The Role of intracellular amyloid β-peptide in the pathophysiology of GNE myopathy and Alzheimer's disease

The Amyloid β-peptide (Aβ) is accumulated in several diseases including GNE myopathy and Alzheimer’s disease (AD). GNE myopathy is a skeletal muscle disorder caused by biallelic mutations in the UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) gene, which codifies for a key enzyme in sialic acid biosynthesis. The mutated GNE gene encodes a hypofunctional enzyme causing a decrease in cellular sialic acid. We have found that hyposialylation favors Aβ endocytosis in skeletal muscle cells, which is dependent on clathrin and heparan sulfate proteoglycan. Furthermore, we have observed that intracellular Aβ induces apoptosis in skeletal muscle cells through the impairment of Akt signaling pathway. Accordingly, Akt phosphorylation is reduced and apoptosis is enhanced in GNE myopathy myoblasts. Finally, through a genome-wide screen in Saccharomyces cerevisiae we have identified novel modulators of Aβ toxicity including components of the mitochondrial respiratory chain and members of the Ca2+ signaling pathways. In conclusion, this study provides a better understanding of the Aβ relevance in the pathophysiology of GNE myopathy and AD.El pèptid β-amiloide (Aβ) s’acumula en diverses malalties com la miopatia de GNE i la Malaltia d’Alzheimer (MA). La miopatia de GNE és una malaltia del múscul esquelètic causada per mutacions al gen UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE), que codifica un enzim clau en la biosíntesis de l’àcid siàlic. El GNE mutat produeixen un enzim hipofuncional i una conseqüent disminució de l’àcid siàlic cel·lular. Hem vist que la hiposialització afavoreix l’endocitosi de l’Aβ en cèl·lules musculars, la qual és dependent de clatrina i de l’heparà sulfat proteoglicà. A més, hem observat que l’Aβ intracel·lular indueix l’apoptosi en cèl·lules musculars mitjançant la inhibició de l’Akt. Així, la fosforilació de l’Akt es troba reduïda i l’apoptosi induïda en mioblasts d’un pacient amb la miopatia de GNE. Finalment, a través d’un cribratge del genoma complert de Saccharomyces cerevisiae, hem identificat nous moduladors de la toxicitat per Aβ que inclouen components de la cadena respiratòria mitocondrial i membres de la via de senyalització del Ca2+. En resum, aquest estudi ofereix una millor comprensió del rol de l’Aβ en la miopatia de GNE i la MA

The Role of intracellular amyloid β-peptide in the pathophysiology of GNE myopathy and Alzheimer's disease

The Amyloid β-peptide (Aβ) is accumulated in several diseases including GNE myopathy and Alzheimer’s disease (AD). GNE myopathy is a skeletal muscle disorder caused by biallelic mutations in the UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) gene, which codifies for a key enzyme in sialic acid biosynthesis. The mutated GNE gene encodes a hypofunctional enzyme causing a decrease in cellular sialic acid. We have found that hyposialylation favors Aβ endocytosis in skeletal muscle cells, which is dependent on clathrin and heparan sulfate proteoglycan. Furthermore, we have observed that intracellular Aβ induces apoptosis in skeletal muscle cells through the impairment of Akt signaling pathway. Accordingly, Akt phosphorylation is reduced and apoptosis is enhanced in GNE myopathy myoblasts. Finally, through a genome-wide screen in Saccharomyces cerevisiae we have identified novel modulators of Aβ toxicity including components of the mitochondrial respiratory chain and members of the Ca2+ signaling pathways. In conclusion, this study provides a better understanding of the Aβ relevance in the pathophysiology of GNE myopathy and AD.El pèptid β-amiloide (Aβ) s’acumula en diverses malalties com la miopatia de GNE i la Malaltia d’Alzheimer (MA). La miopatia de GNE és una malaltia del múscul esquelètic causada per mutacions al gen UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE), que codifica un enzim clau en la biosíntesis de l’àcid siàlic. El GNE mutat produeixen un enzim hipofuncional i una conseqüent disminució de l’àcid siàlic cel·lular. Hem vist que la hiposialització afavoreix l’endocitosi de l’Aβ en cèl·lules musculars, la qual és dependent de clatrina i de l’heparà sulfat proteoglicà. A més, hem observat que l’Aβ intracel·lular indueix l’apoptosi en cèl·lules musculars mitjançant la inhibició de l’Akt. Així, la fosforilació de l’Akt es troba reduïda i l’apoptosi induïda en mioblasts d’un pacient amb la miopatia de GNE. Finalment, a través d’un cribratge del genoma complert de Saccharomyces cerevisiae, hem identificat nous moduladors de la toxicitat per Aβ que inclouen components de la cadena respiratòria mitocondrial i membres de la via de senyalització del Ca2+. En resum, aquest estudi ofereix una millor comprensió del rol de l’Aβ en la miopatia de GNE i la MA

