Regulation Of TLR2,TLR4 And NLRP3 On Inflammatory Injury Induced By Staphylococcus Aureus In Mice

Staphylococcus aureus(S.aureus)is a common gram-positive bacterium,which colonizes on human skin and mucous membrane.At the same time,it is also an important pathogen causing clinical infection,such as bacteremia,osteomyelitis,endocarditis and necrotizing pneumonia.The intensity of host inflammatory response induced by S.aureus infection is usually closely related to the expression of inflammatory mediators and organ injury.As the first line of defense of the host,innate immunity plays a key role in S.aureus infection.The major virulence factor lipoprotein(BLP)of S.aureus induces inflammation in the host by recognizing TLR2,TLR4 and NLRP3 receptors.TLR2 receptors were found to play a major role in this process,but their specific role in the different stages of the host inflammatory response remains unclear.In order to further explore the regulatory mechanism of innate immune receptors TLR2,TLR4 and NLRP3 in the inflammatory response induced by S.aureus infection,This study in C57BL/6J,TLR2^-/-,TLR4^-/-and NLRP3^-/-mice and separation of M1 bone marrow macrophage(BMDM)as the research object,by q PCR,ELISA and Western Blot and immunofluorescence technique,tested the mice serum and lung,and M1 BMDM cell factors(TNF-?,RANTES,IL-1?and IL-10),natural immune receptor(TLR2 and TLR4 and NLRP3)and the expression of damage factor(HMGB1 and HABP2),The changes of p38,JNK,ERK and p65 phosphorylation levels were analyzed,and the mortality of C57BL/6J,TLR2^-/-,TLR4^-/-and NLRP3^-/-mice infected with S.aureus were observed.The results showed that S.aureus lipoprotein could promote the expression of TNF-?,IL-1?,RANTES and IL-10 in M1 BMDM.The expression of TLR2,TLR4 and NLRP3 genes were increased,and the phosphorylation levels of p38,JNK,ERK and p65 were enhanced.After 9h of SA113 infection,TLR2^-/-,TLR4^-/-and NLRP3^-/-M1 BMDM significantly down-regulated the levels of TNF-?,IL-1?,RANTES and IL-10 in TLR2^-/-,TLR4^-/-and NLRP3^-/-M1 BMDM(P<0.05).The phosphorylation levels of JNK and p38 induced by TLR2^-/-M1 BMDM in mice were significantly lower than those in C57BL/6J mice.TNF-?,IL-1?and RANTES were significantly upregulated in TLR2^-/-,TLR4^-/-and NLRP3^-/-macrophages compared with M1 BMDM of C57BL/6J mice 24h after infection with wild-type S.aureus(P<0.05).Compared with C57BL/6J,TLR4^-/-and NLRP3^-/-mice,TLR2^-/-mice had the fastest mortality rate,and all mice died within 48 hours.The secretion of TNF-?,IL-1?,RANTES and IL-10 in serum and lung of TLR2^-/-,TLR4^-/-and NLRP3^-/-mice was significantly reduced after 3h and 6h of SA113infection compared with C57BL/6J mice(P<0.05).The expression of HMGB1 and HABP2 proteins in the lungs of TLR2^-/-mice was significantly decreased.The secretion of TNF-?,RANTES,and IL-10 in serum and lung of TLR2^-/-mice was significantly decreased compared with C57BL/6J mice after 24h of SA113 infection(P<0.05),and significantly inhibited(P<0.05)the expression levels of HMGB1 and HABP2 proteins.In conclusion,S.aureus lipoprotein is the main immunoactive substance that induces the activation of innate immune response by M1 BMDM.When SA113 infected mice,TLR2 mainly played a role in the early inflammatory response to recognize S.aureus.Therefore,when TLR2 was deleted,the activation of MAPK and NF-?B inflammatory signaling pathway was down-regulated,leading to down-regulated secretion of inflammatory cytokines.Together,the two acted to down-regulate lung tissue and lung injury.Studies have shown that when the host is infected with S.aureus for a long time,the body can establish a tolerance phenomenon,and then regulate the inflammatory response.Based on the comprehensive analysis of the above studies and the results obtained in this study,the research group concluded that when TLR2 was absent,mice did not have the conditions to establish such tolerance and could not properly regulate the intensity of inflammatory response,leading to the explosion of cytokines,which led to the increase of lung injury and mortality in mice.