Hypertensive Response to Ischemic Stroke in the Normotensive Wistar Rat.

Background and Purpose- Over 80% of ischemic stroke patients show an abrupt increase in arterial blood pressure in the hours and days following ischemic stroke. Whether this poststroke hypertension is beneficial or harmful remains controversial and the underlying physiological basis is unclear. Methods- To investigate the dynamic cardiovascular response to stroke, adult Wistar rats (n=5-8 per group, 393±34 g) were instrumented with telemeters to blood pressure, intracranial pressure, renal sympathetic nerve activity, and brain tissue oxygen in the predicted penumbra (Po2). After 2 weeks of recovery, cardiovascular signals were recorded for a 3-day baseline period, then ischemic stroke was induced via transient middle cerebral artery occlusion, or sham surgery. Cardiovascular signals were then recorded for a further 10 days, and the functional sensorimotor recovery assessed using the cylinder and sticky dot tests. Results- Baseline values of all variables were similar between groups. Compared to sham, in the 2 days following stroke middle cerebral artery occlusion produced an immediate, transient rise above baseline in mean blood pressure (21±3 versus 2±4 mm Hg; P<0.001), renal sympathetic nerve activity (54±11% versus 7±4%; P=0.006), and cerebral perfusion pressure (12±5 versus 1±4; P≤0.001). Intracranial pressure increased more slowly, peaking 3 days after middle cerebral artery occlusion (14±6 versus -1±1 mm Hg; P<0.001). Treating with the antihypertensive agent nifedipine after stroke (1.5-0.75 mg/kg per hour SC) ameliorated poststroke hypertension (12±3 mm Hg on day 1; P=0.041), abolished the intracranial pressure increase (3±1; P<0.001) and reduced cerebral perfusion pressure (10±3 mm Hg; P=0.017). Preventing poststroke hypertension affected neither the recovery of sensorimotor function nor infarct size. Conclusions- These findings suggest that poststroke hypertension is immediate, temporally matched to an increase in sympathetic outflow, and elevates cerebral perfusion pressure for several days after stroke, which may enhance cerebral perfusion. Preventing poststroke hypertension does not appear to worsen prognosis after stroke in young, normotensive, and otherwise healthy rats. Visual Overview- An online visual overview is available for this article.

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 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.

Efficacy against subcutaneous or intracranial murine GL261 gliomas in relation to the concentration of the vascular-disrupting agent, 5,6-dimethylxanthenone-4-acetic acid (DMXAA), in the brain and plasma.

PURPOSE: Glioblastomas are amongst the most highly vascularised tumours, and the pursuit of anti-angiogenic approaches such as bevacizumab has provided short-term benefits. The purpose of this study was to determine whether the vascular-disrupting agent, dimethylxanthenone-4-acetic acid (DMXAA), could provide longer-lasting therapeutic benefits in a murine model of glioblastoma. METHODS: Luciferase-expressing murine GL261 glioma cells were inoculated subcutaneously or intracranially into C57Bl/6 mice. Mice with tumours were administered DMXAA, and tumours measured using callipers or by optical imager. Concentrations of DMXAA in plasma and brain were measured by LC-MS/MS. RESULTS: DMXAA (25 mg/kg) caused widespread necrosis at 24 h, a 9-day growth delay and complete regressions in 50 % of the mice with subcutaneous GL261 tumours. Co-administered lenalidomide (100 mg/kg) increased the growth delay to 20 days and the percentage of cures to 83 %. The same dose of DMXAA with or without lenalidomide had minimal effects on intracranial GL261 tumours. Concentrations of DMXAA extracted from brain tissue were approximately 25-fold lower than those measured in plasma 15 min to 4 h after DMXAA administration. The presence of intracranial GL261 tumours did not alter the concentrations of DMXAA entering the brain. CONCLUSIONS: DMXAA does not appear to cross the blood-brain barrier efficiently. Thus, whilst excellent activity was obtained against subcutaneous GL261 gliomas, minimal effects were observed against intracranial GL261 tumours. These results emphasise the need to use appropriate orthotopic models for the evaluation of new approaches for the treatment of brain cancers.

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.

Reduced NMDA-induced apoptosis in neurons lacking ataxia telangiectasia mutated protein.

Apoptosis is an important route to neuronal death in experimental models of stroke, the leading neurological cause of death and disability. Here we explore a role for ataxia telangiectasia mutated protein (ATM), an activator of p53, in a primary cortical culture model of stroke. NMDA-induced apoptosis was reduced in cultures derived from mice with targeted deletions in the ATM gene. In addition, NMDA-induced caspase-3 activity was abolished in cultures lacking two functioning copies of the ATM gene. These data provide evidence to suggest that, in primary cortical culture, NMDA-induced apoptosis is partially mediated through ATM. They provide further evidence to support the hypothesis that DNA damage is one route to apoptosis following neuronal injury.