Biology subjects

Rollo, B.

Publications and source records attributed to Rollo, B..

5 recordsLinked to original sources

bioRxiv2024

REST and RCOR genes display distinct expression profiles in neurons and astrocytes using 2D and 3D human pluripotent stem cell models

Repressor element-1 silencing transcription factor (REST) is a transcriptional repressor involved in neurodevelopment and neuroprotection. REST forms a complex with the REST corepressors, CoREST1, CoREST2, or CoREST3 (encoded by RCOR1, RCOR2, and RCOR3, respectively). Emerging evidence suggests that the CoREST family can target unique genes independently of REST, in various neural and glial cell types during different developmental stages. However, there is limited knowledge regarding the expression and function of the CoREST family in human neurodevelopment. To address this gap, we employed 2D and 3D human pluripotent stem cell (hPSC) models to investigate REST and RCOR gene expression levels. Our study revealed a significant increase in RCOR3 expression in glutamatergic cortical and GABAergic ventral forebrain neurons, as well as mature functional NGN2-induced neurons. Additionally, a simplified astrocyte transdifferentiation protocol resulted in a significant decrease in RCOR2 expression following differentiation. REST expression was notably reduced in mature neurons and cerebral organoids, along with RCOR2 in the latter. In summary, our findings provide the first insights into the cell-type-specific expression patterns of RCOR genes in human neuronal and glial differentiation. Specifically, RCOR3 expression increases in neurons, while RCOR2 levels decrease in astrocytes. The dynamic expression patterns of REST and RCOR genes during hPSC neuronal and glial differentiation underscore the potential distinct roles played by REST and CoREST proteins in regulating the development of these cell types in humans. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=138 SRC="FIGDIR/small/584254v1_ufig1.gif" ALT="Figure 1"> View larger version (19K): org.highwire.dtl.DTLVardef@7be30borg.highwire.dtl.DTLVardef@1771388org.highwire.dtl.DTLVardef@a376aforg.highwire.dtl.DTLVardef@1c4dc83_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIREST and RCOR genes display cell-type specific expression patterns in neural cells C_LIO_LIRCOR3 (encodes CoREST3) is upregulated during neuronal and astrocyte differentiation C_LIO_LIRCOR2 (encodes CoREST2) is downregulated during differentiation of astrocytes C_LIO_LIEvidence of potential cell-type specific functions of the CoREST family C_LI

neuroscience↗

bioRxiv2023

Distinctive in vitro phenotypes in iPSC-derived neurons from patients with gain- and loss-of-function SCN2A developmental and epileptic encephalopathy

SCN2A encodes NaV1.2, an excitatory neuron voltage-gated sodium channel and major monogenic cause of neurodevelopmental disorders, including developmental and epileptic encephalopathies (DEE) and autism. Clinical presentation and pharmocosensitivity vary with nature of SCN2A variant dysfunction with gain-of-function (GoF) cases presenting with pre- or peri-natal seizures and loss-of-function (LoF) patients typically having infantile spasms after 6 months of age. Here, we established and assessed patient induced pluripotent stem cell (iPSC) - derived neuronal models for two recurrent SCN2A DEE variants with GoF R1882Q and LoF R853Q associated with early- and late-onset DEE, respectively. Patient-derived iPSC lines were differentiated using a Neurogenin-2 overexpression yielding populations of cortical-like glutamatergic neurons. Electrophysiological and transcriptomic profiles were assessed after 2-4 weeks in culture. Increased neuronal activity at both cellular and network level was observed for R1882Q iPSC-derived neurons at three weeks of differentiation. In contrast, R853Q neurons showed only subtle changes in excitability after four weeks in vitro. In alignment with the reported efficacy in some GoF SCN2A patients, phenytoin (sodium channel blocker) reduced excitability of neurons to the control levels in R1882Q neuronal cultures. Transcriptomic alterations in neurons were detected for each variant and convergent pathways pointed at the shared mechanisms underlying SCN2A DEE.

neuroscience↗

bioRxiv2021

In vitro neurons learn and exhibit sentience when embodied in a simulated game-world

