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Background Alzheimer’s disease (AD) is a progressive, degenerative, and terminal disease without cure. There is an urgent need for a new strategy to treat AD. The aim of this study was to investigate the effects of intermittent hypoxic treatment (IHT) on cognitive functions in a mouse model of AD and unravel the mechanism of action of IHT. Methods Six-month-old APPswe/PS1dE9 (APP/PS1) male mice were exposed to hypoxic environment (14.3% O2) 4 h/day for 14 days or 28 days. Cognitive functions were measured by Morris water maze test after either 14 days or 42 days of interval. Thereafter the distribution of amyloid plaque and microglial activation were determined by mouse brain immunohistochemistry, while the amyloid beta (Aβ) and inflammatory cytokines were measured by ELISA and Western Blot. Microarray was used for studying gene expressions in the hippocampus. Results IHT for 14 days or 28 days significantly improved the spatial memory ability of the 6-month-old APP/PS1 mice. The memory improvement by 14 days IHT lasted to 14 days, but not to 42 days. The level of Aβ plaques and neurofilament accumulations was reduced markedly after the IHT exposure. IHT reduced the pro-inflammatory cytokines IL-1β, IL-6 levels, and β-secretase cleavage of APP processing which implies reduced Aβ production. Microarray analysis revealed a large number of genes in the hippocampus were significantly altered which are known to be metabolism-regulated genes. Conclusions This study provides evidence of the beneficial effect of IHT on the progression of AD by alleviating memory impairment, reducing Aβ accumulation and inflammation in the brain. IHT can be developed as a novel measure to relieve the progression of AD by targeting multiple pathways in the AD pathogenesis.

Transient or permanent loss of tissue perfusion due to ischaemic stroke can lead to damage to the neurovasculature, and disrupt brain homeostasis, causing long-term motor and cognitive deficits. Despite promising pre-clinical studies, clinically approved neuroprotective therapies are lacking. Most studies have focused on neurons while ignoring the important roles of other cells of the neurovascular unit, such as astrocytes and pericytes. Astrocytes are important for the development and maintenance of the blood–brain barrier, brain homeostasis, structural support, control of cerebral blood flow and secretion of neuroprotective factors. Emerging data suggest that astrocyte activation exerts both beneficial and detrimental effects following ischaemic stroke. Activated astrocytes provide neuroprotection and contribute to neurorestoration, but also secrete inflammatory modulators, leading to aggravation of the ischaemic lesion. Astrocytes are more resistant than other cell types to stroke pathology, and exert a regulative effect in response to ischaemia. These roles of astrocytes following ischaemic stroke remain incompletely understood, though they represent an appealing target for neurovascular protection following stroke. In this review, we summarise the astrocytic contributions to neurovascular damage and repair following ischaemic stroke, and explore mechanisms of neuroprotection that promote revascularisation and neurorestoration, which may be targeted for developing novel therapies for ischaemic stroke.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Preconditioning tissue with sublethal ischaemia or hypoxia can confer tolerance (protection) against subsequent ischaemic challenge. In vitro ischaemic preconditioning (IPC) is typically achieved through oxygen glucose deprivation (OGD), whereas hypoxic preconditioning (HPC) involves oxygen deprivation (OD) alone. Here, we report the effects of preconditioning of OGD, OD or glucose deprivation (GD) in ischaemic tolerance models with PC12 cells and primary rat neurons. PC12 cells preconditioned (4 h) with GD or OGD, but not OD, prior to reperfusion (24 h) then ischaemic challenge (OGD 6 h), showed greater mitochondrial activity, reduced cytotoxicity and decreased apoptosis, compared to sham preconditioned PC12 cells. Furthermore, 4 h preconditioning with reduced glucose (0.565 g/L, reduced from 4.5 g/L) conferred protective effects, but not for higher concentrations (1.125 or 2.25 g/L). Preconditioning (4 h) with OGD, but not OD or GD, induced stabilization of hypoxia inducible factor 1α (HIF1α) and upregulation of HIF1 downstream genes (Vegf, Glut1, Pfkfb3 and Ldha). In primary rat neurons, only OGD preconditioning (4 h) conferred neuroprotection. OGD preconditioning (4 h) induced stabilization of HIF1α and upregulation of HIF1 downstream genes (Vegf, Phd2 and Bnip3). In conclusion, OGD preconditioning (4 h) followed by 24 h reperfusion induced ischaemic tolerance (against OGD, 6 h) in both PC12 cells and primary rat neurons. The OGD preconditioning protection is associated with HIF1α stabilization and upregulation of HIF1 downstream gene expression. GD preconditioning (4 h) leads to protection in PC12 cells, but not in neurons. This GD preconditioning-induced protection was not associated with HIF1α stabilization.

