strong clinical belief that valproate is the drug of choice for generalized epilepsies and carbamazepine for partial epilepsies. To overview the best evidence comparing carbamazepine and valproate monotherapy Our search strategy included: (a) MEDLINE 1966-99, (b) The Cochrane Library 1999 issue 4, (c) The trial register of the Cochrane Epilepsy Group (d) the pharmaceutical industry. Randomized controlled trials comparing carbamazepine and valproate monotherapy for epilepsy. This was an individual patient data review. Outcome measures were time to withdrawal of allocated treatment, time to 12 month remission, and time to first seizure post randomization. Data were analysed using the stratified Logrank test with results expressed as hazard ratios (HR) (95% CI), where HR>1 indicates an event is more likely on valproate. A test for an interaction between treatment and epilepsy type (partial versus generalized) was also undertaken. Results Data were available for 1265 patients from five trials, representing 85% of the patients recruited into the eight trials that met our inclusion criteria. The main overall results (HR 95% CI) were: Time to treatment withdrawal 0.97 (0.79-1.18), 12 month remission 0.87 (0.74-1.02), first seizure 1.09 (0.96-1.25) suggesting no overall difference for these outcomes. The test for an interaction between treatment and epilepsy type was non significant for time to treatment withdrawal and 12 month remission, but significant for time to first seizure. The age distribution of adults classified as having a generalized epilepsy indicate that significant numbers of patients may have had their epilepsy misclassified. We have found some evidence to support the policy of using carbamazepine as the first treatment of choice in partial epilepsies, but no evidence to support the choice of valproate in generalized epilepsies, but confidence intervals are too wide to confirm equivalence. Misclassification of patients may have confounded our results, and has important implications for the design and conduct of future trials.


Chronic ectopic pregnancy--two cases of acute rupture despite negative beta hCG.
We present the cases of two women with chronic ectopic pregnancies who presented with acute tubal rupture and hemoperitoneum despite negative beta-human chorionic gonadotropin (beta hCG) pregnancy tests. The appropriate screening use of beta hCG assays to ascertain pregnancy status, the mechanisms by which ectopic pregnancy may be seen with negative beta hCG tests, and the limitations of various assays are discussed. One patient, not initially believed to be pregnant, underwent computed tomography (CT) scan. Experience with use of CT scan in ectopic pregnancy diagnosis is limited; our case illustrates some of the possible CT scan findings. These cases illustrate the potential for ectopic pregnancy to rupture with low, if not undetectable beta hCG hormone levels, and consequently why it is not recommended to rely on quantitative beta hCG levels to guide the decision to proceed with ultrasound imaging.


A statistical model for the study of self-fertilization in plants.
A model is proposed for the study of plant breeding where the self-fertilization rate is of importance. The model can be used to provide convenient maximum likelihood estimation of the self-fertilization rate and allelic frequencies in a parental population and its pollen pool. The self-fertilization rate can also be allowed to vary with genotype or other characteristics of a plant. A simple example illustrates the methodology.


Sclerostin and Dickkopf-1 in renal osteodystrophy.
The serum proteins sclerostin and Dickkopf-1 (Dkk-1) are soluble inhibitors of canonical wnt signaling and were recently identified as components of parathyroid hormone (PTH) signal transduction. This study investigated the associations between sclerostin and Dkk-1 with histomorphometric parameters of bone turnover, mineralization, and volume in stage 5 chronic kidney disease patients on dialysis (CKD-5D). In a cross-sectional study, 60 CKD-5D patients underwent bone biopsies followed by histomorphometry. Levels of sclerostin, Dkk-1, and intact PTH (iPTH) were determined in blood. Serum levels of sclerostin and iPTH correlated negatively. In unadjusted analyses, sclerostin correlated negatively with histomorphometric parameters of turnover, osteoblastic number, and function. In adjusted analyses, sclerostin remained a strong predictor of parameters of bone turnover and osteoblast number. An observed correlation between sclerostin and cancellous bone volume was lost in regression analyses. Sclerostin was superior to iPTH for the positive prediction of high bone turnover and number of osteoblasts. In contrast, iPTH was superior to sclerostin for the negative prediction for high bone turnover and had similar predictive values than sclerostin for the number of osteoblasts. Serum levels of Dkk-1 did not correlate with iPTH or with any histomorphometric parameter. Our data describe a promising role for serum measurements of sclerostin in addition to iPTH in the diagnosis of high bone turnover in CKD-5D patients, whereas measurements of Dkk-1 do not seem to be useful for this purpose.


Unilateral spatial neglect associated with chronic subdural haematoma: a case report.
A 69-year-old right-handed man who exhibited unilateral spatial neglect in association with a chronic subdural haematoma, presented with mild left arm and leg weakness first noted 4 weeks prior to admission. Neurologic examination on admission revealed a mild left hemiparesis, including the face. Neuropsychologic examination revealed left unilateral spatial neglect, but no language disturbance. Minimal support was necessary to maintain activities of daily living. Computed tomography revealed a large right temporoparietal, extraaxial hypodense fluid collection containing scattered hypodense foci. The haematoma was evacuated via a right parietal burr hole. Following surgery, the patient dramatically improved neurologically and neuropsychologically, as well as in independent performance of daily activities. It is suggested that the improvement in ADL provides a behavioural correlate of improvement in the latter, represented a behavioural correlate of improved cerebral function, and that either direct compression by the chronic subdural haematoma or an interhemispheric pressure difference had caused unilateral spatial neglect. Such neglect is an unusual consequence of chronic subdural haematoma.


Case Report: Molecular Characterization of Bloodstream Infections due to Carbapenem-Resistant Acinetobacter baumannii: A Pediatric Case Series.
Acinetobacter baumannii poses a significant threat to public health due to the high rate of multidrug-resistant strains. However, information on the molecular characterization of carbapenem-resistant Acinetobacter baumannii (CRAB) bloodstream infections in children is scarce. This study aimed to describe the molecular characterization of carbapenem-resistant A. baumannii infections in children from a hospital in Mexico. A retrospective study was conducted during the period 2017-2022. Clinical and demographic data were collected from the clinical records. Mass spectrometry was used for the identification of the strains. To confirm A. baumannii strains, a polymerase chain reaction (PCR) method was applied using a gyrB sequence. The carbapenemase-encoding resistance genes were detected by PCR. Six cases of CRAB were documented, including five in neonates. The median intensive care unit stay was 20 days, and all cases had an invasive medical device. Half of the patients had at least one medical condition. A high prevalence of coresistance was observed in most of the antibiotic groups. Three of the six strains coharbored carbapenemase genes: blaOXA-51, blaOXA-24, and blaIMP. Mortality was reported in two neonate patients. The present study shows a high rate of coharboring blaOXA-51, blaOXA-24, and blaIMP-1, which has a direct impact on therapeutic decisions. Implementation of antimicrobial stewardship programs is urgent to stop the spread of this microorganism.


