Streptococcus oralis Induces Lysosomal Impairment of Macrophages via Bacterial Hydrogen Peroxide

Abstract

Streptococcus oralis , an oral commensal, belongs to the mitis group of streptococci and occasionally causes opportunistic infections, such as bacterial endocarditis and bacteremia. Recently, we found that the hydrogen peroxide (H ₂ O ₂ ) produced by S. oralis is sufficient to kill human monocytes and epithelial cells, implying that streptococcal H ₂ O ₂ is a cytotoxin. In the present study, we investigated whether streptococcal H ₂ O ₂ impacts lysosomes, organelles of the intracellular digestive system, in relation to cell death. S. oralis infection induced the death of RAW 264 macrophages in an H ₂ O ₂ -dependent manner, which was exemplified by the fact that exogenous H ₂ O ₂ also induced cell death. Infection with either a mutant lacking spxB , which encodes pyruvate oxidase responsible for H ₂ O ₂ production, or Streptococcus mutans , which does not produce H ₂ O ₂ , showed less cytotoxicity. Visualization of lysosomes with LysoTracker revealed lysosome deacidification after infection with S. oralis or exposure to H ₂ O ₂ , which was corroborated by acridine orange staining. Similarly, fluorescent labeling of lysosome-associated membrane protein-1 gradually disappeared during infection with S. oralis or exposure to H ₂ O ₂ . The deacidification and the following induction of cell death were inhibited by chelating iron in lysosomes. Moreover, fluorescent staining of cathepsin B indicated lysosomal destruction. However, treatment of infected cells with a specific inhibitor of cathepsin B had negligible effects on cell death; instead, it suppressed the detachment of dead cells from the culture plates. These results suggest that streptococcal H ₂ O ₂ induces cell death with lysosomal destruction and then the released lysosomal cathepsins contribute to the detachment of the dead cells.