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The aim of the present study was to evaluate the effects of exposure to Chilean pulp mill effluent extracts on developing postfertilized medaka embryos before and after sex definition relative to sex steroids (testosterone and 17beta‐estradiol) and a wood phytoestrogen (beta‐sitosterol). Our study included 2 waterborne semichronic exposure experiments, using a 24‐h post fertilization (hpf) unknown‐sex FLFII (female leucophore free) group and a second 72‐hpf FLFII phenotypic sex‐identified group (male autofluorescence leucophore) strain of medaka embryos. Chronic exposure of both FLFII strain embryo groups showed similar delay in time to hatch and decreased hatchability. Teratogenic responses such as vertebral malformation (fusion, incomplete formation, and lack of vertebral formation process) and pericardial edema were observed in both experiments, with a high percentage related to FLFII fluorescent leucophore–identified males. In addition, high mortality associated with severe malformations was observed in male and female embryos exposed to testosterone. Our research has demonstrated that exposure to Chilean mill effluent extracts caused severe male medaka embryotoxicity (in postfertilized embryos) before and after sex definition and, irrespective of the experimental group and effluent treatment, suggests partial removal following secondary treatment. Furthermore, differences in the severity and type of teratogenic effects with previous experiments (d‐rR medaka strain), are associated with the unique phenotypes of this medaka mutant strain. Environ Toxicol Chem 2021;40:2297–2305. © 2021 SETAC [ABSTRACT FROM AUTHOR]

Rationale: The objective of this study was to identify unique chemical tracers of oil sands process-affected water (OSPW) to enable definitive discrimination of tailings pond seepage from natural bitumen-influenced waters from the Canadian Alberta McMurray formation. Methods: The approach involved comparing unknowns from an unprecedented sample set of OSPW (n = 4) and OSPW-affected groundwaters (n = 15) with natural bitumen-influenced groundwaters (n = 20), using high-performance liquid chromatography/electrospray ionisation high-resolution mass spectrometry (HPLC/ESI-HRMS) operated in both polarities. Results: Four unknown chemical entities were identified as potential tracers of OSPW seepage and subsequently subjected to structural elucidation. One potential tracer, tentatively identified as a thiophene-containing carboxylic acid [C15H23O3S]-, was only detected in OSPW and OSPW-affected samples, thereby showing the greatest diagnostic potential. The remaining three unknowns, postulated to be two thiochroman isomers [C17H25O3S]+ and an ethyl-naphthalene isomer [C16H21]+, were detected in one and two background groundwaters, respectively. Conclusions: We advanced the state of knowledge for tracers of tailings seepage beyond heteroatomic classes, to identifying diagnostic substances, with structures postulated. Synthesis of the four proposed structures is recommended to enable structural confirmations. This research will guide and inform the Oil Sands Monitoring Program in its efforts to assess potential influences of oil sands development on the Athabasca River watershed. [ABSTRACT FROM AUTHOR]

Effects‐directed analysis (EDA) is used to identify the principal toxic components within a complex mixture using iterative steps of chemical fractionation guided by bioassay results. Bioassay selection can be limited in EDA because of the volume requirements for many standardized test methods, and therefore, a reduced‐volume acute toxicity test that also provides whole‐organism responses is beneficial. To address this need, a static, 7‐d, water‐only, reduced‐volume method (50 mL, 10 organisms) was developed for Hyalella azteca that substantially decreases the volume requirements of standard‐volume acute test exposures (200–500 mL of test solution, 15–20 organisms) while maintaining water quality and meeting control survival criteria. Standard‐ and reduced‐volume methods were compared by conducting concurrent toxicity tests with 2 inorganic toxicants (KCl and CdCl2) and 2 organic mixtures of naphthenic acid fraction components (NAFCs) to evaluate test performance. There was no difference between methods when comparing the median lethal concentrations (LC50s) for KCl and both NAFC mixtures (p > 0.05). The LC50s for CdCl2 were statistically different (p = 0.0002); however, this was not considered biologically meaningful because the difference between LC50s was <2‐fold. In conclusion, the reduced‐volume H. azteca test method generated results comparable to standard‐volume test methods and is suitable for use in situations where limited testing material is available, such as when conducting EDA. Environ Toxicol Chem 2020;39:2221–2227. © Her Majesty the Queen in Right of Canada 2020. Reproduced with the permission of the Minister of Environment and Climate Change Canada. [ABSTRACT FROM AUTHOR]

Background: The literature on the prognostic relevance of signet-ring cell (SRC) histology in gastric cancer (GC) is controversial which is most likely related to inconsistent SRC classification based on haematoxylin–eosin staining. We hypothesised that mucin stains can consistently identify SRC-GC and predict GC patient outcome. Methods: We performed a comprehensive literature review on mucin stains in SRC-GC and characterised the mucin expression in 851 Caucasian GC and 410 Asian GC using Alcian Blue (AB)-Periodic Acid-Schiff (PAS), MUC2 (intestinal-type mucin), and MUC5AC (gastric-type mucin). The relationship between mucin expression and histological phenotype [poorly cohesive (PC) including proportion of SRCs, non-poorly cohesive (non-PC), or mucinous (MC)], clinicopathological variables, and patient outcome was analysed. Results: Depending on mucin expression and cut-offs, the positivity rates of SRC-GC reported in the literature varied from 6 to 100%. Patients with MUC2 positive SRC-GC or SRC-GC with (gastro)intestinal phenotype had poorest outcome. In our cohort study, PC with ≥ 10% SRCs expressed more frequently MUC2, MUC5AC, and ABPAS (p < 0.001, p = 0.004 and p < 0.001, respectively). Caucasians with AB positive GC or combined ABPAS-MUC2 positive and MUC5AC negative had poorest outcome (all p = 0.002). This association was not seen in Asian patients. Conclusions: This is the first study to suggest that mucin stains do not help to differentiate between SRC-GC and non-SRC-GC. However, mucin stains appear to be able to identify GC patients with different outcome. To our surprise, the relationship between outcome and mucin expression seems to differ between Caucasian and Asian GC patients which warrants further investigations. [ABSTRACT FROM AUTHOR]

This study builds upon the work of a multiagency consortium tasked with determining cost-effective solutions for the effects of pulp mill effluents on fish reproduction. A laboratory fathead minnow egg production test and chemical characterization tools were used to benchmark 81 effluents from 20 mills across Canada, representing the major pulping, bleaching, and effluent treatment technologies. For Kraft and mechanical pulp mills, effluents containing less than 20 mg/L BOD5 were found to have the greatest probability of having no effects. Organic loading, expressed as the total detected solvent-extractable components by gas chromatography/mass spectrometry (GC/MS), also correlated with decreased egg laying. Exceptions were found for specific Kraft, mechanical, and sulfite mills, suggesting yet unidentified causative agents are involved. Recycled fiber mill effluents, tested for the first time, were found to have little potential for reproductive effects despite large variations in BOD5 and GC/MS profiles. Effluent treatment systems across all production types were generally efficient, achieving a combined 82-98% BOD5 removal. Further reductions of final effluent organic loadings toward the target of less than 20 mg/L are recommended and can be realized through biotreatment optimization, the reduction of organic losses associated with production upsets and selecting best available technologies that reduce organic loadings to biotreatment. [ABSTRACT FROM AUTHOR]

Water is a vital resource for natural ecosystems and human life, and assuring a high quality of water and protecting it from chemical contamination is a major societal goal in the European Union. The Water Framework Directive (WFD) and its daughter directives are the major body of legislation for the protection and sustainable use of European freshwater resources. The practical implementation of the WFD with regard to chemical pollution has faced some challenges. In support of the upcoming WFD review in 2019 the research project SOLUTIONS and the European monitoring network NORMAN has analyzed these challenges, evaluated the state-of-the-art of the science and suggested possible solutions. We give 10 recommendations to improve monitoring and to strengthen comprehensive prioritization, to foster consistent assessment and to support solution-oriented management of surface waters. The integration of effect-based tools, the application of passive sampling for bioaccumulative chemicals and an integrated strategy for prioritization of contaminants, accounting for knowledge gaps, are seen as important approaches to advance monitoring. Including all relevant chemical contaminants in more holistic “chemical status” assessment, using effect-based trigger values to address priority mixtures of chemicals, to better consider historical burdens accumulated in sediments and to use models to fill data gaps are recommended for a consistent assessment of contamination. Solution-oriented management should apply a tiered approach in investigative monitoring to identify toxicity drivers, strengthen consistent legislative frameworks and apply solutions-oriented approaches that explore risk reduction scenarios before and along with risk assessment. [ABSTRACT FROM AUTHOR]

