Tony Aerts

Neuropsychological and behavioural correlates of CSF biomarkers in dementia.

To improve clinical, neuropsychological and behavioural characterisation of the cerebrospinal fluid (CSF) biomarkers beta-amyloid((1-42)) protein (Abeta42), protein tau (tau) and tau phosphorylated at threonine 181 (P-tau181) across diagnostic dementia categories, a prospective study was set up. Patients with probable Alzheimers disease (AD) (n=201), AD with cerebrovascular disease (CVD) (AD+CVD) (n=33), frontotemporal dementia (FTD) (n=27), dementia with Lewy bodies (DLB) (n=22) and healthy controls (n=148) were included. All patients underwent neuropsychological examination and behavioural assessment by means of a battery of behavioural assessment scales. CSF was obtained by lumbar puncture and levels of Abeta42, tau and P-tau181 were determined with commercially available ELISA kits. Negative correlations between CSF Abeta42 levels and aggressiveness (Spearman: r=-0.223; p=0.002) and positive correlations with age at inclusion (r=0.195; p=0.006), age at onset (r=0.205; p=0.003) and MMSE scores (r=0.198; p=0.005) were found in AD. In AD+CVD, CSF Abeta42 levels were correlated with MMSE (r=0.482; p=0.006), Hierarchic Dementia Scale (r=0.503; p=0.017) and Boston Naming Test (r=0.516; p=0.012) scores. In controls, age was positively correlated with CSF tau (r=0.465; p<0.001) and P-tau181 levels (r=0.312; p<0.001). CSF tau and P-tau181 levels correlated significantly in all groups, whereas CSF Abeta42 correlated with tau and P-tau181 levels in healthy controls only. Negative correlations between CSF Abeta42 levels and aggressiveness were found in AD patients. CSF Abeta42 seems to be a stage marker for AD (+/-CVD) given the positive correlations with neuropsychological test results suggesting that CSF Abeta42 might be of help for monitoring disease progression. Different correlations between age and CSF biomarker levels were obtained in healthy controls compared to AD patients, indicating that AD-induced pathophysiological processes change age-dependent regulation of CSF biomarker levels.

Light scattering by bovine alpha-crystallin proteins in solution: hydrodynamic structure and interparticle interaction

We have studied diluted bovine eye lens alpha-crystallin solutions by using light scattering. The protein particles were modeled as hard spheres, showing electrostatic repulsion, due to surplus electric charges, and weak attractive interaction. The repulsive potential VR is defined by the radius of the particles, the Debye length kappa-1, and the number of charges at the Gouy layer; the attractive potential has been described by the London-van der Waals potential and is defined by the Hamaker constant A. We have used the diluted gas approximation and the one component macrofluid model to relate the experimental static factor Ki to the theoretical expression of the interaction potential V(x). This resulted in a Hamaker constant A of 0.06 +/- 0.01 KBT and an effective charge q ranging from 18 +/- 1 at low ionic strength (omega = 0.0022 M) to 50 +/- 5 at high ionic strength (omega = 0.1472 M).

STRUCTURAL BASIS OF EYE LENS TRANSPARENCY : LIGHT SCATTERING BY CONCENTRATED SOLUTIONS OF BOVINE ALPHA -CRYSTALLIN PROTEINS

Short range order of the crystallins does account for the transparency of the eye lens. To explain the solution structure of this highly concentrated protein solution on a quantitative basis, the hydrodynamic structure and the interparticle interactions of the proteins have to be known. For that purpose, the light scattering of concentrated solutions of alpha-crystallin has been studied. Starting from the detailed knowledge of the solution parameters of alpha-crystallin in diluted solutions, the structure of concentrated solutions up to 360 mg/ml has been studied using light scattering. Our results indicate that subtle changes in the macromolecular structure such as optical anisotropy or structural asymmetry for part of the alpha-crystallins, which results in solute light-scattering heterogeneity, can dramatically increase the light scattering by the alpha-crystallins and cause solution opacity.

Study of the Chaperoning Mechanism of Bovine Lens α-Crystallin, a Member of the α-Small Heat Shock Superfamily

Abstract We have studied the interaction between lysozyme, destabilized by reducing its -S-S- bonds, and bovine eye lens α -crystallin, a member of the α -small heat shock protein superfamily. We have used gel filtration, photon correlation spectroscopy, and analytical ultracentrifugation to study the binding of lysozyme by α -crystallin at 25°C and 37°C. We can conclude that α -crystallin chaperones the destabilized protein in a two-step process. First the destabilized proteins are bound by the α -crystallin so that nonspecific aggregation of the destabilized protein is prevented. This complex is unstable, and a reorganization and inter-particle exchange of the peptides result in stable and soluble large particles. α -Crystallin does not require activation by temperature for the first step of its chaperone activity as it prevents the formation of nonspecific aggregates at 25°C as well as at 37°C. The reorganization of the peptides, however, gives rise to smaller particles at 37°C than at 25°C. Indirect evidence shows that the association of several α -crystallin/substrate protein complexes leads to the formation of very large particles. These are responsible for the increase of the light scattering.

