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The increase of many deadly diseases like infections by multidrug-resistant bacteria implies re-inventing the wheel on drug discovery. A better comprehension of the metabolisms and regulation of diseases, the increase in knowledge based on the study of disease-born microorganisms’ genomes, the development of more representative disease models and improvement of techniques, technologies, and computation applied to biology are advances that will foster drug discovery in upcoming years. In this paper, several aspects of current methodologies for drug discovery of antibacterial and antifungals, anti-tropical diseases, antibiofilm and antiquorum sensing, anticancer and neuroprotectors are considered. For drug discovery, two different complementary approaches can be applied: classical pharmacology, also known as phenotypic drug discovery, which is the historical basis of drug discovery, and reverse pharmacology, also designated target-based drug discovery. Screening methods based on phenotypic drug discovery have been used to discover new natural products mainly from terrestrial origin. Examples of the discovery of marine natural products are provided. A section on future trends provides a comprehensive overview on recent advances that will foster the pharmaceutical industry.

Background The tetrazolium-based MTT assay has long been regarded as the gold standard of cytotoxicity assays as it is highly sensitive and has been miniaturised for use as a high-throughput screening assay. However, various reports refer to interference by different test compounds, including the glycolysis inhibitor 3-bromopyruvate, with the conversion of the dye to coloured formazan crystals. This study assessed the linear range and reproducibility of three commonly used cell enumeration assays; the neutral red uptake (NRU), resazurin reduction (RES) and sulforhodamine B (SRB) assays, in comparison to the MTT assay. Interference between the MTT assay and three glycolysis inhibitors, 2-deoxyglucose, 3-bromopyruvate and lonidamine, was investigated. Results Data indicate that the NRU, RES and SRB assays showed the smallest variability across the linear range, while the largest variation was observed for the MTT assay. This implies that these assays would more accurately detect small changes in cell number than the MTT assay. The SRB assay provided the most reproducible results as indicated by the coefficient of determination after a limited number of experiments. The SRB assay also produced the lowest variance in the derived 50% inhibitory concentration (IC 50 ), while IC 50 concentrations of 3-bromopyruvate could not be detected using either the MTT or RES assays after 24 hours incubation. Interference in the MTT assay was observed for all three tested glycolysis inhibitors in a cell-free environment. No interferences were observed for the NRU, SRB or RES assays. Conclusions This study demonstrated that the MTT assay was not the best assay in a number of parameters that must be considered when a cell enumeration assay is selected: the MTT assay was less accurate in detecting changes in cell number as indicated by the variation observed in the linear range, had the highest variation when the IC 50 concentrations of the glycolysis inhibitors were determined, and interference between the MTT assay and all the glycolysis inhibitors tested were observed. The SRB assay performed best overall considering all of the parameters, suggesting that it is the most suitable assay for use in preclinical screening of novel therapeutic compounds with oxido-reductive potential.

The process of building new blood vessels (angiogenesis) and controlling the propagation of blood vessels (anti‐angiogenesis) are fundamental to human health, as they play key roles in wound healing and tissue growth. More than 500 million people may stand to benefit from anti‐ or pro‐angiogenic treatments in the coming decades [National Cancer Institute (USA), Cancer Bulltetin, volume 3, no. 9, 2006]. The use of animal models to assay angiogenesis is crucial to the search for therapeutic agents that inhibit angiogenesis in the clinical setting. Examples of persons that would benefit from these therapies are cancer patients, as cancer growth and spread is angiogenesis‐dependent, and patients with aberrant angiogenesis in the eye, which may lead to blindness or defective sight. Recently, anti‐angiogenesis therapies have been introduced successfully in the clinic, representing a turning point in tumor therapy and the treatment of macular degeneration and heralding a new era for the treatment of several commonly occurring angiogenesis‐related diseases. On the other hand, pro‐angiogenic therapies that promote compensatory angiogenesis in hypoxic tissues, such as those subjected to ischemia in myocardial or cerebral hypoxia due to occluding lesions in the coronary or cerebral arteries, respectively, and in cases of poor wound healing, are also being developed. In this review, the current major and newly introduced preclinical angiogenesis assays are described and discussed in terms of their specific advantages and disadvantages from the biological, technical, economical and ethical perspectives. These assays include the corneal micropocket, chick chorioallantoic membrane, rodent mesentery, subcutaneous (s.c.) sponge/matrix/alginate microbead, s.c. Matrigel plug, s.c. disc, and s.c. directed in vivo angiogenesis assays, as well as, the zebrafish system and several additional assays. A note on quantitative techniques for assessing angiogenesis in patients is also included. The currently utilized preclinical assays are not equivalent in terms of efficacy or relevance to human disease. Some of these assays have significance for screening, while others are used primarily in studies of dosage‐effects, molecular structure activities, and the combined effects of two or more agents on angiogenesis. When invited to write this review, I was asked to describe in some detail the rodent mesenteric‐window angiogenesis assay, which has not received extensive coverage in previous reviews.

