Rotenone Induces a Neuropathological Phenotype in Cholinergic-like Neurons Resembling Parkinson's Disease Dementia (PDD).

Parkinson's disease with dementia (PDD) is a neurological disorder that clinically and neuropathologically overlaps with Parkinson's disease (PD) and Alzheimer's disease (AD). Although it is assumed that alpha-synuclein ( α -Syn), amyloid beta (A ß ), and the protein Tau might synergistically induce cholinergic neuronal degeneration, presently the pathological mechanism of PDD remains unclear. Therefore, it is essential to delve into the cellular and molecular aspects of this neurological entity to identify potential targets for prevention and treatment strategies. Cholinergic-like neurons (ChLNs) were exposed to rotenone (ROT, 10 µ M) for 24 h. ROT provokes loss of Δ Ψ m , generation of reactive oxygen species (ROS), phosphorylation of leucine-rich repeated kinase 2 (LRRK2 at Ser935) concomitantly with phosphorylation of α -synuclein ( α -Syn, Ser129), induces accumulation of intracellular A ß (iA ß ), oxidized DJ-1 (Cys106), as well as phosphorylation of TAU (Ser202/Thr205), increases the phosphorylation of c-JUN (Ser63/Ser73), and increases expression of proapoptotic proteins TP53, PUMA, and cleaved caspase 3 (CC3) in ChLNs. These neuropathological features resemble those reproduced in presenilin 1 (PSEN1) E280A ChLNs. Interestingly, anti-oxidant and anti-amyloid cannabidiol (CBD), JNK inhibitor SP600125 (SP), TP53 inhibitor pifithrin- α (PFT), and LRRK2 kinase inhibitor PF-06447475 (PF475) significantly diminish ROT-induced oxidative stress (OS), proteinaceous, and cell death markers in ChLNs compared to naïve ChLNs. In conclusion, ROT induces p- α -Syn, iA ß , p-Tau, and cell death in ChLNs, recapitulating the neuropathology findings in PDD. Our report provides an excellent in vitro model to test for potential therapeutic strategies against PDD. Our data suggest that ROT induces a neuropathologic phenotype in ChLNs similar to that caused by the mutation PSEN1 E280A.

Single-nucleus RNA sequencing demonstrates an autosomal dominant Alzheimer's disease profile and possible mechanisms of disease protection.

Highly penetrant autosomal dominant Alzheimer's disease (ADAD) comprises a distinct disease entity as compared to the far more prevalent form of AD in which common variants collectively contribute to risk. The downstream pathways that distinguish these AD forms in specific cell types have not been deeply explored. We compared single-nucleus transcriptomes among a set of 27 cases divided among PSEN1-E280A ADAD carriers, sporadic AD, and controls. Autophagy genes and chaperones clearly defined the PSEN1-E280A cases compared to sporadic AD. Spatial transcriptomics validated the activation of chaperone-mediated autophagy genes in PSEN1-E280A. The PSEN1-E280A case in which much of the brain was spared neurofibrillary pathology and harbored a homozygous APOE3-Christchurch variant revealed possible explanations for protection from AD pathology including overexpression of LRP1 in astrocytes, increased expression of FKBP1B, and decreased PSEN1 expression in neurons. The unique cellular responses in ADAD and sporadic AD require consideration when designing clinical trials.

PSEN1 E280A Cholinergic-like Neurons and Cerebral Spheroids Derived from Mesenchymal Stromal Cells and from Induced Pluripotent Stem Cells Are Neuropathologically Equivalent.

