Model Of Small Vessel Disease Research Articles

A small molecule p38α MAPK inhibitor, MW150, attenuates behavioral deficits and neuronal dysfunction in a mouse model of mixed amyloid and vascular pathologies

BackgroundInhibition of p38 alpha mitogen activated protein kinase (p38α) has shown great promise as a treatment for Alzheimer's disease (AD) in preclinical tests. However, previous preclinical studies were performed in “pure” models of AD pathology. A vast majority of AD patients have comorbid dementia-contributing pathologies, particularly some form of vascular damage. The present study therefore aimed to test the potential of p38α inhibition to address dysfunction in the context of comorbid amyloid and vascular pathologies. MethodsAn amyloid overexpressing mouse strain (5xFAD) was placed on an 8-week long diet to induce the hyperhomocysteinemia (HHcy) model of small vessel disease. Mice were treated with the brain-penetrant small molecule p38α inhibitor MW150 for the duration of the HHcy diet, and subsequently underwent behavioral, neuroimaging, electrophysiological, or biochemical/immunohistochemical analyses. ResultsMW150 successfully reduced behavioral impairment in the Morris Water Maze, corresponding with attenuation of synaptic loss, reduction in tau phosphorylation, and a partial normalization of electrophysiological parameters. No effect of MW150 was observed on the amyloid, vascular, or neuroinflammatory endpoints measured. ConclusionsThis study provides proof-of-principle that the inhibition of p38α is able to provide benefit even in the context of mixed pathological contributions to cognitive impairment. Interestingly, the benefit was mediated primarily via rescue of neuronal function without any direct effects on the primary pathologies. These data suggest a potential use for p38 inhibitors in the preservation of cognition across contexts, and in particular AD, either alone or as an adjunct to other AD therapies (i.e. anti-amyloid approaches). Future studies to delineate the precise neuronal pathways implicated in the benefit may help define other specific comorbid conditions amenable to this type of approach or suggest future refinement in pharmacological targeting.

MINocyclinE to Reduce inflammation and blood brain barrier leakage in small Vessel diseAse (MINERVA) trial study protocol.

Background:Cerebral small vessel disease (SVD) is a common cause of stroke and cognitive impairment. Recent data has implicated neuroinflammation and increased blood-brain barrier (BBB) permeability in its pathogenesis, but whether such processes are causal and can be therapeutically modified is uncertain. In a rodent model of SVD, minocycline was associated with reduced white matter lesions, inflammation and BBB permeability.Aims:To determine whether blood-brain barrier permeability (measured using dynamic contrast-enhanced MRI) and microglial activation (measured by positron emission tomography using the radioligand 11C-PK11195) can be modified in SVD.Design:Phase II randomised double blind, placebo-controlled trial of minocycline 100 mg twice daily for 3 months in 44 participants with moderate to severe SVD defined as a clinical lacunar stroke and confluent white matter hyperintensities.Outcomes:Primary outcome measures are volume and intensity of focal increases of blood-brain barrier permeability and microglial activation determined using PET-MRI imaging. Secondary outcome measures include inflammatory biomarkers in serum, and change in conventional MRI markers and cognitive performance over 1 year follow up.Discussion:The MINERVA trial aims to test whether minocycline can influence novel pathological processes thought to be involved in SVD progression, and will provide insights into whether central nervous system inflammation in SVD can be therapeutically modulated.

Mizagliflozin, a selective SGLT1 inhibitor, improves vascular cognitive impairment in a mouse model of small vessel disease.

Previously, we showed that sodium/glucose cotransporter 1 (SGLT1) participates in vascular cognitive impairment in small vessel disease. We hypothesized that SGLT1 inhibitors can improve the small vessel disease induced‐vascular cognitive impairment. We examined the effects of mizagliflozin, a selective SGLT1 inhibitor, and phlorizin, a non‐selective SGLT inhibitor, on vascular cognitive impairment in a mouse model of small vessel disease. Small vessel disease was created using a mouse model of asymmetric common carotid artery surgery (ACAS). Two and/or 4 weeks after ACAS, all experiments were performed. Cerebral blood flow (CBF) was decreased in ACAS compared with sham‐operated mice. Phlorizin but not mizagliflozin reversed the decreased CBF of ACAS mice. Both mizagliflozin and phlorizin reversed the ACAS‐induced decrease in the latency to fall in a wire hang test of ACAS mice. Moreover, they reversed the ACAS‐induced longer escape latencies in the Morris water maze test of ACAS mice. ACAS increased SGLT1 and proinflammatory cytokine gene expressions in mouse brains and phlorizin but not mizagliflozin normalized all gene expressions in ACAS mice. Hematoxylin/eosin staining demonstrated that they inhibited pyknotic cell death in the ACAS mouse hippocampus. In PC12HS cells, IL‐1β increased SGLT1 expression and decreased survival rates of cells. Both mizagliflozin and phlorizin increased the survival rates of IL‐1β‐treated PC12HS cells. These results suggest that mizagliflozin and phlorizin can improve vascular cognitive impairment through the inhibition of neural SGLT1 and phlorizin also does so through the improvement of CBF in a mouse model of small vessel disease.