This study compared effects of five hypoxia-inducible factor (HIF) prolyl hydroxylases (PHD) inhibitors on PC12 cells and primary rat neurons following oxygen-glucose deprivation (OGD). At 100 µM, the PHD inhibitors did not cause cytotoxicity and apoptosis. MTT activity was only significantly reduced by FG4592 or Bayer 85–3934 in PC12 cells. The PHD inhibitors at 100 µM significantly increased the LC3-II/LC3-I expression ratio and downregulated p62 in PC12 cells, so did FG4592 (30 µM) and DMOG (100 µM) in neurons. HIF-1α was stabilised in PC12 cells by all the PHD inhibitors at 100 µM except for DMOG, which stabilised HIF-1α at 1 and 2 mM. In primary neurons, HIF-1α was stabilised by FG4592 (30 µM) and DMOG (100 µM). Pretreatment with the PHD inhibitors 24 hours followed by 24 hour reoxygenation prior to 6 hours OGD (0.3% O2) significantly reduced LDH release and increased MTT activity compared to vehicle (1% DMSO) pretreatment. In conclusion, the PHD inhibitors stabilise HIF-1α in normoxia, induce autophagy, and protect cells from a subsequent OGD insult. The new class of PHD inhibitors (FG4592, FG2216, GSK1278863, Bay85-3934) have the higher potency than DMOG. The interplay between autophagy, HIF stabilisation and neuroprotection in ischaemic stroke merits further investigation.

Perspective article.

Editorial

Cerebrospinal fluid (CSF) circulates through the brain and has a unique composition reflecting the biological processes of the brain. Identifying ageing CSF biomarkers can aid in understanding the ageing process and interpreting CSF protein changes in neurodegenerative diseases. In this study, ovine CSF proteins from young (1–2 year old), middle aged (3–6 year old) and old (7–10 year old) sheep were systemically studied. CSF proteins were labelled with iTRAQ tagging reagents and fractionated by 2-dimensional high performance, liquid chromatography. Tryptic peptides were identified using MS/MS fragmentation ions for sequencing and quantified from iTRAQ reporter ion intensities at m/z 114, 115, 116 and 117. Two hundred thirty one peptides were detected, from which 143 proteins were identified. There were 52 proteins with >25% increase in concentrations in the old sheep compared to the young. 33 of them increased >25% but 50% but 1 fold [i.e. haptoglobin (Hp), haemoglobin, neuroendocrine protein 7B2, IgM, fibrous sheath interacting protein 1, vimentin]. There were 18 proteins with >25% decrease in concentrations in the old sheep compared to the young. 17 of them decreased >25% but

Autophagy is a highly regulated, catabolic process, through which macromolecules such as damaged organelles and proteins are degraded and recycled to maintain cellular homeostasis. Various studies have shown the role of autophagy activation in the brain cells such as astrocytes, microglia, neurons and capillary endothelial cells upon an ischaemic insult. The underlying mechanism and the role of autophagy in ischaemic stroke, however, are yet to be fully elucidated. Recent studies have suggested that insufficient or excessive autophagy results in nerve damage and cell death whereas mild/moderate autophagy has a neuroprotective effect. It has been proposed that autophagy may be a therapeutic target in stroke treatment; however, there is a lot of debate as to whether induction or diminution of autophagy plays a role in neuronal survival after cerebral ischaemia and indicate that it has a dual role depending on the time of induction of autophagy. This review has summarized the role of autophagy in ischaemic stroke and explored effects of pharmacological autophagy modulators in ischaemic stroke treatment. Further studies are needed for translating the potential therapeutic approach in stroke treatment aiming at characterizing the timing, amount and specificity of the autophagy modulation.

High altitude cerebral edema (HACE) is a life-threatening illness that develops during the rapid ascent to high altitudes, but its underlying mechanisms remain unclear. Growing evidence has implicated inflammation in the susceptibility to and development of brain edema. In the present study, we investigated the inflammatory response and its roles in HACE in mice following high altitude hypoxic injury. We report that acute hypobaric hypoxia induced a slight inflammatory response or brain edema within 24 h in mice. However, the lipopolysaccharide (LPS)-induced systemic inflammatory response rapidly aggravated brain edema upon acute hypobaric hypoxia exposure by disrupting blood-brain barrier integrity and activating microglia, increasing water permeability via the accumulation of aquaporin-4 (AQP4), and eventually leading to impaired cognitive and motor function. These findings demonstrate that hypoxia augments LPS-induced inflammation and induces the occurrence and development of cerebral edema in mice at high altitude. Here, we provide new information on the impact of systemic inflammation on the susceptibility to and outcomes of HACE.

