Single nuclei RNA-seq reveals a medium spiny neuron glutamate excitotoxicity signature prior to the onset of neuronal death in an ovine Huntington's disease model.

Huntington's disease (HD) is a neurodegenerative genetic disorder caused by an expansion in the CAG repeat tract of the huntingtin (HTT) gene resulting in behavioural, cognitive, and motor defects. Current knowledge of disease pathogenesis remains incomplete, and no disease course-modifying interventions are in clinical use. We have previously reported the development and characterisation of the OVT73 transgenic sheep model of HD. The 73 polyglutamine repeat is somatically stable and therefore likely captures a prodromal phase of the disease with an absence of motor symptomatology even at 5-years of age and no detectable striatal cell loss. To better understand the disease-initiating events we have undertaken a single nuclei transcriptome study of the striatum of an extensively studied cohort of 5-year-old OVT73 HD sheep and age matched wild-type controls. We have identified transcriptional upregulation of genes encoding N-methyl-D-aspartate (NMDA), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and kainate receptors in medium spiny neurons, the cell type preferentially lost early in HD. Further, we observed an upregulation of astrocytic glutamate uptake transporters and medium spiny neuron GABAA receptors, which may maintain glutamate homeostasis. Taken together, these observations support the glutamate excitotoxicity hypothesis as an early neurodegeneration cascade-initiating process but the threshold of toxicity may be regulated by several protective mechanisms. Addressing this biochemical defect early may prevent neuronal loss and avoid the more complex secondary consequences precipitated by cell death.

PRSet: Pathway-based polygenic risk score analyses and software.

Polygenic risk scores (PRSs) have been among the leading advances in biomedicine in recent years. As a proxy of genetic liability, PRSs are utilised across multiple fields and applications. While numerous statistical and machine learning methods have been developed to optimise their predictive accuracy, these typically distil genetic liability to a single number based on aggregation of an individual's genome-wide risk alleles. This results in a key loss of information about an individual's genetic profile, which could be critical given the functional sub-structure of the genome and the heterogeneity of complex disease. In this manuscript, we introduce a 'pathway polygenic' paradigm of disease risk, in which multiple genetic liabilities underlie complex diseases, rather than a single genome-wide liability. We describe a method and accompanying software, PRSet, for computing and analysing pathway-based PRSs, in which polygenic scores are calculated across genomic pathways for each individual. We evaluate the potential of pathway PRSs in two distinct ways, creating two major sections: (1) In the first section, we benchmark PRSet as a pathway enrichment tool, evaluating its capacity to capture GWAS signal in pathways. We find that for target sample sizes of >10,000 individuals, pathway PRSs have similar power for evaluating pathway enrichment as leading methods MAGMA and LD score regression, with the distinct advantage of providing individual-level estimates of genetic liability for each pathway -opening up a range of pathway-based PRS applications, (2) In the second section, we evaluate the performance of pathway PRSs for disease stratification. We show that using a supervised disease stratification approach, pathway PRSs (computed by PRSet) outperform two standard genome-wide PRSs (computed by C+T and lassosum) for classifying disease subtypes in 20 of 21 scenarios tested. As the definition and functional annotation of pathways becomes increasingly refined, we expect pathway PRSs to offer key insights into the heterogeneity of complex disease and treatment response, to generate biologically tractable therapeutic targets from polygenic signal, and, ultimately, to provide a powerful path to precision medicine.

Effect of pyloroplasty on clinical outcomes following esophagectomy.