HB-EGF depolarizes hippocampal arterioles to restore myogenic tone in a genetic model of small vessel disease

Vascular cognitive impairment, the second most common cause of dementia, profoundly affects hippocampal-dependent functions. However, while the growing literature covers complex neuronal interactions, little is known about the sustaining hippocampal microcirculation. Here we examined vasoconstriction to physiological pressures of hippocampal arterioles, a fundamental feature of small arteries, in a genetic mouse model of CADASIL, an archetypal cerebral small vessel disease. Using diameter and membrane potential recordings on isolated arterioles, we observed both blunted pressure-induced vasoconstriction and smooth muscle cell depolarization in CADASIL. This impairment was abolished in the presence of voltage-gated potassium (KV1) channel blocker 4-aminopyridine, or by application of heparin-binding EGF-like growth factor (HB-EGF), which promotes KV1 channel down-regulations. Interestingly, we observed that HB-EGF induced a depolarization of the myocyte plasma membrane within the arteriolar wall in CADASIL, but not wild-type, arterioles. Collectively, our results indicate that hippocampal arterioles in CADASIL mice display a blunted contractile response to luminal pressure, similar to the defect we previously reported in cortical arterioles and pial arteries, that is rescued by HB-EGF. Hippocampal vascular dysfunction in CADASIL could then contribute to the decreased vascular reserve associated with decreased cognitive performance, and its correction may provide a therapeutic option for treating vascular cognitive impairment.

SGLT1 participates in the development of vascular cognitive impairment in a mouse model of small vessel disease

Sodium/glucose cotransporter 1 (SGLT1) participates in ischemia-reperfusion-induced cerebral injury. However, whether SGLT1 participates in the development of small vessel disease induced-vascular cognitive impairment is unknown. We examined the roles of SGLT1 in the development of vascular cognitive impairment in a mouse model of small vessel disease. Small vessel disease was created by placement of an ameroid constrictor around the right common carotid artery (CCA) and placement of a microcoil around the left CCA (ACAS) in wild-type (WT) and SGLT1-knock out (KO) mice. Two and/or 4 weeks after ACAS, all experiments were performed. Hematoxylin/eosin staining demonstrated that the number of pyknotic cell deaths was greater in the ACAS WT than ACAS SGLT1-KO hippocampus. The latency to fall in a wire hang test was significantly shorter in ACAS than sham-operated WT mice, whereas it was similar between ACAS and sham-operated SGLT1-KO mice. The Morris water maze test revealed that ACAS WT mice exhibited longer escape latencies than ACAS SGLT1-KO mice. ACAS significantly increased SGLT1 gene expression in WT mouse brains. Gene expressions of MCP-1, IL-1β, TNF-α, and IL-6 were increased in ACAS WT compared with sham-operated WT mouse brains. Their increased gene expressions were significantly decreased in ACAS SGLT1-KO compared with ACAS WT mice. These results suggest that SGLT1 plays important roles in the development of small vessel dementia.

Abstract 052: Rho Kinase and Endoplasmic Reticulum Stress Mediate Peripheral Vascular Dysfunction in a Model of Small Vessel Disease of the Brain