Alzheimer's disease (AD) is a chronic neurological condition characterized by the severe loss of cholinergic neurons. Currently, the incomplete understanding of the loss of neurons has prevented curative treatments for familial AD (FAD). Therefore, modeling FAD in vitro is essential for studying cholinergic vulnerability. Moreover, to expedite the discovery of disease-modifying therapies that delay the onset and slow the progression of AD, we depend on trustworthy disease models. Although highly informative, induced pluripotent stem cell (iPSCs)-derived cholinergic neurons (ChNs) are time-consuming, not cost-effective, and labor-intensive. Other sources for AD modeling are urgently needed. Wild-type and presenilin (PSEN)1 p.E280A fibroblast-derived iPSCs, menstrual blood-derived menstrual stromal cells (MenSCs), and umbilical cord-derived Wharton Jelly's mesenchymal stromal cells (WJ-MSCs) were cultured in Cholinergic-N-Run and Fast-N-Spheres V2 medium to obtain WT and PSEN 1 E280A cholinergic-like neurons (ChLNs, 2D) and cerebroid spheroids (CSs, 3D), respectively, and to evaluate whether ChLNs/CSs can reproduce FAD pathology. We found that irrespective of tissue source, ChLNs/CSs successfully recapitulated the AD phenotype. PSEN 1 E280A ChLNs/CSs show accumulation of iAPPß fragments, produce eAß42, present TAU phosphorylation, display OS markers (e.g., oxDJ-1, p-JUN), show loss of ΔΨm, exhibit cell death markers (e.g., TP53, PUMA, CASP3), and demonstrate dysfunctional Ca2+ influx response to ACh stimuli. However, PSEN 1 E280A 2D and 3D cells derived from MenSCs and WJ-MSCs can reproduce FAD neuropathology more efficiently and faster (11 days) than ChLNs derived from mutant iPSCs (35 days). Mechanistically, MenSCs and WJ-MSCs are equivalent cell types to iPSCs for reproducing FAD in vitro.

Genetic associations with age at dementia onset in the PSEN1 E280A Colombian kindred.

INTRODUCTION: Genetic associations with Alzheimer's disease (AD) age at onset (AAO) could reveal genetic variants with therapeutic applications. We present a large Colombian kindred with autosomal dominant AD (ADAD) as a unique opportunity to discover AAO genetic associations. METHODS: A genetic association study was conducted to examine ADAD AAO in 340 individuals with the PSEN1 E280A mutation via TOPMed array imputation. Replication was assessed in two ADAD cohorts, one sporadic early-onset AD study and four late-onset AD studies. RESULTS: 13 variants had p<1×10-7 or p<1×10-5 with replication including three independent loci with candidate associations with clusterin including near CLU. Other suggestive associations were identified in or near HS3ST1, HSPG2, ACE, LRP1B, TSPAN10, and TSPAN14. DISCUSSION: Variants with suggestive associations with AAO were associated with biological processes including clusterin, heparin sulfate, and amyloid processing. The detection of these effects in the presence of a strong mutation for ADAD reinforces their potentially impactful role.

Neuroprotective Effect of Combined Treatment with Epigallocatechin 3-Gallate and Melatonin on Familial Alzheimer's Disease PSEN1 E280A Cerebral Spheroids Derived from Menstrual Mesenchymal Stromal Cells.

BACKGROUND: Familial Alzheimer's disease (FAD) is caused by mutations in one or more of 3 genes known as A ß PP, PSEN1, and PSEN2. There are currently no effective therapies for FAD. Hence, novel therapeutics are needed. OBJECTIVE: To analyze the effect of treatment with a combination of epigallocatechin-3-gallate (EGCG) and Melatonin (N-acetyl-5-methoxytryptamine, aMT) in a cerebral spheroid (CS) 3D in vitro model of PSEN 1 E280A FAD. METHODS: We developed a CS in vitro model based on menstrual stromal cells derived from wild-type (WT) and mutant PSEN1 E280A menstrual blood cultured in Fast-N-Spheres V2 medium. RESULTS: Beta-tubulin III, choline acetyltransferase, and GFAP in both WT and mutant CSs spontaneously expressed neuronal and astroglia markers when grown in Fast-N-Spheres V2 medium for 4 or 11 days. Mutant PSEN1 CSs had significantly increased levels of intracellular AßPP fragment peptides and concomitant appearance of oxidized DJ-1 as early as 4 days, and phosphorylated tau, decreased ΔΨm, and increased caspase-3 activity were observed on Day 11. Moreover, mutant CSs were unresponsive to acetylcholine. Treatment with a combination of EGCG and aMT decreased the levels of all typical pathological markers of FAD more efficiently than did EGCG or aMT alone, but aMT failed to restore Ca2 + influx in mutant CSs and decreased the beneficial effect of EGCG on Ca2 + influx in mutant CSs. CONCLUSION: Treatment with a combination of EGCG and aMT can be of high therapeutic value due to the high antioxidant capacity and anti-amyloidogenic effect of both compounds.

Automatic Classification of Subjects of the PSEN1-E280A Family at Risk of Developing Alzheimer's Disease Using Machine Learning and Resting State Electroencephalography.

