Associations between Cerebrovascular Function and the Expression of Genes Related to Endothelial Function in Hormonal Migraine.

There is evidence to suggest that hormonal migraine is associated with altered cerebrovascular function. We aimed to investigate whether the expression of genes related to endothelial function in venous blood (1) might influence cerebrovascular function, (2) differs between hormonal migraineur and non-migraineur women, and (3) changes following resveratrol supplementation. This study utilised data obtained from 87 women (59 hormonal migraineurs and 28 controls) where RNA from venous blood was used to quantify gene expression and transcranial Doppler ultrasound was used to evaluate cerebrovascular function. Spearman's correlation analyses were performed between gene expression, cerebrovascular function, and migraine-related disability. We compared the expression of genes associated with endothelial function between migraineurs and non-migraineurs, and between resveratrol and placebo. The expression of several genes related to endothelial function was associated with alterations in cerebrovascular function. Notably, the expression of CALCA was associated with increased neurovascular coupling capacity (p = 0.013), and both CALCA (p = 0.035) and VEGF (p = 0.014) expression were associated with increased cerebral blood flow velocity in the overall study population. Additionally, VCAM1 expression correlated with decreased pulsatility index (a measure of cerebral arterial stiffness) (p = 0.009) and headache impact test-6 scores (p = 0.007) in the migraineurs. No significant differences in gene expression were observed between migraineurs and controls, or between placebo and resveratrol treatments in migraineurs. Thus, altering the expression of genes related to endothelial function may improve cerebrovascular function and decrease migraine-related disability.

A roadmap for research in post-stroke fatigue: Consensus-based core recommendations from the third Stroke Recovery and Rehabilitation Roundtable.

RATIONALE: Fatigue affects almost half of all people living with stroke. Stroke survivors rank understanding fatigue and how to reduce it as one of the highest research priorities. METHODS: We convened an interdisciplinary, international group of clinical and pre-clinical researchers and lived experience experts. We identified four priority areas: (1) best measurement tools for research, (2) clinical identification of fatigue and potentially modifiable causes, (3) promising interventions and recommendations for future trials, and (4) possible biological mechanisms of fatigue. Cross-cutting themes were aphasia and the voice of people with lived experience. Working parties were formed and structured consensus building processes were followed. RESULTS: We present 20 recommendations covering outcome measures for research, development, and testing of new interventions and priority areas for future research on the biology of post-stroke fatigue. We developed and recommend the use of the Stroke Fatigue Clinical Assessment Tool. CONCLUSIONS: By synthesizing current knowledge in post-stroke fatigue across clinical and pre-clinical fields, our work provides a roadmap for future research into post-stroke fatigue.

A roadmap for research in post-stroke fatigue: Consensus-based core recommendations from the third Stroke Recovery and Rehabilitation Roundtable.

RATIONALE: Fatigue affects almost half of all people living with stroke. Stroke survivors rank understanding fatigue and how to reduce it as one of the highest research priorities. METHODS: We convened an interdisciplinary, international group of clinical and pre-clinical researchers and lived experience experts. We identified four priority areas: (1) best measurement tools for research, (2) clinical identification of fatigue and potentially modifiable causes, (3) promising interventions and recommendations for future trials, and (4) possible biological mechanisms of fatigue. Cross-cutting themes were aphasia and the voice of people with lived experience. Working parties were formed and structured consensus building processes were followed. RESULTS: We present 20 recommendations covering outcome measures for research, development, and testing of new interventions and priority areas for future research on the biology of post-stroke fatigue. We developed and recommend the use of the Stroke Fatigue Clinical Assessment Tool. CONCLUSIONS: By synthesizing current knowledge in post-stroke fatigue across clinical and pre-clinical fields, our work provides a roadmap for future research into post-stroke fatigue.

Mechanisms of Post-Stroke Fatigue: A Follow-Up From the Third Stroke Recovery and Rehabilitation Roundtable.

