Nox2 underpins microvascular inflammation and vascular contributions to cognitive decline.

Chronic microvascular inflammation and oxidative stress are inter-related mechanisms underpinning white matter disease and vascular cognitive impairment (VCI). A proposed mediator is nicotinamide adenine dinucleotide phosphate (NADPH) oxidase 2 (Nox2), a major source of reactive oxygen species (ROS) in the brain. To assess the role of Nox2 in VCI, we studied a tractable model with white matter pathology and cognitive impairment induced by bilateral carotid artery stenosis (BCAS). Mice with genetic deletion of Nox2 (Nox2 KO) were compared to wild-type (WT) following BCAS. Sustained BCAS over 12 weeks in WT mice induced Nox2 expression, indices of microvascular inflammation and oxidative damage, along with white matter pathology culminating in a marked cognitive impairment, which were all protected by Nox2 genetic deletion. Neurovascular coupling was impaired in WT mice post-BCAS and restored in Nox2 KO mice. Increased vascular expression of chemoattractant mediators, cell-adhesion molecules and endothelial activation factors in WT mice post-BCAS were ameliorated by Nox2 deficiency. The clinical relevance was confirmed by increased vascular Nox2 and indices of microvascular inflammation in human post-mortem subjects with cerebral vascular disease. Our results support Nox2 activity as a critical determinant of VCI, whose targeting may be of therapeutic benefit in cerebral vascular disease.

Impaired Glymphatic Function and Pulsation Alterations in a Mouse Model of Vascular Cognitive Impairment.

Large vessel disease and carotid stenosis are key mechanisms contributing to vascular cognitive impairment (VCI) and dementia. Our previous work, and that of others, using rodent models, demonstrated that bilateral common carotid stenosis (BCAS) leads to cognitive impairment via gradual deterioration of the neuro-glial-vascular unit and accumulation of amyloid-ß (Aß) protein. Since brain-wide drainage pathways (glymphatic) for waste clearance, including Aß removal, have been implicated in the pathophysiology of VCI via glial mechanisms, we hypothesized that glymphatic function would be impaired in a BCAS model and exacerbated in the presence of Aß. Male wild-type and Tg-SwDI (model of microvascular amyloid) mice were subjected to BCAS or sham surgery which led to a reduction in cerebral perfusion and impaired spatial learning acquisition and cognitive flexibility. After 3 months survival, glymphatic function was evaluated by cerebrospinal fluid (CSF) fluorescent tracer influx. We demonstrated that BCAS caused a marked regional reduction of CSF tracer influx in the dorsolateral cortex and CA1-DG molecular layer. In parallel to these changes increased reactive astrogliosis was observed post-BCAS. To further investigate the mechanisms that may lead to these changes, we measured the pulsation of cortical vessels. BCAS impaired vascular pulsation in pial arteries in WT and Tg-SwDI mice. Our findings show that BCAS influences VCI and that this is paralleled by impaired glymphatic drainage and reduced vascular pulsation. We propose that these additional targets need to be considered when treating VCI.

Erythrocytes as Biomarkers for Dementia: Analysis of Protein Content and Alpha-Synuclein.

BACKGROUND: Discovering biomarkers for dementia is a pivotal step toward successful early diagnosis and treatment. Although plasma biomarkers have been explored, no consensus has been reached. Alpha-synuclein (AS), a 14 kDa synaptic protein associated with several neurodegenerative diseases, exists natively within erythrocytes (ERC). This protein is characteristic of Lewy body diseases, in which it aggregates into toxic Lewy bodies. As ERC are implicated in dementia, they are a potential target for future biomarkers. OBJECTIVE: The aims of this study were to assess AS levels within ERC and whether AS can be used as a peripheral biomarker to differentiate between dementia and aged matched healthy control subjects. METHODS: A total of 114 samples (60 aging controls, 36 Alzheimer's disease, 12 vascular dementia (VaD) and 6 dementia with Lewy bodies (DLB) subjects) were analyzed. We used Bradford assay to measure protein concentration, indirect ELISA to detect levels of AS, and immunoblotting to identify AS composition. Data were analyzed with nonparametric tests. RESULTS: AS oligomers were present in dementia blood samples, whereas in controls, AS was largely monomeric. There was a significant increase in AS levels in DLB whole blood (p = 0.005; Kruskal-Wallis test), with a sensitivity and specificity of 100.0% and 93.9%. Protein concentrations in ERC isolated at pH 5.7 were significantly increased in dementia patients compared to controls (17.58 versus 40.33µg/ml; p≤0.005; Mann-Whitney test). In the VaD group, the protein concentration in the pH5.7 ERC fraction had sensitivity and specificity of 91.7% and 62.1%. CONCLUSIONS: ERC protein concentration and AS levels have a potential for development of a novel diagnostic dementia blood test.

Tau proteins in the temporal and frontal cortices in patients with vascular dementia.

We previously reported that, in the brains of older patients with vascular dementia (VaD), there is a distinctive accumulation of detergent-extractable soluble amyloid-ß, with a predominance of Aß42 species. It is unclear, however, if tau proteins also accumulate in the brains of older VaD subjects. Using antibody-specific immunoassays, we assessed concentrations of total tau (t-tau) and phosphorylated tau protein, measured at 3 phosphorylated sites (i.e. Thr181, Ser202/Thr205, and Ser262), as well as synaptophysin in the temporal and frontal cortices of 18 VaD, 16 Alzheimer disease (AD), and 16 normal age-matched control subjects. There was selective loss of t-tau protein in VaD compared with controls and AD subjects (p < 0.021 and p < 0.001, respectively). In contrast, phosphorylated tau levels were similar to controls in VaD in both regions, but they were increased in the temporal lobes of patients with AD (p < 0.01 and p < 0.0001 for Ser202/Thr205 and Ser262 phosphorylated sites, respectively). The reduced t-tau in the VaD group was unrelated to any low-level neurofibrillary or amyloid pathology or age at death. These findings suggest that breaches of microvascular or microstructural tissue integrity subsequent to ischemic injury in older age may modify tau protein metabolism or phosphorylation and have effects on the burden of neurofibrillary pathology characteristic of AD.