ZNF667 alleviates the inflammatory damage in intervertebral disc degeneration via inhibiting NF-κB signaling pathway.

Objectives: With the aging population, the incidence of intervertebral disc degeneration (IDD) is increasing every year. The pathogenesis of IDD is complex, and there are currently no effective treatment options. This study aims to investigate the specific function and underlying mechanism of zinc finger protein 667 (ZNF667) in the inflammatory damage of nucleus pulposus cells in IDD.

Methods: Differential expression genes (DEG) associated with IDD were screened from IDD-related datasets in the Gene Expression Omnibus (GEO) (GSE124272 and GSE150408), and ZNF667, which is closely related to gene transcriptional regulation, was selected and analyzed in several IDD-related datasets (GSE124272, GSE150408, GSE56081, GSE147383, GSE23130). Nucleus pulposus tissues were collected from 3 IDD patients and 3 trauma-induced vertebral fracture patients (serving as controls). Hematoxylin and eosin (HE) staining was performed for pathological examination, and immunohistochemistry (IHC) was used to assess ZNF667 expression in the nucleus pulposus tissues. Gene set enrichment analysis (GSEA) was then employed to elucidate the potential mechanisms of ZNF667. For in vitro validation, human primary nucleus pulposus cells were treated with 10 ng/mL of interleukin-1β (IL-1β) to establish an IDD cell model, and subsequently transfected with a ZNF667 overexpression plasmid. Flow cytometry was used to evaluate cell apoptosis, enzyme-linked immunosorbent assay (ELISA) measured the levels of inflammatory factors-cyclooxygenase-2 (COX-2), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in the cell culture supernatant, real-time polymerase chain reaction (RT-PCR) quantified mRNA expression, and Western blotting assessed protein expression levels of ZNF667, myeloid differentiation factor 88 (MyD88), P65, and phosphorylated P65 (p-P65).

Results: Analysis of both the GEO datasets and clinical tissue samples revealed that ZNF667 expression is reduced in IDD. In IDD patients, the extracellular matrix and nucleus pulposus cells are significantly diminished, and the arrangement of fibrochondrocytes is disordered. GSEA results showed that ZNF667 may be involved in biological processes such as angiogenesis, epithelial-mesenchymal transition (EMT), oxidative phosphorylation, peroxisome function, steroid biosynthesis, and the NF-κB-mediated TNF-α signaling pathway. In vitro, ZNF667 was expressed at low levels in the IL-1β-induced IDD cell model, and overexpression of ZNF667 reversed the IL-1β-induced increase in cell apoptosis, the upregulation of inflammation factors (COX-2, IL-6, TNF-α), and the increased expression of NF-κB pathway-related proteins (MyD88 and the p-P65/P65 ratio) (all <0.05).

Conclusions: ZNF667 can alleviate nucleus pulposus cell apoptosis and inflammatory responses by inhibiting the NF-κB signaling pathway, thereby exerting a protective effect on intervertebral discs. This finding not only provides new insights into the pathogenesis of IDD but also suggests a potential therapeutic target for its treatment.