The Impact of the VEGF/VEGFR2/PI3K/AKT Signaling Axis on the Proliferation and Migration Abilities of Human Dental Pulp Stem Cells.

Abstract

The potential therapeutic benefits of human dental pulp stem cells (HDPSCs) in dental regenerative medicine have been demonstrated. However, little is known about the molecular mechanisms regulating the biological characteristics of HDPSCs. The experiment aims to explore whether VEGF activates signaling pathways such as FAK, PI3K, Akt, and p38 in HDPSCs, and to investigate the molecular mechanisms by which VEGF influences proliferation and migration of HDPSCs. Normal and inflamed human dental pulp (HDP) samples were collected, and the levels of VEGF in HDP were assessed. HDPSCs were cultured and purified. HDPSCs were stimulated with lipopolysaccharide (LPS) at gradient concentrations, and real-time quantitative polymerase chain reaction (qPCR) was used to assess changes in VEGF mRNA. Gradient concentrations of VEGF were used to stimulate HDPSCs, and cell migration ability was evaluated through scratch assays and Transwell chamber experiments. Phosphorylation levels of FAK, AKT, and P38 were assessed using Western blotting. Inhibitors of VEGFR2, FAK, AKT, P38, and VEGF were separately applied to HDPSCs, and cell migration ability and phosphorylation levels of FAK, AKT, and P38 were determined. The results indicated significant differences in VEGF levels between normal and inflamed HDP tissues, with levels in the inflamed state reaching 435% of normal levels (normal: 87.91 ng/mL, inflamed: 382.76 ng/mL, P < 0.05). LPS stimulation of HDPSCs showed a significant increase in VEGF mRNA expression with increasing LPS concentrations (LPS concentrations of 0.01, 0.1, 1, and 10 μg/mL resulted in VEGF mRNA expressions of 181.2%, 274.2%, 345.8%, and 460.9%, respectively, P < 0.05). VEGF treatment significantly enhanced the migration ability of HDPSCs in Transwell chamber experiments, with migration rates increasing with VEGF concentrations (VEGF concentrations of 0, 1, 10, 20, 50, and 100 ng/mL resulted in migration rates of 8.41%, 9.34%, 21.33%, 28.41%, 42.87%, and 63.15%, respectively, P < 0.05). Inhibitors of VEGFR2, FAK, AKT, P38, and combined VEGF stimulation demonstrated significant migration inhibition, with migration rates decreasing to 8.31%, 12.64%, 13.43%, 18.32%, and 74.17%, respectively. The migration rate with combined VEGF stimulation showed a significant difference (P < 0.05). The analysis of phosphorylation levels revealed that VEGF stimulation significantly activated phosphorylation of FAK, AKT, and P38, with phosphorylation levels increasing with VEGF concentrations (P < 0.05). The VEGF/VEGFR2 signaling axis regulated the migration ability of HDPSCs through the FAK/PI3K/AKT and P38MAPK pathways. This finding highlighted not only the crucial role of VEGF in injury repair of HDPSCs but also provided important clues for a comprehensive understanding of the potential applications of this signaling axis in dental regenerative medicine.