An Oral Bacteria Mixture Induces Inflammation in Endothelial Cells

Objectives: Oral bacteria are associated with cardiovascular disease (CVD); however, their roles in its pathogenesis are unclear. We evaluated the ability of a mixture of oral bacteria to induce inflammation in human aortic endothelial cells (HAECs) using Streptococcus mutans as a control. We examined the cytotoxicity of the mixture of oral bacteria, and its effect on the production of cytokines and pattern-recognition receptors by HAECs.
Methods: Bacteria were collected from stimulated human saliva (n = 3) by centrifugation. The morphology and viability of HAECs treated with the mixture of oral bacteria or S. mutans were examined at a multiplicity of infection (MOI) of 1 following co-culture for 2, 4, 8, or 24 h. To assess induction of inflammation, HAECs were co-cultured with the mixture of oral bacteria or S. mutans (37°C, 5% CO₂, 4 h; MOI = 1), exposed to antibiotics (1 h), and cultured for a further 24 h. Cytokine production by HAECs was quantified by enzyme-linked immunosorbent assays, and the expression levels of pattern-recognition receptors were evaluated by real-time polymerase chain reaction.
Results: The mixture of oral bacteria induced morphological changes in HAECs, which became of irregular shape or round. HAEC viability was < 90% following co-culture for 8 h with the mixture of oral bacteria; in contrast, S. mutans had no effect. The mixture of oral bacteria induced production of IL-6, IL-8, and MCP-1 proteins, and TLR2 and NOD2 mRNAs, by HAECs (ANOVA, Dunnett’s, p < 0.05); S. mutans did not induce production of any of these factors.
Conclusions: The mixture of oral bacteria induced production of IL-6, IL-8, and MCP-1 proteins, and TLR2 and NOD2 mRNAs, in HAECs. This greater induction of inflammation compared to a single oral bacterial species may be due to the presence of diverse bacterial species in the mixture.