A multi-disciplinary commentary on preclinical research to investigate vascular contributions to dementia.

Although dementia research has been dominated by Alzheimer's disease (AD), most dementia in older people is now recognised to be due to mixed pathologies, usually combining vascular and AD brain pathology. Vascular cognitive impairment (VCI), which encompasses vascular dementia (VaD) is the second most common type of dementia. Models of VCI have been delayed by limited understanding of the underlying aetiology and pathogenesis. This review by a multidisciplinary, diverse (in terms of sex, geography and career stage), cross-institute team provides a perspective on limitations to current VCI models and recommendations for improving translation and reproducibility. We discuss reproducibility, clinical features of VCI and corresponding assessments in models, human pathology, bioinformatics approaches, and data sharing. We offer recommendations for future research, particularly focusing on small vessel disease as a main underpinning disorder.

Antibiotic resistance gene sequencing is necessary to reveal the complex dynamics of immigration from sewers to activated sludge.

Microbial community composition has increasingly emerged as a key determinant of antibiotic resistance gene (ARG) content. However, in activated sludge wastewater treatment plants (AS-WWTPs), a comprehensive understanding of the microbial community assembly process and its impact on the persistence of antimicrobial resistance (AMR) remains elusive. An important part of this process is the immigration dynamics (or community coalescence) between the influent and activated sludge. While the influent wastewater contains a plethora of ARGs, the persistence of a given ARG depends initially on the immigration success of the carrying population, and the possible horizontal transfer to indigenously resident populations of the WWTP. The current study utilized controlled manipulative experiments that decoupled the influent wastewater composition from the influent microbial populations to reveal the fundamental mechanisms involved in ARG immigration between sewers and AS-WWTP. A novel multiplexed amplicon sequencing approach was used to track different ARG sequence variants across the immigration interface, and droplet digital PCR was used to quantify the impact of immigration on the abundance of the targeted ARGs. Immigration caused an increase in the abundance of over 70 % of the quantified ARGs. However, monitoring of ARG amplicon sequence variants (ARG-ASVs) at the immigration interface revealed various immigration patterns such as (i) suppression of the indigenous mixed liquor ARG-ASV by the immigrant, or conversely (ii) complete immigration failure of the influent ARG-ASV. These immigration profiles are reported for the first time here and highlight the crucial information that can be gained using our novel multiplex amplicon sequencing techniques. Future studies aiming to reduce AMR in WWTPs should consider the impact of influent immigration in process optimisation and design.

Cannabis use is associated with low plasma endocannabinoid Anandamide in individuals with psychosis.

BACKGROUND: Cannabis use suppresses the endocannabinoid system in healthy individuals. However, the association between cannabis use with the endocannabinoid system is understudied in individuals with psychosis despite the high rate of cannabis use in these individuals. METHODS: We enrolled 83 individuals who were admitted to an inpatient psychiatric unit with psychotic presentations, and measured their plasma levels of main endocannabinoids, Anandamide (AEA) and 2-Acylglycerol (2-AG), and endocannabinoid related compounds, Palmitoylethanolamine, and N-oleoylethanolamine. Cannabis use was assessed with urine toxicology and frequency of cannabis use was assessed using self-reported questionnaires. The Positive and Negative Syndrome Scale was used to assess the severity of psychotic symptoms. RESULTS: Overall, we had 38 individuals in cannabis positive group (CN+) and 45 individuals in cannabis negative group (CN-). Compared to CN-, CN+ group had lower plasma levels of AEA, which remained significant after controlling for age, gender, race/ethnicity, and use of other drugs. CONCLUSION: Cannabis use is associated with low plasma AEA levels in individuals with psychosis, which is in the same line with reported suppressive effects of cannabis on the endocannabinoid system in healthy individuals. Further studies are needed to investigate the clinical significance of this finding.

Liraglutide protects ß-cells in novel human islet spheroid models of type 1 diabetes.

