Sustained stimulation with tumor necrosis factor-alpha regulates interleukin-8 production and gene transcription in...

Gingival epithelial cells (GECs), the first line of defense in the oral mucosa, are constantly exposed to microbial and inflammatory stimuli. Tumor necrosis factor-alpha (TNF-α) is a major cytokine involved in periodontal inflammation; however, the mechanisms by which GECs respond to sustained TNF-α exposure remain unclear. This study investigated the cellular and transcriptional responses of human GECs to sustained TNF-α stimulation and explored the associated mechanisms.

Human gingival epithelial Ca9-22 cells were stimulated with TNF-α (2 ng/mL) every 24 h for 4 days. Interleukin-8 (IL-8) secretion was measured using an enzyme-linked immunosorbent assay. Global transcriptomic changes were analyzed by RNA sequencing, followed by differential gene expression and functional enrichment analyses using the DESeq2 and g:Profiler software.

A single TNF-α stimulation significantly increased IL-8 production, whereas repeated stimulations progressively attenuated IL-8 secretion, indicating an adaptive (tolerance-like) response. RNA sequencing identified 2674 differentially expressed genes among the control, TNF-α∗1, and TNF-α∗4 groups. Early stimulation (TNF-α∗1) predominantly upregulated genes associated with immune and antiviral responses, whereas sustained stimulation (TNF-α∗4) induced gene expression patterns related to DNA replication and epithelial structural organization.

These findings indicate that under sustained TNF-α stimulation, human GECs initially induce gene expression related to inflammatory responses but subsequently exhibit transcriptional adaptation, which may contribute to regulating mucosal homeostasis during chronic inflammation.