The structure and function of actin cytoskeleton in mature glutamatergic dendritic spines

Dendritic spines are actin-rich protrusions from the dendritic shaft, considered to be the locus where most synapses occur, as they receive the vast majority of excitatory connections in the central nervous system (CNS). Interestingly, hippocampal spines are plastic structures that contain a dense array of molecules involved in postsynaptic signaling and synaptic plasticity. Since changes in spine shape and size are correlated with the strength of excitatory synapses, spine morphology directly reflects spine function. Therefore several neuropathologies are associated with defects in proteins located at the spines. The present work is focused on the spine actin cytoskeleton attending to its structure and function mainly in glutamatergic neurons. It addresses the study of the structural plasticity of dendritic spines associated with long-term potentiation (LTP) and the mechanisms that underlie learning and memory formation. We have integrated the current knowledge on synaptic proteins to relate this plethora of molecules with actin and actin-binding proteins. We further included recent findings that outline key uncharacterized proteins that would be useful to unveil the real ultrastructure and function of dendritic spines. Furthermore, this review is directed to understand how such spine diversity and interplay contributes to the regulation of spine morphogenesis and dynamics. It highlights their physiological relevance in the brain function, as well as it provides insights for pathological processes affecting dramatically dendritic spines, such as Alzheimer's disease.This work was supported by the Plan Estatal de I+D+i 2013-2016 and the ISCIII Subdirección General de Evaluación y Fomento de la Investigación (Grants PI13/00408, PI10/00587 and Red HERACLES RD12/0042/0014) and FEDER Funds; FEDER Funds; Generalitat de Catalunya (SGR09-1369); and Fundació la Marató de TV3 (100310)

The structure and function of actin cytoskeleton in mature glutamatergic dendritic spines

Dendritic spines are actin-rich protrusions from the dendritic shaft, considered to be the locus where most synapses occur, as they receive the vast majority of excitatory connections in the central nervous system (CNS). Interestingly, hippocampal spines are plastic structures that contain a dense array of molecules involved in postsynaptic signaling and synaptic plasticity. Since changes in spine shape and size are correlated with the strength of excitatory synapses, spine morphology directly reflects spine function. Therefore several neuropathologies are associated with defects in proteins located at the spines. The present work is focused on the spine actin cytoskeleton attending to its structure and function mainly in glutamatergic neurons. It addresses the study of the structural plasticity of dendritic spines associated with long-term potentiation (LTP) and the mechanisms that underlie learning and memory formation. We have integrated the current knowledge on synaptic proteins to relate this plethora of molecules with actin and actin-binding proteins. We further included recent findings that outline key uncharacterized proteins that would be useful to unveil the real ultrastructure and function of dendritic spines. Furthermore, this review is directed to understand how such spine diversity and interplay contributes to the regulation of spine morphogenesis and dynamics. It highlights their physiological relevance in the brain function, as well as it provides insights for pathological processes affecting dramatically dendritic spines, such as Alzheimer's disease.This work was supported by the Plan Estatal de I+D+i 2013-2016 and the ISCIII Subdirección General de Evaluación y Fomento de la Investigación (Grants PI13/00408, PI10/00587 and Red HERACLES RD12/0042/0014) and FEDER Funds; FEDER Funds; Generalitat de Catalunya (SGR09-1369); and Fundació la Marató de TV3 (100310)

Increased amyloid β-peptide uptake in skeletal muscle is induced by hyposialylation and may account for apoptosis in GNE myopathy

GNE myopathy is an autosomal recessive muscular disorder of young adults characterized by progressive skeletal muscle weakness and wasting. It is caused by a mutation in the UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) gene, which encodes a key enzyme in sialic acid biosynthesis. The mutated hypofunctional GNE is associated with intracellular accumulation of amyloid β-peptide (Aβ) in patient muscles through as yet unknown mechanisms. We found here for the first time that an experimental reduction in sialic acid favors Aβ1-42 endocytosis in C2C12 myotubes, which is dependent on clathrin and heparan sulfate proteoglycan. Accordingly, Aβ1-42 internalization in myoblasts from a GNE myopathy patient was enhanced. Next, we investigated signal changes triggered by Aβ1-42 that may underlie toxicity. We observed that p-Akt levels are reduced in step with an increase in apoptotic markers in GNE myopathy myoblasts compared to control myoblasts. The same results were experimentally obtained when Aβ1-42 was overexpressed in myotubes. Hence, we propose a novel disease mechanism whereby hyposialylation favors Aβ1-42 internalization and the subsequent apoptosis in myotubes and in skeletal muscle from GNE myopathy patients.This work was supported by Fundació la Marató de TV3 (100310); Supported by the Plan Estatal de I+D+I 2013-2016 and the ISCIII-Subdirección General de Evaluación y Fomento de la Investigación (Grants PI13/00408, PI08/00574 and Red HERACLES RD12/0042/0014) and FEDER; Generalitat de Catalunya (SGR09-1369)

