Subinhibitory concentrations of tetracyclines induce lipopolysaccharide shedding by Porphyromonas gingivalis and modulate the host inflammatory response

Abstract

Antibiotics at below minimal inhibitory concentrations (MICs) may induce various biological responses in bacteria. In this study, we hypothesized that subinhibitory concentrations (subICs) of tetracycline and doxycycline induce the shedding of lipopolysaccharide (LPS) by Porphyromonas gingivalis and, as a consequence, may contribute to enhancing the host inflammatory response associated with periodontitis. A polymyxin-based enzyme-linked immunosorbent assay was used to quantify LPS shedding by P.gingivalis grown in the presence of subICs of tetracycline and doxycycline. A macrophage model was used to show that tetracycline- and doxycycline-mediated LPS shedding by P.gingivalis can induce cytokine secretion. The secretion of interleukin (IL)-1β, IL-8, and tumor necrosis factor-α was quantified by enzyme-linked immunosorbent assay. LPS was shed spontaneously in a time-dependent way by P.gingivalis during growth. LPS shedding was significantly increased by growth in the presence of subICs of tetracycline and doxycycline corresponding to 1/20 of their MICs (0.025μg/mL for tetracycline and 0.0125μg/mL for doxycycline). This shedding was not associated with an increased rate of bacterial cell lysis. Stimulating macrophages with a P.gingivalis culture supernatant induced the secretion of IL-1β, IL-8 and tumor necrosis factor-α when the bacteria were grown in the presence of 1/20 MIC of the antibiotics. Our study showed that growing P.gingivalis in the presence of subICs of either tetracycline or doxycycline induces LPS shedding. Shed LPS may in turn increase cytokine secretion in a macrophage model.