Magdolna Pákáski

Interactions between the amyloid and cholinergic mechanisms in Alzheimer's disease

Alzheimers disease (AD) is a progressive, neurodegenerative disease characterized by memory and cognitive loss, the formation of senile plaques containing amyloid-beta (Abeta) peptide, degeneration of the cholinergic neurons and the development of neurofibrillary tangles. The build-up of Abeta is considered to be a central feature in the pathogenesis of AD. However, other critical molecular and neurochemical alterations too occur, such as a cholinergic dysfunction. As concerns the pathomechanism of the disease, both the amyloid cascade hypothesis and the cholinergic hypothesis of AD are widely accepted. This review surveys recent in vitro and in vivo experimental evidence relating to these two hypotheses. Bidirectional pathways linking them as regards the cholinergic neurotoxicity of Abeta and the regulatory mechanisms of cholinergic receptor activation or enzyme inhibition in the processing of the amyloid precursor protein are also discussed. Further work is warranted to elucidate the exact effects in the interactions between the cholinergic and amyloid hypotheses of the candidate drugs used in AD therapy.

Gene expression profile analysis of lymphocytes from Alzheimer's patients.

Since the function and metabolism of peripheral lymphocytes is known to be altered in Alzheimers disease (AD), a pilot study was carried out to examine differences in gene expression profiles of these cells in 16 AD patients and aged control probands. Using a cDNA microarray representing 3200 distinct human genes, we identified 20 candidate genes whose expression is altered in AD lymphocytes compared with the control probands. Among these were the &agr;2C-adrenoreceptor gene, known to regulate blood pressure and learning, the defensin, histocompability complex enhancer-binding protein, carboxypeptidase M, and the Fc fragment of IgE known to be involved in cellular and humoral immune responses. Others, like human cell death protein, TRAIL, and galectin-4 participate in the regulation of apoptosis. Real-time quantitative reverse transcription-polymerase chain reaction analysis was performed in order to confirm the expression changes in AD lymphocytes, and it could detect down-regulation of defensin and &agr;2c-adrenoceptor genes, while other genes seemed unaltered in their expression, including heat-shock protein (hsp90), cholesteryl ester transfer protein, and apolipoprotein B100 (apoB). The altered expression profile of these genes might be connected with the previously reported AD-specific lymphocyte abnormalities. It remains to be elucidated, however, how these genes are related to the pathomechanism of dementia and whether the gene expression differences of AD lymphocytes reflect disease traits or stage processes.

Reversible and irreversible acetylcholinesterase inhibitors cause changes in neuronal amyloid precursor protein processing and protein kinase C level in vitro.

The alternative routes of cleavage of the amyloid precursor protein (APP) result in the generation and secretion of both soluble APP and beta-amyloid, the latter being the main component of the amyloid deposits in the brains of individuals with Alzheimers disease (AD). This study examined the question of whether acetylcholinesterase (AChE) inhibitors can alter the processing of APP and the level of protein kinase C (PKC) in primary rat basal forebrain cultures. Western blotting was used to test two AChE inhibitors (reversible and irreversible) for their ability to enhance the release of APP and PKC content. These inhibitors were ambenonium (AMB) and metrifonate (MTF), at different concentrations. A significant increase was found in the cell-associated APP level in a basal forebrain neuronal culture, and there was an elevation of the APP release into the medium. Increases were similarly observed in the PKC levels after AMB or MTF treatment. The results suggest that these AChE inhibitors promote the non-amyloidogenic route of APP processing, which may be due to their stimulatory effects on PKC. The PKC activation may enhance the alpha-secretase activity and consequently the production of the N-terminal APP. Since both a decreased level of APP secretion and a low activity and level of PKC may be involved in the pathogenesis of AD, it is concluded that the administration of AChE inhibitors to AD patients may facilitate the memory processes and exert a neuroprotective effect.

Temporal parameters of spontaneous speech in Alzheimer's disease

This paper reports on four temporal parameters of spontaneous speech in three stages of Alzheimers disease (mild, moderate, and severe) compared to age-matched normal controls. The analysis of the time course of speech has been shown to be a particularly sensitive neuropsychological method to investigate cognitive processes such as speech planning and production. The following parameters of speech were measured in Hungarian native-speakers with Alzheimers disease and normal controls: articulation rate, speech tempo, hesitation ratio, and rate of grammatical errors. Results revealed significant differences in most of these speech parameters among the three Alzheimers disease groups. Additionally, the clearest difference between the normal control group and the mild Alzheimers disease group involved the hesitation ratio, which was significantly higher in the latter group. This parameter of speech may have diagnostic value for mild-stage Alzheimers disease and therefore could be a useful aid in medical practice.