Knee pain is commonly seen in orthopedic and rehabilitation outpatient clinical settings, and in the aging population. Bursitis of the knee joint, especially when the volume of the synovial fluid is large enough, can compress and distend the nearby soft tissues, causing pain in the knee joint. Out of all the bursae surrounding the knee joint, supra-patellar bursitis is most often associated with knee pain. Treatment strategies in managing supra-patellar bursitis include the aspiration of joint synovial fluid and then followed by steroid injection into the bursa. When supra-patellar bursitis is caused by degenerative disorders, the concept of viscosupplementation treatment may be effective by injecting hyaluronic acid into the bursa. However, the rheology or the changes in the concentrations of proteins (biomarkers) that are related to the development of bursitis in the synovial fluid is virtually unexplored. Therefore, this study aimed to identify the concentration changes in the synovial fluid total protein amount and individual proteins associated with supra-patellar bursitis using the Bradford protein assay and western immunoglobulin methods. A total of 20 patients were divided into two groups with 10 patients in each group. One group received the high molecular weight hyaluronic acid product of Synvisc Hylan G-F 20 and the other group received the low molecular weight hyaluronic acid product of Hya-Joint Synovial Fluid Supplement once per week injection into the bursa for a total of 3weeks. Significant decreases in the synovial fluid total protein concentrations were observed after the second dosage of high molecular weight hyaluronic acid injections. Apolipoprotein A-I, interleukin 1 beta, alpha 1 antitrypsin, and matrix metalloproteinase 1 proteins revealed a trend of decreasing western immunoblotting band densities after hyaluronic acid injections. The decreases in apolipoprotein A-I and interleukin 1 beta protein band densities were significant in the high molecular weight hyaluronic acid injection group. Transthyretin, complement 5, and matrilin 3 proteins revealed a trend of increasing western immunoblotting band densities after hyaluronic acid injections. Transthyretin revealed significant increases in protein band densities in both the high and low molecular weight hyaluronic acid injection groups. This study may provide the rationale for targeting several biomarkers associated with lipid transport, inflammation, and anti-aging as possible disease modifying therapies for the treatment of supra-patellar bursitis and even degenerative joint disorders.

BackgroundDeterminants for undetected dementia and late-life depression have been not well studied.AimsTo investigate risk factors for undetected dementia and depression in older communities.MethodUsing the method of the 10/66 algorithm, we interviewed a random sample of 7072 participants aged ⩽60 years in six provinces of China during 2007–2011. We documented doctor-diagnosed dementia and depression in the interview. Using the validated 10/66 algorithm we diagnosed dementia (n= 359) and depression (n= 328).ResultsWe found that 93.1% of dementia and 92.5% of depression was undetected. Both undetected dementia and depression were significantly associated with low levels of education and occupation, and living in a rural area. The risk of undetected dementia was also associated with ‘help available when needed‘, and inversely, with a family history of mental illness and having functional impairment. Undetected depression was significantly related to female gender, low income, having more children and inversely with having heart disease.ConclusionsOlder adults in China have high levels of undetected dementia and depression. General socioeconomic improvement, associated with mental health education, targeting high-risk populations are likely to increase detection of dementia and depression in older adults, providing a backdrop for culturally acceptable service development.

Previous attempts to identify neuroprotective targets by studying the ischemic cascade and devising ways to suppress it have failed to translate to efficacious therapies for acute ischemic stroke. We hypothesized that studying the molecular determinants of endogenous neuroprotection in two well-established paradigms, the resistance of CA3 hippocampal neurons to global ischemia and the tolerance conferred by ischemic preconditioning (IPC), would reveal new neuroprotective targets. We found that the product of the tuberous sclerosis complex 1 gene (TSC1), hamartin, is selectively induced by ischemia in hippocampal CA3 neurons. In CA1 neurons, hamartin was unaffected by ischemia but was upregulated by IPC preceding ischemia, which protects the otherwise vulnerable CA1 cells. Suppression of hamartin expression with TSC1 shRNA viral vectors both in vitro and in vivo increased the vulnerability of neurons to cell death following oxygen glucose deprivation (OGD) and ischemia. In vivo, suppression of TSC1 expression increased locomotor activity and decreased habituation in a hippocampal-dependent task. Overexpression of hamartin increased resistance to OGD by inducing productive autophagy through an mTORC1-dependent mechanism. Copyright © 2013 Nature America, Inc.

Dimethyloxalylglycine (DMOG) is an inhibitor of prolyl-4-hydroxylase domain (PHD) enzymes that regulate the stability of hypoxia-inducible factor (HIF). We investigated the effect of DMOG on the outcome after permanent and transient middle cerebral artery occlusion (p/tMCAO) in the rat. Before and after pMCAO, rats were treated with 40 mg/kg, 200 mg/kg DMOG, or vehicle, and with 40 mg/kg or vehicle after tMCAO. Serial magnetic resonance imaging (MRI) was performed to assess infarct evolution and regional cerebral blood flow (rCBF). Both doses significantly reduced infarct volumes, but only 40 mg/kg improved the behavior after 24 hours of pMCAO. Animals receiving 40 mg/kg were more likely to maintain rCBF values above 30% from the contralateral hemisphere within 24 hours of pMCAO. DMOG after tMCAO significantly reduced the infarct volumes and improved behavior at 24 hours and 8 days and also improved the rCBF after 24 hours. A consistent and significant upregulation of both mRNA and protein levels of vascular endothelial growth factor (VEGF) and endothelial nitric oxide synthase (eNOS) was associated with the observed neuroprotection, although this was not consistently related to HIF-1α levels at 24 hours and 8 days. Thus, DMOG afforded neuroprotection both at 24 hours after pMCAO and at 24 hours and 8 days after tMCAO. This effect was associated with an increase of VEGF and eNOS and was mediated by improved rCBF after DMOG treatment. © 2011 ISCBFM All rights reserved.

