Imaging cerebrovascular alterations in experimental models of ageing and vascular cognitive impairment

Vascular cognitive impairment describes a heterogeneous condition in which cognitive decline is precipitated by underlying cerebrovascular dysfunction. Ageing, as well as vascular diseases such as hypertension, stroke, cerebral small vessel disease and cerebral amyloid angiopathy, are risk factors for vascular cognitive impairment. The precise mechanisms by which these conditions impact the cerebral vasculature to drive cognitive decline, however, are unknown. Previous research has indicated that vascular risk factors can lead to microvascular oxidative stress, inflammation and endothelial dysfunction that can lead to tissue hypoperfusion, the development of white and grey matter vascular lesions (microinfarcts and microbleeds) and cognitive impairment. It was hypothesised that ageing, a prominent risk factor for cognitive decline, would induce impairments on neurovascular coupling resulting from neurovascular unit disruption. It was further hypothesised that induction of chronic cerebral hypoperfusion would mediate neurovascular dysfunction and vascular lesion development through increased oxidative stress, resulting in cognitive decline. Finally, it was also hypothesised that neurovascular impairments resulting from ageing and chronic cerebral hypoperfusion would be exacerbated in the presence of amyloid deposition. Four studies were performed in order to test these hypotheses. Vascular risk factors can be reproduced using experimental mouse models and provide a valuable basis in which to test hypotheses and therapeutic interventions. As such, a primary aim of this thesis was to develop and validate sensitive MRI approaches that would allow the detection of vascular alterations in vivo. In the first series of studies, MRI techniques to assess resting cerebral blood flow, vessel number, vascular lesions and inflammation in experimental mice were validated using established in vivo and ex vivo techniques, so that these techniques could be used in subsequent studies for vascular assessments in vivo. Arterial spin labelling was developed to assess resting cerebral blood flow, and was able to detect reductions in blood flow following cerebral hypoperfusion that correlated well with those obtained from laser speckle imaging. Q-map imaging was able to detect reductions in vessel number in acute lesions, and in non-lesioned mice measures of vessel number correlated well with histopathological measures. Structural T2 imaging was performed in order to detect ischaemic and haemorrhagic lesions in chronically hypoperfused mice, and was validated using H&E and Perls’ staining. Finally, contrast-enhanced T2* imaging was used to detect iron oxide uptake by macrophages in the brains of hypoperfused mice, which was further validated by the identification of iron-containing macrophages in immunostained brain sections. The second study was conducted to test the hypothesis that ageing would impair neurovascular unit function and structure, and that these impairments would be exacerbated in the presence of amyloid pathology. The aim of the study was to incorporate previously developed in vivo imaging approaches in the assessment of vascular function and alterations in neurovascular unit structure in both wild type and TgSwDI mice. As predicted, ageing caused a pronounced deficit on measures of neurovascular coupling, however this was not exacerbated by accumulation of amyloid in TgSwDI mice and was not associated with alterations in baseline blood flow measured by arterial spin labelling. Structural assessment of the neurovascular unit revealed a loss of contact between astrocytic endfeet and vasculature, which was significantly associated with the impairment on neurovascular coupling, in addition to other markers of breakdown of the neurovascular unit such as loss of pericyte coverage and microglial activation. Age and thalamic vascular amyloid accumulation were also associated with an increase in the NADPH oxidase (NOX) subunit p47, indicative of increased oxidative stress. Data from this experiment indicate that ageing can profoundly impair neurovascular coupling, mediated by gliosis and loss of astrocytic contacts with vasculature. The third study aimed to test the hypothesis that chronic cerebral hypoperfusion (a prominent early feature of vascular cognitive impairment) would impair vascular function and induce the development of vascular lesions and cognitive decline. The impact of hypoperfusion on neurovascular coupling, ischaemic and haemorrhagic lesion burden and cognition was investigated in wild type and TgSwDI mice. Hypoperfusion induced deficits on neurovascular coupling, increased lesion burden and inflammation assessed with T2 and contrast-enhanced T2* imaging, and caused impairment on measures of learning and memory. Hypoperfusion was also associated with an increase in the levels of NOX2, NOX4 and 3-NT at 3 months following surgery, indicating persistent reactive oxygen species production and oxidative damage in hypoperfused mice. The findings from this study indicate that vascular dysfunction and cognitive impairment following hypoperfusion may be mediated by increased NADPH oxidase activity and resulting oxidative stress. The previous studies indicated that markers of oxidative stress were induced in response to ageing, vascular amyloid accumulation and cerebral hypoperfusion. The final study sought to determine whether increased NOX activity mediates downstream pathological effects on vascular function, vascular lesion development and cognitive decline following hypoperfusion. NOX activity was inhibited pharmacologically by administration of apocynin to hypoperfused TgSwDI mice for 3 months following surgery. Treatment with apocynin significantly restored neurovascular coupling to a level similar to sham-operated mice, and there was a trend toward reduction of ischaemic vascular lesions. However, it was unable to rescue the prominent inflammatory response or decline in cognitive ability, as apocynin-treated mice were no different on these measures to non-treated hypoperfused mice. The data indicate that whilst inhibiting NOX may have potential therapeutic value in improving vascular function, additional interventions, for example to reduce inflammation, may also be required in order to prevent cognitive decline. Overall, the work outlined within the thesis indicate that vascular risk factors of ageing, cerebral amyloid angiopathy and cerebral hypoperfusion may converge on common pathways involving oxidative stress and increased inflammation in order to drive vascular dysfunction and lead to cognitive decline. Inhibition of NOX activity was able to rescue vascular function, however the results indicate that this was not sufficient to protect against cognitive impairment, suggesting additional therapeutic targets may need to be sought in order to fully preserve vascular health and prevent cognitive decline

