Solution structures of 2 : 1 and 1 : 1 DNA polymerase-DNA complexes probed by ultracentrifugation and small-angle X-ray scattering.
We report small-angle X-ray scattering (SAXS) and sedimentation velocity (SV) studies on the enzyme-DNA complexes of rat DNA polymerase beta (Pol beta) and African swine fever virus DNA polymerase X (ASFV Pol X) with one-nucleotide gapped DNA. The results indicated formation of a 2 : 1 Pol beta-DNA complex, whereas only 1 : 1 Pol X-DNA complex was observed. Three-dimensional structural models for the 2 : 1 Pol beta-DNA and 1 : 1 Pol X-DNA complexes were generated from the SAXS experimental data to correlate with the functions of the DNA polymerases. The former indicates interactions of the 8 kDa 5'-dRP lyase domain of the second Pol beta molecule with the active site of the 1 : 1 Pol beta-DNA complex, while the latter demonstrates how ASFV Pol X binds DNA in the absence of DNA-binding motif(s). As ASFV Pol X has no 5'-dRP lyase domain, it is reasonable not to form a 2 : 1 complex. Based on the enhanced activities of the 2 : 1 complex and the observation that the 8 kDa domain is not in an optimal configuration for the 5'-dRP lyase reaction in the crystal structures of the closed ternary enzyme-DNA-dNTP complexes, we propose that the asymmetric 2 : 1 Pol beta-DNA complex enhances the function of Pol beta.


Corticosteroids correct aberrant CFTR localization in the duct and regenerate acinar cells in autoimmune pancreatitis.
Corticosteroids are now widely accepted as a treatment for autoimmune pancreatitis (AIP). However, the molecular mechanism by which steroid treatment improves AIP remains largely unknown. The aim of this study was to elucidate cellular mechanisms by which corticosteroids improve both pancreatic exocrine function and histopathology in AIP. Pancreatic exocrine function was evaluated by the secretin-stimulated function test and pancreatic biopsy specimens were processed for histologic analysis at the time of diagnosis and 3 months after initiation of steroid treatment. Expression and localization of proteins was assayed by immunohistochemistry. Analysis of immunoglobulin (Ig)G4-positive plasma cells was used to verify inflammation in AIP. The number of IgG4-positive plasma cells in pancreatic sections was decreased by steroid treatment, indicating reduced inflammation. Fluid, bicarbonate (HCO(3)(-)), and digestive enzyme secretions all were impaired in most patients with AIP. Corticosteroids improved both HCO(3)(-) and digestive enzyme secretion. A large fraction of the cystic fibrosis transmembrane conductance regulator (CFTR), which plays a central role in pancreatic duct HCO(3)(-) secretion, was mislocalized to the cytoplasm of duct cells before treatment. Corticosteroids corrected the localization of CFTR to the apical membrane, accounting for the improved HCO(3)(-) secretion. Steroid treatment resulted in regeneration of acinar cells, accounting for restored digestive enzyme secretion. Corticosteroids reduce inflammation and restore both digestive enzyme and HCO(3)(-) secretion in patients with AIP by regenerating acinar cells and correcting CFTR localization in pancreatic duct cells. Mislocalization of CFTR may explain aberrant HCO(3)(-) secretion in other forms of pancreatitis.


Right paraduodenal hernia in an adult patient: diagnostic approach and surgical management.
Paraduodenal hernia, a rare congenital anomaly which arises from an error of rotation of the midgut, is the most common type of intraabdominal hernia. There are two variants, right and left paraduodenal hernia, the right being less common. We report the case of a 41-year-old patient with a right paraduodenal hernia with a 6-month history of intermittent episodes of intestinal obstruction. Diagnosis was established by CT scan and upper gastrointestinal series with small bowel follow-through. In a planned laparotomy, herniation of the small bowel loops through the fossa of Waldeyer was found. Division of the lateral right attachments of the colon opened the hernia sac widely, replacing the pre- and postarterial segments of the intestine in the positions they would normally occupy at the end of the first stage of rotation during embryonic development. Six months after the surgery, after an uneventful recovery, the patient remains free of symptoms.


Depression in hemodialysis patients.
Depression is the most frequent psychological complication of haemodialysis (HD) patients (pts) and has been associated with impaired Quality of Life (QoL). The aim of our study was to investigate the prevalence of depression in HD pts in relation to sociodemographic factors and the relationship between depression and QoL. 200 pts from Clinic for haemodialysis in Sarajevo, B&H were participating in the study. Mean age was 57.26+/-13.78 years and mean HD duration was 64.26+/-58.18 months. From the test material we applied BDI and SF-36. 51% of our pts have shown depression (BDI>11) in various degrees (30%-mild depression, 8.5%-moderate depression and 12.5%-severe depression). As we could expect, the most emphasized symptoms of depression were somatic symptoms. 55.5% of pts have shown QoL lower then average. Sociodemographic data such as gender, marital status and HD duration did not influence significantly on pt's QoL and occurrence of depression (p>0.05). As the age of the pts increased, level of depression increased too and QoL significantly decreased (p<0.05). Employed pts have shown significantly better QoL and lower level of depression in relation to unemployed pts (p<0.05). As the educational level of pts increased, QoL increased too and level of depression significantly decreased (p<0.05). Pts in 1st HD shift were significantly more depressed and have significantly worse mental health in compare to pts in 3rd HD shift (p<0.05). Our results showed a high prevalence of depressive symptoms among the study group that was linked to trend of poor QoL.


Age-dependency of ocular parameters: a cross sectional study of young and elderly healthy subjects.
To investigate the effect of aging on ocular parameters, including intraocular pressure (IOP), measured with different tonometry methods in healthy young (HY) and healthy elderly (HE) subjects and to study the effect of corneal parameters on tonometry methods. In this prospective, cross-sectional study, fifty eyes of 50 HY subjects (28 females, 22-31 years of age) and 43 eyes of 43 HE subjects (22 females, 64-79) were included. IOP was measured with four tonometry methods in a standardized order: ocular response analyser (ORA), dynamic contour tonometry (DCT), applanation resonance tonometry (ART) and Goldmann applanation tonometry (GAT). Other measurements included axial length (AL), central corneal thickness (CCT), corneal curvature (CC), anterior chamber volume (ACV), corneal hysteresis (CH) and corneal resistance factor (CRF). The mean IOP (HY/HE; mmHg ± standard deviation (SD)) was 12.2 ± 2.2/14.1 ± 3.5 with GAT. IOP was significantly higher (difference ± standard error) in HE compared to HY measured with an ORA (+3.1 mmHg ± 0.6), GAT (+1.9 ± 0.6) and DCT (+1.6 ± 0.6). No significant difference was found in IOP measured with ART. CH and ACV were significantly lower in HE compared to HY. There was no difference between the groups in CCT, CC, AL or CRF. No tonometry method was dependant on CCT or CC. IOP measured with an ORA and via DCT and GAT was higher in HE compared to HY Swedish subjects, while IOP measured with ART did not differ between the groups. In these homogeneous groups, tonometry methods were independent of CCT and CC.


The value of proteasome inhibition in cancer. Can the old drug, disulfiram, have a bright new future as a novel proteasome inhibitor?
The major approach to the development of anticancer drugs involves searching for new compounds, efficient against malignancies, which are not, as yet, used clinically. This strategy is time-consuming and expensive. Recent studies have disclosed a surprising, but mechanistically consistent, anticancer activity of disulfiram (antabuse), a drug used for about 50 years in the treatment of alcoholism. Disulfiram has been successfully used to suppress hepatic metastases originating from ocular melanoma. The pharmacokinetics of disulfiram and its pharmacological profile in cancer cell lines and in cancer cells obtained from patients is well known. Disulfiram is a readily available and inexpensive substance whose adverse effects are negligible, compared to classical cancerostatics. In addition, the inhibitory potency of disulfiram against the proteasome conforms to current anticancer strategies and represents a new, promising approach to proteasome inhibition.


A model of progressive photo-oxidative degeneration and inflammation in the pigmented C57BL/6J mouse retina.
Light-induced degeneration in rodent retinas is an established model for of retinal degeneration, including the roles of oxidative stress and neuroinflammatory activity. In these models, photoreceptor death is elicited via photo-oxidative stress, and is exacerbated by recruitment of subretinal macrophages and activation of immune pathways including complement propagation. Existing light damage models have relied heavily on albino rodents, and mostly using acute light stimuli. These albino models have proven valuable in uncovering the pathogenic mechanisms of such pathways in the context of retinal disease. However, their inherent albinism hinders comparability to normal retinal physiology, and also makes gene technology analysis time-consuming due to the predominance of the pigmented mouse strains in these applications. In this study, we characterise a new light damage model utilising C57BL/6J mice over a 7 day period of chronic light exposure. We use high-efficiency LED technology to deliver a sustained intensity of 100 k lux with negligible modulation of ambient temperature. We show that in the C57BL/6J mouse, chronic light exposure elicits the cardinal features of light damage including photoreceptor degeneration, atrophy of the choriocapillaris, decreased retinal function and increases in oxidative stress markers 4-HNE and 8-OHG, which emerge progressively over the 7 day period of exposure. These changes are accompanied by robust recruitment of IBA1+ and F4/80 + microglia/macrophages to the ONL and subretinal space, followed the strong up-regulation of monocyte-chemoattractants Ccl2, Ccl3, and Ccl12, as well as increases in expression of complement component C3. These findings are in agreement with prior damage models conducted in albino rodents such as Balb/c mice, and support the use of this new model in further investigating the causative features of oxidative stress and inflammation in retinal disease.


Comparison of binding of [3H]misonidazole and [14C]misonidazole in multicell spheroids.
Uptake of [2-ring-14C]misonidazole and [3H]misonidazole with tritium in the side chain has been compared in 1-mm EMT-6/UW spheroids using liquid scintillation counting and autoradiography. The uptake of both labeled sensitizers as a function of incubation time was virtually identical. Uptake by the spheroids exceeded levels in the medium by 11/2 to 2 hr and was well modeled as a first-order binding process, with rate constants of 0.00324 hr-1 for 3H and 0.00388 hr-1 for 14C. The similar uptake of the two versions of this sensitizer labeled in different positions suggests that the metabolic actions which allow the drug to bind in hypoxic cells do not principally involve metabolites which separate the number 2 carbon of the imidazole ring from the side chain. The pattern of silver grains in autoradiographs was similar for both labeled sensitizers, with most labeled drug bound in an intermediate zone of cells between the necrotic center and the actively proliferating rim of the spheroids. The superior resolution possible with the tritiated compound showed that both nucleus and cytoplasm in viable looking cells were labeled while pycnotic cells were not labeled.