Particle Engineering via Supramolecular Assembly of Macroscopic Hydrophobic Building Blocks.
Tailoring the hydrophobicity of supramolecular assembly building blocks enables the fabrication of well-defined functional materials. However, the selection of building blocks used in the assembly of metal-phenolic networks (MPNs), an emerging supramolecular assembly platform for particle engineering, has been essentially limited to hydrophilic molecules. Herein, we synthesized and applied biscatechol-functionalized hydrophobic polymers (poly(methyl acrylate) (PMA) and poly(butyl acrylate) (PBA)) as building blocks to engineer MPN particle systems (particles and capsules). Our method allowed control over the shell thickness (e.g., between 10 and 21 nm), stiffness (e.g., from 10 to 126 mN m-1), and permeability (e.g., 28-72% capsules were permeable to 500 kDa fluorescein isothiocyanate-dextran) of the MPN capsules by selecting hydrophobic polymer building blocks (PMA or PBA) and controlling the polymer concentration in the MPN assembly solution (0.25-2.0 mM) without additional/engineered assembly processes. Molecular dynamics simulations provided insights in the structural states of the hydrophobic building blocks during assembly and mechanism of film formation. Furthermore, the hydrophobic MPNs facilitated the preparation of fluorescent-labeled and bioactive capsules through postfunctionalization and particle-cell association engineering by controlling the hydrophobicity of the building blocks. Engineering MPN particle systems via building block hydrophobicity is expected to expand their use.


Genetic demonstration of bidirectionality in the high affinity purine base transporter of mutant mouse S49 cells.
A mutant cell line was selected from wild type S49 lymphoblasts that expressed a novel high affinity purine base transport system not found in parental cells or any other mammalian cell line (Aronow, B., Toll, D., Patrick, J., Hollingsworth, P., McCartan, K., and Ullman, B. (1986) Mol. Cell. Biol. 6, 2957-2962). In order to determine whether this nucleobase transport system was bidirectional, mutant cell lines possessing this high affinity base transport capability were derived from a nucleoside transport-deficient derivative of an adenylosuccinate synthetase-deficient S49 cell line. The resulting progeny excreted significantly greater amounts of purine into the cell culture medium than parental cells. This purine was identified as hypoxanthine. These results demonstrate genetically that the high affinity purine base transport system can mediate both the influx and efflux of hypoxanthine.


Risk and Resilience Factors for Psychobehavioral Symptom Trajectories Among Child Welfare-Involved Youth.
The primary aim of the study was to investigate the development of psychobehavioral symptoms (internalizing, externalizing, and posttraumatic stress [PTS]) of child welfare-involved adolescents over a 3-year period and to identify predictors of the trajectories. Using three waves of data from the National Survey of Child and Adolescent Well-Being (NSCAW-II), latent growth curve modeling was conducted on a sample of 350 adolescents who were between 11 and 13 years of age at baseline. Findings indicated that adolescents' internalizing symptoms decreased over time, whereas externalizing symptoms remained stable over time. PTS symptoms decreased over time in girls, but not boys. Physical abuse predicted higher levels of externalizing behaviors and sexual abuse predicted a slower decrease in PTS symptoms. Greater child prosocial skills and higher quality of caregiver-child relationship were associated with lower initial levels of externalizing symptoms. In addition, a higher quality caregiver-child relationship and greater satisfaction with peer relationships were associated with lower initial levels of internalizing and PTS symptoms. Findings suggest that intervention strategies should consider gender, maltreatment type, and interpersonal and social skills to effectively prevent the development of psychobehavioral symptoms among child welfare-involved youth.


Genetic polymorphisms in folate and alcohol metabolism and breast cancer risk: a case-control study in Thai women.
Dietary folate as well as polymorphic variants in one-carbon metabolism genes may modulate risk of breast cancer through aberrant DNA methylation and altered nucleotide synthesis and repair. Alcohol is well recognized as a risk factor for breast cancer, and interactions with one-carbon metabolism has also been suggested. The purpose of this study is to test the hypothesis that genetic polymorphisms in the folate and alcohol metabolic pathway are associated with breast cancer risk. Twenty-seven single nucleotide polymorphisms (SNPs) in the MTR, MTRR, MTHFR, TYMS, ADH1C, ALDH2, GSTP1, NAT1, NAT2, CYP2E1 DRD2, DRD3, and SLC6A4 were genotyped. Five hundred and seventy patients with histopathogically confirmed breast cancer and 497 controls were included in the present study. Association of genotypes with breast cancer risk was evaluated using multivariate logistic regression to estimate odds ratios (OR) and their 95% confidence intervals (95% CI). Increased risk was observed for homozygotes at the MTR SNPs (rs1770449 and rs1050993) with the OR = 2.21 (95% CI 1.18-4.16) and OR = 2.24 (95% CI 1.19-4.22), respectively. A stratified analysis by menopausal status indicated the association between the NAT2 SNP (rs1799930) and breast cancer was mainly evident in premenopausal women (OR 2.70, 95% CI 1.20-6.07), while the MTRR SNP (rs162049) was significant in postmenopausal women (OR 1.61, 95% CI 1.07-2.44). Furthermore, SNPs of the genes that contribute to alcohol behavior, DRD3 (rs167770), DRD2 (rs10891556), and SLC6A4 (rs140701), were also associated with an increased risk of breast cancer. No gene-gene or gene-environment interactions were observed in this study. Our results suggest that genetic polymorphisms in folate and alcohol metabolic pathway influence the risk of breast cancer in Thai population.


Fluorescence of reduced nicotinamides using one- and two-photon excitation.
We examined the steady-state and time-resolved emission of NADH and NAMH resulting from one-photon and two-photon excitation. Similar emission spectra were observed for both modes of excitation. The fundamental anisotropy of NADH is near 0.54 for two-photon excitation from 690 to 740 nm, which is 46% higher than the value of 0.37 observed for one-photon excitation. This observation of a higher anisotropy with two-photon excitation was consistent with INDO/SDCI calculations of the one- and two-photon transitions. Minor differences in the multi-exponential decays of NADH were observed for one- and two-photon excitation, but presently available resolution does not allow us to conclude the decays are distinct. NADH-LADH-IBA complex formation led to an order of magnitude larger of the average lifetimes of NADH fluorescence resulting from one- and two-photon excitation. Fluorescence intensity and fluorescence anisotropy decays of NADH was double-exponential for both modes of excitation and show that the observed heterogeneity of the fluorescence decay kinetics of reduced nicotinamides arises from the inherent photoprocess of the dihydronicotinamide chromophore and not due to any intramolecular interactions with adenine part of NADH. Such interactions are responsible for the depolarization of NADH fluorescence observed for excitation wavelength below 300 nm for OPE and 600 nm for TPE, respectively. NADH displays a low cross-section for two-photon excitation which suggests that fluorescence from NADH will be moderately difficult to observe with two-photon fluorescence microscopy, and may not interfere with observations of TPIF of other extrinsic probes used to label cells.


Propofol inhibits SIRT2 deacetylase through a conformation-specific, allosteric site.
meta-Azi-propofol (AziPm) is a photoactive analog of the general anesthetic propofol. We photolabeled a myelin-enriched fraction from rat brain with [(3)H]AziPm and identified the sirtuin deacetylase SIRT2 as a target of the anesthetic. AziPm photolabeled three SIRT2 residues (Tyr(139), Phe(190), and Met(206)) that are located in a single allosteric protein site, and propofol inhibited [(3)H]AziPm photolabeling of this site in myelin SIRT2. Structural modeling and in vitro experiments with recombinant human SIRT2 determined that propofol and [(3)H]AziPm only bind specifically and competitively to the enzyme when co-equilibrated with other substrates, which suggests that the anesthetic site is either created or stabilized in enzymatic conformations that are induced by substrate binding. In contrast to SIRT2, specific binding of [(3)H]AziPm or propofol to recombinant human SIRT1 was not observed. Residues that line the propofol binding site on SIRT2 contact the sirtuin co-substrate NAD(+) during enzymatic catalysis, and assays that measured SIRT2 deacetylation of acetylated α-tubulin revealed that propofol inhibits enzymatic function. We conclude that propofol inhibits the mammalian deacetylase SIRT2 through a conformation-specific, allosteric protein site that is unique from the previously described binding sites of other inhibitors. This suggests that propofol might influence cellular events that are regulated by protein acetylation state.


