The intellectual disability found in people with Down syndrome is associated with numerous changes in early brain development, including the proliferation and differentiation of neural progenitor cells (NPCs) and the formation and maintenance of myelin in the brain. To study how early neural precursors are affected by trisomy 21, we differentiated two isogenic lines of induced pluripotent stem cells derived from people with Down syndrome into brain-like and spinal cord-like NPCs and promoted a transition towards oligodendroglial fate by activating the Sonic hedgehog (SHH) pathway. In the spinal cord-like trisomic cells, we found no difference in expression of OLIG2 or NKX2.2, two transcription factors essential for commitment to the oligodendrocyte lineage. However, in the brain-like trisomic NPCs, OLIG2 is significantly upregulated and is associated with reduced expression of NKX2.2. We found that this gene dysregulation and block in NPC transition can be normalized by increasing the concentration of a SHH pathway agonist (SAG) during differentiation. These results underscore the importance of regional and cell type differences in gene expression in Down syndrome and demonstrate that modulation of SHH signaling in trisomic cells can rescue an early perturbed step in neural lineage specification.

The intellectual disability (ID) in Down syndrome (DS) is thought to result from a variety of developmental deficits such as alterations in neural progenitor division, neurogenesis, gliogenesis, cortical architecture, and reduced cortical volume. However, the molecular processes underlying these neurodevelopmental changes are still elusive, preventing an understanding of the mechanistic basis of ID in DS. In this study, we used a pair of isogenic (trisomic and euploid) induced pluripotent stem cell (iPSC) lines to generate cortical spheroids (CS) that model the impact of trisomy 21 on brain development. Cortical spheroids contain neurons, astrocytes, and oligodendrocytes and they are widely used to approximate early neurodevelopment. Using single cell RNA sequencing (scRNA-seq), we uncovered cell type-specific transcriptomic changes in the trisomic CS. In particular, we found that excitatory neuron populations were most affected and that a specific population of cells with a transcriptomic profile resembling layer IV cortical neurons displayed the most profound divergence in developmental trajectory between trisomic and euploid genotypes. We also identified candidate genes potentially driving the developmental asynchrony between trisomic and euploid excitatory neurons. Direct comparison between the current isogenic CS scRNA-seq data and previously published datasets revealed several recurring differentially expressed genes between DS and control samples. Altogether, our study highlights the power and importance of cell type-specific analyses within a defined genetic background, coupled with broader examination of mixed samples, to comprehensively evaluate cellular phenotypes in the context of DS.

SummaryThe intellectual disability found in people with Down syndrome (DS) is associated with a decrease in white matter in the central nervous system. To study the mechanism of this myelination deficit, we differentiated two isogenic lines of induced pluripotent stem cells (iPSCs) derived from people with DS into brain-like and spinal cord-like neural progenitor cells (NPCs) and promoted a transition towards oligodendroglial fate by activating the Sonic hedgehog (SHH) pathway. In the spinal cord-like trisomic cells, we found no difference in expression of OLIG2 or NKX2.2, two transcription factors essential for commitment to the oligodendrocyte (OL) lineage. However, in the brain-like trisomic NPCs, OLIG2 is significantly upregulated and is associated with reduced expression of NKX2.2. We found that this gene dysregulation and block in NPC transition can be normalized by increasing the concentration of a SHH pathway agonist (SAG) during differentiation. These results underscore the importance of regional and cell type differences in gene expression in DS and demonstrate that modulation of SHH signaling in trisomic cells can rescue an early perturbed step in neural lineage specification in DS.

The intellectual disability (ID) in Down syndrome (DS) is thought to result from a variety of developmental deficits such as alterations in neural progenitor division, neurogenesis, gliogenesis, cortical architecture, and reduced cortical volume. However, the molecular processes underlying these neurodevelopmental changes are still elusive, preventing an understanding of the mechanistic basis of ID in DS. In this study, we used a pair of isogenic (trisomic and euploid) induced pluripotent stem cell (iPSC) lines to generate cortical spheroids (CS) that model the impact of trisomy 21 on brain development. CS contain neurons, astrocytes, and oligodendrocytes and they are widely used to approximate early neurodevelopment. Using single cell RNA sequencing (scRNA-seq), we uncovered cell type-specific transcriptomic changes in the trisomic CS. In particular, we found that excitatory neuron populations were most affected and that a specific population of cells with a transcriptomic profile resembling layer IV cortical neurons displayed the most profound divergence in developmental trajectory between trisomic and euploid genotypes. We also identified candidate genes potentially driving the developmental asynchrony between trisomic and euploid excitatory neurons. Direct comparison between the current isogenic CS scRNA-seq data and previously published datasets revealed several recurring differentially expressed genes between DS and control samples. Altogether, our study highlights the power and importance of cell type-specific analyses within a defined genetic background, coupled with broader examination of mixed samples, to comprehensively evaluate cellular phenotypes in the context of DS.