INTRODUCTION: The role of concurrent pyloroplasty with esophagectomy is unclear. Available literature on the impact of pyloroplasty during esophagectomy on complications and weight loss is varied. Data on the need for further pyloric intervention are scarce. Our study compares the clinical outcomes after esophagectomy with or without pyloroplasty and investigates the role of post-operative pyloric dilatation. METHODS: Consecutive patients (n = 207) undergoing Ivor Lewis esophagectomy performed by two surgeons at our institution were included. Data on patient demographics, mortality rate, anastomotic leak, respiratory complications (Clavien-Dindo grade ≥ 3), anastomotic stricture rate, and percentage weight loss at 1 and 2 year post-operatively were evaluated. For weight analysis at 1 and 2 year post-operatively, patients were excluded if they had been diagnosed with recurrence or died prior to the 1 or 2 year timepoints. RESULTS: Ninety-two patients did not have a pyloroplasty, and 115 patients had a pyloroplasty. There were no complications resulting from pyloroplasty. There was no significant demographic difference between the groups except for age. Mortality rate, anastomotic leak, respiratory complications, anastomotic stricture rate, and percentage weight loss at 1 and 2 years were statistically similar between the two groups. However, 14.1% of patients without pyloroplasty required post-operative endoscopic pyloric balloon dilatation to treat respiratory complications or gastroparesis. Subgroup analysis of patients without pyloroplasty indicated that patients requiring dilatation had greater weight loss at 1 year (15.8% vs 9.4%, p = 0.02) and higher respiratory complications rate (27.3% vs 4.7%, p = 0.038). CONCLUSIONS: Overall results from our study that pyloroplasty during Ivor Lewis esophagectomy is safe and useful to prevent the need for post-operative pyloric dilatation.

Risk factors for and treatment of anastomotic strictures after Ivor Lewis esophagectomy.

INTRODUCTION: Anastomotic strictures following esophagectomy occur frequently and impact on nutrition and quality of life. Although strictures are often attributed to ischemia and anastomotic leaks, the role of anastomosis size and pyloroplasty is not well evaluated. Our study aims to assess the rate of and risk factors for anastomotic stricture following esophagectomy, and the impact of treatment with regular endoscopic balloon dilatations. METHODS: Consecutive patients (n = 207) undergoing Ivor Lewis esophagectomy performed by two surgeons at our institution were included. Data on patient demographics, surgical outcomes and anastomotic strictures were recorded. Relationship of anastomotic strictures with circular stapler size, pyloroplasty and anastomotic leak was analyzed. Treatment of strictures with endoscopic balloon dilatation was reviewed and percentage weight loss at 1 year was evaluated. RESULTS: Anastomotic strictures occurred in 17.4% of patients. Patient demographics between those with and without stricture were similar. Stricture rate was similar in patients with or without pyloroplasty (13.9% vs 21.7%, respectively, p = 0.14) and in those with or without an anastomotic leak (25.0% vs 16.6%, respectively, p = 0.345). Stricture risk increased with smaller sized stapler (25 mm = 33.3%, 28 mm = 15.3%, 31 mm = 4.8%; p = 0.027). The median number of dilatations required to fully treat strictures was 2 (IQR: 1-3). The median length of time from surgery to first dilatation was 2.9 months (IQR: 2.0-4.7) and to last dilatation was 6.1 months (IQR: 4.8-10.0). Median maximum dilatation diameter was 20 mm (IQR: 18.0-20.0). There were no complications from dilatations. Percentage weight loss at 1 year in patients with strictures was similar to those without strictures (8.7% vs 11.1%, respectively, p = 0.090). CONCLUSIONS: Post-esophagectomy anastomotic strictures are common and not necessarily related to anastomotic leaks or absence of pyloroplasty. Smaller anastomosis size was strongly linked with stricture formation. A driven approach with regular endoscopic balloon dilation is safe and effective in treating these strictures with no excess weight loss at 1 year once treated.

How well do connectivity tools agree over the full life cycle? A case study of Irish Sea plaice Pleuronectes platessa Linnaeus, 1758.

Adult plaice in the Irish Sea have distinct traits that reflect the spawning locations that could suggest a number of different populations. However, do connectivity pathways support this concept? Different tools are directed at measuring exchange or connectivity between different life-history stages, and the challenge is to integrate the signals to obtain full life-cycle estimates. Collectively, the different methods reveal stable connectivity between known spawning and nursery grounds, with sufficient exchange to maintain a single population with weak genetic structure.

Combining SOS1 and MEK Inhibitors in a Murine Model of Plexiform Neurofibroma Results in Tumor Shrinkage.

