Carlos Velez-Pardo

Brain imaging and fluid biomarker analysis in young adults at genetic risk for autosomal dominant Alzheimer's disease in the presenilin 1 E280A kindred: a case-control study

BACKGROUND We have previously characterised functional brain abnormalities in young adults at genetic risk for late-onset Alzheimers disease. To gain further knowledge on the preclinical phase of Alzheimers disease, we sought to characterise structural and functional MRI, CSF, and plasma biomarkers in a cohort of young adults carrying a high-penetrance autosomal dominant mutation that causes early-onset Alzheimers disease. METHODS Between January and August, 2010, 18-26-year-old presenilin 1 (PSEN1) E280A mutation carriers and non-carriers from the Colombian Alzheimers Prevention Initiative Registry in Medellín Antioquia, Colombia, had structural MRI, functional MRI during associative memory encoding and novel viewing and control tasks, and cognitive assessments. Consenting participants also had lumbar punctures and venepunctures. Outcome measures were task-dependent hippocampal or parahippocampal activations and precuneus or posterior cingulate deactivations, regional grey matter reductions, CSF Aβ(1-42), total tau and phospho-tau(181) concentrations, and plasma Aβ(1-42) concentrations and Aβ(1-42):Aβ(1-40) ratios. Structural and functional MRI data were compared using automated brain mapping algorithms and search regions related to Alzheimers disease. Cognitive and fluid biomarkers were compared using Mann-Whitney tests. FINDINGS 44 participants were included: 20 PSEN1 E280A mutation carriers and 24 non-carriers. The carrier and non-carrier groups did not differ significantly in their dementia ratings, neuropsychological test scores, or proportion of apolipoprotein E (APOE) ɛ4 carriers. Compared with non-carriers, carriers had greater right hippocampal and parahippocampal activation (p=0·001 and p<0·014, respectively, after correction for multiple comparisons), less precuneus and posterior cingulate deactivation (all p<0·010 after correction), and less grey matter in several parietal regions (all p<0·002 uncorrected and corrected p=0·009 in the right parietal search region). In the 20 participants (ten PSEN1 E280A mutation carriers and ten non-carriers) who had lumbar punctures and venepunctures, mutation carriers had higher CSF Aβ(1-42) concentrations (p=0·008) and plasma Aβ(1-42) concentrations (p=0·01) than non-carriers. INTERPRETATION Young adults at genetic risk for autosomal dominant Alzheimers disease have functional and structural MRI findings and CSF and plasma biomarker findings consistent with Aβ(1-42) overproduction. Although the extent to which the underlying brain changes are either neurodegenerative or developmental remain to be determined, this study shows the earliest known biomarker changes in cognitively normal people at genetic risk for autosomal dominant Alzheimers disease. FUNDING Banner Alzheimers Foundation, Nomis Foundation, Anonymous Foundation, Forget Me Not Initiative, Boston University Department of Psychology, Colciencias, National Institute on Aging, National Institute of Neurological Disorders and Stroke, and the State of Arizona.

Aβ[25–35] Peptide and Iron Promote Apoptosis in Lymphocytes by an Oxidative Stress Mechanism: Involvement of H2O2, Caspase-3, NF-κB, p53 and c-Jun

Abstract The Aβ deposition in the neuritic plaques is one of the major neuropathological hallmarks of the Alzheimer disease (AD). Studies in vitro have demonstrated that the Aβ 25–35 fragment, which contains the cytotoxic functional sequence of the amyloid peptide, induces neurotoxicity and cell death by apoptosis. Despite intense investigations, a complete picture of the precise molecular cascade leading to cell death in a single cellular model is still lacking. In this study, we provide evidence that Aβ 25–35 induce apoptosis either alone or in presence of iron in peripheral blood lymphocytes cells (PBL) in a concentration-dependent fashion by an oxidative stress mechanism involving: (1) the production of hydrogen peroxide (H 2 O 2 ), reflected by rhodamine-positive fluorescent cells, (2) activation and/or translocation of NF-κB, p53 and c-Jun transcription factors showed by immunocytochemical diaminobenzidine positive nuclei, (3) activation of NF-κB complex by electrophoretic mobility shift assay/immuno-blotting/and ammonium pyrrolidinedithiocarbamate (PDTC) inhibition, (4) caspase-3 activation, reflected by caspase Ac-DEVD-cho inhibition, (5) mRNA synthesis de novo according to actinomycin D cell death inhibition. These results are consistent with the notion that the Aβ 25–35 /H 2 O 2 generation precede the apoptotic process and that once H 2 O 2 is generated, it is able to trigger a specific cell death signalisation. Thus, taken together these results, we present a well-ordered cascade of the major molecular events leading PBL to apoptosis. These results may contribute to explain the importance of Aβ alone or in the presence of redox-available iron in association with Aβ plaques (and neurofibrillary tangles) in AD brains and the significant role played by H 2 O 2 as a second messenger of death signal in some degenerative diseases linked to oxidative stress stimuli.