Health Related Quality of Life of Patients with Bladder Cancer in the RAZOR Trial: A Multi-Institutional Randomized Trial Comparing Robot versus Open Radical Cystectomy.
We evaluated health related quality of life following robotic and open radical cystectomy as a treatment for bladder cancer. Using the Randomized Open versus Robotic Cystectomy (RAZOR) trial population we assessed health related quality of life by using the Functional Assessment of Cancer Therapy (FACT)-Vanderbilt Cystectomy Index and the Short Form 8 Health Survey (SF-8) at baseline, 3 and 6 months postoperatively. The primary objective was to assess the impact of surgical approach on health related quality of life. As an exploratory analysis we assessed the impact of urinary diversion type on health related quality of life. Analyses were performed in subsets of the per-protocol population of 302 patients. There was no statistically significant difference between the mean scores by surgical approach at any time point for any FACT-Vanderbilt Cystectomy Index subscale or composite score (p >0.05). The emotional well-being score increased over time in both surgical arms. Patients in the open arm showed significantly better SF-8 sores in the physical and mental summary scores at 6 months compared to baseline (p <0.05). Continent diversion (versus noncontinent) was associated with worse FACT-bladder-cystectomy score at 3 (p <0.01) but not at 6 months, and the SF-8 physical component was better in continent-diversion patients at 6 months (p=0.019). Our data suggests lack of significant differences in the health related quality of life in robotic and open cystectomies. As robotic procedures become more widespread it is important to discuss this finding during counseling.


Pelvic organ prolapse surgery: changes in perioperative management improving hospital pathway.
There is growing interest in improving operating theatre efficacy and shortening stay in hospital. We made a retrospective patient record review looking at changes in the perioperative management of pelvic organ prolapse surgery during the period 2012 to 2014 in four hospitals in the Gothenburg area in Sweden. We studied 440 pelvic organ prolapse surgery records; procedures performed during 2012, 2013 and 2014. Anterior colporrhaphy, posterior colporrhaphy and combined procedures were included. Patient characteristics, surgery, anesthetic technique, theatre time, surgical time, time in recovery room, and hospital stay was analyzed. We found major changes in perioperative management at the same time as local anesthesia and sedation (LAS) increased. Operating room efficiency was increased; increased number of patients bypassed the recovery room and hospital stay was overall shortened, an increasing number of patients were discharged day of surgery. Patients discharge day of surgery increased from 24% to 72% between 2012 and 2014. Surgery under LAS increased from 87% to 96% of the procedures in the private hospital and from 18% to 83% in the university hospital between 2012 and 2014. There is a clear change in perioperative care of patients undergoing pelvic organ prolapse surgery to a more enhanced perioperative management. Routine and practices differs however between hospitals. The use of local anesthesia and sedation improve theatre turn around and shortens hospital stay.


Comparison of outcomes between surgical treatment of colorectal cancer metastatic to the liver and the peritoneum: Review of the literature.
Liver and peritoneum are two of the most common sites of colorectal metastases. We searched for articles comparing outcomes of surgical management for metastatic colorectal cancer to the liver and peritoneum. Cytoreductive surgery/heated intraperitoneal chemotherapy has a similar safety profile and survival outcomes as hepatectomy for colorectal metastases after stratifying by resection status and should be incorporated earlier in the management algorithm for colorectal cancer patients with peritoneal metastases METHODS: We performed a wide search on PubMed, EMBASE, and Google Scholar for articles comparing outcomes of surgical management for metastatic colorectal cancer to the liver and peritoneum. We focused on studies comparing their perioperative clinical outcomes as well as their oncological outcomes. The following words were included in the search: comparison, outcomes, metastasectomy, colorectal cancer, liver, peritoneal surface disease, hepatectomy, and cytoreduction. One hundred and twenty studies were evaluated. Six of these studies met the criteria for this review.


Opportunities to enhance ward audit: a multi-site qualitative study.
Hospitals in many countries are encouraged to develop audits to assess and improve the quality of care. Ward audit is a specific form of audit and feedback that is commonly used but little studied. The aim of this study is to describe the content and application of hospital ward audit in order to identify potential enhancements to such audits. Multiple qualitative methods were used to study a diversity sample of four English National Health Service organisations over a 16-month period. We undertook semi-structured interviews (n = 32), documentary analysis (n = 44) and 25 h of observations of healthcare workers involved in the design and implementation of ward audit. Data were analysed using framework analysis. Findings were presented iteratively to stakeholders who used them to develop a description of the content and delivery of ward audit. Ward audit consisted of seven stages: impetus; method; preparation of staff; assessing practice; analysis; feedback; and decide on action to improve. Two key stages were the monthly assessment of practice using case note data extraction, and the resulting feedback to clinical staff, ward managers, matrons and directors of nursing. At three organisations, the case note data were extracted by staff and there was evidence that this resulted in misrepresentation of the clinical performance audited. The misrepresentation appeared to be associated with the anticipation of punitive feedback from directors of nursing and matrons, as well as time pressures and a lack clarity about the method of audit data collection. Punitive feedback was reported to occur if no data were collected, if data demonstrated poor performance or if performance did not improve. Organisations invest considerable clinical resources in ward audit, but such audits may have unintended, potentially negative, consequences due to the impacts from punitive feedback. We discuss potential enhancements to ward audit (e.g. providing feedback recipients with suggested actions for improvement) and discuss implications for theory. There is a need to reduce the use of punitive feedback.


Differential Impacts of Land-Based Sources of Pollution on the Microbiota of Southeast Florida Coral Reefs.
Coral reefs are dynamic ecosystems known for decades to be endangered due, in large part, to anthropogenic impacts from land-based sources of pollution (LBSP). In this study, we utilized an Illumina-based next-generation sequencing approach to characterize prokaryotic and fungal communities from samples collected off the southeast coast of Florida. Water samples from coastal inlet discharges, oceanic outfalls of municipal wastewater treatment plants, treated wastewater effluent before discharge, open ocean samples, and coral tissue samples (mucus and polyps) were characterized to determine the relationships between microbial communities in these matrices and those in reef water and coral tissues. Significant differences in microbial communities were noted among all sample types but varied between sampling areas. Contamination from outfalls was found to be the greatest potential source of LBSP influencing native microbial community structure among all reef samples, although pollution from inlets was also noted. Notably, reef water and coral tissue communities were found to be more greatly impacted by LBSP at southern reefs, which also experienced the most degradation during the course of the study. The results of this study provide new insights into how microbial communities from LBSP can impact coral reefs in southeast Florida and suggest that wastewater outfalls may have a greater influence on the microbial diversity and structure of these reef communities than do contaminants carried in runoff, although the influences of runoff and coastal inlet discharge on coral reefs are still substantial.IMPORTANCE Coral reefs are known to be endangered due to sewage discharge and to runoff of nutrients, pesticides, and other substances associated with anthropogenic activity. Here, we used next-generation sequencing to characterize the microbial communities of potential contaminant sources in order to determine how environmental discharges of microbiota and their genetic material may influence the microbiomes of coral reef communities and coastal receiving waters. Runoff delivered through inlet discharges impacted coral microbial communities, but impacts from oceanic outfalls carrying treated wastewater were greater. Geographic differences in the degree of impact suggest that coral microbiomes may be influenced by the microbiological quality of treated wastewater.