Individuals with neurofibromatosis type 1 develop rat sarcoma virus (RAS)-mitogen-activated protein kinase-mitogen-activated and extracellular signal-regulated kinase (RAS-MAPK-MEK)-driven nerve tumors called neurofibromas. Although MEK inhibitors transiently reduce volumes of most plexiform neurofibromas in mouse models and in neurofibromatosis type 1 (NF1) patients, therapies that increase the efficacy of MEK inhibitors are needed. BI-3406 is a small molecule that prevents Son of Sevenless (SOS)1 interaction with Kirsten rat sarcoma viral oncoprotein (KRAS)-GDP, interfering with the RAS-MAPK cascade upstream of MEK. Single agent SOS1 inhibition had no significant effect in the DhhCre;Nf1 fl/fl mouse model of plexiform neurofibroma, but pharmacokinetics (PK)-driven combination of selumetinib with BI-3406 significantly improved tumor parameters. Tumor volumes and neurofibroma cell proliferation, reduced by MEK inhibition, were further reduced by the combination. Neurofibromas are rich in ionized calcium binding adaptor molecule 1 (Iba1)+ macrophages; combination treatment resulted in small and round macrophages, with altered cytokine expression indicative of altered activation. The significant effects of MEK inhibitor plus SOS1 inhibition in this preclinical study suggest potential clinical benefit of dual targeting of the RAS-MAPK pathway in neurofibromas. SIGNIFICANCE STATEMENT: Interfering with the RAS-mitogen-activated protein kinase (RAS-MAPK) cascade upstream of mitogen activated protein kinase kinase (MEK), together with MEK inhibition, augment effects of MEK inhibition on neurofibroma volume and tumor macrophages in a preclinical model system. This study emphasizes the critical role of the RAS-MAPK pathway in controlling tumor cell proliferation and the tumor microenvironment in benign neurofibromas.

Association of GDF15 levels with body mass index and endocrine status in ß-thalassaemia.

OBJECTIVE: GDF15 has emerged as a stress-induced hormone, acting on the brain to reduce food intake and body weight while affecting neuroendocrine function. Very high GDF15 levels are found in thalassaemia, where growth, energy balance and neuroendocrine function are impaired. We examined the relationships between GDF15 and anthropometric measures and endocrine status in ß-thalassaemia. DESIGN: Cross sectional study. PATIENTS: All ß-thalassaemia patients attending the thalassaemia unit of Colombo North Teaching Hospital for blood transfusions. MEASUREMENTS: Anthropometric data, appetite scores, circulating GDF15, IGF, thyroid and reproductive hormone levels in 103 ß-thalassaemia patients were obtained. RESULTS: GDF15 levels were markedly elevated in thalassaemia patients (24.2-fold with ß-thalassaemia major compared with healthy controls). Among patients with ß-thalassaemia major, the relationship between GDF15 and body mass index (BMI) was curvilinear with all individuals with GDF15 levels above 24,000 pg/mL having a BMI below 20 kg/m2 . After adjustment for BMI, age and Tanner stage, serum IGF1 concentrations correlated negatively with GDF15 in all thalassaemia patients (ß = -.027, p = .02). We found a significant positive relationship between GDF15 and gonadotropin (in both sexes) and testosterone (in males). CONCLUSIONS: GDF15 levels were markedly elevated in patients with ß-thalassaemia and its association with BMI is consistent with the known effect of GDF15 to reduce body weight. The inverse association between GDF15 with IGF1 levels may reflect a neuroendocrine impact of GDF15 or an indirect effect via impaired nutritional state. The positive association with testosterone in males and gonadotropins in both sexes, was surprising and should prompt further GDF15 studies on the hypothalamic pituitary gonadal axis.

Bridging a diagnostic Kawasaki disease classifier from a microarray platform to a qRT-PCR assay.