CADASIL, a monogenic condition due to Notch3 mutations is a small vessel disease of the brain resulting in premature vascular dementia and stroke. CADASIL patients have cerebral vascular narrowing, progressive SMCs loss and granular osmiophilic material (GOM) deposition, however it is unclear whether peripheral arteries also exhibit dysfunction. We studied mice harbouring the CADASIL-causing Notch3 mutation (TgNotch3 R169C ), which recapitulates human CADASIL, to evaluate whether peripheral small arteries exhibit vascular dysfunction. Mesenteric arteries (MA) from male TgNotch3 R169C and wildtype (TgNotch3 WT ) mice (6 months old) were investigated. Vascular structure and function were assessed by myography. No changes were observed in blood pressure and cardiac function in CADASIL mice, measured by tail cuff and echocardiography respectively. Expression of Notch3 and target genes was augmented in MA from CADASIL mice ( Notch3 : 2.0±0.5 vs. 6.0±1.3; HeyL: 1.1±0.4 vs. 2.9±0.6, p<0.01). CADASIL mice exhibited impaired endothelium-dependent (Emax 109.9±7.4 vs. 81.3±5.4) and -independent (pD 2 7.8±0.1 vs 6.8±0.3) relaxation and increased reactivity in response to AngII (Emax 33.7±6.8 vs. 72.8±4.4), phenylephrine (Emax: 70.6±7.2 vs. 92.1±4.2), and U46619 (Emax: 123.4±4.4 vs. 215.1±24.4) (p<0.05 vs TgNotch3 WT ); effects attenuated by fasudil (Rho-kinase inhibitor) and 4-PBA (ER stress inhibitor) (p<0.05 vs. TgNotch3 R169C ). MA from CADASIL mice also had hypotrophic remodelling. U46619-induced calcium influx was increased in VSMCs (AUC: 1.3 fold increase) from TgNotch3 R169C and gene expression of Rho GEFs (LARG: 1.1±0.1 vs. 2.1±0.3; PDZ: 0.9±0.1 vs. 2.9±0.4) was increased in MA from TgNotch3 R169C vs. TgNotch3 WT mice (p<0.05). eNOS phosphorylation (Thr 495 ) and BiP expression (ER stress marker) were increased in vessels from CADASIL mice (eNOS: 1.8 fold increase; Bip: 1.7 fold increase; p<0.05 vs. TgNotch3 WT ). In conclusion, our data demonstrated that the vasculopathy associated with Notch3 mutation is also present in peripheral small vessels where the interplay between Notch3, Rho-kinase and ER stress may be important. We identify potential new therapeutic targets in CADASIL, for which there are no disease-specific treatments.

Effects of dietary salt on gene and protein expression in brain tissue of a model of sporadic small vessel disease.

Background: The effect of salt on cerebral small vessel disease (SVD) is poorly understood. We assessed the effect of dietary salt on cerebral tissue of the stroke-prone spontaneously hypertensive rat (SHRSP) – a relevant model of sporadic SVD – at both the gene and protein level. Methods: Brains from 21-week-old SHRSP and Wistar-Kyoto rats, half additionally salt-loaded (via a 3-week regime of 1% NaCl in drinking water), were split into two hemispheres and sectioned coronally – one hemisphere for mRNA microarray and qRT-PCR, the other for immunohistochemistry using a panel of antibodies targeting components of the neurovascular unit. Results: We observed differences in gene and protein expression affecting the acute phase pathway and oxidative stress (ALB, AMBP, APOH, AHSG and LOC100129193, up-regulated in salt-loaded WKY versus WKY, >2-fold), active microglia (increased Iba-1 protein expression in salt-loaded SHRSP versus salt-loaded WKY, p<0.05), vascular structure (ACTB and CTNNB, up-regulated in salt-loaded SHRSP versus SHRSP, >3-fold; CLDN-11, VEGF and VGF down-regulated >2-fold in salt-loaded SHRSP versus SHRSP) and myelin integrity (MBP down-regulated in salt loaded WKY rats versus WKY, >2.5-fold). Changes of salt-loading were more pronounced in SHRSP and occurred without an increase in blood pressure in WKY rats. Conclusion: Salt exposure induced changes in gene and protein expression in an experimental model of SVD and its parent rat strain in multiple pathways involving components of the glio-vascular unit. Further studies in pertinent experimental models at different ages would help clarify the short- and long-term effect of dietary salt in SVD.

Abstract P242: Microvascular Changes in Brain During Salt-sensitive Hypertension: Molecular Mechanisms and Implications for Small Vessel Disease