Inspiratory muscle fatigue following running to volitional fatigue: the influence of baseline strength.
Respiratory muscle fatigue has been demonstrated following short-term exercise to volitional fatigue, as well as following prolonged submaximal exercise. There is some suggestion that the respiratory muscles of 'athletic' individuals have superior strength and greater fatigue resistance but it is not known whether inspiratory muscle strength influences fatigueability of the inspiratory muscles. The present study examined this question in 24 moderately trained young men. Inspiratory muscle strength was measured at residual volume using a hand held Mouth Pressure Meter before and after an incremental, multistage shuttle run to volitional fatigue. Following the run, there was a significant fall in inspiratory mouth pressures (-10.5 +/- SD 8.2%; p < 0.001 Pre- vs Post Pipeak). The subjects with the weakest inspiratory muscles exhibited significantly greater fatigue than those with the strongest (-17.0 +/- SD 7.8% c.f. 6.8 +/- SD 4.4% for the 25th and 75th percentiles respectively p < 0.01). These data support existing evidence that the respiratory muscles fatigue following high intensity exercise. In addition, they provide new evidence that this phenomenon occurs in moderately trained young men and that the severity of the fatigue is related to the baseline strength of the inspiratory muscles.


Histone deacetylase 6 in health and disease.
Histone deacetylase (HDAC)6 is a member of the class IIb HDAC family. This enzyme is zinc-dependent and mainly localized in the cytoplasm. HDAC6 is a unique isoenzyme with two functional catalytic domains and specific physiological roles. Indeed, HDAC6 deacetylates various substrates including α-tubulin and HSP90α, and is involved in protein trafficking and degradation, cell shape and migration. Consequently, deregulation of HDAC6 activity was associated to a variety of diseases including cancer, neurodegenerative diseases and pathological autoimmune response. Therefore, HDAC6 represents an interesting potential therapeutic target. In this review, we discuss structural features of this histone deacetylase, regulation of its expression and activity, biological functions, implication in human disease initiation and progression. Finally will describe novel and selective HDAC6 inhibitors.


Incidence of atrial fibrillation in cryptogenic stroke with patent foramen ovale closure: protocol for the prospective, observational PFO-AF study.
After closure of patent foramen ovale (PFO) due to stroke, atrial fibrillation (AF) occurs in up to one in five patients. However, data are sparse regarding the possible pre-existence of AF in these patients prior to PFO closure, and about recurrence of AF in the long term after the procedure. No prospective study to date has investigated these topics in patients with implanted cardiac monitor (ICM). The PFO-AF study (registered with ClinicalTrials.gov under the number NCT04926142) will investigate the incidence of AF occurring within 2 months after percutaneous closure of PFO in patients with prior stroke. AF will be identified using systematic ICM. Secondary objectives are to assess incidence and burden of AF in the 2 months prior to, and up to 2 years after PFO closure. Prospective, multicentre, observational study including 250 patients with an indication for PFO closure after stroke, as decided by interdisciplinary meetings with cardiologists and neurologists. Patients will undergo implantation of a Reveal Linq device (Medtronic). Percutaneous PFO closure will be performed 2 months after device implantation. Follow-up will include consultation, ECG and reading of ICM data at 2, 12 and 24 months after PFO closure. The primary endpoint is occurrence of AF at 2 months, defined as an episode of AF or atrial tachycardia/flutter lasting at least 30 s, and recorded by the ICM and/or any AF or atrial tachycardia/flutter documented on ECG during the first 2 months of follow-up. The study was approved by the Ethics Committee 'Comité de Protection des Personnes (CPP) Sud-Méditerranéen III' on 2 June 2021 and registered with ClinicalTrials.gov (NCT04926142). Findings will be presented in national and international congresses and peer-reviewed journals. NCT04926142.


Role of RNA secondary structure and processing in stability of the nifH1 transcript in the cyanobacterium Anabaena variabilis.
In the cyanobacterium Anabaena variabilis ATCC 29413, aerobic nitrogen fixation occurs in micro-oxic cells called heterocysts. Synthesis of nitrogenase in heterocysts requires expression of the large nif1 gene cluster, which is primarily under the control of the promoter for the first gene, nifB1. Strong expression of nifH1 requires the nifB1 promoter but is also controlled by RNA processing, which leads to increased nifH1 transcript stability. The processing of the primary nifH1 transcript occurs at the base of a predicted stem-loop structure that is conserved in many heterocystous cyanobacteria. Mutations that changed the predicted secondary structure or changed the sequence of the stem-loop had detrimental effects on the amount of nifH1 transcript, with mutations that altered or destabilized the structure having the strongest effect. Just upstream from the transcriptional processing site for nifH1 was the promoter for a small antisense RNA, sava4870.1. This RNA was more strongly expressed in cells grown in the presence of fixed nitrogen and was downregulated in cells 24 h after nitrogen step down. A mutant strain lacking the promoter for sava4870.1 showed delayed nitrogen fixation; however, that phenotype might have resulted from an effect of the mutation on the processing of the nifH1 transcript. The nifH1 transcript was the most abundant and most stable nif1 transcript, while nifD1 and nifK1, just downstream of nifH1, were present in much smaller amounts and were less stable. The nifD1 and nifK1 transcripts were also processed at sites just upstream of nifD1 and nifK1. In the filamentous cyanobacterium Anabaena variabilis, the nif1 cluster, encoding the primary Mo nitrogenase, functions under aerobic growth conditions in specialized cells called heterocysts that develop in response to starvation for fixed nitrogen. The large cluster comprising more than a dozen nif1 genes is transcribed primarily from the promoter for the first gene, nifB1; however, this does not explain the large amount of transcript for the structural genes nifH1, nifD1, and nifK1, which are also under the control of the distant nifB1 promoter. Here, we demonstrate the importance of a predicted stem-loop structure upstream of nifH1 that controls the abundance of nifH1 transcript through transcript processing and stabilization and show that nifD1 and nifK1 transcripts are also controlled by transcript processing.


[Endoscopic polypectomy of the colon in children].
Colon Polyps constitute are one of the most frequent causes of rectal bleeding in infancy. The advent of endoscopic polypectomy through colonoscopy has resulted very useful in their treatment. From 1979 to 1984, 73 polypectomies were performed in 64 patients with colonic polyps, between 2 and 14 years old, 26 girls and 38 boys. 103 polyps were resected, 71 pedunculated and 32 sessiles. The most frequent location was the recto-sigmoid (85.6%), with 14.4% situated above it. 55.7% of the removed polyps were more than 1 cm., juvenile histologic type predominating (90%). No complication was observed either during or after this procedure. It is demonstrated that, endoscopic polypectomy in children is a safe, simple and effective method which avoids complications that may or surgery might produce.


Hepatitis C virus infection and oral lichen planus: a report from The Netherlands.
The reported frequency of anti-hepatitis C virus (HCV) antibodies in groups of cutaneous lichen planus (CLP) and/or oral lichen planus (OLP) patients varies from about 4% to 65%. Most of these studies have been performed in countries with a high overall prevalence of HCV infection in the general population, such as the southern European countries and Japan. Limited data are available from areas with a low prevalence of HCV infection. Therefore, we investigated the prevalence of HCV infection in a series of 55 patients with OLP in the Netherlands, which apparently has a low prevalence of HCV infection. None of the 55 patients revealed serological evidence of antibodies to HCV. The present data suggest that HCV infection in OLP patients in the Netherlands is probably not very common. A larger study group with a sex- and age-matched control group is required to advise against routine serological examination for HCV infection in Dutch OLP patients.