BACKGROUND: Kawasaki disease (KD) is a systemic vasculitis that mainly affects children under 5 years of age. Up to 30% of patients develop coronary artery abnormalities, which are reduced with early treatment. Timely diagnosis of KD is challenging but may become more straightforward with the recent discovery of a whole-blood host response classifier that discriminates KD patients from patients with other febrile conditions. Here, we bridged this microarray-based classifier to a clinically applicable quantitative reverse transcription-polymerase chain reaction (qRT-PCR) assay: the Kawasaki Disease Gene Expression Profiling (KiDs-GEP) classifier. METHODS: We designed and optimized a qRT-PCR assay and applied it to a subset of samples previously used for the classifier discovery to reweight the original classifier. RESULTS: The performance of the KiDs-GEP classifier was comparable to the original classifier with a cross-validated area under the ROC curve of 0.964 [95% CI: 0.924-1.00] vs 0.992 [95% CI: 0.978-1.00], respectively. Both classifiers demonstrated similar trends over various disease conditions, with the clearest distinction between individuals diagnosed with KD vs viral infections. CONCLUSION: We successfully bridged the microarray-based classifier into the KiDs-GEP classifier, a more rapid and more cost-efficient qRT-PCR assay, bringing a diagnostic test for KD closer to the hospital clinical laboratory. IMPACT: A diagnostic test is needed for Kawasaki disease and is currently not available. We describe the development of a One-Step multiplex qRT-PCR assay and the subsequent modification (i.e., bridging) of the microarray-based host response classifier previously described by Wright et al. The bridged KiDs-GEP classifier performs well in discriminating Kawasaki disease patients from febrile controls. This host response clinical test for Kawasaki disease can be adapted to the hospital clinical laboratory.

Associations between breast milk intake volume, macronutrient intake and infant growth in a longitudinal birth cohort: the Cambridge Baby Growth and Breastfeeding Study (CBGS-BF).

Growth patterns of breastfed infants show substantial inter-individual differences, partly influenced by breast milk (BM) nutritional composition. However, BM nutritional composition does not accurately indicate BM nutrient intakes. This study aimed to examine the associations between both BM intake volumes and macronutrient intakes with infant growth. Mother-infant dyads (n 94) were recruited into the Cambridge Baby Growth and Breastfeeding Study (CBGS-BF) from a single maternity hospital at birth; all infants received exclusive breast-feeding (EBF) for at least 6 weeks. Infant weight, length and skinfolds thicknesses (adiposity) were repeatedly measured from birth to 12 months. Post-feed BM samples were collected at 6 weeks to measure TAG (fat), lactose (carbohydrate) (both by 1H-NMR) and protein concentrations (Dumas method). BM intake volume was estimated from seventy infants between 4 and 6 weeks using dose-to-the-mother deuterium oxide (2H2O) turnover. In the full cohort and among sixty infants who received EBF for 3+ months, higher BM intake at 6 weeks was associated with initial faster growth between 0 and 6 weeks (ß + se 3·58 + 0·47 for weight and 4·53 + 0·6 for adiposity gains, both P < 0·0001) but subsequent slower growth between 3 and 12 months (ß + se - 2·27 + 0·7 for weight and -2·65 + 0·69 for adiposity gains, both P < 0·005). BM carbohydrate and protein intakes at 4-6 weeks were positively associated with early (0-6 weeks) but tended to be negatively related with later (3-12 months) adiposity gains, while BM fat intake showed no association, suggesting that carbohydrate and protein intakes may have more functional relevance to later infant growth and adiposity.

Infrared thermography as a tool for the measurement of negative emotions in dairy cows.

In commercial dairy cows, the conditions in which they are kept may lead to negative emotional states associated with the development of chronic physiological and behavioural abnormalities that may compromise their health, welfare and productivity. Such states include fear, stress or anxiety. Behavioural rather than physiological tests are more likely to be used to indicate these states but can be limited by their subjectivity, need for specialised infrastructure and training (of the operator and sometimes the animal) and the time-consuming nature of data collection. Popularly used physiological measures such as blood cortisol may be more appropriate for acute rather than chronic assessments but are easily confounded, for example by a response to the act of measurement per se. More sophisticated physiological measures such as functional magnetic resonance imaging (fMRI) and electroencephalography (EEG) may be impractical due to cost and time and, like blood cortisol, have the confounding associated with the act of measurement. By contrast, infrared thermography of external body surfaces is remote, non-invasive, easily repeated and follows an objective methodology, allowing longitudinal data acquisition for the inference of changes in chronic emotional state over time. The objective of this review was to investigate the potential of infrared thermography to measure cow emotions. In lactating dairy cows, maximum IRT of the eyes and coronary band of the limbs seem to be most representative of thermoregulatory changes, which are repeatable and correlate with behavioural and physiological indicators of emotional state. IRT methodologies have the potential to become a fundamental tool for the objective assessment of welfare state in dairy cows.

