Effects of Porphyromonas gingivalis lipopolysaccharide on the behavior of human dental pulp stem cells in vitro and their inflammation-related transcriptomics profile

Abstract Objective: This study aims to investigate the effects of Porphyromonas gingivalis lipopolysaccharide (P. g LPS) on the migration, proliferation, and odontoblast differentiation of human dental pulp stem cells (DPSCs) in vitro, and to explore the transcriptomics related to LPS-induced inflammation. Methods: Human DPSCs were cultured and treated with 0.5-2 µg/mL of LPS for 24, 48, and 72 h, respectively. The expression of Arg-1, IL-1β, and TNF-α in DPSCs and cell viability was assessed. The time and concentration of LPS with the least influence on cell growth were selected to establish an in vitro pulpitis model. The CCK8 assay and Transwell method were used to detect the influence of LPS on cell proliferation and migration. LPS-stimulated DPSCs were induced to differentiate into odontoblasts, and the effect of LPS on cell differentiation was analyzed. RNA-Seq analysis was performed to investigate the transcriptomics profile related to LPS inflammation Results: Based on the results of cell viability and expression of inflammatory factors, DPSCs pre-treated with 1µg/mL of LPS for 24 hours were utilized as the in vitropulpitis model in this study. Under these conditions, the migration and cell proliferation rate of DPSCs were significantly suppressed (P<0.01). Following induction by odontoblast differentiation medium for 21 days, the expression of odontoblast marker genes (DSPP, DMP1, and RUNX2) and alkaline phosphatase in LPS-stimulated cells was significantly inhibited (P<0.01), and the number of calcium staining-positive cell nodules reduced significantly compared to that in the control group. Transcriptomic analysis revealed enrichment of the TLR4/NF-κB signaling pathway and NOD-like receptor signaling pathway under LPS stimulation; KEGG analysis showed enrichment of PI3K-Akt signaling pathway, chemokine action pathway, TNF, NF-kappaB signaling pathway in DPSCs under inflammatory conditions. Conclusions: DPSCs have the capacity to secrete inflammatory cytokines and contribute to tissue inflammatory reactions. Under inflammatory conditions in this study, their migration, proliferation, and differentiation abilities are inhibited. LPS participates in the regulation of cell signaling networks through multiple mechanisms. Clinical significance: This study suggests that DPSCs may still maintain their proliferation and differentiation capabilities under severe inflammation in clinical practice. Therefore, controlling inflammation and enhancing the biological properties of stem cells are important strategies for promoting the healing of dental pulp.