MiR-7 controls cholesterol biosynthesis through posttranscriptional regulation of DHCR24 expression

Dysregulation of cholesterol homeostasis is associated with several pathologies including cardiovascular diseases and neurological disorders such as Alzheimer's disease (AD). MicroRNAs (miRNAs) have emerged as key post-transcriptional regulators of cholesterol metabolism. We previously established the role of miR-7 in regulating insulin resistance and amyloidosis, which represents a common pathological feature between type 2 diabetes and AD. We show here an additional metabolic function of miR-7 in cholesterol biosynthesis. We found that miR-7 blocks the last steps of the cholesterol biosynthetic pathway in vitro by targeting relevant genes including DHCR24 and SC5D posttranscriptionally. Intracranial infusion of miR-7 on an adeno-associated viral vector reduced the expression of DHCR24 in the brain of wild-type mice, supporting in vivo miR-7 targeting. We also found that cholesterol regulates endogenous levels of miR-7 in vitro, correlating with transcriptional regulation through SREBP2 binding to its promoter region. In parallel to SREBP2 inhibition, the levels of miR-7 and hnRNPK (the host gene of miR-7) were concomitantly reduced in brain in a mouse model of Niemann Pick type C1 disease and in murine fatty liver, which are both characterized by intracellular cholesterol accumulation. Taken together, the results establish a novel regulatory feedback loop by which miR-7 modulates cholesterol homeostasis at the posttranscriptional level, an effect that could be exploited for therapeutic interventions against prevalent human diseases.This work was supported by the “Talento Program” from the Madrid Government, Spain (2017-T1/BMD-5333 and 2021-5A/BMD-20964), (RTI2018-095061-B-I00) and (PID2021-128264OB-I00) funded by MCIN/AEI/10.13039/501100011033 and “ERDF A way of making Europe” by the European Union (to CMR); Consejería de Educación e Investigación from the Madrid Government, Spain: “Convocatoria de ayudas para la contratación de ayudantes de investigación” (PEJ-2018-AI/BMD-9724) (to CMR and MT-P); “Convocatoria de ayudas para la contratación de investigadores postdoctorales” (PEDJ-2018-POST/BDM-8900) (to CMR and AP-G) and “Convocatoria de ayudas para la contratación de investigadores predoctorales” (PEJD-2019-PRE/BMD-14499) (to CMR and YM-M) from the Madrid Government, Spain; (RTI2018-098113-B-I00) (to RB and DGC) and (PID2021-122766OB-I00) (to AMV) and (PID2020-112830RB-I00) (to MD-L) funded by MCIN/AEI/10.13039/501100011033 and “ERDF A way of making Europe” by the European Union; (PI18/01152 and PI21/01173) funded by Instituto de Salud Carlos III, (ISCIII) (to OP); Centro de Investigación Biomédica en Red de Diabetes y Enfermedades Metabólicas Asociadas (CIBERdem), Instituto de Salud Carlos III, Madrid, Spain (research contract of P-R) and 2021-5A/BMD-20964 (research contract of VP-M). We thank the Quantification and Molecular Characterization Unit and the Lipid and Lipoprotein Unit (IRYCIS) for support

Alteraciones de mGluR5 y el sistema endocannabinoide en la patología psiquiátrica de la enfermedad de Niemann Pick Tipo C