To enable accurate, high-throughput and longer-term studies of the immunopathogenesis of type 1 diabetes (T1D), we established three in-vitro islet-immune injury models by culturing spheroids derived from primary human islets with proinflammatory cytokines, activated peripheral blood mononuclear cells or HLA-A2-restricted preproinsulin-specific cytotoxic T lymphocytes. In all models, ß-cell function declined as manifested by increased basal and decreased glucose-stimulated insulin release (GSIS), and decreased intracellular insulin content. Additional hallmarks of T1D progression such as loss of the first-phase insulin response (FFIR), increased proinsulin-to-insulin ratios, HLA-class I expression, and inflammatory cytokine release were also observed. Using these models, we show that liraglutide, a glucagon-like peptide 1 receptor agonist, prevented loss of GSIS under T1D-relevant stress, by preserving the FFIR and decreasing immune cell infiltration and cytokine secretion. Our results corroborate that liraglutide mediates an anti-inflammatory effect that aids in protecting ß-cells from the immune-mediated attack that leads to T1D.

A review of experimental models of focal cerebral ischemia focusing on the middle cerebral artery occlusion model.

Cerebral ischemic stroke is a leading cause of death and disability, but current pharmacological therapies are limited in their utility and effectiveness. In vitro and in vivo models of ischemic stroke have been developed which allow us to further elucidate the pathophysiological mechanisms of injury and investigate potential drug targets. In vitro models permit mechanistic investigation of the biochemical and molecular mechanisms of injury but are reductionist and do not mimic the complexity of clinical stroke. In vivo models of ischemic stroke directly replicate the reduction in blood flow and the resulting impact on nervous tissue. The most frequently used in vivo model of ischemic stroke is the intraluminal suture middle cerebral artery occlusion (iMCAO) model, which has been fundamental in revealing various aspects of stroke pathology. However, the iMCAO model produces lesion volumes with large standard deviations even though rigid surgical and data collection protocols are followed. There is a need to refine the MCAO model to reduce variability in the standard outcome measure of lesion volume. The typical approach to produce vessel occlusion is to induce an obstruction at the origin of the middle cerebral artery and reperfusion is reliant on the Circle of Willis (CoW). However, in rodents the CoW is anatomically highly variable which could account for variations in lesion volume. Thus, we developed a refined approach whereby reliance on the CoW for reperfusion was removed. This approach improved reperfusion to the ischemic hemisphere, reduced variability in lesion volume by 30%, and reduced group sizes required to determine an effective treatment response by almost 40%. This refinement involves a methodological adaptation of the original surgical approach which we have shared with the scientific community via publication of a visualised methods article and providing hands-on training to other experimental stroke researchers.

On-Site Prescription Dispensing Improves Antidepressant Adherence among Uninsured Depressed Patients.

The successful treatment of depressive disorders critically depends on adherence to prescribed treatment regimens. Despite increasing rates of antidepressant medication prescription, adherence to the full treatment course remains poor. Rates of antidepressant non-adherence are higher for uninsured patients and members of some marginalized racial and ethnic communities due to factors such as inequities in healthcare and access to insurance. Among patients treated in a free, student-run and faculty-supervised clinic serving uninsured patients in a majority Hispanic community in East Harlem, adherence rates are lower than those observed in patients with private or public New York State health insurance coverage. A prior study of adherence in these patients revealed that difficulty in obtaining medications from an off-site hospital pharmacy was a leading factor that patients cited for non-adherence. To alleviate this barrier to obtaining prescriptions, we tested the effectiveness of on-site, in-clinic medication dispensing for improving antidepressant medication adherence rates among uninsured patients. We found that dispensing medications directly to patients in clinic was associated with increased visits at which patients self-reported proper adherence and increased overall adherence rates. Furthermore, we found evidence that higher rates of antidepressant medication adherence were associated with more favorable treatment outcomes. All patients interviewed reported increased satisfaction with on-site dispensing. Overall, this study provides promising evidence that on-site antidepressant dispensing in a resource-limited setting improves medication adherence rates and leads to more favorable treatment outcomes with enhanced patient satisfaction.

Biomarkers of endothelial dysfunction in cocaine overdose and overdose-related cardiovascular events.