Controlled release of MT-1207 using a novel gastroretentive bilayer system comprised of hydrophilic and hydrophobic polymers.

In the present study, novel gastroretentive bilayer tablets were developed that are promising for the once-a-day oral delivery of the drug candidate MT-1207. The gastroretentive layer consisted of a combination of hydrophilic and hydrophobic polymers, namely polyethylene oxide and Kollidon® SR. A factorial experiment was conducted, and the results revealed a non-effervescent gastroretentive layer that, unlike most gastroretentive layers reported in the literature, was easy to prepare, and provided immediate tablet buoyancy (mean floating lag time of 1.5 s) that lasted over 24 h in fasted state simulated gastric fluid (FaSSGF) pH 1.6, irrespective of the drug layer, thereby allowing a 24-hour sustained release of MT-1207 from the drug layer of the tablets. Furthermore, during in vitro buoyancy testing of the optimised bilayer tablets in media of different pH values (1.0, 3.0, 6.0), the significant difference (one-way ANOVA, p < 0.001) between the respective total floating times indicated that stomach pH effects on tablet buoyancy are important to be considered during the development of non-effervescent gastroretentive formulations and the choice of dosing regimen. To the best of our knowledge, this has not been reported before, and it should probably be factored in when designing dosing regimens. Finally, a pharmacokinetic study in Beagle dogs indicated a successful in vivo 24-hour sustained release of MT-1207 from the optimised gastroretentive bilayer tablet formulations with the drug plasma concentration remaining above the estimated minimum effective concentration of 1 ng/mL at the 24-hour timepoint and also demonstrated the gastroretentive capabilities of the hydrophilic and hydrophobic polymer combination. The optimised formulations will be forwarded to clinical development.

Impact of aerial humidity on seasonal malaria: an ecological study in Zambia.

BACKGROUND: Seasonal patterns of malaria cases in many parts of Africa are generally associated with rainfall, yet in the dry seasons, malaria transmission declines but does not always cease. It is important to understand what conditions support these periodic cases. Aerial moisture is thought to be important for mosquito survival and ability to forage, but its role during the dry seasons has not been well studied. During the dry season aerial moisture is minimal, but intermittent periods may arise from the transpiration of peri-domestic trees or from some other sources in the environment. These periods may provide conditions to sustain pockets of mosquitoes that become active and forage, thereby transmitting malaria. In this work, humidity along with other ecological variables that may impact malaria transmission have been examined. METHODS: Negative binomial regression models were used to explore the association between peri-domestic tree humidity and local malaria incidence. This was done using sensitive temperature and humidity loggers in the rural Southern Province of Zambia over three consecutive years. Additional variables including rainfall, temperature and elevation were also explored. RESULTS: A negative binomial model with no lag was found to best fit the malaria cases for the full year in the evaluated sites of the Southern Province of Zambia. Local tree and granary night-time humidity and temperature were found to be associated with local health centre-reported incidence of malaria, while rainfall and elevation did not significantly contribute to this model. A no lag and one week lag model for the dry season alone also showed a significant effect of humidity, but not temperature, elevation, or rainfall. CONCLUSION: The study has shown that throughout the dry season, periodic conditions of sustained humidity occur that may permit foraging by resting mosquitoes, and these periods are associated with increased incidence of malaria cases. These results shed a light on conditions that impact the survival of the common malaria vector species, Anopheles arabiensis, in arid seasons and suggests how they emerge to forage when conditions permit.

Expression of selected genes encoding mechanistic pathways, nutrient and amino acid transporters in jejunum and ileum of broiler chickens fed a reduced protein diet supplemented with arginine, glutamine and glycine under stress stimulated by dexamethasone.