Chronic cerebral hypoperfusion:a key mechanism leading to vascular cognitive impairment and dementia. Closing the translational gap between rodent models and human vascular cognitive impairment and dementia

Increasing evidence suggests that vascular risk factors contribute to neurodegeneration, cognitive impairment and dementia. While there is considerable overlap between features of vascular cognitive impairment and dementia (VCID) and Alzheimer’s disease (AD), it appears that cerebral hypoperfusion is the common underlying pathophysiological mechanism which is a major contributor to cognitive decline and degenerative processes leading to dementia. Sustained cerebral hypoperfusion is suggested to be the cause of white matter attenuation, a key feature common to both AD and dementia associated with cerebral small vessel disease (SVD). White matter changes increase the risk for stroke, dementia and disability. A major gap has been the lack of mechanistic insights into the evolution and progress of VCID. However, this gap is closing with the recent refinement of rodent models which replicate chronic cerebral hypoperfusion. In this review, we discuss the relevance and advantages of these models in elucidating the pathogenesis of VCID and explore the interplay between hypoperfusion and the deposition of amyloid β (Aβ) protein, as it relates to AD. We use examples of our recent investigations to illustrate the utility of the model in preclinical testing of candidate drugs and lifestyle factors. We propose that the use of such models is necessary for tackling the urgently needed translational gap from preclinical models to clinical treatments.</jats:p

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

ACKNOWLEDGMENTS Schematic diagrams in Figures 2, 8 are created withBiorender.com. FUNDING We gratefully acknowledge the grant support from the Alzheimer’s Society (152 (PG-157); 290 (AS-PG-15b-018); 228 (AS-DTC-2014-017), 314 (AS –PhD-16-006), and Alzheimer’s Research United Kingdom (ART-PG2010-3; ARUK-PG2013- 22; ARUK-PG2016B-6), and The University of Edinburgh Centre for Cognitive Ageing and Cognitive Epidemiology, part of the cross council Lifelong Health and Wellbeing Initiative (G0700704/84698). ML and JB are funded by an Alzheimer’s Society Scotland Doctoral Training Programme and RS Macdonald Trust. ML was also funded by a China Scholarship Council (CSC)/University of Edinburgh scholarship.Peer reviewedPublisher PD

