Genome-wide association identifies nine common variants associated with fasting proinsulin levels and provides new insights into the pathophysiology of type 2 diabetes.
Proinsulin is a precursor of mature insulin and C-peptide. Higher circulating proinsulin levels are associated with impaired β-cell function, raised glucose levels, insulin resistance, and type 2 diabetes (T2D). Studies of the insulin processing pathway could provide new insights about T2D pathophysiology. We have conducted a meta-analysis of genome-wide association tests of ∼2.5 million genotyped or imputed single nucleotide polymorphisms (SNPs) and fasting proinsulin levels in 10,701 nondiabetic adults of European ancestry, with follow-up of 23 loci in up to 16,378 individuals, using additive genetic models adjusted for age, sex, fasting insulin, and study-specific covariates. Nine SNPs at eight loci were associated with proinsulin levels (P < 5 × 10(-8)). Two loci (LARP6 and SGSM2) have not been previously related to metabolic traits, one (MADD) has been associated with fasting glucose, one (PCSK1) has been implicated in obesity, and four (TCF7L2, SLC30A8, VPS13C/C2CD4A/B, and ARAP1, formerly CENTD2) increase T2D risk. The proinsulin-raising allele of ARAP1 was associated with a lower fasting glucose (P = 1.7 × 10(-4)), improved β-cell function (P = 1.1 × 10(-5)), and lower risk of T2D (odds ratio 0.88; P = 7.8 × 10(-6)). Notably, PCSK1 encodes the protein prohormone convertase 1/3, the first enzyme in the insulin processing pathway. A genotype score composed of the nine proinsulin-raising alleles was not associated with coronary disease in two large case-control datasets. We have identified nine genetic variants associated with fasting proinsulin. Our findings illuminate the biology underlying glucose homeostasis and T2D development in humans and argue against a direct role of proinsulin in coronary artery disease pathogenesis.


Prevalence and Correlates of Olfactory Dysfunction in Old Age: A Population-Based Study.
Olfactory dysfunction (OD) in old age is associated with poor health outcomes. Interrelationships among different correlates of OD can offer insights into the underlying mechanisms, but to date remain understudied. Odor identification performance and self-reported olfactory functioning were studied in 2,234 people aged 60-90 years, who were free of neurodegenerative disease and enrolled in the Swedish National Study on Aging and Care in Kungsholmen (SNAC-K) study, Stockholm, Sweden. OD was defined as the inability to identify more than 10 out of 16 odors (free or cued identification) in a standardized odor identification task. OD prevalence was estimated, and associations with demographic, genetic, vascular, clinical, and behavioral factors, as well as their interactions were examined using multiple logistic regression analyses. Overall prevalence of OD was 24.8% (CI: 23.1; 26.6). Self-reports were characterized by low sensitivity (35%), but high specificity (87%). Advancing age (OR = 15.50, CI = 9.40; 26.10 between the first and last age group), and history of coronary heart disease (OR = 1.35, 95% CI = 1.04; 1.75) were the principal factors associated with an increased probability of OD, whereas female gender (OR = 0.53, 95% CI = 0.43; 0.66) and more years of education (OR = 0.97, CI 0.94; 0.99) were linked to a lower probability. Exploratory interaction analyses indicated that prevalence of OD was particularly elevated among Apolipropotein E (APOE) ε4 allele carriers who were also obese, and that being physically active counteracted the negative impact of cerebrovascular disease on OD. Demographic and genetic factors, but also prior and current health insults, are linked to OD in old age. Modulatory effects of behavioral factors highlight their value as possible prevention targets.


Supporting tobacco control: stimulating local newspaper coverage with a technical assistance website for local coalitions.
A large and growing literature confirms that well-designed web-based programs can be effective in preventing or treating several chronic diseases. This study examined how the Internet can deliver information and train community activists and specifically tested the effects of web-based technical assistance on local tobacco control coalitions' efforts to use media advocacy to advance their agendas. The authors compared a highly interactive, Enhanced website (intervention) to a noninteractive, Basic text-based website (comparison) in Colorado communities. A total of 24 tobacco control coalitions led by local county health departments and nursing services were enrolled in the project and randomly assigned to use either the intervention or comparison website. A total of 73 local daily and weekly newspapers were identified in the service areas of 23 of the 24 coalitions. A posttest assessment of newspaper coverage was conducted to locate all newspaper articles with tobacco control information published between January 1 and April 9, 2004, the last 3 months of the intervention. Although there was no evidence of a treatment effect on the frequency of newspaper articles on tobacco-related issues, there was, however, evidence that newspapers in counties where the coalition had access to the Enhanced website printed more stories focused on local/regional issues and more anti-tobacco local/regional stories than in the counties where coalitions had access to the Basic website. Coalitions can improve their influence on local media for community tobacco control when high-quality online technical assistance, training, and resources are available to them.


Optimal stroke preventive strategy for patients aged 80 years or older with atrial fibrillation: a systematic review with traditional and network meta-analysis.
An optimal antithrombotic strategy for patients aged 80 years or older with atrial fibrillation (AF) remains elusive. Using a systematic review with traditional and network meta-analysis, we investigated outcomes in AF patients ≥80 years treated with different antithrombotic strategies. We searched eligible randomised controlled trials (RCTs) and observational studies from MEDLINE, EMBASE, Cochrane Library and Web of Science databases from inception to 16 December 2021. Research comparing treatment outcomes of novel oral anticoagulants (NOACs), aspirin, vitamin K antagonists (VKAs) or no oral anticoagulant/placebo therapy in patients ≥80 years with AF were included. Outcomes were stroke or systemic embolism (SSE), major bleeding, all-cause mortality, intracranial bleeding (ICH) and gastrointestinal bleeding. Traditional and network meta-analyses were performed. Net clinical benefit integrating SSE and major bleeding was calculated. Fifty-three studies were identified for analysis. In the meta-analysis of RCTs, risk of SSE (risk ratio [RR]: 0.82; 95% confidence interval [CI]: 0.73-0.99) and ICH (RR: 0.38; 95% CI: 0.28-0.52) was significantly reduced when NOACs were compared with VKAs. Network meta-analysis of RCTs demonstrated that edoxaban (P-score: 0.8976) and apixaban (P-score: 0.8528) outperformed other antithrombotic therapies by showing a lower major bleeding risk and better net clinical benefit. Both traditional and network meta-analyses from RCTs combining with observational studies showed consistent results. In patients aged 80 years or older with AF, NOACs have better outcomes than VKAs regarding efficacy and safety profiles. Edoxaban and apixaban may be preferred treatment options since they are safer than other antithrombotic strategies.


There is a need for a multidisciplinary approach to patient safety.
In the lead paper, Zimmerman et al. explore the role of problem and solution ownership in solution adoption by front-line healthcare staff. The front-line ownership (FLO) approach suggested by the authors proposes a shift in leadership style such that managers act as enablers of patient safety improvement efforts by front-line staff (i.e., a bottom-up management style). From a systems perspective, it would be necessary to examine contributing factors to patient safety problems, including process, workflow, task requirements etc., in order to effectively utilize a solution-oriented approach such as FLO. These factors would be difficult to identify by clinical staff alone. There is a need for a multidisciplinary approach to patient safety management in healthcare.


Effect of Oxalic Acid Treatment on Conductive Coatings Formed by Ni@Ag Core-Shell Nanoparticles.
Low-cost metallic nanoink based on nickel-silver core-shell nanoparticles (Ni@Ag NPs) was used for the formation of conductive metallic coatings with low sintering temperature, which can be successfully applied for replacement of currently used silver-based nanoinks in printed electronics. The effect of oxalic acid (OA) on the sintering temperature and conductivity of coatings formed by Ni@Ag NPs was evaluated. It was found that the addition of OA to the ink formulation and post-printing treatment of deposited films with this acid provided a noticeable decrease in the sintering temperature required for obtaining conductive patterns that is especially important for utilizing the polymeric substrates. The obtained resistivity of metallic coatings after sintering at temperature as low as 100 °C was found to be 30 µΩ·cm, only ~4 times higher compared to the resistivity of bulk Ni that is promising for future application of such materials for fabrication of low-cost flexible printed patterns.


Promoting equity in health through research and understanding.
Developing strategies to reduce inequities in health requires an understanding of how inequities occur, determining the salient factors in their production, and deciding which ones are most amenable to change. The recognition of several principles regarding the manifestations and genesis of inequities can help to decide on strategies. In making decisions, it is important to consider whether the aim is to reduce disparities in the occurrence of ill health or to reduce disparities in the severity (including co-morbidity, disability, dysfunction and fatality) of ill health. Evidence shows that the major impact on equity of health services, particularly regarding their potential to reduce severity, is attributable to the strength of primary care resources and services in communities and countries. Virtually every influence on the genesis of inequities is determined by the political context in which policy is made. The issue of health services is not different in this regard from other types of strategies. There is no longer any doubt about the pervasive influence of social factors on health. Almost two centuries of descriptive research provides convincing evidence of associations between social structures and relationships and health status in all countries and in all societies; if there is anything new from more recent research, it is that the association is not limited to differences between the lowest social strata and other social strata. Rather, the association is noted throughout the social spectrum. That is, there is a social gradient in health such that, for many if not most manifestations of ill health, the lower the social stratum, the worse the health. The challenge for the future is to understand why this is the case, to create a consensus that these inequalities are unnecessary and unacceptable, and to devise strategies that are both effective and possible. This paper will focus on the first of these aims, in a context that facilitates attention to the second and third aims.


Isolation and characterization of an endogenous C-terminal fragment of the alpha-neo-endorphin/dynorphin precursor from bovine caudate nucleus.
Antibodies have been raised to a synthetic peptide corresponding to the C-terminal 15-amino acid residues of prodynorphin, the common precursor to the neo-endorphins and dynorphins. The amino acid sequence of the antigen was based on the sequence deduced from mRNA isolated and cloned from porcine hypothalamus (Kakidani, H., Y. Furutani, H. Takahashi, M. Noda, Y. Morimoto, T. Hirose, M. Asai, S. Inayama, S. Nakanishi, and S. Numa (1982) Nature 298: 245-248). Using a radioimmunoassay developed from these antibodies we have isolated an endogenous prodynorphin C-fragment from bovine caudate nucleus. The isolated peptide displayed characteristics on gel filtration similar to those of synthetic prodynorphin C-fragment predicted from the porcine mRNA sequence but had low cross-reactivity in the radioimmunoassay. Sequencing and amino acid analysis showed a substitution of serine for asparagine at position 6 in the porcine sequence. Dynorphin B (rimorphin), which is adjacent to prodynorphin C-fragment in the precursor, was isolated from the same extract. Amino acid analysis and elution position on a gel filtration column confirmed its structure as that previously characterized from bovine pituitary extracts. The release of prodynorphin C-fragment and the C-terminus of dynorphin B from the porcine precursor would require cleavage at a single arginine residue. However, a terminal arginine was not present on either of these prodynorphin peptides isolated from bovine caudate. The data would suggest that processing at a single arginine residue results in elimination of the arginine, a feature in common with processing at paired basic residues.