BACKGROUND: The study of genetic variant carriers provides an opportunity to identify neurophysiological changes in preclinical stages. Electroencephalography (EEG) is a low-cost and minimally invasive technique which, together with machine learning, provide the possibility to construct systems that classify subjects that might develop Alzheimer's disease (AD). OBJECTIVE: The aim of this paper is to evaluate the capacity of the machine learning techniques to classify healthy Non-Carriers (NonCr) from Asymptomatic Carriers (ACr) of PSEN1-E280A variant for autosomal dominant Alzheimer's disease (ADAD), using spectral features from EEG channels and brain-related independent components (ICs) obtained using independent component analysis (ICA). METHODS: EEG was recorded in 27 ACr and 33 NonCr. Statistical significance analysis was applied to spectral information from channels and group ICA (gICA), standardized low-resolution tomography (sLORETA) analysis was applied over the IC as well. Strategies for feature selection and classification like Chi-square, mutual informationm and support vector machines (SVM) were evaluated over the dataset. RESULTS: A test accuracy up to 83% was obtained by implementing a SVM with spectral features derived from gICA. The main findings are related to theta and beta rhythms, generated in the parietal and occipital regions, like the precuneus and superior parietal lobule. CONCLUSION: Promising models for classification of preclinical AD due to PSEN-1-E280A variant can be trained using spectral features, and the importance of the beta band and precuneus region is highlighted in asymptomatic stages, opening up the possibility of its use as a screening methodology.

(-)-Epigallocatechin-3-Gallate Diminishes Intra-and Extracellular Amyloid-Induced Cytotoxic Effects on Cholinergic-like Neurons from Familial Alzheimer's Disease PSEN1 E280A.

Alzheimer's disease (AD) is a complex neurodegenerative disease characterized by functional disruption, death of cholinergic neurons (ChNs) because of intracellular and extracellular Aß aggregates, and hyperphosphorylation of protein TAU (p-TAU). To date, there are no efficient therapies against AD. Therefore, new therapies for its treatment are in need. The goal of this investigation was to evaluate the effect of the polyphenol epigallocatechin-3-gallate (EGCG) on cholinergic-like neurons (ChLNs) bearing the mutation E280A in PRESENILIN 1 (PSEN1 E280A). To this aim, wild-type (WT) and PSEN1 E280A ChLNs were exposed to EGCG (5-50 µM) for 4 days. Untreated or treated neurons were assessed for biochemical and functional analysis. We found that EGCG (50 µM) significantly inhibited the aggregation of (i)sAPPßf, blocked p-TAU, increased ∆Ψm, decreased oxidation of DJ-1 at residue Cys106-SH, and inhibited the activation of transcription factor c-JUN and P53, PUMA, and CASPASE-3 in mutant ChLNs compared to WT. Although EGCG did not reduce (e)Aß42, the polyphenol reversed Ca2+ influx dysregulation as a response to acetylcholine (ACh) stimuli in PSEN1 E280A ChLNs, inhibited the activation of transcription factor NF-κB, and reduced the secretion of pro-inflammatory IL-6 in wild-type astrocyte-like cells (ALCs) when exposed to mutant ChLNs culture supernatant. Taken together, our findings suggest that the EGCG might be a promising therapeutic approach for the treatment of FAD.

Multi-Target Effects of the Cannabinoid CP55940 on Familial Alzheimer's Disease PSEN1 E280A Cholinergic-Like Neurons: Role of CB1 Receptor.

BACKGROUND: Alzheimer's disease (AD) is characterized by structural damage, death, and functional disruption of cholinergic neurons (ChNs) as a result of intracellular amyloid-ß (Aß) aggregation, extracellular neuritic plaques, and hyperphosphorylation of protein tau (p-Tau) overtime. OBJECTIVE: To evaluate the effect of the synthetic cannabinoid CP55940 (CP) on PSEN1 E280A cholinergic-like nerve cells (PSEN1 ChLNs)-a natural model of familial AD. METHODS: Wild type (WT) and PSEN1 ChLNs were exposed to CP (1µM) only or in the presence of the CB1 and CB2 receptors (CB1Rs, CB2Rs) inverse agonist SR141716 (1µM) and SR144528 (1µM) respectively, for 24 h. Untreated or treated neurons were assessed for biochemical and functional analysis. RESULTS: CP in the presence of both inverse agonists (hereafter SR) almost completely inhibits the aggregation of intracellular sAßPPßf and p-Tau, increases ΔΨm, decreases oxidation of DJ-1Cys106-SH residue, and blocks the activation of c-Jun, p53, PUMA, and caspase-3 independently of CB1Rs signaling in mutant ChLNs. CP also inhibits the generation of reactive oxygen species partially dependent on CB1Rs. Although CP reduced extracellular Aß42, it was unable to reverse the Ca2+ influx dysregulation as a response to acetylcholine stimuli in mutant ChLNs. Exposure to anti-Aß antibody 6E10 (1:300) in the absence or presence of SR plus CP completely recovered transient [Ca2+]i signal as a response to acetylcholine in mutant ChLNs. CONCLUSION: Taken together our findings suggest that the combination of cannabinoids, CB1Rs inverse agonists, and anti-Aß antibodies might be a promising therapeutic approach for the treatment of familial AD.

