Faculty Bibliography

The increase in the incidence of neurodegenerative diseases, in particular Alzheimer's Disease (AD), is a consequence of the world's population aging but unfortunately, existing treatments are only effective at delaying some of the symptoms and for a limited time. Despite huge efforts by both academic researchers and pharmaceutical companies, no disease-modifying drugs have been brought to the market in the last decades. Recently, several studies shed light on Carbonic Anhydrases (CAs, EC 4.2.1.1) as possible new targets for AD treatment. In the present review we summarized preclinical and clinical findings regarding the role of CAs and their inhibitors/activators on cognition, aging and neurodegeneration and we discuss future challenges and opportunities in the field.

Background: Alzheimer's disease (AD) is thought to involve a cerebrovascular component and AD shares many risk factors with cardiovascular diseases. Amyloid beta (Abeta) is neurotoxic and damages brain microcirculation. Abeta production is up-regulated by hypoxia. Oxygen availability and extraction not only depends on the increase in cerebral blood flow (CBF) but also on the decrease of capillary transit-time heterogeneity (CTH). Structural or functional damage caused to the brain microcirculation will reduce the oxygen availability by a reduction in CBF and increased CTH (capillary dysfunction). Carbonic anhydrase inhibitors (CAIs) have shown to reduce the Abeta neurovascular mitochondrial toxicity, but the effect of this treatment in brain hemodynamics has not been evaluated. The present work aims to investigate the effect of long-term treatment with carbonic anhydrase inhibitors (CAIs) on the neurovascular response in Tg-SweDI mice.
Method(s): All experiments were approved by the Danish Animal Inspectorate. Tg-SwDI mice (4 months old) were fed for 140 - 150 days with a diet supplemented with acetazolamide (G1), methazolamide (G2) or no medication (G3). The groups were randomized and researchers were blind during the data acquisition. A control group of wild-type (WT; G4) mice was included. We performed imaging in awake head-restrained TgSwDI and WT mice through a cranial window placed onto the somatosensory area of the barrel cortex. We estimated relative changes in brain hemodynamics during whiskers stimulation (10 s). We estimated CBF and cerebral blood volume (rCBV) using laser Doppler flowmetry and optical intrinsic signal imaging, respectively. Two-photon imaging was employed to estimate intravascular oxygen partial pressure (ptO2), mean transit-time (MTT), CTH and capillary hemodynamics.
Result(s): Our preliminary analysis in awake WT mice (G4; N = 7) shows that functional activation produced an increase in regional CBF and CBV of 9.41 +/-3.7% and 1.49+/-0.35%, respectively. During activation, MTT and CTH respectively decreased 14.06 +/-5.92% and 23.28 +/- 9.60%. Arterial and venous ptO2 increased by 1.33 +/- 0.33% and 2.60 +/-0.56%. The estimated OEF showed a decrease of 12.4% +/-2.9%. A blinded analysis is currently assessed to evaluate the effect of CAIs on the brain hemodynamic response to functional activation.
Conclusion(s): Our work describes the signature of capillary dysfunction in an AD mouse model. Our results also enable us to elucidate the effect of the carbonic anhydrase inhibitors in capillary dysfunction in AD

A mechanistic understanding of the pathology of psychiatric disorders has been hampered by extensive heterogeneity in biology, symptoms, and behavior within diagnostic categories that are defined subjectively. We investigated whether leveraging individual differences in information-processing impairments in patients with post-traumatic stress disorder (PTSD) could reveal phenotypes within the disorder. We found that a subgroup of patients with PTSD from two independent cohorts displayed both aberrant functional connectivity within the ventral attention network (VAN) as revealed by functional magnetic resonance imaging (fMRI) neuroimaging and impaired verbal memory on a word list learning task. This combined phenotype was not associated with differences in symptoms or comorbidities, but nonetheless could be used to predict a poor response to psychotherapy, the best-validated treatment for PTSD. Using concurrent focal noninvasive transcranial magnetic stimulation and electroencephalography, we then identified alterations in neural signal flow in the VAN that were evoked by direct stimulation of that network. These alterations were associated with individual differences in functional fMRI connectivity within the VAN. Our findings define specific neurobiological mechanisms in a subgroup of patients with PTSD that could contribute to the poor response to psychotherapy.