Hypertension is a major risk factor for small vessel disease (SVD), a leading contributor to stroke and dementias. Mechanisms that underlie SVD in brain are poorly defined, with no specific therapy at present. Because parenchymal and pial arterioles are targets of the SVD process, we examined microvascular changes in a model using deoxycorticosterone-salt (DOCA) to activate the brain renin-angiotensin system (RAS), with resulting salt-sensitive (sodium- and fluid-dependent) hypertension. Male C57Bl/6 mice were treated with DOCA and given the choice of drinking H 2 O or H 2 O with 0.9% NaCl for 3 wks. Along with a modest elevation in mean arterial pressure (79±2 vs 95±3 mmHg, P&lt;0.05), DOCA impaired endothelium-dependent dilation of both isolated parenchymal (baseline diameter of 15±1 μm) and pial arterioles (37±1 μm) in a pathway specific manner. Endothelium-dependent hyperpolarization was intact while eNOS-mediated vasodilation was markedly impaired along with reductions in phosphorylation in AKT (an upstream activator of eNOS). Local inhibition of angiotensin II type 1 (AT1-R) or mineralocorticoid receptors (MR) or Rho kinase (including ROCK2), restored endothelial function in DOCA-treated mice. Inner diameter of maximally dilated parenchymal arterioles was reduced approximately 20% by DOCA (P&lt;0.05 vs sham). DOCA increased mRNA expression of RAS components (eg, AGT, ACE) in both brain and cerebral vessels. In NZ44 reporter mice that express GFP driven by the AT1A-R promoter, DOCA increased cerebrovascular GFP protein expression about 3-fold (P&lt;0.05). Thus, DOCA activates both the brain and the cerebrovascular RAS, impairs select pathways affecting parenchymal and pial arteriolar function, while producing inward microvascular remodeling. AT1R, MR and ROCK2 are key contributors to cerebral microvascular dysfunction in this clinically relevant model of SVD.

Potassium channelopathy-like defect underlies early-stage cerebrovascular dysfunction in a genetic model of small vessel disease

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), caused by dominant mutations in the NOTCH3 receptor in vascular smooth muscle, is a genetic paradigm of small vessel disease (SVD) of the brain. Recent studies using transgenic (Tg)Notch3(R169C) mice, a genetic model of CADASIL, revealed functional defects in cerebral (pial) arteries on the surface of the brain at an early stage of disease progression. Here, using parenchymal arterioles (PAs) from within the brain, we determined the molecular mechanism underlying the early functional deficits associated with this Notch3 mutation. At physiological pressure (40 mmHg), smooth muscle membrane potential depolarization and constriction to pressure (myogenic tone) were blunted in PAs from TgNotch3(R169C) mice. This effect was associated with an ∼ 60% increase in the number of voltage-gated potassium (KV) channels, which oppose pressure-induced depolarization. Inhibition of KV1 channels with 4-aminopyridine (4-AP) or treatment with the epidermal growth factor receptor agonist heparin-binding EGF (HB-EGF), which promotes KV1 channel endocytosis, reduced KV current density and restored myogenic responses in PAs from TgNotch3(R169C) mice, whereas pharmacological inhibition of other major vasodilatory influences had no effect. KV1 currents and myogenic responses were similarly altered in pial arteries from TgNotch3(R169C) mice, but not in mesenteric arteries. Interestingly, HB-EGF had no effect on mesenteric arteries, suggesting a possible mechanistic basis for the exclusive cerebrovascular manifestation of CADASIL. Collectively, our results indicate that increasing the number of KV1 channels in cerebral smooth muscle produces a mutant vascular phenotype akin to a channelopathy in a genetic model of SVD.

Dilated Perivascular Spaces in Small-Vessel Disease: A Study in CADASIL

Background and Aim: Dilated perivascular spaces (dPVS) have previously been associated with aging and hypertension-related cerebral microangiopathy. However, their risk factors, radiological features and clinical relevance have been poorly evaluated in CADASIL (cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy), a unique model to investigate the pathophysiology of ischemic small-vessel disease. The purpose of this study was to investigate these different aspects in a large cohort of patients with this disorder. Methods: Demographic and MRI data of 344 patients from a prospective cohort study were analyzed. The severity of dPVS was evaluated separately in the anterior temporal lobes, subinsular areas, basal ganglia and white matter, using validated semiquantitative scales. Logistic and multiple linear regression models were used to determine the risk factors associated with the severity of dPVS in these different regions and their relationships with cognition, disability and the MRI markers of the disease (white matter hyperintensities (WMH) lacunar infarcts, microbleeds and brain parenchymal fraction (BPF)). Results: The severity of dPVS was found to increase with age regardless of cerebral area (p < 0.001). In contrast with dPVS in other locations, the severity of dPVS in the temporal lobes or subinsular areas was also found strongly and specifically related to the extent of WMH (p < 0.001). Conversely, no significant association was detected with lacunar volume, number of microbleeds or BPF. A high degree of dPVS in the white matter was associated with lower cognitive performances independently of age and other MRI markers of the disease including BPF (p ≤ 0.04). Conclusions: In CADASIL, the progression of the hereditary microangiopathy with aging may promote the dilation of perivascular spaces throughout the whole brain but with variable extent according to cerebral location. In temporal lobes and subinsular areas, dPVS are common MRI features and may share a similar pathogenesis with the extension of WMH during the course of the disease. dPVS may also participate in the development of cognitive decline in this model of small-vessel disease, and their large number in white matter may alert clinicians to a higher risk of cognitive decline in CADASIL.