Imipramine and citalopram facilitate amyloid precursor protein secretion in vitro

Comorbid depression of Alzheimers disease (AD) is a common mood disorder in the elderly and a broad spectrum of antidepressants have been used for its treatment. Abeta peptides and other derivatives of the amyloid precursor protein (APP) have been implicated as central to the pathogenesis of AD. However, the functional relationship of APP and its proteolytic derivatives to antidepressant therapy is not known. In this study, Western blotting was used to test the ability of the tricyclic antidepressant (TCA) imipramine or the selective serotonin reuptake inhibitor (SSRI) citalopram to change the release of APP and the protein kinase C (PKC) content. Both antidepressants increased APP secretion in primary rat neuronal cultures. Imipramine or citalopram enhanced the level of secreted APP by 3.2- or 3.4-fold, respectively. Increases in PKC level were observed only after imipramine treatment. These in vitro data suggest that both TCA and SSRI are able to interfere with the APP metabolism. Imipramine promotes the non-amyloidogenic route of APP processing via stimulatory effects on PKC. We propose that PKC is not involved in the mechanism underlying the effects of citalopram on the APP metabolism. Since the secreted APP is not further available for the pathological cleavage of beta- and gamma-secretases, antidepressant medication might be beneficial in AD therapy.

Impact of Venlafaxine on Gene Expression Profile in Lymphocytes of the Elderly with Major Depression – Evolution of Antidepressants and the Role of the “Neuro-Immune” System

Antidepressive drugs offer considerable symptomatic relief in mood disorders and, although commonly discovered by screening with single biological targets, most interact with multiple receptors and signaling pathways. Antidepressants require a treatment regimen of several weeks before clinical efficacy is achieved in patient populations. While the biochemical mechanisms underlying the delayed temporal profile remain unclear, molecular adaptations over time are likely involved. The selective serotonin and noradrenaline reuptake inhibitor, venlafaxine, offers a dual antidepressive action. Its pharmacological behavior, however, is unknown at the genetic level, and it is difficult to monitor in human brain samples. Because the hypothalamic-pituitary-adrenal axis is often severely disrupted in mood disorders, lymphocytes may serve as models of neuropsychiatric conditions. As such, we examined the role of venlafaxine on the gene expression profile of human lymphocytes. DNA microarray was used to measure the expression patterns of multiple genes in human lymphocytes from depressed patients treated with this mood stabilizer. In this self-controlled study, RNAs of control and treated samples were purified, converted into cDNA and labeled with either Cy3 or Cy5, mixed and hybridized to DNA microarrays containing human oligonucleotides corresponding to more than 8,000 genes. Genes that were differentially regulated in response to treatment were selected for follow up on the basis on novelty, gene identity, and level of over-expression/repression, and selected transcripts were profiled by real-time PCR (data have been normalized to β-actin). Using software analysis of the microarray data, a number of transcripts were differentially expressed between control and treated samples, of which only 57 were found to significantly vary with the “P” value of 0.05 or lower as a result of exposure to venlafaxine. Of these, 31 genes were more highly expressed and 26 transcripts were found to be significantly less abundant. Most selected genes were verified with QRT-PCR to alter. As such, independent verification using QRT-PCR demonstrated the reliability of the method. Genes implicated in ionic homeostasis were differentially expressed, as were genes associated with cell survival, neural plasticity, signal transduction, and metabolism. Understanding how gene expression is altered over a clinically relevant time course of administration of venlafaxine may provide insight into the development of antidepressant efficacy as well as the underlying pathology of mood disorders. These changes in lymphocytes are thought to occur in the brain, and a “neuro-immune system” is proposed by this study.

