Lactate‐Induced H3K18 Lactylation by p300 Promotes Odontogenic Differentiation of Human Dental Pulp Stem Cells via WNT5A /Ca 2+ Signalling Pathway: A Laboratory Investigation

ABSTRACT Aim To delineate the mechanisms underlying the role of lactate‐related lactylation, a novel post‐translational modification, in the odontogenic differentiation of human dental pulp stem cells (hDPSCs). Methodology Dynamic levels of key glycolytic enzymes and histone lactylation along odontogenic differentiation in hDPSCs were evaluated by Western Blot and quantitative PCR (qPCR). Odontogenic differentiation was assessed via Alkaline Phosphatase and Alizarin Red S staining after treatment with sodium lactate, both with and without p300 knockdown. CUT&Tag sequencing of histone H3K18 lactylation (H3K18la) was performed, and genes identified through this analysis were subjected to pathway enrichment. WNT5A‐knockdown hDPSCs were generated to elucidate the impact of both WNT5A and WNT5A/Ca 2+ signalling axis in odontogenesis. Finally, we validated the critical function of lactylation in dentine regeneration using a subcutaneous ectopic transplantation model in nude mice with and without sodium lactate via intraperitoneal injection. Results The similar dynamic trends were observed in glycolytic enzyme expression, intracellular lactate and lactylation levels in hDPSCs. Exogenous lactate dose‐dependently increased H3K18 lactylation and significantly promoted hDPSC odontogenic differentiation. Mechanistically, the key lactyltransferase p300 was demonstrated to be a regulator that enhanced lactate‐induced H3K18la modification and odontogenic differentiation in hDPSCs. CUT&Tag sequencing identified WNT5A as a target gene whose promoter exhibited increased H3K18la enrichment upon lactate treatment. Lactate upregulated WNT5A expression and activated the WNT5A/Ca 2+ signalling pathway via p300‐mediated H3K18la, consequently accelerating the odontogenic differentiation. Further in vivo evidence revealed that lactate administration promoted pulp‐like tissue regeneration, accompanied by increased hDPSC odontogenic marker expression and H3K18la levels in nude mice with subcutaneously transplanted dentine slices. Conclusions The lactate‐driven mechanism acts through the p300‐mediated H3K18la‐WNT5A/Ca 2+ signalling axis to promote the odontogenic differentiation of hDPSCs. The crosstalk between metabolism, epigenetic modifications and signalling pathways of hDPSCs might offer novel strategies for dental pulp regeneration.