Current Alzheimer Research - Volume 10, Issue 7, 2013

Previous studies have investigated the activity and protein levels of BACE1, the β-secretase, in the brain and cerebrospinal fluid (CSF) of Alzheimer’s disease (AD) patients, however, results remain contradictory. We present here a highly specific and sensitive BACE1 ELISA, which allows measuring accurately BACE1 levels in human samples. We find that BACE1 levels in CSF of AD patients and other neurological disorder (OND) patients are slightly increased when compared to those of a non-neurological disorder control group (NND). BACE1 levels in CSF were well correlated with total-tau and hyperphosphorylated tau levels in the CSF, suggesting that the recorded alterations in BACE1 levels correlate with cell death and neurodegeneration.

The microtubule-associated protein (MAP) tau has been implicated in the pathology of numerous neurodegenerative diseases. In the past decade, the hyperphosphorylated and aggregated states of tau protein have been important targets in the drug discovery field for the potential treatment of Alzheimer’s disease. Although several compounds have been reported to reduce the hyperphosphorylated state of tau or impact the stabilization of tau, their therapeutic activities are remain to be validated. Recently, reduction of total cellular tau protein has emerged as an alternate intervention point for drug development and a potential treatment of tauopathies. We have developed and optimized homogenous assays, using the AlphaLISA and HTRF assay technologies, for the quantification of total cellular tau protein levels in the SH-SY5Y neuroblastoma cell line. The signal-to-basal ratios were 375 and 5.3, and the Z’ factors were 0.67 and 0.60 for the AlphaLISA and HTRF tau assays, respectively. The clear advantages of these homogeneous tau assays over conventional total tau assays, such as ELISA and Western blot, are the elimination of plate wash steps and miniaturization of the assay into 1536-well plate format for the ultra–high-throughput screening of large compound libraries.

The aggregation of beta-amyloid (Aβ) into plaques in the extracellular compartment of the brain is a pathological hallmark of Alzheimer’s disease (AD). Although the pathways for misprocessing of Aβ leading to plaque formation are not well understood, they may be related to synapse turnover and neuron activity. In this study, we have utilised transgenic mice co-expressing mutations in the amyloid precursor protein and presenilin 1 genes (APP/PS1) to determine how long-term denervation of the olfactory bulb, a CNS area affected early by AD-like pathology, may affect Aβ plaque formation. The olfactory bulb of pre-symptomatic mice was denervated by ablating the olfactory epithelium unilaterally with Triton X-100 solution. Mice were subjected to nasal washes for a total of 4 or 8 times, at 3-week intervals either with 1% Triton X-100 solution or phosphate buffered saline (sham denervation). Denervation of the olfactory bulb resulted in a statistically significant (p<0.05) decrease in amyloid plaque load in the ipsilateral olfactory bulb, and bilaterally also in the neocortex and hippocampus at 8-9 months age. Amyloid precursor protein was predominantly expressed by mitral cells in the olfactory bulb, which are normally postsynaptic to olfactory axons. The number of APP positive mitral cells was significantly increased in the denervated olfactory bulb of wild type but not of the APP/PS1 mice, which consistently showed high immunoreactivity for APP pre- and post-denervation. In summary, our results show that Aβ plaque deposition in the central nervous system can be modified transsynaptically by deafferentation.

The soluble amyloid β precursor protein α (sAβPPα) released after α-secretase cleavage of the amyloid β precursor protein (AβPP) has several functions including modulation of neuronal excitability and synaptic plasticity; it has been suggested that some of these effects are mediated by activation of NF-κB via induction of PI3K/Akt signaling pathway. We have recently described the presence of several consensus binding sites of c-Rel transcription factor in the promoter region of the GNB2L1 gene, coding for the Receptor for Activated C Kinase -1 (RACK-1). We investigated whether sAβPPα could influence the expression of RACK-1 through NF-κB involvement. Our data demonstrate that sAβPPα regulates RACK-1 gene expression through PI3K/Akt-dependent pathway, inducing c-Rel nuclear translocation and NF-κB activation. Since RACK-1 is the scaffold of protein kinase C βII (PKCβII), we turned our attention to this kinase in order to evaluate whether sAβPPα could also influence PKCβII signalling demonstrating that sAβPPα induces PKCβII translocation and interaction with its scaffold with consequent RACK-1/PKCβII complex increase in membrane. Altogether these results suggest the existence of an interesting loop between the functions of the metabolic products of AβPP and the role of PKC and that the impact of a dysregulated AβPP metabolism occurring in several conditions (from physiological aging to injury response) may have consequences on the potential protective functions of the non amyloidogenic sAβPPα.

