Objective: Dystrophin protein—highly expressed in the central nervous system—is critical for synaptogenesis and axonal guidance. The absence of brain dystrophin is considered the molecular correlate of the cognitive deficits seen in boys with Duchenne Muscular Dystrophy, an X-linked genetic disease caused by the lack of functional dystrophin. Single nucleus RNA-sequencing (snRNA-seq) permits unbiased transcriptomic investigation of cells of interest. We investigated the transcriptome of hippocampi of the mdx52 model of DMD to better understand how dystrophin deficiency affects brain development in early postnatal period.
Methods: Hippocampi were isolated from 4 male pups each from mdx52 and BL6 mice on postnatal day zero and single nuclei were partitioned using the Chromium system from 10x Genomics. Following library preparation, samples was sequenced on a NovaSeq6000. An average of 25,000 single nuclei with a read-depth of 100,000 transcripts were performed from each hippocampus. Raw sequencing reads were processed with Cell Ranger and cell type clustering was performed using Seurat v5. Cell clusters were labelled using cell-type specific gene expression markers. Cell Chat analyses was performed to investigate transcriptome-based aberrations.
Results: A total of 18 cell clusters including neurons, interneurons, astrocytes, oligodendrocytes, choroid plexus, epithelial cells, and microglia were identified. There was no statistical difference in the cell-type composition between the mdx52 and BL6 mice groups. Differential expression analysis demonstrated transcriptomic changes in a subset of GABAergic interneurons in the mdx52 hippocampi. Cell Chat analysis demonstrated abnormalities in the extracellular matrix-related pathways.
Conclusions: Compelling transcriptomic changes in post-natal hippocampi in mouse model of dystrophinopathy includes differences in a subset of inhibitory interneurons as well as distinct differences in cell-cell interactions.
Our work was generously funded by the MDA Idea Development Award.