White matter hyperintensities (WMHs) are frequently seen on brain magnetic resonance imaging scans of older people. Usually interpreted clinically as a surrogate for cerebral small vessel disease, WMHs are associated with increased likelihood of cognitive impairment and dementia (including Alzheimer's disease [AD]). WMHs are also seen in cognitively healthy people. In this collaboration of academic, clinical, and pharmaceutical industry perspectives, we identify outstanding questions about WMHs and their relation to cognition, dementia, and AD. What molecular and cellular changes underlie WMHs? What are the neuropathological correlates of WMHs? To what extent are demyelination and inflammation present? Is it helpful to subdivide into periventricular and subcortical WMHs? What do WMHs signify in people diagnosed with AD? What are the risk factors for developing WMHs? What preventive and therapeutic strategies target WMHs? Answering these questions will improve prevention and treatment of WMHs and dementia.

Alzheimer's disease (AD) is the most prevalent form of dementia. Cerebrovascular dysfunction is one of the earliest events in the pathogenesis of AD, as well as in vascular and mixed dementias. Cerebral amyloid angiopathy (CAA), the deposition of amyloid around cerebral vessels, is observed in up to 90% of AD patients and in approximately 50% of elderly individuals over 80 years of age. CAA is a strong contributor to vascular dysfunction in AD. CAA-laden brain vessels are characterized by dysfunctional hemodynamics and leaky blood-brain barrier (BBB), contributing to clearance failure and further accumulation of amyloid-β (Aβ) in the cerebrovasculature and brain parenchyma. Mitochondrial dysfunction is increasingly recognized as an important early initiator of the pathogenesis of AD and CAA. The objective of this review is to discuss the effects of Aβ on cerebral microvascular cell function, focusing on its impact on endothelial mitochondria. After introducing CAA and its etiology and genetic risk factors, we describe the pathological relationship between cerebrovascular amyloidosis and brain microvascular endothelial cell dysfunction, critically analyzing its roles in disease progression, hypoperfusion, and BBB integrity. Then, we focus on discussing the effect of Aβ challenge on endothelial mitochondrial dysfunction pathways, and their contribution to the progression of neurovascular dysfunction in AD and dementia. Finally, we report potential pharmacological and non-pharmacological mitochondria-targeted therapeutic strategies which may help prevent or delay cerebrovascular failure.

Mounting evidence suggests that mitochondrial dysfunction plays a causal role in the etiology and progression of Alzheimer's disease (AD). We recently showed that the carbonic anhydrase inhibitor (CAI) methazolamide (MTZ) prevents amyloid β (Aβ)-mediated onset of apoptosis in the mouse brain. In this study, we used MTZ and, for the first time, the analog CAI acetazolamide (ATZ) in neuronal and cerebral vascular cells challenged with Aβ, to clarify their protective effects and mitochondrial molecular mechanism of action. The CAIs selectively inhibited mitochondrial dysfunction pathways induced by Aβ, without affecting metabolic function. ATZ was effective at concentrations 10 times lower than MTZ. Both MTZ and ATZ prevented mitochondrial membrane depolarization and H₂ O₂ generation, with no effects on intracellular pH or ATP production. Importantly, the drugs did not primarily affect calcium homeostasis. This work suggests a new role for carbonic anhydrases (CAs) in the Aβ-induced mitochondrial toxicity associated with AD and cerebral amyloid angiopathy (CAA), and paves the way to AD clinical trials for CAIs, FDA-approved drugs with a well-known profile of brain delivery.

Traumatic brain injury (TBI) and Alzheimer's disease (AD) are devastating neurological disorders, whose complex relationship is not completely understood. Cerebrovascular pathology, a key element in both conditions, could represent a mechanistic link between Aβ/tau deposition after TBI and the development of post concussive syndrome, dementia and chronic traumatic encephalopathy (CTE). In addition to debilitating acute effects, TBI-induced neurovascular injuries accelerate amyloid β (Aβ) production and perivascular accumulation, arterial stiffness, tau hyperphosphorylation and tau/Aβ-induced blood brain barrier damage, giving rise to a deleterious feed-forward loop. We postulate that TBI can initiate cerebrovascular pathology, which is causally involved in the development of multiple forms of neurodegeneration including AD-like dementias. In this review, we will explore how novel biomarkers, animal and human studies with a focus on cerebrovascular dysfunction are contributing to the understanding of the consequences of TBI on the development of AD-like pathology.