Developing an integrated intervention to address intimate partner violence and psychological distress in Congolese refugee women in Tanzania.
Multi-sectoral, integrated interventions have long been recommended for addressing mental health and its social determinants (e.g., gender-based violence) in settings of ongoing adversity. We developed an integrated health and protection intervention to reduce psychological distress and intimate partner violence (IPV), and tested its delivery by lay facilitators in a low-resource refugee setting. Formative research to develop the intervention consisted of a structured desk review, consultation with experts and local stakeholders (refugee incentive workers, representatives of humanitarian agencies, and clinical experts), and qualitative interviews (40 free list interviews with refugees, 15 key informant interviews). Given existing efforts by humanitarian agencies to prevent gender-based violence in this particular refugee camp, including with (potential) perpetrators, we focused on a complementary effort to develop an integrated intervention with potential to reduce IPV and associated mental health impacts with female IPV survivors. We enrolled Congolese refugee women with elevated psychological distress and past-year histories of IPV (n = 60) who received the intervention delivered by trained and supervised lay refugee facilitators. Relevance, feasibility and acceptability of the intervention were evaluated through quantitative and qualitative interviews with participants. We assessed instrument test-retest reliability (n = 24), inter-rater reliability (n = 5 interviews), internal consistency, and construct validity (n = 60). We designed an 8-session intervention, termed Nguvu ('strength'), incorporating brief Cognitive Processing Therapy (focused on helping clients obtaining skills to overcome negative thoughts and self-perceptions and gain control over the impact these have on their lives) and Advocacy Counseling (focused on increasing autonomy, empowerment and strengthening linkages to community supports). On average, participants attended two-thirds of the sessions. In qualitative interviews, participants recommended adaptations to specific intervention components and provided recommendations regarding coordination, retention, safety concerns and intervention participation incentives. Analysis of the performance of outcome instruments overall revealed acceptable reliability and validity. We found it feasible to develop and implement an integrated, multi-sectoral mental health and IPV intervention in a refugee camp setting. Implementation challenges were identified and may be informative for future implementation and evaluation of multi-sectoral strategies for populations facing ongoing adversity. ISRCTN65771265, June 27, 2016.


Nanoscale spatial limitations of large-area substrate conformal imprint lithography.
We demonstrate a soft-imprint nanofabrication technique offering nanometer resolution over an area as large as a 150 mm diameter wafer. It makes use of a composite imprint stamp composed of a quaternary siloxane-modified poly-di-methyl-siloxane patterned rubber layer with a relatively high Young's modulus that is laminated on a thin glass support. The in-plane stiffness of the stamp avoids pattern deformation over large areas, while out-of-plane flexibility allows conformal contact to be made over the entire substrate area. The stamp is used in conjunction with a novel tetra-methyl-ortho-siloxane/methyl-tri-methoxy-siloxane sol-gel imprint resist material developed to replicate nanoscale features in rigid silica at room temperature. We demonstrate better than 10 nm resolution in imprinted line gratings and individual pillars with aspect ratio as high as 5:1. Gaps as small as 6 nm can be reproduced. The patterns can be used as an etch mask to pattern 150 mm diameter silicon and quartz substrates while maintaining sub-10 nm resolution.


High-intensity intermittent activities at school: controversies and facts.
In comparison to continuous aerobic type activity, little is known about high-intensity intermittent physical activity in children. Repeated short-term high-intensity activities (> maximal aerobic speed and <10 s) are more characteristic of the spontaneous physical activity of children. Recent studies have shown during repetitive bouts of sprints separated by short recovery intervals, that prepubescent children compared with adults are more able to maintain their performance without substantial fatigue. Moreover, repetitive runs at high velocities (near and higher than the maximal aerobic speed) separated by short recovery periods may elicit a high oxygen consumption in children. Several studies using interval training programmes for 7 weeks, twice a week for 30 min in physical education lessons showed that children's aerobic performance (maximal O2 uptake, maximal aerobic speed) could be enhanced. Training based on these repeated short-term high-intensity exercises could also improve children's anaerobic performance (short-term muscle power, strength and speed). Current evidence suggests that recovery from high-intensity exercises is faster in children than in adults and that repeated runs at high velocities separated by short recovery intervals can improve both aerobic and anaerobic performance. Although continuous aerobic type activity is more scientifically established as a training mode, repeated short-term high-intensity exercises in physical education programmes should be considered to enhance aerobic, as well as, anaerobic fitness in children.


Infant feeding practices and asthma in children aged 6 months to 5 years using a propensity score approach.
We examined the association between exclusive breastfeeding, early introduction of feeding formula, early weaning, and asthma in children aged six months to five years in a sample of non-institutionalized US children using a propensity score approach. Our study used data from the National Survey of Children's Health (2012-2018) of 3,820 children with physician-diagnosed asthma aged 6 months to 5 years. Propensity score matching (PSM) was applied to control selection bias with age, sex, race, birth weight, Federal Poverty Level, parent's education, and parent smoking history used as covariates in PSM. The total number in the matched sample was 6,904 (3,452 non-asthmatics; 3,452 asthmatics). Matched and unmatched samples were analysed using the χ2 test and multiple logistic regression. Exclusive breastfeeding was protective against asthma in the pre-matching (AOR 0.72; 95% CI: 0.54-0.97; p = 0.03) and post-matching (AOR 0.66; 95% CI: 0.55-0.81; p < 0.001) samples. Formula feeding before 6 months was associated with asthma in unmatched (AOR 1.38; 95% CI: 1.15-1.66; p < 0.001) and matched (AOR 1.31; 95% CI: 1.16-1.47; p < 0.001) sample. Early weaning before 6 months was associated with asthma in unmatched (AOR 1.62; 95% CI: 1.35-1.54; p < 0.001) and matched sample (AOR 1.37; 95% CI: 1.23-1.54; p < 0.001). Public health systems should continue to recommend the implementation of the World Health Organization exclusive breastfeeding guideline in developed countries. Asthma interventions in children under two years should continue to emphasize exclusive breastfeeding to reduce the incidence of infant asthma.


A multi-omics approach for understanding the effects of moderate wine consumption on human intestinal health.
The human gut is a highly diverse microbial ecosystem. Although showing a well-defined core of dominant taxa, an interindividual variability exists in microbiome arrangement patterns, and the presence and proportion of specific species, determining individual metabolic features-metabotypes-which govern the health effects of dietary interventions (i.e. polyphenol consumption). Starting with a 19-volunteer human intervention study, divided into low, medium, and high wine-polyphenol-metabolizers, we detected interindividual discrepancies on the effect of wine consumption in gut bacterial alpha-diversity, but a significant homogenization of beta-diversity among moderate wine consumers, independently of their metabotype. In addition, the abundance of key health-related taxa such as Akkermansia sp. increased after moderate wine intake in the group of high polyphenol-metabolizers. Regarding the metabolic activity, significant (p < 0.05) positive correlations in the production of SCFAs were observed after wine intake. Finally, we were able to correlate the microbiome and the metabolome of the three metabotypes, and to identify some metabolites-biomarker species, highlighting the genera Phascolarctobacterium, Pelotomaculum and Prevotella, as positively correlated with polyphenol concentration, and Prevotella, Zymophilus and Eubacterium as positively correlated with SCFAs concentration in faeces. Our results contribute to the evidence of the need of including the microbiome variable in personalized nutrition programs, as different metabotyes respond differently to dietary interventions.


Intrafollicular location of marginal zone/CD1d(hi) B cells is associated with autoimmune pathology in a mouse model of lupus.
Marginal zone (MZ) B cells contain a large number of autoreactive clones and the expansion of this compartment has been associated with autoimmunity. MZ B cells also efficiently transport blood-borne antigen to the follicles where they activate T cells and differentiate into plasma cells. Using the B6.NZM2410.Sle1.Sle2.Sle3 (B6.TC) model of lupus, we show that the IgM+ CD1d(hi)/MZ B-cell compartment is expanded, and a large number of them reside inside the follicles. Contrary to the peripheral B-cell subset distribution and their activation status, the intrafollicular location of B6.TC IgM+ CD1d(hi)/MZ B cells depends on both bone marrow- and stromal-derived factors. Among the factors responsible for this intrafollicular location, we have identified an increased response to CXCL13 by B6.TC MZ B cells and a decreased expression of VCAM-1 on stromal cells in the B6.TC MZ. However, the reduced number of MZ macrophages observed in B6.TC MZs was independent of the IgM+ CD1d(hi)/B-cell location. B7-2 but not B7-1 deficiency restored IgM+ CD1d(hi)/MZ B-cell follicular exclusion in B6.TC mice, and it correlated with tolerance to dsDNA and a significant reduction of autoimmune pathology. These results suggest that follicular exclusion of IgM+ CD1d(hi)/MZ B cells is an important B-cell tolerance mechanism, and that B7-2 signaling is involved in breaching this tolerance checkpoint.


The impact of open versus closed format ICU admission practices on the outcome of high risk surgical patients: a cohort analysis.
In the year 2000, the organizational structure of the ICU in the Zaandam Medical Centre (ZMC) changed from an open to a closed format ICU. The objective of this study was to evaluate the effect of this organizational change on outcome in high risk surgical patients. The medical records of all consecutive high risk surgical patients admitted to the ICU from 1996 to 1998 (open format) and from 2003 to 2005 (closed format), were reviewed. High-risk patients were defined according to the Identification of Risk in Surgical patients (IRIS) score. Parameters studied were: mortality, morbidity, ICU length of stay (LOS) and hospital LOS. Mortality of ICU patients was 25.7% in the open format group and 15.8% in the closed format group (p = 0.01). Morbidity decreased from 48.6% to 46.1% (p = 0.6). The average length of hospital stay was 17 days in the open format group, and 21 days in the closed format group (p = 0.03). High risk surgical patients in the ICU are patients that have undergone complex and often extensive surgery. These patients are in need of specialized treatment and careful monitoring for maximum safety and optimal care. Our results suggest that closed format is a more favourable setting than open format to minimize the effects of high risk surgery, and to warrant safe outcome in this patient group.


