The elementary steps of transcription as catalyzed by E. coli RNA polymerase during one and two rounds of the nucleotide addition cycle (NAC) were resolved in rapid kinetic studies. Modelling of ...stopped-flow kinetic data of pyrophosphate release in a coupled enzyme assay during one round of the NAC indicates that the rate of pyrophosphate release is significantly less than that for nucleotide incorporation. Upon modelling of the stopped-flow kinetic data for pyrophosphate release during two rounds of the NAC, it was observed that the presence of the next nucleotide for incorporation increases the rate of release of the first pyrophosphate equivalent; incorrect nucleotides for incorporation had no effect on the rate of pyrophosphate release. Although the next nucleotide for incorporation increases the rate of pyrophosphate release, it is still significantly less than the rate of incorporation of the first nucleotide. The results from the stopped-flow kinetic studies were confirmed by using quench-flow followed by thin-layer chromatography (QF-TLC) with only the first nucleotide for incorporation labeled on the gamma phosphate with .sup.32 P to monitor pyrophosphate release. Collectively, the results are consistent with an NTP-driven model for the NAC in which the binding of the next cognate nucleotide for incorporation causes a synergistic conformational change in the enzyme that triggers the more rapid release of pyrophosphate, translocation of the enzyme along the DNA template strand and nucleotide incorporation.
Global estimates of unsafe abortions have been produced for 1995, 2003, and 2008. However, reconceptualisation of the framework and methods for estimating abortion safety is needed owing to the ...increased availability of simple methods for safe abortion (eg, medical abortion), the increasingly widespread use of misoprostol outside formal health systems in contexts where abortion is legally restricted, and the need to account for the multiple factors that affect abortion safety.
We used all available empirical data on abortion methods, providers, and settings, and factors affecting safety as covariates within a Bayesian hierarchical model to estimate the global, regional, and subregional distributions of abortion by safety categories. We used a three-tiered categorisation based on the WHO definition of unsafe abortion and WHO guidelines on safe abortion to categorise abortions as safe or unsafe and to further divide unsafe abortions into two categories of less safe and least safe.
Of the 55· 7 million abortions that occurred worldwide each year between 2010–14, we estimated that 30·6 million (54·9%, 90% uncertainty interval 49·9–59·4) were safe, 17·1 million (30·7%, 25·5–35·6) were less safe, and 8·0 million (14·4%, 11·5–18·1) were least safe. Thus, 25·1 million (45·1%, 40·6–50·1) abortions each year between 2010 and 2014 were unsafe, with 24·3 million (97%) of these in developing countries. The proportion of unsafe abortions was significantly higher in developing countries than developed countries (49·5% vs 12·5%). When grouped by the legal status of abortion, the proportion of unsafe abortions was significantly higher in countries with highly restrictive abortion laws than in those with less restrictive laws.
Increased efforts are needed, especially in developing countries, to ensure access to safe abortion. The paucity of empirical data is a limitation of these findings. Improved in-country data for health services and innovative research to address these gaps are needed to improve future estimates.
UNDP/UNFPA/UNICEF/WHO/World Bank Special Programme of Research, Development and Research Training in Human Reproduction; David and Lucile Packard Foundation; UK Aid from the UK Government; Dutch Ministry of Foreign Affairs; Norwegian Agency for Development Cooperation.
