Campbell, B. C.V. et al. (2019) Penumbral imaging and functional outcome in patients with anterior circulation ischaemic stroke treated with endovascular thrombectomy versus medical therapy: a meta-analysis of individual patient-level data.ABSTRACT Background: CT-perfusion (CTP) and MRI may assist patient selection for endovascular thrombectomy. We aimed to establish whether imaging assessments of ischaemic core and penumbra volumes were associated with functional outcomes and treatment effect.

Campbell, B. C. V. et al. (2018) Effect of general anaesthesia on functional outcome in patients with anterior circulation ischaemic stroke having endovascular thrombectomy versus standard care: a meta-analysis of individual patient data. Lancet Neurology, 17(1), pp. 47-53. (doi:10.1016/S1474-4422(17)30407-6) This is the author's final accepted version.There may be differences between this version and the published version. You are advised to consult the publisher's version if you wish to cite from it.http://eprints.gla.ac.uk/149670/ variables. An alternative approach using propensity-score stratification was also used. To account for between-trial variance we used mixed-effects modeling with a random effect for trial incorporated in all models. Bias was assessed using the Cochrane tool.Findings: Of 1764 patients in 7 trials, 871 were allocated to endovascular thrombectomy. After exclusion of 74 patients (72 who did not undergo the procedure and 2 with missing data on anaesthetic strategy), 236/797 (30%) of endovascular patients were treated under GA. At baseline, GA patients were younger and had shorter time to randomisation but similar pre-treatment clinical severity compared to non-GA. Endovascular thrombectomy improved functional outcome at 3 months versus standard care in both GA (adjusted common odds ratio (cOR) 1·52, 95%CI 1·09-2·11, p=0·014) and non-GA (adjusted cOR 2·33, 95%CI 1·75-3·10, p<0·001) patients. However, outcomes were significantly better for those treated under non-GA versus GA (covariate-adjusted cOR 1·53, 95%CI 1·14-2·04, p=0·004; propensitystratified cOR 1·44 95%CI 1·08-1·92, p=0·012). The risk of bias and variability among studies was assessed to be low.Interpretation: Worse outcomes after endovascular thrombectomy were associated with GA, after adjustment for baseline prognostic variables. These data support avoidance of GA whenever possible. The procedure did, however, remain effective versus standard care in patients treated under GA, indicating that treatment should not be withheld in those who require anaesthesia for medical reasons. Funding:The HERMES collaboration was funded by an unrestricted grant from Medtronic to the University of Calgary. Research in contextEvidence before this study between abolition of the thrombectomy treatment effect in MR CLEAN and no effect in THRACE. Three single-centre randomised trials of general anaesthesia versus conscious sedation found either no difference in functional outcome between groups or a slight benefit of general anaesthesia. Added value of this studyThese data from contemporary, high quality randomised trials form the largest study to date of the association between general anesthesia and the benefit of endovascular thrombectomy versus standard care. We used two different approaches to adjust for baseline imbalances (multivariable logistic regression and propensity-score stratification). We found that GA for endovascular thrombectomy, as practiced in contemporary clinical care across a wide range of expert centres during the rand...

BackgroundMalignant glioma cells are particularly motile and can travel diffusely through the brain parenchyma, apparently without following anatomical structures to guide their migration. The neural adhesion/recognition protein L1 (L1CAM; CD171) has been implicated in contributing to stimulation of motility and metastasis of several non-neural cancer types. We explored the expression and function of L1 protein as a stimulator of glioma cell motility using human high-grade glioma surgical specimens and established rat and human glioma cell lines.ResultsL1 protein expression was found in 17 out of 18 human high-grade glioma surgical specimens by western blotting. L1 mRNA was found to be present in human U-87/LacZ and rat C6 and 9L glioma cell lines. The glioma cell lines were negative for surface full length L1 by flow cytometry and high resolution immunocytochemistry of live cells. However, fixed and permeablized cells exhibited positive staining as numerous intracellular puncta. Western blots of cell line extracts revealed L1 proteolysis into a large soluble ectodomain (~180 kDa) and a smaller transmembrane proteolytic fragment (~32 kDa). Exosomal vesicles released by the glioma cell lines were purified and contained both full-length L1 and the proteolyzed transmembrane fragment. Glioma cell lines expressed L1-binding αvβ5 integrin cell surface receptors. Quantitative time-lapse analyses showed that motility was reduced significantly in glioma cell lines by 1) infection with an antisense-L1 retroviral vector and 2) L1 ectodomain-binding antibodies.ConclusionOur novel results support a model of autocrine/paracrine stimulation of cell motility in glioma cells by a cleaved L1 ectodomain and/or released exosomal vesicles containing L1. This mechanism could explain the diffuse migratory behavior of high-grade glioma cancer cells within the brain.

