Neural stem cells in the adult olfactory bulb core generate mature neurons in vivo

Although previous studies suggest that neural stem cells (NSCs) exist in the adult olfactory bulb (OB), their location, identity, and capacity to generate mature neurons in vivo has been little explored. Here, we injected enhanced green fluorescent protein (EGFP)-expressing retroviral particles into the OB core of adult mice to label dividing cells and to track the differentiation/maturation of any neurons they might generate. EGFP-labeled cells initially expressed adult NSC markers on days 1 to 3 postinjection (dpi), including Nestin, GLAST, Sox2, Prominin-1, and GFAP. EGFP+ -doublecortin (DCX) cells with a migratory morphology were also detected and their abundance increased over a 7-day period. Furthermore, EGFP-labeled cells progressively became NeuN+ neurons, they acquired neuronal morphologies, and they became immunoreactive for OB neuron subtype markers, the most abundant representing calretinin expressing interneurons. OB-NSCs also generated glial cells, suggesting they could be multipotent in vivo. Significantly, the newly generated neurons established and received synaptic contacts, and they expressed presynaptic proteins and the transcription factor pCREB. By contrast, when the retroviral particles were injected into the subventricular zone (SVZ), nearly all (98%) EGFP+ -cells were postmitotic when they reached the OB core, implying that the vast majority of proliferating cells present in the OB are not derived from the SVZ. Furthermore, we detected slowly dividing label-retaining cells in this region that could correspond to the population of resident NSCs. This is the first time NSCs located in the adult OB core have been shown to generate neurons that incorporate into OB circuits in vivo

Estudio de células madre neurales del bulbo olfatorio y su relación por el nicho astroglial

Tesis doctoral inédita leída en la Universidad Autónoma de Madrid. Facultad de Medicina, Departamento de Anatomía, Hstología y Neurociencia. Fecha de lectura: 21 de Septiembre de 201

TNF and IL6/Jak2 signaling pathways are the main contributors of the glia-derived neuroinflammation present in Lafora disease, a fatal form of progressive myoclonus epilepsy

Lafora disease (LD; OMIM#254780) is a rare form of progressive myoclonus epilepsy (prevalence <1:1,000,000) characterized by the accumulation of insoluble deposits of aberrant glycogen (polyglucosans), named Lafora bodies, in the brain but also in peripheral tissues. LD is the most severe form of the group of progressive myoclonus epilepsies, since patients present a rapid deterioration and dementia with amplification of seizures, leading to death after a decade from the onset of the first symptoms. We have recently described that reactive glia-derived neuroinflammation should be considered a novel hallmark of LD since we observed a florid upregulation of differentially expressed genes in both LD mouse lines, which were mainly related to mediators of inflammatory response. In this work, we define an upregulation of the expression of mediators of the TNF and IL6/JAK2 signaling pathways in LD. In addition, we describe the activation of the non-canonical form of the inflammasome. Furthermore, we describe the infiltration of peripheral immune cells in the brain parenchyma, which could aggravate glia-derived neuroinflammation. Finally, we describe CXCL10 and S100b as blood biomarkers of the disease, which will allow the study of the progression of the disease using serum blood samples. We consider that the identification of these initial inflammatory changes in LD will be very important to implement possible anti-inflammatory therapeutic strategies to prevent the development of the disease

Exploring the role of galectins in cancer: in vitro and in vivo approaches

Galectins have been linked to tumorigenesis since 1975, even before this family of proteins was given its name. Since then, hundreds of papers have analyzed the role of different galectins in cancer development and progression, deciphering their involvement in many different pathological events, from the regulation of cell cycle, to angiogenesis, metastasis, and immune attack evasion. Importantly, the tumor galectin profile is often altered in many cancers and aberrant levels of some of the members of this family have been considered in diagnosis and frequently correlated with patient prognosis and clinicopathological characteristics. In this chapter, we summarize most frequent techniques employed in cancer research to interrogate the role of galectins, using Gal-1 to illustrate one member of the family and pancreatic cancer as an experimental model. We will cover from techniques employed to detect their expression (tissue and blood samples) to the most frequent tools used to change expression levels and the cell line-based in vitro studies and murine preclinical models used to explore their role in tumor progression and/or clinical translation

Deciphering the Polyglucosan Accumulation Present in Lafora Disease Using an Astrocytic Cellular Model