Sequential cleavages of APP by β-secretase and γ-secretase release β-amyloid (Aβ) and one secreted form of APP (sAPP-β) in Alzheimer’s disease (AD). Alternatively, in non-pathological situations, APP is predominantly cleaved by α-secretase within the amyloid sequence, to release the other soluble form of APP, sAPP-α. However, the functions of the two types of sAPP are still unclear. We performed this study to compare the function of sAPP-α and sAPP-β in differentiation of the glioma cell line U251. We found that sAPP-α suppressed astrocytic differentiation and promoted neuronal differentiation in U251 cells. Additionally, sAPP-α enhanced U251 terminal differentiation into a cholinergic-like neuronal phenotype. In contrast, sAPP-β suppressed neuronal differentiation and promoted the astrocytic differentiation of U251 cells. These findings could not only enrich the knowledge of the potential physiological function of sAPP-α and sAPP-β, but also indicate that they may be connected to the pathological mechanism of AD. Furthermore, these findings suggest that new strategies, such as increasing the level of sAPP-α and/or decreasing the level of sAPP-β in brain, or transplanting stem cells with increased sAPP-α and/or decreased sAPP-β, may have potential value for AD treatment.

The cause and mechanism of development of Alzheimer’s disease (AD) remain unexplained. Hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis, denoted by adrenal cortisol hypersecretion, is a recognised feature of the condition but generally disregarded as causative, due to lack of association between AD and other hypercortisolemic states. However, a meta-analysis of published studies suggests a need for reappraisal. A specific circadian rhythm of cortisol hypersecretion pertains at mild-to-moderate AD stages, entailing increased levels at the circadian peak from a low nadir. This is in contrast to the continuously elevated levels that are characteristic of other hypercortisolemic states, e.g. Cushing’s disease or major depression. This previously overlooked detail provides a starting premise here: that equating the form of hypercortisolism in AD with that in other states is inappropriate, as phasic and chronic elevation elicit different neuroendocrine effects. Theoretical implications are discussed in this review. Given the capacity of glucocorticoids and corticotropin-releasing hormone to induce AD-associated pathologies, I suggest a role for circadian cortisol hypersecretion in the initiation of sporadic AD; and propose a temporal mechanism for AD development featuring neuroinflammation- mediated suppression of central glucocorticoid receptor (GR) signaling. This latter may represent a critical phase in AD development, where the density of functional GR is proposed to underlie the “cognitive reserve”. Supporting evidence for this mechanism is drawn from the brain regional locations of AD neuropathologies, and from risk factors for AD development (aging, ApoE-4 genotype, and hypertension). Thus, it is argued that basal hypercortisolemia merits further scrutiny regarding AD causation and development.

This paper investigates how commonly prescribed pharmacologic treatments for Alzheimer’s disease (AD) affect Event-Related Potential (ERP) biomarkers as tools for predicting AD conversion in individuals with Mild Cognitive Impairment (MCI). We gathered baseline ERP data from two MCI groups (those taking AD medications and those not) and later determined which subjects developed AD (Convert->AD) and which subjects remained cognitively stable (Stable). We utilized a previously developed and validated multivariate system of ERP components to measure medication effects among these four subgroups. Discriminant analysis produced classification scores for each individual as a measure of similarity to each clinical group (Convert->AD, Stable), and we found a large significant main Group effect but no main AD Medications effect and no Group by Medications interaction. This suggested AD medications have negligible influence on this set of ERP components as weighted markers of disease progression. These results provide practical information to those using ERP measures as a biomarker to identify and track AD in individuals in a clinical or research setting.