Overdose of stimulant drugs has been associated with increased risk of adverse cardiovascular events (ACVE), some of which may be ascribed to endothelial dysfunction. The aims of this study were to evaluate biomarkers of endothelial dysfunction in emergency department (ED) patients with acute cocaine overdose and to assess the association between in-hospital ACVE in ED patients with any acute drug overdose. This was a prospective consecutive cohort study over 9 months (2015-2016) at two urban, tertiary-care hospital EDs. Consecutive adults (≥18 years) presenting with suspected acute drug overdose were eligible and separated into three groups: cocaine (n = 47), other drugs (n = 128), and controls (n = 11). Data were obtained from medical records and linked to waste serum specimens, sent as part of routine clinical care, for biomarker analysis. Serum specimens were collected and analyzed using enzyme-linked immunosorbent assay kit for three biomarkers of endothelial dysfunction: (a) endothelin-1 (ET-1), (b) regulated upon activation normal T cell expressed and secreted (RANTES), and (c) soluble intercellular adhesion molecule-1 (siCAM-1). Mean siCAM was elevated for cocaine compared with controls and other drugs (p < .01); however, mean RANTES and ET-1 levels were not significantly different for any drug exposure groups. Receiver operating characteristics curve analysis for prediction of in-hospital ACVE revealed excellent performance of siCAM-1 (area under curve, 0.86; p < .001) but lack of predictive utility for either RANTES or ET-1. These results suggest that serum siCAM-1 is a viable biomarker for acute cocaine overdose and that endothelial dysfunction may be an important surrogate for adverse cardiovascular events following any drug overdose.

Cannabinoid use in psychotic patients impacts inflammatory levels and their association with psychosis severity.

Inflammatory abnormalities are well-documented in individuals with chronic psychotic disorders. Particular attention has focused on interleukin-6 (IL-6) and its correlation with psychotic symptom severity. Cannabis use is associated with an increased risk of psychosis and also has immunomodulating properties. It has been hypothesized that inflammatory disturbances are a common underlying pathology between cannabis use and psychosis. We measured inflammatory markers in individuals admitted to a psychiatric unit with acute psychosis who had toxicology positive for natural and/or synthetic cannabinoids (n = 59) compared to patients with negative cannabinoid toxicology (n = 60). Psychosis severity was assessed using the Positive and Negative Syndrome Scale (PANSS). While PANSS scores were similar between groups, cannabinoid-positive participants were more likely to receive pro re nata (PRN or as-needed) medications for agitation in the psychiatric emergency room, particularly synthetic cannabinoid-positive participants. In unadjusted models, cannabinoid-positive participants had lower interferon-γ (IFN-γ) levels (p = 0.046), but this finding was not significant after adjusting for covariates and multiple comparisons. Among cannabinoid-positive participants, IL-6 levels negatively correlated with PANSS total score (p = 0.040), as well as positive (p = 0.035) and negative (p = 0.024) subscales. Results suggest inflammatory alterations among psychotic individuals with comorbid cannabinoid use.

Longitudinal Multimodal fMRI to Investigate Neurovascular Changes in Spontaneously Hypertensive Rats.

Hypertension is an important risk factor for age-related cognitive decline and neuronal pathologies. Studies have shown a correlation between hypertension, disruption in neurovascular coupling and cerebral autoregulation, and cognitive decline. However, the mechanisms behind this are unclear. To further understand this, it is advantageous to study neurovascular coupling as hypertension progresses in a rodent model. Here, we use a longitudinal functional MRI (fMRI) protocol to assess the impact of hypertension on neurovascular coupling in spontaneously hypertensive rats (SHRs). Eight female SHRs were studied at 2, 4, and 6 months of age, as hypertension progressed. Under an IV infusion of propofol, animals underwent fMRI, functional MR spectroscopy, and cerebral blood flow (CBF) quantification to study changes in neurovascular coupling over time. Blood pressure significantly increased at 4 and 6 months (P < .0001). CBF significantly increased at 4 months old (P < .05), in the acute stage of hypertension. The size of the active region decreased significantly at 6 months old (P < .05). Change in glutamate signal during activation, and N-acetyl-aspartate (NAA) signal, remained constant. This study shows that, while cerebral autoregulation is impaired in acute hypertension, the blood oxygenation-level-dependent (BOLD) response remains unaltered until later stages. At this stage, the consistent NAA and glutamate signals show that neuronal death has not occurred, and that neuronal activity is not affected at this stage. This suggests that neuronal activity and viability is not lost until much later, and changes observed here in BOLD activity are due to vascular effects.