Naphthenic acid fraction components (NAFCs) are constituents of oil sands process-affected water (OSPW), which is generated as a result of unconventional oil production via surface mining in the Athabasca oil sands region. NAFCs are often considered to be major drivers of OSPW toxicity to various taxa, including fishes. However, the molecular targets of these complex mixtures are not fully elucidated. Here we examined the effects in walleye ( Sander vitreus ) embryos after exposure to NAFCs extracted from fresh OSPW. Eleutheroembryos (exposed to 0, 4.2 or 8.3 mg/L NAFCs from 1 day post-fertilization to hatch) were subsampled, measured for growth and deformities, and molecular responses were assessed via real-time polymerase chain reaction (PCR). Fourteen genes were evaluated, with a focus on the aryl-hydrocarbon receptor (AhR) − cytochrome P450 pathway ( arnt , cyp1a1 ), the oxidative stress axis ( cat, gst, sod, gpx1b ), apoptosis ( e.g . casp3 , bax and p53 ), growth factor signaling ( e.g . insulin-like growth factors igf1, igf1b, and igf1bp ), and tissue differentiation ( vim ). NAFC exposure was associated with an increase in the expression of cyp1a1 , and a decrease in gpx1b and ribosomal protein rps40 . These results indicate that NAFC effects on walleye early-life stages may be mediated through oxidative stress via pathways that include AhR. [ABSTRACT FROM AUTHOR]

• Wood frogs can be exposed to NAFCs during critical life history phases. • NAFC-exposed tadpoles demonstrated altered chemical antipredator communication. • Suboptimal behavioral patterns were detected at sublethal NAFC concentrations. • Effects were detected in direction changes but not line crosses in a common assay. • Sublethal thresholds must be identified to inform OSPW release guidelines. Environmental pollutants can disrupt chemical communication between aquatic organisms by interfering with the production, transmission, and/or detection of, as well as responses to, chemical cues. Here, we test the hypothesis that early-life exposure to naphthenic acid fraction compounds (NAFCs) from oil sands tailings disrupts antipredator-associated chemical communication in larval amphibians. Wild adult wood frogs (Rana sylvatica) captured during their natural breeding period were combined (1 female:2 males) in six replicate mesocosms filled with either uncontaminated lakewater or with NAFCs isolated from an active tailings pond in Alberta, Canada, at nominal 5 mg/L concentrations. Egg clutches were incubated and tadpoles maintained in their respective mesocosms for ∼40 days post-hatch. Tadpoles (Gosner stage 25-31) were then transferred individually to trial arenas filled with uncontaminated water and exposed to one of six chemical alarm cue (AC) stimuli solutions following a 3 × 2 × 2 design (3 AC types × 2 stimulus carriers × 2 rearing exposure groups). Relative to control tadpoles, NAFC-exposed tadpoles demonstrated higher baseline activity levels (line crosses and direction changes) when introduced to uncontaminated water. Antipredator responses differed in graded fashion with AC type, with control ACs eliciting the greatest latency to resume activity, water the least, and NAFC-exposed ACs intermediate. Pre- to post-stimulus difference scores were non-significant in control tadpoles, while NAFC-exposed tadpoles demonstrated significantly greater variation. While this suggests that exposure to NAFCs from fertilization through hatching may have interfered with AC production, it is unclear whether the quality or quantity of cues was affected. There was also no clear evidence that NAFC carrier water interfered with ACs or the alarm response in unexposed control tadpoles. These results emphasize the importance of understanding how behavioral and physiological effects of early-life NAFC exposure on critical antipredator responses may persist across life history stages. [ABSTRACT FROM AUTHOR]

The acid-extractable organic compounds (AEOs), including naphthenic acids (NAs), present within oil sands process-affected water (OSPW) receive great attention due to their known toxicity. While recent progress in advanced separation and analytical methodologies for AEOs has improved our understanding of the composition of these mixtures, little is known regarding any variability (i.e., spatial, temporal) inherent within, or between, tailings ponds. In this study, 5 samples were collected from the same location of one tailings pond over a 2-week period. In addition, 5 samples were collected simultaneously from different locations within a tailings pond from a different mine site, as well as its associated recycling pond. In both cases, the AEOs were analyzed using SFS, ESI-MS, HRMS, GC×GC-ToF/MS, and GC- & LC-QToF/MS (GC analyses following conversion to methyl esters). Principal component analysis of HRMS data was able to distinguish the ponds from each other, while data from GC×GC-ToF/MS, and LC- and GC-QToF/MS were used to differentiate samples from within the temporal and spatial sample sets, with the greater variability associated with the latter. Spatial differences could be attributed to pond dynamics, including differences in inputs of tailings and surface run-off. Application of novel chemometric data analyses of unknown compounds detected by LC- and GC-QToF/MS allowed further differentiation of samples both within and between data sets, providing an innovative approach for future fingerprinting studies. [ABSTRACT FROM AUTHOR]

Aquatic environments are often contaminated with complex mixtures of chemicals that may pose a risk to ecosystems and human health. This contamination cannot be addressed with target analysis alone but tools are required to reduce this complexity and identify those chemicals that might cause adverse effects. Effect-directed analysis (EDA) is designed to meet this challenge and faces increasing interest in water and sediment quality monitoring. Thus, the present paper summarizes current experience with the EDA approach and the tools required, and provides practical advice on their application. The paper highlights the need for proper problem formulation and gives general advice for study design. As the EDA approach is directed by toxicity, basic principles for the selection of bioassays are given as well as a comprehensive compilation of appropriate assays, including their strengths and weaknesses. A specific focus is given to strategies for sampling, extraction and bioassay dosing since they strongly impact prioritization of toxicants in EDA. Reduction of sample complexity mainly relies on fractionation procedures, which are discussed in this paper, including quality assurance and quality control. Automated combinations of fractionation, biotesting and chemical analysis using so-called hyphenated tools can enhance the throughput and might reduce the risk of artifacts in laboratory work. The key to determining the chemical structures causing effects is analytical toxicant identification. The latest approaches, tools, software and databases for target-, suspect and non-target screening as well as unknown identification are discussed together with analytical and toxicological confirmation approaches. A better understanding of optimal use and combination of EDA tools will help to design efficient and successful toxicant identification studies in the context of quality monitoring in multiply stressed environments. [ABSTRACT FROM AUTHOR]

There is a growing need for environmental screening of natural waters in the Athabasca region of Alberta, Canada, particularly in the differentiation between anthropogenic and naturally-derived organic compounds associated with weathered bitumen deposits. Previous research has focused primarily upon characterization of naphthenic acids in water samples by negative-ion electrospray ionization methods. Atmospheric pressure photoionization is a much less widely used ionization method, but one that affords the possibility of observing low polarity compounds that cannot be readily observed by electrospray ionization. This study describes the first usage of atmospheric pressure photoionization Fourier transform ion cyclotron resonance mass spectrometry (in both positive-ion and negative-ion modes) to characterize and compare extracts of oil sands process water, river water, and groundwater samples from areas associated with oil sands mining activities. When comparing mass spectra previously obtained by electrospray ionization and data acquired by atmospheric pressure photoionization, there can be a doubling of the number of components detected. In addition to polar compounds that have previously been observed, low-polarity, sulfur-containing compounds and hydrocarbons that do not incorporate a heteroatom were detected. These latter components, which are not amenable to electrospray ionization, have potential for screening efforts within monitoring programs of the oil sands. [Figure not available: see fulltext.] [ABSTRACT FROM AUTHOR]

