Resistance mechanisms and cross-resistance for a pyridine-pyrimidine amide inhibitor of microtubule polymerization.

Despite their clinical importance, drug resistance remains problematic for microtubule targeting drugs. D4-9-31, a novel microtubule destabilizing agent, has pharmacology that suggests it can overcome common resistance mechanisms and has been shown to remain efficacious in cell and animal models with acquired taxane resistance. To better understand resistance mechanisms and the breadth of cross-resistance with D4-9-31, this study examines the A2780 ovarian cancer cell line as it develops acquired resistance with continuous exposure to D4-9-31. Analyzing cellular responses to D4-9-31 reveals that D4-9-31 resistance is associated with increased mitochondrial respiration, but no cross-resistance to other microtubule targeting agents is observed. Sequencing of transcripts of parental cells and resistant counterparts reveals mutations and altered expression of microtubule-associated genes, but not in genes commonly associated with resistance to microtubule targeting drugs. Additionally, our findings suggest distinct mechanisms drive short- and long-term drug resistance.

Blockade of PD1 and TIM3 restores innate and adaptive immunity in patients with acute alcoholic hepatitis.

BACKGROUND & AIMS: Susceptibility to bacterial infection is a feature of alcohol-related liver disease. Programmed cell death 1 (PD1), the T-cell immunoglobulin and mucin domain-containing protein 3 (TIM3, also known as hepatitis A virus cellular receptor 2), and their respective ligands-CD274 (also known as PD ligand 1 [PDL1]) and galectin-9-are inhibitory receptors that regulate the balance between protective immunity and host immune-mediated damage. However, their sustained hyperexpression promotes immune exhaustion and paralysis. We investigated the role of these immune inhibitory receptors in driving immune impairments in patients with alcoholic liver disease. METHODS: In a prospective study, we collected blood samples from 20 patients with acute alcoholic hepatitis (AAH), 16 patients with stable advanced alcohol-related cirrhosis, and 12 healthy individuals (controls). Whole blood or peripheral blood mononuclear cells were assessed for expression of PD1, PDL1, TIM3, galectin-9, and Toll-like receptors on subsets of innate and adaptive immune effector cells. We measured antibacterial immune responses to lipopolysaccharide (endotoxin) using ELISpot assays, and used flow cytometry to quantify cytokine production, phagocytosis, and oxidative burst in the presence or absence of blocking antibodies against PD1 or TIM3. RESULTS: Antibacterial innate and adaptive immune responses were greatly reduced in patients with AAH, compared with controls, and patients with alcohol-related cirrhosis had less severe dysfunctions in innate immune effector cells and preserved functional T-cell responses. Fewer T cells from patients with AAH produced interferon gamma in response to lipopolysaccharide, compared with controls. In addition, patients with AAH had greater numbers of interleukin 10-producing T cells, and reduced levels of neutrophil phagocytosis and oxidative burst in response to Escherichia coli stimulation, compared with controls. T cells from patients with AAH, but not alcohol-related cirrhosis, expressed higher levels of PD1 and PDL1, or TIM3 and galectin-9, than T cells from controls. Antibodies against PD1 and TIM3 restored T-cell production of interferon gamma, reduced the numbers of interleukin 10-producing T cells, and increased neutrophil antimicrobial activities. Circulating levels of endotoxin in plasma from patients with AAH caused over expression of immune inhibitory receptors on T cells via Toll-like receptor 4 binding to CD14(+) monocytes. CONCLUSIONS: Antibacterial immune responses are impaired in patients with AAH. Lymphocytes from these patients express high levels of immune inhibitory receptors, produce lower levels of interferon gamma, and have increased IL10 production due to chronic endotoxin exposure. These effects can be reversed by blocking PD1 and TIM3, which increase the antimicrobial activities of T cells and neutrophils.

The Cpx envelope stress response regulates and is regulated by small noncoding RNAs.