Stroke mostly occurs in elderly people and patient outcomes after stroke are highly influenced by age. A better understanding of the causes of stroke in the elderly might have important practical implications not only for clinical management, but also for preventive strategies and future health-care policies. In this Review, we explore the evidence from both human and animal studies relating to the effect of old age-in terms of susceptibility, patient outcomes and response to treatment-on ischemic stroke. Several aging-related changes in the brain have been identified that are associated with an increase in vulnerability to ischemic stroke in the elderly. Furthermore, risk factor profiles for stroke and mechanisms of ischemic injury differ between young and elderly patients. Elderly patients with ischemic stroke often receive less-effective treatment and have poorer outcomes than younger individuals who develop this condition. Neuroprotective agents for ischemic stroke have been sought for decades but none has proved effective in humans. One contributing factor for this translational failure is that most preclinical studies have used young animals. Future research on ischemic stroke should consider age as a factor that influences stroke prevention and treatment, and should focus on the management of acute stroke in the elderly to reduce the incidence and improve outcomes in this vulnerable group.

This study investigated the effects of 2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetic acid (IOX3), a selective small molecule inhibitor of hypoxia-inducible factor (HIF) prolyl hydroxylases, on mouse brains subject to transient focal cerebral ischaemia. Male, 8- to 12-week-old C57/B6 mice were subjected to 45 min of middle cerebral artery occlusion (MCAO) either immediately or 24 h after receiving IOX3. Mice receiving IOX3 at 20 mg/kg 24 h prior to the MCAO had better neuroscores and smaller blood-brain barrier (BBB) disruption and infarct volumes than mice receiving the vehicle, whereas those having IOX3 at 60 mg/kg showed no significant changes. IOX3 treatment immediately before MCAO was not neuroprotective. IOX3 up-regulated HIF-1α, and increased EPO expression in mouse brains. In an in vitro BBB model (RBE4 cell line), IOX3 up-regulated HIF-1α and delocalized ZO-1. Pre-treating IOX3 on RBE4 cells 24 h before oxygen-glucose deprivation had a protective effect on endothelial barrier preservation with ZO-1 being better localized, while immediate IOX3 treatment did not. Our study suggests that HIF stabilization with IOX3 before cerebral ischaemia is neuroprotective partially because of BBB protection, while immediate application could be detrimental. These results provide information for studies aimed at the therapeutic activation of HIF pathway for neurovascular protection from cerebral ischaemia. We show that IOX3, a selective small molecule (280.66 Da) HIF prolyl hydroxylase inhibitor, could up-regulate HIF-1α and increase erythropoietin expression in mice. We further demonstrate that HIF stabilization with IOX3 before cerebral ischaemia is neuroprotective partially because of blood-brain barrier (BBB) protection, while immediate application is detrimental both in vivo and in vitro. These findings provide new insights into the role of HIF stabilization in ischaemic stroke.

Despite advances in the diagnosis and treatment of acute ischaemic stroke in the past two decades, stroke has remained the third cause of mortality and the single leading cause of disability worldwide. The immediate goal of acute ischaemic stroke therapy is to salvage the ischaemic penumbra through recanalisation of the occluded cerebral blood vessel. This is currently achieved through thrombolytics, which are pharmacological agents that can break up a clot blocking the flow of blood. To date, the only approved thrombolytic for treatment of acute ischaemic stroke is recombinant tissue plasminogen activator (alteplase, rt-PA), however, alteplase is substantially underused because of concerns regarding adverse bleeding risk. This limitation has fuelled the search for other thrombolytic agents, which display greater fibrin dependence and selectivity, but lack detrimental effects within the central nervous system. Development of alternative fibrinolytic agents that might be easier and safer to administer could lead to wider acceptance and use of thrombolytic therapy for stroke. Although other thrombolytic agents (e.g. streptokinase) have failed to show benefit over alteplase, there is still on-going research in search of alternative agents with higher target specificity and better safety profile. The potential thrombolytic agents with trials in progress include desmoteplase, tenecteplase, reteplase, plasmin and microplasmin. This review summarises current therapies with thrombolytics (e.g. alteplase and urokinase), their limitations and side effects, and also discusses ongoing clinical studies with the various potential emerging thrombolytic agents.

This study investigated the function of each of the hypoxia inducible factor (HIF) prolyl-4-hydroxylase enzymes (PHD1–3) in the first 24 h following transient focal cerebral ischaemia by using mice with each isoform genetically suppressed. Male, 8- to 12-week old PHD1−/−, PHD2+/− and PHD3−/− mice and their wild-type (WT) littermate were subjected to 45 min of middle cerebral artery occlusion (MCAO). During the experiments, regional cerebral blood flow (rCBF) was recorded by laser Doppler flowmetry. Behaviour was assessed at both 2 h and 24 h after reperfusion with a common neuroscore. Infarct volumes, blood–brain barrier (BBB) disruption, cerebral vascular density, apoptosis, reactive oxygen species (ROS), HIF1α, and glycogen levels were then determined using histological and immunohistochemical techniques. When compared to their WT littermates, PHD2+/− mice had significantly increased cerebral microvascular density and more effective restoration of CBF upon reperfusion. PHD2+/− mice showed significantly better functional outcomes and higher activity rates at both 2 h and 24 h after MCAO, associated with significant fewer apoptotic cells in the penumbra and less BBB disruption; PHD3−/− mice had impaired rCBF upon early reperfusion but comparable functional outcomes; PHD1−/− mice did not show any significant changes following the MCAO. Production of ROS, HIF1α staining and glycogen content in the brain were not different in any comparison. Life-long genetic inhibition of PHD enzymes produces different effects on outcome in the first 24 h after transient cerebral ischaemia. These need to be considered in optimizing therapeutic effects of PHD inhibitors, particularly when isoform specific inhibitors become available.