Summary Background Information about the incidence of induced abortion is needed to motivate and inform efforts to help women avoid unintended pregnancies and to monitor progress toward that end. We ...estimate subregional, regional, and global levels and trends in abortion incidence for 1990 to 2014, and abortion rates in subgroups of women. We use the results to estimate the proportion of pregnancies that end in abortion and examine whether abortion rates vary in countries grouped by the legal status of abortion. Methods We requested abortion data from government agencies and compiled data from international sources and nationally representative studies. With data for 1069 country-years, we estimated incidence using a Bayesian hierarchical time series model whereby the overall abortion rate is a function of the modelled rates in subgroups of women of reproductive age defined by their marital status and contraceptive need and use, and the sizes of these subgroups. Findings We estimated that 35 abortions (90% uncertainty interval UI 33 to 44) occurred annually per 1000 women aged 15–44 years worldwide in 2010–14, which was 5 points less than 40 (39–48) in 1990–94 (90% UI for decline −11 to 0). Because of population growth, the annual number of abortions worldwide increased by 5·9 million (90% UI −1·3 to 15·4), from 50·4 million in 1990–94 (48·6 to 59·9) to 56·3 million (52·4 to 70·0) in 2010–14. In the developed world, the abortion rate declined 19 points (−26 to −14), from 46 (41 to 59) to 27 (24 to 37). In the developing world, we found a non-significant 2 point decline (90% UI −9 to 4) in the rate from 39 (37 to 47) to 37 (34 to 46). Some 25% (90% UI 23 to 29) of pregnancies ended in abortion in 2010–14. Globally, 73% (90% UI 59 to 82) of abortions were obtained by married women in 2010–14 compared with 27% (18 to 41) obtained by unmarried women. We did not observe an association between the abortion rates for 2010–14 and the grounds under which abortion is legally allowed. Interpretation Abortion rates have declined significantly since 1990 in the developed world but not in the developing world. Ensuring access to sexual and reproductive health care could help millions of women avoid unintended pregnancies and ensure access to safe abortion. Funding UK Government, Dutch Ministry of Foreign Affairs, Norwegian Agency for Development Cooperation, The David and Lucile Packard Foundation, UNDP/UNFPA/UNICEF/WHO/World Bank Special Programme of Research, Development and Research Training in Human Reproduction.
A 3-month feeding trial with tongue sole Cynoglossus semilaevis broodstock was conducted before and during the spawning season to investigate the effects of dietary arachidonic acid (ARA) on the ...production of sex steroid hormones and gonadal gene expression of key proteins in steroidogenesis. Three isonitrogenous and isolipidic experimental diets were formulated to contain different ARA levels: the control diet without ARA supplementation (C, 0.58% ARA of total fatty acids (TFA)) and two diets with low (5.14% of TFA, ARA-L) or high ARA (15.44% of TFA, ARA-H) supplementation. The diets were randomly assigned to 9 tanks of 3-year-old tongue sole (10 females and 15 males in each tank). Fish were reared in a flowing seawater system and fed to apparent satiation twice daily. At the end of the feeding trial, tissue samples from mature females (MF, with spontaneous ovulation), immature females (IMF, early vitellogenesis), and mature males (MM, expressing milt) were collected to assay the production of sex steroid hormones, gonadal gene expression of sex steroid-synthesizing proteins, as well as the fatty acid profiles of gonad, liver and muscle lipids. Results showed that ARA supplementation significantly reduced the estradiol production in females, but stimulated the testosterone production in males. ARA supplementation significantly reduced the mRNA expression of aromatase in ovaries but significantly increased the gene expression of 3β-hydroxysteroid dehydrogenase (3β-HSD) in testes. In mature ovaries, diet ARA-L significantly reduced the gene expression of follicle stimulating hormone receptor (FSHR), 3β-HSD, and 17β-HSD; however, in immature ovaries, it significantly increased the gene expression of FSHR, steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (P450ssc), 3β-HSD, and 17β-HSD. In all gonads, 17α-hydroxylase (P450c17) responded to dietary ARA differently from other sex steroid-synthesizing proteins. ARA was preferentially accumulated in tongue sole gonad lipids. ARA concentrations were highest in gonad, liver and muscle lipids of MM fish and lowest in MF fish. Compared to female tongue sole, males had higher DHA concentrations in gonad lipids, but lower concentrations in liver lipids. In conclusion, results suggest dietary ARA regulates sex steroid hormone synthesis in tongue sole broodstock, and accumulates in gonad lipids, depending on both fish gender and maturation stage. Dietary ARA supplementation appears more important for male fish than for female fish, and more important for immature females than for mature females.