Curvularia keratitis.
To determine the risk factors and clinical signs of Curvularia keratitis and to evaluate the management and outcome of this corneal phaeohyphomycosis. We reviewed clinical and laboratory records from 1970 to 1999 to identify patients treated at our institution for culture-proven Curvularia keratitis. Descriptive statistics and regression models were used to identify variables associated with the length of antifungal therapy and with visual outcome. In vitro susceptibilities were compared to the clinical results obtained with topical natamycin. During the 30-year period, our laboratory isolated and identified Curvularia from 43 patients with keratitis, of whom 32 individuals were treated and followed up at our institute and whose data were analyzed. Trauma, usually with plants or dirt, was the risk factor in one half; and 69% occurred during the hot, humid summer months along the US Gulf Coast. Presenting signs varied from superficial, feathery infiltrates of the central cornea to suppurative ulceration of the peripheral cornea. A hypopyon was unusual, occurring in only 4 (12%) of the eyes but indicated a significantly (P = .01) increased risk of subsequent complications. The sensitivity of stained smears of corneal scrapings was 78%. Curvularia could be detected by a panfungal polymerase chain reaction. Fungi were detected on blood or chocolate agar at or before the time that growth occurred on Sabouraud agar or in brain-heart infusion in 83% of cases, although colonies appeared only on the fungal media from the remaining 4 sets of specimens. Curvularia was the third most prevalent filamentous fungus among our corneal isolates and the most common dematiaceous mold. Corneal isolates included C senegalensis, C lunata, C pallescens, and C prasadii. All tested isolates were inhibited by 4 micrograms/mL or less of natamycin. Topical natamycin was used for a median duration of 1 month, but a delay in diagnosis beyond 1 week doubled the average length of topical antifungal treatment (P = .005). Visual acuity improved to 20/40 or better in 25 (78%) of the eyes. Curvularia keratitis typically presented as superficial feathery infiltration, rarely with visible pigmentation, that gradually became focally suppurative. Smears of corneal scrapings often disclosed hyphae, and culture media showed dematiaceous fungal growth within 1 week. Natamycin had excellent in vitro activity and led to clinical resolution with good vision in most patients with corneal curvulariosis. Complications requiring surgery were not common but included exophytic inflammatory fungal sequestration, treated by superficial lamellar keratectomy, and corneal perforation, managed by penetrating keratoplasty.


A systematic review of the literature examining the diagnostic efficacy of measurement of fractionated plasma free metanephrines in the biochemical diagnosis of pheochromocytoma.
BACKGROUND: Fractionated plasma metanephrine measurements are commonly used in biochemical testing in search of pheochromocytoma. METHODS: We aimed to critically appraise the diagnostic efficacy of fractionated plasma free metanephrine measurements in detecting pheochromocytoma. Nine electronic databases, meeting abstracts, and the Science Citation Index were searched and supplemented with previously unpublished data. Methodologic and reporting quality was independently assessed by two endocrinologists using a checklist developed by the Standards for Reporting of Diagnostic Studies Accuracy Group and data were independently abstracted. RESULTS: Limitations in methodologic quality were noted in all studies. In all subjects (including those with genetic predisposition): the sensitivities for detection of pheochromocytoma were 96%-100% (95% CI ranged from 82% to 100%), whereas the specificities were 85%-100% (95% CI ranged from 78% to 100%). Statistical heterogeneity was noted upon pooling positive likelihood ratios when those with predisposition to disease were included (p < 0.001). However, upon pooling the positive or negative likelihood ratios for patients with sporadic pheochromocytoma (n = 191) or those at risk for sporadic pheochromocytoma (n = 718), no statistical heterogeneity was noted (p = 0.4). For sporadic subjects, the pooled positive likelihood ratio was 5.77 (95% CI = 4.90, 6.81) and the pooled negative likelihood ratio was 0.02 (95% CI = 0.01, 0.07). CONCLUSION: Negative plasma fractionated free metanephrine measurements are effective in ruling out pheochromocytoma. However, a positive test result only moderately increases suspicion of disease, particularly when screening for sporadic pheochromocytoma.


Insulin resistance determines phagocytic nicotinamide adenine dinucleotide phosphate oxidase overactivation in metabolic syndrome patients.
Metabolic syndrome (MetS) is associated with insulin resistance and increases the cardiovascular risk. Oxidative stress constitutes a potential mechanism that links insulin resistance and cardiovascular disease. The aim of this study was to analyze the relationship of NADPH oxidase activation with insulin resistance, and the effect of this interaction on the cardiovascular risk in MetS patients. NADPH oxidase-dependent superoxide production and expression was evaluated by luminescence and western blot, respectively, in peripheral blood mononuclear cells obtained from 125 patients with MetS. Insulin resistance was defined by the homeostasis model assessment index. Matrix metalloproteinase-9 was quantified by enzyme-linked immunosorbent assay in plasma samples. To ascertain the mechanisms involved in vivo, we performed in-vitro experiments in cultured macrophages. Fifty-six percent of patients with MetS showed insulin resistance. Plasma matrix metalloproteinase-9 levels were higher (P < 0.05) in insulin-resistant patients than in patients with insulin sensitivity. NADPH oxidase-dependent superoxide production was augmented (P < 0.05) in insulin-resistant patients with respect to insulin-sensitive patients. The interaction between insulin resistance and abnormally high NADPH oxidase-mediated superoxide production was associated with the highest matrix metalloproteinase-9 values. Increased NADPH oxidase-dependent superoxide production was significantly associated with higher NADPH oxidase p22phox expression in insulin-resistant than in insulin-sensitive patients. Interestingly, insulin upregulated p22phox in peripheral blood mononuclear cells and in murine macrophages. Insulin resistance is associated with phagocytic NADPH oxidase activation. This association results in the highest cardiovascular risk in MetS patients.


It looks like construct validity, but look again: Comment on Clutterbuck et al. (2021) and recommendations for test developers in the broad "empathy" domain.
Conceptual flaws can undermine even rigorous test development efforts, especially in the broad empathy and social cognition domains, which are characterized by measure proliferation and inconsistently used construct terms. We discuss these issues, focusing on a new instrument of "mentalizing" as a case study. Across several studies, Clutterbuck et al. (2021a) developed the Four-Item Mentalising Index (FIMI). They described it as the first self-report measure of mentalizing ability and suggested that it offers substantial advances for research and assessment. As we demonstrate with conceptual arguments and empirical data, the FIMI embodies several major problems that are common in this area of research. Using the FIMI as a case study, we underline the importance for test developers of attending to the nonnegotiable necessity of discriminant validity analyses, the challenge of choosing appropriate convergent validity measures, and the difficulties of navigating the jingle-jangle jungle of empathy and social cognition construct terms. (PsycInfo Database Record (c) 2022 APA, all rights reserved).


Real-world use and modeled impact of glucose-lowering therapies evaluated in recent cardiovascular outcomes trials: An NCDR® Research to Practice project.
Aims Recent trials (EMPA-REG OUTCOME and Liraglutide Effect and Action in Diabetes: Evaluation of Cardiovascular Outcome Results [LEADER]) have shown improved cardiovascular (CV) mortality with specific currently available glucose-lowering medications (empagliflozin and liraglutide, respectively), but were limited to selected patient populations. We sought to evaluate the current use and potential real-world impact of empagliflozin (and other sodium-glucose co-transporter 2 inhibitors [SGLT2is]) and liraglutide (and other glucagonlike peptide-1 receptor agonist [GLP-1 RAs]) among patients in the Diabetes Collaborative Registry (DCR). Methods and results We evaluated 182,525 patients from the DCR - a large, US-based outpatient registry of individuals with type 2 diabetes from 313 sites that included cardiology, endocrinology and primary care practices. Among these patients, 26.2% met major eligibility criteria for EMPA-REG OUTCOME and 48.0% met major eligibility criteria for LEADER. Of these potentially eligible patients, only a small minority were actually prescribed these agents: 5.2% on an SGLT2i and 6.0% on a GLP-1 RA, respectively. Patients receiving these studied medications or medication classes, in general, had lower CV disease burden compared with those not on these agents. Assuming similar risk reductions as in the clinical trials, if all potentially trial-eligible patients in the DCR were treated for 1 year with empagliflozin (or other SGLT2is, assuming a class effect) or liraglutide (or other GLP-1 RAs, assuming a class effect), this may have prevented 354 CV deaths, 231 heart failure hospitalizations, 329 CV deaths and 247 myocardial infarctions, respectively. Conclusion In a large, US-based outpatient registry, we found a significant number of patients would have been potentially eligible for glucose-lowering agents that demonstrated CV benefit in recent clinical trials. In view of these findings, a broader and better-targeted use of these medications in evidence-based patient populations should be considered.


Population-specific modelling of between/within-subject flow variability in the carotid arteries of the elderly.
Computational fluid dynamics models are increasingly proposed for assisting the diagnosis and management of vascular diseases. Ideally, patient-specific flow measurements are used to impose flow boundary conditions. When patient-specific flow measurements are unavailable, mean values of flow measurements across small cohorts are used as normative values. In reality, both the between-subjects and within-subject flow variabilities are large. Consequently, neither one-shot flow measurements nor mean values across a cohort are truly indicative of the flow regime in a given person. We develop models for both the between-subjects and within-subject variability of internal carotid flow. A log-linear mixed effects model is combined with a Gaussian process to model the between-subjects flow variability, while a lumped parameter model of cerebral autoregulation is used to model the within-subject flow variability in response to heart rate and blood pressure changes. The model parameters are identified from carotid ultrasound measurements in a cohort of 103 elderly volunteers. We use the models to study intracranial aneurysm flow in 54 subjects under rest and exercise and conclude that OSI, a common wall shear-stress derived quantity in vascular CFD studies, may be too sensitive to flow fluctuations to be a reliable biomarker.