Importance of the long-acting partner drug in artemisinin-based combination therapy.
The 1st African Health Research Organization International Malaria Symposium on Clinical Pharmacology of Antimalarial Drugs collected approximately 60 health professionals, scientists, policy makers and nongovernmental organization representatives from Africa, Europe and the USA for updates on recent developments in malaria therapy. It was the first African Health Research Organization symposium in Africa aimed at taking an African leadership in the fight against malaria. The intention is to start a tradition of annually recurring symposia that will eventually grow to become a leading international event in malaria research. Apart from scientific presentations, substantial time was dedicated to discussions and brainstorming, with a view to developing tools that can realistically be used to address challenges relating to all aspects of malaria treatment and control. The meeting was funded partly by the Medicines for Malaria Venture and Dafra Pharma International Ltd., Belgium, both of which were represented by some of the speakers.


Transmembrane topology and histidine protein kinase activity of AgrC, the agr signal receptor in Staphylococcus aureus.
The agr P2 operon in Staphylococcus aureus codes for the elements of a density-sensing cassette made up of a typical two-component signalling system and its corresponding inducer. It is postulated that the autoinducer, a post-translationally modified octapeptide generated from the AgrD peptide, interacts with a receptor protein, coded by agrC, to transmit a signal via AgrA regulating expression of staphylococcal virulence genes through expression of agr RNA III. We show by analysis of PhoA fusions that AgrC is a transmembrane protein, and confirm using Western blotting that a 46 kDa protein corresponding to AgrC is present in the bacterial membrane. This protein is autophosphorylated on a histidine residue only in response to supernatants from an agr+ strain, and can also respond to the purified native octapeptide. A recombinant fusion protein where most of the N-terminal region of AgrC is replaced by the Escherichia coli maltose-binding protein is also autophosphorylated in response to stimulation by agr+ supernatants or purified octapeptide. We conclude that AgrC is the sensor molecule of a typical two-component signal system in S. aureus, and that the ligand-binding site of AgrC is probably located in the third extracellular loop of the protein.


Histopathologcial and clonal study of combined lobular and ductal carcinoma of the breast.
Lobular carcinoma in situ (LCIS) clinically constitutes a risk factor for the subsequent development of either invasive lobular carcinoma (ILC) or invasive ductal carcinoma (IDC). In order to approach the possibility of this common precursor of both ILC and IDC, we investigated combined lobular and ductal carcinomas. Thirty-two cases of lobular carcinoma were picked up out of 773 cases of operated breast carcinomas. The histopathological detailed re-examination using immunostain of E-cadherin and β-catenin revealed a rather high frequency of combined lobular carcinomas than previous reports. Clinicopathologically, combined lobular carcinomas were younger and smaller than pure lobular carcinomas, and the cytological atypia was relatively low. These results suggested that combined lobular carcinomas could be detected in the earlier stage of breast cancer. Furthermore, the lobular and ductal components of combined carcinomas coexisted in the neighborhood and were distributed contiguously. The immunohistochemical phenotypes of both components were accorded in most combined cases. A genetic analysis using methylation-specific PCR on the HUMARA gene demonstrated that the same allele was inactivated in both lobular and ductal components in all detectable cases of combined carcinoma. Therefore, it is reasonable to assume that both lobular and ductal components of combined carcinomas are clonal and derived from the LCIS as the common precursor lesion, which may contradict the conventional concept that the lobular and ductal carcinomas arise from distinct differentiation pathways.


Antiretinal antibodies in central serous chorioretinopathy: prevalence and clinical implications.
To investigate the possible role of autoimmune reactions directed against retinal tissue in central serous chorioretinopathy (CSC), by analysing the presence of serum antiretinal antibodies (ARAs) and establishing their clinical relevance. Sixty-three patients with CSC were included, and clinical characteristics were collected. Serum samples of all patients with CSC, 101 uveitis patients and 60 healthy donors were analysed for the presence of ARAs by indirect immunofluorescence. Furthermore, all CSC serum samples were analysed on Western blot. Correlations between laboratory findings and clinical features of CSC were determined by logistic regression. Antiretinal antibodies (ARAs) were present in 54% of the patients with CSC, in 46% of uveitis patients (p = 0.153) and in 17% of healthy controls (p < 0.001). The majority of ARAs in CSC were directed against photoreceptors (27%), which occurred significantly more often compared to uveitis patients (15%, p = 0.039) and to healthy controls (5%, p = 0.003). No associations between clinical CSC characteristics and the presence of ARAs were found. Serum ARAs are present in more than half of the patients with CSC, and especially, ARAs directed against photoreceptors were detected more frequently compared to both healthy controls and uveitis patients. Further research is warranted to unravel the role of ARAs in the pathogenesis of CSC.


Gene expression and parental dominance in hybrid plants.
Genomic imprinting, where the genes from one parent have different expression properties to those of the other parent, occurs in plants. It has potentially significant consequences because of the importance of hybrids in plant evolution and plant breeding, and provides a mechanism that can hide genetic variation for many generations. The study of nuclear organization shows that chromosome and genome position relates to imprinting in F1 hybrids, with peripheral genomes tending to be expressed preferentially. In some inbred, polyploid hybrids, such as Triticale (a wheat x rye hybrid), treatment with the demethylation agent azacytidine releases hidden variation, which was perhaps lost because of imprinting phenomena.


Atomic-scale intermolecular interaction of hydrogen with a single VOPc molecule on the Au(111) surface.
Molecular dynamics of hydrogen molecules (H2) on surfaces and their interactions with other molecules have been studied with the goal of improvement of hydrogen storage devices for energy applications. Recently, the dynamic behavior of a H2 at low temperature has been utilized in scanning tunnelling microscopy (STM) for sub-atomic resolution imaging within a single molecule. In this work, we have investigated the intermolecular interaction between H2 and individual vanadyl phthalocyanine (VOPc) molecules on Au(111) substrates by using STM and non-contact atomic force microscopy (NC-AFM). We measured tunnelling spectra and random telegraphic noise (RTN) on VOPc molecules to reveal the origin of the dynamic behavior of the H2. The tunnelling spectra show switching between two states with different tunnelling conductance as a function of sample bias voltage and RTN is measured near transition voltage between the two states. The spatial variation of the RTN indicates that the two-state fluctuation is dependent on the atomic-scale interaction of H2 with the VOPc molecule. Density functional theory calculations show that a H2 molecule can be trapped by a combination of a tip-induced electrostatic potential well and the potential formed by a VOPc underneath. We suggest the origin of the two-state noise as transition of H2 between minima in these potentials with barrier height of 20-30 meV. In addition, the bias dependent AFM images verify that H2 can be trapped and released at the tip-sample junction.