A rapid and sensitive liquid chromatography quadrupole time of flight method has been established for the determination of total naphthenic acid concentrations in aqueous samples. This is the first methodology that has been adopted for routine, high resolution, high throughput analysis of total naphthenic acids at trace levels in unprocessed samples. A calibration range from 0.02 to 1.0 μg mL −1 total Merichem naphthenic acids was validated and demonstrated excellent accuracy (97–111% recovery) and precision (1.9% RSD at 0.02 μg mL −1 ). Quantitative validation was also demonstrated in a non-commercial oil sands process water (OSPW) acid extractable organics (AEOs) fraction containing a higher percentage of polycarboxylic acid isomers than the Merichem technical mix. The chromatographic method showed good calibration linearity of ≥0.999 RSQ to 0.005 μg mL −1 total naphthenic acids with a precision <3.1% RSD and a calculated detection limit of 0.0004 μg mL −1 employing Merichem technical mix reference material. The method is well suited to monitoring naturally occurring and industrially derived naphthenic acids (and other AEOs) present in surface and ground waters in the vicinity of mining developments. The advantage of the current method is its direct application to unprocessed environmental samples and to examine natural naphthenic acid isomer profiles. It is noted that where the isomer profile of samples differs from that of the reference material, results should be considered semi-quantitative due to the lack of matching isomer content. The fingerprint profile of naphthenic acids is known to be transitory during aging and the present method has the ability to adapt to monitoring of these changes in naphthenic acid content. The method's total ion scan approach allows for data previously collected to be examined retrospectively for specific analyte mass ions of interest. A list of potential naphthenic acid isomers that decrease in response with aging is proposed and a quantitative assay of an adamantane carboxylic acid is reported. [ABSTRACT FROM AUTHOR]

Naphthenic acids (NAs) are constituents of oil sands process-affected water (OSPW). These compounds can be both toxic and persistent and thus are a primary concern for the ultimate remediation of tailings ponds in northern Alberta’s oil sands regions. Recent research has focused on the toxicity of NAs to the highly vulnerable early life-stages of fish. Here we examined fathead minnow embryonic survival, growth and deformities after exposure to extracted NA fraction components (NAFCs), from fresh and aged oil sands process-affected water (OSPW), as well as commercially available NA mixtures. Commercial NA mixtures were dominated by acyclic O 2 species, while NAFCs from OSPW were dominated by bi- and tricyclic O 2 species. Fathead minnow embryos less than 24 h old were reared in tissue culture plates terminating at hatch. Both NAFC and commercial NA mixtures reduced hatch success, although NAFCs from OSPW were less toxic (EC50 = 5–12 mg/L, nominal concentrations) than commercial NAs (2 mg/L, nominal concentrations). The toxicities of NAFCs from aged and fresh OSPW were similar. Embryonic heart rates at 2 days post-fertilization (dpf) declined with increasing NAFC exposure, paralleling patterns of hatch success and rates of cardiovascular abnormalities (e.g., pericardial edemas) at hatch. Finfold deformities increased in exposures to commercial NA mixtures, not NAFCs. Thus, commercial NA mixtures are not appropriate surrogates for NAFC toxicity. Further work clarifying the mechanisms of action of NAFCs in OSPW, as well as comparisons with additional aged sources of OSPW, is merited. [ABSTRACT FROM AUTHOR]

Although bicyclic acids have been reported to be the major naphthenic acids in oil sands process-affected water (OSPW) and a well-accepted screening assay indicated that some bicyclics were the most acutely toxic acids tested, none have yet been identified. Here we show by comprehensive multidimensional gas chromatography–mass spectrometry (GC × GC–MS), that >100 C 8–15 bicyclic acids are typically present in OSPW. Synthesis or purchase allowed us to establish the GC × GC retention times of methyl esters of numerous of these and the mass spectra and published spectra of some additional types, allowed us to identify bicyclo[2.2.1]heptane, bicyclo[3.2.1]octane, bicyclo[4.3.0]nonane, bicyclo[3.3.1]nonane and bicyclo[4.4.0]decane acids in OSPW and a bicyclo[2.2.2]octane acid in a commercial acid mixture. The retention positions of authentic bicyclo[3.3.0]octane and bicyclo[4.2.0]octane carboxylic acid methyl esters and published retention indices, showed these were also possibilities, as were bicyclo[3.1.1]heptane acids. Bicyclo[5.3.0]decane and cyclopentylcyclopentane carboxylic acids were ruled out in the samples analysed, on the basis that the corresponding alkanes eluted well after bicyclo[4.4.0]decane (latest eluting acids). Bicyclo[4.2.1]nonane, bicyclo[3.2.2]nonane, bicyclo[3.3.2]decane, bicyclo[4.2.2]decane and spiro[4.5]decane carboxylic acids could not be ruled out or in, as no authentic compounds or literature data were available. Mass spectra of the methyl esters of the higher bicyclic C 12–15 acids suggested that many were simply analogues of the acids identified above, with longer alkanoate chains and/or alkyl substituents. Our hypothesis is that these acids represent the biotransformation products of the initially somewhat more bio-resistant bicyclanes of petroleum. Although remediation studies suggest that many bicyclic acids can be relatively quickly removed from suitably treated OSPW, examination by GC × GC–MS may show which isomers are affected most. Knowledge of the structures will allow the toxicity of any residual isomers to be calculated and measured. [ABSTRACT FROM AUTHOR]

RATIONALE: Characterising complex mixtures of organic compounds in polar fractions of heavy petroleum is challenging, but is important for pollution studies and for exploration and production geochemistry. Oil sands process-affected water (OSPW) stored in large tailings ponds by Canadian oil sands industries contains such mixtures. METHODS: A polar OSPW fraction was obtained by silver ion solid-phase extraction with methanol elution. This was examined by numerous methods, including electrospray ionisation (ESI) Fourier transform ion cyclotron resonance mass spectrometry (FTICRMS) and ultra-high-pressure liquid chromatography (uHPLC)/Orbitrap MS, in multiple ionisation and MS/MS modes. Compounds were also synthesised for comparison. RESULTS: The major ESI ionisable compounds detected (+ion mode) were C15-28 SO3 species with 3-7 double bond equivalents (DBE) and C27-28 SO5 species with 5 DBE. ESI-MS/MS collision-induced losses were due to water, methanol, water plus methanol and water plus methyl formate, typical of methyl esters of hydroxy acids. Once the fraction was re-saponified, species originally detected by positive ion MS, could be detected only by negative ion MS, consistent with their assignment as sulphur-containing hydroxy carboxylic acids. The free acid of a keto dibenzothiophene alkanoic acid was added to an unesterified acid extract of OSPW in known concentrations as a putative internal standard, but attempted quantification in this way proved unreliable. CONCLUSIONS: The results suggest the more polar acidic organic SO3 constituents of OSPW include C15-28 S-containing, alicyclic and aromatic hydroxy carboxylic acids. SO5 species are possibly sulphone analogues of these. The origin of such compounds is probably via further biotransformation (hydroxylation) of the related S-containing carboxylic acids identified previously in a less polar OSPW fraction. The environmental risks, corrosivity and oil flow assurance effects should be easier to assess, given that partial structures are now known, although further identification is still needed. [ABSTRACT FROM AUTHOR]

Abstract: Pulp and paper wood feedstocks have been previously implicated as a source of chemicals with the ability to interact with or disrupt key neuroendocrine endpoints important in the control of reproduction. We tested nine Canadian conifers commonly used in pulp and paper production as well as 16 phytochemicals that have been observed in various pulp and paper mill effluent streams for their ability to interact in vitro with the enzymes monoamine oxidase (MAO), glutamic acid decarboxylase (GAD), and GABA-transaminase (GABA-T), and bind to the benzodiazepine-binding site of the GABA(A) receptor (GABA(A)-BZD). These neuroendocrine endpoints are also important targets for treatment of neurological disorders such as anxiety, epilepsy, or depression. MAO and GAD were inhibited by various conifer extracts of different polarities, including major feedstocks such as balsam fir, black spruce, and white spruce. MAO was selectively stimulated or inhibited by many of the tested phytochemicals, with inhibition observed by a group of phenylpropenes (e.g. isoeugenol and vanillin). Selective GAD inhibition was also observed, with all of the resin acids tested being inhibitory. GABA(A)-BZD ligand displacement was also observed. We compiled a table identifying which of these phytochemicals have been described in each of the species tested here. Given the diversity of conifer species and plant chemicals with these specific neuroactivities, it is reasonable to propose that MAO and GAD inhibition reported in effluents is phytochemical in origin. We propose disruption of these neuroendocrine endpoints as a possible mechanism of reproductive inhibition, and also identify an avenue for potential research and sourcing of conifer-derived neuroactive natural products. [Copyright &y& Elsevier]