Quaternary structure of bovine α-crystallin: influence of temperature

Abstract The tertiary and quaternary structure of α -crystallin is still a matter of controversy. We have characterized the native α -crystallin quaternary structure by isolating it at the in vivo temperature and solvent conditions. It can be represented by a distribution of expanded particles with a weight average molar mass of 550 000 g/mol. On decreasing (to 4°C) or increasing (up to 50°C) the temperature, the size distribution increases to larger particles. Only at lower temperatures (4°C), a stable population of particles is obtained with weight average molar mass of 700 000 g/mol. In all conditions, α -crystallin behaves as a very expanded particle with a maximum hydrodynamic volume of 3.15 ml/g. The transitions in quaternary structure are rather slow: it takes several hours to evolve from a population of aggregates, characteristic for given solvent conditions, to another distribution in size and quaternary structure on changing the environment. The quaternary structure of α -crystallin is an uncharacteristic parameter of the particle: a broad distribution of values can be obtained on changing the environment. Any realistic model should include this property. Our studies favor an open loose structure, where peptides can be added or removed without drastic changes of secondary and tertiary structure of the peptides.

Brain Region-Specific Monoaminergic Correlates of Neuropsychiatric Symptoms in Alzheimer's Disease

Neuropsychiatric symptoms (NPS) in Alzheimers disease (AD) are present during the disease course of nearly all AD patients and consist of psychosis, agitation/aggression, and depression, among others. Given their detrimental consequences regarding life expectancy, cognition, and socio-economic costs, it is essential to elucidate their neurochemical etiology to facilitate the development of novel and effective pharmacotherapeutics. This study attempted to identify brain region-specific monoaminergic correlates of NPS by measuring the levels of eight monoamines and metabolites in nine relevant postmortem brain regions of 40 behaviorally characterized AD patients, i.e., dopamine (DA), serotonin (5-HT), (nor)epinephrine and their respective metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid, 5-hydroxy-3-indoleacetic acid (5-HIAA), and 3-methoxy-4-hydroxyphenylglycol (MHPG), using RP-HPLC-ECD. Likewise, Mini-Mental State Examination (MMSE) score correlates of monoaminergic neurotransmitter alterations were calculated. As a result, MMSE scores, used as a measure of dementia severity, correlated positively with hippocampal 5-HIAA levels as well as with 5-HT levels of the superior temporal gyrus and cerebellar cortex. Furthermore, hippocampal 5-HIAA levels inversely correlated with agitation scores, whereas thalamic MHPG levels comparably did with the presence of hallucinations. Finally, in the cerebellar cortex, DOPAC/DA ratios, indicative of DA turnover, correlated with physically agitated behavior while MHPG levels correlated with affective disturbances. These findings support the assumption that specific NPS features in AD might be (in)directly related to brain region-specific monoaminergic neurotransmitter alterations. Additionally, the effect of AD pathology on neurochemical alterations in the cerebellum requires further examination due to its important but underestimated role in the neurochemical pathophysiology of NPS in AD.

Novel and sensitive reversed-phase high-pressure liquid chromatography method with electrochemical detection for the simultaneous and fast determination of eight biogenic amines and metabolites in human brain tissue

A fast and simple RP-HPLC method with electrochemical detection (ECD) and ion pair chromatography was developed, optimized and validated in order to simultaneously determine eight different biogenic amines and metabolites in post-mortem human brain tissue in a single-run analytical approach. The compounds of interest are the indolamine serotonin (5-hydroxytryptamine, 5-HT), the catecholamines dopamine (DA) and (nor)epinephrine ((N)E), as well as their respective metabolites, i.e. 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), 5-hydroxy-3-indoleacetic acid (5-HIAA) and 3-methoxy-4-hydroxyphenylglycol (MHPG). A two-level fractional factorial experimental design was applied to study the effect of five experimental factors (i.e. the ion-pair counter concentration, the level of organic modifier, the pH of the mobile phase, the temperature of the column, and the voltage setting of the detector) on the chromatographic behaviour. The cross effect between the five quantitative factors and the capacity and separation factors of the analytes were then analysed using a Standard Least Squares model. The optimized method was fully validated according to the requirements of SFSTP (Société Française des Sciences et Techniques Pharmaceutiques). Our human brain tissue sample preparation procedure is straightforward and relatively short, which allows samples to be loaded onto the HPLC system within approximately 4h. Additionally, a high sample throughput was achieved after optimization due to a total runtime of maximally 40min per sample. The conditions and settings of the HPLC system were found to be accurate with high intra and inter-assay repeatability, recovery and accuracy rates. The robust analytical method results in very low detection limits and good separation for all of the eight biogenic amines and metabolites in this complex mixture of biological analytes.