Three new DNA helicases from Saccharomyces cerevisiae.
At least six DNA helicases have been identified during fractionation of extracts from the yeast Saccharomyces cerevisiae. Three of those, DNA helicases B, C, and D, have been further purified and characterized. DNA helicases B and C co-purified with DNA polymerase delta through several chromatographic steps, but were separated from the polymerase by hydrophobic chromatography. DNA helicase D co-purified with Replication Factor C over seven chromatographic steps, and was only separated from it by glycerol gradient centrifugation in the presence of 0.2 M NaCl. All three helicases are DNA dependent ATPases with Km values for ATP of 190 microM, 325 microM, and 60 microM for DNA helicases B, C, and D, respectively. Their DNA helicase activities are comparable. They are 5'-3' helicases and have pH optima of 6.5-7 and Mg2+ optima of 1-2 mM. However, they differ in the nucleotide requirement for helicase action. Whereas all three helicases preferred ATP, dATP, UTP, CTP, and dCTP as cofactors, DNA helicase C also used GTP, but not dTTP. On the other hand, DNA helicase D used dTTP, but not GTP, and DNA helicase B used neither nucleotide as cofactor. These studies allowed us to conclude that DNA helicases B, C, and D are not only distinct enzymes, but also different from two previously identified yeast DNA helicases, the RAD3 protein and ATPase III.


Perceived barriers and facilitators to mental health help-seeking in young people: a systematic review.
Adolescents and young adults frequently experience mental disorders, yet tend not to seek help. This systematic review aims to summarise reported barriers and facilitators of help-seeking in young people using both qualitative research from surveys, focus groups, and interviews and quantitative data from published surveys. It extends previous reviews through its systematic research methodology and by the inclusion of published studies describing what young people themselves perceive are the barriers and facilitators to help-seeking for common mental health problems. Twenty two published studies of perceived barriers or facilitators in adolescents or young adults were identified through searches of PubMed, PsycInfo, and the Cochrane database. A thematic analysis was undertaken on the results reported in the qualitative literature and quantitative literature. Fifteen qualitative and seven quantitative studies were identified. Young people perceived stigma and embarrassment, problems recognising symptoms (poor mental health literacy), and a preference for self-reliance as the most important barriers to help-seeking. Facilitators were comparatively under-researched. However, there was evidence that young people perceived positive past experiences, and social support and encouragement from others as aids to the help-seeking process. Strategies for improving help-seeking by adolescents and young adults should focus on improving mental health literacy, reducing stigma, and taking into account the desire of young people for self-reliance.


Fertility preservation and breast cancer: a review.
Breast cancer is the most common malignancy in women, and its incidence increases with age, with the majority of patients diagnosed after menopause. However, in 15-25% of cases, patients are premenopausal at the time of diagnosis, and about 7% of them are below the age of 40. Therefore, a considerable amount of young women are diagnosed with breast cancer during their reproductive life. Within this group, most cancer cases require cytotoxic chemotherapy and/or hormone therapy, which are responsible for a decrease in the patients' reproductive function, along with their age. The efficacy of such treatments, among other factors, has led to a high five-year-survival rate, which results in an increasing number of young women who survive breast cancer before having fulfilled their reproductive wishes, especially considering the current trend to delay pregnancy until the late 30s or early 40s in developed countries. The combination of these factors justifies the importance of fertility preservation and reproductive counselling at the time of breast cancer diagnosis in young women. A wide range of fertility preservation techniques has been developed, such as ovarian suppression, oocyte and embryo cryopreservation, immature oocyte retrieval and in vitro maturation, and ovarian tissue cryopreservation. Early counselling and referral of these patients to fertility specialists are fundamental factors in order to maximise their chances of pregnancy. This review aims to update the knowledge about the influence of breast cancer in fertility, the influence of pregnancy and fertility preservation techniques in breast cancer patients and assessment of ovarian reserve for a better treatment choice. A special section dedicated to BRCA-mutation carriers has been included because of their specific features. A comprehensive literature search has been conducted, including publications from the last five years.


Detection of MCT and MCTC types of human mast cells by immunohistochemistry using new monoclonal anti-tryptase and anti-chymase antibodies.
We developed an improved immunohistochemical technique for distinguishing human mast cells of the MCT (tryptase-positive, chymase-negative) and MCTC (tryptase-positive, chymase-positive) types utilizing a biotinylated murine anti-chymase monoclonal antibody (MAb), termed B7, and an alkaline phosphatase-conjugated murine anti-tryptase MAb, termed G3. The B7 MAb also was used to show the selective presence of chymase in mast cells. The distribution of MCT and MCTC cells in Carnoy's fluid-fixed tissue sections of human lung, skin, small intestine, and tonsils was analyzed by the new technique and the results compared to those obtained with the older method using a rabbit polyclonal antichymase antibody and a mouse anti-tryptase MAb in indirect immunoperoxidase and indirect immunoalkaline phosphatase protocols, respectively. In tissues known to contain predominantly mature mast cells, there were no quantitative differences between the two techniques, although the staining intensity achieved with the anti-chymase MAb was greater and without development of high background, compared to results achieved with the polyclonal antibody. MCT cells were the predominant type seen in the alveoli of the lung (93%) and in the small intestinal mucosa (81%). MCTC cells predominanted in the skin (99%) and in the small intestinal submucosa (77%) and, to a lesser degree, in tonsils (60%). However, in newborn foreskin tissue which contains predominantly immature forms of mast cells, 75% of all mast cells were stained uniformly and intensely with B7, whereas only 43% were stained with the polyclonal anti-chymase antibody. Therefore, the use of MAb provides for better standardization of reagents and more accurate assessment of the distribution of human MCT and MCTC cells in tissues than previously available methods.


Investigation of the role of typhoid toxin in acute typhoid fever in a human challenge model.
Salmonella Typhi is a human host-restricted pathogen that is responsible for typhoid fever in approximately 10.9 million people annually1. The typhoid toxin is postulated to have a central role in disease pathogenesis, the establishment of chronic infection and human host restriction2-6. However, its precise role in typhoid disease in humans is not fully defined. We studied the role of typhoid toxin in acute infection using a randomized, double-blind S. Typhi human challenge model7. Forty healthy volunteers were randomized (1:1) to oral challenge with 104 colony-forming units of wild-type or an isogenic typhoid toxin deletion mutant (TN) of S. Typhi. We observed no significant difference in the rate of typhoid infection (fever ≥38 °C for ≥12 h and/or S. Typhi bacteremia) between participants challenged with wild-type or TN S. Typhi (15 out of 21 (71%) versus 15 out of 19 (79%); P = 0.58). The duration of bacteremia was significantly longer in participants challenged with the TN strain compared with wild-type (47.6 hours (28.9-97.0) versus 30.3(3.6-49.4); P ≤ 0.001). The clinical syndrome was otherwise indistinguishable between wild-type and TN groups. These data suggest that the typhoid toxin is not required for infection and the development of early typhoid fever symptoms within the context of a human challenge model. Further clinical data are required to assess the role of typhoid toxin in severe disease or the establishment of bacterial carriage.


Laparoscopic treatment of hepatic hydatid cysts with a liposuction device.
We describe herein a surgical technique, whereby we use a liposuction device for the laparoscopic treatment of hepatic hydatid cysts (HHC). Ten patients with 12 hepatic hydatid cysts were treated with this technique. All patients received pre- and postoperative antiscolecidal medications. The laparoscopic technique consisted of partial aspiration of the cyst fluid and replacement of the aspirated fluid with 10% Betadine. The Betadine solution was left in situ for 10 minutes. Evacuation of the cyst contents was carried out with the liposuction device. The residual cavity was unroofed by partial excision of the ectocyst. A drain was left alongside the cyst. No intra- or postoperative complications were encountered. All patients were mobilized freely, were allowed to eat a regular meal 6 hours after recovery from anesthesia, and were discharged on the third postoperative day. All patients resumed their normal household and work activities by the tenth postoperative day. The patients were regularly followed up every 2 months for 2 years. At follow-up in the surgical clinic, no evidence of recurrence was noted either clinically, serologically, or by imaging techniques. We conclude that the laparoscopic treatment of HHC is feasible and advantageous. We believe that the use of a liposuction device facilitates rapid and efficient evacuation of the viscid organic contents of the cyst and helps in the obliteration of the residual cavity.


Examining levels of risk behaviors among black men who have sex with Men (MSM) and the association with HIV acquisition.
Seroadaptation is defined as the practice of modifying sexual behavior based on one's own HIV serostatus, the perceived HIV serostatus of sexual partners, and differences in risk of HIV transmission by sexual acts. Because this definition implies intent, we use the term "seroprotection" to describe HIV negative participants reporting condomless anal sex (CAS) either exclusively with seronegative partners, or only as the insertive partner with HIV positive or unknown serostatus partners. Little is known about seroprotection in Black men who have sex with men (MSM). We evaluated the independent association of seroprotection and HIV acquisition among the 1144 HIV-negative Black MSM enrolled in HPTN 061 using Cox models; we stratified by city of enrollment, and controlled for number of partners, age, and drug use. Behaviors reported at 0, 6, and 12 months were assigned to three mutually exclusive categories: (1) No CAS; (2) Seroprotection; and (3) CAS without seroprotection. In 2,861 six-month intervals; 28 HIV seroconversions occurred. No CAS was reported at 33.3% of visits, seroprotection at 46.6% of visits, and CAS without seroprotection at 20.1% of visits. The seroconversion rate per 100 person-years for no CAS was 0.98 (95% CI: 0.27, 2.51), compared with 2.39 (95% CI: 1.03, 4.71) and 13.33 (95% CI: 7.62, 21.66) for seroprotection and CAS without seroprotection, respectively. Compared to CAS without seroprotection, intervals without CAS were associated with an 87% reduction (aHR: 0.13, 95% CI: 0.03-0.46) in HIV acquisition and intervals with seroprotection with a 78% reduction (aHR: 0.22, 95% CI: 0.09-0.57). No CAS is the safest behavior to prevent HIV acquisition. Seroprotective behaviors significantly reduced risk, but HIV incidence was still >2/100 person-years, suggesting that additional strategies, such as pre-exposure prophylaxis, are warranted for this population.


