Progress Toward a Comprehensive Cellular Phenotype of CHD2 Epileptic Encephalopathy
Abstract number :
3.027
Submission category :
1. Basic Mechanisms / 1B. Epileptogenesis of genetic epilepsies
Year :
2024
Submission ID :
294
Source :
www.aesnet.org
Presentation date :
12/9/2024 12:00:00 AM
Published date :
Authors :
Presenting Author: Sang Ho Yoon, PhD – University of California Irvine
Robert F Hunt, PhD – University of California Irvine
Rationale: Mutations in the chromatin regulator Chromodomain helicase DNA-binding 2 (CHD2) are increasingly found in neurodevelopmental and neuropsychiatric conditions, including epilepsy, intellectual disability and autism spectrum disorders. To accurately recapitulate the early developmental stages of synaptic and neuronal network dysfunction in these patients, direct access to patient neurons is indispensable. Here, we describe a stem cell-based platform for phenotyping CHD2+/- human hippocampal neurons across genomic, anatomical, electrophysiological and behavioral domains.
Methods: To identify optimal conditions for producing hippocampal neurons from human induced pluripotent stem cells (hiPSCs), we exposed developing neural progenitors to 13 different combinations morphogens, based on prior efforts to generate human-derived hippocampal neurons. We screened the cultures for markers of medial pallium progenitors over 15 weeks of neural development after neuron plating. We used CRISPR-Cas9 engineering to produce isogenic iPSC lines (iControl, CHD2+/- and CHD2-/-).
Results: Resulting neural progenitors expressed PAX6, SOX2, FOXG1, POU3F1, DCX and had a significant increase in CA1-specific transcripts, as compared to pan-neuronal neural progenitor cells. By 15 weeks in vitro or following transplantation into dorsal hippocampus of young-adult NOD SCID recipients, the resulting neurons expressed markers of CA1. Expression of GRIK4 (CA3), PCP4 (CA2), PROX1 (DG), GABA (interneurons) or NKX2-1 (medial ganglionic eminence) was very low or absent. Transplanted human neurons did not migrate far from the injection site, but they integrated into mouse hippocampus with anatomical features that are strikingly similar to native born mouse neurons. Detailed transcriptomic, anatomical and electrophysiological analysis of CHD2 mutations in mouse and human-derived neurons are currently underway.
Conclusions: Our work provides an innovative pre-clinical strategy for generating a highly specialized type of human neuron that can be used to identify disease mechanisms and test new therapies.
Funding: This work was supported by funding from NIH R01-NS126399.
Basic Mechanisms