Studies have shown that ageing alone can cause increases in the concentrations of many cerebrospinal fluid (CSF) proteins. Therefore, CSF protein concentrations must be interpreted with caution before concluding that the increased concentrations of certain proteins can be used as disease-specific biomarkers. Age-related reduction in CSF turnover has been shown to have a significant concentrating effect on CSF proteins from young to old. As a result, CSF protein concentrations need to be corrected with age-specific turnovers first before performing any data comparisons between different ages. This study applied the concept of CSF/plasma concentration ratios of plasma-derived proteins that is frequently used in the investigation of brain barrier integrity to calculate the amount of protein that enters the CSF from the plasma side in different age groups. Based on our calculations, proteins with molecular weights greater than 91.92 kDa for the young, 109.51 kDa for the middle-aged and 120 kDa for the old should not be able to cross the brain barriers of the blood-brain and blood-CSF barriers to enter the CSF from the plasma side. For proteins that can be derived from the choroid plexus (CP), brain, and plasma, the amount that crosses the barriers to enter the CSF from the plasma side will contribute to their measured total protein concentrations in the CSF. CP and brain production of these proteins can be calculated when turnover corrected CSF protein concentrations are further corrected by the amount of protein that crosses the barriers. In this study, CP and brain produced concentrations of transthyretin, retinol binding protein, alpha-1-antitrypsin, gelsolin, and lactotransferrin were calculated. The production of these proteins decreased with age with alpha-1-antitrypsin protein revealing the most substantial decrease of 86% from young (0.14±0.01 mg·dL(-1)) to old (0.02 mg·dL(-1)). In conclusion, measured CSF protein concentrations for proteins that can be derived from the CP, brain, and plasma need to be corrected by age-specific CSF turnovers and by the amount of protein that crosses the brain barriers first before their concentrations can be compared logically between different ages.

Background Previous studies indicated overall relatively low prevalence of dementia in older people in China, which may be biased by studied samples or methods. We determined the prevalence of dementia cases and subcases in China and examined their socio-economic correlates. Methods Using the Geriatric Mental State interview, we examined random samples of 2917 participants aged ≥ 65 years in urban and rural Anhui, China in 2001–2003, and 3327 in four other provinces in 2008–2009. Dementia cases and subcases were diagnosed by Geriatric Mental State-Automated Geriatric Examination for Computer Assisted Taxonomy. Results Age-standardised prevalence for cases and subcases of dementia in the Anhui elders was 7.20% (95%CI 6.29%–8.20%) and 10.5% (9.38%–11.6%), and in the four provinces, 9.86% (8.80%–10.9%) and 8.51% (7.51%–9.52%). The matched figures among the participants who were literate were 3.05% (2.08%–4.02%) and 10.0% (8.38%–11.6%), and 4.92% (3.89%–5.96%) and 6.76% (5.55%–7.96%), respectively. There were higher prevalence rates of dementia cases and subcases in the rural elders than in the urban. Both the Anhui and four-province studies showed an obvious association of dementia with higher and lower incomes among elders who had lower educational levels or had the lowest occupational class. The highest risk of dementia was found in those who were illiterate but had the highest income or had the job of business/nonmanual labouring. Conclusions People in China have a higher prevalence of dementia than previously reported. Its U-shaped relationship with income and the excess subcases prevalence predicates a significant burden of disease, both now and for the future, suggesting preventive strategy for dementia in China. Copyright © 2011 John Wiley & Sons, Ltd.

The best neuroprotectant for acute ischaemic stroke would always be the rapid return of oxygen and glucose to physiological levels. This is currently provided by thrombolysis which restores blood flow to the ischaemic region. The attempt to confer neuroprotection by targeting the brain parenchyma has shown promise in experimental stroke models, but has unequivocally failed to translate to the clinic. Neuroprotective therapy primarily targets the biochemical cascade that produces cell death following cerebral ischaemia. However, these agents may also alter signal transduction that controls cerebral blood flow, for example glutamate, which may affect the outcome after ischaemia. In these cases, neuroprotection may potentially be due to the improved access to oxygen and glucose rather than biochemical prevention of cell death. Improvement in cerebral blood flow is an important but often overlooked effect of neuroprotective therapy, analogous to the protective effects of drug-induced hypothermia. This short review will discuss cerebral blood flow alteration and protection of the brain in the context of ischaemic preconditioning, oxygen sensing and thrombolysis. Future neuroprotection studies in cerebral ischaemia require stringent monitoring of cerebral blood flow, plus other physiological parameters. This will increase the chances that any protection observed may be able to translate to human therapy.

