Improving the microenvironment to augment endogenous regenerative potential has emerged as a fundamental concept for stimulating and expediting periodontal tissue repair and regeneration. Previous studies have demonstrated that TPPU, a soluble epoxide hydrolase inhibitor (sEHi), mediates the suppression of inflammatory bone loss in periodontitis models. However, the underlying mechanisms remain largely elusive. In this study, we constructed a human umbilical vein endothelial cell (HUVEC) and periodontal ligament stem cell (PDLSC) coculture system in vitro and tested the anti-inflammatory effect of TPPU under inflammatory conditions. The roles of HIF-1α and Endomucin (EMCN) in the anti-inflammatory effects of TPPU were analyzed. The effects of TPPU on osteogenesis and osteoclastogenesis in cocultured cells were examined. The in vivo periodontitis model further verified the effects of TPPU on inhibiting neutrophil adhesion and inflammation and inhibiting osteoclasts. Our in vitro experiments demonstrated that TPPU enhances the interaction between mesenchymal stem cells and vascular endothelial cells to enhance anti-inflammatory and osteogenic differentiation effects and revealed a new anti-inflammatory mechanism of TPPU involving the upregulation of EMCN in endothelial cells to prevent lymphocyte recruitment. We also confirmed that TPPU inhibits osteoclast activity. Our in vivo findings showed that TPPU inhibits osteoclast activity and neutrophil adhesion and enhances periodontal tissue repair and regeneration. TPPU promotes local regeneration in periodontitis by inhibiting inflammation and bone resorption. Thus, targeting soluble epoxide hydrolase represents an endogenous regenerative strategy for periodontitis treatment.

Type H vessels have recently been identified to modulate osteogenesis. Epoxyeicostrioleic acids (EETs) have an essential contribution to vascular homeostasis. However, whether increased EETs with soluble epoxide hydrolase (sEH) inhibitor TPPU enhance the coupling of angiogenesis and osteogenesis remains largely unknown. The effects of TPPU on cross-talk between co-cultured human umbilical vein endothelial cells (HUVECs) and human dental pulp stem cells (hDPSCs), and on long bone growth and calvarial defect repair in mice were investigated in vitro and in vivo. TPPU enhanced osteogenic differentiation of co-cultured HUVECs and hDPSCs in vitro and increased type H vessels, and long bone growth and bone repair of calvarial defect. Mechanistically, TPPU promoted cell proliferation and angiogenesis, reclined cell apoptosis, and significantly increased CD31hi EMCNhi endothelial cells (ECs) and SLIT3 and HIF-1α expression levels in co-cultured HUVECs and hDPSCs. Knockdown of Slit3 in hDPSCs or Hif-1α in HUVECs impaired the formation of CD31hi EMCNhi ECs and reversed TPPU-induced osteogenesis. We defined a previously unidentified effect of TPPU coupling angiogenesis and osteogenesis. TPPU induced type H vessels by upregulating the expression of hDPSCs-derived SLIT3, which resulted in the activation of ROBO1/YAP1/HIF-1α signalling pathway in ECs. Targeting metabolic pathways of EETs represents a new strategy to couple osteogenesis and angiogenesis, sEH is a promising therapeutic target for bone regeneration and repair.