Combined heterogeneous distribution of salt and aroma in food enhances salt perception.
Aroma-taste interactions and heterogeneous spatial distribution of tastants were used as strategies for taste enhancement. This study investigated the combination of these two strategies through the effect of heterogeneous salt and aroma distribution on saltiness enhancement and consumer liking for hot snacks. Four-layered cream-based products were designed with the same total amount of sodium and ham aroma but varied in their spatial distribution. Unflavoured products containing the same amount of salt and 35% more salt were used as references. A consumer panel (n = 82) rated the intensity of salty, sweet, sour, bitter and umami tastes as well as ham and cheese aroma intensity for each product. The consumers also rated their liking for the products in a dedicated sensory session. The results showed that adding salt-associated aroma (ham) led to enhancement of salty taste perception regardless of the spatial distribution of salt and aroma. Moreover, products with a higher heterogeneity of salt distribution were perceived as saltier (p < 0.01), whereas heterogeneity of ham aroma distribution had only a marginal effect on both aroma and salty taste perception. Furthermore, heterogeneous products were well liked by consumers compared to the homogeneous products.


A model of psychosocial hypertension showing reversibility and progression of cardiovascular complications.
The sequence of pathophysiological changes that can result from the stimulating effects of a sustained disturbance of the social environment was studied in ten colonies of socially deprived mice. Sixteen formerly isolated males were placed with 16 normal females in population cages consisting of seven intercommunicating boxes. Six of these socially disturbed 32-member colonies were terminated after periods of interaction ranging from 2 days to 9 months. The remaining four were terminated a month or more after the males had been returned to individual isolation. Indirect blood pressure measurements, body and heart weights, and sections of hearts and aortas were studied in the males. Following the shorter exposures, blood pressure reverted to normal in a few days. Exposure of 6 months or more were associated with unchanged body weights and sustained increases in heart weight and blood pressure readings. In addition, there was a significant development of aortic arteriosclerosis and myocardial fibrosis. These changes persisted despite prolonged return to isolation.


Investigating liver stiffness and viscosity for fibrosis, steatosis and activity staging using shear wave elastography.
Quantitative shear wave elastography was shown to be an effective tool for the non-invasive diagnosis and staging of chronic liver diseases. The liver shear modulus, estimated from the propagation velocity of shear waves, is correlated to the degree of fibrosis and can therefore be used for the non-invasive staging of fibrosis. We performed a clinical prospective study in a total of 120 patients with various chronic liver diseases to compare the accuracy of supersonic shear imaging (SSI), a technique based on acoustic radiation and ultrafast ultrasound imaging, to 1D transient elastography (FibroScan) for the staging and grading of fibrosis as assessed by liver biopsy. Since shear wave propagation spectroscopy can also provide additional mechanical information on soft tissues, such as viscosity, we also investigated those new mechanical parameters as possible predictors of fibrosis, steatosis, and disease activity. SSI was successfully performed in 98.3% of patients and it was shown to be as accurate as FibroScan for the staging of fibrosis both for the whole population (N=120) and for the subgroup with viral hepatitis (n=70) (AUC=0.85 [0.77-0.96] and 0.89 [0.81-0.97] for significant fibrosis, AUC=0.90 [0.83-0.97] and 0.87 [0.75-0.98] for cirrhosis, with respect to SSI [n=68/70] and FibroScan [n=66/68]). Viscosity could also be used to stage the degree of fibrosis (AUC=0.76 [0.64-0.87] for significant fibrosis and AUC=0.87 [0.74-0.99] for cirrhosis), for the subgroup of patients with viral hepatitis (n=67/70) but was a poor predictor of disease activity and steatosis levels. Supersonic shear imaging is a robust technique for the staging of liver fibrosis. Liver viscosity was found to be correlated with fibrosis but not to steatosis or disease activity.


Decoding Three Different Preference Levels of Consumers Using Convolutional Neural Network: A Functional Near-Infrared Spectroscopy Study.
This study decodes consumers' preference levels using a convolutional neural network (CNN) in neuromarketing. The classification accuracy in neuromarketing is a critical factor in evaluating the intentions of the consumers. Functional near-infrared spectroscopy (fNIRS) is utilized as a neuroimaging modality to measure the cerebral hemodynamic responses. In this study, a specific decoding structure, called CNN-based fNIRS-data analysis, was designed to achieve a high classification accuracy. Compared to other methods, the automated characteristics, constant training of the dataset, and learning efficiency of the proposed method are the main advantages. The experimental procedure required eight healthy participants (four female and four male) to view commercial advertisement videos of different durations (15, 30, and 60 s). The cerebral hemodynamic responses of the participants were measured. To compare the preference classification performances, CNN was utilized to extract the most common features, including the mean, peak, variance, kurtosis, and skewness. Considering three video durations, the average classification accuracies of 15, 30, and 60 s videos were 84.3, 87.9, and 86.4%, respectively. Among them, the classification accuracy of 87.9% for 30 s videos was the highest. The average classification accuracies of three preferences in females and males were 86.2 and 86.3%, respectively, showing no difference in each group. By comparing the classification performances in three different combinations (like vs. so-so, like vs. dislike, and so-so vs. dislike) between two groups, male participants were observed to have targeted preferences for commercial advertising, and the classification performance 88.4% between "like" vs. "dislike" out of three categories was the highest. Finally, pairwise classification performance are shown as follows: For female, 86.1% (like vs. so-so), 87.4% (like vs. dislike), 85.2% (so-so vs. dislike), and for male 85.7, 88.4, 85.1%, respectively.


Left cervical aortic arch with proximal obstruction.
Cervical aortic arch is a rare congenital anomaly. We present a case of left-sided cervical aortic arch, found in a patient with mitral regurgitation, mitral stenosis, and a regurgitant bicuspid aortic valve. There was atypical obstruction proximally within the arch. The obstructive segment was resected, and corrected by performing an end-to-side anostomosis. Mitral valvoplasty was performed in the same surgical procedure. To the best of our knowledge, a cervical arch has not previously been described with such atypical obstruction, and in association with multiple lesions involving the left heart.


Reduced lymphocyte transformation in early cancer of the breast.
The cell mediated immune response has been measured in vitro by lymphocyte transformation in 53 patients with malignant tumours of the breast and an equal number of patients in the same age group operated upon for benign tumours. The response to phytohaemagglutinin (PHA) was reduced in the patients with malignant tumours when the lymphocytes were grown either in the patient's plasma or in autologous plasma. This was observed when the response was measured both by the uptake of radioactive DNA precursor and by autoradiography. Reduced lymphocyte transformation was present even very early in the disease and certainly preoperatively. The reduction appeared to be even more marked in the advanced cases. An interesting finding was that the plasma from cancer patients contained a factor which reduced the PHA transformation of lymphocytes from a healthy donor.


Influence of fiber manufacturing tolerances on the spectral response of a bend loss based all-fiber edge filter.
It is shown that manufacturing tolerances of the fiber parameters bend radius and numerical aperture (NA) significantly influence the fiber bend loss performance and spectral response of a fiber based edge filter. A theoretical model, validated by experimental results, is used to determine the changes in key spectral parameters for an edge filter, resulting from changes within their manufacturing tolerance range, for both the bend radius and NA. Finally, it is shown that bend-radius tuning during fabrication of such filters is a means of mitigating the effect of manufacturing variations.


Effect of progesterone and its synthetic analogs on reproduction and embryonic development of a freshwater invertebrate model.
The presence of a mixture of progestogens at ng/L concentration levels in surface waters is a worldwide problem. Only a few studies explore the effect of progestogen treatment in a mixture as opposed to individual chemicals to shed light on how non-target species respond to these contaminants. In the present study, we used an invertebrate model species, Lymnaea stagnalis, exposed to a mixture of four progestogens (progesterone, levonorgestrel, drospirenone, and gestodene) in 10ng/L concentration for 3 weeks. Data at both physiological and cellular/molecular level were analyzed using the ELISA technique, stereomicroscopy combined with time lapse software, and capillary microsampling combined with mass spectrometry. The treatment of adult Lymnaeas caused reduced egg production, and low quality egg mass on the first week, compared to the control. Starting from the second week, the egg production, and the quality of egg mass were similar in both groups. At the end of the third week, the egg production and the vitellogenin-like protein content of the hepatopancreas were significantly elevated in the treated group. At the cellular level, accelerated cell proliferation was observed during early embryogenesis in the treated group. The investigation of metabolomic changes resulted significantly elevated hexose utilization in the single-cell zygote cytoplasm, and elevated adenylate energy charge in the egg albumen. These changes suggested that treated snails provided more hexose in the eggs in order to improve offspring viability. Our study contributes to the knowledge of physiological effect of equi-concentration progestogen mixture at environmentally relevant dose on non-target aquatic species.


All-aqueous multiphase microfluidics.
Immiscible aqueous phases, formed by dissolving incompatible solutes in water, have been used in green chemical synthesis, molecular extraction and mimicking of cellular cytoplasm. Recently, a microfluidic approach has been introduced to generate all-aqueous emulsions and jets based on these immiscible aqueous phases; due to their biocompatibility, these all-aqueous structures have shown great promises as templates for fabricating biomaterials. The physico-chemical nature of interfaces between two immiscible aqueous phases leads to unique interfacial properties, such as an ultra-low interfacial tension. Strategies to manipulate components and direct their assembly at these interfaces needs to be explored. In this paper, we review progress on the topic over the past few years, with a focus on the fabrication and stabilization of all-aqueous structures in a multiphase microfluidic platform. We also discuss future efforts needed from the perspectives of fluidic physics, materials engineering, and biology for fulfilling potential applications ranging from materials fabrication to biomedical engineering.


Toxicity Minimized Cryoprotectant Addition and Removal Procedures for Adherent Endothelial Cells.
Ice-free cryopreservation, known as vitrification, is an appealing approach for banking of adherent cells and tissues because it prevents dissociation and morphological damage that may result from ice crystal formation. However, current vitrification methods are often limited by the cytotoxicity of the concentrated cryoprotective agent (CPA) solutions that are required to suppress ice formation. Recently, we described a mathematical strategy for identifying minimally toxic CPA equilibration procedures based on the minimization of a toxicity cost function. Here we provide direct experimental support for the feasibility of these methods when applied to adherent endothelial cells. We first developed a concentration- and temperature-dependent toxicity cost function by exposing the cells to a range of glycerol concentrations at 21°C and 37°C, and fitting the resulting viability data to a first order cell death model. This cost function was then numerically minimized in our state constrained optimization routine to determine addition and removal procedures for 17 molal (mol/kg water) glycerol solutions. Using these predicted optimal procedures, we obtained 81% recovery after exposure to vitrification solutions, as well as successful vitrification with the relatively slow cooling and warming rates of 50°C/min and 130°C/min. In comparison, conventional multistep CPA equilibration procedures resulted in much lower cell yields of about 10%. Our results demonstrate the potential for rational design of minimally toxic vitrification procedures and pave the way for extension of our optimization approach to other adherent cell types as well as more complex systems such as tissues and organs.


