Delayed citalopram administration reduces brain inflammation and enhances skilled motor function after ischaemic stroke in 'MacGreen' mice.

Recent evidence suggests that treatment with antidepressants may promote functional recovery. However, the timeframe in which these pharmacological agents can influence stroke recovery is not well understood. This research investigated whether delayed administration of citalopram, used clinically in the management of post-stroke depression, could improve long-term functional recovery following experimental stroke. MacGreen mice carrying an enhanced green fluorescent protein reporter gene in monocyte and macrophage populations were subjected to 45 min occlusion of the middle cerebral artery. Animals were administered citalopram (10 mg/kg/day, n = 20) or saline (n = 20) starting 3 days after stroke for 28 days. Neurological deficits and skilled motor performance in the staircase task were recorded for 9 weeks post stroke. Grey and white matter structural lesions were quantified at Week 9, and enhanced green fluorescent protein immunohistochemistry was used to evaluate the effect of citalopram on inflammation. Twenty-five animals were included in the final analysis. Citalopram-treated animals (n = 13) showed a significant increase in impaired forepaw use in the staircase task compared with saline-treated animals (n = 12) 2, 3 and 7 weeks post stroke but no difference in neurological score at any time point examined. Citalopram treatment was associated with decreased monocyte/macrophage cell density and increased white matter tract integrity within the ipsilateral cortex. In conclusion, delayed administration of citalopram decreased brain inflammation and produced functional gains in our mouse model of stroke. Beneficial effects on skilled motor functions were long-lasting.

Anti-Epileptic Medication Exposure Influences Functional Status in New Zealand Stroke Patients: A Retrospective Population-Level Study.

BACKGROUND: Patients who develop seizures after stroke have disproportionately poorer outcomes and increased mortality. OBJECTIVE: Our objective was to investigate whether exposure to anti-epileptic medications influenced long-term functional status after stroke. METHODS: We used linked health administrative data from a cohort of adult stroke patients in New Zealand. Demographics and prescription information were obtained from the National Minimum Dataset and Pharmaceutical Collection, respectively. Activities of daily living (ADL) scores for the same patients were obtained using the International Resident Assessment Instrument. Beta regression was used to investigate the relationship between anti-epileptic drug (AED) exposure and functional status. RESULTS: The study included 3606 patients with a single ischaemic stroke between 2012 and 2017. In total, 15% were dispensed an AED in the 3 months before or after stroke. The adjusted odds ratio (OR) for AED exposure was 1.29 (95% confidence interval [CI] 1.15-1.45). Overall AED exposure, categorical body mass index (BMI), ethnicity, length of hospital stay, and exposure to paracetamol, opioids, anti-psychotics, and anti-nausea medications were significantly associated with changes in the mean ADL score percentages. Considering the exposure timeframe, the ORs for AED exposure only after stroke and for exposure both before and after stroke were 1.52 (95% CI 1.31-1.78) and 1.09 (95% CI 0.93-1.27), respectively. CONCLUSION: Stroke patients with AED exposure had greater odds of a higher ADL score, indicating a poorer long-term functional status than those unexposed to AEDs. The timeframe of exposure impacted on functional status, with patients exposed only after stroke having increased odds of higher ADL scores than those exposed both before and after stroke.

Higher BMI Confers a Long-Term Functional Status Advantage in Elderly New Zealand European Stroke Patients.