Calcitriol and the parathyroid hormone-ionized calcium curve: a comparison of methodologic approaches.
Investigations of the effects of calcitriol on the inverse sigmoidal relationship between parathyroid hormone (PTH) and ionized calcium (iCa) have yielded contradictory conclusions, possibly because of variations in experimental and analytical approaches. To clarify the existing literature, PTH-iCa curves were constructed by inducing hypo- and hypercalcemia through alterations in dialysate calcium concentration in eight hemodialysis patients with mild to moderate secondary hyperparathyroidism. The effects of low (metabolic acidosis) versus normal bicarbonate dialysis were compared before and after 4 wk of intravenous calcitriol, in a cross-over design. The PTH-iCa curves were primarily evaluated by using a four-parameter model. In addition, a variety of alternative published analytic approaches were examined and PTH-iCa curve slopes were further evaluated after normalization by (maximum - minimum PTH). The latter partially corrects for gland mass and cell-secretory capacity, and therefore yields a purer measure of the sensitivity of secretory activity to changes in iCa. The results of the study indicate that calcitriol decreased basal, maximal, and minimal PTH and non-normalized slope (all P < 0.05), but did not affect set point or normalized slope, independent of the specific analytic approach. Acute metabolic acidosis did not affect the PTH-iCa curve. Thus, intravenous calcitriol appears to decrease parathyroid gland functional mass, as reflected by decreases in maximal and minimal PTH levels, but does not affect the sensitivity of the parathyroid gland to changes in iCa, as set point and normalized slope were unaffected.


Hysterography in patients with suspected uterine cancer: radiographic and histologic correlations and clinical implications.
The present study correlated the hysterographic and nonradiation-affected uterine histologic findings in 91 patients with suspected uterine cancer. Hysterography helped diagnose correctly a benign or malignant lesion, and helped identify the correct primary site of a malignant lesion in 11 patients. In 65 patients with a final diagnosis of endometrial carcinoma, all five cases of true endocervical involvement were identified, 69% were determined to have either no residual tumor or no myometrial invasion, and 88% were predicted correctly to have a myometrial invasion of greater than one-half. Prediction of the volume, distribution, and point of maximum invasion permitted the uterus to be opened so as to permit the best histologic assessment of the depth and extent of the lesion.


A computer touch screen system and training procedure for use with primate infants: Results from pigtail monkeys (Macaca nemestrina).
Computerized cognitive and perceptual testing has resulted in many advances towards understanding adult brain-behavior relations across a variety of abilities and species. However, there has been little migration of this technology to the assessment of very young primate subjects. We describe a training procedure and software that was developed to teach infant monkeys to interact with a touch screen computer. Eighteen infant pigtail macaques began training at 90-postnatal days and five began at 180-postnatal days. All animals were trained to reliably touch a stimulus presented on a computer screen and no significant differences were found between the two age groups. The results demonstrate the feasibility of using computers to assess cognitive and perceptual abilities early in development.


ME3277, a GPIIb/IIIa antagonist reduces cerebral infarction without enhancing intracranial hemorrhage in photothrombotic occlusion of rabbit middle cerebral artery.
GPIIb/IIIa antagonists are expected to have a beneficial effect on acute cerebral infarction, however, the occurrence of intracranial hemorrhage has not been as widely investigated. A rabbit focal thrombotic occlusion model of the middle cerebral artery was established by creating a photochemical reaction between green light and Rose Bengal. Hemorrhagic transformation was common in the area of cerebral infarction. Using this model, the effect of a GPIIb/IIIa antagonist, ME3277 (low dose, (L); 0.15 mg/kg + 0.125 mg/kg x h, middle dose, (M); 0.3 mg/kg + 0.25 mg/kg x h and high dose, (H); 0.6 mg/kg + 0.5 mg/kg x h), aspirin (20 mg/kg) and sodium ozagrel (thromboxane A2 synthase inhibitor, 1 mg/kg + 2 mg/ kg x h) were evaluated. Drugs were intravenously administrated 30 minutes after the photochemical reaction for 24 hours. Aspirin inhibited the ex vivo platelet aggregation induced by arachidonic acid and collagen but not by adenosine diphosphate (ADP), while sodium ozagrel only inhibited the arachidonic acid-induced aggregation. ME3277 dose-dependently inhibited the platelet aggregation induced by all the inducers (approximately 60% in L, 80% in M, and 90% in H). At 24 hours of middle cerebral artery (MCA) occlusion, infarct volume was significantly reduced by aspirin and each dose of ME3277. These agents improved neurologic deficits, with ME3277 being more potent than aspirin. Sodium ozagrel did not alter the infarct volume nor neurologic deficits. No drug was found to worsen hemorrhage volume despite increasing bleeding time (2-3 fold) in the skin. In this model, the occluded artery was spontaneously recanalized and re-thrombosed frequently. One mechanism by which antiplatelet agents reduced infarct volume was inhibition of rethrombosis of the MCA. These results suggest that treatment with a GPIIb/IIIa antagonist is a useful intervention for acute cerebral infarction prolonging dose bleeding time to 3 times the basal value.


Localization of a human reduced folate carrier protein in the mitochondrial as well as the cell membrane of leukemia cells.
IgG polyclonal antiserum was generated in New Zealand White rabbits immunized with a 16-mer peptide consisting of a specific amino acid sequence at residues corresponding to the sixth to seventh predicted transmembrane domain of the human reduced folate carrier (RFC). Using Western immunoblotting to examine the cytosolic and membrane fractions of the human CCRF-CEM T-cell lymphoblastic leukemia cell line, polyclonal antihuman RFC antiserum recognized two bands in the cytosolic fraction (approximately 60 kDa and approximately 70 kDa) on 10% polyacrylamide gels. In the membrane fraction, an approximately 60-kDa protein was identified. Comparative studies of a panel of human tumor cell lines including the HT1080 fibrosarcoma, 8805 malignant fibrous histiocytoma, and the MCF breast cancer cell lines revealed similar findings. Likewise, a recombinant approximately 60-kDa membrane protein was identified after expression of baculovirus-infected Sf9 insect cells containing cDNA of the human RFC. In the CEM-7A cell line, a variant of the CCRF-CEM cell line that overexpresses the RFC, 21-fold overexpression of the approximately 60-kDa membrane protein (RFC) was shown by Western analysis. To characterize further the cellular distribution of the human RFC, immunohistochemical analyses were performed in CCRF-CEM T-cell lymphoblastic leukemia cells. Predominantly membrane localization of the antibody reacting sites was detected; however, a cytoplasmic component was noted as well. By confocal microscopy and by immunogold electron microscopy, the cytoplasmic expression was found to be largely of mitochondrial origin. These findings were corroborated by Western immunoblotting of mitochondrial membrane isolates from the CCRF-CEM cell line, which demonstrate an approximately 60-kDa protein. The localization of the human RFC to the mitochondrial membrane is a novel finding, and it suggests a role for the mitochondrial membrane in the transport of folates.


Iteration of Partially Specified Target Matrices: Application to the Bi-Factor Case.
The current study proposes a new bi-factor rotation method, Schmid-Leiman with iterative target rotation (SLi), based on the iteration of partially specified target matrices and an initial target constructed from a Schmid-Leiman (SL) orthogonalization. SLi was expected to ameliorate some of the limitations of the previously presented SL bi-factor rotations, SL and SL with target rotation (SLt), when the factor structure either includes cross-loadings, near-zero loadings, or both. A Monte Carlo simulation was carried out to test the performance of SLi, SL, SLt, and the two analytic bi-factor rotations, bi-quartimin and bi-geomin. The results revealed that SLi accurately recovered the bi-factor structures across the majority of the conditions, and generally outperformed the other rotation methods. SLi provided the biggest improvements over SL and SLt when the bi-factor structures contained cross-loadings and pure indicators of the general factor. Additionally, SLi was superior to bi-quartimin and bi-geomin, which performed inconsistently across the types of factor structures evaluated. No method produced a good recovery of the bi-factor structures when small samples (N = 200) were combined with low factor loadings (0.30-0.50) in the specific factors. Thus, it is recommended that larger samples of at least 500 observations be obtained.


The reinforcement of short interresponse times.
Five contingencies were superimposed successively on a variable-interval schedule of reinforcement. In each of the resulting conditions, a different short, interresponse time was reinforced and an interresponse-time distribution was obtained from each of three pigeons. The lower bound of the reinforced interresponse times ranged from 0.3 to 2.4 sec. The resulting distributions were combined, according to a rationale based upon concurrent operants, induction, and a property of variable-interval schedules, to describe the interresponse-time distributions from a variable-interval schedule.


Ovarian Cancer Metastasis to the Breast: A Case Report and Review of the Literature.
Although ovarian cancer often presents as a widespread disease, metastases to the breast and/or axillary lymph nodes are a very rare event, accounting for only 0.03-0.6% of all breast cancers. Its early recognition and accurate distinction from primary breast cancer are of crucial importance to choose an adequate systemic therapy over unnecessary surgeries. We presented the case of a 53-year-old woman who was diagnosed with breast metastases 2 years after the diagnosis of advanced primary serous ovarian cancer. The patient underwent primary cytoreductive surgery and platinum-based chemotherapy in combination with bevacizumab, followed by bevacizumab maintenance for 18 months. After 2 years of negative follow-ups, the disease unexpectedly spread to the left breast and axillary lymph nodes. No axillary lymph node dissection or breast surgery was performed. The patient received axillary radiotherapy and multiple chemotherapy lines: gemcitabine/cisplatin, liposomal doxorubicin, topotecan, olaparib/cediranib, paclitaxel, and cisplatin. Unfortunately, none of these treatments improved her prognosis and she died 3 years after the disease recurrence. Ovarian cancer metastasis to the breast reveals a disseminated disease with a poor prognosis. Currently, no valid treatment options are available as the disease shows multidrug chemoresistance. In the era of precision medicine, the characterization of genetic and molecular markers may play a role in offering new promising targeted therapies.


Effect of terminal amino acids on the stability and specificity of PNA-DNA hybridisation.
The effect of various charged or hydrophobic amino acids on the hybridisation of fully complementary and mismatch PNA-DNA duplexes was investigated via UV melting curve analysis. The results described here show that the thermal stability and binding specificity of PNA probes can be modified by conjugation to amino acids and these effects should be considered in experimental design when conjugating PNA sequences to solubility enhancing groups or cell transport peptides. Where stabilisation of a duplex is important, without there being a corresponding need for specific binding to fully complementary targets, the conjugation of multiple lysine residues to the C-terminus of PNA may be the best probe design. If, however, the key is to obtain maximum discrimination between fully complementary and mismatch targets, a replacement of glutamic acid for lysine as the routine solubility enhancing group is recommended.