Studies have shown that ageing and several neurological diseases of the central nervous system are often accompanied with increase in concentrations of many cerebrospinal fluid (CSF) proteins. However, few studies have actually looked into the mechanisms behind the increase in CSF protein concentrations. In this study, CSF secretion rates and turnovers were measured using the in situ perfused choroid plexus (CP) technique in a group of sheep between 1 and 10 years of age. CSF protein concentrations were determined using quantitative proteomic techniques. CSF turnover in hours correlated significantly with age, changing from 10.5+/-2.7h in the young to 17.1+/-2.4h in the old. The amount of CSF replaced per hour decreased from 2.46+/-0.42mL in the young to 1.17+/-0.16mL in the old. The age-related reduction in CSF turnover was calculated to have a concentrating effect of approximately 1.32 times in middle-aged and 2.10 times in old CSF proteins. After CSF turnover normalization, CSF albumin (a plasma-derived protein) concentration still increased significantly with age; however, both brain-derived and partially brain-derived protein concentrations in the CSF decreased with age after normalization. Regression analysis between turnovers and albumin concentrations has shown that reduced CSF turnover was the cause of increased CSF albumin concentrations with age. Therefore, CSF protein concentrations should be normalized according to their age-specific turnovers first before their concentrations can be compared logically between different ages.

Dysfunction of the choroid plexuses (CPs) and the blood-cerebrospinal fluid barrier (BCSFB) might contribute to age-related cognitive decline and neurodegenerative disease. We used the CPs from young (1-2 years), middle-aged (3-6 years) and old (7-10 years) sheep to explore effects of aging on various aspects of CP and BCSFB functions. Total protein in the cerebrospinal fluid (CSF) was significantly higher in old compared to young sheep and CSF secretion by the CP perfused in situ was significantly lower in both old and middle-aged when compared to young sheep, which correlated with reduced (22)Na(+) uptake and efflux by the CP. Steady-state extractions of a low and medium size molecular weight extracellular space marker, (14)C-mannitol and (3)H-polyethylene glycol, respectively, were significantly higher in CPs from old compared to young animals; however, there was no significant difference in steady-state extraction of a high molecular weight marker, (125)I-bovine serum albumin. This indicates increased passive BCSFB permeability for small and medium sized molecules in old sheep. CP redox activity was significantly lower in the old animals as assessed by the MTT assay, however, there was no significant difference in ATP content and energy charge of the CP with age suggesting adequate baseline energy reserve capacity. These data indicate that normal aging processes alter protein content in the CSF, CSF secretion, integrity of the BCSFB and Na(+) flux in the epithelial layer, which could impact on CSF homeostasis and turnover.

Patients after repair of tetralogy of Fallot are at increased risk of arrhythmic death. Clinical data suggest that pulmonary regurgitation predisposes to these arrhythmias, although the cellular electrophysiologic effects of pulmonary regurgitation are unknown. We induced pulmonary regurgitation in lambs, and 3 mo later, having quantified the pulmonary regurgitant (PR) fraction, studied right ventricular mechanical and electrophysiologic properties in vivo and in vitro. The PR fraction was greater in PR (75 +/- 10%) than in sham-operated animals (8 +/- 4%; p0.01). In vivo, monophasic action potential duration and activation time, at rest and during acute right ventricular stretch, were similar in both groups. However, the dispersion of activation time was greater in PR animals at rest (13 +/- 1.1 versus 8 +/- 1.1 ms; p0.05). Furthermore, the dispersion of activation increased during right ventricular stretch in PR, but not in sham-operated animals. In vitro, myocardial force-frequency responses were similar in both groups, indicating preserved systolic performance, but mechanical restitution studies showed a prolonged refractory period (447 +/- 22 versus 370 +/- 26 ms; p0.05) and a decreased recovery time constant (184 +/- 19 versus 265 +/- 20 ms; p0.001) in PR animals, indicating altered calcium cycling. Furthermore, the myocardial conduction velocity was reduced in PR animals (31 +/- 3.58 versus 47.9 +/- 5.1 cm/s; p0.01), resulting from a 2-fold increase in intracellular resistance (437.25 +/- 125.93 versus 194 +/- 43.27 Omega. cm; p = 0.025). Chronic PR leads to inhomogeneity of right ventricular activation, alters myocardial calcium cycling, reduces conduction velocity, and increases intracellular resistivity. These may contribute to the development of arrhythmias associated with PR, including those in patients after tetralogy repair.

Association of passive smoking with cognitive impairment in older adults is unclear. We carried out a systematic literature review and a new study to determine the association. There were 3 cross-sectional studies published, showing a significant association of passive smoking with cognitive impairment (a relative risk (RR) of about 1.30-1.90). In the new cohort study, we interviewed 1081 never-smoking participants aged ≥ 65 years in China using a standard method of the Geriatric Mental State–Automated Geriatric Examination for Computer Assisted Taxonomy and found a significant association with dose response; multivariate adjusted RR was 1.02 (95% confidence interval 0.67-1.55) in >0 to 49 exposure level years of passive smoking, 1.57 (1.00-2.47) in 50 to 99, and 2.12 (1.24-3.63) in ≥100, trend P = .008. The relationship seems not to be a reverse causality of the effect. Passive smoking could be considered an important risk factor for cognitive impairment in older adults. Avoiding exposure to passive smoking would help to preserve cognitive decline in later life.