Improved reperfusion following alternative surgical approach for experimental stroke in mice.

Background: Following ischemic stroke, recanalisation and restoration of blood flow to the affected area of the brain is critical and directly correlates with patient recovery.  In vivo models of ischemic stroke show high variability in outcomes, which may be due to variability in reperfusion.  We previously reported that a surgical refinement in the middle cerebral artery occlusion (MCAO) model of stroke, via repair of the common carotid artery (CCA), removes the reliance on the Circle of Willis for reperfusion and reduced infarct variability.  Here we further assess this refined surgical approach on reperfusion characteristics following transient MCAO in mice. Methods: Mice underwent 60 min of MCAO, followed by either CCA repair or ligation at reperfusion.  All mice underwent laser speckle contrast imaging at baseline, 24 h and 48 h post-MCAO. Results: CCA ligation reduced cerebral perfusion in the ipsilateral hemisphere compared to baseline (102.3 ± 4.57%) at 24 h (85.13 ± 16.09%; P < 0.01) and 48 h (75.04 ± 12.954%; P < 0.001) post-MCAO. Repair of the CCA returned perfusion to baseline (94.152 ± 2.44%) levels and perfusion was significantly improved compared to CCA ligation at both 24 h (102.83 ± 8.41%; P < 0.05) and 48 h (102.13 ± 9.34%; P < 0.001) post-MCAO. Conclusions: Our findings show CCA repair, an alternative surgical approach for MCAO, results in improved ischemic hemisphere perfusion during the acute phase.

A longitudinal, multi-parametric functional MRI study to determine age-related changes in the rodent brain.

As the population ages, the incidence of age-related neurological diseases and cognitive decline increases. To further understand disease-related changes in brain function it is advantageous to examine brain activity changes in healthy aging rodent models to permit mechanistic investigation. Here, we examine the suitability, in rodents, of using a novel, minimally invasive anaesthesia protocol in combination with a functional MRI protocol to assess alterations in neuronal activity due to physiological aging. 11 Wistar Han female rats were studied at 7, 9, 12, 15 and 18 months of age. Under an intravenous infusion of propofol, animals underwent functional magnetic resonance imaging (fMRI) and functional magnetic resonance spectroscopy (fMRS) with forepaw stimulation to quantify neurotransmitter activity, and resting cerebral blood flow (CBF) quantification using arterial spin labelling (ASL) to study changes in neurovascular coupling over time. Animals showed a significant decrease in size of the active region with age (P â€‹< â€‹0.05). fMRS results showed a significant decrease in glutamate change with stimulation (ΔGlu) with age (P â€‹< â€‹0.05), and ΔGlu became negative from 12 months onwards. Global CBF remained constant for the duration of the study. This study shows age related changes in the blood oxygen level dependent (BOLD) response in rodents that correlate with those seen in humans. The results also suggest that a reduction in synaptic glutamate turnover with age may underlie the reduction in the BOLD response, while CBF is preserved.

Sexual dimorphism following in vitro ischemia in the response to neurosteroids and mechanisms of injury.