Artificial Intelligence-Based 3D Angiography for Visualization of Complex Cerebrovascular Pathologies.
By means of artificial intelligence, 3D angiography is a novel postprocessing method for 3D imaging of cerebral vessels. Because 3D angiography does not require a mask run like the current standard 3D-DSA, it potentially offers a considerable reduction of the patient radiation dose. Our aim was an assessment of the diagnostic value of 3D angiography for visualization of cerebrovascular pathologies. 3D-DSA data sets of cerebral aneurysms (n CA = 10), AVMs (n AVM = 10), and dural arteriovenous fistulas (dAVFs) (n dAVF = 10) were reconstructed using both conventional and prototype software. Corresponding reconstructions have been analyzed by 2 neuroradiologists in a consensus reading in terms of image quality, injection vessel diameters (vessel diameter [VD] 1/2), vessel geometry index (VGI = VD1/VD2), and specific qualitative/quantitative parameters of AVMs (eg, location, nidus size, feeder, associated aneurysms, drainage, Spetzler-Martin score), dAVFs (eg, fistulous point, main feeder, diameter of the main feeder, drainage), and cerebral aneurysms (location, neck, size). In total, 60 volumes have been successfully reconstructed with equivalent image quality. The specific qualitative/quantitative assessment of 3D angiography revealed nearly complete accordance with 3D-DSA in AVMs (eg, mean nidus size3D angiography/3D-DSA= 19.9 [SD, 10.9]/20.2 [SD, 11.2] mm; r = 0.9, P = .001), dAVFs (eg, mean diameter of the main feeder3D angiography/3D-DSA= 2.04 [SD, 0.65]/2.05 [SD, 0.63] mm; r = 0.9, P = .001), and cerebral aneurysms (eg, mean size3D angiography/3D-DSA= 5.17 [SD, 3.4]/5.12 [SD, 3.3] mm; r = 0.9, P = .001). Assessment of the geometry of the injection vessel in 3D angiography data sets did not differ significantly from that of 3D-DSA (vessel geometry indexAVM: r = 0.84, P = .003; vessel geometry indexdAVF: r = 0.82, P = .003; vessel geometry indexCA: r = 0.84, P <.001). In this study, the artificial intelligence-based 3D angiography was a reliable method for visualization of complex cerebrovascular pathologies and showed results comparable with those of 3D-DSA. Thus, 3D angiography is a promising postprocessing method that provides a significant reduction of the patient radiation dose.


Collaborative practice in a global health context: Common themes from developed and developing countries.
This paper reports on a study commissioned by the World Health Organization (WHO) to explore common themes of collaborative practice. The WHO requested global clarification of (1) the nature of collaborative practice, (2) its perceived importance, and (3) strategies for systematizing collaborative practice throughout national health systems. While there are many interpretations of collaborative practice around the world, there was a need to ascertain common underlying themes that illustrate good practice in both developed and developing countries to inform an international Framework for Action. A multiple case study design was used to examine collaborative practice in primary health care and commonalities across countries. Staff at each of WHO's six regional offices invited key informants in one or two primary health care organizations where collaborative practice was the model of care to complete case studies. Ten case studies were received from ten different countries, representing all six WHO regions. The results are described according to the study's three areas of focus: describing collaborative practice globally, the shared importance of collaborative practice, and systematizing collaborative practice. Collaborative practice requires a strong political framework that encourages interprofessional education and teamworking. Shared governance models and enabling legislation are required. At a practical level, interprofessional health care teams function most efficiently with shared clinical pathways and a common patient record.


Overcoming roadblocks in the development of vaccines for leishmaniasis.
The leishmaniases represent a group of parasitic diseases caused by infection with one of several species of Leishmania parasites. Disease presentation varies because of differences in parasite and host genetics and may be influenced by additional factors such as host nutritional status or co-infection. Studies in experimental models of Leishmania infection, vaccination of companion animals and human epidemiological data suggest that many forms of leishmaniasis could be prevented by vaccination, but no vaccines are currently available for human use. We describe some of the existing roadblocks to the development and implementation of an effective leishmaniasis vaccine, based on a review of recent literature found on PubMed, BioRxiv and MedRxiv. In addition to discussing scientific unknowns that hinder vaccine candidate identification and selection, we explore gaps in knowledge regarding the commercial and public health value propositions underpinning vaccine development and provide a route map for future research and advocacy. Despite significant progress, leishmaniasis vaccine development remains hindered by significant gaps in understanding that span the vaccine development pipeline. Increased coordination and adoption of a more holistic view to vaccine development will be required to ensure more rapid progress in the years ahead.


One-dimensional diamondoid polyaniline-like nanothreads from compressed crystal aniline.
Low-dimensional nanomaterials such as highly ordered polyaniline (PANI) have attracted considerable interest due to their expected extraordinary electronic and optoelectronic properties. In spite of several attempts, the attainment of atomically well-ordered PANI is a long-standing challenge. Pressure-induced polymerization of aromatic molecules in the crystal phase has been demonstrated as a practicable route for the synthesis of highly ordered polymers but this approach has never been tested to produce PANI. Here we show the synthesis of diamondoid polyaniline-like nanothreads at 33 GPa and 550 K by compressing aniline in crystal phase-II. Infrared spectroscopy, transmission electron microscopy, X-ray diffraction data, and DFT calculations support the formation of this totally new polyaniline-like nanothread. The NH2-enriched carbon nanothread combines the outstanding mechanical properties of carbon nanotubes with the versatility of NH2 groups decorating the exterior of the nanothreads representing potential active sites for doping and as linkers for molecules with biological interest and inorganic nanostructures. The synergy of all of these properties emphasizes the strong potential of this material to be applied in a broad range of areas, from chemistry to materials engineering.


Drug-Drug Interactions (DDIs) in Psychiatric Practice, Part 9: Interactions Mediated by Drug-metabolizing Cytochrome P450 Enzymes.
This column is the ninth in a series exploring drug-drug interactions (DDIs) with a special emphasis on psychiatric medications. The first 3 columns in this DDI series discussed why patients being treated with psychiatric medications are at increased risk for taking multiple medications and thus experiencing DDIs, how to recognize such DDIs, strategies for avoiding and/or minimizing adverse outcomes from such DDIs, and pharmacokinetic considerations concerning DDIs in psychiatric practice. The fourth and fifth columns in this series presented a pair of parallel tables, one of which outlined the primary, known mechanism(s) of action of all commonly used psychiatric medications and one of which summarized major types of pharmaco-dynamic DDIs based on mechanism of action. Clinicians can use these 2 tables together to predict pharmacodynamically mediated DDIs. The sixth column discussed key pharmacodynamic interactions involving ethanol, opioids, and monoamine oxidase inhibitors. The seventh and eighth columns presented the concept of relative receptor binding and included tables summarizing the relative receptor binding affinity of currently available antipsychotics and antidepressants, respectively. This ninth and final column in this series discusses pharmacokinetic DDIs with a focus on psychiatric medications and contains 3 tables. The first table is an abbreviated version of a table available online showing which drugs are substrates for which cytochrome P450 (CYP) enzymes and which drugs are inhibitors or inducers of specific CYP enzymes. The abbreviated version of the table presented in this column focuses on psychiatric medications. This table and the larger website version can allow prescribers to anticipate which drug combinations may pose the risk of a CYP enzyme-mediated DDI. The second table summarizes which antidepressants inhibit specific CYP enzymes and which antidepressants do not or are unlikely to inhibit specific CYP enzymes. The third table presents psychiatric medications whose clearance is not principally dependent on CYP enzyme-mediated oxidative metabolism as a necessary step in their clearance from the body. The latter 2 tables inform prescribers as to which drugs they may prefer to use to avoid CYP enzyme-mediated DDIs. The overall goal of this series of columns is to present a simple way of conceptualizing neuropsychiatric medications in terms of their pharmacodynamics and pharmacokinetics to allow prescribers to take these facts into consideration when they need to use ≥2 drugs in combination to optimally treat a patient.


The Inhibitory Effects of Static Magnetic Field on Escherichia coli from two Different Sources at Short Exposure Time.
This study was intended to investigate the effectiveness of static magnetic field on the growth of Escherichia coli (E. coli) provided from two sources, the urine samples of patients with urinary tract infections and the reference strain E. coli ATCC 25922. Bacterial samples in Nutrient Broth were subjected to a range of magnetic intensities (2, 4, 6, 9, 14, 16, 18, and 20 mT) at various exposure times (0, 15, 30, 45, 60, 75, and 90 min). The survival rate was measured in the presence and absence of the magnetic field over time. The cell counts of uropathogenic E. coli did not statistically differed from those of the standard strain if exposed to the magnetic field. The fluctuation was observed in cell viabilities at different magnetic intensities below 18 mT. Both groups presented a significant decline in survival rate as exposed to 18 and 20 mT.


Dual MET-EGFR combinatorial inhibition against T790M-EGFR-mediated erlotinib-resistant lung cancer.
Despite clinical approval of erlotinib, most advanced lung cancer patients are primary non-responders. Initial responders invariably develop secondary resistance, which can be accounted for by T790M-EGFR mutation in half of the relapses. We show that MET is highly expressed in lung cancer, often concomitantly with epidermal growth factor receptor (EGFR), including H1975 cell line. The erlotinib-resistant lung cancer cell line H1975, which expresses L858R/T790M-EGFR in-cis, was used to test for the effect of MET inhibition using the small molecule inhibitor SU11274. H1975 cells express wild-type MET, without genomic amplification (CNV = 1.1). At 2 microM, SU 11274 had significant in vitro pro-apoptotic effect in H1975 cells, 3.9-fold (P = 0.0015) higher than erlotinib, but had no effect on the MET and EGFR-negative H520 cells. In vivo, SU11274 also induced significant tumour cytoreduction in H1975 murine xenografts in our bioluminescence molecular imaging assay. Using small-animal microPET/MRI, SU11274 treatment was found to induce an early tumour metabolic response in H1975 tumour xenografts. MET and EGFR pathways were found to exhibit collaborative signalling with receptor cross-activation, which had different patterns between wild type (A549) and L858R/T790M-EGFR (H1975). SU11274 plus erlotinib/CL-387,785 potentiated MET inhibition of downstream cell proliferative survival signalling. Knockdown studies in H1975 cells using siRNA against MET alone, EGFR alone, or both, confirmed the enhanced downstream inhibition with dual MET-EGFR signal path inhibition. Finally, in our time-lapse video-microscopy and in vivo multimodal molecular imaging studies, dual SU11274-erlotinib concurrent treatment effectively inhibited H1975 cells with enhanced abrogation of cytoskeletal functions and complete regression of the xenograft growth. Together, our results suggest that MET-based targeted inhibition using small-molecule MET inhibitor can be a potential treatment strategy for T790M-EGFR-mediated erlotinib-resistant non-small-cell lung cancer. Furthermore, optimised inhibition may be further achieved with MET inhibition in combination with erlotinib or an irreversible EGFR-TKI.