Monoamine neurotoxins-induced apoptosis in lymphocytes by a common oxidative stress mechanism: involvement of hydrogen peroxide (H2O2), caspase-3, and nuclear factor kappa-B (NF-κB), p53, c-Jun transcription factors

The destruction of dopaminergic and serotonergic nerve cells by selective 6-hydroxydopamine (6-OHDA), 5,6-dihydroxytryptamine (5,6-DHT) and 5,7-dihydroxytryptamine (5,7-DHT), respectively, is a commonly used tool to investigate the mapping of neuronal pathways, elucidation of function and to mimic human neurodegenerative disease such as Parkinsons and Alzheimers diseases. Despite intense investigations, a complete picture of the precise molecular cascade leading to cell death in a single cellular model is still lacking. In this study, we provide evidence that 6-OHDA, 5,6- and 5,7-DHT toxins-induced apoptosis in peripheral blood lymphocytes cells in a concentration-dependent fashion by a common oxidative mechanism involving: (1) the oxidation of toxins into quinones and production of the by-product hydrogen peroxide, reflected by desipramine-a monoamine uptake blocker-and antioxidants inhibition, (2) activation and/or translocation of nuclear factor-kappaB, p53 and c-Jun transcription factors, showed by immunocytochemical diaminobenzidine-positive stained nuclei, (3) caspase-3 activation, reflected by caspase Ac-DEVD-CHO inhibition, (4) mRNA and protein synthesis de novo according to cycloheximide and actinomycin D cell death inhibition. These results are consistent with the notion that uptake and intracellular autoxidation of those toxins precede the apoptotic process and that once H(2)O(2) is generated, it is able to trigger a specific cell death signalisation. Thus, taken together these results, we present an ordered cascade of the major molecular events leading peripheral blood lymphocytes to apoptosis. These results may contribute to explain the importance of H(2)O(2) as a second messenger of death signal in some degenerative diseases linked to oxidative stress stimuli.

CA1 Hippocampal Neuronal Loss in Familial Alzheimer's Disease Presenilin-1 E280A Mutation Is Related to Epilepsy

Summary:  Purpose: Alzheimer disease (AD) and epilepsy are brain disorders frequently associated with neuronal cell loss in mesial temporal lobe structures, but presenting different patterns of damage. Recently it was proposed that a causal relation may exist between AD pathology and the appearance of epilepsy in some cases with AD. This study aimed to determine the neuronal loss in CA1 hippocampal region from patients bearing the presenilin‐1 [E280A] mutation (PS1[E280A]) associated with seizures.

The Bad, the Good, and the Ugly about Oxidative Stress

Alzheimers disease (AD), Parkinsons disease (PD), and cancer (e.g., leukemia) are the most devastating disorders affecting millions of people worldwide. Except for some kind of cancers, no effective and/or definitive therapeutic treatment aimed to reduce or to retard the clinic and pathologic symptoms induced by AD and PD is presently available. Therefore, it is urgently needed to understand the molecular basis of these disorders. Since oxidative stress (OS) is an important etiologic factor of the pathologic process of AD, PD, and cancer, understanding how intracellular signaling pathways respond to OS will have a significant implication in the therapy of these diseases. Here, we propose a model of minimal completeness of cell death signaling induced by OS as a mechanistic explanation of neuronal and cancer cell demise. This mechanism might provide the basis for therapeutic design strategies. Finally, we will attempt to associate PD, cancer, and OS. This paper critically analyzes the evidence that support the “oxidative stress model” in neurodegeneration and cancer.