The Escherichia coli genome encodes approximately 30 two-component systems that are required for sensing and responding to a variety of environmental and physiological cues. Recent studies have revealed numerous regulatory connections between two-component systems and small noncoding RNAs (sRNAs), which posttranscriptionally regulate gene expression by base pairing with target mRNAs. In this study, we investigated the role of sRNAs in the CpxAR two-component system, which detects and mediates an adaptive response to potentially lethal protein misfolding in the Gram-negative bacterial envelope. Here, we showed for the first time that sRNAs are members of the Cpx regulon. We found that CpxR binds to the promoter regions and regulates expression of two sRNA genes, cyaR and rprA. We also investigated the roles that these sRNAs play in the Cpx response. Cpx repression of cyaR expression creates a feed-forward loop, in which CpxAR increases expression of the inner membrane protein YqaE both directly at the transcriptional level and indirectly at the translational level. Moreover, we found that RprA exerts negative feedback on the Cpx response, reducing Cpx activity in a manner that is dependent on the response regulator CpxR but independent of all of RprA's previously described targets. sRNAs therefore permit the fine-tuning of Cpx pathway activity and its regulation of target genes, which could assist bacterial survival in the face of envelope stress.

Proteomic characterization of histotroph during the preimplantation phase of the estrous cycle in cattle.

Uterine secretions, or histotroph, are a critical component for early embryo survival, functioning as the sole supply of vitamins, minerals, enzymes, and other myriad of nutrients required by the developing conceptus before implantation. Histotroph is therefore a promising source for biomarkers of uterine function and for enhancing our understanding of the environment supporting early embryo development and survival. Utilizing label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) shotgun proteomics, we characterized the uterine proteome at two key preimplantation stages of the estrous cycle in high fertility cattle. We identified 300 proteins on Day 7 and 510 proteins on Day 13 including 281 proteins shared between days. Five proteins were more abundant (P < 0.05) on Day 7 compared with Day 13 and included novel histotroph proteins cytokeratin 10 and stathmin. Twenty-nine proteins were more abundant (P < 0.05) including 13 unique on Day 13 compared with Day 7 and included previously identified legumain, metalloprotease inhibitor-2, and novel histotroph proteins chromogranin A and pyridoxal kinase. Functional analysis of the 34 differentially expressed proteins (including 14 novel to histotroph) revealed distinct biological roles putatively involved in early pregnancy, including remodelling of the uterine environment in preparation for implantation; nutrient metabolism; embryo growth, development and protection; maintenance of uterine health; and maternal immune modulation. This study is the first reported LC-MS/MS based global proteomic characterization of the uterine environment in any domesticated species before implantation and provides novel information on the temporal alterations in histotroph composition during critical stages for early embryo development and uterine function during the early establishment of pregnancy.

Enhancing the Australian Gridded Climate Dataset rainfall analysis using satellite data.

Rainfall estimation over large areas is important for a thorough understanding of water availability, influencing societal decision-making, as well as being an input for scientific models. Traditionally, Australia utilizes a gauge-based analysis for rainfall estimation, but its performance can be severely limited over regions with low gauge density such as central parts of the continent. At the Australian Bureau of Meteorology, the current operational monthly rainfall component of the Australian Gridded Climate Dataset (AGCD) makes use of statistical interpolation (SI), also known as optimal interpolation (OI) to form an analysis from a background field of station climatology. In this study, satellite observations of rainfall were used as the background field instead of station climatology to produce improved monthly rainfall analyses. The performance of these monthly datasets was evaluated over the Australian domain from 2001 to 2020. Evaluated over the entire national domain, the satellite-based SI datasets had similar to slightly better performance than the station climatology-based SI datasets with some individual months being more realistically represented by the satellite-SI datasets. However, over gauge-sparse regions, there was a clear increase in performance. For a representative sub-domain, the Kling-Gupta Efficiency (KGE) value increased by + 8% (+ 12%) during the dry (wet) season. This study is an important step in enhancing operational rainfall analysis over Australia.

Effectiveness and Safety of Nonvitamin K Oral Anticoagulants Rivaroxaban and Apixaban in Patients with Venous Thromboembolism: A Meta-Analysis of Real-World Studies.

