Alessio Crestini

A novel PSEN2 mutation associated with a peculiar phenotype.

Background: Mutations of presenilin 2 gene are a rare cause of familial Alzheimer disease (AD). We describe an Italian family with hereditary dementia associated with a novel mutation in the presenilin 2 gene. Methods: Clinical investigations of the diseased subjects; interviews with relatives; studies of medical records; pedigree analysis; and neuroradiologic, neuropathologic, and molecular genetic studies were carried out in the pedigree. Results: Genetic analysis showed a novel PSEN2 A85V mutation present in the proband and in all analyzed affected members, in a subject presenting with an amnesic mild cognitive impairment, and in a young, still asymptomatic subject. The proband showed a clinical phenotype indicative of Lewy body dementia and the neuropathologic examination demonstrated the presence of unusually abundant and widespread cortical Lewy bodies in addition to the hallmark lesions of AD. Other affected members exhibited a clinical phenotype typical of AD. Conclusions: Our findings add complexity to the spectrum of atypical phenotypes associated with presenilin mutations and should then be taken into account when considering the nosography of neurodegenerative diseases. They also support previous data that specific mutations of genes associated with familial Alzheimer disease may influence the presence and extent of Lewy bodies.

Genetic study of Sardinian patients with Alzheimer's disease

We describe the genetic analysis of an Alzheimers disease (AD) sample derived from a genetically isolated population. Genetic assessment included the analysis of genes involved in AD, such as the genes for amyloid precursor protein (APP), presenilin 1 (PSEN1) and presenilin 2 (PSEN2). We also assessed genes for some proteins that constitute the gamma-secretase complex: nicastrin (NCSTN), presenilin enhancer-2 (PEN2), in addition to the AD risk factor apolipoprotein E (APOE). Using polymerase chain reaction and single strand conformational polymorphism method, screens for APP, PSEN1 and PSEN2 genes revealed one mutation in PSEN1. Furthermore, we found an intronic +17G>C polymorphism in PEN2 which, in homozygous form, was greater in early onset Alzheimers disease (EOAD) compared to controls, and one haplotype in the NCSTN gene which was linked to EOAD and familial AD (FAD). Finally, the genotyping of APOE confirmed that the varepsilon4 allele could be a risk factor for the onset of AD, in particular for FAD subjects. In conclusion, these results show the existence of Sardinian genetic peculiarities, essential in studies regarding genetically inherited and multifactorial disorders, as AD.

Hypoxia induces up-regulation of progranulin in neuroblastoma cell lines

Progranulin (PGRN) is a widely expressed multifunctional protein, involved in regulation of cell growth and cell cycle progression with a possible involvement in neurodegeneration. We looked for PGRN regulation in three different human neuroblastoma cell lines, following exposure to two different stimuli commonly associated to neurodegeneration: hypoxia and oxidative stress. For gene and protein expression analysis we carried out a quantitative RT-PCR and western blotting analysis. We show that PGRN is strongly up-regulated by hypoxia, through the mitogen-actived protein kinase (MAPK)/extracellular signal-regulated kinase (MEK) signaling cascade. PGRN is not up-regulated by H(2)O(2)-induced oxidative stress. These results suggest that PGRN in the brain could exert a protective role against hypoxic stress, one of principal risk factors involved in frontotemporal dementia pathogenesis.

PEN-2 gene mutation in a familial Alzheimer's disease case

AbstractGenetic evidence indicates a central role of cerebral accumulation of β–amyloid (Aβ) in the pathogenesis of Alzheimer’s disease (AD). Beside presenilin 1 and 2, three other recently discovered proteins (Aph 1, PEN 2 and nicastrin) are associated with γ–secretase activity, the enzymatic complex generating Aβ. Alterations in genes encoding these proteins were candidates for a role in AD. The PEN 2 gene was examined for unknown mutations and polymorphisms in sporadic and familial Alzheimer patients. Samples from age–matched controls (n = 253), sporadic AD (SAD, n = 256) and familial AD (FAD, n = 140) were screened with DHPLC methodology followed by sequencing. Scanning the gene identified for the first time a missense mutation (D90N) in a patient with FAD. Three intronic polymorphisms were also identified, one of which had a higher presence of the mutated allele in AD subjects carrying the allele ε4 of apolipoprotein E than controls. The pathogenic role of the PEN–2 D90N mutation in AD is not clear, but the findings might lead to new studies on its functional and genetic role.

Nicastrin gene in familial and sporadic Alzheimer's disease.

Nicastrin is a protein recently discovered associated to presenilins and involved in the production of amyloid beta peptide that accumulates in Alzheimers disease (AD) brain. In this study the nicastrin gene was examined for unknown mutations and polymorphisms in 104 patients with familial AD (52 early-onset and 52 late-onset), 174 sporadic AD and 191 healthy neurological controls of Italian origin. The scanning of the nicastrin gene identified a missense mutation (N417Y) in two patients with sporadic AD, in an early-onset familial AD and in a young control subject. Furthermore, we found two silent mutations and four intronic polymorphisms, three of them co-segregating in a single haplotype. We found some differences in the distribution of genotype alterations in the AD population compared to the controls. However, our data together with other evidence did not support the pathological role of missense mutation N417Y.