Tesis Doctoral inédita leída en la Universidad Autónoma de Madrid, Facultad de Ciencias, Departamento de Biología Molecular. Fecha de Lectura: 16-12-2022Esta tesis tiene embargado el acceso al texto completo hasta el 16-06-2024Niemann Pick Tipo C (NPC) es una enfermedad de depósito lisosomal causada por mutaciones en la proteína transportadora de colesterol NPC1, cuya deficiencia provoca la acumulación de este lípido en los compartimentos endolisosomales y con ello su disfunción. Aunque las mutaciones afectan a todas las células del organismo, las neuronas son especialmente sensibles a la deficiencia de esta proteína. Por ello, NPC es considerada principalmente una enfermedad neurológica. Además de problemas motores que llevan a la ataxia, los pacientes de NPC sufren alteraciones cognitivas, trastornos psiquiátricos y convulsiones epilépticas que dificultan el correcto diagnóstico de esta enfermedad rara y que afectan a la calidad de vida. Esta sintomatología apunta a las alteraciones sinápticas como eventos patológicos clave en el desarrollo de la NPC. Relevante en este contexto es el receptor metabotrópico de glutamato 5 (mGluR5), acoplado a proteínas G e implicado en la regulación fina de la plasticidad sináptica. Sus alteraciones se han relacionado con trastornos psiquiátricos como depresión, bipolaridad, ansiedad, esquizofrenia o autismo. Además de su implicación directa en estos trastornos, este receptor regula al sistema endocannabinoide (SECB) cuyas anomalías se han relacionado con alteraciones psiquiátricas similares. Dada su importancia para la plasticidad sináptica, la función de mGluR5 está muy regulada. La distribución celular del receptor, que puede estar presente en la membrana plasmática o en el interior celular, es uno de los mecanismos de control, así como su localización dinámica en las balsas lipídicas de la membrana enriquecidas en colesterol. En esta tesis doctoral se postuló que las anomalías en colesterol en NPC alteran mGluR5 afectando a la plasticidad sináptica y contribuyendo a los trastornos psiquiátricos típicos de la enfermedad. Los resultados obtenidos utilizando diferentes modelos para NPC, que incluyen un ratón mutante para NPC1, modelos celulares neuronales deficientes en esta proteína y tejido de pacientes NPC, confirman esta hipótesis. Demuestran que el tráfico alterado de colesterol en neuronas, provocado por la deficiencia de NPC1, afecta a los niveles, distribución y funcionalidad de mGluR5 al interferir en el correcto funcionamiento de la vía endolisosomal. El receptor se acumula en los compartimentos endolisosomales donde no se degrada eficientemente e hiperactiva vías de señalización, provocando alteraciones en la plasticidad sináptica y en sistemas dependientes como el SECB. Estas anomalías contribuyen a los trastornos psiquiátricos en el ratón modelo para NPC que pueden revertirse mediante la modulación farmacológica de mGluR5 y del SECB. Consideramos que la presente tesis doctoral muestra por primera vez que las alteraciones psiquiátricas características de NPC son debidas, al menos parcialmente, a alteraciones en mGluR5 y proporciona nuevas estrategias terapéuticas no invasivas para mejorar la calidad de vida de los pacientes que sufren esta fatal enfermeda

Inhibition of fatty acid amide hydrolase prevents pathology in neurovisceral acid sphingomyelinase deficiency by rescuing defective endocannabinoid signaling.

Acid sphingomyelinase deficiency (ASMD) leads to cellular accumulation of sphingomyelin (SM), neurodegeneration, and early death. Here, we describe the downregulation of the endocannabinoid (eCB) system in neurons of ASM knockout (ASM‐KO) mice and a ASMD patient. High SM reduced expression of the eCB receptor CB1 in neuronal processes and induced its accumulation in lysosomes. Activation of CB1 receptor signaling, through inhibition of the eCB‐degrading enzyme fatty acid amide hydrolase (FAAH), reduced SM levels in ASM‐KO neurons. Oral treatment of ASM‐KO mice with a FAAH inhibitor prevented SM buildup; alleviated inflammation, neurodegeneration, and behavioral alterations; and extended lifespan. This treatment showed benefits even after a single administration at advanced disease stages. We also found CB1 receptor downregulation in neurons of a mouse model and a patient of another sphingolipid storage disorder, Niemann–Pick disease type C (NPC). We showed the efficacy of FAAH inhibition to reduce SM and cholesterol levels in NPC patient‐derived cells and in the brain of a NPC mouse model. Our findings reveal a pathophysiological crosstalk between neuronal SM and the eCB system and offer a new treatment for ASMD and other sphingolipidoses.We are grateful to Steven Laffoon and the Wylder Nation Foundation (www.wyldernation.org) for the continuous support. We thank Dr. Melissa Wasserstein at the Children's Hospital at Montefiore for insightful comments and Dr. A. Rábano at the Fundación Cien and Dr. E. Martínez‐Sáez at the Hospital Vall d′Hebron for donating brain samples. We also thank the microscope facility at the CBMSO for their assistance with the confocal imaging and the institutional support of the Fundación Ramón Areces to the CBMSO. This work was funded by the Wylder Nation Foundation and by grants from the Spanish Ministry of Science and Education (SAF2017‐87698‐R) (AEI/FEDER, UE) to M.D.L and a MERIT award from the National Institutes of Health (2R37HD028607) to E.H.S.Peer reviewe