Background: Rivaroxaban and apixaban are the most widely used nonvitamin K oral anticoagulants (NOACs) in patients with venous thromboembolism (VTE). This meta-analysis evaluates the effectiveness and safety of both NOACs versus standard of care (SoC) in real-world practice. Methods: Real-world evidence (RWE) studies were identified through a systematic literature review conducted between January 2012 and July 2020, using Embase, MEDLINE, and the websites of cardiological, hematological, and oncological associations. Eligible RWE studies recruited adult patients with deep vein thrombosis and/or pulmonary embolism and presented a comparison between rivaroxaban and apixaban versus SoC, consisting either of vitamin K antagonists, heparins, or combinations thereof. Hazard ratios (HRs) for the comparison between NOACs and SoC were extracted from the relevant studies or estimated based on the reported binary data. The between-treatment contrasts were reported as HRs with associated 95% confidence intervals. Results: A total of 65 RWE studies were identified and considered relevant for the meta-analysis. Compared with SoC, both rivaroxaban and apixaban were associated with reduced risks of recurrent VTE and a lower rate of major bleeding events. Patients treated with rivaroxaban were at a lower risk of all-cause death compared with those receiving SoC (HR = 0.56 [0.39-0.80]), while evidence for apixaban from the identified studies was insufficient to demonstrate a statistically significant change in mortality (HR = 0.66 [0.30-1.47]). Conclusion: This analysis indicates that in real-world practice, rivaroxaban and apixaban are associated with a lower risk of recurrent VTE and major bleeding events compared with SoC. Survival benefit in patients treated with rivaroxaban was also observed.

Effects of lipid-encapsulated conjugated linoleic acid supplementation on milk production, bioenergetic status and indicators of reproductive performance in lactating dairy cows.

Conjugated linoleic acid (CLA) reduces mammary milk fat synthesis in a dose-dependent manner. Our objective was to determine the effects of lipid-encapsulated CLA (LE-CLA) supplementation on milk production, reproductive performance and metabolic responses in lactating dairy cows fed a grass silage-based diet. Seventy-two Holstein-Friesian cows (32 primiparous and 40 multiparous) were used in a completely randomized block design. Cows received either 80 g of LE-CLA daily or 60 g of calcium salts of palm fatty acids daily (control) from parturition until 60 days in milk. LE-CLA contained a 50:50 mix of cis-9,trans-11 CLA and trans-10,cis-12 CLA, resulting in a daily intake of 6 g of each isomer. Milk production and dry matter intake were recorded daily, and blood samples were collected 3-times a week. Blood samples were analysed for circulating concentrations of glucose, non-esterified fatty acids (NEFA), ß-hydroxybutyrate (BHBA), insulin and insulin-like growth factor-I (IGF-I). Progesterone was measured in blood samples collected after the first post-partum insemination. Ovarian ultrasound examinations commenced at 8-10 d post partum and were carried out 3-times a week until first ovulation. LE-CLA treatment resulted in decreased milk fat concentration, with consequent improvements in energy balance and body condition score (BCS). The peak concentration of NEFA in blood was reduced by LE-CLA, but circulating concentrations of insulin, glucose, IGF-I, BHBA and progesterone were not affected. There was no effect of LE-CLA supplementation on the post-partum interval to first ovulation. Services per conception tended to be reduced. The reduction in milk energy output and improvement in energy status and BCS in LE-CLA-supplemented cows provides a strong rationale for further studies with greater cow numbers to test effects on reproductive performance.

Hereditary haemorrhagic telangiectasia and pulmonary arteriovenous malformations.

Hereditary haemorrhagic telangiectasia (HHT) also known as Osler-Weber-Rendu syndrome is an autosomal dominant disorder affecting 1 in 8000 individuals. The eponym recognises the 19th-century physicians William Osler, Henri Jules Louis Marie Rendu and Frederick Parkes Weber who each independently described the disease. It is characterised by epistaxis, telangiectasia and visceral arteriovenous malformations. Individuals with HHT have been found to have abnormal plasma concentrations of transforming growth factor beta and vascular endothelial growth factor secondary to mutations in ENG, ACVRL1 and MADH4. Pulmonary artery malformations (PAVMs) are abnormal communications between pulmonary arteries and veins and are found in up to 50% of individuals with HHT. The clinical features suggestive of PAVMs are stigmata of right to left shunting such as dyspnoea, hypoxaemia, cyanosis, cerebral embolism and unexplained haemoptysis or haemothorax. The authors present the case of a 33-year-old woman presenting with progressive dyspnoea during the COVID-19 pandemic. She had a typical presentation of HHT with recurrent epistaxis, telangiectasia and pulmonary arteriovenous malformations. Although rare, PAVM should be considered in individuals presenting to the emergency department with dyspnoea and hypoxaemia. Delayed diagnosis can result in fatal embolic and haemorrhagic complications.