Comparison of clustering methods for high-dimensional single-cell flow and mass cytometry data.
Recent technological developments in high-dimensional flow cytometry and mass cytometry (CyTOF) have made it possible to detect expression levels of dozens of protein markers in thousands of cells per second, allowing cell populations to be characterized in unprecedented detail. Traditional data analysis by "manual gating" can be inefficient and unreliable in these high-dimensional settings, which has led to the development of a large number of automated analysis methods. Methods designed for unsupervised analysis use specialized clustering algorithms to detect and define cell populations for further downstream analysis. Here, we have performed an up-to-date, extensible performance comparison of clustering methods for high-dimensional flow and mass cytometry data. We evaluated methods using several publicly available data sets from experiments in immunology, containing both major and rare cell populations, with cell population identities from expert manual gating as the reference standard. Several methods performed well, including FlowSOM, X-shift, PhenoGraph, Rclusterpp, and flowMeans. Among these, FlowSOM had extremely fast runtimes, making this method well-suited for interactive, exploratory analysis of large, high-dimensional data sets on a standard laptop or desktop computer. These results extend previously published comparisons by focusing on high-dimensional data and including new methods developed for CyTOF data. R scripts to reproduce all analyses are available from GitHub (https://github.com/lmweber/cytometry-clustering-comparison), and pre-processed data files are available from FlowRepository (FR-FCM-ZZPH), allowing our comparisons to be extended to include new clustering methods and reference data sets. © 2016 The Authors. Cytometry Part A published by Wiley Periodicals, Inc. on behalf of ISAC.


History as reflective practice: A model for integrating historical studies into nurse education.
The role of history in developing professional identity in nursing is well known, and the discipline of nursing history research continues to flourish. Yet this work often struggles to find its way into undergraduate university nurse education courses. We put forward a model for "history as reflective practice" in which we suggest that historical studies can be used as a form of evidence to develop critical thinking and clinical reasoning, as well as situate nursing practice within its social and political context. In this model, we draw on historical scholarship related to the profession, practice and person, focusing on work which demonstrates nursing's contribution to broader systems of health care. Drawing on Lewenson and Lynaugh's 'history by stealth' approach, curriculum mapping and constructive alignment techniques are used to identify the moments in an existing programme where historical scholarship is relevant to an intended learning outcome. We then use an interdisciplinary team to develop learning activities and assessment tasks drawing on both primary and secondary sources that are then embedded within existing subjects. This model encourages students to consider history as a way of knowing and as a form of evidence within their reflective practice. Furthermore, it creates knowledge that continues to foster and acknowledge nurses', and nursing's, contribution to the development of human health.


Building focused cardiac ultrasound capacity in a lower middle-income country: A single centre study to assess training impact.
In low- and middle-income countries (LMICs) where echocardiography experts are in short supply, training non-cardiologists to perform Focused Cardiac Ultrasound (FoCUS) could minimise diagnostic delays in time-critical emergencies. Despite advocacy for FoCUS training however, opportunities in LMICs are limited, and the impact of existing curricula uncertain. The aim of this study was to assess the impact of FoCUS training based on the Focus Assessed Transthoracic Echocardiography (FATE) curriculum. Our primary objective was to assess knowledge gain. Secondary objectives were to evaluate novice FoCUS image quality, assess inter-rater agreement between expert and novice FoCUS and identify barriers to the establishment of a FoCUS training programme locally. This was a pre-post quasi-experimental study at a tertiary hospital in Nairobi, Kenya. Twelve novices without prior echocardiography training underwent FATE training, and their knowledge and skills were assessed. Pre- and post-test scores were compared using the Wilcoxon signed-rank test to establish whether the median of the difference was different than zero. Inter-rater agreement between expert and novice scans was assessed, with a Cohen's kappa >0.6 indicative of good inter-rater agreement. Knowledge gain was 37.7%, with a statistically significant difference between pre-and post-test scores (z = 2.934, p = 0.001). Specificity of novice FoCUS was higher than sensitivity, with substantial agreement between novice and expert scans for most FoCUS target conditions. Overall, 65.4% of novice images were of poor quality. Post-workshop supervised practice was limited due to scheduling difficulties. Although knowledge gain is high following a brief training in FoCUS, image quality is poor and sensitivity low without adequate supervised practice. Substantial agreement between novice and expert scans occurs even with insufficient practice when the prevalence of pathology is low. Supervised FoCUS practice is challenging to achieve in a real-world setting in LMICs, undermining the effectiveness of training initiatives.


Considering methodological options for reviews of theory: illustrated by a review of theories linking income and health.
Review of theory is an area of growing methodological advancement. Theoretical reviews are particularly useful where the literature is complex, multi-discipline, or contested. It has been suggested that adopting methods from systematic reviews may help address these challenges. However, the methodological approaches to reviews of theory, including the degree to which systematic review methods can be incorporated, have received little discussion in the literature. We recently employed systematic review methods in a review of theories about the causal relationship between income and health. This article discusses some of the methodological issues we considered in developing the review and offers lessons learnt from our experiences. It examines the stages of a systematic review in relation to how they could be adapted for a review of theory. The issues arising and the approaches taken in the review of theories in income and health are considered, drawing on the approaches of other reviews of theory. Different approaches to searching were required, including electronic and manual searches, and electronic citation tracking to follow the development of theories. Determining inclusion criteria was an iterative process to ensure that inclusion criteria were specific enough to make the review practical and focused, but not so narrow that key literature was excluded. Involving subject specialists was valuable in the literature searches to ensure principal papers were identified and during the inductive approaches used in synthesis of theories to provide detailed understanding of how theories related to another. Reviews of theory are likely to involve iterations and inductive processes throughout, and some of the concepts and techniques that have been developed for qualitative evidence synthesis can be usefully translated to theoretical reviews of this kind. It may be useful at the outset of a review of theory to consider whether the key aim of the review is to scope out theories relating to a particular issue; to conduct in-depth analysis of key theoretical works with the aim of developing new, overarching theories and interpretations; or to combine both these processes in the review. This can help decide the most appropriate methodological approach to take at particular stages of the review.


Bone blood flow and age in the rat.
1. The proportion of the cardiac output delivered to the skeleton and individual bones has been determined in rats of differing ages and weights with radioactive microspheres. 2. There was a decrease in the blood flow to the tibia--fibula, jaw, femur, pelvis, humerus, radius and ulna, and scapula with increase in age and weight of rats, but no change was seen in blood flow to the ribs, spine, skull or whole skeleton. 3. The proportion of the cardiac output delivered to the whole skeleton was approximately 3%. 4. The derived values for skeletal blood flow agree well with estimates by other authors using the technique of bone uptake and blood clearance of 45Ca.


A cleavable ligand column for the rapid isolation of large quantities of homogeneous and functional neurotensin receptor 1 variants from E. coli.
G protein-coupled receptors (GPCRs) are key players of cell signaling, thus representing important drug targets for the treatment of human diseases. Since inherent difficulties in receptor production and handling have precluded the application of many in vitro experiments, major questions about GPCR mechanisms and dynamics remain elusive to date. We recently used directed evolution in Escherichia coli on neurotensin receptor 1 (NTR1) for the generation of GPCR variants with greatly elevated functional expression levels and with excellent stability in detergent micelles. In this work we outline a highly efficient purification method for our evolved receptor variants, which is based on the application of an inexpensive, disposable high-affinity ligand column as the initial purification step. The ligand resin allows isolation of correctly folded GPCR variants directly from whole E. coli cell lysates at the scale of 10mg and it permits preparations of agonist- and antagonist-bound receptor samples. The purification principle presented here was key to the first structures of signaling-active NTR1 variants (Egloff et al., 2014). Since E. coli is uniquely suitable for the production of fully deuterated proteins, our method provides the basis for an array of NMR experiments that were not feasible for GPCRs to date, but which will shed light on novel aspects of receptor function and dynamics.


Sex knowledge in males and females recovering from a myocardial infarction: a brief communication.
The purpose of this article was to explore sexual knowledge in persons who had suffered from myocardial infarction (MI). Seventy-six Swedish persons completed the "Sex after MI Knowledge Test" questionnaire. Overall, 53% of the men and 45% of the women scored maximum in the test. In a comparison between sexes, the men scored significantly more often a correct answer compared to the women for two out of the 25 items. The levels of correct answers were less then 50% for 14 out of the 25 items in both sexes. In conclusion we found that people who had suffered MI had poor levels of knowledge about sex and that there were some differences concerning lesser knowledge among the females in comparison to males. In regard to application, using a validated instrument facilitates an interactive communication between the patient and health care professionals, and opens up for a tailored education in line with the patient's and his or her partner's needs.