The monoaminergic footprint of depression and psychosis in dementia with Lewy bodies compared to Alzheimer’s disease

IntroductionDepression and psychosis are two of the most severe neuropsychiatric symptoms (NPS) in dementia with Lewy bodies (DLB) and Alzheimer’s disease (AD). Both NPS have negative effects on cognitive performance and life expectancy. The current study aimed to investigate and compare monoaminergic etiologies between both neurodegenerative conditions, given the lack of an efficient pharmacological treatment until present.MethodsEleven behaviorally relevant brain regions of the left frozen hemisphere of 10 neuropathologically confirmed AD patients with/without depression (AD + D/-D; 5 were psychotic within AD + D), 10 confirmed DLB patients, all of whom were depressed (DLB + D; 5 psychotic patients), and, finally, 10 confirmed control subjects were regionally dissected. All patients were retrospectively assessed before death using the Behavioral Pathology in Alzheimer’s Disease Rating Scale (Behave-AD) and Cornell Scale for Depression in Dementia amongst others. The concentrations of dopamine (DA), serotonin (5-HT), (nor)adrenaline and respective metabolites, i.e. 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), 5-hydroxy-3-indoleacetic acid (5-HIAA), and, 3-methoxy-4-hydroxyphenylglycol (MHPG), were determined using reversed-phase high-performance liquid chromatography with electrochemical detection.ResultsDLB subjects had the overall lowest monoamine and metabolite concentrations regarding 33 out of 41 significant monoaminergic group alterations. Moreover, MHPG levels were significantly decreased in almost 8 out of 11 brain regions of DLB- compared to AD patients. We also observed the lowest 5-HT and 5-HIAA levels, and 5-HIAA/5-HT turnover ratios in DLB + D compared to AD + D subjects. Additionally, a 4- and 7-fold increase of DOPAC/DA and HVA/DA turnover ratios, and, a 10-fold decrease of thalamic DA levels in DLB + D compared to AD + D patients and control subjects was noticed. Regarding psychosis, hippocampal DA levels in the overall DLB group significantly correlated with Behave-AD AB scores. In the total AD group, DA levels and HVA/DA ratios in the amygdala significantly correlated with Behave-AD AB scores instead.ConclusionsMonoaminergic neurotransmitter alterations contribute differently to the pathophysiology of depression and psychosis in DLB as opposed to AD, with a severely decreased serotonergic neurotransmission as the main monoaminergic etiology of depression in DLB. Similarly, psychosis in DLB might, in part, be etiologically explained by dopaminergic alterations in the hippocampus, whereas in AD, the amygdala might be involved.

Serum MHPG Strongly Predicts Conversion to Alzheimer's Disease in Behaviorally Characterized Subjects with Down Syndrome.

BACKGROUND Down syndrome (DS) is the most prevalent genetic cause of intellectual disability. Early-onset Alzheimers disease (AD) frequently develops in DS and is characterized by progressive memory loss and behavioral and psychological signs and symptoms of dementia (BPSD). Predicting and monitoring the progression of AD in DS is necessary to enable adaptive caretaking. OBJECTIVE Reliable blood biomarkers that aid the prediction of AD are necessary, since cerebrospinal fluid sampling is rather burdensome, particularly for people with DS. Here, we investigate serum levels of eight biogenic amines and their metabolites in relation to dementia staging and probable BPSD items. METHODS Using RP-HPLC with electrochemical detection, (nor)adrenergic (NA/A and MHPG), serotonergic (5-HT and 5-HIAA), and dopaminergic (DA, HVA, and DOPAC) compounds were quantified in the serum of DS subjects with established AD at baseline (n = 51), DS subjects without AD (n = 50), non-demented DS individuals that converted to AD over time (n = 50), and, finally, healthy non-DS controls (n = 22). RESULTS Serum MHPG levels were significantly lower in demented and converted DS subjects (p < 0.0001) compared to non-demented DS individuals and healthy controls. Those subjects with MHPG levels below median had a more than tenfold increased risk of developing dementia. Furthermore, significant correlations were observed between monoaminergic serum values and various probable BPSD items within each DS group. CONCLUSION Decreased serum MHPG levels show great potential as biomarker to monitor and predict conversion to AD in DS. Moreover, significant monoaminergic alterations related to probable BPSD items, suggesting that monoaminergic dysregulation is an underlying biological mechanism, and demonstrating the need to develop a validated rating scale for BPSD in DS.

Physicochemical studies on bovine eye lens proteins II. Comparative physical study of the low-molecular-weight α-crystallins from calf lens cortical and nuclear fiber cells

The alpha L of cortical and nuclear fiber cells have been studied using hydrodynamical and physicochemical techniques. From the sedimentation and the diffusion coefficients in identical conditions, it can be concluded that alpha L,N is appreciably larger than alpha L,C but both have a similar structure in solution: a spherical particle with a high hydration. The alpha L,N not only contains several degraded alpha A- and alpha B-peptides but also a typical pattern of beta-peptides. The fluorescence spectrum indicates a shift of the hydrophobic tryptophan residues from a hydrophobic environment in alpha L,C to a more solvent-exposed and polar neighbourhood for alpha L,N. Also solubility studies on alpha L,C and alpha L,N in different solvent conditions and temperatures, indicate more apolar interactions between the peptides of the nuclear alpha L, than its cortical counterpart. The more hydrophobic interaction pattern of the peptides in alpha L,N can also be reconciled with a lower mean hydration potential, indicative of a higher hydrophobicity of the degraded alpha A-peptides.