Single cell analysis of the cellular heterogeneity and interactions in the injured mouse spinal cord

The wound healing process that occurs after spinal cord injury is critical for maintaining tissue homeostasis and limiting tissue damage, but eventually results in a scar-like environment that is not conducive to regeneration and repair. A better understanding of this dichotomy is critical to developing effective therapeutics that target the appropriate pathobiology, but a major challenge has been the large cellular heterogeneity that results in immensely complex cellular interactions. In this study, we used single cell RNA sequencing to assess virtually all cell types that comprise the mouse spinal cord injury site. In addition to discovering novel subpopulations, we used expression values of receptor-ligand pairs to identify signaling pathways that are predicted to regulate specific cellular interactions during angiogenesis, gliosis, and fibrosis. Our dataset is a valuable resource that provides novel mechanistic insight into the pathobiology of not only spinal cord injury, but also other traumatic disorders of the CNS. Overall design: Single cell RNA sequencing of uninjured and injured spinal cord at 1 dpi, 3 dpi, and 7 dpi of wild-type mice. In total, we sequenced three uninjured biological replicates (from five animals), three biological replicates at 1 dpi (from five animals), two biological replicates at 3 dpi (from three animals), and two biological replicates at 7dpi (from three animals). One biological replicate from each time point consisted of cells from a single animal's spinal cord. All other biological replicates consisted of cells pooled from two spinal cords. For these pooled samples, one of two spinal cords were processed to enrich for astrocytes using Miltenyi's Anti-ACSA-2 MicroBead kit. All samples were initially processed with Miltenyi Neural Tissue Dissociation Kit-P.