Neurons may be particularly sensitive to disruptions in transcription factor trafficking. Survival and injury signals must traverse dendrites or axons, in addition to soma, to affect nuclear transcriptional responses. Transcription factors exhibit continued nucleocytoplasmic shuttling; the predominant localization is regulated by binding to anchoring proteins that mask nuclear localization/export signals and/or target the factor for degradation. Two functional groups of karyopherins, importins and exportins, mediate RanGTPase-dependent transport through the nuclear pore. A growing number of recent studies, in Alzheimer, Parkinson, and Lewy body diseases, amyotrophic lateral sclerosis, and human immunodeficiency virus encephalitis, implicate aberrant cytoplasmic localization of transcription factors and their regulatory kinases in degenerating neurons. Potential mechanisms include impaired nuclear import, enhanced export, suppression of degradation, and sequestration in protein aggregates or organelles and may reflect unmasking of alternative cytoplasmic functions, both physiologic and pathologic. Some "nuclear" factors also function in mitochondria, and importins are also involved in axonal protein trafficking. Detrimental consequences of a decreased nuclear to cytoplasmic balance include suppression of neuroprotective transcription mediated by cAMP- and electrophile/antioxidant-response elements and gain of toxic cytoplasmic effects. Studying the pathophysiologic mechanisms regulating transcription factor localization should facilitate strategies to bypass deficits and restore adaptive neuroprotective transcriptional responses.

The microtubule (MT) system is important for many aspects of neuronal function, including motility, differentiation, and cargo trafficking. Parkinson’s disease (PD) is associated with increased oxidative stress and alterations in the integrity of the axodendritic tree. To study dynamic mechanisms underlying the neurite shortening phenotype observed in many PD models, we employed the well-characterized oxidative parkinsonian neurotoxin, 6-hydroxydopamine (6OHDA). In both acute and chronic sub-lethal settings, 6OHDA-induced oxidative stress elicited significant alterations in MT dynamics, including reductions in MT growth rate, increased frequency of MT pauses/retractions, and increased levels of tubulin acetylation. Interestingly, 6OHDA decreased the activity of tubulin deacetylases, specifically sirtuin 2 (SIRT2), through more than one mechanism. Restoration of tubulin deacetylase function rescued the changes in MT dynamics and prevented neurite shortening in neurondifferentiated, 6OHDA-treated cells. These data indicate that impaired tubulin deacetylation contributes to altered MT dynamics in oxidatively-stressed cells, conferring key insights for potential therapeutic strategies to correct MT-related deficits contributing to neuronal aging and disease.

We describe a novel fully automated high-throughput time-lapse microscopy system and evaluate its performance for precisely tracking the motility of several glioma and osteoblastic cell lines. Use of this system revealed cell motility behavior not discernable with conventional techniques by collecting data (1) from closely spaced time points (minutes), (2) over long periods (hours to days), (3) from multiple areas of interest, (4) in parallel under several different experimental conditions. Quantitation of true individual and average cell velocity and path length was obtained with high spatial and temporal resolution in ''scratch'' or ''wound healing'' assays. This revealed unique motility dynamics of drug-treated and adhesion molecule-transfected cells and, thus, this is a considerable improvement over current methods of measurement and analysis. Several fluorescent vital labeling methods commonly used for end-point analyses (GFP expression, DiO lipophilic dye, and Qtracker nanocrystals) were found to be useful for timelapse studies under specific conditions that are described. To illustrate one application, fluorescently labeled tumor cells were seeded onto cell monolayers expressing ectopic adhesion molecules, and this resulted in consistently reduced tumor cell migration velocities. These highly quantitative time-lapse analysis methods will promote the creation of new cell motility assays and increase the resolution and accuracy of existing assays.

Eristostatin, an RGD-containing disintegrin isolated from the venom of Eristicophis macmahoni, inhibits lung or liver colonization of melanoma cells in a mouse model. In this study, transwell migration and in vitro wound closure assays were used to determine the effect of eristostatin on the migration of melanoma cells. Eristostatin significantly impaired the migration of five human melanoma cell lines. Furthermore, it specifically inhibited cell migration on fibronectin in a concentration-dependent manner, but not that on collagen IV or laminin. In contrast, eristostatin was found to have no effect on cell proliferation or angiogenesis. These results indicate that the interaction between eristostatin and melanoma cells may involve fibronectin-binding integrins that mediate cell migration. Mutations to alanine of seven residues within the RGD loop of eristostatin and four residues outside the RGD loop of eristostatin resulted in significantly less potency in both platelet aggregation and wound closure assays. For six of the mutations, however, decreased activity was found only in the latter assay. We conclude that a different mechanism and/or integrin is involved in these two cell activities.

Age-related neurodegenerative diseases are associated with alterations in gene expression in affected neurons. One of the mechanisms that could account for this is altered subcellular localization of transcription factors, which has been observed in human post-mortem brains of each of the major neurodegenerative diseases, including Parkinson’s disease (PD). The specific mechanisms are yet to be elucidated; however a potential mechanism involves alterations in nuclear transport. In this study, we examined the nucleocytoplasmic trafficking of select transcription factors in response to a PD-relevant oxidative injury, 6-hydroxydopamine (6OHDA). Utilizing a well-established model of ligand-regulated nucleocytoplasmic shuttling, the glucocorticoid receptor, we found that 6OHDA selectively impaired nuclear import through an oxidative mechanism without affecting nuclear export or nuclear retention. Interestingly, impaired nuclear import was selective as Nrf2 (nuclear factor E2-related factor 2) nuclear localization remained intact in 6OHDA-treated cells. Thus, oxidative stress specifically impacts the subcellular localization of some but not all transcription factors, which is consistent with observations in post-mortem PD brains. Our data further implicate a role for altered microtubule dependent trafficking in the differential effects of 6OHDA on transcription factor import. Oxidative disruption of microtubule-dependent nuclear transport may contribute to selective declines in transcriptional responses of aging or diseased dopaminergic cells.