Exposure to compounds present in petroleum and wastewaters from oil and gas extraction sites in the Alberta Oil Sands Region can impair reproductive health. It has been established that acid extractable organics found in oil sands process-affected water (OSPW) such as naphthenic acids (NA-fraction components; NAFC) can adversely affect reproductive outcomes. We have shown that NAFC exposure results in a significant upregulation of GDF15 in placental trophoblasts, a cellular stress marker known to be involved in human embryonic development and necessary for the maintenance of pregnancy. However, little is known regarding the mechanism(s) underlying NAFC-induced increases in GDF15 production during early placentation. The goal of this study was to examine the effects of NAFC exposure on the regulation of critical transcription factors of GDF15 in extravillous trophoblast cells. Of these transcription factors, inflammatory mediators including prostaglandins have been reported to inhibit proliferation and migration of trophoblast cells in vitro. Hence, the secondary goal of this study was to determine whether inflammation mediated through prostaglandin production is critical to GDF15 secretion. HTR-8/SVneo cells were exposed to an NAFC for 6 and 24 h to assess the expression of key transcriptional regulators, GDF15 secretion, and prostaglandin (PGE 2) output. Treatment with NAFC (125 mg/L only) significantly increased GDF15 expression and secretion in association with upregulation of the transcription factors KLF4 , EGR1 , ATF3 and TP53. Similarly, PTGS2 (i.e. COX2) expression and PGE 2 output were significantly increased at the same concentration. However, co-treatment with a COX2 selective antagonist (SC236) only partially blocked the NAFC-induced increase in PGE 2 output and did not block GDF15 expression or secretion. These findings suggest that while NAFC may affect GDF15 production, it is not exclusively a result of prostaglandin-mediated inflammation. This study provides new insights into the mechanisms by which NAFC may adversely affect placental trophoblast cell function in mammals. The graphical abstract was created with BioRender.com. [Display omitted] • The effect of naphthenic acid fraction components (NAFC) on placenta is unknown. • NAFC increased GDF15 and its transcription factors in placental trophoblasts. • NAFC increased prostaglandin output, indicative of an inflammatory response. • GDF15 induction does not appear to be linked to increased prostaglandin synthesis. • Consequences of increased GDF15 in early placentation remain to be determined. [ABSTRACT FROM AUTHOR]

Abstract: Previous studies have shown that mummichog (Fundulus heteroclitus; a lunar, asynchronous-spawning killifish of the western Atlantic) exposed to 17α-ethynylestradiol (EE2) exhibit decreased plasma reproductive steroid levels, decreased gonadal steroid production, increased plasma vitellogenin, decreased fecundity and impaired fertilization. The objective of this study was to determine the potential mechanisms by which EE2 depresses gonadal steroidogenesis and influences estrogen signalling in the mummichog. Adult recrudesced fish were exposed to the potent synthetic estrogen, ethinylestradiol (EE2; 0–270ng/L) for 14 days. Following exposure, gonadal tissue was removed and incubated for 24h with stimulators of steroidogenesis, including forskolin; 25-OH cholesterol; or pregnenolone. Testosterone production was decreased in basal, forskolin-stimulated and pregnenolone-stimulated EE2-exposed males, indicating effects on the steroidogenic pathway both at and downstream of cholesterol mobilization to P450 side-chain cleavage (P450scc) and/or P450scc conversion of cholesterol to pregnenolone. Hepatic transcript levels of estrogen receptor alpha (ERα) and vitellogenin were increased in EE2-treated males compared to control recrudescing males and females confirming an estrogenic response. Hepatic heat shock protein 90 (Hsp90), a chaperoning molecule involved in estrogen signalling, was not affected by EE2 exposure at either the transcript or protein level. However, higher levels of Hsp90 observed in the membrane fractions of female fish raise interesting questions regarding the influence of gender on Hsp90''s role in estrogen signalling. These results demonstrate that EE2 can alter steroid production at specific sites within the steroidogenic pathway and can stimulate hepatic estrogen signalling, providing important information regarding the molecular mechanisms underlying the endocrine response of the mummichog to exogenous estrogen. [Copyright &y& Elsevier]

Abstract: We tested the hypothesis that gross morphological abnormalities are a sensitive indicator of exposure to waterborne androgenic and anti-androgenic compounds during embryonic, larval and juvenile stages of development in the common estuarine killifish, the mummichog (Fundulus heteroclitus; Pisces: Cyprinodontidae). Static exposures with daily renewal were carried out with 10–100,000ng/L of the androgen agonist, 17α-methyltestosterone (MT), or the androgen antagonist, cyproterone acetate (CA), for 60 days post-fertilization (PF) in duplicate exposures. Measured concentrations were 78.4–155.8% of nominal concentrations for MT and 13.5–168.1% for CA. No dose-related or consistent effects of MT or CA were observed before hatch. In 60 days PF juveniles, incidence of skeletal abnormalities (scoliosis, lordosis, head, facial and fin), soft tissue abnormality (anal swelling) and hemorrhaging were significantly increased by MT but only at high concentrations (≥1000ng/L). The 10,000 and 100,000ng/L concentrations of MT produced a wider range of abnormalities than 1000ng/L. Over 90% of fish exposed to 10,000 or 100,000ng/L were abnormal with an average of over 3.5 abnormalities per fish. CA did not increase the incidence of any type of abnormality. Survival of juveniles to the end of the exposure was reduced by MT at concentrations of 1000ng/L and greater in the first experiment and at concentrations of 10,000ng/L and greater in the second experiment. Juvenile length was reduced by high concentrations of MT (≥10,000ng/L) in the first experiment and by most concentrations in the second experiment. We conclude that morphological abnormalities in early-life stages of mummichogs are not a sensitive indicator of exposure to androgenic or anti-androgenic waterborne EDSs at environmentally relevant concentrations. [Copyright &y& Elsevier]

The oil sands area of northern Alberta has river sediments that contain natural bitumen. Eggs and fish in these rivers may be exposed to bitumen-related chemicals early in life. This paper assesses a short embryo-larval fish exposure to oil sands sediment and follows the fish behaviour as they mature in clean water and examines their breeding success as adults (5 months afterwards). The three different oil sands river sediments tested were: a sediment collected outside of the bitumen deposit (tested at 3 g/L, Reference sediment from upstream Steepbank River site), and two sediments collected within the deposit (each tested at low (1 g/L) and high (3 g/L) concentrations). The sediments within the bitumen deposit were from the Ells and Steepbank (Stp) Rivers, and both contained significant total PAHs (>170 ng/g wet weight sediment) and alkylated PAHs (>4480 ng/g). Fish were exposed to these sediments for 21 days (as eggs and larval fish), and then transferred permanently to clean water to mature and breed. There was a significant decrease in the number of egg clutches produced by fish exposed early in life to Stp downstream high sediment (compared to Reference sediment). There was also a decrease in overall cumulative egg production, with fish from Stp downstream high sediment producing just over 1000 eggs in total while fish exposed to Ref sediment produced nearly 6900 eggs. The fish with reduced egg production were also less social than expected as they matured, and they had a lower % of early vitellogenic eggs in their ovaries. Overall, the exposure shows that a single, brief exposure during early life stages to natural bitumen can affect fish in adulthood. Naturally occurring bitumen-derived PAHs can reduce fish reproductive output by complex mechanisms, measurable as lower ovary maturity and changes in social behaviour., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Crown Copyright © 2024. Published by Elsevier Ltd. All rights reserved.)