OBJECTIVE: Obesity is a risk factor for ischaemic stroke but provides a survival advantage. The relationship between body mass index (BMI) and long-term function is less clear. The presence of an obesity paradox can inform clinical care and identify vulnerable patients who need additional support post-stroke. MATERIALS AND METHODS: This study used linked health administrative data of a population based cohort of adult patients who experienced an ischaemic stroke between 2012 and 2017 in New Zealand. Patient demographics were obtained from the National Minimum Dataset (NMDS). BMI and Activities of Daily Living scores (ADLs) for the same patients were obtained from the International Resident Assessment Instrument (InterRAI™). RESULTS: Linked data was obtained for 3731 patients. Ninety-five percent of the cohort were aged 65 or older and the average age of stroke was 84.5 years. The majority of patients (55%) identified as New Zealand European. Beta regression indicated BMI and European ethnicity were negatively associated with ADL score. Univariate analysis confirmed patients with underweight stroke had significantly higher ADL scores than other BMI categories (p<0.001), however functional status for patients with overweight and obesity were comparable. Further, Asian and Pacific Peoples had higher ADL scores than Europeans (p<0.05). A higher BMI was advantageous to all ADL subscores. CONCLUSION: An abridged obesity paradox was evident in our cohort of stroke patients where a BMI in the overweight, but not obese range conferred a long-term functional status advantage. Collectively these results suggest underweight and non-European patients may require additional supportive clinical care post-stroke.

Inconsistencies in histone acetylation patterns among different HD model systems and HD post-mortem brains.

Huntington's disease (HD) is a neurodegenerative disorder caused by a CAG trinucleotide repeat expansion in exon 1 of the huntingtin gene. Emerging evidence shows that additional epigenetic factors can modify disease phenotypes. Harnessing the ability of the epigenome to modify the disease for therapeutic purposes is therefore of interest. Epigenome modifiers, such as histone deacetylase inhibitors (HDACi), have improved pathology in a range of HD models. Yet in clinical trials, HDACi have failed to alleviate HD symptoms in patients. This study investigated potential reasons for the lack of translation of the therapeutic benefits of HDACi from lab to clinic. We analysed histone acetylation patterns of immuno-positive nuclei from brain sections and tissue microarrays from post-mortem human control and HD cases alongside several well-established HD models (OVT73 transgenic HD sheep, YAC128 mice, and an in vitro cell model expressing 97Q mutant huntingtin). Significant increases in histone H4 acetylation were observed in post-mortem HD cases, OVT73 transgenic HD sheep and in vitro models; these changes were absent in YAC128 mice. In addition, nuclear labelling for acetyl-histone H4 levels were inversely proportional to mutant huntingtin aggregate load in HD human cortex. Our data raise concerns regarding the utility of HDACi for the treatment of HD when regions of pathology exhibit already elevated histone acetylation patterns and emphasize the importance of searching for alternative epigenetic targets in future therapeutic strategies aiming to rescue HD phenotypes.

Therapeutic Relevance of Elevated Blood Pressure After Ischemic Stroke in the Hypertensive Rats.

Over 80% of patients exhibit an acute increase in blood pressure (BP) following stroke. Current clinical guidelines make no distinction in BP management between patients with or without prior hypertension. Spontaneously hypertensive (SH) rats were preinstrumented with telemeters to record BP, intracranial pressure, and brain tissue oxygen in the predicted ischemic penumbra for 3 days before and 10 days after transient middle cerebral artery occlusion (n=8 per group) or sham (n=5). Before stroke, BP was either left untreated or chronically treated to a normotensive level (enalapril 10 mg/kg per day). Poststroke elevations in BP were either left uncontrolled, controlled (to the prestroke baseline level), or overcontrolled (to a normotensive level) via subcutaneous infusion of labetalol. Baseline values of intracranial pressure and brain tissue oxygen were similar between all groups, whereas BP was lower in treated SH rats (144±3 versus 115±5 mm Hg; P<0.001). Following middle cerebral artery occlusion, a similar rise in BP was observed in untreated (+16±2 mm Hg; P=0.005) and treated SH rats (+13±5 mm Hg; P=0.021). Intervening to prevent BP from increasing after stroke did not worsen outcome. However, reducing BP below prestroke baseline levels was associated with higher intracranial pressure (days 1-3; P<0.001), reduced cerebral perfusion pressure (days 2-4; P<0.001), higher mortality, slower functional recovery and larger infarct volumes. Although treating to maintain BP at the prestroke baseline level was not detrimental, our results suggest that when setting BP targets after stroke, consideration must be given to the potential negative impact of inadvertent excessive BP lowering in subjects with undiagnosed or poorly controlled hypertension.