The knowledge of brain syndromes is essential for stroke physicians and neurologists, particularly those that can be extremely difficult and challenging to diagnose due to the great variability of symptom presentation and yet of clinical significance in terms of potential devastating effect with poor outcome. The diagnosis and understanding of stroke syndromes has improved dramatically over the years with the advent of modern imaging, while the management is similar to general care as recommended by various guidelines in addition to care of such patients on specialized units with facilities for continuous monitoring of vital signs and dedicated stroke therapy. Such critical care can be provided either in the acute stroke unit, the medical intensive care unit or the neurological intensive care unit. There may be no definitive treatment at reversing stroke syndromes, but it is important to identify the signs and symptoms for an early diagnosis to prompt quick treatment, which can prevent further devastating complications following stroke. The aim of this article is to discuss some of the important clinical stroke syndromes encountered in clinical practice and discuss their management.

No pharmacological intervention has been shown convincingly to improve neurological outcome in stroke patients after the brain tissue is infarcted. While conventional therapeutic strategies focus on preventing brain damage, stem cell treatment has the potential to repair the injured brain tissue. Stem cells not only produce a source of trophic molecules to minimize brain damage caused by ischaemia/reperfusion and promote recovery, but also potentially turn to new cells to replace those lost in ischaemic core. Although preclinical studies have shown promise, stem cell therapy for stroke treatment in human is still at an early stage and it is difficult to draw conclusions from current clinical trials about the efficacy of the different treatments used in humans. This article reviews the potential of various types of stem cells, from embryonic to adult to induced pluripotent stem cells, in stroke therapy, highlights new evidence from the ongoing clinical trials and discusses some of the problems associated with translating stem cell technology to a clinical therapy for stroke.

Albumin is the most abundant protein in both CSF and plasma, and albumin quotient is often used to assess the functions of brain barriers especially that of the blood-CSF barrier [i.e. the choroid plexus (CP) which also secretes CSF]. In this study, we took albumin as a model molecule to investigate ageing-related alterations in the CSF-CP system in sheep. We found significant ageing-related increases in the weight of lateral CP [122.4 +/- 14.0 mg in the young, 198.6 +/- 35.4 mg in the middle aged, 286.1 +/- 25.1 mg in the old (p < 0.05)], in the CSF albumin as well as the albumin quotient. Albumin protein spots in old CSF displayed wider on 2D western immunoblotting images, and had higher densities on images of 2D large gels stained with Pro-Q Emerald 488 compared to the young samples, suggesting ageing-related post-translational modification in the albumin. CSF secretion was reduced with age: 0.148 +/- 0.013 mL/min/g in the young, 0.092 +/- 0.02 mL/min/g in the middle aged, 0.070 +/- 0.013 mL/min/g in the old (p < 0.05). The (125)I-BSA extraction was not different among the sheep groups, nor was altered by temperature reduction, monensin, nocodazole, anti-transforming growth factor beta receptor II antibody, as well as unlabelled albumins. In conclusion, elevation of albumin in old CSF is associated with reduced CSF secretion by the CP, which size increases with age. (125)I-BSA extract, reflecting the extracellular space rather than the active albumin uptake in the CP, is not different between ages. These early changes in health ageing may result in the accumulation and modifications of CSF proteins leading to neurotoxicity.

The pathogenesis of stroke is multifactorial, and inflammation is thought to have a critical function in lesion progression at early time points. Detection of inflammatory processes associated with cerebral ischemia would be greatly beneficial in both designing individual therapeutic strategies and monitoring outcome. We have recently developed a new approach to imaging components of the inflammatory response, namely endovascular adhesion molecule expression on the brain endothelium. In this study, we show specific imaging of vascular cell adhesion molecule (VCAM)-1 expression in a mouse model of middle cerebral artery occlusion (MCAO), and a reduction in this inflammatory response, associated with improved behavioral outcome, as a result of preconditioning. The spatial extent of VCAM-1 expression is considerably greater than the detectable lesion using diffusion-weighted imaging (25% versus 3% total brain volume), which is generally taken to reflect the core of the lesion at early time points. Thus, VCAM-1 imaging seems to reveal both core and penumbral regions, and our data implicate VCAM-1 upregulation and associated inflammatory processes in the progression of penumbral tissue to infarction. Our findings indicate that such molecular magnetic resonance imaging (MRI) approaches could be important clinical tools for patient evaluation, acute monitoring of therapy, and design of specific treatment strategies.

Prostaglandin D2 synthase (PGDS) is a glycoprotein that is exclusively brain derived and is one of the most abundant proteins in the cerebrospinal fluid (CSF). Due to its high CSF specificity, it can be used as a tool for the diagnosis of central nervous system (CNS) disorders. However, several studies have yielded contradictory CSF PGDS concentrations in various CNS neurodegenerative disorders. Sheep CSF samples from different ages were used in this study and 2-dimensional electrophoresis (2-DE) was applied in PGDS identification and concentration calculation. SYPRO Ruby Protein Gel Stain was the staining method used to stain the 2-DE gel protein spots. Pro-Q Emerald 488 Staining for Glycoproteins was used for the staining of glycoproteins. A total of nine PGDS isoforms were identified and CSF total PGDS concentration was calculated to increase linearly by 44% from young (0.9323 ± 0.0637 mg dL−1) to old (1.3669 ± 0.0558 mg dL−1). However, the proportion of CSF total PGDS as a percentage of CSF total protein was discovered to decrease exponentially with age. This was due to the influence of larger age-related increase in CSF albumin concentration (>200% from young to old) as albumin is the most abundant protein in the CSF (>60% of total CSF proteins). Active deglycosylation was not observed in PGDS isoforms during healthy ageing. Some PGDS isoforms were observed to have age-related increase in glycation. These findings suggest that CSF PGDS concentration is increased during healthy ageing and must be taken into consideration when using PGDS as a potential biomarker in diagnosing CNS neurodegenerative disorders. Whether age-related increase in the glycation of some CSF PGDS isoforms will result in detrimental effects on the PGDS protein function needs further investigations.

