Studies On Expression Of Toll-like Receptors And Cytokines In RAW264.7 Cells And Dendritic Cells Induced By Streptococcus Equi

Dendritic Cells(DCs) and Macrophages(M?s) are major host cells infected by Streptococcus equi subspecies equi(S.equi). Bacterium are recognized by pattern recognition receptors(PRRs) in cells through different signaling pathways, starting the expression of cytokines, and activating the innate immune response. In order to investigate the effect of Toll-like receptors(TLRs)-mediated signaling pathway on host cells infected by S.equi, this study has carried out mainly works as two aspects:(1)The mRNA expression levels of TLRs, Myeloid differentiation factor 88(MyD88) and cytokines in RAW264.7 cells infected by S.equi were detected by real-time fluorescence quantitative PCR. The results showed that at 6 h post-infection, the mRNA expression levels of TLR1, TLR2, TLR6 and MyD88 in S.equi-infected RAW264.7 cells had no change compared with those of uninfected group; at 12 h post-infection, the mRNA expression levels of TLR1, TLR2 and TLR6 had no obviously change, while the mRNA expression level of MyD88 was increased; at 24 h post-infection, the mRNA expression levels of TLR1, TLR2, TLR6, IL-10 and IL-12 were significantly increased, but IL-1, IL-6 and TNF-? mRNA levels were significantly decreased. These results indicated that the TLRs-mediated signaling pathway was participated in the immune response against S.equi.(2)DCs were cultured with GM-CSF and IL-4, and real-time fluorescence quantitative PCR was used to detect mRNA expression levels of TLRs, MyD88 and cytokines in DCs infected by S.equi at 16, 20, 26 h. The results showed that at 16, 20, 26 h post-infection, the mRNA expression levels of TLR1, TLR2, TLR6 and MyD88 in DCs infected by S. equi were significantly higher than those of the uninfected group. At 16, 26 h post-infection, the mRNA expression level of IL-1 had no change, IL-10 and TNF-? were significantly increased. IL-6 was significantly increased at 16 h post-infection, but no change was seen at 26 h post-infection. At 16 h post-infection, the mRNA expression level of IL-12 had no obviously change, while it was significantly increased at 26 h post-infection. The expression levels of CD80, CD86, MHCII and cytokines in DCs infected by S.equi were detected by flow cytometry and real-time fluorescence quantitative PCR when TLR2 was blocked by treating with anti-TLR2 IgG, respectively. The results showed that the expression levels of CD80, CD86, MHCII had no change, but the mRNA expression levels of IL-6 and IL-12 were significantly decreased after treatment of DCs with anti-TLR2 IgG. These results indicated that S. equi was able to regulate the expression levels of TLRs, MyD88 and cytokines, and induced the productions of IL-6 and IL-12 in DCs via the TLR2 signalling pathway.In summary, our studies found that S.equi could affect the mRNA expression of TLRs, MyD88 and cytokines in RAW264.7 cells and DCs, and the productions of IL-6 and IL-12 in DCs were regulated via the TLRs-mediated signalling pathway. These findings provide experimental basis for the interactions between S.equi and target cells.