The detrimental effect of Mecp2 null astrocytes on synapses: exploring the molecular mechanisms to find novel druggable targets for Rett syndrome

Rett syndrome (RTT) is a rare devastating neurodevelopmental disorder that 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. Initial studies supported a role for MeCP2 exclusively in neurons, which are profoundly defective in RTT. Indeed, RTT is considered a synaptopathy, characterized by dendritic spine dysgenesis, impaired spine plasticity, and disrupted excitatory/inhibitory balance. However, recent data supported the involvement also of astrocytes, which can affect neuronal maturation through non-cell autonomous mechanisms. Nevertheless, many aspects of RTT astrocyte dysfunctions remain still unknown and astrocytes’ heterogeneity in RTT has been only marginally explored. According to the crucial role of astrocytes in promoting synapse formation and functioning, we have investigated the influence of Mecp2 null astrocytes on synaptic phenotype. By using in vitro co-cultures, we have demonstrated that the lack of Mecp2 in cortical astrocytes dramatically affected the number of synaptic puncta in wild-type (WT) neurons. To gain insights into the involved molecular mechanisms, we performed bulk RNA-sequencing on WT neurons co-cultured with KO astrocytes; by profiling the molecular pathways activated in WT neurons by the paracrine effects triggered by KO cortical astrocytes, we confirmed that KO astrocytes influenced neuronal pathways mainly associated with synaptic maturation and functions, and elicited inflammatory responses. qPCR and luminex assay indicated the up-regulated release of a subset of cytokines by Mecp2 null astrocytes, with a possible synaptotoxic effect. We thus validated the role of one of these secreted factors, being aware that their identification might reveal novel therapeutic approaches for the treatment of RTT.