Reducing crude protein and supplementation with synthetic amino acids in poultry nutrition is a recent trend to avoid wastage of protein and ammonia in production systems. Stress has been shown to impair intestinal barrier and increase inflammatory response. This study was performed on intestinal tissues of broiler chickens to understand the mechanism of stress induced by a synthetic glucocorticoid, dexamethasone (DEX) and the effect of supplementation of arginine, glutamine and glycine in reduced protein diets. Intestinal tissue samples from a previous study were utilized. Male Ross 308 chickens received a basal diet for the first seven days and then fed with crude protein that was reduced to 194 g/kg in grower experimental diets supplemented with glutamine, glycine and additional arginine at 10, 10 and 5 g/kg respectively. Half of the 96 individual birds were injected with DEX (0.5 mg/kg body weight) or saline on days 14, 16, 18 and 20 of age. mRNA expression for jejunum and ileum for amino acid transporters (y+LAT-1, Bo,+ AT, EAAT-3 and CAT-1), mechanistic genes (SGLT-1, mTOR, IAP and FABP-2) and pro-inflammatory genes (MUC-2, NF-κB, iNOS, IL-8 and IL-1ß) were analysed using real-time PCR. The results showed that DEX decreased y+ LAT1 in jejunum, Bo ,+ AT and EAAT-3 in ileum. Arginine increased CAT-1 in the jejunum and ileum under DEX treatment. Through an interaction, DEX reduced IAP in jejunum of glycine and arginine supplemented group and reduced mTOR in jejunum independently. DEX reduced MUC-2 and iNOS in jejunum and increased iNOS and IL8 in the ileum. Amino acid supplementation did not appear to ameliorate these effects; however, there were some positive effects of glycine on NF-κB and arginine through increased CAT-1. Mechanistic understanding of amino acid supplementation in broiler diets warrants further research particularly when dietary protein is reduced below the level tested in the present study.

In vitro and in vivo evaluation of a sustained-release once-a-day formulation of the novel antihypertensive drug MT-1207.

Hypertension is one of the most common chronic cardiovascular disorders. Sustained-release formulations are developed to maintain drug therapeutic levels throughout the treatment of hypertension, to promote patient compliance and improve patient outcomes. We have developed and tested in in vivo trials a once-a-day tablet formulation for the novel antihypertensive drug MT-1207. The tablets based upon a hydrophilic polymer matrix underwent post-compression parameter and physicochemical characterisations, along with in vitro drug release testing. The most promising formulation containing 31% w/w HPMC K15M gave a 24-hour release of MT-1207 with an almost constant release rate up to 20 hours. Follow in in vivo studies were carried out in Beagle dogs for the optimised sustained-release tablets in comparison to immediate-release tablets. The results showed that a sustained release of MT-1207 from the new formulation was achieved with a drug t1/2 2-2.5 times longer than the immediate-release tablets. Moreover, the AUC0-24h values of both sustained- and immediate-release tablets were identical at the same dose of 30 mg, indicating that the same amount of drug was absorbed in each case. For treatments based upon MT-1207, this development is significant for future commercial exploitation via scale-up and further trials, and for improved patient outcomes.

Methylation of the C19MC microRNA locus in the placenta: association with maternal and chilhood body size.

OBJECTIVES: To study DNA methylation at the C19MC locus in the placenta and its association with (1) parental body size, (2) transmission of haplotypes for the C19MC rs55765443 SNP, and (3) offspring's body size and/or body composition at birth and in childhood. SUBJECTS AND METHODS: Seventy-two pregnant women-infant pairs and 63 fathers were included in the study. Weight and height of mothers, fathers and newborns were registered during pregnancy or at birth (n = 72). Placental DNA methylation at the C19MC imprinting control region (ICR) was quantified by bisulfite pyrosequencing. Genotyping of the SNP was performed using restriction fragment length polymorphisms. The children's body size and composition were reassessed at age 6 years (n = 32). RESULTS: Lower levels of placental C19MC methylation were associated with increased body size of mother, specifically with higher pregestational and predelivery weights and height of the mother (ß from -0.294 to -0.371; R2 from 0.04 to 0.10 and all p < 0.019), and with higher weight, height, waist and hip circumferences, and fat mass of the child (ß from -0.428 to -0.552; R2 from 0.33 to 0.56 and all p < 0.009). Parental transmission of the SNP did not correlate with an altered placental methylation status at the C19MC ICR. CONCLUSIONS: Increased maternal size is associated with reduced placental C19MC methylation, which, in turn, relate to larger body size of the child.