Aims: This study describes the pharmacokinetic (PK)/target engagement (TE) relationship of tozorakimab, an anti-interleukin (IL)-33 antibody, by building a mechanistic population PK/TE model using phase 1 biomarker data., Methods: The analysis included tozorakimab PK and TE in serum assessed in 60 tozorakimab-treated participants, including healthy adults and patients with mild chronic obstructive pulmonary disease. Scenarios evaluated three dose frequencies (once every 2, 4 or 6 weeks) administered subcutaneously at seven doses of tozorakimab (30, 60, 90, 120, 150, 300 or 600 mg). For each dose, simulations were performed with 5000 virtual individuals to predict systemic TE. Inhibition of IL-33/soluble ST2 (sST2) complex levels at trough PK at steady state was assessed in each dosing scenario. The PK/TE modelling analyses were performed using a nonlinear mixed-effect modelling approach., Results: The final two-compartment PK model with tozorakimab binding IL-33 in the central compartment adequately described the systemic PK and TE of tozorakimab at population and individual levels. The mean PK parameter estimates of absorption rate, central volume of distribution and clearance were 0.48 (90% confidence interval [CI]: 0.40-0.59, 1/day), 12.64 (90% CI: 8.60-18.62, L) and 0.87 (90% CI: 0.65-1.16, L/day), respectively. Consistent with the observed value, tozorakimab bioavailability was 45%. For all three dose frequencies, predicted inhibition of systemic IL-33/sST2 levels was more than 95% at doses greater than 90 mg., Conclusions: The PK/TE model reliably quantified the relationship between PK and systemic TE of tozorakimab, with potential utility for predicting clinical dose-response relationships and supporting clinical dose selection., (© 2024 AstraZeneca. British Journal of Clinical Pharmacology published by John Wiley & Sons Ltd on behalf of British Pharmacological Society.)

Opioid receptors within the CNS regulate pain sensation and mood and are key targets for drugs of abuse. Within the adult rodent hippocampus (HPC), μ-opioid receptor agonists suppress inhibitory parvalbumin-expressing interneurons (PV-INs), thus disinhibiting the circuit. However, it is uncertain if this disinhibitory motif is conserved in other cortical regions, species, or across development. We observed that PV-IN mediated inhibition is robustly suppressed by opioids in hippocampus proper but not neocortex in mice and nonhuman primates, with spontaneous inhibitory tone in resected human tissue also following a consistent dichotomy. This hippocampal disinhibitory motif was established in early development when PV-INs and opioids were found to regulate primordial network rhythmogenesis. Acute opioid-mediated modulation was partially occluded with morphine pretreatment, with implications for the effects of opioids on hippocampal network activity important for learning and memory. Together, these findings demonstrate that PV-INs exhibit a divergence in opioid sensitivity across brain regions that is remarkably conserved across evolution and highlights the underappreciated role of opioids acting through immature PV-INs in shaping hippocampal development., Competing Interests: DECLARATION OF INTERESTS The authors declare no competing interests.

Responding to global standards and legislative updates in Canada, including Bill S-5 (2023), toxicity testing is shifting towards more ethical, in vitro methods. Traditional two-dimensional (2D) monolayer cell cultures, limited in replicating the complex in vivo environment, have prompted the development of more relevant three-dimensional (3D) spheroidal hepatocyte cultures. This study introduces the first 3D spheroid model for McA-RH7777 cells, assessing xenobiotic receptor activation, cellular signaling, and toxicity against dexamethasone and naphthenic acid (NA)-fraction components; NAFCs. Our findings reveal that 3D McA-RH7777 spheroids demonstrate enhanced sensitivity and more uniform dose-response patterns in gene expression related to xenobiotic metabolism (AhR and PPAR) for both single compounds and complex mixtures. Specifically, 3D cultures showed significant gene expression changes upon dexamethasone exposure and exhibited varying degrees of sensitivity and resistance to the apoptotic effects induced by NAFCs, in comparison to 2D cultures. The optimization of 3D culture conditions enhances the model's physiological relevance and enables the identification of genomic signatures under varied exposures. This study highlights the potential of 3D spheroid cultures in providing a more accurate representation of the liver's microenvironment and advancing our understanding of cellular mechanisms in toxicity testing.

Saltwater crocodiles ( Crocodylus porosus ) are farmed in Australia primarily for their skins and meat. Commercially, they are raised in group pens as hatchlings and grower crocodiles and then moved to unitised (individual) pens for the final finishing stage when they are several years old. They will exhibit aggressive behaviour towards each other in captivity. Unitised pens can prevent animal injury and teeth marks on the skins but may result in other social restrictions. Research into behavioural housing preferences could assist the industry and inform the process of guideline development for optimal crocodile management and welfare. This study assessed the impacts of two housing systems, unitised or group pens, in 20 commercial finishing crocodiles through measuring behavioural profiles of individuals from video recordings, including housing preference when given a choice. Both pens included water and an above-water shelf, but the crocodiles in unitised pens could also access underneath the shelf. A threat perception test was applied to assess anxiety when housed individually or in groups. However, it was difficult to apply a standardised stimulus to all animals that reliably elicited a behavioural response. Further work would be needed to validate this test for commercial reptiles as the outcomes were not robust. The behavioural observation results showed clear differences in where the crocodiles spent their time across the day and in their activity levels between the pen types. However, interpretation of this variation was confounded by the physical and social differences between the pen types given the inconsistency in shelf access. Behaviours exhibited also differed given there were social opportunities in the group pens where individuals were observed engaged in both aggressive and non-aggressive contact interactions. In the free choice environment, crocodiles spent similar amounts of time in both unitised and group pens, suggesting there were features of both pen types that were attractive to the animals. However, skins were damaged from teeth marks highlighting the physical and economical risks of group housing. Further work could validate behavioural tests to quantify affective state impacts in different housing environments and whether social interactions do provide benefits for improving crocodile welfare., Competing Interests: The authors declare that this study received funding from Primary Resource Investments Pty Ltd. The funder had the following involvement in the study: care and management of the animals, downloading of video footage and temperature logger data, and provided consent to publish., (Copyright © 2024 Campbell, Hewitt, Lee, Timmerhues and Small.)

Killing of farmed saltwater crocodiles involve stunning with a penetrative captive bolt device, followed by a cut across the nape of the neck and physical destruction of the brain to ensure death. This study was a welfare-based assessment of the use of a penetrative captive bolt device in saltwater crocodiles, to determine whether it satisfies the criteria of a humane stunning method and can be regarded as a direct killing method without the need for the application of an adjunct method. Methods used were electroencephalogram (EEG), observation of post-stun behavior, and postmortem examination of gross pathology of the cranium and brain. 30 of 30 animals, demonstrated immediate and irreversible loss of consciousness. There was extensive damage to the brain in all animals, deemed to be inconsistent with cortical function and possible recovery. The CASH Special 0.22 penetrative captive bolt pistol (1.25 grain cartridge), applied to the top of the cranial plate, produced immediate and irreversible unconsciousness in all the animals studied. This method satisfies animal welfare expectations, providing crocodile processors with a technique that contributes to a humane killing process.

Purpose: To explore the factors that influence clinicians (occupational therapists, physiotherapists, vascular surgeons, and rehabilitation medicine physicians) when prescribing prosthetic rehabilitation. Additionally, the study aimed to gain insight into clinicians' perspectives regarding the role of patient cognition in prosthetic rehabilitation., Materials and Methods: This research constitutes one segment of a broader action research study which was undertaken in 2022. A total of thirty-four key clinicians involved in the amputation and prosthetic rehabilitation pathway within a local health district in Australia were engaged through a combination of group and individual interviews as well as surveys., Results: Five essential considerations when prescribing prosthetic rehabilitation emerged. These included patient's goals, medical history, quality of life, cognitive abilities, and the support available on discharge. This study also revealed variations in opinions among different disciplines concerning appropriateness of prosthetic rehabilitation for the patient cohort. Despite this, there was a desire to build a consensus around a shared approach of identification for patients and clinicians., Conclusion: The identification of these key pillars for clinician consideration has simplified a complex area of care. These pillars could be used to guide pertinent conversations regarding prosthetic rehabilitation and are closely linked with the patient's cognition.

