Oxidative stress and the use of antioxidants in stroke

Transient or permanent interruption of cerebral blood flow by occlusion of a cerebral artery gives rise to an ischaemic stroke leading to irreversible damage or dysfunction to the cells within the affected tissue along with permanent or reversible neurological deficit. Extensive research has identified excitotoxicity, oxidative stress, inflammation and cell death as key contributory pathways underlying lesion progression. The cornerstone of treatment for acute ischaemic stroke remains reperfusion therapy with recombinant tissue plasminogen activator (rt-PA). The downstream sequelae of events resulting from spontaneous or pharmacological reperfusion lead to an imbalance in the production of harmful reactive oxygen species (ROS) over endogenous anti-oxidant protection strategies. As such, anti-oxidant therapy has long been investigated as a means to reduce the extent of injury resulting from ischaemic stroke with varying degrees of success. Here we discuss the production and source of these ROS and the various strategies employed to modulate levels. These strategies broadly attempt to inhibit ROS production or increase scavenging or degradation of ROS. While early clinical studies have failed to translate success from bench to bedside, the combination of anti-oxidants with existing thrombolytics or novel neuroprotectants may represent an avenue worthy of clinical investigation. Clearly, there is a pressing need to identify new therapeutic alternatives for the vast majority of patients who are not eligible to receive rt-PA for this debilitating and devastating disease

Gene Therapy for Cardiovascular Disease

The last decade has seen substantial advances in the development of gene therapy strategies and vector technology for the treatment of a diverse number of diseases, with a view to translating the successes observed in animal models into the clinic. Perhaps the overwhelming drive for the increase in vascular gene transfer studies is the current lack of successful long-term pharmacological treatments for complex cardiovascular diseases. The increase in cardiovascular disease to epidemic proportions has also led many to conclude that drug therapy may have reached a plateau in its efficacy and that gene therapy may represent a realistic solution to a long-term problem. Here, we discuss gene delivery approaches and target diseases

Improved gene delivery to human saphenous vein cells and tissue using a peptide-modified adenoviral vector

The establishment of efficient gene delivery to target human tissue is a major obstacle for transition of gene therapy from the pre-clinical phases to the clinic. The poor long-term patency rates for coronary artery bypass grafting (CABG) is a major clinical problem that lacks an effective and proven pharmacological intervention. Late vein graft failure occurs due to neointima formation and accelerated atherosclerosis. Since CABG allows a clinical window of opportunity to genetically modify vein ex vivo prior to grafting it represents an ideal opportunity to develop gene-based therapies. Adenoviral vectors have been frequently used for gene delivery to vein ex vivo and pre-clinical studies have shown effective blockade in neointima development by overexpression of candidate therapeutic genes. However, high titers of adenovirus are required to achieve sufficient gene delivery to provide therapeutic benefit. Improvement in the uptake of adenovirus into the vessel wall would therefore be of benefit. Here we determined the ability of an adenovirus serotype 5 vector genetically-engineered with the RGD-4C integrin targeting peptide inserted into the HI loop (Ad-RGD) to improve the transduction of human saphenous vein smooth muscle cells (HSVSMC), endothelial cells (HSVEC) and intact saphenous vein compared to a non-modified virus (Ad-CTL). We exposed each cell type to virus for 10, 30 or 60 mins and measured transgene at 24 h post infection. For both HSVSMC and HSVEC Ad-RGD mediated increased transduction, with the largest increases observed in HSVSMC. When the experiments were repeated with intact human saphenous vein (the ultimate clinical target for gene therapy), again Ad-RGD mediated higher levels of transduction, at all clinically relevant exposures times (10, 30 and 60 mins tissue:virus exposure). Our study demonstrates the ability of peptide-modified Ad vectors to improve transduction to human vein graft cells and tissue and has important implications for gene therapy for CABG

Combined antiapoptotic and antioxidant approach to acute neuroprotection for stroke in hypertensive rats

We hypothesized that targeting key points in the ischemic cascade with combined neuroglobin (Ngb) overexpression and c-jun N-terminal kinase (JNK) inhibition (SP600125) would offer greater neuroprotection than single treatment after in vitro hypoxia/reoxygenation and in a randomized, blinded in vivo experimental stroke study using a clinically relevant rat strain. Male spontaneously hypertensive stroke-prone rats underwent transient middle cerebral artery occlusion (tMCAO) and were divided into the following groups: tMCAO; tMCAO+control GFP-expressing canine adenovirus-2, CAVGFP; tMCAO+Ngb-expressing CAV-2, CAVNgb; tMCAO+SP600125; tMCAO+CAVNgb+SP600125; or sham procedure. Rats were assessed till day 14 for neurologic outcome before infarct determination. In vitro, combined lentivirus-mediated Ngb overexpression+SP600125 significantly reduced oxidative stress and apoptosis compared with single treatment(s) after hypoxia/reoxygenation in B50 cells. In vivo, infarct volume was significantly reduced by CAVNgb, SP600125, and further by CAVNgb+SP600125. The number of Ngb-positive cells in the peri-infarct cortex and striatum was significantly increased 14 days after tMCAO in animals receiving CAVNgb. Neurologic outcome, measured using a 32-point neurologic score, significantly improved with CAVNgb+SP600125 compared with single treatments at 14 days after tMCAO. Combined Ngb overexpression with JNK inhibition reduced hypoxia/reoxygenation-induced oxidative stress and apoptosis in cultured neurons and reduced infarct and improved neurologic outcome more than single therapy after in vivo experimental stroke in hypertensive rats

