Sulfur Metabolism in Porphyromonas gingivalis: Implications for Blood-Brain Barrier Permeability in an In Vitro Model

Abstract

Abstract:

Porphyromonas gingivalis, a key causative agent in chronic periodontitis, has been increasingly associated with systemic illnesses, such as neurological disorders. The objective of this work was to examine the impact of P. gingivalis and its interaction with L-cysteine on the integrity of the blood-brain barrier (BBB) utilising the hCMEC/D3 cell line as an in vitro model. Our hypothesis was that P. gingivalis, especially when combined with L-cysteine, would impair the integrity of the blood-brain barrier (BBB) via affecting sulphur metabolism.

Methods: We utilised both wild-type (W50) and mutant (PG343) strains of P. gingivalis to create conditioned medium, both with and without the addition of L-cysteine. The integrity of BBB was evaluated by fluorescent tracer permeability experiments, transendothelial electrical resistance (TEER) measures, and immunofluorescence imaging of the actin cytoskeleton. In addition, bismuth chloride tests were conducted to analyse the integrity of bacterial membranes, while MTT assays were employed to investigate the impact of mercaptoethanol on the vitality of hCMEC/D3 cells.

Results: The conditioned medium of wild-type P. gingivalis caused a considerable increase in BBB permeability, and this effect was enhanced by L-cysteine. The TEER tests exhibited consistent stability, indicating that alterations in permeability might transpire via mechanisms unrelated to significant disruption of tight junctions. Immunofluorescence imaging showed significant alterations in cell shape and actin arrangement in hCMEC/D3 cells when exposed to both P. gingivalis conditioned media and L-cysteine. The mutant strain PG343, which has a deficiency in L-cysteine metabolism, did not have a significant impact on blood-brain barrier (BBB) permeability or transendothelial electrical resistance (TEER) values. Analysis of bismuth chloride revealed that the wild-type P. gingivalis, when exposed to L-cysteine, exhibited heightened membrane permeability. However, this effect was not observed in the mutant strain. The MTT assays revealed that the vitality of hCMEC/D3 cells was affected by mercaptoethanol in a concentration-dependent manner.

Conclusion: our findings confirm the idea that sulphur metabolism is important for the virulence of P. gingivalis and its possible impact on the integrity of the blood-brain barrier (BBB). P. gingivalis and L-cysteine's synergistic effect on endothelial cell morphology and BBB permeability proposes a potential mechanism by which this oral pathogen could contribute to neurological disorders. These findings emphasise the complex connections between periodontal infections, their byproducts, and the blood-brain barrier, emphasising the necessity for additional research on the molecular mechanisms involved in these processes.

Keywords: Porphyromonas gingivalis, blood-brain barrier, L-cysteine, sulfur metabolism, neurological disorders, periodontal disease.