BACKGROUND: Post-stroke fatigue (PSF) is a significant and highly prevalent symptom, whose mechanisms are poorly understood. The third Stroke Recovery and Rehabilitation Roundtable paper on PSF focussed primarily on defining and measuring PSF while mechanisms were briefly discussed. This companion paper to the main paper is aimed at elaborating possible mechanisms of PSF. METHODS: This paper reviews the available evidence that potentially explains the pathophysiology of PSF and draws parallels from fatigue literature in other conditions. We start by proposing a case for phenotyping PSF based on structural, functional, and behavioral characteristics of PSF. This is followed by discussion of a potentially significant role of early inflammation in the development of fatigue, specifically the impact of low-grade inflammation and its long-term systemic effects resulting in PSF. Of the many neurotransmitter systems in the brain, the dopaminergic systems have the most evidence for a role in PSF, along with a role in sensorimotor processing. Sensorimotor neural network dynamics are compromised as highlighted by evidence from both neurostimulation and neuromodulation studies. The double-edged sword effect of exercise on PSF provides further insight into how PSF might emerge and the importance of carefully titrating interventional paradigms. CONCLUSION: The paper concludes by synthesizing the presented evidence into a unifying model of fatigue which distinguishes between factors that pre-dispose, precipitate, and perpetuate PSF. This framework will help guide new research into the biological mechanisms of PSF which is a necessary prerequisite for developing treatments to mitigate the debilitating effects of post-stroke fatigue.

Active immunotherapy reduces NOTCH3 deposition in brain capillaries in a CADASIL mouse model.

Cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is the most common monogenic form of familial small vessel disease; no preventive or curative therapy is available. CADASIL is caused by mutations in the NOTCH3 gene, resulting in a mutated NOTCH3 receptor, with aggregation of the NOTCH3 extracellular domain (ECD) around vascular smooth muscle cells. In this study, we have developed a novel active immunization therapy specifically targeting CADASIL-like aggregated NOTCH3 ECD. Immunizing CADASIL TgN3R182C150 mice with aggregates composed of CADASIL-R133C mutated and wild-type EGF1-5 repeats for a total of 4 months resulted in a marked reduction (38-48%) in NOTCH3 deposition around brain capillaries, increased microglia activation and lowered serum levels of NOTCH3 ECD. Active immunization did not impact body weight, general behavior, the number and integrity of vascular smooth muscle cells in the retina, neuronal survival, or inflammation or the renal system, suggesting that the therapy is tolerable. This is the first therapeutic study reporting a successful reduction of NOTCH3 accumulation in a CADASIL mouse model supporting further development towards clinical application for the benefit of CADASIL patients.

Exploring the effects of resveratrol supplementation on cerebrovascular function in hormonal migraineurs: A pilot study.

Background: Past research suggests that hormonal migraineurs may have poorer cerebrovascular function than women who do not suffer from migraine. Resveratrol, a vasoactive phytoestrogen, has been shown to improve cerebrovascular function in several populations but has never been tested in hormonal migraineurs. Aim: To investigate the effects of 3-month resveratrol supplementation on the cerebrovascular function of hormonal migraineurs. Methods: We conducted a randomised, double-blind, placebo-controlled, crossover intervention pilot study with resveratrol (150 mg/d for 3 months) in ten hormonal migraineurs (mean age: 37.2 ± 2.6 years). Participants visited the University of Newcastle's Clinical Nutrition Research Centre where quality of life and disability, and cerebrovascular function were assessed. Quality of life and disability were examined using Migraine-Specific Quality of Life, Headache Impact Test-6 and the Migraine Disability Assessment. Cerebrovascular function was determined using transcranial Doppler ultrasound to bilaterally measure blood flow velocity in the middle and posterior cerebral arteries at rest and in response to a hypercapnic stimulus. Cerebrovascular responsiveness to a cognitive task battery was also measured bilaterally in the middle cerebral arteries. Results: Compared to placebo, blood flow velocity in the right posterior cerebral artery was significantly higher (P = 0.041) following resveratrol supplementation. No other significant differences in cerebrovascular function between resveratrol and placebo treatments were observed. Baseline correlation analyses revealed higher blood flow velocities in the middle and posterior cerebral arteries were associated with better quality of life and less disability. However, higher cerebrovascular responsiveness to hypercapnia in the posterior circulation was associated with higher migraine-related disability and poorer migraine-related quality of life. Conclusion: In this pilot we found evidence that resveratrol may increase blood flow velocity in the right posterior cerebral artery in hormonal migraineurs. Larger cohorts are required confirm this effect and its potential relationship to migraine in premenopausal women.

