Abnormal neuronal and synaptic morphology in Down syndrome brains reproduces in 2D and 3D human cellular models

Down syndrome (DS) is the most common autosomal aneuploidy compatible with postnatal survival and is caused by full or partial trisomy of chromosome 21. In this study, human isogenic induced pluripotent stem cells (iPSCs) were differentiated into 2D neurons and cortico-striatal assembloids (hCSAs). Aberrant neuronal morphology was observed: trisomic (T21) neurons formed aggregates of cell bodies interconnected with thick neurite bundles radiating outwards, connected and overlapped with other neurons in between the bundles while disomic (D21) neurons were evenly distributed across the surface and formed strong neuronal networks. Detailed analysis revealed significantly shorter neurites with larger diameter, fewer branches, and fewer terminal points in T21 neurons compared to D21 neurons in both 2D cultures and hCSAs. We observed similar phenotypes in foetal and postnatal human brain tissue. Furthermore, we observed aberrant mitochondrial morphology with an excess of some mitochondrial proteins (AIF, TOMM20) and significantly lower expression of synaptic markers (SYN-1, PSD95 and GEPH) throughout in vitro differentiation of T21 neurons. Finally, our data showed that T21 spheroids were significantly smaller throughout differentiation compared to D21 spheroids. T21 spheroids also exhibited significantly higher expression of neural stem cells (SOX2), significantly lower expression of proliferating cells (Ki67) and significantly higher expression of apoptotic cells (CCaspase-3). Overall, our study demonstrates that trisomy 21 leads to aberrant neuronal morphology in both 2D neurons and hCSAs, consistent with observations in the human brain.