Problem: Although there is robust evidence for the benefits of midwifery group practice (MGP) caseload care, there are limited opportunities for women to access this model in Australia. There is also limited knowledge on how to sustain these services., Background: MGP can benefit childbearing women and babies and improve satisfaction for women and midwives. However, sustainability of the model is challenging. While MGPs are often supported and celebrated, in Australia some services have closed, while others struggle to adequately staff MGPs., Aim: To investigate midwives and managers opinions on the management, culture, and sustainability of MGP., Methods: A national survey of MGP midwives and managers was distributed (2021 and 2022). Quantitative data were analysed using descriptive statistics, and qualitative data were analysed using content analysis., Results: A total of 579 midwives and 90 managers completed the survey. The findings suggest that many MGPs do not support new graduates and students to work in MGP. Over half (59.8%) the participants (midwives and managers) reported that the women and families were the best aspect about working in MGP, while 44.3% said the effects on midwives' lifestyle and families were the worst aspect., Discussion: The relationship with women remains the major motivator for providing MGP care. However, work-life imbalance is a deterrent, exacerbated by staffing shortages. Staffing might be improved by adequate renumeration, strengthening orientation, and attracting new graduates and students through experience in MGP., Conclusions: There is a need to attract midwives to MGP and improve work-life balance and sustainability., Competing Interests: Declaration of Competing Interest None declared., (Copyright © 2024 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related deaths globally, including Australia. The absence of a consensus clinical practice guideline (CPG) specific to HCC management poses challenges in reducing morbidity, mortality, and improving patient recovery. This systematic review aims to evaluate the existing evidence and assess the potential of published guidelines, including those with an international scope, to provide guidance for healthcare professionals in Australia., Methods: Electronic search of MEDLINE, Embase, Cochrane Library, Google Scholar, and PubMed was conducted. Peer-reviewed English language articles from 2005 to June 2022 were included if they described management of HCC as part of an evidence-based overall management plan or CPG. The quality of the included CPGs was assessed by the Appraisal of Guidelines for Research and Evaluation II (AGREE II) tool., Results: Twenty-one articles from 16 regions throughout the world were included in this review. All included guidelines (n = 21, 100%) recommended evaluating cirrhosis, hepatitis B, and hepatitis C as potential risk factors of HCC. Obesity and non-alcoholic fatty liver disease were recommended by 19 CPGs (91%) as risk factor for HCC. Fourteen guidelines (67%) endorsed using the BCLC staging system. Eighteen guidelines (86%) recommended a multidisciplinary approach for the management of HCC. Eighteen guidelines (86%) advised that surveillance using ultrasound should be implemented in all cirrhotic patients every 6 months regardless of the cause of cirrhosis. AGREE II mean overall assessment score was 90% indicating that all guidelines included were highly recommended in majority of domains., Conclusions: The included CPGs provided a comprehensive approach, emphasizing the evaluation of risk factors, utilization of the BCLC staging system, and the importance of a multidisciplinary approach. Regular surveillance using ultrasound for cirrhotic patients was widely recommended. An understanding of contemporary international CPGs can prioritize aspects of the management of HCC to assist healthcare professionals to develop a national guideline to enable standardized, comprehensive, and evidence-based care for patients with HCC., (© 2023. Crown.)

Tozorakimab is a human monoclonal antibody that neutralizes interleukin (IL)-33. IL-33 is a broad-acting epithelial "alarmin" cytokine upregulated in lung tissue of patients with chronic obstructive pulmonary disease (COPD). This first-in-human, phase I, randomized, double-blind, placebo-controlled study (NCT03096795) evaluated the safety, tolerability, pharmacokinetics (PKs), immunogenicity, target engagement, and pharmacodynamics (PDs) of tozorakimab. This was a 3-part study. In part 1, 56 healthy participants with a history of mild atopy received single escalating doses of either intravenous or subcutaneous tozorakimab or placebo. In part 2, 24 patients with mild COPD received multiple ascending doses of subcutaneous tozorakimab or placebo. In part 3, 8 healthy Japanese participants received a single intravenous dose of tozorakimab or placebo. The safety data collected included treatment-emergent adverse events (TEAEs), vital signs, and clinical laboratory parameters. Biological samples for PKs, immunogenicity, target engagement, and PD biomarker analyses were collected. No meaningful differences in the frequencies of TEAEs were observed between the tozorakimab and placebo arms. Three tozorakimab-treated participants with COPD experienced treatment-emergent serious adverse events. Subcutaneous or intravenous tozorakimab demonstrated linear, time-independent PKs with a mean half-life of 11.7-17.3 days. Treatment-emergent anti-drug antibody frequency was low. Engagement of tozorakimab with endogenous IL-33 in serum and nasal airways was demonstrated. Tozorakimab significantly reduced serum IL-5 and IL-13 levels in patients with COPD compared with placebo. Overall, tozorakimab was well tolerated, with a linear, time-independent serum PK profile. Additionally, biomarker studies demonstrated proof of mechanism. Overall, these data support the further clinical development of tozorakimab in COPD and other inflammatory diseases., (© 2023 AstraZeneca and The Authors. Clinical Pharmacology & Therapeutics published by Wiley Periodicals LLC on behalf of American Society for Clinical Pharmacology and Therapeutics.)

Background: Longer-term symptoms (long COVID) may be present in seemingly recovered patients for several months and can be debilitating., Aim: To investigate the prevalence and type of symptoms in those with a prior COVID-19 diagnosis., Methods: This prospective, longitudinal observational study commenced in July 2020 investigating the longer-term health impacts of COVID-19. Participants were recruited via public health units and media publicity. Surveys were completed upon enrolment, and at 1, 3, 6 and 12 months. Outcome measures included incidence of activity limitations and symptoms against health and vaccination status, age and gender., Results: Overall, 339 participants were recruited. At 3 months after COVID-19, 66.8% reported symptoms, and 44.8% were still experiencing symptoms at 12 months. Fatigue was most common at every point (between 53.1% and 33.1%). Pain symptoms increased in relative prevalence over time, whereas respiratory/pulmonary-type symptoms decreased substantially after 3 months. Females and younger people were more likely to experience symptoms in the early stages of long COVID (P < 0.01) and those with more comorbidities in the latter stages (P < 0.001). Vaccination showed a statistically significant protective effect against symptoms (P < 0.01-0.001)., Conclusion: Long-term COVID-19 symptoms exist among recovered patients up to 12 months after contracting the virus. Fatigue is a primary contributor, while chronic pain became more problematic after 6 months. Vaccination was a factor in preventing long-term symptoms and aiding faster recovery from symptoms. Further work exploring additional contributors to symptom prevalence would assist in developing appropriate follow-up care., (© 2023 The Authors. Internal Medicine Journal published by John Wiley & Sons Australia, Ltd on behalf of Royal Australasian College of Physicians.)

Background: Despite robust evidence on the benefits of midwifery group practice (MGP), there remains difficulties with implementing and sustaining the model. However, contemporary data on the MGP workforce and how each model has been operationalised are limited. This constrains an understanding of the factors that help or hinder implementation and sustainability of MGP., Aim: To describe the characteristics of Australian MGPs and the factors that help or hinder sustainability., Methods: A national cross-sectional survey was undertaken in Australia between March 2021 and July 2022, inclusive. Quantitative data were analysed using descriptive analysis while qualitative data were analysed using content analysis., Findings: Of 669 survey responses, 579 were midwives and 90 were managers. The mean years of experience for clinical midwives was eight years, and 47.8% (almost twice the national average) completed a Bachelor of Midwifery (BMid). Half (50.2%) the models provided care for women of all risk. Midwives resigned from MGP because of the MGP work conditions (30%) and how the service was managed or supported (12.7%). Managers resigned from MGP because of role changes, conflict with their manager, and limited support. Almost half (42.6%) of MGP managers also managed other areas, leading to heavy workloads, competing demands, and burnout., Conclusion: The BMid appears to be a common educational pathway for MGP midwives, and many MGP services are providing care to women with complexities. Flexible practice agreements, organisational support and appropriate workloads are vital for recruitment, retention, and sustainability of MGP., Competing Interests: Declaration of Competing Interest None declared., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The Guidelines for Qualifications of Neurodiagnostic Personnel (QNP) document has been created through the collaboration of the American Clinical Neurophysiology Society (ACNS), the American Society of Neurophysiological Monitoring (ASNM), the American Association of Neuromuscular & Electrodiagnostic Medicine (AANEM), and ASET The Neurodiagnostic Society (ASET). The quality of patient care is optimized when neurophysiological procedures are performed and interpreted by appropriately trained and qualified practitioners at every level. These societies recognize that neurodiagnostics is a large field with practitioners who have entered the field through a variety of training paths. This document suggests job titles, associated job responsibilities, and the recommended levels of education, certification, experience, and ongoing education appropriate for each job. This is important because of the growth and development of standardized training programs, board certifications, and continuing education in recent years. This document matches training, education, and credentials to the various tasks required for performing and interpreting neurodiagnostic procedures. This document does not intend to restrict the practice of those already working in neurodiagnostics. It represents recommendations of these societies with the understanding that federal, state, and local regulations, as well as individual hospital bylaws, supersede these recommendations. Because neurodiagnostics is a growing and dynamic field, the authors fully intend this document to change over time., (Copyright © 2023 The Author(s). Published by Wiley Periodicals LLC on behalf of American Association of Neuromuscular & Electrodiagnostic Medicine, Inc.)

