P. Gingivalis LPS Drives Osteoclastogenesis and Impairs Osteoblast Differentiation by Suppressing Sema3A/Nrp1 Via TLR-4

Abstract

Objectives

This study demonstrates that the Sema3A/Nrp1 pathway confers protection against Porphyromonas gingivalis LPS (P-LPS)-induced osteolysis and identifies key molecular mechanisms involved.

Methods

The effect of P-LPS on Sema3A/Nrp1 expression assessed in the preosteoclast cell line (RAW267.4) and primary mouse bone marrow stromal cells (BMSCs) using Real-time-PCR, western blot and ELISA. The effect of recombinant Sema3A on RAW267.4 cell and BMSC proliferation under P-LPS treatment was determined by CCK8 assay. Osteoclastic and osteoblastic markers gene was analysed by Real-time-PCR. ALP activity and mineralization were tested in BMSCs for osteoblast differentiation. TRAP staining and activity were measured for osteoclast differentiation. Micro-CT was applied to analysed P-LPS-induced calvarial osteolytic model.

Results

P-LPS induced significantly downregulated Sema3A/Nrp1 expression in both the preosteoclast cell line (RAW267.4) and primary mouse bone marrow stromal cells (BMSCs). Moreover, P-LPS exposure elevated osteoclastic gene expression, increased TRAP activity and promoted RANKL-induced osteoclast differentiation. However, these effects of promotion were attenuated by Sema3A administration. In addition, P-LPS impaired osteoblast differentiation in BMSCs, as evidenced by depressed osteogenic markers, attenuated β-catenin transcriptional activity, and diminished mineralization, all of which were rescued by Sema3A supplementation. Intriguingly, Sema3A alleviated P-LPS-induced repression of proliferation in BMSCs but did not affect the proliferation of RAW267.4 cells. Furthermore, P-LPS downregulated the expression of Sema3A/Nrp1 via Toll-like receptor 4 (TLR-4). Additionally, an in vivo study revealed that Sema3A administration markedly ameliorated P-LPS-mediated inflammatory osteolysis.

Conclusion

Our data identify the Sema3A-Nrp1 axis as a P-LPS–sensitive regulator of bone homeostasis, offering therapeutic potential to reverse infection-induced osteolysis through dual modulation of osteoclast and osteoblast activities. Sema3A attenuated PLPS- driven bone loss via concurrent osteoclast inhibition and osteoblast stimulation

Clinical Relevance

Our work identifies the above dual-targeting mechanism as a foundation for novel therapies against infection-related oral osteolysis.