BACKGROUND: Cerebral ischemic stroke is a significant cause of morbidity and mortality. Sex differences exist following stroke in terms of incidence, symptoms, outcomes and response to some treatments. Importantly, molecular mechanisms of injury, activated following ischemia may differ between the sexes and if so may account, at least in part, for sex differences seen in treatment response. Here we aimed to determine, using single-sex organotypic hippocampal slice cultures, whether the effectiveness of a potential treatment option, i.e. sex steroids, exhibited any sexual dimorphism and whether sex affected the mechanisms of apoptosis activated following ischemia. RESULTS: Following exposure to ischemia, male-derived tissue exhibited higher levels of cell death than female-derived tissue. Various sex steroid hormones, i.e. progesterone, allopregnanolone, and estradiol, were protective in terms of reducing the amount of cell death in male- and female-derived tissue whereas medoxyprogesterone acetate (MPA) was only protective in female-derived tissue. The protective effect of progesterone was abolished in the presence of finasteride, a 5α-reductase inhibitor, suggesting it was largely mediated via its conversion to allopregnanolone. To test the hypothesis that sex differences exist in the activation of specific elements of the apoptotic pathway activated following ischemia we administered Q-VD-OPH, a caspase inhibitor, or PJ34, an inhibitor of poly (ADP ribose) polymerase (PARP). Caspase inhibition was only effective, in terms of reducing cell death, in female-derived tissue, whereas PARP inhibition was only protective in male-derived tissue. However, in both sexes, the protective effects of progesterone and estradiol were not observed in the presence of either caspase or PARP inhibition. CONCLUSIONS: Sex differences exist in both the amount of cell death produced and those elements of the cell death pathway activated following an ischemic insult. There are also some sex differences in the effectiveness of steroid hormones to provide neuroprotection following an ischemic insult-namely MPA was only protective in female-derived tissue. This adds further support to the notion sex is an important factor to consider when investigating future drug targets for CNS disorders, such as ischemic stroke.

Imaging Functional Recovery Following Ischemic Stroke: Clinical and Preclinical fMRI Studies.

Disability and effectiveness of physical therapy are highly variable following ischemic stroke due to different brain regions being affected. Functional magnetic resonance imaging (fMRI) studies of patients in the months and years following stroke have given some insight into how the brain recovers lost functions. Initially, new pathways are recruited to compensate for the lost region, showing as a brighter blood oxygen-level-dependent (BOLD) signal over a larger area during a task than in healthy controls. Subsequently, activity is reduced to baseline levels as pathways become more efficient, mimicking the process of learning typically seen during development. Preclinical models of ischemic stroke aim to enhance understanding of the biology underlying recovery following stroke. However, the pattern of recruitment and focusing seen in humans has not been observed in preclinical fMRI studies that are highly variable methodologically. Resting-state fMRI studies show more consistency; however, there are still confounding factors to address. Anesthesia and method of stroke induction are the two main sources of variability in preclinical studies; improvements here can reduce variability and increase the intensity and reproducibility of the BOLD response detected by fMRI. Differences in task or stimulus and differences in analysis method also present a source of variability. This review compares clinical and preclinical fMRI studies of recovery following stroke and focuses on how refinement of preclinical models and MRI methods may obtain more representative fMRI data in relation to human studies.

Dietary Intakes Differ by Body Composition Goals: An Observational Study of Professional Rugby Union Players in New Zealand.

Preseason in rugby union is a period of intensive training where players undergo conditioning to prepare for the competitive season. In some cases, this includes modifying body composition through weight gain or fat loss. This study aimed to describe the macronutrient intakes of professional rugby union players during pre-season training. It was hypothesized that players required to gain weight would have a higher energy, carbohydrate and protein intake compared to those needing to lose weight. Twenty-three professional rugby players completed 3 days of dietary assessment and their sum of eight skinfolds were assessed. Players were divided into three groups by the team coaches and medical staff: weight gain, weight maintain and weight loss. Mean energy intakes were 3,875 ± 907 kcal·d-1 (15,965 ± 3,737 kJ·d-1) (weight gain 4,532 ± 804 kcal·d-1; weight maintain 3,825 ± 803 kcal·d-1; weight loss 3,066 ± 407 kcal·d-1) and carbohydrate intakes were 3.7 ± 1.2 g·kg-1·d-1 (weight gain 4.8 ± 0.9 g.kg-1·d-1; weight maintain 2.8 ± 0.7 g·kg-1·d-1; weight loss 2. 6 ± 0.7 g·kg-1·d-1). The energy and carbohydrate intakes are similar to published intakes among rugby union players. There were significant differences in energy intake and the percent of energy from protein between the weight gain and the weight loss group.

Body Image Among Elite Rugby Union Players.