As neuroinflammation is an early event in the pathogenesis of Alzheimer’s disease, new selective antiinflammatory drugs could lead to promising preventive strategies. We evaluated the safety, tolerability, pharmacokinetics and pharmacodynamics of CHF5074, a new microglial modulator, in a 12-week, double-blind, placebo-controlled, parallel groups, ascending dose study involving 96 MCI patients. Subjects were allocated into three successive study cohorts to receive ascending, titrated doses of CHF5074 (200, 400 or 600 mg/day) or placebo. Vital signs, cardiac safety, neuropsychological performance and safety clinical laboratory parameters were assessed on all subjects. Plasma samples were collected throughout the study for measuring drug concentrations, soluble CD40 ligand (sCD40L) and TNF-α. At the end of treatment, cerebrospinal fluid (CSF) samples were optionally collected after the last dose to measure drug levels, β- amyloid1-42 (Aβ42), tau, phospho-tau181, sCD40L and TNF-α. Ten patients did not complete the study: one in the placebo group (consent withdrawn), two in the 200-mg/day treatment group (consent withdrawn and unable to comply) and seven in the 400-mg/day treatment group (five AEs, one consent withdrawn and one unable to comply). The most frequent treatment-emergent adverse events were diarrhea, dizziness and back pain. There were no clinically significant treatmentrelated clinical laboratory, vital sign or ECG abnormalities. CHF5074 total body clearance depended by gender, age and glomerular filtration rate. CHF5074 CSF concentrations increased in a dose-dependent manner. At the end of treatment, mean sCD40L and TNF-α levels in CSF were found to be inversely related to the CHF5074 dose (p=0.037 and p=0.001, respectively). Plasma levels of sCD40L in the 600-mg/day group were significantly lower than those measured in the placebo group (p=0.010). No significant differences between treatment groups were found in neuropsychological tests but a positive dose-response trend was found on executive function in APOE4 carriers. This study shows that CHF5074 is well tolerated in MCI patients after a 12-week titrated treatment up to 600 mg/day and dose-dependently affects central nervous system biomarkers of neuroinflammation.

Objectives: The current study evaluated whether the functional connectivity pattern of the thalamo-cortical network in patients with Alzheimer’s disease (AD) and mild cognitive impairment (MCI) would show disease severityrelated alterations. Methods: Resting-state functional magnetic resonance imaging (MRI) data were obtained from 35 patients with AD, 27 patients with MCI and 27 subjects with normal cognition (NC). First, the altered functional connectivity pattern in AD patients was evaluated in comparison to NC subjects. Second, the MCI subjects were included to evaluate how different stages of disease affect the functional connectivity pattern of the thalamus. Finally, a correlation analysis was performed between the strength of the functional connectivity of the identified regions and various clinical variables to evaluate the relationship between the strength of functional connectivity and the cognitive abilities of MCI and AD patients. Results: When compared to NC subjects, AD patients showed decreased functional connectivity between the left thalamus and brain regions including the precuneus/posterior cingulate cortex, right middle frontal gyrus and left inferior frontal gyrus. Decreased functional connectivity was also found between the right thalamus and right middle frontal gyrus and left inferior parietal lobule/angular gyrus. In addition, increased functional connectivity was observed between the bilateral thalamus and brain regions including the middle frontal gyrus, middle temporal gyrus, inferior temporal gyrus, superior parietal lobule, postcentral gyrus and precuneus. Functional connectivity between the bilateral thalamus and the identified brain regions of MCI subjects was intermediate in comparison to the functional connectivity of AD and NC subjects. A significant correlation between the fitted functional connectivity strength and the clinical variables was also detected. Conclusion: Our results revealed disease severity-related alterations of the thalamo-default mode network and thalamocortical connectivity in AD and MCI patients. These results support the hypothesis of network disconnection in AD.