Study On The Expression Of TLR2 And TLR4 Genes In Mastitis Breast Tissue Of Small Tail Han Sheep(Ovis Aries)

Toll-like receptor family(TLRs) is a very important pattern recognition receptors(PRRs) discovered in recent years, which perception molecular pattern(PAMPs) and identification of pathogenic microorganismsin innate immunity, by stimulating the signal cascade induced of inflammatory cytokines and cytokine production, and plays an important role in the anti-infection. To explore the expression of Toll-like receptor 2,4(TLR2,TLR4)in the Small Tail Han sheep(Ovis aries)mastitis tissue which caused by Escherichia coli and Staphyloccocus aureus. In the study, E.coli(O113 type) and S.aureus were used to infect Small Tail Han sheep breast and built the mastitis pathological model. The relative expression of TLR2 and TLR4 genes in the normal breast tissue and the breast tissue was in 48 h and 96 h post-infection with E.coli and S.aureus were detected by the methods of q RT-PCR. The expression and distribution of TLR2 and TLR4 protein in the normal breast tissue and the breast tissue infected with E.coli and S.aureus were examined by Western-blot and immunehistochemical SP method.The results showed that the TLR2 and TLR4 expressed in both normal breast tissue and the breast tissue infected with E.coli and S.aureus, but the TLR2 and TLR4 m RNA expressed were highest in 48 h post-infection, and the expression of TLR2 and TLR4 m RNA in 96 h post-infection had significantly lower than 48 h(P<0.05), and the relative levels of TLR2 and TLR4 were lowest in normal control group(P<0.05). Among them, the TLR4 m RNA expressed in the breast tissue infected with E.coli was volume significantly higher than the TLR2(P < 0.01). After the infection of S.aureus TLR2 and TLR4 m RNA expression in breast tissue volume difference was not significant. Western blot(WB) testing found that compared with the control group, after two bacterias infection in breast tissue relative expression of TLR4 protein was significantly enhanced, and 48 h relative expression level was significantly higher than 96 h(P <0.05).The results of immunohistochemistry demonstrated that TLR2 and TLR4 expressed in the breast epithelium in normal breast tissue, and compared with the control group, the positive expression was significantly enhanced after infected and expressed were highest in 48h post-infection(P <0.05). Among them, the breast tissue TLR4 expression was stronger than TLR2 positive expression(P <0.05) after the E. coli infection, after infected with S. aureus TLR2 and TLR4 protein mainly expressed in dropped mammary epithelial cells and lymphocyte predominant inflammatory cells. The results of this study showed that the expression of TLR2 and TLR4 was obviously increased after infected with E.coli and S. aureus in breast tissue of Small Tail Han sheep, which will establish the foundation the mechanism of TLR4 in sheep mastitis process during further study.