Gibson, C, Hindle, C, McLay-Cooke, R, Slater, J, Brown, R, Smith, B, Baker, D, Healey, P, and Black, K. Body image among elite rugby union players. J Strength Cond Res 33(8): 2217-2222, 2019-There is limited information on the risk of eating disorders (EDs) and body image of elite male athletes. However, research studies suggest that there are some athletes who have poor body image and they may be at increased risk of developing EDs. Therefore, the current study investigated risk of EDs, body image, and the relationship with age, in elite rugby union players during their preseason training period. This cross-sectional study was undertaken at the start of the preseason among elite rugby union players in New Zealand. Twenty-six professional rugby union players completed a 49-item questionnaire on body image and disordered eating. A "body image score" was calculated from questionnaire subscales including "drive for thinness," "bulimia," and "body dissatisfaction," with total scores above 20 indicative of poor body image. Body image scores varied from 8 to 39 out of a possible 0-100. Disordered eating behaviors were reported, including binge eating at least once a week (15%, n = 4/26), pathogenic weight control use (4%, n = 1/26), and avoidance of certain foods (77%, n = 20/26). There was a statistically significant inverse association between the bulimia subscale and age (p = 0.034). At the start of the preseason training period, many elite rugby union players experience disturbances in body image. The prevalence of disordered eating behaviors is of concern, and needs to be minimized due to the negative impact on health and performance. A focus on assessment and education of younger male rugby players may be required to reduce disordered eating patterns.

Wastewater microbial community structure and functional traits change over short timescales.

Wastewater contains microorganisms coming from various sources, e.g. feces discharges, soil infiltrations and sewer biofilms and sediments. The primary objective of this work was to determine if end-of-pipe wastewater microbial community structures exhibits short-timescale variation, and assess possible microbial origins. To this end, we measured hourly physicochemical characteristics of wastewater influent for 2 days and analyzed the microbial community at 4-h intervals using 16S rRNA gene amplicon sequencing. Results showed large variations in the microbial community composition at phylum and genus levels, i.e. Proteobacteria ranged from 44 to 63% of the total relative abundance and Arcobacter ranged from 11 to 22%. Diurnal patterns were observed in the alpha-diversity, beta-diversity and the prevalence of several taxa. Wastewater physicochemical characteristics explained 61% of the total microbial community variance by Canonical Correspondence Analysis (CCA), with flow rate being the main explanatory variable exhibiting a clear diurnal profile. Comparison with public databases using closed reference OTUs revealed that only 7.3% of the sequences were shared with human gut microbiota and 21.7% with soil microbiota, the majority being from the sewer biofilms and sediments. The functional trait, weighted average ribosomal RNA operon (rrn) copy number per genome, was found to be relatively high in the wastewater microbiota (average 3.6, soil 2.1, and human gut 2.6) and significantly correlated with flow, inferring active microbial enrichments in the sewer. The prevalence of Methylophilaceae, methanol oxidation genes and denitrification genes were related to high influent methanol and NO3- concentration in the influent wastewater. These functional organisms and genes indicate important carbon and nutrient removal related functions in the sewer. Together, the observed temporal patterns of the microbial community and functional traits suggest that high wastewater flow causes greater transport of active sewer microorganisms which are functionally important.

Middle Cerebral Artery Occlusion Allowing Reperfusion via Common Carotid Artery Repair in Mice.

The ischemic stroke is a major cause of adult long-term disability and death worldwide. The current treatments available are limited, with only tissue plasminogen activator (tPA) as an approved drug treatment to target ischemic strokes. Current research in the field of ischemic stroke focuses on better understanding the pathophysiology of stroke, to develop and investigate novel pharmaceutical targets. Reliable experimental stroke models are crucial for the progression of potential treatments. The middle cerebral artery occlusion (MCAO) model is clinically relevant and the most frequently used surgical model of ischemic stroke in rodents. However, the outcomes of this model, such as lesion volume, are associated with high levels of variability, particularly in mice. The alternative MCAO model described here allows the reperfusion of the common carotid artery (CCA) and the increased perfusion of the middle cerebral artery (MCA) territory, using a tissue pad with fibrinogen-based sealant to repair the vessel, and the improved welfare of the mice by avoiding external carotid artery (ECA) ligation. This reduces the reliance on the Circle of Willis, which is known to be highly anatomically variable in mice. Representative data show that using this alternative surgical approach decreases the variability in lesion volumes between the traditional MCAO approach and the alternative approach described here.