Familial Alzheimer's disease sustained by presenilin 2 mutations: Systematic review of literature and genotype–phenotype correlation

Familial Alzheimers disease (FAD), despite representing a rare condition, is attracting a growing interest in the scientific community. Improved phenotyping of FAD cases may have a relevant impact both in clinical and research contexts. We performed a systematic review of studies describing the phenotypic features of FAD cases sustained by PSEN2 mutations, the less common cause of monogenic AD. Special attention was given to the clinical manifestations as well as to the main findings coming from the most commonly and widely adopted diagnostic procedures. Basing on the collected data, we also attempted to conduct a genotype-phenotype correlation analysis. Overall, the mutations involving the PSEN2 gene represent an extremely rare cause of FAD, having been reported to date in less than 200 cases. They are mainly associated, despite some peculiar and heterogeneous features, to a typical AD phenotype. Nevertheless, the frequent occurrence of psychotic symptoms may represent a potential distinctive element. The scarcity of available phenotypic descriptions strongly limits the implementation of genotype-phenotype correlations.

Circulating microRNAs in Neurodegenerative Diseases

Neurodegenerative diseases, such as Alzheimers disease (AD), Parkinsons disease (PD), and amyotrophic lateral sclerosis (ALS), are caused by a combination of events that impair normal neuronal function. Although they are considered different disorders, there are overlapping features among them from the clinical, pathological, and genetic points of view. Synaptic dysfunction and loss, neurite retraction, and the appearance of other abnormalities such as axonal transport defects normally precede the neuronal loss that is a relatively late event. The diagnosis of many neurodegenerative diseases is mainly based on patients cognitive function analysis, and the development of diagnostic methods is complicated by the brains capacity to compensate for neuronal loss over a long period of time. This results in the late clinical manifestation of symptoms, a time when successful treatment is no longer feasible. Thus, a noninvasive diagnostic method based on early events detection is particularly important. In the last years, some biomarkers expressed in human body fluids have been proposed. microRNAs (miRNAs), with their high stability, tissue- or cell type-specific expression, lower cost, and shorter time in the assay development, could constitute a good tool to obtain an early disease diagnosis for a wide number of human pathologies, including neurodegenerative diseases. The possibilities and challenges of using these small RNA molecules as a signature for neurodegenerative disorders is a highly promising approach for developing minimally invasive screening tests and to identify new therapeutic targets.

Changes in Cholesterol Metabolism are Associated With PS1 and PS2 Gene Regulation in SK-N-BE

Several lines of evidence suggest that the cholesterol content of neuronal membranes influences amyloid precursor protein (APP) processing; however, its role in transcriptional regulation of the cofactor for γ-secretase, the key enzyme for the production of the Aβ peptide, is poorly understood. This study investigates whether the changes in cellular cholesterol metabolism modulate the expression of genes involved in the γ-secretase complex function. The abundance of mRNA transcripts for presenilin 1 and 2 (PS1 and PS2), APP, and nicastrin were evaluated in neuroblastoma cells exposed either to serum-depleted medium or to low-density lipoproteins (LDL). Cholesterol esterification was markedly inhibited by mevinolin and U18666A; but was not significantly affected by any other of the tested treatments, γ-Secretase genes and cofactors were not co-regulated and were not influenced by statin inhibition of cholesterol synthesis. Nicastrin and the APP isoforms showed constitutive expression. In the absence of exogenous lipids, cell PS1 and PS2 expression was induced by LDL and by lysosomal sequestration of cholesterol. However, a different pattern of induction of presenilin gene expression was observed in the latter condition, suggesting that lysosomal cholesterol levels are strong inducers of PS2 transcription. Taken together, these results indicate that lipid metabolism has a complex influence of γ-secretase transcriptional pathways and, in particular, exogenous cholesterol and compartmentalization in neuroblastoma cells play a relevant role in regulating the transcription of presenilins, while modulation of the cholesterol biosynthesis pathway seems to exert a minor influence on the expression of γ-secretase genes and cofactors.

EFFECTS OF SIMULATED MICROGRAVITY ON THE DEVELOPMENT AND MATURATION OF DISSOCIATED CORTICAL NEURONS

SummaryAlthough a wealth of evidence supports the hypothesis that some functions of the nervous system may be altered during exposure to microgravity, the possible changes in basic neuronal physiology are not easy to assess. Indeed, few studies have examined whether microgravity affects the development of neurons in culture. In the present study, a suspension of dissociated cortical cells from rat embryos were exposed to 24 h of simulated microgravity before plating in a normal adherent culture system. Both preexposed and control cells were used after a period of 7–10 d in vitro. The vitality and the level of reactive oxygen species of cultures previously exposed did not differ from those of normal cultures. Cellular characterization by immunostaining with a specific antibody displayed normal neuronal phenotype in control cells, whereas pretreatment in simulated microgravity revealed an increase of glial fibrillary acidic protein fluorescence in the elongated stellate glial cells. Electrophysiological recording indicated that the electrical properties of neurons preexposed were comparable with those of controls. Overall, our results indicate that a short time of simulated microgravity preexposure does not affect dramatically the ability of dissociated neural cells to develop and differentiate in an adherent culture system.

Altered oxidative stress profile in the cortex of mice fed an enriched branched-chain amino acids diet: possible link with amyotrophic lateral sclerosis?

Branched‐chain amino acids (BCAAs), valine, isoleucine, and leucine, are widely used among athletes as dietary integrators. Although the occurrence of untoward effects of BCCA supplementation, with particular regard to neurological disturbances, cannot be excluded, no specific studies have been performed so far. The aim of this work was to evaluate the effects of a diet enriched in BCAAs on the expression of oxidative stress pathway genes in the brain of C57Bl/6J mice. Animals were fed a standard or a BCAA diet for 95 days starting from postnatal day 21 until sacrifice. BCAA treatment, at doses comparable to human usage, significantly down‐regulated the expression of some antioxidant genes, while up‐regulating the expression of some oxygen transporters. In conclusion, it appears that BCAAs administered by diet could alter some specific oxidative stress pathways in the brain. Caution should thus be exercised in the widespread use of BCAAs as dietary integrators in sports practice.