Diego Mastroeni

GAB2 Alleles Modify Alzheimer's Risk in APOE ε4 Carriers

The apolipoprotein E (APOE) epsilon4 allele is the best established genetic risk factor for late-onset Alzheimers disease (LOAD). We conducted genome-wide surveys of 502,627 single-nucleotide polymorphisms (SNPs) to characterize and confirm other LOAD susceptibility genes. In epsilon4 carriers from neuropathologically verified discovery, neuropathologically verified replication, and clinically characterized replication cohorts of 1411 cases and controls, LOAD was associated with six SNPs from the GRB-associated binding protein 2 (GAB2) gene and a common haplotype encompassing the entire GAB2 gene. SNP rs2373115 (p = 9 x 10(-11)) was associated with an odds ratio of 4.06 (confidence interval 2.81-14.69), which interacts with APOE epsilon4 to further modify risk. GAB2 was overexpressed in pathologically vulnerable neurons; the Gab2 protein was detected in neurons, tangle-bearing neurons, and dystrophic neuritis; and interference with GAB2 gene expression increased tau phosphorylation. Our findings suggest that GAB2 modifies LOAD risk in APOE epsilon4 carriers and influences Alzheimers neuropathology.

Alzheimer's disease is associated with reduced expression of energy metabolism genes in posterior cingulate neurons

Alzheimers disease (AD) is associated with regional reductions in fluorodeoxyglucose positron emission tomography (FDG PET) measurements of the cerebral metabolic rate for glucose, which may begin long before the onset of histopathological or clinical features, especially in carriers of a common AD susceptibility gene. Molecular evaluation of cells from metabolically affected brain regions could provide new information about the pathogenesis of AD and new targets at which to aim disease-slowing and prevention therapies. Data from a genome-wide transcriptomic study were used to compare the expression of 80 metabolically relevant nuclear genes from laser-capture microdissected non-tangle-bearing neurons from autopsy brains of AD cases and normal controls in posterior cingulate cortex, which is metabolically affected in the earliest stages; other brain regions metabolically affected in PET studies of AD or normal aging; and visual cortex, which is relatively spared. Compared with controls, AD cases had significantly lower expression of 70% of the nuclear genes encoding subunits of the mitochondrial electron transport chain in posterior cingulate cortex, 65% of those in the middle temporal gyrus, 61% of those in hippocampal CA1, 23% of those in entorhinal cortex, 16% of those in visual cortex, and 5% of those in the superior frontal gyrus. Western blots confirmed underexpression of those complex I–V subunits assessed at the protein level. Cerebral metabolic rate for glucose abnormalities in FDG PET studies of AD may be associated with reduced neuronal expression of nuclear genes encoding subunits of the mitochondrial electron transport chain.

Epigenetic changes in Alzheimer's disease: Decrements in DNA methylation

DNA methylation is a vital component of the epigenetic machinery that orchestrates changes in multiple genes and helps regulate gene expression in all known vertebrates. We evaluated immunoreactivity for two markers of DNA methylation and eight methylation maintenance factors in entorhinal cortex layer II, a region exhibiting substantial Alzheimers disease (AD) pathology in which expression changes have been reported for a wide variety of genes. We show, for the first time, neuronal immunoreactivity for all 10 of the epigenetic markers and factors, with highly significant decrements in AD cases. These decrements were particularly marked in PHF1/PS396 immunoreactive, neurofibrillary tangle-bearing neurons. In addition, two of the DNA methylation maintenance factors, DNMT1 and MBD2, have been reported also to interact with ribosomal RNAs and ribosome synthesis. Consistent with these findings, DNMT1 and MBD2, as well as p66α, exhibited punctate cytoplasmic immunoreactivity that co-localized with the ribosome markers RPL26 and 5.8s rRNA in ND neurons. By contrast, AD neurons generally lacked such staining, and there was a qualitative decrease in RPL26 and 5.8s rRNA immunoreactivity. Collectively, these findings suggest epigenetic dysfunction in AD-vulnerable neurons.