Brain urea increase is an early Huntington's disease pathogenic event observed in a prodromal transgenic sheep model and HD cases.

The neurodegenerative disorder Huntington's disease (HD) is typically characterized by extensive loss of striatal neurons and the midlife onset of debilitating and progressive chorea, dementia, and psychological disturbance. HD is caused by a CAG repeat expansion in the Huntingtin (HTT) gene, translating to an elongated glutamine tract in the huntingtin protein. The pathogenic mechanism resulting in cell dysfunction and death beyond the causative mutation is not well defined. To further delineate the early molecular events in HD, we performed RNA-sequencing (RNA-seq) on striatal tissue from a cohort of 5-y-old OVT73-line sheep expressing a human CAG-expansion HTT cDNA transgene. Our HD OVT73 sheep are a prodromal model and exhibit minimal pathology and no detectable neuronal loss. We identified significantly increased levels of the urea transporter SLC14A1 in the OVT73 striatum, along with other important osmotic regulators. Further investigation revealed elevated levels of the metabolite urea in the OVT73 striatum and cerebellum, consistent with our recently published observation of increased urea in postmortem human brain from HD cases. Extending that finding, we demonstrate that postmortem human brain urea levels are elevated in a larger cohort of HD cases, including those with low-level neuropathology (Vonsattel grade 0/1). This elevation indicates increased protein catabolism, possibly as an alternate energy source given the generalized metabolic defect in HD. Increased urea and ammonia levels due to dysregulation of the urea cycle are known to cause neurologic impairment. Taken together, our findings indicate that aberrant urea metabolism could be the primary biochemical disruption initiating neuropathogenesis in HD.

A sampling strategy for the determination of infrared temperature of relevant external body surfaces of dairy cows.

Infrared thermography is a tool to investigate the welfare of cattle. This study aimed to identify a sampling strategy for recording infrared thermograms in dairy cows, in order to most efficiently determine biologically relevant changes in the maximum infrared temperature (IRT) of the eyes and coronary band of forelimbs. Thirty-one dairy cows were used for the study. They were assessed with four replicates of thermograms for each of the head and lower forelimb per cow for 6 mostly consecutive days (sessions). The data obtained were subjected to random effects Analysis of Variance which was used to estimate the variance components for this sampling model, using maximum IRT of both eyes; (left + right eye)/2 and both limbs; (left + right coronary band of forelimb)/2 as dependant variables. The variance components were used to calculate least significant differences (LSD) between two theoretical treatment groups under different sampling scenarios. Analysis showed that there was minimal improvement in precision beyond 2 thermograms within a session but there was improvement with increasing the number of sessions from 2 to 3. The LSD of both eyes and both limbs reached a biologically relevant difference (0.4 and 0.9 °C, respectively) at a minimum number of 14 - 16 cows monitored for 2 consecutive thermography sessions, or 10 - 12 cows for 3 sessions. We conclude that no more than 2 replicate IRT measures are required per session but that measuring on 3 consecutive days should be considered, depending on whether time or number of cows used is the primary limitation.

Assessing response to dry-off in dairy cows kept outdoors using spontaneous behaviours and infrared thermography-a pilot study.

We used spontaneous behaviours to assess response to dry-off involving abrupt dietary and milking frequency changes, followed by regrouping, after the last milking in 15 clinically healthy Holstein-Frisian cows kept outdoors. Moreover, we explored the potential of infrared thermography to detect eye temperature variations possibly induced by dry-off. On days - 1, 0, 1 and 2 relative to dry-off, we recorded whether cows vocalised during feed delivery; ate fresh feed within 5 min; and mean maximum eye temperature at approximately 1 h after feed delivery. On days 1 and 2, cows were more likely to eat fresh feed compared to days - 1 and 0. No difference in likelihood of vocalising was found. Compared to day - 1, eye temperature was substantially higher on days 0 and 2. Collectively, the results suggest that cows responded, both behaviourally and physiologically, to the abrupt dry-off management. The interpretation of the current findings deserves further investigation using larger sample sizes, more controlled environments and further behavioural, physiological, cognitive and clinical measures.