Experience with hookwire localisation excision biopsy at a medical centre in Malaysia.
With an increasing number of women undergoing breast screening, an effective method of removing non-palpable lumps detected by mammography or sonography is by hookwire localisation excision biopsy (HWLB). The aim of this paper was to audit the practice of HWLB at the University Malaya Medical Centre. Patients with benign or suspicious preoperative diagnoses of a non-palpable lump and who underwent HWLB were included in this study. Pathological examination of the surgical specimens was conducted and a correlation with preoperative assessment modalities was reported. A total of 59 HWLBs were carried out in 57 female patients. The mean age of the patients was 51.5 years. The overall malignancy rate was 32.3 percent (19 out of 59 cases) with a benign to malignant ratio of 3.1 to 1. Ten of these cases were ductal carcinoma-in-situ. Out of 25 patients who were suspicious on preoperative assessment, 16 malignancies were found, while in the 33 patients thought to be benign on preoperative assessment, there were three malignancies, giving a sensitivity of 84.2 percent and a specificity of 76.9 percent (p is less than 0.05). The mean tissue volume excised in 53 available records was 50.0 cm(3), with pathological tissue comprising only 15.4 percent of the total excised volume. Clear margins were obtained in 42.1 percent of the patients. The overall operative complication rate was 10.2 percent. Malignancy was reported in one third of women undergoing HWLB, of which 16 had suspicious features on radiological assessment and/or fine needle aspiration cytology/core needle biopsy preoperatively. Non-palpable lumps should be excised by HWLB for a definitive diagnosis in case of any suspicion on preoperative assessment, as the prognosis is excellent.


Crystal structure of rhodocytin, a ligand for the platelet-activating receptor CLEC-2.
Binding of the snake venom protein rhodocytin to CLEC-2, a receptor on the surface of human platelets, initiates a signaling cascade leading to platelet activation and aggregation. We have previously solved the structure of CLEC-2. The 2.4 A resolution crystal structure of rhodocytin presented here demonstrates that it is the first snake venom or other C-type lectin-like protein to assemble as a non-disulfide linked (alphabeta)(2) tetramer. Rhodocytin is highly adapted for interaction with CLEC-2 and displays a concave binding surface, which is highly complementary to the experimentally determined binding interface on CLEC-2. Using computational dynamic methods, surface electrostatic charge and hydrophobicity analyses, and protein-protein docking predictions, we propose that the (alphabeta)(2) rhodocytin tetramer induces clustering of CLEC-2 receptors on the platelet surface, which will trigger major signaling events resulting in platelet activation and aggregation.


Terminal digit preference: a source of measurement error in breast cancer diameter reporting.
Objectives: Women diagnosed with breast cancer are offered treatment and therapy based on tumor characteristics, including tumor diameter. There is scarce knowledge whether tumor diameter is accurately reported, or whether it is unconsciously rounded to the nearest half-centimeter (terminal digit preference). This study aimed to assess the precision (number of digits) of breast cancer tumor diameters and whether they are affected by terminal digit preference. Furthermore, we aimed to assess the agreement between mammographic and histopathologic tumor diameter measurements.Material and Methods: This national registry study included reported mammographic and registered histopathologic tumor diameter information from the Cancer Registry of Norway for invasive breast cancers diagnosed during 2012-2016. Terminal digit preference was assessed using histograms. Agreement between mammographic and histopathologic measurements was assessed using the intraclass correlation coefficient (ICC) and Bland-Altman plots.Results: Mammographic, histopathologic, or both tumor measurements were available for 7792, 13,541 and 6865 cases, respectively. All mammographic and 97.2% of histopathologic tumor diameters were recorded using whole mm. Terminal digits of zero or five were observed among 38.7% and 34.8% of mammographic and histopathologic measurements, respectively. There was moderate agreement between the two measurement methods (ICC = 0.52, 95% CI: 0.50-0.53). On average, mammographic measurements were 1.26 mm larger (95% limits of agreement: -22.29-24.73) than histopathologic measurements. This difference increased with increasing tumor size.Conclusion: Terminal digit preference was evident among breast cancer tumor diameters in this nationwide study. Further studies are needed to investigate the potential extent of under-staging and under-treatment resulting from this measurement error.


Frequent use of paracetamol and risk of allergic disease among women in an Ethiopian population.
The hypothesis that paracetamol might increase the risk of asthma and other allergic diseases have gained support from a range of independent studies. However, in studies based in developed countries, the possibility that paracetamol and asthma are associated through aspirin avoidance is difficult to exclude. To explore this hypothesis among women in a developing country, where we have previously reported aspirin avoidance to be rare. In 2005/6 a population based cohort of 1065 pregnant women was established in Butajira, Ethiopia and baseline demographic data collected. At 3 years post birth, an interview-based questionnaire administered to 945 (94%) of these women collected data on asthma, eczema, and hay fever in the past 12 month, frequency of paracetamol use and potential confounders. Allergen skin tests to Dermatophagoides pteronyssinus and cockroach were also performed. The independent effects of paracetamol use on allergic outcomes were determined using multiple logistic regression analysis. The prevalence of asthma, eczema and hay fever was 1.7%, 0.9% and 3.8% respectively; of any one of these conditions 5.5%, and of allergen sensitization 7.8%. Paracetamol use in the past month was reported by 29%, and associations of borderline significance were seen for eczema (adjusted OR (95% CI) = 8.51 (1.68 to 43.19) for 1-3 tablets and 2.19 (0.36 to 13.38) for ≥4 tablets, compared to no tablets in the past month; overall p = 0.055) and for 'any allergic condition' (adjusted OR (95% CI) = 2.73 (1.22 to 6.11) for 1-3 tablets and 1.35 (0.67 to 2.70) for ≥4 tablets compared to 0 in the past month; overall p = 0.071). This study provides further cross-sectional evidence that paracetamol use increases the risk of allergic disease.


Polysubstance Use Patterns among Justice-Involved Individuals Who Use Opioids.
The current study explores pre-incarceration polysubstance use patterns among a justice-involved population who use opioids. Design: Setting: Data from prison and jail substance use programing in the state of Kentucky from 2015-2017 was examined. Participants: A cohort of 6,569 individuals who reported both pre-incarceration use of opioids and reported the use of more than one substance per day. Measurements: To determine the different typologies of polysubstance use involving opioids, latent profile analysis of the pre-incarceration thirty-day drug use of eight substances was conducted. Multinomial logistic regression predicted latent profile membership. Findings: Six unique profiles of polysubstance use involving opioids and other substances were found; Primarily Alcohol (9.4%), Primarily Heroin (19.0%), Less Polysubstance Use (34.3%), Tranquilizer Polysubstance Use (16.3%), Primarily Buprenorphine (7.8%), and Stimulant-Opioid (13.2%). Profiles differed by rural/urban geography, injection drug use, physical, and mental health symptoms. Conclusion: Findings indicate the heterogeneity of opioid use among a justice-involved population. More diverse polysubstance patterns may serve as a proxy to identifying individuals with competing physical and mental health needs. Future interventions could be tailored to polysubstance patterns during the period of justice-involvement.


Long-term effects of metals in sewage sludge on soils, microorganisms and plants.
This paper reviews the evidence for impacts of metals on the growth of selected plants and on the effects of metals on soil microbial activity and soil fertility in the long-term. Less is known about adverse long-term effects of metals on soil microorganisms than on crop yields and metal uptake. This is not surprising, since the effects of metals added to soils in sewage sludge are difficult to assess, and few long-term experiments exist. Controlled field experiments with sewage sludges exist in the UK, Sweden, Germany and the USA and the data presented here are from these long-term field experiments only. Microbial activity and populations of cyanobacteria, Rhizobium leguminosarum bv. trifolii, mycorrhizae and the total microbial biomass have been adversely affected by metal concentrations which, in some cases, are below the European Community's maximum allowable concentration limits for metals in sludge-treated soils. For example, N2-fixation by free living heterotrophic bacteria was found to be inhibited at soil metal concentrations of (mg kg-1): 127 Zn, 37 Cu, 21 Ni, 3.4 Cd, 52 Cr and 71 Pb. N2-fixation by free-living cyanobacteria was reduced by 50% at metal concentrations of (mg kg-1): 114 Zn, 33 Cu, 17 Ni, 2.9 Cd, 80 Cr and 40 Pb. Rhizobium leguminosarum bv. trifolii numbers decreased by several orders of magnitude at soil metal concentrations of (mg kg-1): 130-200 Zn, 27-48 Cu, 11-15 Ni, and 0.8-1.0 Cd. Soil texture and pH were found to influence the concentrations at which toxicity occurred to both microorganisms and plants. Higher pH, and increased contents of clay and organic carbon reduced metal toxicity considerably. The evidence suggests that adverse effects on soil microbial parameters were generally found at surprizingly modest concentrations of metals in soils. It is concluded that prevention of adverse effects on soil microbial processes and ultimately soil fertility, should be a factor which influences soil protection legislation.


Outside the envelope: rare events disrupt the relationshipbetween climate factors and species interactions.
The order in which species arrive during community assembly can be an important driver of community composition and function. However, the strength of these priority effects can be variable, in part because of strong site and year effects. To understand how priority effects vary in importance with abiotic conditions, we initiated identical community assembly experiments in which we varied the timing of arrival of native and exotic grass species in each of 4 yr across three grassland sites in northern California. This uniquely replicated experiment tested the power of priority to determine initial community structure in a restoration context across a natural range of conditions. There were large and significant differences in both total seeded cover and the strength of priority across sites and years of initiation, confirming the suspicion that most ecological experiments may lack spatial and temporal generality. On the other hand, much of the variation in strength of priority could be related to climate. Strikingly, however, the model fit across the three sites and the first 3 yr of the study (the first nine experiments) was radically altered when we included the fourth year, which was characterized by an unusual weather pattern with higher temporal variability in rainfall (a rainfall pattern predicted to increase with climate change). This year produced relatively low strength of priority, supporting the suggestion that highly variable climates may be associated with lower strength of priority effects. Experiments that examine community assembly over a range of naturally occurring abiotic conditions enhance our ability to predict when priority effects will be important, allowing us to explore shifting patterns of community assembly in the face of climate change and optimize restoration strategies based on environmental conditions.


Functional in vitro assembly of the integral membrane bacterial thermosensor DesK.
The Bacillus subtilis DesK histidine kinase (HK) is an integral membrane thermosensor that forms part of a regulatory circuit which controls the physical state of membrane lipids. In the pursuit of biochemical and structural approaches to study lipid fluidity-dependent DesK thermosensing, we found that standard expression methods failed to produce enough amounts of a fully functional protein. Here, we describe a high-yield purification method based in an Escherichia coliin vitro transcription-translation system. The enzymatic activities of the full-length protein, either solubilized with detergents or co-translationally inserted into liposomes, have been characterized and compared with those measured for the constitutively active cytoplasmic domain of DesK, lacking the transmembrane sensor domain. As expected, the autokinase activity of liposome-inserted DesK was greatly increased when the incubation temperature was decreased from 37 to 25 degrees C. This is the first report of the spontaneous in vitro membrane insertion of a fully functional bacterial HK thermosensor. Moreover, this single step procedure should greatly aid the isolation of a wide range of membrane-associated HKs for biochemical and biophysical studies.