Key traits for ruminant livestock across diverse production systems in the context of climate change: perspectives from a global platform of research farms.

Ruminant livestock are raised under diverse cultural and environmental production systems around the globe. Ruminant livestock can play a critical role in food security by supplying high-quality, nutrient-dense food with little or no competition for arable land while simultaneously improving soil health through vital returns of organic matter. However, in the context of climate change and limited land resources, the role of ruminant-based systems is uncertain because of their reputed low efficiency of feed conversion (kilogram of feed required per kilogram of product) and the production of methane as a by-product of enteric fermentation. A growing human population will demand more animal protein, which will put greater pressure on the Earth's planetary boundaries and contribute further to climate change. Therefore, livestock production globally faces the dual challenges of mitigating emissions and adapting to a changing climate. This requires research-led animal and plant breeding and feeding strategies to optimise ruminant systems. This study collated information from a global network of research farms reflecting a variety of ruminant production systems in diverse regions of the globe. Using this information, key changes in the genetic and nutritional approaches relevant to each system were drawn that, if implemented, would help shape more sustainable future ruminant livestock systems.

In vitro efficacy of a copper iodine complex PPE disinfectant for SARS-CoV-2 inactivation.

Background: The ability to protect workers and healthcare professionals from infection by SARS-CoV-2, the virus that causes coronavirus disease 2019 (COVID-19), is of great concern. Hospitals, nursing homes and employers are adopting infection control strategies based on guidance from leading public health organizations such as the CDC, OSHA, FDA, and other government bodies. Certain hard surface disinfectants are effective against SARS-CoV-2 but are not suitable for use on skin or personal protective equipment (PPE) that comes into contact with skin. Furthermore, near-ubiquitous alcohol-based hand sanitizers are acceptable for use on skin, but they are not suitable for use on PPE. PPE, especially masks, are also commonly being used for longer durations than normal. There is a need for new products and techniques that can effectively disinfect PPE during wear time without having detrimental effects on surrounding skin. Clyraguard spray is a novel copper iodine complex designed to be used on non-critical PPE. Methods: In this study, the Clyraguard copper iodine complex was tested for its ability to inactivate SARS-CoV-2 in solution. Results: These data indicate the product to be effective in reducing SARS-CoV-2 titers in a time-dependent manner, with the virus being reduced below the detection limits within 30 minutes. Conclusions: These results suggest that Clyraguard may be an effective tool for mitigating cross-contamination of non-critical PPE that may come into contact with SARS-CoV-2.

Efficacy of a novel iodine complex solution, CupriDyne, in inactivating SARS-CoV-2.

The coronavirus known as SARS-CoV-2, which causes COVID-19 disease, is presently responsible for a global pandemic wherein more than 3.5 million people have been infected and more than 250,000 killed to-date. There is currently no vaccine for COVID-19, leaving governments and public health agencies with little defense against the virus aside from advising or enforcing best practices for virus transmission prevention, which include hand-washing, physical distancing, use of face covers, and use of effective disinfectants. In this study, a novel iodine complex called CupriDyne® was assessed for its ability to inactivate SARS-CoV-2. CupriDyne was shown to be effective in inactivating the virus in a time-dependent manner, reducing virus titers by 99% (2 logs) after 30 minutes, and reducing virus titers to below the detection limit after 60 minutes. The novel iodine complex tested herein offers a safe and gentle alternative to conventional disinfectants for use on indoor and outdoor surfaces.

Toward Improved Environmental Stability of Polymer:Fullerene and Polymer:Nonfullerene Organic Solar Cells: A Common Energetic Origin of Light- and Oxygen-Induced Degradation.

With the emergence of nonfullerene electron acceptors resulting in further breakthroughs in the performance of organic solar cells, there is now an urgent need to understand their degradation mechanisms in order to improve their intrinsic stability through better material design. In this study, we present quantitative evidence for a common root cause of light-induced degradation of polymer:nonfullerene and polymer:fullerene organic solar cells in air, namely, a fast photo-oxidation process of the photoactive materials mediated by the formation of superoxide radical ions, whose yield is found to be strongly controlled by the lowest unoccupied molecular orbital (LUMO) levels of the electron acceptors used. Our results elucidate the general relevance of this degradation mechanism to both polymer:fullerene and polymer:nonfullerene blends and highlight the necessity of designing electron acceptor materials with sufficient electron affinities to overcome this challenge, thereby paving the way toward achieving long-term solar cell stability with minimal device encapsulation.