Denosumab: A comprehensive review.
The clinical sequelae from bone metastases, termed skeletal-related events (SREs), are among the most frequent and debilitating complications in patients with advanced cancer. Bone metastases are characterized by pathologically increased osteoclast activity, and accumulating evidence indicates that tumor cells interact within the bone to stimulate the receptor activator of nuclear factor kB (RANK)-RANK ligand (RANKL) pathway. RANKL is an essential mediator of osteoclast formation, function, and survival. Because of the central role of RANKL in cancer-induced bone destruction, the inhibition of RANKL has the potential to result in the reduction of pathologic bone resorption. Denosumab is a fully human monoclonal antibody specific for RANKL that inhibits the formation, activation, and survival of osteoclasts. This in turn decreases bone resorption and reduces cancer-induced bone destruction. In this review, we give an overview of the drug Denosumab with its history, mechanism of action, clinical trial data, adverse effects, and future challenges.


The formation of cholest-5-ene-3 ,26-diol as an intermediate in the conversion of cholesterol into bile acids by liver mitochondria.
1. When [(14)C]cholesterol was incubated with rat liver mitochondria, radioactive 26-hydroxycholesterol, 3beta-hydroxychol-5-enoic acid and other bile acids were isolated from the incubation mixture. 2. In the absence of added 26-hydroxycholesterol, the specific radioactivity of the 26-hydroxycholesterol formed from [(14)C]cholesterol during the incubation was higher than that of the 3beta-hydroxychol-5-enoic acid. Addition of increasing amounts of 26-hydroxycholesterol led to a progressive fall in the specific radioactivity, and to a progressive increase in the mass, of the 3beta-hydroxychol-5-enoic acid recovered at the end of the incubation. 3. It is concluded that 26-hydroxycholesterol is an intermediate in the formation of 3beta-hydroxychol-5-enoic acid from cholesterol. 4. Comparison of the specific radioactivity of the 26-hydroxycholesterol formed in the incubation mixture with that of the added [(14)C]cholesterol indicates that endogenous cholesterol in mitochondria is accessible to cholesterol 26-hydroxylase.


Crossing borders to bind proteins--a new concept in protein recognition based on the conjugation of small organic molecules or short peptides to polypeptides from a designed set.
A new concept for protein recognition and binding is highlighted. The conjugation of small organic molecules or short peptides to polypeptides from a designed set provides binder molecules that bind proteins with high affinities, and with selectivities that are equal to those of antibodies. The small organic molecules or peptides need to bind the protein targets but only with modest affinities and selectivities, because conjugation to the polypeptides results in molecules with dramatically improved binder performance. The polypeptides are selected from a set of only sixteen sequences designed to bind, in principle, any protein. The small number of polypeptides used to prepare high-affinity binders contrasts sharply with the huge libraries used in binder technologies based on selection or immunization. Also, unlike antibodies and engineered proteins, the polypeptides have unordered three-dimensional structures and adapt to the proteins to which they bind. Binder molecules for the C-reactive protein, human carbonic anhydrase II, acetylcholine esterase, thymidine kinase 1, phosphorylated proteins, the D-dimer, and a number of antibodies are used as examples to demonstrate that affinities are achieved that are higher than those of the small molecules or peptides by as much as four orders of magnitude. Evaluation by pull-down experiments and ELISA-based tests in human serum show selectivities to be equal to those of antibodies. Small organic molecules and peptides are readily available from pools of endogenous ligands, enzyme substrates, inhibitors or products, from screened small molecule libraries, from phage display, and from mRNA display. The technology is an alternative to established binder concepts for applications in drug development, diagnostics, medical imaging, and protein separation.


The relationships between dimensions of hostility and cardiovascular reactivity as a function of task characteristics.
The present study examined the independent relationships between dimensions of hostility and cardiovascular responses to a laboratory task with and without harassment. Fifty-three males, aged 18 to 26, with a negative parental history of cardiovascular disease were selected on the basis of their scores on the Cook and Medley Hostility (Ho) scale (greater than 24 or less than 14). Factor-analysis of six separate measures of hostility/anger resulted in a two-factor solution; Factor 1 representing antagonistic hostility and Factor 2 representing neurotic hostility. Results showed that high factor scores on antagonistic hostility were significantly associated with greater systolic blood pressure (SBP) and forearm blood flow (FBF) changes and poorer SBP recovery to harassment. In addition, high factor scores on neurotic hostility significantly predicted greater FBF changes to harassment. Additional correlational analysis showed that cardiovascular responses were positively associated with self-reported negative affects but only for subjects with high scores on either dimension. These results are in agreement with recent evidence suggesting that only antagonistic hostility may be related to increased severity of coronary artery disease and that the degree of interpersonal conflict moderates the association between coronary-prone behaviors and cardiovascular responses.


Predicting fine-scale distributions of peripheral aquatic species in headwater streams.
Headwater species and peripheral populations that occupy habitat at the edge of a species range may hold an increased conservation value to managers due to their potential to maximize intraspecies diversity and species' adaptive capabilities in the context of rapid environmental change. The southern Appalachian Mountains are the southern extent of the geographic range of native Salvelinus fontinalis and naturalized Oncorhynchus mykiss and Salmo trutta in eastern North America. We predicted distributions of these peripheral, headwater wild trout populations at a fine scale to serve as a planning and management tool for resource managers to maximize resistance and resilience of these populations in the face of anthropogenic stressors. We developed correlative logistic regression models to predict occurrence of brook trout, rainbow trout, and brown trout for every interconfluence stream reach in the study area. A stream network was generated to capture a more consistent representation of headwater streams. Each of the final models had four significant metrics in common: stream order, fragmentation, precipitation, and land cover. Strahler stream order was found to be the most influential variable in two of the three final models and the second most influential variable in the other model. Greater than 70% presence accuracy was achieved for all three models. The underrepresentation of headwater streams in commonly used hydrography datasets is an important consideration that warrants close examination when forecasting headwater species distributions and range estimates. Additionally, it appears that a relative watershed position metric (e.g., stream order) is an important surrogate variable (even when elevation is included) for biotic interactions across the landscape in areas where headwater species distributions are influenced by topographical gradients.


Performance appraisal: a primer for the lower level health care and rehabilitation worker.
This article reviews the use of performance appraisal in health care and rehabilitation organizations, particularly as such an evaluation measure could be used to examine the results achieved by managerial and supervisory level personnel. This entails a reversal of the prevailing view, that is, appraisal of lower level workers by "superiors" to a counter view as seen from those at "lower levels" looking upwards. Hierarchies, of course, are facing increasing challenges. Performance appraisal can serve the needs of the people at the lower end as well as those defined as the "management." Ultimately, performance appraisal can assure that all people in an organization will achieve benefits, not just those at the managerial level.


Transcriptome analysis of the ischemia-reperfused remodeling myocardium: temporal changes in inflammation and extracellular matrix.
cDNA microarray analysis was performed to screen 15,000 genes and expressed sequence tags (ESTs) to identify changes in the ischemia-reperfused (I-R) rat myocardial transcriptome in the early (day 2) and late (day 7) inflammatory phases of acute myocardial infarction. Lists of candidate genes that were affected by I-R transiently (2 or 7 days only) or on a more sustained basis (2 and 7 days) were derived. The candidate genes represented three major functional categories: extracellular matrix, apoptosis, and inflammation. To expand on the findings from microarray studies that dealt with the two above-mentioned time points, tissues collected from days 0, 0.25, 2, 3, 5, and 7 after reperfusion were examined. Acute myocardial infarction resulted in upregulation of IL-6 and IL-18. Genes encoding extracellular matrix proteins such as types I and III collagen were upregulated in day 2, and that response progressively grew stronger until day 7 after I-R. Comparable response kinetics was exhibited by the candidate genes of the apoptosis category. Caspases-2, -3, and -8 were induced in response to acute infarction. Compared with the myocardial tissue from the sham-operated rats, tissue collected from the infarct region stained heavily positive for the presence of active caspase-3. Laser microdissection and pressure catapulting technology was applied to harvest infarct and adjacent noninfarct control tissue from a microscopically defined region in the rat myocardium. Taken together, this work presents the first evidence gained from the use of DNA microarrays to understand the molecular mechanisms implicated in the early and late inflammatory phases of the I-R heart.


The Relationship Between Human Papillomavirus, OFD1 and Primary Ciliogenesis in the Progression of Oropharyngeal Cancer: A Retrospective Cohort Study.
Infection with human papillomavirus (HPV) has been indicated to be a important risk factor for oropharyngeal squamous cell carcinoma (OPSCC). Primary ciliogenesis defects contribute to tumorigenesis, and OFD1 at centriolar satellites is a crucial suppressor of primary ciliogenesis. To identify novel markers associated with HPV-induced carcinogenesis, the interactions between HPV infection and primary ciliogenesis in the tumorigenesis and progression of OPSCC were investigated in this study. The 1530 OPSCC patients recruited in this research were treated from 2000 to 2017. Immunohistochemistry and RT-PCR were performed on tissue samples to compare the expression of p16, TSLP, TGFβ1, IFNγ, OFD1, and their relationship with clinical characteristics of patients. We speculate that the positive expression of p16 is related to early primary OPSCC, and the survival rate of p16 positive patients after radiotherapy and surgery is higher. Expression of TSLP on dendritic cells in HPV-positive OPSCC correlated with the expression of OFD1. HPV-positive OPSCC showed increased expression of OFD1 combined with reduced ciliogenesis. Hence, TSLP induced by HPV infection may reduce the invasive potential of OPSCC cells by promoting OFD1 expression, thereby inhibiting primary ciliogenesis. Our study demonstrated that HPV may be related to the progression of OPSCC by regulating OFD1 expression and primary ciliogenesis, making this protein a potential therapeutic target.


Neuroendocrine carcinomas (carcinoid tumor) of the testis. A clinicopathologic and immunohistochemical study of ten cases.
We studied 10 cases of primary pure testicular neuroendocrine carcinoma. Patients were between 16 and 48 years old and had testicular swelling with pain or a painless testicular mass and no history of neuroendocrine carcinoma or other malignant neoplasm. All underwent orchiectomy. The tumors were low (n = 9) and intermediate (n = 1) grades with a variegated histologic appearance characterized by a nesting pattern, cords of neoplastic cells with rosettes, or sheets of neoplastic cells. Mitotic activity was lacking in 9 cases. In 1 case, mitotic figures ranged from 7 to 8 per 10 high-power fields, and cellular atypia and comedo-like necrosis were present. Immunohistochemical studies using a keratin cocktail, chromogranin, synaptophysin, epidermal growth factor, p53, placental-like alkaline phosphatase, and CD117 (c-kit) were performed in all cases. Keratin, chromogranin, and synaptophysin were positive in all tumors. Clinical follow-up information was obtained for 6 patients (range, 12-60 months): 5 with low-grade tumors were alive 24 to 60 months after diagnosis; 1 with an intermediate-grade tumor died of tumor 12 months after initial diagnosis. The behavior of these tumors, while in the testicular region, correlates well with the histologic grade. We propose replacing the term testicular carcinoid with neuroendocrine carcinoma, which better reflects the nature of these neoplasms.