Epigenetic Differences in Cortical Neurons from a Pair of Monozygotic Twins Discordant for Alzheimer's Disease

DNA methylation [1], [2] is capable of modulating coordinate expression of large numbers of genes across many different pathways, and may therefore warrant investigation for their potential role between genes and disease phenotype. In a rare set of monozygotic twins discordant for Alzheimers disease (AD), significantly reduced levels of DNA methylation were observed in temporal neocortex neuronal nuclei of the AD twin. These findings are consistent with the hypothesis that epigenetic mechanisms may mediate at the molecular level the effects of life events on AD risk, and provide, for the first time, a potential explanation for AD discordance despite genetic similarities.

Consistent decrease in global DNA methylation and hydroxymethylation in the hippocampus of Alzheimer's disease patients

Epigenetic dysregulation of gene expression is thought to be critically involved in the pathophysiology of Alzheimers disease (AD). Recent studies indicate that DNA methylation and DNA hydroxymethylation are 2 important epigenetic mechanisms that regulate gene expression in the aging brain. However, very little is known about the levels of markers of DNA methylation and hydroxymethylation in the brains of patients with AD, the cell-type specificity of putative AD-related alterations in these markers, as well as the link between epigenetic alterations and the gross pathology of AD. The present quantitative immunohistochemical study investigated the levels of the 2 most important markers of DNA methylation and hydroxymethylation, that is, 5-methylcytidine (5-mC) and 5-hydroxymethylcytidine (5-hmC), in the hippocampus of AD patients (n = 10) and compared these to non-demented, age-matched controls (n = 10). In addition, the levels of 5-hmC in the hippocampus of a pair of monozygotic twins discordant for AD were assessed. The levels of 5-mC and 5-hmC were furthermore analyzed in a cell-type and hippocampal subregion-specific manner, and were correlated with amyloid plaque load and neurofibrillary tangle load. The results showed robust decreases in the hippocampal levels of 5-mC and 5-hmC in AD patients (19.6% and 20.2%, respectively). Similar results were obtained for the twin with AD when compared to the non-demented co-twin. Moreover, levels of 5-mC as well as the levels of 5-hmC showed a significant negative correlation with amyloid plaque load in the hippocampus (r(p) = -0.539, p = 0.021 for 5-mC and r(p) = -0.558, p = 0.016 for 5-hmC). These human postmortem results thus strengthen the notion that AD is associated with alterations in DNA methylation and hydroxymethylation, and provide a basis for further epigenetic studies identifying the exact genetic loci with aberrant epigenetic signatures.

Neuroinflammation in Alzheimer's disease and Parkinson's disease: are microglia pathogenic in either disorder?

Microglial activation similar to that which occurs in peripheral macrophages during inflammatory attack was first demonstrated in the Alzheimers disease (AD) brain two decades ago. Localization to pathologically vulnerable regions of AD cortex, localization to sites of specific AD pathology such as amyloid-beta peptide (Abeta) deposits, and the ability of activated microglia to release toxic inflammatory factors suggested that the activation of microglia in AD might play a pathogenic role. However, proving this hypothesis in a disease in which so many profound pathologies occur (e.g., Abeta deposition, neurofibrillary tangle formation, inflammation, neuronal loss, neuritic loss, synaptic loss, neuronal dysfunction, vascular alterations) has proven difficult. Although investigations of microglia in Parkinsons disease (PD) are more recent and therefore less extensive, demonstration of a pathogenic role for microglial activation may actually be much simpler in PD than AD because the root pathological event in PD, loss of dopamine (DA)-secreting substantia nigra neurons, is already well established. Indeed, indirect but converging evidence of a pathogenic role for activated microglia in PD has already begun to emerge. The nigra reportedly has the highest density of microglia in brain, and, in PD, nigral microglia are not only highly activated but also highly clustered around dystrophic DA neurons. 6-OHDA and MPTP models of PD in rodents induce substantia nigra microglial activation. More cogent, injections of the classic microglial/macrophage activator lipopolysaccharide into or near the rodent nigra cause a specific loss of DA neurons there. Culture models with human microglia and human cellular targets replicate this phenomenon. Notably, nearly all the proposed etiologies of PD, including brain bacterial and viral exposure, pesticides, drug contaminants, and repeated head trauma, are known to cause brain inflammation. A mechanism by which activated microglia might specifically target DA neurons remains a critical missing link in the proof of a pathogenic role for activated microglia in PD. If such a link could be established, however, clinical intervention trials with agents that dampen microglial activation might be warranted in PD.