Cerebrospinal fluid (CSF) studies consistently show that CSF levels of amyloid-beta 1-42 (Aβ42) are reduced and tau levels increased prior to the onset of cognitive decline related to Alzheimer's disease (AD). However, the preclinical prediction accuracy for low CSF Aβ42 levels, a surrogate for brain Aβ42 deposits, is not high. Moreover, the pathology data suggests a course initiated by tauopathy contradicting the contemporary clinical view of an Aβ initiated cascade. CSF Aβ42 and tau data from 3 normal aging cohorts (45-90 years) were combined to test both cross-sectional (n = 766) and longitudinal (n = 651) hypotheses: 1) that the relationship between CSF levels of Aβ42 and tau are not linear over the adult life-span; and 2) that non-linear models improve the prediction of cognitive decline. Supporting the hypotheses, the results showed that a u-shaped quadratic fit (Aβ2) best describes the relationship for CSF Aβ42 with CSF tau levels. Furthermore we found that the relationship between Aβ42 and tau changes with age-between 45 and 70 years there is a positive linear association, whereas between 71 and 90 years there is a negative linear association between Aβ42 and tau. The quadratic effect appears to be unique to Aβ42, as Aβ38 and Aβ40 showed only positive linear relationships with age and CSF tau. Importantly, we observed the prediction of cognitive decline was improved by considering both high and low levels of Aβ42. Overall, these data suggest an earlier preclinical stage than currently appreciated, marked by CSF elevations in tau and accompanied by either elevations or reductions in Aβ42. Future studies are needed to examine potential mechanisms such as failing CSF clearance as a common factor elevating CSF Aβxx analyte levels prior to Aβ42 deposition in brain.

Evidence supporting the hypothesis that reduced cerebrospinal fluid (CSF) clearance is involved in the pathophysiology of Alzheimer's disease (AD) comes from primarily from rodent models. However, unlike rodents where predominant extra-cranial CSF egress is via olfactory nerves traversing the cribriform plate, human CSF clearance pathways are not well characterized. Using dynamic Positron Emission Tomography (PET) with 18F-THK5117 a tracer for tau pathology, the ventricular CSF time activity was used as a biomarker for CSF clearance. We tested three hypotheses: 1. Extra-cranial CSF is detected at the superior turbinates; 2. CSF clearance is reduced in AD; and 3. CSF clearance is inversely associated with amyloid deposition. Methods: 15 subjects, 8 with AD and 7 normal control volunteers were examined with 18F-THK5117. 10 subjects additionally received 11C-PiB PET scans and 8 were PiB positive. Ventricular time activity curves (TAC) of 18F-THK5117 were used to identify highly correlated TAC from extra-cranial voxels. Results: For all subjects, the greatest density of CSF positive extra-cranial voxels was in the nasal turbinates. Tracer concentration analyses validated the superior nasal turbinate CSF signal intensity. AD patients showed ventricular tracer clearance reduced by 23% and 66% fewer superior turbinate CSF egress sites. Ventricular CSF clearance was inversely associated with amyloid deposition. Conclusion: The human nasal turbinate is part of the CSF clearance system. Lateral ventricle and superior nasal turbinates CSF clearance abnormalities are found in AD. Ventricular CSF clearance reductions are associated with increased brain amyloid depositions. These data suggest that PET measured CSF clearance is a biomarker of potential interest in AD and other neurodegenerative diseases.

Background: It is now accepted that mitochondrial dysfunction is a key early event in the progression of neuronal and vascular degeneration in Alzheimer's disease (AD) and that therapies aimed at preventing mitochondrial failure may represent promising new strategies in the pursue of a cure for this devastating disease. Carbonic anhydrases (CAs) are a family of enzymes that catalyze the rapid interconversion of carbon dioxide and water to bicarbonate and protons (or vice versa), maintaining acid-base balance in blood and other tissues. CA isoforms are present in the mitochondria. CA inhibitors (CAIs), such as metazolamide (MTZ) and acetazolamide (ATZ) are clinically used for glaucoma, epilepsy (rarely), and high altitude sickness. Methods: We analyzed the effects of two main CAIs used in clinical settings (MTZ and ATZ) on the mechanism of mitochondrial damage and neurovascular degeneration induced by amyloid beta (Abeta), using cerebral vascular and neural cells as well as the TgSwDI (Swedish- Dutch-Iowa) transgenic mouse model of cerebral amyloidosis. Results: Both CAIs consistently prevented specific pathways of mitochondrial dysfunction induced by Abeta in cerebral microvascular endothelial, neuronal and glial cells, without affecting ATP production, pH, and Calcium flux. Increase of hydrogen peroxide, loss of mitochondrial membrane potential, release of Cytochrome C, caspase activation, and apoptotic cell death were inhibited by CAIs. ATZ was effective at concentrations lower than MTZ. Both drugs, given with diet, were able to ameliorate behavioral paradigms in relatively young TgSwDI mice. Conclusions: CAIs might represent a potentially successful strategy to prevent early mitochondrial dysfunction and neurovascular loss in AD. Further studies in animal models and clinical settings are needed to confirm our hypothesis