Rett syndrome (RTT) is a rare devastating neurodevelopmental disorder that with an incidence of ~ 1:10,000 represents the most common genetic cause of severe intellectual disability in girls. Mutations in the X-linked methyl-CpG-binding protein 2 (MECP2) gene have been reported in over 95% cases of classical forms of RTT. Although initial studies supported a role for MeCP2 exclusively in neurons, recent data indicate a function also in astrocytes, which emerged as critical players involved in RTT pathogenesis through non-cell autonomous effects. Nevertheless, many aspects of RTT astrocyte dysfunctions remain unknown. With this PhD project, we wanted to use different in vivo and in vitro approaches that could help fill this gap of knowledge, in particular as regards to the effects that RTT astrocytes exert on neuronal health, and the involved molecular mechanisms. We herein demonstrate that astrocytes in the Mecp2 null mouse brain show progressive and area-specific alterations in morphology, with cortical areas exhibiting the most affected phenotype compared to the hippocampal cells; no phenotype, instead, is observed in cerebellar astrocytes, at least at early time points. These data, that have been reinforced by in vitro studies, represent the first evidence in the RTT field of the importance of considering astrocyte regionality. Considering the link between astrocyte morphogenesis and synaptogenesis, we have further investigated the effects of KO astrocytes on synaptic phenotypes. Our studies highlight a clear defect in synaptic maturation particularly in the Mecp2 null motor cortex. However, our in vivo exploration of synaptic terminals have not permit to find a stringent correlation between astrocyte morphology and synaptic defects probably due to the use of a globally null mouse model. Conversely, by using in vitro co-cultures, we have demonstrated for the first time that the lack of Mecp2 in astrocytes dramatically influences the synaptogenesis of WT neurons, by affecting both pre- and post-synaptic terminals by the release of one or more thermo-labile neurotoxic factors. To identify the involved molecular mechanisms, we have used RNAseq profiling which molecular pathways are activated in wild-type neurons by the paracrine effects triggered by KO astrocytes. Bioinformatic analyses have confirmed that KO astrocytes influence neuronal pathways mainly associated with the pre- and post-synaptic compartments, the mitochondrial functionality and inflammatory responses, therefore suggesting the possibility of an aberrant release of pro-inflammatory molecules. Collectively, the data presented in this thesis advance our understanding on the pathophysiology of RTT and point astrocytes and their paracrine factors as active players for the synaptic defects observed in RTT. We believe that the identification of the involved factors might reveal novel therapeutic approaches for the treatment of RTT.

SEARCHING FOR NOVEL MOLECULAR TARGETS IN ASTROCYTES FOR THE TREATMENT OF RETT SYNDROME / E. Albizzati ; tutor: N. Landsberger ; supervisor: A. Frasca ; coordinatore: M. Locati. - : . Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, 2021 Jun 11. ((33. ciclo, Anno Accademico 2020.

SEARCHING FOR NOVEL MOLECULAR TARGETS IN ASTROCYTES FOR THE TREATMENT OF RETT SYNDROME

E. Albizzati
2021-06-11

Abstract

Rett syndrome (RTT) is a rare devastating neurodevelopmental disorder that with an incidence of ~ 1:10,000 represents the most common genetic cause of severe intellectual disability in girls. Mutations in the X-linked methyl-CpG-binding protein 2 (MECP2) gene have been reported in over 95% cases of classical forms of RTT. Although initial studies supported a role for MeCP2 exclusively in neurons, recent data indicate a function also in astrocytes, which emerged as critical players involved in RTT pathogenesis through non-cell autonomous effects. Nevertheless, many aspects of RTT astrocyte dysfunctions remain unknown. With this PhD project, we wanted to use different in vivo and in vitro approaches that could help fill this gap of knowledge, in particular as regards to the effects that RTT astrocytes exert on neuronal health, and the involved molecular mechanisms. We herein demonstrate that astrocytes in the Mecp2 null mouse brain show progressive and area-specific alterations in morphology, with cortical areas exhibiting the most affected phenotype compared to the hippocampal cells; no phenotype, instead, is observed in cerebellar astrocytes, at least at early time points. These data, that have been reinforced by in vitro studies, represent the first evidence in the RTT field of the importance of considering astrocyte regionality. Considering the link between astrocyte morphogenesis and synaptogenesis, we have further investigated the effects of KO astrocytes on synaptic phenotypes. Our studies highlight a clear defect in synaptic maturation particularly in the Mecp2 null motor cortex. However, our in vivo exploration of synaptic terminals have not permit to find a stringent correlation between astrocyte morphology and synaptic defects probably due to the use of a globally null mouse model. Conversely, by using in vitro co-cultures, we have demonstrated for the first time that the lack of Mecp2 in astrocytes dramatically influences the synaptogenesis of WT neurons, by affecting both pre- and post-synaptic terminals by the release of one or more thermo-labile neurotoxic factors. To identify the involved molecular mechanisms, we have used RNAseq profiling which molecular pathways are activated in wild-type neurons by the paracrine effects triggered by KO astrocytes. Bioinformatic analyses have confirmed that KO astrocytes influence neuronal pathways mainly associated with the pre- and post-synaptic compartments, the mitochondrial functionality and inflammatory responses, therefore suggesting the possibility of an aberrant release of pro-inflammatory molecules. Collectively, the data presented in this thesis advance our understanding on the pathophysiology of RTT and point astrocytes and their paracrine factors as active players for the synaptic defects observed in RTT. We believe that the identification of the involved factors might reveal novel therapeutic approaches for the treatment of RTT.
LANDSBERGER, NICOLETTA
FRASCA, ANGELISA
LOCATI, MASSIMO
Settore BIO/11 - Biologia Molecolare
SEARCHING FOR NOVEL MOLECULAR TARGETS IN ASTROCYTES FOR THE TREATMENT OF RETT SYNDROME / E. Albizzati ; tutor: N. Landsberger ; supervisor: A. Frasca ; coordinatore: M. Locati. - : . Dipartimento di Biotecnologie Mediche e Medicina Traslazionale, 2021 Jun 11. ((33. ciclo, Anno Accademico 2020.
Doctoral Thesis
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Descrizione: Tesi di dottorato completa di Elena Albizzati
Tipologia: Tesi di dottorato completa
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2434/846870
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