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Spatiotemporal characterization of human early intervertebral disc formation at single-cell resolution

ABSTRACT: Intervertebral disc (IVD) performs as a fibrocartilaginous joint to anchor adjacent vertebrae (VB) and is essential for the mechanical stabilization of axial skeleton. Although several studies have demonstrated the cellular heterogeneity of adult mature IVD, a single-cell transcriptomic atlas mapping early IVD formation is still lacking. Here, we generated the first spatiotemporal and single-cell based transcriptomic atlas of human IVD formation at embryonic stage, as well as a comparative mouse transcript landscape. The notochord (NC)/nucleus pulposus (NP) cells, vertebral chondrocytes, and annulus fibrosus (AF) cells were characterized. Our single-cell transcriptomic atlas that resolves spatiotemporally regulated cellular heterogeneity underlying early IVD development will be leveraged to develop preventative and regenerative strategies for IVD degeneration.

ORGANISM(S): Mus musculus

PROVIDER: GSE216511 | GEO | 2025/10/25

REPOSITORIES: GEO

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Project description:Very little is known about how intervertebral disc (IVD) is formed or maintained. Members of the TGF-ß superfamily are secreted signaling proteins that regulate many aspects of development including cellular differentiation. We recently showed that deletion of Tgfbr2 in Col2a expressing tissue results in alterations in development of IVD annulus fibrosus. The results suggested TGF-ß has an important role in regulating development of the axial skeleton, however, the mechanistic basis of TGF-ß action in these specialized joints is not known. One of the hurdles to understanding development of IVD is a lack of known markers. To identify genes that are enriched in the developing IVD and to begin to understand the mechanism of TGF-ß action in IVD development, we undertook a global analysis of gene expression comparing gene expression profiles in developing vertebrae and IVD. We also compared expression profiles in tissues from wild type and Tgfbr2 mutant mice. Lists of IVD and vertebrae enriched genes were generated. Expression patterns for several genes were verified either through in situ hybridization or literature/ database searches resulting in a list of genes that can be used as markers of IVD. Cluster analysis using genes listed under the Gene Ontology terms multicellular organism development and pattern specification indicated that mutant IVD more closely resembled vertebrae than wild type IVD. We propose TGF-ß has two functions in IVD development: 1) to prevent chondrocyte differentiation in the presumptive IVD and 2) to promote differentiation of annulus fibrosus from sclerotome. We have identified genes that are enriched in the IVD and regulated by TGF-ß that warrant further investigation as regulators of IVD development. Thirteen samples were analyzed. This includes three biological replicates of laser captured IVD from E13.5 day control mice, three biological replicates of laser captured vertebrae from the same E13.5 day control mice, three biological relicates of laser captured vertebrae from E13.5 day Col2aCre;Tgfbr2lox/lox mice, and four biological replicates of laser captured IVD from E13.5 day Col2aCre;Tgfbr2lox/lox mice.

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Spatiotemporal characterization of human early intervertebral disc formation at single-cell resolution

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