Delayed Varenicline Administration Reduces Inflammation and Improves Forelimb Use Following Experimental Stroke.

BACKGROUND: Pharmacological activation of the cholinergic anti-inflammatory pathway (CAP), specifically by activating α7 nicotinic acetylcholine receptors, has been shown to confer short-term improvements in outcome. Most studies have investigated administration within 24 hours of stroke, and few have investigated drugs approved for use in human patients. We investigated whether delayed administration of varenicline, a high-affinity agonist at α7 nicotinic receptors and an established therapy for nicotine addiction, decreased brain inflammation and improved functional performance in a mouse model of experimental stroke. METHODS: CSF-1R-EGFP (MacGreen) mice were subjected to transient middle cerebral artery occlusion and administered varenicline (2.5 mg/kg/d for 7 days) or saline (n = 10 per group) 3 days after stroke. Forelimb asymmetry was assessed in the Cylinder test every 2 days after surgery, and structural lesions were quantified at day 10. Enhanced green fluorescent protein (EGFP) and growth associated protein 43 (GAP43) immunohistochemistry were used to evaluate the effect of varenicline on inflammation and axonal regeneration, respectively. RESULTS: Varenicline-treated animals showed a significant increase in impaired forelimb use compared with saline-treated animals 10 days after stroke. Varenicline treatment was associated with reduced EGFP expression and increased GAP43 expression in the striatum of MacGreen mice. CONCLUSION: Our results show that delayed administration of varenicline promotes recovery of function following experimental stroke. Motor function improvements were accompanied by decreased brain inflammation and increased axonal regeneration in nonpenumbral areas. These results suggest that the administration of an exogenous nicotinic agonist in the subacute phase following stroke may be a viable therapeutic strategy for stroke patients.

Cyclic-RGDyC functionalized liposomes for dual-targeting of tumor vasculature and cancer cells in glioblastoma: An in vitro boron neutron capture therapy study.

The efficacy of boron neutron capture therapy depends on the selective delivery of 10B to the target. Integrins αvß3 are transmembrane receptors over-expressed in both glioblastoma cells and its neovasculature. In this study, a novel approach to dual-target glioblastoma vasculature and tumor cells was investigated. Liposomes (124 nm) were conjugated with a αvß3 ligand, cyclic arginine-glycine-aspartic acid-tyrosine-cysteine peptide (c(RGDyC)-LP) (1% molar ratio) through thiol-maleimide coupling. Expression of αvß3 in glioblastoma cells (U87) and human umbilical vein endothelial cells (HUVEC), representing tumor angiogenesis, was determined using Western Blotting with other cells as references. The results showed that both U87 and HUVEC had stronger expression of αvß3 than other cell types, and the degree of cellular uptake of c(RGDyC)-LP correlated with the αvß3-expression levels of the cells. In contrast, control liposomes without c(RGDyC) showed little cellular uptake, regardless of cell type. In an in vitro boron neutron capture therapy study, the c(RGDyC)-LP containing sodium borocaptate generated more rapid and significant lethal effects to both U87 and HUVEC than the control liposomes and drug solution. Interestingly, neutron irradiated U87 and HUVEC showed different types of subsequent cell death. In conclusion, this study has demonstrated the potential of a new dual-targeting strategy using c(RGDyC)-LP to improve boron neutron capture therapy for glioblastoma.

Varenicline improves motor and cognitive deficits and decreases depressive-like behaviour in late-stage YAC128 mice.