Paraquat induces apoptosis in human lymphocytes: Protective and rescue effects of glucose, cannabinoids and insulin-like growth factor-1

In order to establish causal or protective treatments for Parkinsons disease (PD), it is necessary to identify the cascade of deleterious events that lead to the dysfunction and death of dopaminergic neurons. Paraquat (PQ) is a pesticide used as xenobiotic compound to model PD. However, the mechanism(s) of PQ-induced cell death and the mechanism(s) of cytoprotection in a single cell model are still unknown. In this study, lymphocytes were treated with (0.1–1 mM) PQ. Apoptotic morphology was assessed with acridine orange/ethidium bromide staining. Further evaluation included (i) superoxide radicals, reflected by nitroblue tetrazolium reduction to formazan, (ii) the production of hydrogen peroxide, reflected by rhodamine-positive fluorescent cells, (iii) the generation of hydroxyl radicals, reflected by dimethylsulfoxide and melatonin √OH scavengers, (iv) activation and/or translocation of NF-κB, p53 and c-Jun transcription factors showed by immunocytochemical staining, and by ammonium pyrrolidinedithiocarbamate, pifithrin-α and SP600125 inhibition and (V) caspase-3 activation, reflected by caspase Ac-DEVD-cho inhibition. To elucidate the mechanism of cytoprotection, lymphocytes were treated with PQ in the presence of cannabinoids, insulin-like growth factor-1 and glucose. We provide evidence that PQ induces apoptosis in lymphocytes in a concentration- and time-dependent fashion by an oxidative stress mechanism involving O2√ − , H2O2/(√OH) generation, simultaneous activation of NF-κB/p53/c-Jun transcription factors, mitochondrial depolarization and caspase-3 activation leading to morphological apoptosis. Moreover, dying lymphocytes are protected and rescued from PQ noxious stimuli by direct antioxidant effect by cannabinoids, receptor mediated signaling by IGF-1, and/or energetic protection by glucose. It is concluded that PQ-induced apoptosis in lymphocytes by a mechanism involving reactive oxygen species generation, mitochondrial dysfunction, transcriptional factors and caspase-3 activation. However, this cell death routine can be reversed by the action of cannabinoids, IGF-1 and glucose. These data may provide innovating therapeutic strategies to intervene environmentally or genetically susceptible PD population to oxidative stress.

Ultrastructure evidence of necrotic neural cell death in familial Alzheimer's disease brains bearing presenilin-1 E280A mutation1

Recently, it has been demonstrated that there is no obvious correlation between DNA fragmentation (according to Terminal dUTP Nick-End Labeling technique) and the severity of amyloid-beta (Abeta) deposition and neurofibrillary tangle (NFT) formation in patients bearing mutations in presenilin 1[E280A]. Indeed, it was observed in 10 out of 48 brain sections TUNEL-positive labeling, while none showed classical apoptotic morphology. Based on these findings, we were interested to determine whether cortical cells from temporal and hippocampus post mortem brain sections die either by an apoptotic or necrotic process in FAD-brain sections labeled TUNEL positive compared with normal brain subjects labeled TUNEL-negative using electron microscopy (EM). We found that FAD-brain sections labeled TUNEL positive display the typical morphological characteristics of cell death by necrosis i.e. the nuclear chromatin form flocculent aggregates with poorly defined edges and electron lucent (it does not appears black on EM); the chromatin aggregates are irregularly scattered through the nucleus; mitochondria are swoolen with flocculent matrix densities. No apoptotic bodies were observed in any of the brain areas studied. These results may indicate that necrosis is the most generalized cell death process occurring in terminal PS1E280A brains and the DNA fragmentation of nuclei labeled by TUNEL technique may reflect DNA vulnerability. Thus, cell death by necrosis and the accompanying histopathological observations such as severe deposition of amyloid plaques and NFTs, severe gliosis, cortical depopulation, influx of lymphocytes indicative of a chronic inflammation may have an important impact on future therapeutic strategies in the treatment of PS1E280A patients.