This study is beneficial to the broodstock diet formulation.
•Dietary ARA enhanced the testosterone production in mature tongue sole testis but reduced the estradiol production in ovary.•Dietary ARA regulated gene expressions of sex steroid-synthesizing proteins depending on fish gender and maturation stage.•ARA is preferentially accumulated in gonads, and testes showed higher ARA contents than ovaries.•This is the first study observing the effects of fish gender and maturation stage on regulation of steroidogenesis by ARA.
Background
Neoadjuvant chemotherapy (NAC) downstages axillary disease in 55 % of node-positive (N1) breast cancer. The feasibility and accuracy of sentinel lymph node biopsy (SLNB) after NAC for ...percutaneous biopsy-proven N1 patients who are clinically node negative (cN0) by physical examination after NAC is under investigation. ACOSOG Z1071 reported a false-negative rate of <10 % if ≥3 nodes are removed with dual tracer, including excision of the biopsy-proven positive lymph node (BxLN). We report our experience using radioactive seed localization (RSL) to retrieve the BxLN with SLNB (RSL/SLNB) for cN0 patients after NAC.
Methods
We performed a retrospective review of a single-institution, prospectively maintained registry for the years 2013 to 2014. Patients with BxLN who received NAC and had RSL/SLNB were identified. All BxLNs were marked with a radiopaque clip before NAC to facilitate RSL.
Results
Thirty patients with BxLN before NAC were cN0 after NAC and underwent RSL/SLNB. Median age was 55 years. Disease stage was IIA–IIIB. Twenty-nine of 30 had ductal cancer (12 triple negative and 16 HER-2 positive). One to 11 nodes were retrieved. Twenty-nine of 30 BxLN were successfully localized with RSL. Note was made of the BxLN-containing isotope and/or dye in 22 of 30. Nineteen patients had no residual axillary disease; 11 had persistent disease. All who remained node positive had disease in the BxLN.
Conclusions
RSL/SLNB is a promising approach for axillary staging after NAC in patients whose disease becomes cN0. The status of the BxLN after NAC predicted nodal status, suggesting that localization of the BxLN may be more accurate than SLNB alone for staging the axilla in the cN0 patient after NAC.
From 1798 to 1801, during the Haitian Revolution, President John Adams and Toussaint Louverture forged diplomatic relations that empowered white Americans to embrace freedom and independence for ...people of color in Saint-Domingue. The United States supported the Dominguan revolutionaries with economic assistance and arms and munitions; the conflict was also the U.S. Navy's first military action on behalf of a foreign ally. This cross-cultural cooperation was of immense and strategic importance as it helped to bring forth a new nation: Haiti.Diplomacy in Black and Whiteis the first book on the Adams-Louverture alliance. Historian and former diplomat Ronald Angelo Johnson details the aspirations of the Americans and Dominguans-two revolutionary peoples-and how they played significant roles in a hostile Atlantic world. Remarkably, leaders of both governments established multiracial relationships amid environments dominated by slavery and racial hierarchy. And though U.S.-Dominguan diplomacy did not end slavery in the United States, it altered Atlantic world discussions of slavery and race well into the twentieth century.Diplomacy in Black and Whitereflects the capacity of leaders from disparate backgrounds to negotiate political and societal constraints to make lives better for the groups they represent. Adams and Louverture brought their peoples to the threshold of a lasting transracial relationship. And their shared history reveals the impact of decisions made by powerful people at pivotal moments. But in the end, a permanent alliance failed to emerge, and instead, the two republics born of revolution took divergent paths.