This study investigated the transfer of T4 from cerebrospinal fluid (CSF) into the choroid plexuses (CP) and ventricular brain regions, and the role of P-glycoprotein (P-gp), multidrug resistance protein 1 (mrp1) and organic anion transporting polypeptides (oatps). During in vivo ventriculo-cisternal (V-C) perfusion in the anesthetized rabbit (meditomidine hydrochloride 0.5 mg kg(-1), pentobarbitone 10 mg kg(-1) i.v.), 125I-T4 was perfused continuously into ventricular CSF with reference molecules 14C-mannitol and blue dextran. Over 2 h, 36.9+/-4.6% 125I-T4 was recovered in cisternal CSF. Addition of P-gp substrate verapamil increased CSF 125I-T4 recovery to 51.4+/-2.8%, although mrp1 and oatp substrates had no significant effect. In brain, 125I-T4 showed greatest accumulation in the CP (1.52+/-0.31 ml g(-1)), followed by ventricular regions (caudate putamen, ependyma, hippocampus, 0.05-0.14 ml g(-1)). At the CP, verapamil and probenecid (but not indomethacin) significantly increased 125I-T4 accumulation, implicating a role for P-gp and oatps. Of other brain regions, all three drugs increased accumulation in caudate putamen 3-5 times, and indomethacin and probenecid increased accumulation in ependyma 4-5 times. The role of P-gp was investigated further in isolated incubated CPs using 5 microg/ml C219 anti-P-gp antibody. Both 125I-T4 and 3H-cyclosporin accumulation increased by 80%, suggesting that P-gp is functional in the CP and has a role in removal of T4. Combined with the in vivo results, these studies suggest that P-gp provides a local homeostatic mechanism, maintaining CSF T4 levels. We conclude that P-gp and oatps contribute to the transfer of 125I-T4 between the CSF, CP and brain, hence regulating 125I-T4 availability in CSF.

Objective To assess the association of socioeconomic deprivation (SED) with survival after stroke in China. Design A systematic literature review and a new population-based cohort study. Setting and participants In urban and rural communities in Anhui, China, 2978 residents aged ≥60 years took part in baseline investigation and were followed up for 5 years; five published studies were identified for a systematic review. Primary and secondary outcome measures 167 of 2978 participants (5.6%) had doctor-diagnosed stroke at baseline or 1 year later. All-cause mortality in the follow-up. Results In the Anhui cohort follow-up of 167 patients with stroke, 64 (38.3%) died. Multivariate adjusted hazard ratio (HR) of mortality in patients with educational level of less than or equal to primary school was 1.88 (95% CI 1.05 to 3.36) compared to those having more than primary school education. Increased HR of mortality in patients living in a rural area was at borderline significant (1.64, 0.97 to 2.78), but the HR in patients with lower levels of occupation and income was not significant. Published studies showed a significant increase in stroke mortality in relation to some SED indicators. Pooled relative risk (RR) of mortality in patients with low education was 3.07 (1.27 to 7.34), in patients with low income 1.58 (1.50 to 1.65) and in patients living in rural areas 1.47 (1.37 to 1.58). Conclusions The evidence suggests the presence of a mortality gradient after stroke for material as well as social forms of deprivation in China. Inequalities in survival after stroke persist and need to be taken into account when implementing intervention programmes.

Biomarkers for neurodegenerative disorders are potentially present in cerebrospinal fluid (CSF) and can be detected using proteomic technologies. Since CSF is high in salt and low in protein, its study by proteomic methods requires appropriate sample preparation. In this study, we applied four different sample treatments to the same ovine CSF sample. Precipitation with acetone or using a 2-D Clean-Up Kit (GE Healthcare BioSciences, Little Chalfont, UK) preserved more proteins, and produced more gel spots than spin columns from Sigma and Bio-Rad. A 53-kDa spot, identified by MS/MS as transthyretin (TTR) tetramer, was not detected in samples treated with the 2-D Clean-Up Kit, though it was always present on all gels prepared using the other three methods. Western immunoblotting confirmed the low recovery of tetrameric TTR by the 2-D Clean-Up Kit and showed that the tetrameric form of TTR predominated in ovine but not in rat CSF. In one ovine CSF sample haemoglobin was found, indicating blood contamination. We conclude that acetone precipitation is a simple and efficient way to prepare ovine CSF for 2-DE. The use of the 2-D Clean-Up Kit leads to the disappearance of tetrameric TTR only from ovine CSF proteome.