Integrating neurons into digital systems to leverage their innate intelligence may enable performance infeasible with silicon alone, along with providing insight into the cellular origin of intelligence. We developed DishBrain, a system which exhibits natural intelligence by harnessing the inherent adaptive computation of neurons in a structured environment. In vitro neural networks from human or rodent origins, are integrated with in silico computing via high-density multielectrode array. Through electrophysiological stimulation and recording, cultures were embedded in a simulated game-world, mimicking the arcade game Pong. Applying a previously untestable theory of active inference via the Free Energy Principle, we found that learning was apparent within five minutes of real-time gameplay, not observed in control conditions. Further experiments demonstrate the importance of closed-loop structured feedback in eliciting learning over time. Cultures display the ability to self-organise in a goal-directed manner in response to sparse sensory information about the consequences of their actions.

synthetic biology↗

bioRxiv2020

Antisense oligonucleotide therapy for KCNT1 encephalopathy

Developmental and epileptic encephalopathies (DEE) are characterized by pharmacoresistant seizures with concomitant intellectual disability. Epilepsy of infancy with migrating focal seizures (EIMFS) is one of the most severe of these syndromes. De novo mutations in ion channels, including gain-of-function variants in KCNT1, have been found to play a major role in the etiology of EIMFS. Here, we test a potential precision therapeutic approach in KCNT1-associated DEE using a gene silencing antisense oligonucleotide (ASO) approach. The homozygous p.P924L (L/L) mouse model recapitulates the frequent, debilitating seizures and developmental compromise that are seen in patients. After a single intracerebroventricular bolus injection of a Kcnt1 gapmer ASO in symptomatic mice at postnatal day 40, seizure frequency was significantly reduced, behavioral abnormalities improved, and overall survival was extended compared to mice treated with a control ASO (non-hybridizing sequence). ASO administration at neonatal age was also well-tolerated and effective in controlling seizures and extending the lifespan of treated animals. The data presented here provides a proof of concept for ASO-based gene silencing as a promising therapeutic approach in KCNT1-associated epilepsies.

neuroscience↗

bioRxiv2020

Antisense oligonucleotide therapy for SCN2A gain-of-function epilepsy

The clinical spectrum associated with SCN2A de novo mutations (DNMs) continues to expand and includes autism spectrum disorder with or without seizures, in addition to early and late seizure onset developmental and epileptic encephalopathies (DEEs). Recent biophysical studies on SCN2A variants suggest that the majority of early seizure onset DEE DNMs cause gain of function. Gain of function in SCN2A, the principal sodium channel of excitatory pyramidal neurons, would result in heightened neuronal activity and is likely to underlie the pathology seen in early seizure onset DEE patients. Supratherapeutic dosing of the non-selective sodium channel blocker phenytoin, is effective in controlling seizures in these patients but does not impact neurodevelopment, raising the idea that more profound and specific reduction in SCN2A function could significantly improve clinical outcome. To test the potential therapeutic benefit of reducing SCN2A in early seizure onset DEE we centrally administered an antisense oligonucleotide (ASO) targeting mouse Scn2a (Scn2a ASO) to a mouse model of human SCN2A early seizure onset DEE. Mice were genetically engineered to harbour the human equivalent SCN2A p.R1882Q mutation (Q/+), one of the most recurrent mutations in early seizure onset DEE. Q/+ mice presented with spontaneous seizures at postnatal day (P) 1 and did not survive beyond P30. Intracerebroventricular Scn2a ASO administration into Q/+ mice between P1-2 (that reduced Scn2a mRNA levels by 50%) significantly extended lifespan and markedly reduced spontaneous seizures occurrence. Across a range of cognitive and motor behavioural tests, Scn2a ASO treated Q/+ mice were largely indistinguishable from wildtype (+/+) mice. Further improvements in survival and behaviour were seen by adjustment of dosing regimens during development. Scn2a ASO efficacy was also evident at the cellular level. Whole cell patch clamp recording showed that Scn2a ASO administration reversed changes in neuronal excitability in layer 2/3 pyramidal neurons of Q/+ mice to levels seen in +/+ mice. Safety was assessed in +/+ mice and showed a developmental stage dependent tolerability and a favourable therapeutic index. This study suggests that a human SCN2A gapmer ASO could profoundly and safely impact early seizure onset DEE patients and heralds a new era of precision therapy in neurodevelopmental disorders.

neuroscience↗