OBJECTIVE: Studies in the post mortem human brain and in genetic mouse model suggest that dysfunctional cholinergic neurotransmission, through a loss of agonist rather than receptors may be a significant contributing factor to HD pathology. If correct, pharmacological replacement may therefore be a potential treatment strategy. We have investigated whether chronic administration of the selective nicotinic partial agonist varenicline improved motor, cognitive and affective symptoms in a transgenic mouse model of Huntington's disease. METHOD: The performance of 15 month old YAC128 mice and age-matched wild-type littermates was assessed in the rotarod, T-maze, novel object recognition, novelty suppressed feeding and forced swim tests prior to and after treatment with varenicline (5 mg/kg/day for 28 days via miniosmotic pump). Thymidine analogues, whilst DARPP32 and EM48 immunohistochemistry were used to assess the effect of varenicline on progenitor cell proliferation and survival, medium spiny neurons and aggregate formation respectively. RESULTS: Chronic treatment with varenicline significantly improved motor coordination, delay-dependent memory and reduced depressive-like behaviour in late stage YAC128 mice. Varenicline also produced genotype-independent improvements in recognition memory and reduced anxiety. In addition, varenicline displayed anxiolytic effects and improved spatial memory in the absence of compromised function. Functional improvements were accompanied by neuropathological changes including increased aggregate formation, neuroprotection and increased progenitor cell proliferation and survival. INTERPRETATION: Our findings provide evidence that administration of an exogenous nicotinic agonist may be of clinical benefit in the treatment of late-stage Huntington's disease.

Varenicline improves motor and cognitive symptoms in early Huntington's disease.

The aim of this study was to describe the effects of varenicline, a smoking cessation aid that acts as a nicotinic agonist, on cognitive function in patients with early clinical Huntington's disease (HD) who were current smokers. Three gene-positive patients transitioning to symptomatic HD were evaluated using the Unified Huntington's Disease Rating Scale part I and III (motor and behavioral subscales) at baseline and after 4 weeks of treatment. Cognitive function was assessed using a touch screen computer-based neurocognitive test battery (IntegNeuro®). Varenicline (1 mg twice daily) significantly improved performance in executive function and emotional recognition tasks. Our case reports describe no clinically significant adverse effects and suggest that varenicline improves aspects of cognitive function in patients with early HD. A randomized controlled study is now underway.

Inhibition of glutamate regulated calcium entry into leukemic megakaryoblasts reduces cell proliferation and supports differentiation.

Human megakaryocytes release glutamate and express glutamate-gated Ca(2+)-permeable N-methyl-D-aspartate receptors (NMDARs) that support megakaryocytic maturation. While deregulated glutamate pathways impact oncogenicity in some cancers, the role of glutamate and NMDARs in megakaryocytic malignancies remains unknown. The aim of this study was to determine if NMDARs participate in Ca(2+) responses in leukemic megakaryoblasts and if so, whether modulating NMDAR activity could influence cell growth. Three human cell lines, Meg-01, Set-2 and K-562 were used as models of leukemic megakaryoblasts. NMDAR components were examined in leukemic cells and human bone marrow, including in megakaryocytic disease. Well-established NMDAR modulators (agonists and antagonists) were employed to determine NMDAR effects on Ca(2+) flux, cell viability, proliferation and differentiation. Leukemic megakaryoblasts contained combinations of NMDAR subunits that differed from normal bone marrow and the brain. NMDAR agonists facilitated Ca(2+) entry into Meg-01 cells, amplified Ca(2+) responses to adenosine diphosphate (ADP) and promoted growth of Meg-01, Set-2 and K-562 cells. Low concentrations of NMDAR inhibitors (riluzole, memantine, MK-801 and AP5; 5-100µM) were weakly cytotoxic but mainly reduced cell numbers by suppressing proliferation. The use-dependent NMDAR inhibitor, memantine (100µM), reduced numbers and proliferation of Meg-01 cells to less than 20% of controls (IC50 20µM and 36µM, respectively). In the presence of NMDAR inhibitors cells acquired morphologic and immunophenotypic features of megakaryocytic differentiation. In conclusion, NMDARs provide a novel pathway for Ca(2+) entry into leukemic megakaryoblasts that supports cell proliferation but not differentiation. NMDAR inhibitors counteract these effects, suggesting a novel opportunity to modulate growth of leukemic megakaryoblasts.