KDM4B, a potential prognostic biomarker revealed by large-scale public databases and clinical samples in uterine corpus endometrial carcinoma.
Uterine corpus endometrial carcinoma (UCEC) is the fourth most common cancer among women worldwide. The 5 year survival rate for patients with advanced UCEC is only 17%. Lysine-specific demethylase 4B (KDM4B), a histone demethylase, is overexpressed or dysregulated in various cancers and is associated with tumor progression and poor prognosis. We performed bioinformatics analysis and in vitro assays to assess the role of KDM4B in UCEC. Additionally, the role of KDM4B in the tumor immune microenvironment was explored. The mRNA and protein levels of KDM4B in UCEC were evaluated using The Cancer Genome Atlas (TCGA), The Human Protein Atlas (HPA), and Gene Expression Omnibus (GEO) databases. Immunohistochemistry and western blotting were performed to verify the protein expression level of KDM4B in two batches of clinical samples. Kaplan-Meier survival, univariate, and multivariate analyses were performed to assess the correlation between KDM4B expression and the prognosis of patients with UCEC. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses were performed to predict the function and mechanism of KDM4B, and four immune-related databases (TIMER, CIBERSORT, TISIDB, and EPIC) were used to explore their relevance in a tumor immune microenvironment. First, KDM4B was significantly overexpressed in UCEC tissues at the mRNA and protein levels. Immunohistochemistry and western blotting confirmed the abnormal overexpression of KDM4B. Additionally, upregulation of KDM4B was associated with different clinicopathological prognostic factors. Second, the overexpression of KDM4B was also associated with shorter overall survival (OS) and progression-free survival (PFS). Univariate and multivariate analyses confirmed that KDM4B was an independent prognostic factor for poor survival. Then, GO and KEGG analysis revealed that KDM4B is enriched in biological processes and cellular signaling pathways closely related to the immune response. Finally, KDM4B expression was closely associated with immune cell infiltration and expression of immune checkpoint molecules, indicating that it may be a new immune-related potential oncogene in UCEC. KDM4B may be a new potential oncogene that is clinically significant in patients with UCEC. KDM4B may not only be used to assess the clinical prognosis of patients with UCEC but may also be a target for immunotherapy or targeted gene therapy.


Interaction of Hsp27 with native phosphorylase kinase under crowding conditions.
Interaction of the wild type (wt) heat shock protein Hsp27 and its three-dimensional (3D) mutant (mimicking phosphorylation at Ser15, 78, and 82) with rabbit skeletal muscle phosphorylase kinase (PhK) has been studied under crowding conditions modeled by addition of 1 M trimethylamine N-oxide (TMAO). According to the data of sedimentation velocity and dynamic light scattering, crowding provokes the formation of large-sized associates of both PhK and Hsp27. Under crowding conditions, small associates of PhK and Hsp27 interact with each other thus leading to dissociation of large homooligomers of each protein. Taking into account high concentrations of PhK in the cell, we speculate that native PhK might modulate the oligomeric state and chaperone-like activity of Hsp27.


A role for phosphodiesterase 3B in acquisition of brown fat characteristics by white adipose tissue in male mice.
Obesity is linked to various diseases, including insulin resistance, diabetes, and cardiovascular disorders. The idea of inducing white adipose tissue (WAT) to assume characteristics of brown adipose tissue (BAT), and thus gearing it to fat burning instead of storage, is receiving serious consideration as potential treatment for obesity and related disorders. Phosphodiesterase 3B (PDE3B) links insulin- and cAMP-signaling networks in tissues associated with energy metabolism, including WAT. We used C57BL/6 PDE3B knockout (KO) mice to elucidate mechanisms involved in the formation of BAT in epididymal WAT (EWAT) depots. Examination of gene expression profiles in PDE3B KO EWAT revealed increased expression of several genes that block white and promote brown adipogenesis, such as C-terminal binding protein, bone morphogenetic protein 7, and PR domain containing 16, but a clear BAT-like phenotype was not completely induced. However, acute treatment of PDE3B KO mice with the β3-adrenergic agonist, CL316243, markedly increased the expression of cyclooxygenase-2, which catalyzes prostaglandin synthesis and is thought to be important in the formation of BAT in WAT and the elongation of very long-chain fatty acids 3, which is linked to BAT recruitment upon cold exposure, causing a clear shift toward fat burning and the induction of BAT in KO EWAT. These data provide insight into the mechanisms of BAT formation in mouse EWAT, suggesting that, in a C57BL/6 background, an increase in cAMP, caused by ablation of PDE3B and administration of CL316243, may promote differentiation of prostaglandin-responsive progenitor cells in the EWAT stromal vascular fraction into functional brown adipocytes.


Surface coating of PLGA microparticles with protamine enhances their immunological performance through facilitated phagocytosis.
Surface modifications of poly(lactide-co-glycolide) microparticles with different polycationic electrolytes have mainly been studied for conjugation to antigens and/or adjuvants. However, the in vivo immunological effects of using surface charged particles have not been address yet. In this study, microparticles were coated or not with protamine, a cationic and arginine-rich electrolyte that confers microparticles with a positively surface charge. We then evaluated the potential of protamine-coatings to assist the induction of immune responses in mice. Interestingly, enhanced antibodies and T-cell responses were observed in mice treated with the coated particles. In vitro studies suggested that the improved immunological performance was mediated by an increased uptake. Indeed, protamine-coated particles that carried a plasmid were even internalised into non-phagocytic cells and to cause their transfection. These results open the way for further research into a novel technology that combines the use protamine for facilitated cell penetration of that and biodegradable microparticles for prolonged antigen or drug release.


Immune dysregulation in myelodysplastic syndrome.
Myelodysplastic syndrome (MDS) represents one of the most challenging health-related problems in the elderly. Characterized by dysplastic morphology in the bone marrow in association with ineffective hematopoiesis, pathophysiological causes of this disease are diverse including genetic abnormalities within myeloid progenitors, altered epigenetics, and changes in the bone marrow microenvironment. The concept that T-cell mediated autoimmunity contributes to bone marrow failure has been widely accepted due to hematologic improvement after immunosuppressive therapy (IST) in a subset of patients. Currently, IST for MDS primarily involves anti-thymocyte globulin (ATG)-based regimens in which responsiveness is strongly associated with younger (under 60 years) age at disease onset. In such cases, progressive cytopenia may occur as a consequence of expanded self-reactive CD8(+) cytotoxic T lymphocytes (CTLs) that suppress hematopoietic progenitors. Although most hematologists agree that IST can offer durable hematologic remission in younger patients with MDS, an international clinical study and a better understanding of the molecular mechanisms contributing to the expansion of self-reactive CTLs is crucial. In this review, data accumulated in the US, Europe, and Asia will be summarized to provide insight and direction for a multi-center international trial.


Guidelines to Compose an Ideal Bacteriophage Cocktail.
Properly designed bacteriophage therapeutics are the cornerstone for a successful outcome of bacteriophage therapy. Here we present an overview of the different strategies and steps that can be taken to develop a bacteriophage cocktail that complies with relevant quality and safety requirements. It is based on empirical bacteriophage therapy knowledge from over a century of experience, more recently performed studies, and emerging technologies. We emphasize the selection of adequate bacteriophages and describe a modified Appelmans' method to improve the overall performance of therapeutic bacteriophages individually and collectively in the cocktail. We present two versions of the method, which differ from each other by the employed techniques to evaluate phage activity and synergy: photometric assessment of bacterial growth versus measurement of bacterial respiration via the Omnilog® system.