The enrichment of cancer stem cells using composite alginate/polycaprolactone nanofibers.
Cancer stem cells (CSCs) are potential platforms to high-throughput screen anti-cancer drugs. However, they are difficult to isolate from cancer cells. Therefore, we proposed to fabricate 3-D scaffolds for CSC enrichment. Alginate is a biocompatible polysaccharide with poor cell adhesion, whereas polycaprolactone (PCL) is relative cell adhesive. These two materials were coelectrospun as composite scaffolds. Cells collected from alginate and composite fibers demonstrated high stemness, epithelial-mesenchymal transition, invasion, drug resistance, and angiogenesis. Interestingly, cells collected from composite fibers with low ratio of PCL were significantly improved their CSC properties compared to those from pure alginate fibers because few PCL fibers spatially separated cell populations to concentrate CSCs. These results suggested that alginate fibers effectively enriched CSCs and composite fibers created an uneven microenvironment to regulate cell morphology and distribution, by which cell-cell interaction was thus manipulated. These tunable scaffolds are potential to isolate CSCs from different tissues to facilitate the cancer research.


Endoscopic subcutaneous neurovascular lower-extremity myofascial flap harvesting for genitourinary reconstruction.
Neurovascular myofascial flaps have been widely utilized when healthy tissues outside the anatomic confines of the pelvis are necessary for complex genitourinary reconstructions. Myofascial flaps have great potential in providing support and tone to the bladder neck in a model of total urinary incontinence. Two muscles were evaluated for minimally invasive harvesting utilizing an adult rabbit and human cadaveric model: the gracilis, because of its neurovascular supply and length, and the sartorius, because of its location. Three adult New Zealand White rabbits were prepared and draped in sterile fashion after general anesthesia. Bilateral 2-cm incisions were made over the origins of the gracilis and sartorius muscles. Dissection times were entered in a database on a personal computer for paired Student t-tests. Special techniques for dissection included utilization of a 5-mm subcutaneous dissector/retractor and reticulated scissors and dissector. Once the tissues were harvested, rotational angles of the functional arcs around the origins of the gracilis and sartorius flaps were measured, as were the lengths of the muscular flaps from the proximal pedicle to the free distal end. Initial positioning of the myofascial flap at the bladder neck was investigated in this nonsurvival study. Blood loss was estimated before the animals were humanely sacrificed. The possibilities of minimally invasive genitourinary reconstruction are numerous. Both flaps can be dissected endoscopically and have rotational angles allowing transfer to more cephalad positions in the groin or intra-abdominally. Further studies are needed to establish the efficacy of the myofascial graft for minimally invasive genitourinary reconstructions.


Analyses of the secretomes of Erwinia amylovora and selected hrp mutants reveal novel type III secreted proteins and an effect of HrpJ on extracellular harpin levels.
SUMMARY Erwinia amylovora is a plant pathogenic enterobacterium that causes fire blight disease of apple, pear and other rosaceous plants. A type III (T3) secretion system, encoded by clustered, chromosomal hrp genes (hypersensitive response and pathogenicity), is essential for infection, but only a few proteins are known that are secreted through this pathway (the T3 'secretome'). We developed an efficient protocol for purification and concentration of extracellular proteins and used it to characterize the T3 secretome of E. amylovora Ea273 by comparing preparations from the wild-type strain with those from mutants defective in hrp secretion, regulation, or in genes encoding putative T3-secreted proteins. Proteins were resolved by gel electrophoresis and identified using mass spectrometry and a draft sequence of the E. amylovora genome. Twelve T3-secreted proteins were identified, including homologues of known effector and helper proteins, and HrpJ, a homologue of YopN of Yersinia pestis. Several previously uncharacterized T3-secreted proteins were designated as Eops for Erwinia outer proteins. Analysis of the secretome of a non-polar hrpJ mutant demonstrated that HrpJ is required for accumulation of wild-type levels of secreted harpins. HrpJ was found to be essential for pathogenesis, and to play a major role in elicitation of the hypersensitive reaction in tobacco.


Supraglottitis in children: evaluation and management.
Supraglottitis is a life-threatening illness characterized by inflammation of the supraglottic structures including the epiglottis, arytenoids, aryepiglottic folds, and ventricular bands. This paper is a comprehensive review of the etiology, epidemiology, pathology, differential diagnosis, natural course, complications, and management of supraglottitis. A high index of suspicion is required to properly interpret the historical, physical, and radiologic findings. As soon as the diagnosis of supraglottitis is established, the airway sould be secured by either an endotracheal tube or tracheostomy followed by antibiotic and supportive therapy.


Murine peritoneal macrophages support murine cytomegalovirus replication.
Because there have been different conclusions regarding the susceptibility of murine macrophages to murine cytomegalovirus (MCMV) infection and replication, we have undertaken a detailed comparison of MCMV infection of macrophages with that of a permissive cell line, mouse embryo cells. Although both cell lines undergo productive infections with MCMV, there are marked differences in certain aspects of the viral replication which may account for some of the different conclusions regarding the MCMV cycle in macrophages. Although both cell lines produce MCMV after infection, the time course of the infection differed markedly between the cell types. Similarly, the proportion of infected macrophages that are releasing infection virions is much smaller than the proportion of a comparably infected mouse embryo cell culture. Tissue culture passage of MCMV first enhanced (after one passage) and then reduced the infectivity of the virus for macrophages in vitro. The delayed time course and lesser production at early intervals after infection of macrophage cultures could not be attributed to demonstrable inhibitors or to replication in contaminating fibroblasts in the macrophage cultures.


Investigation of longitudinal spatial coherence for electromagnetic optical fields.
For light fields, the coherence in longitudinal direction is governed by both the frequency spectra and angular spectra they possess. In this work, we develop and report a theoretical formulation to demonstrate the effect of the angular spectra of electromagnetic light fields in quantifying their longitudinal spatial coherence. The experimental results obtained by measuring the electromagnetic longitudinal spatial coherence and degree of cross-polarization of uniformly polarized light fields for different angular spectra validate the theoretical findings.


A Novel Multiparametric Drug-Scoring Method for High-Throughput Screening of 3D Multicellular Tumor Spheroids Using the Celigo Image Cytometer.
Three-dimensional (3D) tumor models have been increasingly used to investigate and characterize cancer drug compounds. The ability to perform high-throughput screening of 3D multicellular tumor spheroids (MCTS) can highly improve the efficiency and cost-effectiveness of discovering potential cancer drug candidates. Previously, the Celigo Image Cytometer has demonstrated a novel method for high-throughput screening of 3D multicellular tumor spheroids. In this work, we employed the Celigo Image Cytometer to examine the effects of 14 cancer drug compounds on 3D MCTS of the glioblastoma cell line U87MG in 384-well plates. Using parameters such as MCTS diameter and invasion area, growth and invasion were monitored for 9 and 3 d, respectively. Furthermore, fluorescent staining with calcein AM, propidium iodide, Hoechst 33342, and caspase 3/7 was performed at day 9 posttreatment to measure viability and apoptosis. Using the kinetic and endpoint data generated, we created a novel multiparametric drug-scoring system for 3D MCTS that can be used to identify and classify potential drug candidates earlier in the drug discovery process. Furthermore, the combination of quantitative and qualitative image data can be used to delineate differences between drugs that induce cytotoxic and cytostatic effects. The 3D MCTS-based multiparametric scoring method described here can provide an alternative screening method to better qualify tested drug compounds.


Transcatheter mitral valve replacement: an update.
A significant number of patients with mitral valve disease are at high to prohibitive risk for surgical repair or replacement. Transcatheter mitral valve interventions have evolved over the last few years. We review the recent growing evidence on transcatheter mitral valve replacement (TMVR). Tendyne is the first TMVR device commercially approved in Europe. All TMVR systems remain under investigation in the United States. Several early studies have reported the feasibility and safety outcomes for TMVR implanted via transapical or transseptal approach. TMVR using balloon expandable valve for degenerated bioprosthetic valves, surgical rings and mitral annular calcification appears feasible in selected patients. Early experience with TMVR systems is promising. Larger ongoing studies will help understand longer term outcomes and offer insights into patient selection.


Laser in situ keratomileusis for spherical hyperopia and hyperopic astigmatism using the NIDEK EC-5000 excimer laser.
To assess the efficacy, predictability, and safety of LASIK for the surgical correction of low to moderate hyperopia and hyperopic astigmatism using the NIDEK EC-5000 excimer laser. In a multicenter United States Food and Drug Administration (FDA) regulated study of LASIK, 7 centers enrolled 293 eyes with manifest refraction sphere that ranged from +0.50 to +6.00 diopters (D) with or without astigmatism up to 3.00 D. The intended outcome was plano in all eyes. Patients were treated bilaterally. One year postoperative outcomes are reported. The mean spherical equivalent refraction (MRSE) for all eyes changed from +3.51 +/- 1.45 [corrected] D (range: +0.50 to +6.63 D) preoperatively to +0.35 +/- 0.54 D (range: +1.63 [corrected] to +2.00 D) 1 year [corrected] Overall, 61% (170/279) of eyes achieved distance uncorrected visual acuity (UCVA) of 20/20 or better, 82% (228/279) of eyes saw 20/25 or better, and 99% (277/279) of eyes saw 20/40 or better. Refractive accuracy was demonstrated as 63.1% (176/279) of eyes achieved a MRSE within +/- 0.50 D and 90.3% (252/279) of eyes within +/- 1.00 D. Less than 2% (4/279) of eyes lost 2 lines of distance best spectacle-corrected visual acuity. Stability of refraction was demonstrated by 6 months, with a mean hyperopic shift of < 0.03 D from 3 to 6 months. The NIDEK EC-5000 corrected hyperopia and hyperopic astigmatism with UCVA, refractive accuracy, 1-year stability, and safety that surpassed all FDA criteria.