The glymphatic system and multiple sclerosis: An evolving connection

Multiple sclerosis (MS) is a complex autoimmune disorder that affects the central nervous system, resulting in demyelination and an array of neurological manifestations. Recently, there has been significant scientific interest in the glymphatic system, which operates as a waste-clearance system for the brain. This article reviews the existing literature, and explores potential links between the glymphatic system and MS, shedding light on its evolving significance in the context of MS pathogenesis. The authors consider the pathophysiological implications of glymphatic dysfunction in MS, the impact of disrupted sleep on glymphatic function, and the bidirectional relationship between MS and sleep disturbances. By offering an understanding of the intricate interplay between the glymphatic system and MS, this review provides valuable insights which may lead to improved diagnostic techniques and more effective therapeutic interventions

The counter regulatory axis of the renin angiotensin system in the brain and ischaemic stroke: insight from preclinical stroke studies and therapeutic potential

Stroke is the 2nd leading cause of death worldwide and the leading cause of physical disability and cognitive issues. Although we have made progress in certain aspects of stroke treatment, the consequences remain substantial and new treatments are needed. Hypertension has long been recognised as a major risk factor for stroke, both haemorrhagic and ischaemic. The renin angiotensin system (RAS) plays a key role in blood pressure regulation and this, plus local expression and signalling of RAS in the brain, both support the potential for targeting this axis therapeutically in the setting of stroke. While historically, focus has been on suppressing classical RAS signalling through the angiotensin type 1 receptor (AT1R), the identification of a counter-regulatory axis of the RAS signalling via the angiotensin type 2 receptor (AT2R) and Mas receptor has renewed interest in targeting the RAS. This review describes RAS signalling in the brain and the potential of targeting the Mas receptor and AT2R in preclinical models of ischaemic stroke. The animal and experimental models, and the route and timing of intervention, are considered from a translational perspective

Use of in vivo phage display to engineer novel adenoviruses for targeted delivery to the cardiac vasculature

We performed in vivo phage display in the stroke prone spontaneously hypertensive rat, a cardiovascular disease model, and the normotensive Wistar Kyoto rat to identify cardiac targeting peptides, and then assessed each in the context of viral gene delivery. We identified both common and strain-selective peptides, potentially indicating ubiquitous markers and those found selectively in dysfunctional microvasculature of the heart. We show the utility of the peptide, DDTRHWG, for targeted gene delivery in human cells and rats in vivo when cloned into the fiber protein of subgroup D adenovirus 19p. This study therefore identifies cardiac targeting peptides by in vivo phage display and the potential of a candidate peptide for vector targeting strategies

Direct imaging of glymphatic flow using H217O MRI

The recently proposed glymphatic pathway for solute transport and waste clearance from the brain has been the focus of intense debate. By exploiting an isotopically enriched MRI tracer, H217O, we directly imaged glymphatic water transport in the rat brain in vivo for the first time. Our results reveal glymphatic transport that is dramatically faster and more extensive than previously thought and unlikely to be explained by diffusion alone. Moreover, we confirm the critical role of aquaporin-4 channels in glymphatic transport

Variability of functional outcome measures used in animal models of stroke and vascular cognitive impairment – a review of contemporary studies

Despite promising preclinical data, few novel stroke therapies have shown efficacy in man. Efforts to improve standards in conduct and reporting of preclinical research are ongoing. In clinical trials, inconsistency in outcome measures led to regulatory agencies and funders mandating use of a core set of functional outcomes. Our aim was to describe functional outcome measures in preclinical stroke and vascular cognitive impairment (VCI) studies. From 14 high impact journals (January 2005–December 2015 inclusive), 91,956 papers were screened with 1302 full texts analyzed for stroke (ischemic and hemorrhagic) and 56 for VCI studies. In total, 636 (49%) stroke and 37 (66%) VCI papers reported functional outcome measures. There were 74 different functional assessments reported in stroke and 20 in VCI studies. Neurological deficit scores (74%) and Morris water maze (60%) were most commonly used in stroke and VCI, respectively. However, inconsistencies in methods used to assess and score recovery were noted. Neurological and behavioural functional outcome measures are increasingly used in preclinical stroke or VCI studies; however, there is substantial variation in methods. A strict standardized outcome set may not be suitable for translational work, but greater consistency in choice, application and reporting of outcomes may improve the science

Unscheduled changes in pre-clinical stroke model housing contributes to variance in physiological and behavioural data outcomes: a post hoc analysis

Ischaemic stroke presents a significant problem worldwide with no neuroprotective drugs available. Many of the failures in the search for neuroprotectants are attributed to failure to translate from pre-clinical models to humans, which has been combatted with rigorous pre-clinical stroke research guidelines. Here, we present post hoc analysis of a pre-clinical stroke trial, conducted using intraluminal filament transient middle cerebral artery occlusion in the stroke-prone spontaneously hypertensive rat, whereby unscheduled changes were implemented in the animal housing facility. These changes severely impacted body weight post-stroke resulting in a change from the typical body weight of 90.6% of pre-surgery weight post-stroke, to on average 80.5% of pre-surgery weight post-stroke. The changes also appeared to impact post-stroke blood pressure, with an increase from 215.4 to 240.3 mmHg between housing groups, and functional outcome post-stroke, with a 38% increased latency to contact in the sticky label test. These data highlight the importance of tightly controlled housing conditions when using physiological or behavioural measurements as a primary outcome