Single-cell RNA-seq identifies unique transcriptional landscapes of human nucleus pulposus and annulus fibrosus cells

Abstract

Intervertebral disc (IVD) disease (IDD) is a complex, multifactorial disease. While various aspects of IDD progression have been reported, the underlying molecular pathways and transcriptional networks that govern the maintenance of healthy nucleus pulposus (NP) and annulus fibrosus (AF) have not been fully elucidated. We defined the transcriptome map of healthy human IVD by performing single-cell RNA-sequencing (scRNA-seq) in primary AF and NP cells isolated from non-degenerated lumbar disc. Our systematic and comprehensive analyses revealed distinct genetic architecture of human NP and AF compartments and identified 2,196 differentially expressed genes. Gene enrichment analysis showed that SFRP1, BIRC5, CYTL1, ESM1 and CCNB2 genes were highly expressed in the AF cells; whereas, COL2A1, DSC3, COL9A3, COL11A1, and ANGPTL7 were mostly expressed in the NP cells. Further, functional annotation clustering analysis revealed the enrichment of receptor signaling pathways genes in AF cells, while NP cells showed high expression of genes related to the protein synthesis machinery. Subsequent interaction network analysis revealed a structured network of extracellular matrix genes in NP compartments. Our regulatory network analysis identified FOXM1 and KDM4E as signature transcription factor of AF and NP respectively, which might be involved in the regulation of core genes of AF and NP transcriptome.

Spatially defined single-cell transcriptional profiling characterizes diverse chondrocyte subtypes and nucleus pulposus progenitors in human intervertebral discs

“…2e), and highly expressed IBSP and CYTL1, which are NP-negative biomarkers (Supplementary Table 3). 17,52 Moreover, subclusters C1 and C2 showed a higher percentage of cells arrested in the G2/ M phase than that in others, which was indicative of relatively higher proliferative activity (Fig. 2f).…”

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

“…Taken together, our data indicate a series of very distinct cell populations that are closely orchestrated living in this unique environment. Our data, whether in the single-cell RNAseq or the bulk RNAseq, show that we could identify most NP specific markers and genes previously reported to be expressed at higher levels in the NP than in the AF [ 86 , 87 ].…”

Extracellular matrix gene expression signatures as cell type and cell state identifiers

“…Beyond development, planarian scRNA-seq revealed that muscles produce most of the matrisome, and inhibiting one key matrisome gene (hemicentin) resulted in severe epidermal ruffling and displacement of cells during homeostatic tissue turnover, suggesting an important role for tissue regeneration [28] . Furthermore, in healthy human lumbar discs, the core matrisome can be used to distinguish primary annulus fibrosus and nucleus pulposus cells based on 90 out of the 274 core matrisome genes being differentially expressed in the opposite direction using scRNA-seq [35] . Similarly, 115 matrisome genes are characteristically expressed in the six cell types that make up the human cutaneous neurofibroma microenvironment [36] .…”