A Randomised, Double-Blind, Placebo-Controlled Crossover Trial of Resveratrol Supplementation for Prophylaxis of Hormonal Migraine.

Resveratrol, a vasoactive phytoestrogen, has beneficial effects on cerebrovascular function. Previous research has shown that hormonal migraineurs have poorer cerebrovascular function than non-migraineur women. We aimed to investigate if resveratrol supplementation for three months could reduce the hormonal migraine burden index (HMBI: the number of days with menstrual migraine per month), reduce migraine-related disability and improve migraine-related quality of life. A randomised, double-blind, placebo-controlled, crossover, intervention trial was conducted in 62 hormonal migraineurs (mean age: 37.5 ± 0.8 years). Participants consumed 75 mg of resveratrol or matching placebo capsules twice daily for three months before crossing over to the other treatment arm. Participants completed a daily diary and the Headache Impact Test-6™, Migraine Disability Assessment and Migraine-Specific Quality of Life questionnaires at months 0, 3 and 6. The HMBI was the primary outcome and was calculated using data extracted from the participant's diary. No differences in the HMBI (p = 0.895), the Headache Impact Test-6™, the Migraine Disability Assessment and Migraine-Specific Quality of Life were found between the resveratrol and placebo treatments. Resveratrol supplementation for three months did not affect the HMBI, the migraine-related disability or quality of life measures in our cohort of hormonal migraineurs.

Cerebrovascular Function in Hormonal Migraine: An Exploratory Study.

Background: Migraineurs, particularly young premenopausal women, are at increased risk of cerebrovascular disease; however, there is currently limited evidence as to whether hormonal migraine is associated with poor cerebrovascular function. Objectives: The objectives of this study were to: (1) investigate the potential association of cerebrovascular function with hormonal migraine and (2) determine whether abnormalities of cerebrovascular function in hormonal migraineurs are associated with migraine-related disability and/or quality of life. Method: A cross-sectional study was undertaken in 50 hormonal migraineurs (mean age: 38.7 ± 1.2 years) and 29 controls (mean age: 35.6 ± 1.8 years). Data were collected at a single point in time from all participants during the inter-ictal period when they were free from migraine and not menstruating. Transcranial Doppler ultrasound was used to measure resting blood flow velocity and cerebrovascular responsiveness (CVR) to hypercapnia and cognitive stimulation (neurovascular coupling) in the left and right middle cerebral artery (MCA). Additionally, hormonal migraineurs completed three questionnaires to assess migraine-related disability and quality of life as well as migraine frequency and intensity: Headache Impact Test-6™, Migraine-Specific Quality of Life and Migraine Disability Assessment. Results: Hormonal migraineurs had lower resting mean blood flow velocity (MBFV) (P = 0.009) and neurovascular coupling during cognitive stimulation (P = 0.010) in the left MCA than controls. No such differences were found in the right MCA. Additionally, heart rate (P = 0.004) was higher in hormonal migraineurs than controls. However, no differences in CVR to hypercapnia were found between hormonal migraineurs and controls. Multi-variate analysis revealed age to be a significant (P = 0.012) predictor of MBFV in the left MCA. Negative correlations between headache frequency and CVR to hypercapnia in the left (P = 0.026) and right MCA (P = 0.044) were found. Additionally, negative correlations between neurovascular coupling during the 2-Back 1.5 s task in the right MCA and the MSQoL emotional (P = 0.013) and role-function restrictive (P = 0.039) domains were found. Conclusions: This is the first study to show that hormonal migraineurs have poorer cerebrovascular function, as represented by lower resting MBFV and impaired neurovascular coupling in the left MCA. Future studies should investigate whether improving cerebrovascular function can prevent hormonal migraine and improve quality of life. Clinical Trial Registration: ACTRN12618001230246.

Differences in proliferation rate between CADASIL and control vascular smooth muscle cells are related to increased TGFß expression.

Cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) is a familial fatal progressive degenerative disorder. One of the pathological hallmarks of CADASIL is a dramatic reduction of vascular smooth muscle cells (VSMCs) in cerebral arteries. Using VSMCs from the vasculature of the human umbilical cord, placenta and cerebrum of CADASIL patients, we found that CADASIL VSMCs had a lower proliferation rate compared to control VSMCs. Exposure of control VSMCs and endothelial cells (ECs) to media derived from CADASIL VSMCs lowered the proliferation rate of all cells examined. By quantitative RT-PCR analysis, we observed increased Transforming growth factor-ß (TGFß) gene expression in CADASIL VSMCs. Adding TGFß-neutralizing antibody restored the proliferation rate of CADASIL VSMCs. We assessed proliferation differences in the presence or absence of TGFß-neutralizing antibody in ECs co-cultured with VSMCs. ECs co-cultured with CADASIL VSMCs exhibited a lower proliferation rate than those co-cultured with control VSMCs, and neutralization of TGFß normalized the proliferation rate of ECs co-cultured with CADASIL VSMCs. We suggest that increased TGFß expression in CADASIL VSMCs is involved in the reduced VSMC proliferation in CADASIL and may play a role in situ in altered proliferation of neighbouring cells in the vasculature.

Role of the Long Non-Coding RNA MAPT-AS1 in Regulation of Microtubule Associated Protein Tau (MAPT) Expression in Parkinson's Disease.

Studies investigating the pathogenic role of the microtubule associated protein tau (MAPT) gene in Parkinson's disease (PD) have indicated that DNA methylation of the promoter region is aberrant in disease, leading to dysregulated MAPT expression. We examined two potential regulators of MAPT gene expression in respect to PD, a promoter-associated long non-coding RNA MAPT-AS1, and DNA methyltransferases (DNMTs), enzymes responsible for new and maintenance of DNA methylation. We assessed the relationship between expression levels of MAPT and the candidate MAPT-AS1, DNMT1, DNMT3A and DNMT3B transcripts in four brain regions with varying degrees of cell loss and pathology (putamen, anterior cingulate cortex, visual cortex and cerebellum) in N = 10 PD and N = 10 controls. We found a significant decrease in MAPT-AS1 expression in PD (p = 7.154 x 10-6). The transcript levels of both MAPT-AS1 (p = 2.569 x 10-4) and DNMT1 (p = 0.001) correlated with those of MAPT across the four brain regions, but not with each other. Overexpression of MAPT-AS1 decreased MAPT promoter activity by ∼2.2 to 4.3 fold in an in vitro luciferase assay performed in two cell lines (p ≤ 2.678 x 10-4). Knock-down expression of MAPT-AS1 led to a 1.3 to 6.3 fold increase in methylation of the endogenous MAPT promoter (p ≤ 0.011) and a 1.2 to 1.5 fold increased expression of the 4-repeat MAPT isoform transcript (p ≤ 0.013). In conclusion, MAPT-AS1 and DNMT1 have been identified as potential epigenetic regulators of MAPT expression in PD across four different brain regions. Our data also suggest that increased MAPT expression could be associated with disease state, but not with PD neuropathology severity.

Effect of PSEN1 mutations on MAPT methylation in early-onset Alzheimer's disease.

The MAPT gene is a risk locus for multiple neurodegenerative diseases, including idiopathic Parkinson's and Alzheimer's disease. We examined whether altered DNA methylation of the MAPT promoter, with its potential to modulate gene expression, was a common phenomenon in Alzheimer's disease patients with differing aetiologies. We measured MAPT promoter methylation in a brain tissue cohort of early-onset Alzheimer's disease (EOAD) with defined causative mutations in the PSEN1 gene (Normal = 10, PSEN1 AD = 10), and idiopathic late-onset Alzheimer's disease (Normal = 12, LOAD = 12). We found a brain-region-specific decrease in MAPT promoter methylation in PSEN1 AD patients. Overexpression of PSEN1 reduced MAPT promoter activity in an in vitro luciferase study, and led to an increase in methylation of the endogenous MAPT promoter. Overexpression of PSEN1 with a deletion of exon 9 mutation (Δex9) led to a smaller reduction in MAPT promoter activity relative to wild-type PSEN1 in the luciferase assay, consistent with a decreased ability to modulate endogenous MAPT gene methylation. Our study indicates a novel effect of PSEN1 on MAPT methylation, and suggests a mutation-specific effect of the PSEN1 Δex9 mutation.