15 páginas, 7 figuras, 1 tablaLafora disease (LD) is a neurological disorder characterized by progressive myoclonus epilepsy. The hallmark of the disease is the presence of insoluble forms of glycogen (polyglucosan bodies, or PGBs) in the brain. The accumulation of PGBs is causative of the pathophysiological features of LD. However, despite the efforts made by different groups, the question of why PGBs accumulate in the brain is still unanswered. We have recently demonstrated that, in vivo, astrocytes accumulate most of the PGBs present in the brain, and this could lead to astrocyte dysfunction. To develop a deeper understanding of the defects present in LD astrocytes that lead to LD pathophysiology, we obtained pure primary cultures of astrocytes from LD mice from the postnatal stage under conditions that accumulate PGBs, the hallmark of LD. These cells serve as novel in vitro models for studying PGBs accumulation and related LD dysfunctions. In this sense, the metabolomics of LD astrocytes indicate that they accumulate metabolic intermediates of the upper part of the glycolytic pathway, probably as a consequence of enhanced glucose uptake. In addition, we also demonstrate the feasibility of using the model in the identification of different compounds that may reduce the accumulation of polyglucosan inclusions.This work was supported by a grant from the Spanish Ministry of Science and Innovation PID2020-112972RB-I00, a grant from la Fundació La Marató TV3 (202032), and a grant from the National Institutes of Health P01NS097197, which established the Lafora Epilepsy Cure Initiative (LECI), to P.S.Peer reviewe

Selective PARP 2 inhibition hampers pancreatic cancer tumorigenesis and progression

Trabajo presentado en el 18th ASEICA International Congress, celebrado en Santiago de Compostela (España), del 16 al 18 de noviembre de 202

Proneural factors Ascl1 and Neurog2 contribute to neuronal subtype identities by establishing distinct chromatin landscapes

12 páginas, 7 figurasDevelopmental programs that generate the astonishing neuronal diversity of the nervous system are not completely understood and thus present a major challenge for clinical applications of guided cell differentiation strategies. Using direct neuronal programming of embryonic stem cells, we found that two main vertebrate proneural factors, Ascl1 and neurogenin 2 (Neurog2), induce different neuronal fates by binding to largely different sets of genomic sites. Their divergent binding patterns are not determined by the previous chromatin state, but are distinguished by enrichment of specific E-box sequences that reflect the binding preferences of the DNA-binding domains. The divergent Ascl1 and Neurog2 binding patterns result in distinct chromatin accessibility and enhancer activity profiles that differentially shape the binding of downstream transcription factors during neuronal differentiation. This study provides a mechanistic understanding of how transcription factors constrain terminal cell fates, and it delineates the importance of choosing the right proneural factor in neuronal reprogramming strategies.This work is supported by NICHD (R01HD079682) and Project ALS (A13-0416) to E.O.M. and by NYSTEM pre-doctoral training grant (C026880) to B.A. S.M. is supported by NIGMS (R01GM125722) and the National Science Foundation ABI Innovation Grant No. DBI1564466. Any opinions, findings and conclusions or recommendations expressed in this material are those of the authors and do not necessarily reflect the views of the National Science Foundation. M.R. is supported by NYU MSTP (T32GM007308) and Developmental Genetics T32 (T32HD007520) grants. N.F. and M.M.E. are supported by ERC Starting Grant (2011-281920). The authors would like to thank Link Tejavibulya and Apeksha Ashokkumar for their help with molecular biology; Mohammed Khalfan for his help with scRNA-seq analysis. Michael Cammer from the NYU Medical Center Microscopy Core for the ImageJ script used in calcium imaging analysis; and NYU Genomics Core facility. Finally, we would like to thank Steve Small, Nikos Konstantinidis, Pinar Onal, Orly Wapinski, Sevinç Ercan, Chris Rushlow, Claude Desplan and Mazzoni lab members for their helpful suggestions on the manuscript.Peer reviewe

Reactive Glia-Derived Neuroinflammation: a Novel Hallmark in Lafora Progressive Myoclonus Epilepsy That Progresses with Age

