Introduction: Low back pain is a major disabling health condition in humans, with a lifetime prevalence as high as 84%.  IVDD is a widely recognized contributor to low back pain. The major constituents of the normal NP are type-II collagen and aggrecan proteoglycan. The loss of these extracellular matrix proteins occurs in IVDD. The current treatments of low back pain are limited to relieving back or leg symptoms. They do not focus on replenishing the NP loss and restoring the native disk structure. NPCs are the main cell type residing in the NP, and they are responsible for maintaining tissue homeostasis (8). NPCs proliferate slowly and lack self-regeneration capacity, adding to the intractability of the disease. Current studies on the pathophysiology of NPCs are usually supported by mRNA and epigenomic analyses. However, bulk-tissue level resolution masks the complexity of alterations across cells and within cell types. The uncharacterized cell types and markers residing in the NP raise interest in terms of unexplored cellular heterogeneity.

Methods: We aimed to provide a single-cell view of IVDD pathology, profiling 39,732 cells from NP tissues across eight individuals with different grades of progressive degeneration. Notably, we comprehensively characterized the transcriptome feature of NPCs and immune cells, and we decoded the cell percentage, the heterogeneity of cell subtypes during degeneration, providing a unique cellular-level insight into transcriptional alterations associated with IVDD pathology.

Results: Six novel human NP cell (NPCs) populations were identified by their distinct molecular signatures. The potential functional differences among NPC  subpopulations were analyzed at the single-cell level. Predictivetranscripts, transcriptional factors, and signal pathways with respect to degeneration grades were analyzed. We reported that fibroNPCs, one of our identified cell types, is the subpopulation for end-stage degeneration. CD90+NPCs were observed to be progenitor cells in degenerative NP tissues. NP-infiltrating immune cells comprise a previously unrecognized diversity of cell types, including granulocytic myeloid-derived suppressor cells (G-MDSCs). We uncovered integrin αM (CD11b) and oxidized low density lipoprotein receptor 1 (OLR1) as surface markers of NP-derived G-MDSCs. The G MDSCs were also found to be enriched in mildly degenerated (grade II and III) NP tissues compared to severely degenerated (grade IV and V) NP tissues. Their immunosuppressive function and alleviation effects on NPCs’ matrix degradation were revealed in vitro.  

Discussion: These findings might aid the understanding of molecular complexity and cellular heterogeneity in intervertebral disc degeneration. The IVDs have been identified as immune privilege organs; the steady-state of immune privilege is fundamental to organ homeostasis. This single-cell study also showed there are multiple immune cell lines inside the NP, which may play a role in IVDD progression.