Chemokines direct neural progenitor cell migration following striatal cell loss.

In this study we demonstrate the chemokines MCP-1, MIP-1alpha and GRO-alpha play a role in directing adult subventricular zone (SVZ)-derived progenitor cell migration following striatal cell death. MCP-1, MIP-1alpha and GRO-alpha were significantly upregulated in the striatum 2-3 days following QA-induced lesioning, correlating with maximum SVZ-derived progenitor cell recruitment into the lesioned striatum. We established that SVZ-derived progenitor cells express receptors for each chemokine, and demonstrated MCP-1, MIP-1alpha and GRO-alpha to be potent chemoattractants for SVZ-derived progenitor cells in vitro. Immunofluorescence revealed MCP-1, MIP-1alpha and GRO-alpha are predominantly expressed in the striatum by NG2-positive cells that appear to infiltrate from the bloodstream 6 h following QA lesioning. These results indicate that upregulation of MCP-1, MIP-1alpha and GRO-alpha following striatal cell death leads to chemoattraction of SVZ-derived progenitor cells into the damaged striatum and raises a potential role for blood-derived cells in directing the recruitment of SVZ-derived progenitors following brain injury.

APOE epsilon3 gene transfer attenuates brain damage after experimental stroke.

Apolipoprotein E (apoE, protein; APOE, gene) is the major lipid-transport protein in the brain and plays an important role in modulating the outcome and regenerative processes after acute brain injury. The aim of the present study was to determine if gene transfer of the epsilon3 form of APOE improves outcome in a murine model of transient focal cerebral ischaemia. Mice received an intrastriatal injection of vehicle, a second-generation adenoviral vector containing the green fluorescent protein gene (Ad-GFP) or a vector containing the APOE epsilon3 gene (Ad-APOE) 3 days before 60 mins focal ischaemia. Green fluorescent protein expression was observed in cells throughout the striatum and subcortical white matter indicating successful gene transfer and expression. ApoE levels in the brain were significantly increased after Ad-APOE compared with Ad-GFP or vehicle treatment. Ad-APOE treatment reduced the volume of ischaemic damage by 50% compared with Ad-GFP or vehicle treatment (13+/-3 versus 29+/-4 versus 27+/-5 mm(3)). The extent of postischaemic apoE immunoreactivity was enhanced in Ad-APOE compared with Ad-GFP or vehicle treated mice. These results show the ability of APOE gene transfer to markedly improve outcome after cerebral ischaemia and suggest that modulating apoE levels may be a potential strategy in human stroke therapy.

Increased expression of macrophage receptor with collagenous structure (MARCO) in mouse cortex following middle cerebral artery occlusion.

Ischaemia induces activation of resident microglia and infiltration of peripheral monocyte/macrophage cells into the central nervous system. The role of scavenger receptors, receptors critical to the recognition and clearance of cell debris, has not been investigated during cerebral ischaemia. MARCO is an inducible member of the scavenger receptor family unique to cells of monocytic lineage and is a cell surface marker that plays a critical role in the differentiation of monocytes to dendritic cells. To understand the role of MARCO in cerebral ischaemia, we investigated its expression in mice following middle cerebral artery (MCA) occlusion. No MARCO mRNA expression was observed in naive mouse brain. There was no significant increase in expression of MARCO mRNA following transient occlusion (60min) of the MCA at any time point up to 24 h. However, a significant, marked increase in MARCO mRNA expression was observed at 24 h in the cortex of mouse brains after a permanent occlusion of the MCA. The increased expression of MARCO mRNA at 24 h after prolonged ischaemia is consistent with its putative role in the clearance of debris and/or degenerating cells after severe ischaemia and supports previous publications showing the presence of dendritic cells around permanently occluded lesions.