Revisiting NeuroD1-mediated Glia-to-Neuron Trans-differentiation Using Transgenic Mice

Direct astrocyte-to-neuron conversion holds promise for in situ neuroregeneration to treat neurodegenerative diseases. However, commonly-used adeno-associated vectors carrying GFAP mini-promoters drive leaky expression of neurogenic factors in endogenous neurons, raising concerns about trans-differentiation validity. Developing inducible, lineage-traceable transgenic mice, we evaluate NeuroD1 effects on astrocyte-to-neuron conversion in spinal cord and brain injury models. Lineage-tracing and single-cell transcriptomics reveal that ectopic NeuroD1 expression promotes robust conversion of reactive astrocytes in lesion cores to neurons at the acute phase of injury, but not in peri-injury astrocytes. We performed spinal cord injury in iNeuroD1GFP adult mice at postnatal day 60 (P60) and administered TAM (100 mg/kg/day)44 from days post-injury (dpi) 5 to 8. At dpi10,spinal cords were rapidly extruded from iNeuroD1GFP mice (n=3) after anesthesia. Each cord was separated into lesion core and distal regions, and analyzed using snRNA-seq