17β-Estradiol protects lymphocytes against dopamine and iron-induced apoptosis by a genomic-independent mechanism: Implication in Parkinson's disease

Dopamine (DA) in combination with iron (Fe(2+)) has been demonstrated to induce apoptosis in neuronal-like PC12 cells by an oxidative stress mechanism. To get a better insight of cell death and protective mechanisms in DA/Fe(2+)-induced toxicity, we investigated the effects of DA/Fe(2+) and the antioxidant action of 17 beta-estradiol (E2) in peripheral blood lymphocytes (PBL). We found that DA/Fe(2+)-induces apoptosis in PBL via a hydrogen peroxide (H(2)O(2))-mediated oxidative mechanism, which in turn triggers a cascade of molecular events requiring RNA and de novo protein synthesis. We have also demonstrated that E2 prevents significantly DA/Fe(2+)-induced apoptosis in PBL by directly inhibiting the intracellular accumulation of peroxides generated by DA/Fe(2+)-reaction. This protective activity is independent of the presence or activation of the estrogen receptors (ERs). These data further support and validate our previous hypothesis that DA/Fe(2+)/H(2)O(2) could be a general mediator of oxidative stress through a common cell death mechanism in both neuronal and nonneuronal cells. These findings may be particularly relevant to the potential approaches to rescue and prolong the survival of neurons by estrogens in patients with Parkinsons disease (PD).

TPEN induces apoptosis independently of zinc chelator activity in a model of acute lymphoblastic leukemia and ex vivo acute leukemia cells through oxidative stress and mitochondria caspase-3- and AIF-dependent pathways.

Acute lymphoblastic leukemia is still an incurable disease with resistance to therapy developing in the majority of patients. We investigated the effect of TPEN, an intracellular zinc chelator, in Jurkat and in ex vivo acute lymphoblastic leukemia (ALL) cells resistant to chemotherapy. Changes of nuclei morphology, reactive oxygen species generation, presence of hypodiploid cells, phosphatidylserine translocation, mitochondrial membrane depolarization, immunohistochemical identification of cell death signalling molecules, and pharmacological inhibition were assayed to detect the apoptotic cell death pathways. We found that TPEN induces apoptosis in both types of cells by a molecular oxidative stress pathway involving O2•− > H2O2 ≫ NF-κB (JNK/c-Jun) >p53> loss ΔΨm> caspase-3, AIF > chromatin condensation/DNA fragmentation. Interestingly, TPEN induced apoptosis independently of glucose; leukemic cells are therefore devoid of survival capacity by metabolic resistance to treatment. Most importantly, TPEN cytotoxic effect can eventually be regulated by the antioxidant N-acetyl-cysteine and zinc ions. Our data suggest that TPEN can be used as a potential therapeutic prooxidant agent against refractory leukemia. These data contribute to understanding the importance of oxidative stress in the treatment of ALL.

Vitamin K3 and vitamin C alone or in combination induced apoptosis in leukemia cells by a similar oxidative stress signalling mechanism

BackgroundSecondary therapy-related acute lymphoblastic leukemia might emerge following chemotherapy and/or radiotherapy for primary malignancies. Therefore, other alternatives should be pursued to treat leukemia.ResultsIt is shown that vitamin K3- or vitamin C- induced apoptosis in leukemia cells by oxidative stress mechanism involving superoxide anion radical and hydrogen peroxide generation, activation of NF-κB, p53, c-Jun, protease caspase-3 activation and mitochondria depolarization leading to nuclei fragmentation. Cell death was more prominent when Jurkat and K562 cells are exposed to VC and VK3 in a ratio 1000:1 (10 mM: 10 μM) or 100:1 (300 μM: 3 μM), respectively.ConclusionWe provide for the first time in vitro evidence supporting a causative role for oxidative stress in VK3- and VC-induced apoptosis in Jurkat and K562 cells in a domino-like mechanism. Altogether these data suggest that VK3 and VC should be useful in the treatment of leukemia.