Vitamin E dietary supplementation improves neurological symptoms and decreases c-Abl/p73 activation in niemann-pick C mice

Niemann-Pick C (NPC) disease is a fatal neurodegenerative disorder characterized by the accumulation of free cholesterol in lysosomes. We have previously reported that oxidative stress is the main upstream stimulus activating the proapoptotic c-Abl/p73 pathway in NPC neurons. We have also observed accumulation of vitamin E in NPC lysosomes, which could lead to a potential decrease of its bioavailability. Our aim was to determine if dietary vitamin E supplementation could improve NPC disease in mice. NPC mice received an alpha-tocopherol (α-TOH) supplemented diet and neurological symptoms, survival, Purkinje cell loss, α-TOH and nitrotyrosine levels, astrogliosis, and the c-Abl/p73 pathway functions were evaluated. In addition, the effect of α-TOH on the c-Abl/p73 pathway was evaluated in an in vitro NPC neuron model. The α-TOH rich diet delayed loss of weight, improved coordination and locomotor function and increased the survival of NPC mice. We found increased Purkinje neurons and α-TOH levels and reduced astrogliosis, nitrotyrosine and phosphorylated p73 in cerebellum. A decrease of c-Abl/p73 activation was also observed in the in vitro NPC neurons treated with α-TOH. In conclusion, our results show that vitamin E can delay neurodegeneration in NPC mice and suggest that its supplementation in the diet could be useful for the treatment of NPC patients.This study was supported by grants from the Fondo Nacional de Desarrollo Científico y/nTecnológico (FONDECYT) (grant numbers 1120512 to A.R.A, and 1110310 to S.Z); Fondo de/nFomento al Desarrollo Científico y Tecnológico FONDEF D10I1077 (to A.R.A and S.Z.);/nFondo Nacional de Desarrollo de Areas Prioritarias, FONDAP, Project no. 15090007, Center for/nGenome Regulation (CGR) to S.Z.; Generalitat de Catalunya (SGR2009-1369) and La Marató de TV3/n(No. 100310

Increased amyloid β-peptide uptake in skeletal muscle is induced by hyposialylation and may account for apoptosis in GNE myopathy

GNE myopathy is an autosomal recessive muscular disorder of young adults characterized by progressive skeletal muscle weakness and wasting. It is caused by a mutation in the UDP-N-acetylglucosamine 2-epimerase/N-acetylmannosamine kinase (GNE) gene, which encodes a key enzyme in sialic acid biosynthesis. The mutated hypofunctional GNE is associated with intracellular accumulation of amyloid β-peptide (Aβ) in patient muscles through as yet unknown mechanisms. We found here for the first time that an experimental reduction in sialic acid favors Aβ1-42 endocytosis in C2C12 myotubes, which is dependent on clathrin and heparan sulfate proteoglycan. Accordingly, Aβ1-42 internalization in myoblasts from a GNE myopathy patient was enhanced. Next, we investigated signal changes triggered by Aβ1-42 that may underlie toxicity. We observed that p-Akt levels are reduced in step with an increase in apoptotic markers in GNE myopathy myoblasts compared to control myoblasts. The same results were experimentally obtained when Aβ1-42 was overexpressed in myotubes. Hence, we propose a novel disease mechanism whereby hyposialylation favors Aβ1-42 internalization and the subsequent apoptosis in myotubes and in skeletal muscle from GNE myopathy patients.This work was supported by Fundació la Marató de TV3 (100310); Supported by the Plan Estatal de I+D+I 2013-2016 and the ISCIII-Subdirección General de Evaluación y Fomento de la Investigación (Grants PI13/00408, PI08/00574 and Red HERACLES RD12/0042/0014) and FEDER; Generalitat de Catalunya (SGR09-1369)

Rare variants in calcium homeostasis modulator 1 (CALHM1) found in early onset alzheimer's disease patients alter calcium homeostasis