Estimated US Cancer Deaths Prevented With Increased Use of Lung, Colorectal, Breast, and Cervical Cancer Screening.
Increased use of recommended screening could help achieve the Cancer Moonshot goal of reducing US cancer deaths. To estimate the number of cancer deaths that could be prevented with a 10-percentage point increase in the use of US Preventive Services Task Force (USPSTF)-recommended screening. This decision analytical model study is an extension of previous studies conducted for the USPSTF from 2018 to 2023. This study simulated contemporary cohorts of US adults eligible for lung, colorectal, breast, and cervical cancer screening. Annual low-dose computed lung tomography among eligible adults aged 50 to 80 years; colonoscopy every 10 years among adults aged 45 to 75 years; biennial mammography among female adults aged 40 to 74 years; and triennial cervical cytology screening among female adults aged 21 to 29 years, followed by human papillomavirus testing every 5 years from ages 30 to 65 years. Estimated number of cancer deaths prevented with a 10-percentage point increase in screening use, assuming screening commences at the USPSTF-recommended starting age and continues throughout the lifetime. Outcomes were presented 2 ways: (1) per 100 000 and (2) among US adults in 2021; and they were expressed among the target population at the age of screening initiation. For lung cancer, estimates were among those who will also meet the smoking eligibility criteria during their lifetime. Harms from increased uptake were also reported. A 10-percentage point increase in screening use at the age that USPSTF recommended screening commences was estimated to prevent 226 lung cancer deaths (range across models within the cancer site, 133-332 deaths), 283 (range, 263-313) colorectal cancer deaths, 82 (range, 61-106) breast cancer deaths, and 81 (1 model; no range available) cervical cancer deaths over the lifetimes of 100 000 persons eligible for screening. These rates corresponded with an estimated 1010 (range, 590-1480) lung cancer deaths prevented, 11 070 (range, 10 280-12 250) colorectal cancer deaths prevented, 1790 (range, 1330-2310) breast cancer deaths prevented, and 1710 (no range available) cervical cancer deaths prevented over the lifetimes of eligible US residents at the recommended age to initiate screening in 2021. Increased uptake was also estimated to generate harms, including 100 000 (range, 45 000-159 000) false-positive lung scans, 6000 (range, 6000-7000) colonoscopy complications, 300 000 (range, 295 000-302 000) false-positive mammograms, and 348 000 (no range available) colposcopies over the lifetime. In this decision analytical model study, a 10-percentage point increase in uptake of USPSTF-recommended lung, colorectal, breast, and cervical cancer screening at the recommended starting age was estimated to yield important reductions in cancer deaths. Achieving these reductions is predicated on ensuring equitable access to screening.


Associations Between Neighborhood Social Capital, Health Literacy, and Self-Rated Health Among People With Chronic Illness.
Health literacy skills are important for health and self-management for people with chronic illness. Neighborhood social capital can provide resources, such as access to information and informal social control over unhealthy behavior. The benefit of these resources, and the access people have to these resources, might depend on levels of health literacy. We investigated whether neighborhood social capital is differentially related to the health of people with chronic illness according to health literacy skills. This study focused on health literacy skills in 4 domains related to the ability to access and understand health information and to the ability to perform self-management. We found a significant positive interaction between social capital and health literacy skills for accessing and understanding health information. This suggests that health literacy enhances people's ability to gain access to and use neighborhood resources to benefit health. There was no interaction effect between social capital and health literacy skills in the other 2 domains. More research is needed to investigate how people with chronic illness can benefit from knowledge, support, and other social resources for health and self-management also whether they have limited health literacy skills.


Impact of involved free light chain (FLC) levels in patients achieving normal FLC ratio after initial therapy in light chain amyloidosis (AL).
Achievement of a normal FLC ratio (FLCr) following treatment indicates hematologic response and suggests better outcomes in light chain amyloidosis (AL). We examined if elevated involved free light chain (hiFLC) impacts outcomes in patients achieving normal FLCr. We retrospectively analyzed 345 AL patients who were diagnosed within a 10-year period (2006-2015) and had 2 consecutive normal FLCr values after 1st line treatment. Among these, patients with hiFLC at 1st reading of normal FLCr (hiFLC1; n = 166; 48.1%) were compared to those who did not (n = 179; 51.9%). Patients with AL who have hiFLC1 after initial therapy had higher rates of multi-organ involvement (63.3 vs 46.4%; P = .002) and patients in advanced Mayo stage (42.9 vs 32.2%; P = .04) at diagnosis. The median progression free survival [PFS; 38.2 (95%CI; 26.4, 55.4) vs 67.1 (95%CI; 55.8, 88) months; P = .0002] and overall survival [OS; 94.4 (95%CI; 78, 107.1) vs not reached (NR, 95%CI; 116.1, NR) months; P < .0001] were lower in those who had hiFLC1. A more stringent comparison for patients with 2 consecutive hiFLC (hIFLC2; n = 111; 32.2%) versus not (n = 2234; 67.8%) showed consistent results [PFS; 27.1 (95%CI; 23, 53.8) vs 63.3 (95%CI; 55.4, 77) months; P < .0001 and OS; 78 (95% CI; 54.6, 98.8) vs NR (95%CI; NR, NR); P < .0001]. This poor prognostic impact of hiFLC on survival was independent of serum creatinine, Mayo stage, negative immunofixation status and inclusion of transplant in initial therapy on multivariate analysis. Hence, persistent elevation of iFLC predicts poor prognosis even among patients achieving normal ratio after initial therapy in AL.


In vivo and in vitro L-DOPA and reticuline exposure increases ganglionic morphine levels.
Given the presence of morphine, its metabolites and precursors, i.e., reticuline, in mammalian and invertebrate tissues, it has become imperative to determine if exposing tissues to putative opiate alkaloid and dopamine precursors would result in increasing endogenous morphine levels. Endogenous morphine levels were determined by high performance liquid chromatography coupled to electrochemical detection and radioimmunoassay, following incubation of Mytilus edulis pedal ganglia with reticuline or L-DOPA. Injection of L-DOPA or reticuline into healthy animals was via the foot. Ganglia incubated in vitro with reticuline or L-DOPA for 1 hour exhibited a concentration and time dependent statistically significant increase in their endogenous morphine levels (5.0 +/- 0.47, 3.6 +/- 0.45 ng/ganglion, respectively). Injection of intact, healthy animals with reticuline or L-DOPA also results in significantly higher endogenous ganglionic morphine levels. Taken together, we show that L-DOPA is being converted to morphine, demonstrating that pedal ganglia can synthesize morphine from these putative precursors in vitro and in vivo. This is the first demonstration of morphine being synthesized in a normal, healthy free living animal.


Synthesis, production, and biotechnological applications of exopolysaccharides and polyhydroxyalkanoates by archaea.
Extreme environments, generally characterized by atypical temperatures, pH, pressure, salinity, toxicity, and radiation levels, are inhabited by various microorganisms specifically adapted to these particular conditions, called extremophiles. Among these, the microorganisms belonging to the Archaea domain are of significant biotechnological importance as their biopolymers possess unique properties that offer insights into their biology and evolution. Particular attention has been devoted to two main types of biopolymers produced by such peculiar microorganisms, that is, the extracellular polysaccharides (EPSs), considered as a protection against desiccation and predation, and the endocellular polyhydroxyalkanoates (PHAs) that provide an internal reserve of carbon and energy. Here, we report the composition, biosynthesis, and production of EPSs and PHAs by different archaeal species.