Characterization of short range DNA looping in endotoxin-mediated transcription of the murine inducible nitric-oxide synthase (iNOS) gene.
The local structural properties and spatial conformations of chromosomes are intimately associated with gene expression. The spatial associations of critical genomic elements in inducible nitric-oxide synthase (iNOS) transcription have not been previously examined. In this regard, the murine iNOS promoter contains 2 NF-kappaB binding sites (nt -86 and nt -972) that are essential for maximal transactivation of iNOS by LPS. Although AP-1 is commonly listed as an essential transcription factor for LPS-mediated iNOS transactivation, the relationship between AP-1 and NF-kappaB in this setting is not well studied. In this study using a model of LPS-stimulated ANA-1 murine macrophages, we demonstrate that short range DNA looping occurs at the iNOS promoter. This looping requires the presence of AP-1, c-Jun, NF-kappaB p65, and p300-associated acetyltransferase activity. The distal AP-1 binding site interacts via p300 with the proximal NF-kappaB binding site to create this DNA loop to participate in iNOS transcription. Other geographically distant AP-1 and NF-kappaB sites are certainly occupied, but selected sites are critical for iNOS transcription and the formation of the c-Jun, p65, and p300 transcriptional complex. In this "simplified" model of murine iNOS promoter, numerous transcription factors recognize and bind to various response elements, but these locales do not equally contribute to iNOS gene transcription.


Case Volume-Dependent Changes in Operative Morbidity following Free Flap Breast Reconstruction: A 15-Year Single-Center Analysis.
Operative morbidity is a common yet modifiable feature of complex surgical procedures. With increasing case volume, improvement in morbidity has been reported through designated procedural processes and greater repetition. Defined as a volume-outcome association, improvement in breast reconstruction morbidity with increasing free flap volume requires further characterization. A retrospective analysis was conducted among consecutive free flap patients using a two-microsurgeon model between January of 2002 and December of 2017. Patient demographics and operative characteristics were obtained from medical records. Complications including unplanned surgical intervention (take-back) and flap loss were obtained from prospectively kept databases. Individual surgeon operative volume was estimated by considering overall practice volume and correcting for the number of surgeons at any given time. During the study period, 3949 patients met inclusion criteria. A total of 6607 breasts underwent reconstruction with 6675 free flaps. Mean patient age was 50 ± 9.4 years and mean body mass index was 28.8 ± 5.0 kg/m2. Bilateral reconstruction was performed on 2633 patients (66.5 percent), with 4626 breasts (70.5 percent) reconstructed in the immediate setting. Overall, breast and donor-site complications were reported in 507 breasts (7.7 percent) and 607 cases (15.4 percent), respectively. Take-back was required in 375 cases (9.5 percent), with complete flap loss occurring in 57 cases (0.9 percent). Based on annual flaps per surgeon, the incidence of complications decreased with increasing volume (slope = -0.12; p = 0.056). Through procedural efficiency and execution of defined clinical processes using a two-microsurgeon model, increases in microsurgical breast reconstruction case volume result in decreased morbidity. Therapeutic, IV.


Action at the mu receptor is sufficient to explain the supraspinal analgesic effect of opiates.
Mu, delta and kappa opiate receptors have been implicated in the production of analgesia. In order to study the relative contributions of these receptor types to supraspinally mediated analgesia, apparent pA2 values and rank order potencies were determined for i.c.v. injected highly selective opioid agonists in the mouse using a thermal nociceptive test. Drugs used included the prototypical mu agonist morphine, putative mu agonists [D-Ala2, N-methyl-Phe4, Gly5-ol] enkephalin and BAM 22P, putative delta agonists [D-Pen2, D-Pen5] enkephalin, [D-Thr2, Thr6, Leu5] enkephalin and [D-Ser2, Leu5, Thr6] enkephalin and the putative kappa agonists [trans-3,4-dichloro-N-methyl-N[2-(1-pyrrolidinyl)-cyclohexyl]- benzeneacetamide methane sulfonate hydrate] and Dynorphin A (1-13). We were unable to demonstrate significant analgesic potencies for i.c.v. injected Dynorphin A (1-13) or BAM 22P in the absence of marked behavioral abnormalities. The rank order potency of the remaining compounds studied was found to be: [D-Ala2, N-methyl-Phe4, Gly5-ol] enkephalin greater than [D-Thr2, Thr6, Leu5] enkephalin greater than [D-Ser2, Leu5, Thr6] enkephalin greater than Morphine greater than [D-Pen2, D-Pen5] enkephalin greater than [trans-3, 4-dichloro-N-methyl-N[2-(1-pyrrolidinyl)-cyclohexyl]-benzeneacetamide methane sulfonate hydrate]. Apparent pA2 values of morphine, [D-Ala2, N-methyl-Phe4, Gly5-ol] enkephalin and [D-Pen2, D-Pen5] enkephalin in naloxone antagonism trials did not differ significantly. These results indicate that although both mu- and delta-selective ligands produce potent analgesia, a single receptor (mu) is sufficient to explain the supraspinal effects of opiate drugs.


Ethics and cancer: a survey of the literature.
We have identified 776 articles, books, book chapters, and letters published since 1945 dealing with both ethics and cancer. Of this number, 473 (61%) were discussions in medical and scientific journal articles; they were published in 188 journals, demonstrating the scattered nature of this literature. The recent increase of writing in ethics and cancer has been dramatic, the median year of publication being 1979. Although the majority of articles addressed more than one ethical issue, the most common topic was whether patients should be told the truth about their diagnosis and prognosis (found in 241 of the items identified). The ethics of cancer research has only recently assumed importance in the medical literature, the median year of publication being 1981. The great interest in the ethics of cancer research is evident in that 255 (33%) of the 776 items dealt with research, 104 of these focusing on randomized trials. Besides truth-telling and research, 78 other issues are being discussed, among them informed consent for adults (119 items), the physician's responsibility for psychologic management of patients (79 items) and their families (68 items), responsibilities of nurses (58 items), the use and testing of laetrile (55 items), and euthanasia (38 items). Surgery was discussed in 90 items, pediatrics in 71, and radiotherapy in 44. The authors of this literature relied upon at least nine identifiable ethical positions. Only a limited number of authors were aware of the philosophic literature and of the complexities inherent in defining such terms as "benefit," "harm," and "quality of life."


Comprehensive profiling of bioactive compounds in germinated black soybeans via UHPLC-ESI-QTOF-MS/MS and their anti-Alzheimer's activity.
Black soybeans contain several bioactive compounds and commonly consumed due to their health-related activities but rarely cultivated as edible sprouts. The present study investigated the changes that occurred during germination in two new genotypes black soybeans. Raw and germinated seeds were tested against in vitro Alzheimer's disease (AD) biomarkers, including oxidative stress, inflammatory factors and cholinesterase enzymes as well as γ-aminobutyric acid (GABA) levels. Sprouts significantly inhibited the cholinesterase enzymes and inflammatory factors (protein denaturation, proteinase and lipoxygenase) than seeds. An increase in phenolic, flavonoid and GABA (10-folds) content and antioxidant capacity (ABTS, DPPH, and FRAP) was observed in germinated seeds. However, anthocyanin content was decreased in sprouts. UHPLC-ESI-QTOF-MS2 metabolites profiling approach identified 22 compounds including amino acids, peptides, fatty acids, and polyphenols. Among identified compounds, daidzein, genistein, gallic acid, spermidine, L-asparagine, and L-lysine exhibited the highest increase after germination. The current study reveals that germination of black soybeans have promising potential to inhibit/prevent AD and can be used to develop functional foods.


Gestational diabetes and the risk of offspring obesity.
Intrauterine exposure to the metabolic alterations of maternal diabetes may increase the risk of later obesity. We determined whether offspring of mothers with diet-treated, gestational diabetes mellitus (GDM) have an increased risk of childhood obesity and examined the relationship between childhood obesity and metabolic markers of GDM. At a health maintenance organization in Seattle, WA, we reviewed medical records to obtain the life-time height and weight measurements of 524, 8- to 10-year-old children whose mothers had been screened for GDM. Maternal plasma glucose and triglyceride levels were obtained in midgestation 1 hour after ingestion of 50 g of glucose. Those with glucose screening levels >/=7.77 mmol/L (140 mg/dL) underwent a 3-hour, 100-g, oral glucose tolerance test to determine GDM status. Cord serum insulin levels also were obtained at birth. Obesity was defined as an average body mass index between 5 and 10 years of age at or above the 85th percentile for age and sex. The prevalence of obesity was 19% in the 58 offspring of mothers with diet-treated GDM and 24% in the 257 offspring of mothers with negative glucose screen values. There also was no difference in mean body mass index (adjusted for age and sex) between these two groups of offspring. Among all 524 offspring, there was no significant increase in the rate of offspring obesity according to the quartile of maternal screening glucose, triglyceride, oral glucose tolerance test, or cord serum insulin level. Prenatal exposure to the metabolic effects of mild, diet-treated GDM does not increase the risk of childhood obesity.


Selective potentiating effect of RS14203 on a serotoninergic pathway in anesthetized rats.
The usefulness of selective inhibitors of type 4 phosphodiesterase (PDE4) in the treatment of inflammation and pulmonary diseases is limited by their side effects: nausea and vomiting. We studied the effect of three structurally diverse PDE4 inhibitors on the vagal nerve afferent and efferent fibers in anesthetized rats. The effects of RS14203, (R)-rolipram, and CT-2450 were evaluated on the von Bezold-Jarisch reflex (vagal afferent fibers) and in a model of vagal electrical stimulation (vagal efferent fibers). All three PDE4 inhibitors were administered at 1, 10, or 100 microg/kg (iv) 15 min prior to the induction of bradycardia by an iv injection of 2-methyl-5-HT (von Bezold-Jarisch reflex) or by vagal electrical stimulation. At 100 microg/kg, RS14203 significantly potentiated the 2-methyl-5-HT response. No statistically significant effects were observed with (R)-rolipram or CT-2450 at the doses studied. RS14203, (R)-rolipram, or CT-2450 (1-100 microg/kg iv) did not affect the bradycardia induced by vagal electrical stimulation. Consequently, our results show that RS14203 selectively facilitates serotoninergic neurotransmission in vagal afferent fibers. The emetic action of RS14203 may be mediated by this mechanism.


High-Fidelity Identification of Single Nucleotide Polymorphism by Type V CRISPR Systems.
Accurate and sensitive detection of single nucleotide polymorphism (SNP) holds significant clinical implications, especially in the field of cancer diagnosis. Leveraging its high accuracy and programmability, the CRISPR system emerges as a promising platform for advancing the identification of SNPs. In this study, we compared two type V CRISPR/Cas systems (Cas12a and Cas14a) for the identification of cancer-related SNP. Their identification performances were evaluated by characterizing their mismatch tolerance to the BRAF gene. We found that the CRISPR/Cas14a system exhibited superior accuracy and robustness over the CRISPR/Cas12a system for SNP detection. Furthermore, blocker displacement amplification (BDA) was combined with the CRISPR/Cas14a system to eliminate the interference of the wild type (WT) and increase the detection accuracy. In this strategy, we were able to detect BRAF V600E as low as 103 copies with a sensitivity of 0.1% variant allele frequency. Moreover, the BDA-assisted CRISPR/Cas14a system has been applied to identify the BRAF mutation from human colorectal carcinoma cells, achieving a high sensitivity of 0.5% variant allele frequency, which is comparable to or even superior to those of most commercially available products. This work has broadened the scope of the CRISPR system and provided a promising method for precision medicine.