Expert problem-solving is driven by powerful languages for thinking about problems and their solutions. Acquiring expertise means learning these languages-systems of concepts, alongside the skills to use them. We present DreamCoder, a system that learns to solve problems by writing programs. It builds expertise by creating domain-specific programming languages for expressing domain concepts, together with neural networks to guide the search for programs within these languages. A 'wake-sleep' learning algorithm alternately extends the language with new symbolic abstractions and trains the neural network on imagined and replayed problems. DreamCoder solves both classic inductive programming tasks and creative tasks such as drawing pictures and building scenes. It rediscovers the basics of modern functional programming, vector algebra and classical physics, including Newton's and Coulomb's laws. Concepts are built compositionally from those learned earlier, yielding multilayered symbolic representations that are interpretable and transferrable to new tasks, while still growing scalably and flexibly with experience. This article is part of a discussion meeting issue 'Cognitive artificial intelligence'.

The aim of this study was to assess welfare outcomes of electrical stunning as a means of restraint in farmed grower saltwater crocodiles ( Crocodylus porosus ). Physical handling of a stunned, unconscious crocodile is far safer for the operator than handling a fully conscious animal. Electroencephalogram (EEG) was recorded before and after the application of electrical stunning at 50 Hz or 400 Hz using an electrical stunner applied to the cranial plate (Position 1: P1-50 Hz; n = 31, P1-400 Hz; n = 29) or immediately behind the skull (Position 2: P2-50 Hz; n = 29; P2-400 Hz; n = 30). For all electrical stuns, percentage total EEG power in a 10-s epoch decreased in the alpha and beta frequency bands; and increased in the delta and lower frequencies bands. All electrical stuns resulted in increased strength of signal, based on the quadratic mean EEG power in all frequency bands of the EEG. Greatest change in power occurred in the delta frequency band, with P1-50 Hz. This was greater than with P2-50 Hz; while decibel change using 400 Hz at either position was intermediate and not significantly different from either. Application of either electrical stunner at position 1 resulted in seizure-like activity and activation in low frequencies, but at position 2 this was not consistent across all animals. The ability of the electrical stunning equipment to consistently induce recoverable unconsciousness could be ranked in decreasing order as: P1-50 Hz > P1-400 Hz = P2-50 Hz > P2-400 Hz. Based on behavioural observations, all animals in the study appeared to stunned however evaluation of duration of EEG changes indicates that use of the electrical stunning equipment at 50 Hz would allow some margin for inaccuracies in tong placement, while achieving a consistently reliable stun., Competing Interests: Submission of this manuscript to Animal Welfare has been approved by the funding body., (© The Author(s) 2023.)

Many factors contribute to students' academic success, and some, including first-generation (FG) college student status and academic preparation, are especially relevant in physiology programs. The purpose of this qualitative study was to examine FG college students' perceptions of academic preparedness and intended success strategies with the goal of informing program-level strategies to support FG students in similar undergraduate physiology-related programs. FG freshmen completed short surveys and reflections at the start and end of the semester. Qualitative responses from written reflections were compiled, and content was analyzed. Students were high achievers in high school; 98% expected As and Bs in college, but only 53% had achieved this by the end of the first semester. At the start of the semester, FG students reported feeling prepared academically but were hoping to improve their academic readiness skills, and academic success plans focused on organizational strategies. At the end of the semester, some thought they were as prepared as they expected, but most found they were not as academically prepared for college as they had expected. Several minor themes were identified as areas to address with future cohorts. This study proposes several potential avenues by which to support FG freshmen's academic success in similar physiology programs, including early identification of at-risk students, setting realistic expectations, educating students early and often about evidence-based strategies, and developing academic recovery strategies as needed. NEW & NOTEWORTHY A qualitative investigation of first-generation (FG) college freshmen's perceptions of academic preparedness and intended success strategies informs the development of potential avenues by which to support FG freshmen in physiology programs.

Purpose: Proliferation of T-follicular helper (TFH) CD4+ T cells is a postulated pathogenic mechanism for T-cell non-Hodgkin lymphomas (T-NHL). The inducible T-cell costimulator (ICOS) is highly expressed by TFH, representing a potential target. MEDI-570 is a monoclonal antibody against ICOS, which eliminates ICOS+ cells in preclinical models., Patients and Methods: We report the safety, pharmacokinetics (PK), pharmacodynamics (PD), and clinical activity of MEDI-570 in T-NHL. NCI-9930 is a phase I, first-in-human study of MEDI-570 in relapsed/refractory malignant T-NHL known to express ICOS. MEDI-570 was administered intravenously every 3 weeks for up to 12 cycles. Primary endpoints were safety, dose-limiting toxicities (DLT), and recommended phase II dose (RP2D). Secondary and exploratory endpoints included efficacy parameters and various correlative studies. This study is supported by the National Cancer Institute (NCT02520791)., Results: Twenty-three patients were enrolled and received MEDI-570 at five dose levels (0.01-3 mg/kg). Sixteen (70%) had angioimmunoblastic T-cell lymphoma (AITL); median age was 67 years (29-86) and the median prior lines of therapies was 3 (1-16). Most common grade 3 or 4 adverse events were decreased CD4+ T cells (57%), lymphopenia (22%), anemia (13%), and infusion-related reactions (9%). No DLTs were observed. The RP2D was determined at 3 mg/kg. Analysis of T-cell subsets showed reductions in CD4+ICOS+ T cells reflecting its effects on TFH cells. The response rate in AITL was 44%., Conclusions: MEDI-570 was well tolerated and showed promising clinical activity in refractory AITL. MEDI-570 resulted in sustained reduction of ICOS+ T lymphocytes., (©2023 American Association for Cancer Research.)

Background: Understanding the impact of surgery on patients will enable clinicians to provide evidence-based perioperative management. This study aimed to investigate the quality of life (QoL) impacts following head and neck surgery for advanced stage head and neck cancer., Methods: Head and neck cancer survivors were invited to complete five validated questionnaires to investigate QoL. Associations between QoL and patient variables were analyzed. Variables included age, time since operation, length of surgery, length of stay, Comorbidity Index, estimated 10-year survival, sex, flap type, treatment and cancer type. Outcome measures were also compared to normative outcomes., Results: The majority of participants (N = 27; 55% male; mean (standard deviation) age: 62.6 (13.8) years; mean time since operation: 801 days) had a squamous cell carcinoma (88.9%) and free flap repair (100%). Time since operation was significantly (P < 0.05) associated with higher rates of depression (r = -0.533), psychological needs (r = -0.0415) and physical/daily living needs (r = -0.527). Length of surgery and length of stay were significantly associated with depression (r = 0.442; r = 0.435) and length of stay was significantly associated with speaking difficulties (r = -0.456). There was a significant association between work and education scores with age (r = 0.471), length of surgery (r = 0.424), Comorbidity Index (r = 0.456) and estimated 10-year survival (r = -0.523)., Conclusions: Age, time since operation, length of surgery, length of stay, Comorbidity Index and estimated 10-year survival were the outcomes associated with QoL. Patient-reported outcome measures and psychological support could be included in the standard care pathway for head and neck cancer patients to ensure holistic management of their condition., Competing Interests: The authors have no relevant financial or non-financial conflict of interest to disclose., (Copyright 2023, Douglas et al.)

Researchers have reported limitations with research governance processes across Australia. This study aimed to streamline research governance processes across a local health district. Four basic principles were applied to remove non-value-adding and non-risk-mitigating processes. Average processing times were reduced from 29 to 5 days and end-user satisfaction was improved, all within the same staffing levels., (© 2023 Royal Australasian College of Physicians.)