DNA methylation of the MAPT gene in Parkinson's disease cohorts and modulation by vitamin E in vitro.

Parkinson's disease (PD) is a neurodegenerative disorder for which environmental factors influence disease risk and may act via an epigenetic mechanism. The microtubule-associated protein tau (MAPT) is a susceptibility gene for idiopathic PD. Methylation levels were determined by pyrosequencing of bisulfite-treated DNA in a leukocyte cohort (358 PD patients and 1084 controls) and in two brain cohorts (Brain1, comprising 69 cerebellum controls; and Brain2, comprising 3 brain regions from 28 PD patients and 12 controls). In vitro assays involved the transfection of methylated promoter-luciferase constructs or treatment with an exogenous micronutrient. In normal leukocytes, the MAPT H1/H2 diplotype and sex were predictors of MAPT methylation. Haplotype-specific pyrosequencing confirmed that the H1 haplotype had higher methylation than the H2 haplotype in normal leukocytes and brain tissues. MAPT methylation was negatively associated with MAPT expression in the Brain1 cohort and in transfected cells. Methylation levels differed between three normal brain regions (Brain2 cohort, putamen < cerebellum < anterior cingulate cortex). In PD samples, age at onset was positively associated with MAPT methylation in leukocytes. Moreover, there was hypermethylation in the cerebellum and hypomethylation in the putamen of PD patients compared with controls (Brain2 cohort). Finally, leukocyte methylation status was positively associated with blood vitamin E levels, and the effect was more significant in H2 haplotype carriers; this result was confirmed in cells that were exposed to 100 µM vitamin E. The significant effects of sex, diplotype, and brain region suggest that hypermethylation of the MAPT gene is neuroprotective by reducing MAPT expression. The effect of vitamin E on MAPT represents a possible gene-environment interaction.

Sigma nonopioid intracellular receptor 1 mutations cause frontotemporal lobar degeneration-motor neuron disease.

OBJECTIVE: Frontotemporal lobar degeneration (FTLD) is the most common cause of early-onset dementia. Pathological ubiquitinated inclusion bodies observed in FTLD and motor neuron disease (MND) comprise trans-activating response element (TAR) DNA binding protein (TDP-43) and/or fused in sarcoma (FUS) protein. Our objective was to identify the causative gene in an FTLD-MND pedigree with no mutations in known dementia genes. METHODS: A mutation screen of candidate genes, luciferase assays, and quantitative polymerase chain reaction (PCR) was performed to identify the biological role of the putative mutation. Neuropathological characterization of affected individuals and western blot studies of cell lines were performed to identify the pathological mechanism of the mutation. RESULTS: We identified a nonpolymorphic mutation (c.672*51G>T) in the 3'-untranslated region (UTR) of the Sigma nonopioid intracellular receptor 1 (SIGMAR1) gene in affected individuals from the FTLD-MND pedigree. The c.672*51G>T mutation increased gene expression by 1.4-fold, corresponding with a significant 1.5-fold to 2-fold change in the SIGMAR1 transcript or Sigma-1 protein in lymphocyte or brain tissue. Brains of SIGMAR1 mutation carriers displayed a unique pathology with cytoplasmic inclusions immunopositive for either TDP-43 or FUS but not Sigma-1. Overexpression of SIGMAR1 shunted TDP-43 and FUS from the nucleus to the cytoplasm by 2.3-fold and 5.2-fold, respectively. Treatment of cells with Sigma-1 ligands significantly altered translocation of TDP-43 by up to 2-fold. INTERPRETATION: SIGMAR1 is a causative gene for familial FTLD-MND with a unique neuropathology that differs from other FTLD and MND cases. Our findings also suggest Sigma-1 drugs as potential treatments for the TDP-43/FUS proteinopathies.