PCR detection of enterohemorrhagic Escherichia coli O145 in food by targeting genes in the E. coli O145 O-antigen gene cluster and the shiga toxin 1 and shiga toxin 2 genes.
Shiga toxin-producing Escherichia coli (STEC) strains belonging to serogroup O145 are an important cause of hemorrhagic colitis and hemolytic uremic syndrome worldwide. Cattle and other animals are potential reservoirs for this pathogen. To develop PCR assays for detection and identification of E. coli O145, the wzx (O-antigen flippase) and wzy (O-antigen polymerase) genes in the O145 O-antigen gene cluster that are specific for this serogroup were selected as targets. Oligonucleotide primers complementary to regions in the E. coli O145 wzx and wzy genes were designed to perform PCR assays with DNA from strains of E. coli O145, non-O145 E. coli serogroups, and other bacterial genera. The assays were highly specific for E. coli O145. A multiplex PCR assay targeting the E. coli O145 wzx and wzy genes and the Shiga toxin 1 (stx(1)) and Shiga toxin 2 (stx(2)) genes and a real-time multiplex PCR assay targeting the O145 wzy, stx(1), and stx(2) genes were developed for detection of STEC O145. The assays were used for detecting STEC O145 in seeded ground beef, lettuce, and raw milk initially inoculated with ca. 2, 20, or 200 CFU/25 g or 25 mL after 8 or 20 h of enrichment at 42 degrees C in modified EC broth containing 20 mg/L of novobiocin. STEC O145 was detected in all samples inoculated with 2 CFU/25 g or 25 mL. The detection limit of the multiplex PCR assays was <or=7.9 x 10(4) CFU/mL, which corresponded to <or=400 CFU/PCR reaction. The PCR assays can be employed to identify enterohemorrhagic E. coli serogroup O145 and to detect low levels of the pathogen in food.


Unrelated donor α/β T cell- and B cell-depleted HSCT for the treatment of pediatric acute leukemia.
Unrelated donor (URD) hematopoietic stem cell transplant (HSCT) is associated with an increased risk of severe graft-versus-host disease (GVHD). TCRαβ/CD19 depletion may reduce this risk, whereas maintaining graft-versus-leukemia. Outcome data with TCRαβ/CD19 depletion generally describe haploidentical donors, with relatively few URDs. We hypothesized that TCRαβ/CD19-depletion would attenuate the risks of GVHD and relapse for URD HSCT. Sixty pediatric and young adult (YA) patients with hematologic malignancies who lacked a matched-related donor were enrolled at 2 large pediatric transplantation centers between October 2014 and September 2019. All patients with acute leukemia had minimal residual disease testing, and DP typing was available for 77%. All patients received myeloablative total body irradiation- or busulfan-based conditioning with no posttransplant immune suppression. Engraftment occurred in 98%. Four-year overall survival was 69% (95% confidence interval [CI], 52%-81%), and leukemia-free survival was 64% (95% CI, 48%-76%), with no difference between lymphoid and myeloid malignancies (P = .6297 and P = .5441, respectively). One patient (1.7%) experienced primary graft failure. Relapse occurred in 11 patients (3-year cumulative incidence, 21%; 95% CI, 11-34), and 8 patients (cumulative incidence, 15%; 95% CI, 6.7-26) experienced nonrelapse mortality. Grade III to IV acute GVHD was seen in 8 patients (13%), and 14 patients (26%) developed chronic GVHD, of which 6 (11%) had extensive disease. Nonpermissive DP mismatch was associated with higher likelihood of acute GVHD (odds ratio, 16.50; 95% CI, 1.67-163.42; P = .0166) but not with the development of chronic GVHD. URD TCRαβ/CD19-depleted peripheral HSCT is a safe and effective approach to transplantation for children/YAs with leukemia. This trial was registered at www.clinicaltrials.gov as #NCT02323867.


Etiopathogenesis of hyperostosis frontalis interna: a mystery still.
Hyperostosis frontalis interna is a morphological pattern characterized by single or multiple bony nodules situated on the inner lamina of the frontal bone. It is seldom found in males, but it is a common phenomenon among post-menopausal females in modern societies but relatively rare in antiquity. The etiopathogenesis of the trait is a matter of debate and ranges from genetic predisposition to epigenetic, while endocrine disturbances, aging, and dietary factors are also listed among the causes. We studied the frequency, characteristic features, and etiopathogenesis of the disease in recent cadaveric and dry skull specimens. The frequency of hyperostosis frontalis interna in cadavers and dry skull materials was almost identical, 12.5% and 12.3%, respectively. In cadavers, 87.5% of severe hyperostosis frontalis interna cases were found in females over 65 years-old. Interestingly, in two cadavers we found hyperostotic lesions spreading onto adjacent tissues such as the dura and falx cerebri. We provide some new aspects that may help in better understanding of the etiopathogenesis of hyperostosis frontalis interna. Thereby, we discuss the various etiopathogenesis models found in the literature.


Functional genomics of pH homeostasis in Corynebacterium glutamicum revealed novel links between pH response, oxidative stress, iron homeostasis and methionine synthesis.
The maintenance of internal pH in bacterial cells is challenged by natural stress conditions, during host infection or in biotechnological production processes. Comprehensive transcriptomic and proteomic analyses has been conducted in several bacterial model systems, yet questions remain as to the mechanisms of pH homeostasis. Here we present the comprehensive analysis of pH homeostasis in C. glutamicum, a bacterium of industrial importance. At pH values between 6 and 9 effective maintenance of the internal pH at 7.5 +/- 0.5 pH units was found. By DNA microarray analyses differential mRNA patterns were identified. The expression profiles were validated and extended by 1D-LC-ESI-MS/MS based quantification of soluble and membrane proteins. Regulators involved were identified and thereby participation of numerous signaling modules in pH response was found. The functional analysis revealed for the first time the occurrence of oxidative stress in C. glutamicum cells at neutral and low pH conditions accompanied by activation of the iron starvation response. Intracellular metabolite pool analysis unraveled inhibition of the TCA and other pathways at low pH. Methionine and cysteine synthesis were found to be activated via the McbR regulator, cysteine accumulation was observed and addition of cysteine was shown to be toxic under acidic conditions. Novel limitations for C. glutamicum at non-optimal pH values were identified by a comprehensive analysis on the level of the transcriptome, proteome, and metabolome indicating a functional link between pH acclimatization, oxidative stress, iron homeostasis, and metabolic alterations. The results offer new insights into bacterial stress physiology and new starting points for bacterial strain design or pathogen defense.


Synergistic stimulation of type I interferons during influenza virus coinfection promotes Streptococcus pneumoniae colonization in mice.
Pneumococcal infection of the respiratory tract is often secondary to recent influenza virus infection and accounts for much of the morbidity and mortality during seasonal and pandemic influenza. Here, we show that coinfection of the upper respiratory tract of mice with influenza virus and pneumococcus leads to synergistic stimulation of type I IFNs and that this impairs the recruitment of macrophages, which are required for pneumococcal clearance, due to decreased production of the chemokine CCL2. Type I IFN expression was induced by pneumococcal colonization alone. Colonization followed by influenza coinfection led to a synergistic type I IFN response, resulting in increased density of colonizing bacteria and susceptibility to invasive infection. This enhanced type I IFN response inhibited production of the chemokine CCL2, which promotes the recruitment of macrophages and bacterial clearance. Stimulation of CCL2 by macrophages upon pneumococcal infection alone required the pattern recognition receptor Nod2 and expression of the pore-forming toxin pneumolysin. Indeed, the increased colonization associated with concurrent influenza virus infection was not observed in mice lacking Nod2 or the type I IFN receptor, or in mice challenged with pneumococci lacking pneumolysin. We therefore propose that the synergistic stimulation of type I IFN production during concurrent influenza virus and pneumococcal infection leads to increased bacterial colonization and suggest that this may contribute to the higher rates of disease associated with coinfection in humans.


The African Pediatric Fellowship Program: Training in Africa for Africans.
Africa has a significant burden of childhood disease, with relatively few skilled health care professionals. The African Paediatric Fellowship Programme was developed by the Department of Pediatrics and Child Health at the University of Cape Town to provide relevant training for African child health professionals, by Africans, within Africa. Trainees identified by partner academic institutions spend 6 months to 2 years training in the Department of Pediatrics and allied disciplines. They then return to their home institution to build practice, training, research, and advocacy. From 2008 to 2015, 73 physicians have completed or are completing training in general pediatrics or a pediatric subspecialty. At 1 year posttraining, 98% to 100% are practicing back in their home institution. The impact of the returning fellows is evident from their practice interventions, research collaborations, and positions as stakeholders who can change health care policies. Thirty-three centers in 13 African countries are partners with the program, and the program template is now followed by other partner sites in Africa. Increasing and retaining the skills pool of African child health specialists is building a network of motivated, highly skilled clinicians who are equipped to advance child health in Africa.


Association between dietary carbohydrates and body weight.
The role of dietary carbohydrates in weight loss has received considerable attention in light of the current obesity epidemic. The authors investigated the association of body mass index (weight (kg)/height (m)(2)) with dietary intake of carbohydrates and with measures of the induced glycemic response, using data from an observational study of 572 healthy adults in central Massachusetts. Anthropometric measurements, 7-day dietary recalls, and physical activity recalls were collected quarterly from each subject throughout a 1-year study period. Data were collected between 1994 and 1998. Longitudinal analyses were conducted, and results were adjusted for other factors related to body habitus. Average body mass index was 27.4 kg/m(2) (standard deviation, 5.5), while the average percentage of calories from carbohydrates was 44.9 (standard deviation, 9.6). Mean daily dietary glycemic index was 81.7 (standard deviation, 5.5), and glycemic load was 197.8 (standard deviation, 105.2). Body mass index was found to be positively associated with glycemic index, a measure of the glycemic response associated with ingesting different types of carbohydrates, but not with daily carbohydrate intake, percentage of calories from carbohydrates, or glycemic load. Results suggest that the type of carbohydrate may be related to body weight. However, further research is required to elucidate this association and its implications for weight management.