Cholesterol-24-hydroxylase (CYP46) in the old brain: Analysis of positive populations and factors triggering its expression in astrocytes

Cholesterol-24-hydroxylase (CYP46), a member of the cytochrome P450 superfamily of enzymes, is selectively expressed in the brain and is mainly responsible for cholesterol turnover in the central nervous system. Although increased cyp46A1 gene expression has been linked to cognitive alterations in aging and observed in neurodegenerative diseases and after traumatic brain injury, a detailed characterization of the brain regions and cell types in which CYP46 is expressed in old individuals has not been performed. Using immunohistochemistry and immunofluorescence, we investigated the specific regions and cell populations in the brain, in which cyp46A1 is expressed in 24-month-old mice. We found that CYP46 is localized in the same neuronal populations in young and old brains, mainly in the hippocampus, in cortical layers, and in Purkinje neurons of the cerebellum. No increase in CYP46 levels was found in astrocytes in old mice brains, in primary astrocyte-neuron cocultures aged in vitro, or in primary cultures of senescent astrocytes. However, interleukin-6 treatment strongly induced cyp46A1 expression in reactive astrocytes characterized by high GFAP levels but had no effect in nonactivated astrocytes. Our data suggest that cholesterol-24-hydroxylase expression is triggered in reactive astrocytes in response to proinflammatory signals, probably as part of a response mechanism to injury.Fil: Cataldi, Gabriel Enrique. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra; ArgentinaFil: Elorza, Setiembre Delfina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra; ArgentinaFil: Toledano Zaragoza, Ana. Centro de Biología Molecular Severo Ochoa.; EspañaFil: de Olmos, Soledad. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra; ArgentinaFil: Cragnolini, Andrea Beatriz. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigaciones Biológicas y Tecnológicas. Universidad Nacional de Córdoba. Facultad de Ciencias Exactas, Físicas y Naturales. Instituto de Investigaciones Biológicas y Tecnológicas; ArgentinaFil: Martín, Mauricio Gerardo. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra. Universidad Nacional de Córdoba. Instituto de Investigación Médica Mercedes y Martín Ferreyra; Argentin

Ellagic acid and its metabolites urolithins A/B ameliorate most common disease phenotypes in cellular and mouse models for lysosomal storage disorders by enhancing extracellular vesicle secretion

Niemann Pick diseases types A (NPDA) and C (NPDC) are lysosomal storage disorders (LSDs) leading to cognitive impairment, neurodegeneration, and early death. NPDA and NPDC have different genetic origins, being caused by mutations in the acid sphingomyelinase (ASM) or the cholesterol transport protein NPC1, respectively. However, they share a common pathological hallmark in the accumulation of lipids in the endolysosomal compartment. Here, we tested the hypothesis that polyphenols reduce lipid overload in NPD cells by enhancing the secretion of extracellular vesicles (ECVs). We show that among the polyphenols tested, the ellagic acid metabolites, urolithin A and B, were the safest and most efficient in increasing ECV secretion. They reduced levels of accumulating lipids and lysosomal size and permeabilization in cultured bone marrow-derived macrophages and neurons from ASMko and NPC1 mutant mice, which mimic NPDA and NPDC, respectively. Moreover, oral treatment with ellagic acid reduced lipid levels, ameliorated lysosomal alterations, and diminished microglia activation in the brain of NPD mice. These results support the therapeutic value of ECV secretion and polyphenols for NPDs, which may also help treat other LSDs characterized by intracellular lipid overload