Abstract
Nucleus pulposus cells (NPCs) play a vital role in maintaining the homeostasis of the intervertebral disc (IVD). Previous studies have discovered that NPCs exhibited malfunction due to cellular senescence during disc aging and degeneration; this might be one of the key factors of IVD degeneration. Thus, we conducted this study in order to investigate the altered biofunction and the underlying genes and pathways of senescent NPCs. We isolated and identified NPCs from the tail discs of young (2 months) and old (24 months) SD rats and confirmed the senescent phenotype through SA-β-gal staining. CCK-8 assay, transwell assay, and cell scratch assay were adopted to detect the proliferous and migratory ability of two groups. Then, a rat Gene Chip Clariom™ S array was used to detect differentially expressed genes (DEGs). After rigorous bioinformatics analysis of the raw data, totally, 1038 differentially expressed genes with a fold change > 1.5 were identified out of 23189 probes. Among them, 617 were upregulated and 421 were downregulated. Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis were conducted and revealed numerous number of enriched GO terms and signaling pathways associated with senescence of NPCs. A protein-protein interaction (PPI) network of the DEGs was constructed using the Search Tool for the Retrieval of Interacting Genes (STRING) database and Cytoscape software. Module analysis was conducted for the PPI network using the MCODE plugin in Cytoscape. Hub genes were identified by the CytoHubba plugin in Cytoscape. Derived 5 hub genes and most significantly up- or downregulated genes were further verified by real-time PCR. The present study investigated underlying mechanisms in the senescence of NPCs on a genome-wide scale. The illumination of molecular mechanisms of NPCs senescence may assist the development of novel biological methods to treat degenerative disc diseases.
Highlights
Low back pain (LBP) is a major age-related disease, contributing to patients’ suffering and disability and causing large financial burden to society [1]
Nucleus pulposus cells (NPCs) are responsible for the metabolism homeostasis of the extracellular matrix (ECM) by producing collagen I, collagen II, and proteoglycan, which are the main components of the gelatinous structures of nucleus pulposus (NP) [9]
NPCs were successfully isolated from rat nucleus pulposus tissues
Summary
Low back pain (LBP) is a major age-related disease, contributing to patients’ suffering and disability and causing large financial burden to society [1]. NP is a kind of gelatinous tissue containing extracellular matrix (ECM) comprising highly hydrated proteoglycan, collagen fibers, and aggrecan [6]. It plays a vital role in maintaining the physiological function of IVD because NP could absorb stress when the IVD is confronting diverse mechanical impact [7]. During aging and degeneration of IVD, the normal function of NPCs was disrupted, resulting in aberrant metabolism of ECM, which could accelerate the process of IVDD [10, 11]. Cytotherapy by reactivate degenerated NPCs has been proposed to be an ideal biological therapy method to treat IVDD [11]. The specific mechanism of NPC degeneration is still unknown, which hindered the development of cytotherapy
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