Recently, the interest in the application of cell viability assays has been increasing in various fields. Cell viability assays may be broadly classified as (a) dye exclusion assays, (b) colorimetric assays, (c) fluorometric assays, (d) luminometric assays, and (e) flow cytometric assays. Dye exclusion assays include trypan blue, eosin, congo red, and erythrosine B stain assays, whereas 3‐[4,5‐dimethylthiazol‐2‐yl]‐2,5 diphenyl tetrazolium bromide (MTT), 3‐(4,5‐dimethylthiazol‐2‐yl)‐5‐(3‐carboxymethoxyphenyl)‐2‐(4‐sulfophenyl)‐2H‐tetrazolium (MTS), 2,3‐bis‐(2‐methoxy‐4‐nitro‐5‐sulfophenyl)‐2H‐tetrazolium‐5‐carboxanilide (XTT), 2‐(4‐iodophenyl)‐3‐(4‐nitrophenyl)‐5‐(2,4‐disulfophenyl)‐2H tetrazolium, monosodium salt (WST‐1), 2‐(2‐methoxy‐4‐nitrophenyl)‐3‐(4‐nitrophenyl)‐5‐(2,4‐disulfophenyl)‐2H‐tetrazolium, monosodium salt (WST‐8), lactate dehydrogenase (LDH), sulforhodamine B (SRB), neutral red uptake (NRU), and crystal violet stain (CVS) assays are among the colorimetric assays. Similarly, resazurin and 5‐carboxyfluorescein diacetate acetoxymethyl ester (5‐CFDA‐AM) assays are based on fluorometric measurements, whereas luminometric assays comprise adenosine triphosphate and real‐time viability assays. Major flow cytometric assays include membrane asymmetry, membrane permeability, and mitochondria assays. In this guideline, the mechanisms and the practice of assessment of the most common cell viability assays applied in research labs are discussed in detail. An ideal cell viability assay should be safe, rapid, reliable, efficient, and time‐ and cost‐effective, and should not interfere with the test compound. Overall, it can be concluded that more than one cell viability assay should be applied in order to obtain reliable results. Cell viability assays may be broadly classified as (a) dye exclusion assays, (b) colorimetric assays, (c) fluorometric assays, (d) luminometric assays, and (e) flow cytometric assays. In this guideline, the mechanisms and the practice of assessment of the most common cell viability assays applied in research labs are discussed in detail.

The major works of one of the nineteenth century's most influential philosophers In the early days of the American experiment, as the states spread across the continent and the young nation was reshaped by the Industrial Revolution, no intellectual held more power than Ralph Waldo Emerson.

American Samoa is the only U.S. jurisdiction that does not recognize gender-neutral marriage despite the Supreme Court’s Obergefell decision invalidating laws that limit marriage to male–female couples. Among U.S. territories, American Samoa has five unique features: It is the only territory that the United States acquired through negotiation with ruling sovereigns, whose land is largely communally owned, whose residents lack birthright citizenship, that remains under control of the Secretary of the Interior, and that lacks a federal court. This Essay explains how these characteristics have combined to thwart marriage equality in American Samoa. American Samoa’s denial of marriage equality is surprising because for centuries Samoan culture has respected third-gender individuals, called fa‘afafine. Despite this heritage and the Obergefell opinion recognizing the constitutional right to gender-neutral marriage, American Samoa does not allow fa‘afafine to marry their male partners. After documenting the centuries-old Polynesian tradition of respecting third-gender individuals, this Essay shows how current leaders in American Samoa are using suspect precedent to prohibit marriage equality for the fa‘afafine. In a series of racist opinions from 1901, known as the Insular Cases, the Supreme Court held that the U.S. Constitution does not apply to U.S. territories because their residents cannot be entrusted with rights and self-governance. Although all other U.S. territories acceded to Obergefell, American Samoa’s politicians have relied on Insular logic to block marriage equality from reaching America’s most distant territory. This Essay explains the inherent unfairness of allowing the anachronistic Insular Cases to prevent fa‘afafine from having marriage rights today.