Comparison of the Moral Sensitivity, Judgment, and Actions of Australian and Turkish Veterinary Students in Relation to Animal Ethics Issues.

Veterinarians regularly face animal ethics conflicts, and research has identified the moral reasoning methods that they utilize to solve these. It is unclear whether students' sensitivity to animal ethics conflicts influences their reasoning methods, and the recent development of appropriate tests allows this to be assessed. We compared the moral reasoning methods, intended action and sensitivity of 112 first-year veterinary students in two contrasting veterinary schools, in Australia and Turkey. Students were presented with two animal ethics issues: breeding blind hens to address welfare concerns in intensive housing, for moral reasoning evaluation; and a video of a lame dairy cow walking, for sensitivity assessment. The sensitivity score was not related to the principal moral reasoning methods, which are Personal Interest (PI), Maintaining Norms (MN), and Universal Principles (UP). However, less sensitive students were more concerned about professional criticism of emotional reactions when addressing the hen scenario. Turkish students, mostly males, used more MN reasoning when deciding the hen dilemma. Australian, mostly female, students did not. Overall, female students were more likely to consider the universal moral principles in moral reasoning than male students and were more likely to recommend against breeding blind hens. This suggests that females are more likely to consider the ethical implications of their actions than males. This study demonstrates relationships between ethical sensitivity (ES) and moral reasoning, and cultural and gender effects on moral action choices. Students placing greater importance on professional criticism about having an emotional reaction are more likely to be those who have less ES.

Reduced size at birth and persisting reductions in adiposity in recent, compared with earlier, cohorts of infants born to mothers with gestational diabetes mellitus.

AIMS/HYPOTHESIS: This study aimed to explore the infancy growth trajectories of 'recent' and 'earlier' offspring of mothers with gestational diabetes mellitus (OGDM), each compared with the same control infants, and investigate whether 'recent' OGDM still exhibit a classical phenotype, with macrosomia and increased adiposity. METHODS: Within a prospective observational birth cohort, 98 'earlier' OGDM born between 2001 and 2009 were identified using 75 g oral glucose tolerance testing at 28 weeks gestation, 122 recent OGDM born between 2011 and 2013 were recruited postnatally through antenatal diabetes clinics, and 876 normal birthweight infants of mothers with no history of diabetes were recruited across the full study period as the control group. All infants followed the same study protocol (measurements at birth, 3, 12 and 24 months, including weight, length and skinfold thickness indicating adiposity, and detailed demographic data). In all cases, GDM was defined using the International Association of Diabetes and Pregnancy Study Group criteria. RESULTS: Earlier OGDM had higher birthweight SD scores (SDS) than control infants. Conversely, recent OGDM had similar birthweight- and length SDS to control infants (mean ± SD, 0.1 ± 1.0 and- 0.1 ± 0.9, respectively), but lower mean skinfold thickness SDS (-0.4 ± 0.6 vs 0.0 ± 0.9; p < 0.001). After birth, earlier OGDM showed reduced gains in weight and length between 3 and 12 months. In contrast, recent OGDM had increased weight and skinfold thickness gains until 3 months, followed by reduced gains in those variables from 3 to 12 months, compared with control infants. At 24 months, recent OGDM had lower adiposity than control infants (mean skinfold thickness SDS -0.3 ± 0.7 vs 0.0 ± 0.8; p < 0.001). At all time points recent OGDM had lower growth measurements than earlier OGDM. CONCLUSIONS/INTERPRETATION: Recent OGDM showed different growth trajectories to the earlier group, namely normalisation of birthweight and reduced adiposity at birth, followed by initial rapid weight gain but subsequent reduced adiposity postnatally. While avoidance of macrosomia at birth may be advantageous, the longer-term health implications of these changing growth trajectories are uncertain.