iPSCs-derived nerve-like cells from familial Alzheimer's disease PSEN 1 E280A reveal increased amyloid-beta levels and loss of the Y chromosome.

Alzheimer's disease (AD) is a progressive, degenerative disorder that mainly results in memory loss and a cognitive disorder. Although the cause of AD is still unknown, a minor percentage of AD cases are produced by genetic mutations in the presenilin-1 (PSEN1) gene. Differentiated neuronal cells derived from induced pluripotent stem cells (iPSCs) of patients can recapitulate key pathological features of AD in vitro; however, iPSCs studies focused on the p.E280 A mutation, which afflicts the largest family in the world with familial AD, have not been carried out yet. Although a link between the loss of the Y (LOY) chromosome in peripheral blood cells and risk for AD has been reported, LOY-associated phenotype has not been previously studied in PSEN1 E280 A carriers. Here, we report the reprogramming of fibroblast cells into iPSCs from a familial AD patient with the PSEN1 E280 A mutation, followed by neuronal differentiation into neural precursor cells (NPCs), and the differentiation of NPCs into differentiated neurons that lacked a Y chromosome. Although the PSEN1 E280 A iPSCs and NPCs were successfully obtained, after 8 days of differentiation, PSEN1 E280 A differentiated neurons massively died reflected by release and/ or activation of death markers, and failed to reach complete neural differentiation compared to PSEN 1 wild type cells.

Quantitative EEG analysis disease during resting and memory task in carriers and non-carriers of PS-1 E280A mutation of familial Alzheimer's / Análisis del EEG cuantitativo durante tareas de memoria y reposo en portadores y no portadores de la mutación PS-1 E280A de la enfermedad de Alzheimer

ABSTRACT Background: Alzheimer's disease is the most leading cause of dementia in the world; the mutation PS-1 E280A alters the gene of the Presenilin-1 and causes an early onset familial Alzheimer's disease. This mutation has been found in large kindred of Antioquia, Colombia. The objective of this study was to find differences revealed by electroencephalogram between healthy subjects and asymptomatic carriers that can be used as clinical markers of the disease in this population. Methods: EEG was recorded in 15 asymptomatic E280A carriers and 15 healthy non carriers during resting and a memory task using 64 channels amplifier. Two conditions in the memory task were analyzed: encoding and retrieval, the process of recording and evocating information, respectively. Power spectrum was calculated in delta (0. 5-4. 0 Hz), theta (4. 0-8. 0 Hz), alpha-1 (8. 0-10. 0 Hz), alpha-2 (10. 0-13. 0 Hz), beta (13. 0-25. 0 Hz) and gamma (25. 0-50 Hz) frequency bands for four regions of interest. Changes were evaluated in different conditions by ANOVA analysis.. Results: In resting condition a significant decrease was found in theta (p=0. 0001) and an increase in alpha-2 frequencies (p=0.037) in carriers compare with controls. During encoding of the memory task theta was significantly lower in carriers compared with controls (p=0. 008) and comparing resting versus retrieval process for each group, there was more theta synchronization in carriers. Conclusions: Early changes in theta frequencies were observed in the EEG recordings, it could be use as clinical markers in this population. Also it seems carriers activate additional cortical regions in order to conserve successful cognitive functions before clinical impairment.