Epigenetic mechanisms in Alzheimer's disease

Epigenetic modifications help orchestrate sweeping developmental, aging, and disease-causing changes in phenotype by altering transcriptional activity in multiple genes spanning multiple biologic pathways. Although previous epigenetic research has focused primarily on dividing cells, particularly in cancer, recent studies have shown rapid, dynamic, and persistent epigenetic modifications in neurons that have significant neuroendocrine, neurophysiologic, and neurodegenerative consequences. Here, we provide a review of the major mechanisms for epigenetic modification and how they are reportedly altered in aging and Alzheimers disease (AD). Because of their reach across the genome, epigenetic mechanisms may provide a unique integrative framework for the pathologic diversity and complexity of AD.

Altered neuronal gene expression in brain regions differentially affected by Alzheimer's disease: a reference data set

Alzheimers Disease (AD) is the most widespread form of dementia during the later stages of life. If improved therapeutics are not developed, the prevalence of AD will drastically increase in the coming years as the worlds population ages. By identifying differences in neuronal gene expression profiles between healthy elderly persons and individuals diagnosed with AD, we may be able to better understand the molecular mechanisms that drive AD pathogenesis, including the formation of amyloid plaques and neurofibrillary tangles. In this study, we expression profiled histopathologically normal cortical neurons collected with laser capture microdissection (LCM) from six anatomically and functionally discrete postmortem brain regions in 34 AD-afflicted individuals, using Affymetrix Human Genome U133 Plus 2.0 microarrays. These regions include the entorhinal cortex, hippocampus, middle temporal gyrus, posterior cingulate cortex, superior frontal gyrus, and primary visual cortex. This study is predicated on previous parallel research on the postmortem brains of the same six regions in 14 healthy elderly individuals, for which LCM neurons were similarly processed for expression analysis. We identified significant regional differential expression in AD brains compared with control brains including expression changes of genes previously implicated in AD pathogenesis, particularly with regard to tangle and plaque formation. Pinpointing the expression of factors that may play a role in AD pathogenesis provides a foundation for future identification of new targets for improved AD therapeutics. We provide this carefully phenotyped, laser capture microdissected intraindividual brain region expression data set to the community as a public resource.

Peripheral clearance of amyloid β peptide by complement C3-dependent adherence to erythrocytes

Brain deposits of amyloid beta peptide (Abeta) have been a diagnostic hallmark of Alzheimers disease (AD) for nearly a century. Recent studies have demonstrated that Abeta is also present in peripheral blood. Here, we present evidence that circulating Abeta42 is subject to complement C3b-dependent adherence to complement receptor 1 (CR1) on erythrocytes, a classical set of mechanisms by which pathogens and proteins recognized as foreign are cleared from the bloodstream. Levels of Abeta42 targeted by this pathway differ significantly in AD compared to mild cognitive impairment and nondemented elderly controls.

The epigenetics of aging and neurodegeneration

Epigenetics is a quickly growing field encompassing mechanisms regulating gene expression that do not involve changes in the genotype. Epigenetics is of increasing relevance to neuroscience, with epigenetic mechanisms being implicated in brain development and neuronal differentiation, as well as in more dynamic processes related to cognition. Epigenetic regulation covers multiple levels of gene expression; from direct modifications of the DNA and histone tails, regulating the level of transcription, to interactions with messenger RNAs, regulating the level of translation. Importantly, epigenetic dysregulation currently garners much attention as a pivotal player in aging and age-related neurodegenerative disorders, such as Alzheimers disease, Parkinsons disease, and Huntingtons disease, where it may mediate interactions between genetic and environmental risk factors, or directly interact with disease-specific pathological factors. We review current knowledge about the major epigenetic mechanisms, including DNA methylation and DNA demethylation, chromatin remodeling and non-coding RNAs, as well as the involvement of these mechanisms in normal aging and in the pathophysiology of the most common neurodegenerative diseases. Additionally, we examine the current state of epigenetics-based therapeutic strategies for these diseases, which either aim to restore the epigenetic homeostasis or skew it to a favorable direction to counter disease pathology. Finally, methodological challenges of epigenetic investigations and future perspectives are discussed.