Tumor necrosis factor-α and interleukin-1β expression pathway induced by Streptococcus mutans in macrophage cell line RAW 264.7

Streptococcus mutans, a major etiological agent of dental caries, frequently causes systemic disease, such as subacute bacterial endocarditis, if it enters the bloodstream. In this study, the production pathways of the proinflammatory cytokines, tumor necrosis factor-a (TNF-a) and interleukin-1 beta (IL-1 beta), induced by S.similar to mutans in mouse macrophage were examined using a quantitative real-time polymerase chain reaction and an enzyme-linked immunosorbent assay. The S.similar to mutans stimulated the expression of TNF-a and IL-1 beta mRNA at a multiplicity of infection of 1 : 100, which increased at 2 and 4 h, respectively, to 24 h. It also induced the production of high levels of the TNF-a and IL-1 beta proteins, which increased at 2 h and reached a peak at 4 and 24 h, respectively. Nuclear factor-?B (NF-?B) was activated and reached a maximum level 30 min after the S.similar to mutans treatment. The expression of TNF-a and IL-1 beta mRNA and protein was suppressed by the treatment with pyrrolidine dithiocarbamate, an NF-?B inhibitor. The S.similar to mutans-induced TNF-a expression was suppressed by the presence of SB203580, a p38 mitogen-activated protein (MAP) kinase inhibitor, or SP600125, a Jun N-terminal kinase (JNK) MAP kinase inhibitor. On the other hand, IL-1 beta expression was inhibited by extracellular signal-regulated kinase (ERK)/p38/JNK MAP kinase inhibitor pretreatment. In addition, TNF-a production was suppressed more in the Toll-like receptor 2-/- (TLR2-/-) macrophages than in the TLR4-/- macrophages, whereas IL-1 beta production was suppressed more in the TLR4-/- macrophages than in the TLR2-/- macrophages. These results show that S.similar to mutans stimulates the production of TNF-a and IL-1 beta in the mouse macrophage cell line, RAW 264.7, by activating ERK/p38/JNK, and NF-?B through TLR2 and TLR4, respectively.