High-throughput screening (HTS) of chemical compounds to identify modulators of molecular targets is a mainstay of pharmaceutical development. Increasingly, HTS is being used to identify chemical ...probes of gene, pathway, and cell functions, with the ultimate goal of comprehensively delineating relationships between chemical structures and biological activities. Achieving this goal will require methodologies that efficiently generate pharmacological data from the primary screen and reliably profile the range of biological activities associated with large chemical libraries. Traditional HTS, which tests compounds at a single concentration, is not suited to this task, because HTS is burdened by frequent false positives and false negatives and requires extensive follow-up testing. We have developed a paradigm, quantitative HTS (qHTS), tested with the enzyme pyruvate kinase, to generate concentration-response curves for > 60,000 compounds in a single experiment. We show that this method is precise, refractory to variations in sample preparation, and identifies compounds with a wide range of activities. Concentration-response curves were classified to rapidly identify pyruvate kinase activators and inhibitors with a variety of potencies and efficacies and elucidate structure-activity relationships directly from the primary screen. Comparison of qHTS with traditional single-concentration HTS revealed a high prevalence of false negatives in the single-point screen. This study demonstrates the feasibility of qHTS for accurately profiling every compound in large chemical libraries (>10⁵ compounds). qHTS produces rich data sets that can be immediately mined for reliable biological activities, thereby providing a platform for chemical genomics and accelerating the identification of leads for drug discovery.
High-throughput screening (HTS) assays enable the testing of large numbers of chemical substances for activity in diverse areas of biology. The biological responses measured in HTS assays span ...isolated biochemical systems containing purified receptors or enzymes to signal transduction pathways and complex networks functioning in cellular environments. This Review addresses factors that need to be considered when implementing assays for HTS and is aimed particularly at investigators new to this field. We discuss assay design strategies, the major detection technologies and examples of HTS assays for common target classes, cellular pathways and simple cellular phenotypes. We conclude with special considerations for configuring sensitive, robust, informative and economically feasible HTS assays.
The World Health Organization (WHO) Safe abortion: technical and policy guidance for health systems states that regulatory, policy, and programmatic barriers that hinder access to and timely ...provision of safe abortion care should be removed. Although some regulatory requirements facilitate access, where they act as barriers, they can deter women from seeking safe abortion care. We use data available in the Global Abortion Policies Database as of February 2019 to review policies related to regulatory requirements identified as access barriers in the Safe abortion guidance. We include only countries where such policies apply, i.e., where abortion is lawful on the woman's request, with no requirement for justification and/or for one or more legal grounds. The results demonstrate the variation that exists in regulatory requirements, but little remains known about how they are implemented in practice and the implications on how women access and how providers offer safe abortion services.
•The most common regulatory requirement is to obtain additional authorizations.•Misoprostol alone is included in the official drug list of 66% of countries.•Abortion is often restricted to provision by a physician provider.•Sixteen percent of countries have mandatory waiting periods.
Objectives
This study examined the accuracy of radioactive seed localization (RSL) of lymph nodes (LNs) following neoadjuvant chemotherapy (NAC) for invasive breast carcinoma, recorded pathologic ...features of LNs following NAC, evaluated concordance of response between breast and LNs, and identified clinicopathologic factors associated with higher risk of residual lymph node involvement.
Methods
Clinical records, imaging, and pathology reports and slides were retrospectively reviewed for 174 breast cancer patients who received NAC. Chi-square and Fisher’s exact tests were used to compare differences in risk of residual lymph node disease.
Results
Retrieval of biopsied pre-therapy positive LN was confirmed in 86/93 (88%) cases overall, and in 75/77 (97%) of cases utilizing RSL. Biopsy clip site was the best pathologic feature to confirm retrieval of a biopsied lymph node. Pre-therapy clinical N stage > 0, positive pre-therapy lymph node biopsy, estrogen and progesterone receptor positivity, Ki67 < 50%, HR + /HER2− tumors, and residual breast disease had higher likelihood of residual lymph node disease after NAC (
p
< 0.001).
Conclusions
RSL-guided LN excision improves retrieval of previously biopsied LNs following NAC. The pathologist can use histologic features to confirm retrieval of targeted LNs, and tumor characteristics can be used to predict a higher risk of residual LN involvement.
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