Transmission ratio distortion at the growth hormone gene (GH1) in bovine preimplantation embryos: An in vitro culture-induced phenomenon?

The growth hormone gene (GH1) and its polypeptide product (GH) have a crucial role in reproduction, embryogenesis and general development. A polymorphism present in the fifth exon of the bovine GH1 gene (GH1 p.Leu127Val) has been associated with GH release and milk production in cattle. The objective of the present study was to examine the genotype frequencies of the GH1 p.Leu127Val polymorphism in bovine blastocysts produced in vitro and in vivo to determine if allelic variation of the GH1 gene affects embryo development and survival. A heterozygous (p.Leu127/Val127) sire was used for in vitro fertilization of oocytes of unknown maternal genotype (n = 104) and known maternal genotype (n = 115). PCR amplification and genotyping of the GH1 gene from Day 8 blastocysts derived from these fertilized oocytes demonstrated that there was significant over-representation from the expected Mendelian ratio of GH1 p.Leu127/Leu127 homozygotes from oocytes of known maternal genotype (P = 0.006). Contrary to this, analysis of in vivo-produced bovine blastocysts of known parental GH1 genotype (n = 69) did not reveal an overrepresentation of GH1 p.Leu127/Leu127 homozygotes. These results suggest that developing in vitro-produced embryos are exposed to a selection process, probably due to a less favorable culture environment, that acts to increase the number of GH1 p.Leu127/Leu127 homozygotes, thereby giving rise to the observed transmission ratio distortion (TRD) of GH1 genotypes when compared to in vivo produced embryos.

Occurrence and weight-of-evidence risk assessment of alkyl sulfates, alkyl ethoxysulfates, and linear alkylbenzene sulfonates (LAS) in river water and sediments.

Alkyl sulfates (AS), alkyl ethoxysulfates (AES) and linear alkyl benzene sulfonates (LAS) are all High Production Volume (HPV) and 'down-the-drain' chemicals used globally in detergent and personal care products, resulting in low levels ultimately released to the environment via wastewater effluent. Due to their surfactant properties, they preferentially sorb to sediments. Hence, assessment of their levels and potential perturbations on benthos are of interest. The relative levels of AS/AES decreased with distance from the wastewater treatment plant outfall. However, this was not evident for LAS. Short chained AES and especially AS dominated the homologue distribution for AES. There were no evident patterns in LAS homologue distribution. The overall mean margin of exposure (MoE) for AS/AES and LAS is approximately 40 (range: 3 to 100) suggesting no noteworthy perturbation on biota. The findings in this study are in concordance with previous preliminary hazard screening. Comparative sediment contamination analyses principally based on Chapman and Anderson [Chapman PM, Anderson, J. A decision-making framework for sediment contamination. Integr Environ Assess Mana. 2005; 1: 163-173.] and the U.S. Environmental Protection Agency RAPID assessment methods [USEPA. Rapid bioassessment protocols for use in wadeable streams and rivers: Periphyton, benthic, macroinvertebrates, and fish. 1999. Second Edition. U.S. Environmental Protection Agency Office of Water, Washington, D.C. EPA 841-B-99-002.] did not reveal significant correlations between the surfactant concentrations and ecological status of the sampling locations. Several Lines of Evidence (LoE) of the Weight-of-Evidence (WoE) lead to the conclusion of low aquatic risk associated to the monitored compounds.

An unusual cause of jaundice in a patient with breast cancer.

A 48-year old woman with metastatic breast cancer and extensive bone marrow infiltration was admitted with extreme lethargy, jaundice and deranged liver function tests. She had been started on anastrozole in May 2013 for bony metastases, detected on a bone scan. A CT scan performed at that time had shown no evidence of metastatic or nodal disease elsewhere. Over the subsequent 2 months, the patient had become progressively jaundiced. Outpatient abdominal ultrasound and CT liver had shown a fatty liver with no focal lesions. She was admitted in August 2013 with bilirubin 567, alkaline phosphatase 385, alanine aminotransferase 98, albumin 25 and international normalised ratio 1.9. The patient ultimately had a liver biopsy, which demonstrated features of drug-induced steatohepatitis, and anastrozole was found to have been the probable cause. This case explores the differentials of jaundice in a patient with cancer and describes a rare cause of drug-induced liver injury.