15 páginas, 7 figuras, 3 tablas. Contiene en material suplementario adjunto: 2 figuras y 8 tablas.Lafora disease (LD) is a rare, fatal form of progressive myoclonus epilepsy. The molecular basis of this devastating disease is still poorly understood, and no treatment is available yet, which leads to the death of the patients around 10 years from the onset of the first symptoms. The hallmark of LD is the accumulation of insoluble glycogen-like inclusions in the brain and peripheral tissues, as a consequence of altered glycogen homeostasis. In addition, other determinants in the pathophysiology of LD have been suggested, such as proteostasis impairment, with reduction in autophagy, and oxidative stress, among others. In order to gain a general view of the genes involved in the pathophysiology of LD, in this work, we have performed RNA-Seq transcriptome analyses of whole-brain tissue from two independent mouse models of the disease, namely Epm2a-/- and Epm2b-/- mice, at different times of age. Our results provide strong evidence for three major facts: first, in both models of LD, we found a common set of upregulated genes, most of them encoding mediators of inflammatory response; second, there was a progression with the age in the appearance of these inflammatory markers, starting at 3 months of age; and third, reactive glia was responsible for the expression of these inflammatory genes. These results clearly indicate that neuroinflammation is one of the most important traits to be considered in order to fully understand the pathophysiology of LD, and define reactive glia as novel therapeutic targets in the disease.This work was supported by grants from the Spanish Ministry of Economy and Competitiveness SAF2014-54604-C3-1-R and SAF2017-83151-R, a grant from Fundación Ramón Areces (CIVP18A3935) and a grant from the National Institute of Health (NIH-NINDS) P01NS097197, which established the Lafora Epilepsy Cure Initiative (LECI), to PS. We also acknowledge a grant from the Spanish Ministry of Economy and Competitiveness SAF2014-54604-C3-2-R to EK and a grant from Generalitat Valenciana Prometeo2018/135 to PS and FVP.Peer reviewe

Selective PARP-2 targeted therapy as anew strategy in pancreatic cancer

Trabajo presentado en el Defence is the Best Attack: Immuno-Oncology Breakthroughs. EACR Conference, celebrada en Barcelona (España), del 9 al 11 de mayo de 2023Pancreatic ductal adenocarcinoma (PDA) has been predicted to soon becomethe second leading cause of cancer related deaths. Although poly (ADPribose)polymerase (PARP) inhibitors (PARPi) have emerged as promising anticancerdrugs for BRCA mutated tumors, in pancreatic cancer, phase III clinical resultshave failed. Major limitations of PARPi are due to their lack of selectivity. In thisregard, we have recently demonstrated that Parp-2, but not Parp-1, plays aspecific role in replicative stress, which is a known PDA hallmark. Therefore, wehypothesized that Parp-2 may represent a new potential target to fight againstPDA. Importantly, Parp-2 genetic deletion in Ela-myc transgenic mice resulted in a43% increase in animal survival. Tumor histopathological characterizationshowed that Parp-2 inhibition hampers acinarto-ductal metaplasia, a key eventin pancreatic cancer initiation. Parp-2 loss in vitro significantly decreasedtumoroid generation capacity and induced DNA damage accumulation,increasing replicative stress and apoptosis. Molecular analysis of Ela-myc:Parp-2-/- vs Ela-myc Parp-2+/+ pancreatic tumors in preneoplastic lesions, indicatedthat Parp-2 inhibition resulted in a less immune-evasive environment. Moreover,GSEA at late tumor stages shows p53 pathway enrichment in Ela-myc Parp-2+/+vs Ela-myc:Parp-2-/-. Indeed, p53 inactivation was found in primary cell linesestablished from late stage Ela-myc:Parp-2-/- tumors, suggesting that loss offunction of this pathway is required for tumor progression in the absence ofParp-2. Altogether our data highlight that Parp-2 is a novel target in pancreaticcancer and open new avenues for therapeutic intervention against thisaggressive tumor by using specific Parp-2 pharmacological inhibitors

Age-Related microRNA Overexpression in Lafora Disease Male Mice Provides Links between Neuroinflammation and Oxidative Stress

Lafora disease is a rare, fatal form of progressive myoclonus epilepsy characterized by continuous neurodegeneration with epileptic seizures, characterized by the intracellular accumulation of aberrant polyglucosan granules called Lafora bodies. Several works have provided numerous evidence of molecular and cellular alterations in neural tissue from experimental mouse models deficient in either laforin or malin, two proteins related to the disease. Oxidative stress, alterations in proteostasis, and deregulation of inflammatory signals are some of the molecular alterations underlying this condition in both KO animal models. Lafora bodies appear early in the animal’s life, but many of the aforementioned molecular aberrant processes and the consequent neurological symptoms ensue only as animals age. Here, using small RNA-seq and quantitative PCR on brain extracts from laforin and malin KO male mice of different ages, we show that two different microRNA species, miR-155 and miR-146a, are overexpressed in an age-dependent manner. We also observed altered expression of putative target genes for each of the microRNAs studied in brain extracts. These results open the path for a detailed dissection of the molecular consequences of laforin and malin deficiency in brain tissue, as well as the potential role of miR-155 and miR-146a as specific biomarkers of disease progression in LD