Abstract
BackgroundAutism spectrum disorder (ASD) is a neurodevelopmental disorder with a high co-morbidity of epilepsy and associated with hundreds of rare risk factors. NRXN1 deletion is among the commonest rare genetic factors shared by ASD, schizophrenia, intellectual disability, epilepsy, and developmental delay. However, how NRXN1 deletions lead to different clinical symptoms is unknown. Patient-derived cells are essential to investigate the functional consequences of NRXN1 lesions to human neurons in different diseases.MethodsSkin biopsies were donated by five healthy donors and three ASD patients carrying NRXN1α+/− deletions. Seven control and six NRXN1α+/− iPSC lines were derived and differentiated into day 100 cortical excitatory neurons using dual SMAD inhibition. Calcium (Ca2+) imaging was performed using Fluo4-AM, and the properties of Ca2+ transients were compared between two groups of neurons. Transcriptome analysis was carried out to undercover molecular pathways associated with NRXN1α+/− neurons.ResultsNRXN1α+/− neurons were found to display altered calcium dynamics, with significantly increased frequency, duration, and amplitude of Ca2+ transients. Whole genome RNA sequencing also revealed altered ion transport and transporter activity, with upregulated voltage-gated calcium channels as one of the most significant pathways in NRXN1α+/− neurons identified by STRING and GSEA analyses.ConclusionsThis is the first report to show that human NRXN1α+/− neurons derived from ASD patients’ iPSCs present novel phenotypes of upregulated VGCCs and increased Ca2+ transients, which may facilitate the development of drug screening assays for the treatment of ASD.
Highlights
Autism spectrum disorder (ASD) is a neurodevelopmental disorder with a high co-morbidity of epilepsy and associated with hundreds of rare risk factors
Overproduction of GABAergic neurons with FOXG1 overexpression and accelerated cell cycle were reported in induced pluripotent stem cells of sporadic ASD with macrocephaly [6]
The mutations were validated by SNP array (Additional file 1: Figure S1A). Induced pluripotent stem cell (iPSC) were derived from dermal fibroblasts (Fig. 1a–c) and characterized for pluripotency by expression of alkaline phosphatase (Fig. 1d), NANOG, OCT4, SOX2, SSEA4, and TRA-1-60 (Additional file 1: Figure S2) and tri-germ layer potential by TUJ1, ASM, and AFP (Fig. 1d–j, n, o) and cell cycle markers Ki67 and PH3 (Fig. 1k–m)
Summary
Autism spectrum disorder (ASD) is a neurodevelopmental disorder with a high co-morbidity of epilepsy and associated with hundreds of rare risk factors. NRXN1 deletion is among the commonest rare genetic factors shared by ASD, schizophrenia, intellectual disability, epilepsy, and developmental delay. Conclusions: This is the first report to show that human NRXN1α+/− neurons derived from ASD patients’ iPSCs present novel phenotypes of upregulated VGCCs and increased Ca2+ transients, which may facilitate the development of drug screening assays for the treatment of ASD. An increased ratio of synaptic excitation/inhibition (E/I) affecting neuroplasticity has been proposed as a common pathway for ASD [2]. This has been linked to altered functional and structural connectivity. Overproduction of GABAergic neurons with FOXG1 overexpression and accelerated cell cycle were reported in induced pluripotent stem cells (iPSCs) of sporadic ASD with macrocephaly [6]
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