The Cellular Composition of Bovine Coccygeal Intervertebral Discs: A Comprehensive Single-Cell RNAseq Analysis

Intervertebral disc (IVD) degeneration and its medical consequences is still one of the leading causes of morbidity worldwide. To support potential regenerative treatments for degener-ated IVDs, we sought to deconvolute the cell composition of the nucleus pulposus (NP) and the annulus fibrosus (AF) of bovine intervertebral discs. Bovine calf tails have been extensively used in intervertebral disc research as a readily available source of NP and AF material from healthy and young IVDs. We used single-cell RNA sequencing (scRNAseq) coupled to bulk RNA sequencing (RNAseq) to unravel the cell populations in these two structures and analyze developmental changes across the rostrocaudal axis. By integrating the scRNAseq data with the bulk RNAseq data to stabilize the clustering results of our study, we identified 27 NP structure/tissue specific genes and 24 AF structure/tissue specific genes. From our scRNAseq results, we could deconvolute the heterogeneous cell populations in both the NP and the AF. In the NP, we detected a notochordal-like cell cluster and a progenitor stem cell cluster. In the AF, we detected a stem cell-like cluster, a cluster with a predomi-nantly fibroblast-like phenotype and a potential endothelial progenitor cluster. Taken together, our results illustrate the cell phenotypic complexity of the AF and NP in the young bovine IVDs.