Introduction: Two parallel versions (A and B) of the Oxford Cognitive Screen (OCS) were developed in the United Kingdom (UK) as a stroke-specific screen of five key cognitive domains commonly affected post-stroke. We aimed to develop the Australian versions A and B (OCS-AU), including Australian cut-scores indicative of impairment. We hypothesised there to be no difference in performance between the UK and Australian normative data cohorts., Methods: Our multidisciplinary expert panel used the UK pre-defined process to develop the OCS-AU versions A and B. We then conducted a cross-sectional normative study. We purposively recruited community-dwelling, Australian-born, and educated adults; with no known cognitive impairment; representative of age, sex, education level, and living location; at seven sites (four metropolitan, three regional) across four Australian states. Participants completed one or both OCS-AU versions in a randomised order. Australian cohorts were compared with the corresponding UK cohorts for demographics using Pearson's chi-squared test for sex and education, and Welch two-sample t test for age. For the cut-scores indicating cognitive impairment, the fifth (95th) percentiles and group mean performance score for each scored item were compared using Welch two-sample t tests. The pre-defined criteria for retaining OCS cut-scores had no statistically significant difference in either percentile or group mean scores for each scored item., Results: Participants (n = 83) were recruited: fifty-eight completed version A [age (years) mean = 61,SD = 15; 62% female], 60 completed version B [age (years) mean = 62,SD = 13, 53% female], and 35 completed both [age (years) mean = 64,SD = 11, 54% female]. Education was different between the cohorts for version B (12 years, p = 0.002). Cut-scores for all 16 scored items for the OCS-AU version B and 15/16 for version A met our pre-defined criteria for retaining the OCS cut scores., Conclusions: The OCS-AU provides clinicians with an Australian-specific, first-line cognitive screening tool for people after stroke. Early screening can guide treatment and management., (© 2022 The Authors. Australian Occupational Therapy Journal published by John Wiley & Sons Australia, Ltd on behalf of Occupational Therapy Australia.)

Background: Glioblastoma is a high-grade aggressive neoplasm whose outcomes have not changed in decades. In the current treatment pathway, tumour growth continues and remains untreated for several weeks post-diagnosis. Intensified upfront therapy could target otherwise untreated tumour cells and improve the treatment outcome. POBIG will evaluate the safety and feasibility of single-fraction preoperative radiotherapy for newly diagnosed glioblastoma, assessed by the maximum tolerated dose (MTD) and maximum tolerated irradiation volume (MTIV)., Methods: POBIG is an open-label, dual-centre phase I dose and volume escalation trial that has received ethical approval. Patients with a new radiological diagnosis of glioblastoma will be screened for eligibility. This is deemed sufficient due to the high accuracy of imaging and to avoid treatment delay. Eligible patients will receive a single fraction of preoperative radiotherapy ranging from 6 to 14 Gy followed by their standard of care treatment comprising maximal safe resection and postoperative chemoradiotherapy (60 Gy/30 fr) with concurrent and adjuvant temozolomide). Preoperative radiotherapy will be directed to the part of the tumour that is highest risk for remaining as postoperative residual disease (hot spot). Part of the tumour will remain unirradiated (cold spot) and sampled separately for diagnostic purposes. Dose/volume escalation will be guided by a Continual Reassessment Method (CRM) model. Translational opportunities will be afforded through comparison of irradiated and unirradiated primary glioblastoma tissue., Discussion: POBIG will help establish the role of radiotherapy in preoperative modalities for glioblastoma., Trial Registration: NCT03582514 (clinicaltrials.gov)., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: POBIG is funded by UK Research and Innovation Medical Research Council (UKRI MRC). The research program of GB (Chief Investigator, senior author) is supported by Cancer Research UK RadNet Manchester., (© 2023 The Authors.)

Understanding the toxicity of organic compounds in oil sands process-affected water (OSPW) is necessary to inform the development of environmental guidelines related to wastewater management in Canada's oil sands region. In the present study, we investigated the effects of naphthenic acid fraction compounds (NAFCs), one of the most toxic components of OSPW, on mating behaviour, fertility, and offspring viability in the wood frog (Rana sylvatica). Wild adult wood frogs were exposed separately from the opposite sex to 0, 5, or 10 mg/L of OSPW-derived NAFCs for 24 h and then combined in outdoor lake water mesocosms containing the same NAFC concentrations (n = 2 males and 1 female per mesocosm, n = 3 mesocosms per treatment). Mating events were recorded for 48 h and egg masses were measured to determine adult fertility. NAFC exposure had no significant effect on mating behaviour (probability of amplexus and oviposition, amplexus and oviposition latency, total duration of amplexus and number of amplectic events) or fertility (fertilization success and clutch size). Tadpoles (50 individuals per mesocosm at hatching, and 15 individuals per mesocosm from 42 d post-hatch) were reared in the same mesocosms under chronic NAFC exposure until metamorphic climax (61-85 d after hatching). Offspring exposed to 10 mg/L NAFCs during development were less likely to survive and complete metamorphosis, grew at a reduced rate, and displayed more frequent morphological abnormalities. These abnormalities included limb anomalies at metamorphosis, described for the first time after NAFC exposure. The results of this study suggest that NAFCs reduce wood frog reproductive success through declines in offspring viability and therefore raise the concern that exposure to NAFCs during reproduction and development may affect the recruitment of native amphibian populations in the oil sands region., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

The extraction of bitumen from surface mining in the Athabasca Oil Sands Region (AOSR) produces large quantities of oil sands process-affected water (OSPW) that needs to be stored in settling basins near extraction sites. Chemical constituents of OSPW are known to impair bone health in some organisms, which can lead to increased fracture risk and lower reproductive fitness. Naphthenic acid fraction components (NAFCs) are thought to be among the most toxic class of compounds in OSPW; however, the effect of NAFCs on osteoblast development is largely unknown. In this study, we demonstrate that NAFCs from OSPW inhibit osteoblast differentiation and deposition of extracellular matrix, which is required for bone formation. Extracellular matrix deposition was inhibited in osteoblasts exposed to 12.5-125 mg/L of NAFC for 21 days. We also show that components within NAFCs inhibit the expression of gene markers of osteoblast differentiation and function, namely, alkaline phosphatase (Alp), osteocalcin, and collagen type 1 alpha 1 (Col1a1). These effects were partially mediated by the induction of glucocorticoid receptor (GR) activity; NAFC induces the expression of the GR activity marker genes Sgk1 (12.5 mg/L) and p85a (125 mg/L) and inhibits GR protein (125 mg/L) and Opg RNA (12.5 mg/L) expression. This study provides evidence that NAFC concentrations of 12.5 mg/L and above can directly act on osteoblasts to inhibit bone formation and suggests that NAFCs contain components that can act as GR agonists, which may have further endocrine disrupting effects on exposed wildlife., (© 2022 The Authors. Journal of Applied Toxicology published by John Wiley & Sons Ltd.)

The incidence of head and neck cutaneous squamous cell carcinoma (HNcSCC) is unevenly distributed between men and women. At present, the mechanism behind this disparity remains elusive. This study conducted a systematic review and meta-analysis of proportions to investigate the disparity between sexes for patients with HNcSCC. PubMed, Scopus, EMBASE, MEDLINE, Emcare and CINAHL were searched in November 2021 and June 2022 (N > 50, English, human), and studies which examined the association between sex and HNcSCC were included. Analysis was conducted using RStudio with data and forest plots displaying males as a proportion of total patients with HNcSCC. Two independent researchers performed study selection, data extraction, data analysis and risk of bias. Eighty-two studies (1948 to 2018) comprising approximately 186,000 participants (67% male, 33% female) from 29 countries were included. Significantly more males had HNcSCC overall (71%; CI: 67−74). Males were also significantly more affected by cSCC of the ear (92%; CI: 89−94), lip (74%; CI: 66−81), and eyelid (56%; CI: 51−62). This study found HNcSCC disproportionately affected males overall and across all subtypes. Improving our understanding of sex-specific mechanisms in HNcSCC will better inform our preventive, therapeutic and prognostic practices.