Activation of swelling-activated chloride current by tumor necrosis factor-alpha requires ClC-3-dependent endosomal reactive oxygen production.
ClC-3 is a Cl(-)/H(+) antiporter required for cytokine-induced intraendosomal reactive oxygen species (ROS) generation by Nox1. ClC-3 current is distinct from the swelling-activated chloride current (ICl(swell)), but overexpression of ClC-3 can activate currents that resemble ICl(swell). Because H(2)O(2) activates ICl(swell) directly, we hypothesized that ClC-3-dependent, endosomal ROS production activates ICl(swell). Whole-cell perforated patch clamp methods were used to record Cl(-) currents in cultured aortic vascular smooth muscle cells from wild type (WT) and ClC-3 null mice. Under isotonic conditions, tumor necrosis factor-alpha (TNF-alpha) (10 ng/ml) activated outwardly rectifying Cl(-) currents with time-dependent inactivation in WT but not ClC-3 null cells. Inhibition by tamoxifen (10 microm) and by hypertonicity (340 mosm) identified them as ICl(swell). ICl(swell) was also activated by H(2)O(2) (500 microm), and the effect of TNF-alpha was completely inhibited by polyethylene glycol-catalase. ClC-3 expression induced ICl(swell) in ClC-3 null cells in the absence of swelling or TNF-alpha, and this effect was also blocked by catalase. ICl(swell) activation by hypotonicity (240 mosm) was only partially inhibited by catalase, and the size of these currents did not differ between WT and ClC-3 null cells. Disruption of endosome trafficking with either mutant Rab5 (S34N) or Rab11 (S25N) inhibited TNF-alpha-mediated activation of ICl(swell). Thrombin also activates ROS production by Nox1 but not in endosomes. Thrombin caused H(2)O(2)-dependent activation of ICl(swell), but this effect was not ClC-3- or Rab5-dependent. Thus, activation of ICl(swell) by TNF-alpha requires ClC-3-dependent endosomal H(2)O(2) production. This demonstrates a functional link between two distinct anion currents, ClC-3 and ICl(swell).


Analysis of urinary exosomes applications for rare kidney disorders.
Introduction: Exosomes are nanovesicles that play important functions in a variety of physiological and pathological conditions. They are powerful cell-to-cell communication tool thanks to the protein, mRNA, miRNA, and lipid cargoes they carry. They are also emerging as valuable diagnostic and prognostic biomarker sources. Urinary exosomes carry information from all the cells of the urinary tract, downstream of the podocyte. Rare kidney diseases are a subset of an inherited diseases whose genetic diagnosis can be unclear, and presentation can vary due to genetic, epigenetic, and environmental factors. Areas covered: In this review, we focus on a group of rare and often neglected kidney diseases, for which we have sufficient available literature data on urinary exosomes. The analysis of their content can help to comprehend pathological mechanisms and to identify biomarkers for diagnosis, prognosis, and therapeutic targets. Expert opinion: The foreseeable large-scale application of system biology approach to the profiling of exosomal proteins as a source of renal disease biomarkers will be also useful to stratify patients with rare kidney diseases whose penetrance, phenotypic presentation, and age of onset vary sensibly. This can ameliorate the clinical management.


Ancyromonadida: a new phylogenetic lineage among the protozoa closely related to the common ancestor of metazoans, fungi, and choanoflagellates (Opisthokonta).
Molecular and morphological evidence points to the ancyromonad Ancyromonas as a plausible candidate for the closest relative to the common ancestor of metazoans, fungi, and choanoflagellates (the Opisthokonta). Using 18S rDNA sequences from most of the major eukaryotic lineages, maximum-likelihood, minimum-evolution, and maximum-parsimony analyses yielded congruent phylogenies supporting this hypothesis. Combined with ultrastructural similarities between Ancyromonas and opisthokonts, the evidence presented here suggests that Ancyromonas may form an independent lineage, the Ancyromonadida Cavalier-Smith 1997, closer in its relationship to the opisthokonts than is its nearest protist relatives, the Apusomonadida. However, the very low bootstrap support for deep nodes and hypothesis testing indicate that the resolving power of 18S rDNA sequences is limited for examining this aspect of eukaryotic phylogeny. Alternate branching positions for the Ancyromonas lineage cannot be robustly rejected, revealing the importance of ultrastructure when examining the origins of multicellularity. The future use of a multigene approach may additionally be needed to resolve this aspect of eukaryotic phylogeny.


Effect of addition of CO₂ to raw milk on quality of UHT-treated milk.
The objective of this study was to evaluate the effect of addition of CO(2) to raw milk on UHT milk quality during storage. Control milk (without CO(2) addition) and treated milk (with CO(2) addition up to pH 6.2) were stored in bulk tanks at 4°C for 6d. After storage, both samples were UHT processed using indirect heating (140°C for 5s). Samples were aseptically packed in low-density polyethylene pouches and stored in the dark at room temperature. Raw milk was evaluated upon receipt for physicochemical composition, proteolysis, lipolysis, standard plate count, psychrotrophic bacteria, and Pseudomonas spp. counts, and after 6d of storage for proteolysis, lipolysis, and microbial counts. After processing, UHT milk samples were evaluated for physicochemical composition, proteolysis, and lipolysis. Samples were evaluated for proteolysis and lipolysis twice a month until 120d. Peptides from pH 4.6-soluble N filtrates were performed by reversed-phase HPLC after 1 and 120d of storage. A split-plot design was used and the complete experiment was carried out in triplicate. The results were evaluated by ANOVA and Tukey's test. After 6d of storage, CO(2)-treated raw milk kept its physicochemical and microbiological quality, whereas the untreated milk showed significant quality losses. A significant increase in proteolysis occurred during 120d of storage in both treatments, but the increase occurred 1.4 times faster in untreated UHT milk than in CO(2)-treated UHT milk. In both UHT milks, the proteolysis was a consequence of the action of plasmin and microbial proteases. However, the untreated UHT milk showed higher microbial protease activity than the treated UHT milk. The addition of CO(2) to the raw milk maintained the quality during storage, resulting in UHT milk with less proteolysis and possibly longer shelf life, which is usually limited by age gelation of UHT milk.


A global genomic analysis of Salmonella Concord reveals lineages with high antimicrobial resistance in Ethiopia.
Antimicrobial resistant Salmonella enterica serovar Concord (S. Concord) is known to cause severe gastrointestinal and bloodstream infections in patients from Ethiopia and Ethiopian adoptees, and occasional records exist of S. Concord linked to other countries. The evolution and geographical distribution of S. Concord remained unclear. Here, we provide a genomic overview of the population structure and antimicrobial resistance (AMR) of S. Concord by analysing genomes from 284 historical and contemporary isolates obtained between 1944 and 2022 across the globe. We demonstrate that S. Concord is a polyphyletic serovar distributed among three Salmonella super-lineages. Super-lineage A is composed of eight S. Concord lineages, of which four are associated with multiple countries and low levels of AMR. Other lineages are restricted to Ethiopia and horizontally acquired resistance to most antimicrobials used for treating invasive Salmonella infections in low- and middle-income countries. By reconstructing complete genomes for 10 representative strains, we demonstrate the presence of AMR markers integrated in structurally diverse IncHI2 and IncA/C2 plasmids, and/or the chromosome. Molecular surveillance of pathogens such as S. Concord supports the understanding of AMR and the multi-sector response to the global AMR threat. This study provides a comprehensive baseline data set essential for future molecular surveillance.


Elicitation of anti-H-2 cytotoxic T lymphocytes with antigen-modified H-2 negative stimulator cells.
By a newly developed technique, we have modified H-2k-negative cells in such a way as to render them capable of elicitation of anti-H-2 CTL. Purified H-2Kk and/or partially purified Iak glycoproteins were incorporated into lipid vesicles that contained the hemagglutinin-neuraminidase (HN) glycoprotein and the fusion (F) glycoprotein of Sendai virus. These lipid vesicles were incubated with H-2-negative tumor cells (R1-) in a manner that resulted in rendering these cells capable of eliciting anti-H-2Kk CTL. The presence of Iak on the surface of the modified stimulator cell had a significant effect on the proportion of anti-H-2Kk CTL possessing a particular Lyt phenotype. Primary effector cells elicited by an H-2Kk-Iak difference were primarily Lyt 1-, 2+, whereas primary anti-H-2Kk CTL elicited by an H-2Kk difference alone were predominantly Lyt 1+, 2+.