Effect of polysaccharide extract SPSS1 from Apostichopus japonicas spermary on HepG2 cells via iTRAQ-based proteome analysis.
In this study, polysaccharide extract was prepared from Apostichopus japonicus spermary and purified by ion-exchange chromatography and gel filtration chromatography. Two main fractions named SPSS1 and SPSS2 were obtained and analyzed by ultraviolet spectroscopy and mixed with KBr, respectively. Chemical components analysis proved that SPSS1 and SPSS2 were rich in sulfate. Monosaccharide analysis indicated that in addition to the high content of lactose in both kinds of polysaccharides, the highest content of monosaccharide in SPSS1 was galactose, while in SPSS2 it was fucose. Further, the antitumor study of SPSS1 was carried and the results showed that SPSS1 treatment inhibited the proliferation of HepG2 cells. Through the iTRAQ-based proteome analysis, there were 208 differential proteins between control tumor cells and SPSS1 treatment of tumor cells. Compared to control tumor cells, 135 proteins were upregulated and 73 proteins were downregulated in treatment tumor cells. PRACTICAL APPLICATIONS: Our study suggested that polysaccharide from sea cucumbers had the potential to be further developed as antitumor drugs.


Vitiligo associated with regional enteritis.
Although the cause of vitiligo is unknown, there is considerable evidence to indicate that autoimmunity plays a role. Much of this evidence is based on the increased associated occurrence of vitiligo in a number of diseases also believed to be autoimmune in origin. I have observed a case of almost simultaneous onset of vitiligo and regional enteritis. A search of the literature revealed no specific case report associating these two disorders. The association of these two disorders in the same person may be more than fortuitous, since both diseases may be autoimmune in origin.


A physician-pharmacist collaborative care model to prevent opioid misuse.
Clinical pharmacists in primary care clinics can potentially help manage chronic pain and opioid prescriptions by providing services similar to those provided within their scope of practice to patients with diabetes and hypertension. We evaluated the feasibility and acceptability of a pharmacist-physician collaborative care model for patients with chronic pain. The program consisted of an in-person pharmacist consultation and optional follow-up visits over 4 months in 2 primary care practices. Eligible patients had chronic pain and a long-term prescription for opioids or buprenorphine or were referred by their primary care physician (PCP). Pharmacist recommendations were communicated to PCPs via the electronic medical record (EMR) and direct communication. Mixed-methods evaluation included baseline and follow-up surveys with patients, EMR review of opioid-related clinical encounters, and provider interviews. Between January and October 2018, 47 of the 182 eligible patients enrolled, with 46 completing all follow-up; 43 patients (91%) had received opioids over the past 6 months. The pharmacist recommended adding or switching to a nonopioid pain medication for 30 patients, switching to buprenorphine for pain and complex persistent opioid dependence for 20 patients, and tapering opioids for 3 patients. All physicians found the intervention acceptable but wanted more guidance on prescribing buprenorphine for pain. Most patients found the intervention helpful, but some reported a lack of physician follow-up on recommended changes. The study demonstrated that comanagement of patients with chronic pain is feasible and acceptable. Policy changes to increase pharmacists' authority to prescribe may increase physician willingness and confidence to carry out opioid tapers and prescribe buprenorphine for pain.


The external ion microbeam of the LABEC laboratory in florence: some applications to cultural heritage.
Ion beam analysis (IBA) techniques are a powerful analytical tool used to investigate the composition and structure of precious materials principally because they can be applied in atmosphere. Thus, the sample can be analyzed as is, and heating and charging effects are strongly diminished. Since IBA measurements can be made with low ion currents and acquisition time, the damage risk is limited. At the microbeam line of the LABEC laboratory, it is possible to exploit the potentials of IBA techniques in an external set up to reconstruct the distribution maps of all the detected elements over the analyzed area with spatial resolutions as low as 10 μm. This is an important feature when objects with inhomogeneous structures-on a scale of hundred microns or so-are investigated, as happens in some cases with artworks. The detection set up installed on our external microbeam allows us to use different IBA techniques simultaneously. Thus, in a single measurement run, it is possible to obtain complementary information on both sample composition and structure. Some applications to works of art are presented here as examples of the analytical capabilities of the external scanning microbeam in the cultural heritage field.


The role of fat depletion in the biological benefits of caloric restriction.
One of the most robust observations in the biology of aging is that caloric restriction (CR) extends life in a variety of species. Although CR results in substantial decrease in fat mass, the role of fat in life extension was considered minimal. Indeed, in the fields of obesity and diabetes, the amount of fat has been directly implicated in the metabolic consequences. Since it became apparent that fat is a massive endocrine tissue, some of its roles have been recently revised. Many of the systemic effects of CR can now be explained by the chronic effects related to decreased plasma levels of peptides, cytokines, complement factors and substrates that are produced in fat. Most of the benefits of CR on the neuroendocrine system and those related to the improvement in glucose homeostasis can be attributed to a decrease in adipose cells and their products. If all or most of the life-extending benefits of CR can be attributed to decreased fat stores, the expression of specific candidate substrates and proteins may be explored and manipulated in searching for the most powerful adipose-dependent signals that modulate life expectancy.


A longitudinal examination of risky sexual behaviors among Canadian and Italian adolescents: Considering individual, parental, and friend characteristics.
In this study, two longitudinal models of early adolescent risky sexual behaviors (RSB) were compared using a pooled sample of 267 Canadian and Italian adolescents (55% females; 53% Canadians) assessed yearly from grade 8 to 10. We focused on parenting practices (monitoring, control, limit setting), adolescent problem behaviors (antisocial behaviors, substance use) and their friends' deviance (antisocial behaviors, substance use) as predictors of condom use frequency and lifetime number of sexual partners. The socialization model postulates that youths' problem behaviors and RSB are behaviors learned within the friendship network where deviancy training can occur. The selection model posits that delinquent youth tend to affiliate with each other, and that RSB is one of many behaviors that can form the basis of selection. Using structural equation modeling, this study showed that the socialization model was the most accurate to explain the emergence of RSB. A full mediation of parenting practices, passing through deviant friends and youths' problem behavior, was observed for condom use. The same process applied to number of sexual partners, but a direct effect for parenting practices was also found.


Afrocentric screening program for breast, colorectal, and cervical cancer among immigrant patients in Ontario.
Black and immigrant populations across Canada have lower screening rates than Canadian-born white populations, predisposing them to increased cancer morbidity and mortality. Effective interventions are required to increase cancer screening rates among these populations. To improve breast, colorectal, and cervical cancer screening rates at TAIBU Community Health Centre, which has a mandate to provide primary health care services to the Black and immigrant community in the greater Toronto area. An Afrocentric quality improvement program was developed and implemented, consisting of provider audits, cancer screening education programs, a patient call-back program, and a mammography promotion day. TAIBU Community Health Centre's continuous quality improvement approach was successful in engaging health care providers and patients to increase cancer screening participation sustainably in a racially and socioeconomically diverse setting. Rates of breast, colorectal, and cervical cancer screening offered to eligible patients increased from 17% to 72%, 18% to 67%, and 59% to 70%, respectively, between 2011 and 2018.


Weather and nodule mediated variations in delta 13C and delta 15N values in field-grown soybean (Glycine max L.) with special interest in the analyses of xylem fluids.
The nodulating soybean (Enrei) and its non-nodulating mutant (EN 1282) were grown in outdoor plots for 2 years (1994: extraordinary dry, high temperature, 1995: ordinary year). Carbon and nitrogen accumulation, delta 13C and delta 15N values in plant parts and xylem fluids and delta 15N values in the water-extractable soil N were analysed throughout the growing period. Plant growth in 1994 was rapid during the early growth stages, but no pods were produced. In 1995, plant growth was normal and pods were formed. The delta 13C values of the leaves were less negative in 1994 than in 1995 and the nodulated plants showed less negative delta 13C values than non-nodulated plants in both years. The delta 13C values of the leaves during the vegetative phase were positively correlated to the leaf N concentrations. Leaf N concentrations in their turn were influenced by nodulation and weather conditions and/or soil available N. The delta 15N values in the plants and xylem fluids were lower in the nodulated soybean than in non-nodulated soybean in both years, and estimates of the contribution of N2 fixation in nodulated plants based on plant top delta 15N values were 7-14% in 1994 and 37-63% in 1995. The delta 13C values of xylem fluids did not differ between nodulated and non-nodulated plants. Thus, the expected contribution by phosphopenolpyruvate carboxylase-mediated CO2 fixation in the root nodules to plant C-incorporation could not have been significant.


Aberrant projection induced by otocyst removal maintains normal tonotopic organization in the chick cochlear nucleus.
Nucleus magnocellularis (NM), a second-order nucleus in the chick auditory system, is topographically and tonotopically organized. The basilar papilla (cochlea) projects onto the ipsilateral NM via the auditory nerve. The anteromedial region of NM is innervated by the proximal end of the basilar papilla and responds to high-frequency sounds; more posterolateral regions receive input from more distal locations along the papilla and respond to progressively lower frequencies. NM projects exclusively to the third-order neurons of nucleus laminaris (NL). Otocyst removal prevents the formation of the ipsilateral cochlea and cochlear nerve and results in the development of an aberrant functional projection from the contralateral NM to the "deafferented" NM on the operated side of the brain (Jackson and Parks, 1988). In the present experiment, the otocyst was removed unilaterally and the tonotopic organization of the deafferented NM was physiologically mapped at 17-18 d of embryonic age (E17-E18). Quantitative analyses revealed that the frequency organization of the deafferented NM is almost identical to that in normal embryos. Progressively higher characteristic frequencies were recorded at successively more anterior and more medial locations in the nucleus, and the orientation of the tonotopic axis was indistinguishable from normal. Furthermore, the correlation between characteristic frequency and anatomical location is comparable in the deafferented (r = 0.91) and normal (r = 0.87) NM. The only noticeable discrepancy is that characteristic frequencies in NM on both sides of the brain of operated embryos are higher than the frequencies observed previously at comparable regions of the nucleus in unoperated controls.(ABSTRACT TRUNCATED AT 250 WORDS)


Initiation and change of psychotropic medication regimens among adolescents in inpatient care.
The purpose of this study was to compare clinical and service utilization profiles of adolescents admitted to inpatient treatment with and without a psychotropic medication regimen, and estimate correlates of medication use separately for the two groups. Comprehensive data on clinical characteristics and service utilization of 517 adolescents enrolled in Medicaid who were admitted to three inpatient hospitals (one for-profit and two nonprofit) in a mid-Atlantic state were used. Medication correlates were examined with bivariate statistics (chi-square and t-test) and multivariate logistic regressions. Psychotropic medication was prescribed for the majority (90.5%) of adolescents at discharge. During their inpatient stay, 76.3% of the youths who were not on medication at admission (n = 190) initiated medication use, and 45% admitted with prior medication (n = 327) had a medication change. Adolescents admitted with and without medication differed on all sociodemographic and clinical characteristics except age, suicidality, and hospital. Controlling on demographic, clinical, and service history characteristics, hospital setting was the only correlate significantly associated with both initiation and change. Institutional factors may be more important than clinical factors in appraising medication patterns in inpatient settings. Interventions to ensure the development and implementation of medication management guidelines may need to be tailored to the climate and culture of the provider organization as well as directly to individual practitioners, parents or adolescents themselves.