RESUMEN Introducción: la enfermedad de Alzheimer es la principal causa de demencia en el mundo; la mutación PS-1 E280A altera el gen presenilin-1 y causa una variante familiar de la enfermedad que se caracteriza por una aparición temprana. La mutación se ha descubierto en un grupo de familias de Antioquia, Colombia. El objetivo de este estudio fue encontrar diferencias, a partir de registros electroencefalográficos de personas portadores de la mutación en una etapa asintomática y sujetos sanos para evaluar si pueden ser utilizadas como un marcador temprano de la enfermedad en la población portadora de la mutación. Metodología: se realizaron registros EEG en 15 portadores asintomáticos de la mutación E280A y 15 personas sanas no portadoras durante una tarea de memoria y en reposo utilizando un amplificador de 64 canales. En la tarea de memoria se evaluaron dos condiciones: codificación y evocación; el proceso de memorizar y recuperar la información, respectivamente. La potencia espectral fue calculada en las bandas de frecuencia delta (0,5-4,0 Hz), teta (4,0-8,0 Hz), alfa-1 (8,0-10,0 Hz), alfa-2 (10,0-13,0 Hz), beta (13,0-25,0 Hz) y gamma (25,0-50 Hz) para cuatro regiones de interés. Los cambios del espectro fueron evaluados por análisis de varianza ANOVA. Resultados: bajo la condición de reposo se encontró una disminución importante en la potencia de la banda teta (p=0,0001) y un incremento en la banda alfa-2 (p=0,037) en portadores comparados con controles. Durante la tarea de codificación, los portadores mostraron una disminución significativa en la banda teta (p=0,008). Al comparar reposo contra memoria de evocación se encontró una mayor sincronización en teta en los portadores de la mutación. Conclusión: se encontraron cambios tempranos de la potencia en la banda teta que pueden ser utilizados como un marcador clínico de la enfermedad en esta población. Una hipótesis adicional basada en los resultados es que los portadores necesitan ...

Evaluación de la expresión de la proteína precursora de amiloide en células sanguíneas de pacientes con la mutación E280A en el gen de la presenilina 1 / Expression of amyloid precursor protein in peripheral blood cells of patients with the E280A mutation in the presenilin-1 gene

LA enfermedad de Alzheimer es un problema mundial de salud pública. El estudio de los mecanismos moleculares responsables de su aparición permitirá avances terapéuticos y en las técnicas para la evaluación de las personas afectadas y de las que están en riesgo de desarrollar la enfermedad. La mutación E280A en el gen de la Presenilina 1 (PS1) es responsable de una forma grave de la enfermedad de Alzheimer familiar. Con el fin de conocer su efecto sobre la Proteína Precursora de Amiloide (PPA), se cuantificó la producción de esta por células de sangre periférica (mononucleares y linfocitos B) de individuos portadores de dicha mutación. Se analizaron cultivos celulares (HeLa y CHO) como controles positivos, y sangre periférica procedente de portadores sanos de la mutación, de portadores afectados y de un grupo control sano, no portador. Todas las células (HeLa, CHO, mononucleares y linfocitos B) se analizaron por citometría de flujo para detectar la PPA en la membrana citoplasmática y en el espacio intracelular. Se halló un nivel mayor de expresión de esta proteína en el interior de las células que en la membrana celular. En células HeLa y CHO los niveles de expresión intracelular fueron muy variables mientras que en los mononucleares fueron uniformemente bajos. Otros trabajos han detectado niveles mayores de la proteína en los enfermos, pero nuestros resultados indican que no hay diferencia entre los controles sanos, los portadores sanos y los portadores enfermos, lo cual indica que la mutación E280A de la PS1 no ejerce un papel directo en la expresión de la PPA en células mononucleares de sangre periférica. AUT

Abstract Alzheimer's disease is a public health problem both in Colombia and worldwide. Study of the molecular mechanisms involved in this disease may allow the development of methods for evaluation and treatment of Alzheimer's disease patients and people at risk. Mutation E280A in the presenilin-1 gene leads to the development of an aggressive form of familial Alzheimer's disease. In order to define the role of such mutation on the expression of Amyloid Precursor Protein in peripheral blood mononuclear cells and B lymphocytes we carried out a study in three groups of people, namely: healthy carriers of the mutation, affected carriers and healthy non-carriers as controls. Flow cytometry was used for the detection of Amyloid Presursor Protein in cell membranes and intracellulary; HeLa and CHO cells were used as positive controls. Expression level of Amyloid Precursor Protein was higher in the intracellular compartment than in the cell membrane. The levels of expression in the intracellular compartment of HeLa and CHO cells were variable in contrast with those of peripheral blood mononuclear cells in which they were lower but stable. Contrariwise to the results of other authors, who have detected higher levels of Amyloid Precursor Protein in Alzheimer's disease patients, our results revealed no difference between healthy controls and carriers of the E280A mutation in the presenilin-1 gene, either diseased or healthy. Our results show that this mutation does not directly change the expression of Amyloid Precursor Protein in peripheral blood mononuclear cells