Gene expression profile analysis of the rat cortex following treatment with imipramine and citalopram

The effect of antidepressants is the culmination of a series of molecular actions occurring in the brain. These events are thought to lead to changes in the expression level of numerous, but as yet unknown genes that result in different cellular functions. In our present study we addressed this issue by establishing gene expression profiles of the rat brain after treatment with imipramine and citalopram at therapeutic doses. After 96 h and 4 wk, fronto-temporal cortices from controls and each treated strain were prepared and total RNA was isolated, and assessed using a cDNA microarray system containing 3200 clones. The expression of 6 genes was decreased and 8 were over-expressed by imipramine, whereas 27 were repressed and 7 were up-regulated by citalopram. Members of signal transduction (e.g. phosphatidylinositol transfer protein), structural elements (e.g. tubulin, fibronectin), factors related to protein metabolism in general (e.g. proteasomal subunits, ubiquitin-like proteins, polyadenylation sites), components involved in cell survival (e.g. midkine, stress-inducible protein), and determinants of membrane conductance and ion transport (e.g. vacuolar H+-ATPase), and basics of nuclear functions (e.g. translin, basal transcription factor 3), were some of the genes with altered expression. These data demonstrate that antidepressants interfere with the expression of a large array of genes involved in signalling, survival and protein metabolism. Our results demonstrate for the first time that antidepressants specifically regulate neuronal plasticity through induction of a highly specific transcriptional programme in brain cells.

The Effect of Citalopram on Gene Expression Profile of Alzheimer Lymphocytes

Antidepressants are widely used in the treatment of mood disorders associated with dementia, however little information is available on their effect at the molecular level. In certain neurodegenerative disorders, such as in Alzheimers disease, lymphocytes have been used to assess mirror changes that thought to occur in the brain. Gene expression profiles of lymphocytes from Alzheimer patients have been shown to differ from that seen with controls. To address this issue in light of antidepressant treatment, we used lymphocytes derived from Alzheimers disease patients and control individuals to assess the impact of the selective serotonine reuptake inhibitor citalopram on gene expression using a cDNA microarray representing 3200 distinct human genes. Sequences that are differentially regulated after treatment with citalopram were identified and categorized based on similarities in biological functions. This analysis revealed that the overexpression of genes in control and Alzheimer white blood cells by citalopram are implicated in cell survival. Apart from this, citalopram did not markedly alter genes involved in other molecular functions in control cells. In contrast, alteration of genes implicated in ionic currents, cell-adhesion, immune mechanism, and adrenergic functions, were also observed in Alzheimer lymphocytes. The expression of genes of Alzheimer lymphocytes by citalopram is modulated differently which may correlate with the pathology.

Association between a variant of the sigma-1 receptor gene and Alzheimer's disease

Alzheimers disease (AD) is a progressive neurodegenerative disorder with complex etiology and strong genetic predisposition. A number of investigations support the possible involvement of sigma non-opioid intracellular receptor 1 (SIGMAR1) in the pathophysiology of AD. We aimed to investigate the association between SIGMAR1 polymorphisms and late-onset AD, therefore we genotyped rs1799729 (GC-241-240TT) and rs1800866 (Q2P) in 322 Hungarian late-onset AD patients and 250 ethnically matched, elderly control individuals. The investigated polymorphisms were in nearly complete linkage disequilibrium resulting in the GC-Q and TT-P predominant haplotypes that were subjected to the statistical analyses. Our data demonstrates an association between the SIGMAR1 TT-P variant and the risk for developing AD (p=0.019), and a potential modest interaction effect (p=0.058) of the co-presence of the TT-P haplotype with apolipoprotein E4 allele on the risk for AD. Based on this mild significance, we could not fully support the hypothesis that TT-P haplotype in interaction with APOE E4 allele confers risk for developing AD.

Effect of general anesthetics on amyloid precursor protein and mRNA levels in the rat brain

The incidence of Alzheimer’s disease is elevated after exposure to surgical interventions. Since amyloid precursor protein (APP) and its neurotoxic derivatives play key roles in the development of Alzheimer dementia, the role of general anesthesia is controversial in the development of cognitive decline. As such, the effect of anesthetics on APP protein and mRNA levels was assessed utilizing semiquantitative Western-immunoblot and reverse transcription polymerase chain reaction (RT-PCR) in brains of rats following intraperitoneal treatment with propofol and thiopental. The anesthetics did not change cortical APP protein and mRNA concentration considerably. These results indicate that both propofol and thiopental are considered to be relatively safe with respect to APP metabolism.