Upregulation of mitochondrial Nox4 mediates TGF-β-induced apoptosis in cultured mouse podocytes.
Injury to podocytes leads to the onset of chronic renal diseases characterized by proteinuria. Elevated transforming growth factor (TGF)-β in kidney tissue is associated with podocyte damage that ultimately results in apoptosis and detachment. We investigated the proapoptotic mechanism of TGF-β in immortalized mouse podocytes. Exogenous TGF-β1-induced podocyte apoptosis through caspase-3 activation, which was related to elevated ROS levels generated by selective upregulation of NADPH oxidase 4 (Nox4). In mouse podocytes, Nox4 was predominantly localized to mitochondria, and Nox4 upregulation by TGF-β1 markedly depolarized mitochondrial membrane potential. TGF-β1-induced ROS production and caspase activation were mitigated by an antioxidant, the Nox inhibitor diphenyleneiodonium, or small interfering RNA for Nox4. A TGF-β receptor I blocker, SB-431542, completely reversed the changes triggered by TGF-β1. Knockdown of either Smad2 or Smad3 prevented the increase of Nox4 expression, ROS generation, loss of mitochondrial membrane potential, and caspase-3 activation by TGF-β1. These results suggest that TGF-β1-induced mitochondrial Nox4 upregulation via the TGF-β receptor-Smad2/3 pathway is responsible for ROS production, mitochondrial dysfunction, and apoptosis, which may at least in part contribute to the development and progression of proteinuric glomerular diseases such as diabetic nephropathy.


Pharmacological characterization of a novel phosphodiesterase type 5 (PDE5) inhibitor lodenafil carbonate on human and rabbit corpus cavernosum.
Nitrergic nerves and endothelial cells release nitric oxide (NO) in the corpus cavernosum, a key mediator that stimulates soluble guanylyl cyclase to increase cGMP levels causing penile erection. Phosphodiesterase 5 (PDE5) inhibitors, such as sildenafil, prolong the NO effects by inhibiting cGMP breakdown. Here, we report a novel PDE5 inhibitor, lodenafil carbonate, (Bis-(2-{4-[4-ethoxy-3-(1-methyl-7-oxo-3-propyl-6,7-dihydro-1H-pyrazolo[4,3-d]pyrimidin-5-yl)-benzenesulfonyl]piperazin-1-yl}-ethyl)carbonate) that is a dimer of lodenafil. We therefore aimed to compare the effects of sildenafil, lodenafil and lodenafil carbonate on in vitro human and rabbit cavernosal relaxations, activity of crude PDE extracts from human platelets, as well as stability and metabolic studies in rat, dog and human plasma. Pharmacokinetic evaluations after intravenous and oral administration were performed in male beagles. Functional experiments were conducted using organ bath techniques. Pharmacokinetics was studied in beagles by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), following oral or intravascular administration. All PDE5 inhibitors tested concentration-dependently relaxed (0.001-100 microM) phenylephrine-precontracted rabbit and human corpus cavernosum. The cavernosal relaxations evoked by either acetylcholine (0.01-100 microM) or electrical field stimulation (EFS, 1-20 Hz) were markedly potentiated by sildenafil, lodenafil and lodenafil carbonate. Lodenafil carbonate was more potent to inhibit the cGMP hydrolysis in PDE extracts compared with lodenafil and sildenafil. Following intravascular and single oral administration of lodenafil carbonate, only lodenafil and norlodenafil were detected in vivo. These results indicate that lodenafil carbonate works as a prodrug, being lodenafil the active moiety of lodenafil carbonate.


Epigenome-wide association study reveals longitudinally stable DNA methylation differences in CD4+ T cells from children with IgE-mediated food allergy.
Food allergy is mediated by a combination of genetic and environmental risk factors, potentially mediated by epigenetic mechanisms. CD4+ T-cells are key drivers of the allergic response, and may therefore harbor epigenetic variation in association with the disease phenotype. Here we retrospectively examined genome-wide DNA methylation profiles (~450,000 CpGs) from CD4+ T-cells on a birth cohort of 12 children with IgE-mediated food allergy diagnosed at 12-months, and 12 non-allergic controls. DNA samples were available at two time points, birth and 12-months. control comparisons of CD4+ methylation profiles identified 179 differentially methylated probes (DMP) at 12-months and 136 DMP at birth (FDR-adjusted P value<0.05, delta β>0.1). Approximately 30% of DMPs were coincident with previously annotated SNPs. A total of 92 [corrected] allergy-associated non-SNP DMPs were present at birth when individuals were initially disease-free, potentially implicating these loci in the causal pathway. Pathway analysis of differentially methylated genes identified several MAP kinase signaling molecules. Mass spectrometry was used to validate 15 CpG sites at 3 candidate genes. Combined analysis of differential methylation with gene expression profiles revealed gene expression differences at some but not all allergy associated differentially methylated genes. Thus, dysregulation of DNA methylation at MAPK signaling-associated genes during early CD4+ T-cell development may contribute to suboptimal T-lymphocyte responses in early childhood associated with the development of food allergy.


Mixed-valent [FeIV(mu-O)(mu-carboxylato)2FeIII]3+ core.
The symmetrically ligated complexes 1, 2, and 3 with a (mu-oxo)bis(mu-acetato)diferric core can be one-electron oxidized electrochemically or chemically with aminyl radical cations [*NR3][SbCl6] in acetonitrile yielding complexes which contain the mixed-valent [(mu-oxo)bis(mu-acetato)iron(IV)iron(III)]3+ core: [([9]aneN3)(2FeIII2)(mu-O)(mu-CH3CO2)2](ClO4)2 (1(ClO4)2), [(Me3[9]aneN3)(2FeIII2)(mu-O)(mu-CH3CO2)2](PF6)2 (2(PF6)(2)), and [(tpb)(2FeIII2)(mu-O)(mu-CH3CO2)2] (3) where ([9]aneN3) is the neutral triamine 1,4,7-triazacyclononane and (Me3[9]aneN3) is its tris-N-methylated derivative, and (tpb)(-) is the monoanion trispyrazolylborate. The asymmetrically ligated complex [(Me3[9]aneN3)FeIII(mu-O)(mu-CH3CO2)2FeIII(tpb)](PF6) (4(PF6)) and its one-electron oxidized form [4ox]2+ have also been prepared. Finally, the known heterodinuclear species [(Me3[9]aneN3)CrIII(mu-O)(mu-CH3CO2)2Fe([9]aneN3)](PF6)2 (5(PF6)(2)) can also be one-electron oxidized yielding [5ox]3+ containing an iron(IV) ion. The structure of 4(PF6).0.5CH3CN.0.25(C2H5)2O has been determined by X-ray crystallography and that of [5ox]2+ by Fe K-edge EXAFS-spectroscopy (Fe(IV)-O(oxo): 1.69(1) A; Fe(IV)-O(carboxylato) 1.93(3) A, Fe(IV)-N 2.00(2) A) contrasting the data for 5 (Fe(III)-O(oxo) 1.80 A; Fe(III)-O(carboxylato) 2.05 A, Fe-N 2.20 A). [5ox]2+ has an St = 1/2 ground state whereas all complexes containing the mixed-valent [FeIV(mu-O)(mu-CH3CO2)2FeIII]3+ core have an St = 3/2 ground state. Mössbauer spectra of the oxidized forms of complexes clearly show the presence of low spin FeIV ions (isomer shift approximately 0.02 mm s(-1), quadrupole splitting approximately 1.4 mm s(-1) at 80 K), whereas the high spin FeIII ion exhibits delta approximately 0.46 mm s(-1) and DeltaE(Q) approximately 0.5 mm s(-1). Mössbauer, EPR spectral and structural parameters have been calculated by density functional theoretical methods at the BP86 and B3LYP levels. The exchange coupling constant, J, for diiron complexes with the mixed-valent FeIV-FeIII core (H = -2J S1.S2; S(1) = 5/2; S2 = 1) has been calculated to be -88 cm(-1) (intramolecular antiferromagnetic coupling) and for the reduced diferric form of -75 cm(-1) in reasonable agreement with experiment (J = -120 cm(-1)).


MicroRNA-181a promotes tumor growth and liver metastasis in colorectal cancer by targeting the tumor suppressor WIF-1.
Given the emerging role of microRNA in tumor disease progression, we investigated the association between microRNA expression, liver metastasis and prognosis of colorectal cancer. Colorectal cancer tissues from patients with or without liver metastases were profiled to identify differentially expressed microRNA. Expression profile was further assessed using quantitative reverse transcription PCR and in situ hybridization. Correlation between miR-181a expression, the most differentially expressed microRNA, between patients with and without liver metastasis, and its downstream target genes were investigated using qRT-PCR. Luciferase reporter assay was conducted to establish functional association between miR-181a and its target genes. Manipulation of miR-181a expression and its consequences in tumor growth and metastasis were demonstrated in various in vitro and in vivo models. miR-181a was revealed being the most elevated in CRC with liver metastases. miR-181a expression correlated with advanced stage, distant metastasis, and served as an independent prognostic factor of poor overall survival. Stable transfection of CRC cell lines with miR-181a promoted cell motility and invasion, as well as tumor growth and liver metastasis,while silencing its expression resulted in reduced migration and invasion. Additionally, we identified WIF-1 as direct and functional targets of miR-181a. Ectopic expression of miR-181a suppressed the epithelial markers E-cadherin and β-catenin, while enhanced the mesenchymal markers vimentin. Our data demonstrate that miR-181a expression is associated with CRC liver metastasis and survival. miR-181a has strong tumor-promoting effects through inhibiting the expression of WIF-1, and its potential role in promoting epithelial-mesenchymal transition.