Characterization of follicle and CL development in beef heifers using high resolution three-dimensional ultrasonography.

The aim was to characterize dominant follicle (DF) and CL development through the estrous cycle of cattle using three-dimensional (3D) ultrasonography while making a comparison with conventional two-dimensional (2D) B-mode ultrasound (US) and to relate the measures taken to systemic concentrations of steroid hormones and gonadotropins. After synchronization of estrus, the ovaries of crossbred beef heifers (N = 5) were assessed using daily US with a GE Voluson i US scanner until the end of the first follicle wave, then every other day until emergence of the final (ovulatory) wave, when daily US resumed until ovulation. Follicle and CL growth were recorded and mapped. Measures of diameter (2D) and volume (3D) of the DF from the first and ovulatory waves of the cycles; and CL development were captured and stored for further analysis. Blood flow to the DF and CL were assessed using 3D power Doppler US measuring vascularization index (VI; %), vascularization flow index (0/100) and flow index (0/100). Jugular blood samples were collected every 24 hours for progesterone from the first estrus until the second ovulation. Concentrations of estradiol (E2) and follicle stimulating hormone (FSH) were measured every 8 hours from estrus to second follicle wave emergence; then, E2 only was measured from final follicle wave emergence until ovulation. Data were analyzed using PROC MIXED and PROC REG in SAS. Dominant follicle blood flow tended to decrease during follicle wave emergence and DF VI increased (P < 0.05) 24 hours before ovulation after peak E2. Measures of the DF and CL volume (3D) were highly predictive of 2D diameter measures throughout the cycle (P < 0.0001). Predictive values (r(2)) for day of wave emergence and day from ovulation were similar for 2D and 3D measures; however, 2D measures had higher repeatability when compared with 3D measures. There was no relationship between CL VI and progesterone early in the cycle (r(2) = 0.12; P = 0.1); however, there was a strong positive relationship approaching ovulation (r(2) = 0.77; P < 0.0001). In conclusion, 3D power Doppler measures of blood flow appears to be representative of vascular changes in the DF and CL throughout the estrous cycle. However, the extra time required to acquire and analyze a 3D image and the relatively little additional information obtained over that achievable with 2D imaging in terms of follicle and CL development might preclude its widespread use other than for detailed research purposes.

Worldwide need for a comprehensive database of stability data for pharmaceuticals stored in uncontrolled environments.

Developing a database to compile all known, relevant information on the stability of pharmaceuticals in tropical environments would vastly increase access to this information, improving safety and reducing cost. Many studies have examined the stability of pharmaceuticals stored under conditions that exceed manufacturer specifications, but this information is published across so many journals and Websites that it can be virtually inaccessible to clinics or suppliers needing to make pharmaceutical inventory management decisions. We have made progress toward the completion of a database of this nature, but resources and abilities of international public health organizations will be needed to bring the database fully to fruition.

Development of a standardized Intranet database of formulation records for nonsterile compounding, Part 2.

In part 1 of this series, we outlined the rationale behind the development of a centralized electronic database used to maintain nonsterile compounding formulation records in the Mission Health System, which is a union of several independent hospitals and satellite and regional pharmacies that form the cornerstone of advanced medical care in several areas of western North Carolina. Hospital providers in many healthcare systems require compounded formulations to meet the needs of their patients (in particular, pediatric patients). Before a centralized electronic compounding database was implemented in the Mission Health System, each satellite or regional pharmacy affiliated with that system had a specific set of formulation records, but no standardized format for those records existed. In this article, we describe the quality control, database platform selection, description, implementation, and execution of our intranet database system, which is designed to maintain, manage, and disseminate nonsterile compounding formulation records in the hospitals and affiliated pharmacies of the Mission Health System. The objectives of that project were to standardize nonsterile compounding formulation records, create a centralized computerized database that would increase healthcare staff members' access to formulation records, establish beyond-use dates based on published stability studies, improve quality control, reduce the potential for medication errors related to compounding medications, and (ultimately) improve patient safety.