Calcium signaling in the brain is fundamental to the learning and memory process and there is evidence to suggest that its dysfunction is involved in the pathological pathways underlying Alzheimer’s disease (AD). Recently, the calcium hypothesis of AD has received support with the identification of the non-selective Ca2+-permeable channel CALHM1. A genetic polymorphism (p. P86L) in CALHM1 reduces plasma membrane Ca2+ permeability and is associated with an earlier age-at-onset of AD. To investigate the role of CALHM1 variants in early-onset AD (EOAD), we sequenced all CALHM1 coding regions in three independent series comprising 284 EOAD patients and 326 controls. Two missense mutations in patients (p.G330D and p.R154H) and one (p.A213T) in a control individual were identified. Calcium imaging analyses revealed that while the mutation found in a control (p.A213T) behaved as wild-type CALHM1 (CALHM1-WT), a complete abolishment of the Ca2+ influx was associated with the mutations found in EOAD patients (p.G330D and p.R154H). Notably, the previously reported p. P86L mutation was associated with an intermediate Ca2+ influx between the CALHM1-WT and the p.G330D and p.R154H mutations. Since neither expression of wild-type nor mutant CALHM1 affected amyloid ß-peptide (Aß) production or Aß-mediated cellular toxicity, we conclude that rare genetic variants in CALHM1 lead to Ca2+ dysregulation and may contribute to the risk of EOAD through a mechanism independent from the classical Aß cascade.This study was supported by grants from Instituto de Salud Carlos III (PI12/01311, PI10/000587, Red HERACLES RD12/0042/0014), Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED, Spain), Spanish Ministry of Economy and Competiveness (SAF2012-38140), FEDER Funds, and Generalitat de Catalunya (SGR05-266). Council of the Academy of Finland, EVO grant 5772708 of Kuopio University Hospital, the Strategic Funding of the University on Eastern Finland (UEF-Brain) (to M.H and H.S). M.A.V. is the recipient of an ICREA Academia Awar

Epigallocatechin-3-gallate improves cardiac hypertrophy and short-term memory deficits in a Williams-Beuren syndrome mouse model

Williams-Beuren syndrome (WBS) is a neurodevelopmental disorder caused by a heterozygous deletion of 26-28 genes at chromosome band 7q11.23. The complete deletion (CD) mouse model mimics the most common deletion found in WBS patients and recapitulates most neurologic features of the disorder along with some cardiovascular manifestations leading to significant cardiac hypertrophy with increased cardiomyocytes' size. Epigallocatechin-3-gallate (EGCG), the most abundant catechin found in green tea, has been associated with potential health benefits, both on cognition and cardiovascular phenotypes, through several mechanisms. We aimed to investigate the effects of green tea extracts on WBS-related phenotypes through a phase I clinical trial in mice. After feeding CD animals with green tea extracts dissolved in the drinking water, starting at three different time periods (prenatal, youth and adulthood), a set of behavioral tests and several anatomical, histological and molecular analyses were performed. Treatment resulted to be effective in the reduction of cardiac hypertrophy and was also able to ameliorate short-term memory deficits of CD mice. Taken together, these results suggest that EGCG might have a therapeutic and/or preventive role in the management of WBS.This work was supported by the Spanish Ministry of Economy and Competitiveness (grant SAF2012-40036, and SAF2016-78508-R(AEI/MINEICO/FEDER, UE) to VC; “Programa de Excelencia Maria de Maeztu” MDM-2014-0370, the Generalitat de Catalunya (2014SGR1468 and ICREA Acadèmia) to LAPJ; AGAUR FI-DGR/2013 fellowship to CB

Epigallocatechin-3-gallate improves cardiac hypertrophy and short-term memory deficits in a Williams-Beuren syndrome mouse model

Williams-Beuren syndrome (WBS) is a neurodevelopmental disorder caused by a heterozygous deletion of 26-28 genes at chromosome band 7q11.23. The complete deletion (CD) mouse model mimics the most common deletion found in WBS patients and recapitulates most neurologic features of the disorder along with some cardiovascular manifestations leading to significant cardiac hypertrophy with increased cardiomyocytes' size. Epigallocatechin-3-gallate (EGCG), the most abundant catechin found in green tea, has been associated with potential health benefits, both on cognition and cardiovascular phenotypes, through several mechanisms. We aimed to investigate the effects of green tea extracts on WBS-related phenotypes through a phase I clinical trial in mice. After feeding CD animals with green tea extracts dissolved in the drinking water, starting at three different time periods (prenatal, youth and adulthood), a set of behavioral tests and several anatomical, histological and molecular analyses were performed. Treatment resulted to be effective in the reduction of cardiac hypertrophy and was also able to ameliorate short-term memory deficits of CD mice. Taken together, these results suggest that EGCG might have a therapeutic and/or preventive role in the management of WBS.This work was supported by the Spanish Ministry of Economy and Competitiveness (grant SAF2012-40036, and SAF2016-78508-R(AEI/MINEICO/FEDER, UE) to VC; “Programa de Excelencia Maria de Maeztu” MDM-2014-0370, the Generalitat de Catalunya (2014SGR1468 and ICREA Acadèmia) to LAPJ; AGAUR FI-DGR/2013 fellowship to CB