INHIBITING CSF1R ALLEVIATES CEREBROVASCULAR WHITE MATTER DISEASE AND COGNITIVE IMPAIRMENT

White matter abnormalities, related to poor cerebral perfusion, are a core feature of small vessel cerebrovascular disease, and critical determinants of vascular cognitive impairment and dementia. Despite this importance there is a lack of treatment options. Proliferation of microglia producing an expanded, reactive population and associated neuroinflammatory alterations have been implicated in the onset and progression of cerebrovascular white matter disease, in patients and in animal models, suggesting that targeting microglial proliferation may exert protection. Colony-stimulating factor-1 receptor (CSF1R) is a key regulator of microglial proliferation. We found that the expression of CSF1R/Csf1r and other markers indicative of increased microglial abundance are significantly elevated in damaged white matter in human cerebrovascular disease and in a clinically relevant mouse model of chronic cerebral hypoperfusion and vascular cognitive impairment. Using the mouse model, we investigated long-term pharmacological CSF1R inhibition, via GW2580, and demonstrated that the expansion of microglial numbers in chronic hypoperfused white matter is prevented. Transcriptomic analysis of hypoperfused white matter tissue showed enrichment of microglial and inflammatory gene sets, including phagocytic genes that were the predominant expression modules modified by CSF1R inhibition. Further, CSF1R inhibition attenuated hypoperfusion-induced white matter pathology and rescued spatial learning impairments and to a lesser extent cognitive flexibility. Overall, this work suggests that inhibition of CSF1R and microglial proliferation mediates protection against chronic cerebrovascular white matter pathology and cognitive deficits. Our study nominates CSF1R as a target for the treatment of vascular cognitive disorders with broader implications for treatment of other chronic white matter diseases.<br/

Extracellular vesicles and preeclampsia : current knowledge and future research directions

Preeclampsia (PE) is associated with long-term morbidity in mothers and lifelong morbidities for their children, ranging from cerebral palsy and cognitive delay in preterm infants, to hypertension, diabetes and obesity in adolescents and young adults. There are several processes that are critical for development of materno-fetal exchange, including establishing adequate perfusion of the placenta by maternal blood, and the formation of the placental villous vascular tree. Recent studies provide persuasive evidence that placenta-derived extracellular vesicles (EVs) represent a significant intercellular communication pathway, and that they may play an important role in placental and endothelial cell (both fetal and maternal) function. These functions are known to be altered in PE. EVs can carry and transport a wide range of bioactive molescules that have potential to be used as biomarkers and therapeutic delivery tools for PE. EV content is often parent cell specific, thus providing an insight or “thumbprint” of the intracellular environment of the originating cell (e.g., human placenta). EV have been identified in plasma under both normal and pathological conditions, including PE. The concentration of EVs and their content in plasma has been reported to increase in association with disease severity and/or progression. Placenta-derived EVs have been identified in maternal plasma during normal pregnancy and PE pregnancies. They contain placenta-specific proteins and miRNAs and, as such, may be differentiated from maternally-derived EVs. The aim of this review, thus, is to describe the potential roles of EVs in preecmpatic pregnancies, focussing on EVs secreted from placental cells. The biogenesis, specificity of placental EVs, and methods used to characterise EVs in the context of PE pregnancies will be also discussed

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

Diagnosis and investigation of suspected haemophagocytic lymphohistiocytosis in adults: 2023 Hyperinflammation and HLH Across Speciality Collaboration (HiHASC) consensus guideline

Haemophagocytic lymphohistiocytosis (HLH) is a hyperinflammatory syndrome characterised by persistently activated cytotoxic lymphocytes and macrophages, which, if untreated, leads to multiorgan dysfunction and death. HLH should be considered in any acutely unwell patient not responding to treatment as expected, with prompt assessment to look for what we term the three Fs—fever, falling blood counts, and raised ferritin. Worldwide, awareness of HLH and access to expert management remain inequitable. Terminology is not standardised, classification criteria are validated in specific patient groups only, and some guidelines rely on specialised and somewhat inaccessible tests. The consensus guideline described in this Health Policy was produced by a self-nominated working group from the UK network Hyperinflammation and HLH Across Speciality Collaboration (HiHASC), a multidisciplinary group of clinicians experienced in managing people with HLH. Combining literature review and experience gained from looking after patients with HLH, it provides a practical, structured approach for all health-care teams managing adult (>16 years) patients with possible HLH. The focus is on early recognition and diagnosis of HLH and parallel identification of the underlying cause. To ensure wide applicability, the use of inexpensive, readily available tests is prioritised, but the role of specialist investigations and their interpretation is also addressed

Small vessels, dementia and chronic diseases – molecular mechanisms and pathophysiology

Cerebral small vessel disease (SVD) is a major contributor to stroke, cognitive impairment and dementia with limited therapeutic interventions. There is a critical need to provide mechanistic insight and improve translation between pre-clinical research and the clinic. A 2-day workshop was held which brought together experts from several disciplines in cerebrovascular disease, dementia and cardiovascular biology, to highlight current advances in these fields, explore synergies and scope for development. These proceedings provide a summary of key talks at the workshop with a particular focus on animal models of cerebral vascular disease and dementia, mechanisms and approaches to improve translation. The outcomes of discussion groups on related themes to identify the gaps in knowledge and requirements to advance knowledge are summarized