Apoptosis has been recognized as a type of programmed cell death connected with characteristic morphological and biochemical changes in cells. This programmed cell death plays an important role in the genesis of a number of physiological and pathological processes. Thus, it can be very important to detect the signs of apoptosis in a study of cellular metabolism. The present paper provides an overview of methods often being used for detecting DNA fragmentation as one of the most specific findings in apoptosis. To date, three routine assays have been developed for detecting DNA fragmentation: DNA ladder assay, TUNEL assay, and comet assay. All these methods differ in their principles for detecting DNA fragmentation. DNA ladder assay detects the characteristic “DNA ladder” pattern formed during internucleosomal cleavage of DNA. Terminal deoxynUcleotidyl transferase Nick-End Labeling (TUNEL) assay detects DNA strand breaks using terminal deoxynucleotidyl transferase catalyzing attachment of modified deoxynucleotides on the DNA strand breaks. Comet assay can be used for detecting nucleus breakdown producing single/double-strand DNA breaks. The aim of this review is to describe the present knowledge on these three methods, including optimized approaches, techniques, and limitations.

Much progress has been made toward assessing and improving animal welfare in conservation. However, several glaring knowledge gaps remain where animal-welfare concerns exist but animal-welfare studies have not been performed in politically sensitive contexts. Based on contemporary issues in Australia, we identified 4 topics that require more research: animal-welfare oversight for operations designated as management (as opposed to research); animal-welfare impacts of biological agents used to control invasive animals; welfare of animals hunted recreationally; and animal-welfare impacts associated with indigenous wildlife use. Animal-welfare science may be applied to these sensitive topics through simple quantitative studies (e. g., quantifying the frequency of adverse animal-welfare events). Several such studies have effectively addressed animal-welfare concerns in similarly contentious contexts, including feral camel (Camelus dromedarius) culling in Australia, recreational hunting in Scandinavia, and indigenous whale hunting in the United States. For discussions of animal welfare in conservation to be evidence-based, courageous research is required in the 4 key areas we identified. En la conservación se ha progresado mucho en la evaluacióny el mejoramientodel bienestaranimal. Sin embargo, todavía permanecen varios vacíos evidentes en donde existe preocupación por el bienestar animal, pero los estudios sobre este bienestar no se han realizado en contextos políticamente sensibles. Con base en temas contemporáneos en Australia, identificamos cuatro temas que requieren de más investigación: omisión del bienestar animal por operaciones designadas como manejo (en lugar de investigación); impactos de los agentes biológicos usados para controlar a animales invasores sobre el bienestar animal; bienestar de los animales cazados por recreación; e impactos sobre el bienestar animalasociados con el uso de la faunanativa. La ciencia del bienestar animal puede aplicarse a estos temas sensibles por medio de estudios cuantitativos (p. ej.: cuantificación de la frecuencia de eventos adversos para el bienestar animal). Varios de estos estudios han tratado efectivamente las preocupaciones por el bienestar animal en contextos similarmente polémicos, incluyendo el sacrificio de camellos ferales (Camelus dromedarius) en Australia, la cacería recreativa en Escandinavia, y la caza de ballenas por aborígenes en los Estados Unidos. Para que las discusiones sobre el bienestar animal en la conservación estén basadas en evidencias, se requiere de investigaciones atrevidas en las cuatro áreas clave que identificamos. 在保护中，对动物福利的评估和改善已经取得了很大进展。然而，在动物福利问题尚未解决，却因敏感的 政治环境而难以进行动物福利研究的地方，仍存在几个明显的知识空缺。基于目前澳大利亚面临的问题，我们确 定了四个需要进ー步研究的主题: 为管理(而非研究) 进行特定操作时的动物福利监管;用于控制入侵动物的生 物防治天敌对动物福利的影响; 娱乐性狩猎中的动物福利问题，以及与原生野生动物利用有关的动物福利问题。 我们认为，可以通过简单的定量研究（如量化危害动物福利的事件发生的频率) 将动物福利科学应用于这些敏 感话题。一些这样的研究已经有效地解决了在类似的有争议的情况下的动物福利问题，包括澳大利亚的野骆骑 CCamelus dromedariusj) 选择性捕杀、斯堪的纳维亚的娱乐性狩猎7以及美国原生鲸类的捕杀。为保证对保护中 动物福利向题的讨论建立在证据之上，我们要勇于在这四个关键领域发起研究.

In health equity research, anti-Black racism and power imbalances manifest at every phase of the research process and contribute to the marginalization and exclusion of Black scholars. This essay highlights how power operates as a central component of anti-Black racism, and I describe the importance of centering Black scholars in funding, conducting, and implementing health equity research. Interdisciplinary collaboration between the fields of bioethics, public health ethics, and health equity could generate dialogue and develop recommendations to help balance power dynamics, address anti-Black racism, and, ultimately, make meaningful progress toward health equity.