Cerebrovascular dysfunction and microcirculation rarefaction precede white matter lesions in a mouse genetic model of cerebral ischemic small vessel disease

Cerebral ischemic small vessel disease (SVD) is the leading cause of vascular dementia and a major contributor to stroke in humans. Dominant mutations in NOTCH3 cause cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a genetic archetype of cerebral ischemic SVD. Progress toward understanding the pathogenesis of this disease and developing effective therapies has been hampered by the lack of a good animal model. Here, we report the development of a mouse model for CADASIL via the introduction of a CADASIL-causing Notch3 point mutation into a large P1-derived artificial chromosome (PAC). In vivo expression of the mutated PAC transgene in the mouse reproduced the endogenous Notch3 expression pattern and main pathological features of CADASIL, including Notch3 extracellular domain aggregates and granular osmiophilic material (GOM) deposits in brain vessels, progressive white matter damage, and reduced cerebral blood flow. Mutant mice displayed attenuated myogenic responses and reduced caliber of brain arteries as well as impaired cerebrovascular autoregulation and functional hyperemia. Further, we identified a substantial reduction of white matter capillary density. These neuropathological changes occurred in the absence of either histologically detectable alterations in cerebral artery structure or blood-brain barrier breakdown. These studies provide in vivo evidence for cerebrovascular dysfunction and microcirculatory failure as key contributors to hypoperfusion and white matter damage in this genetic model of ischemic SVD.

Measurement of brain atrophy in subcortical vascular disease: A comparison of different approaches and the impact of ischaemic lesions

Measurement of brain atrophy has been proposed as a surrogate marker in MS and degenerative dementias. Although cerebral small vessel disease predominantly affects white and subcortical grey matter, recent data suggest that whole brain atrophy is also a good indicator of clinical and cognitive status in this disease. Automated methods to measure atrophy are available that are accurate and reproducible in disease-free brains. However, optimal methods in small vessel disease have not been established and the impact of ischaemic lesions on different techniques has not been explored systematically. In this study, three contrasting techniques – Statistical Parametric Mapping 5 (SPM5), SIENAX and BrainVisa – were applied to measure cross-sectional atrophy (brain parenchymal fraction or BPF) in a large ( n = 143) two-centre cohort of patients with CADASIL, a genetic model of small vessel disease. All three techniques showed similar sensitivity to trends in BPF associated with age and lesion load. No single technique was particularly vulnerable to error as a result of lesions. Provided major errors in registration were excluded by visual inspection, manual correction of segmentations had a negligible impact with mean errors of 0.41% for SIENAX and 0.46% for BrainVisa. BPF correlated strongly with global cognitive function and physical disability, independent of the technique used. Correlation coefficients with the Minimental State Examination score were: BrainVisa 0.58, SIENAX 0.58, SPM5 0.60 (for all, p < 0.001). These results suggest that all three methods can be applied reliably in patients with ischaemic lesions. Choice of analysis approach for this kind of clinical question will be determined by factors other than their robustness and precision, such as a desire to explore subtle localised changes using extensions of these processing tools.

Hippocampal volume is an independent predictor of global cognitive performance in pure cerebral small vessel disease

Background: Hippocampal atrophy is a hallmark of Alzheimer's disease (AD), but its importance in cerebral small vessel disease (SVD) is unclear. Attempts to study this in sporadic SVD are hampered by the advanced age of many patients and the presence of co-morbid neurodegenerative pathology. To clarify the role of hippocampal changes in SVD we investigated patients with CADASIL, a model of pure SVD and associated cognitive impairment.

Cerebrovascular Risk Factors and Later-Life Major Depression: Testing a Small-Vessel Brain Disease Model

The topic of vascular depression has received increasing prominence as a putative etiology of depression in later life. The authors examined one aspect of this model by comparing the burden of systemic cerebrovascular risk factors (CVRFs) in 130 psychiatric inpatients with major depression and 64 normal control (NC) subjects, all age > or = 50 years. Depressed subjects did not differ statistically from NCs on cumulative CVRF scores. Diabetes mellitus and atrial fibrillation were both associated with depression, but only atrial fibrillation retained an independent association after medical disability was statistically controlled. Among the depressed subjects, CVRF scores were not significantly associated with overall symptom severity, psychiatric disability, age at onset of depression, melancholic subtype, or psychotic depression. These data did not support the notion that a linear model of small-vessel disease might apply to the great majority of older inpatients with major depression.