Role And Mechanisms Of MKK3 Kinase In Staphylococcus Aureus Cutaneous Infection

Staphylococcus aureus is one of the most common gram-positive zoonotic pathogens that can cause group infection,but also an important pathogen can cause worldwide infection,which is mainly exists in skin and respiratory tract mucosa or other parts of the animal.S.aureus is an opportunistic pathogen,when the immune dysfunction or skin mucosa damage,S.aureus can break through the skin and mucosal barrier,and cause infectious disease such as skin,lung and other parts of the hosts,severe infection can leading to death due to the sepisis.In recent years,due to the large amount use of antibiotics in clinic,resulting in multidrugs-resistant S.aureus strains,which increased its pathogenicity and fatality rates year by year.In the process of infection and anti-infection of our body,innate immune system is the body's first line to defense against pathogenes,which plays an important role in controlling the spread of infection and inflammatory reaction.Mitogen-activated protein kinases family is an important signal transduction system in vivo,MKK3 encoding a protein called mitogen-activated protein kinase kinase 3,which is an important member of mitogen-activated protein kinases family.Numerous studies have shown that MKK3 plays a key role in natural immunity,inflammation and cytokine production,while the biological role of MKK3 in S.aureus infection has not yet been clarified.In this experiment we use wild-type(WT)mice and MKK3-deficient(MKK3-/-)mice by inject a certain dose of S.aureus subcutaneously,to explore the role of MKK3 during S.aureus cutaneous infection.Results shows that compared with WT mice,MKK3-/-mice skin abscess area is more significantly increased,while its microscopic pathological changes of skin tissue is more serious,otherwise inflammatory cell infiltration and edema in dermal parts of the skin tissue is more obvious,and the expression of inflammatory mediators as well as cytokines and chemokines secretion were significant increased(p<0.05),but the bacterial loads in skin tissue were not significant among WT mice and MKK3-/-mice(p>0.05).These results indicate that MKK3-/-mice shows higher susceptibility as well as inflammatory reaction in S.aureus cutaneous infection.That means that MKK3 plays an important role in the process of hosts against S.aureus cutaneous infection.Subsequently,mechanisms of MKK3 involved in regulating inflammatory response have been discussed in this experiment.The apotosis in skin tissue was analyzed by TUNEL staining,while the expression of PCNA in skin tissue was analyzed by immunofluorescence and the expression of p-MKK3 in skin tissue was analyzed by immunohistochemistry.Results have shown that the number of cell apoptosis and cell proliferation is low in the absence of S.aureus cutaneous infection.However after being infected with S.aureus,the cell apoptosis in MKK3-/-mice skin tissue are mainly distributed in the epidermis layer of the skin,and has no significant difference compared with WT mice(p>0.05).Whereas compared with WT mice,the expression of PCNA in skin tissue were significantly increased in MKK3-/-mice(p<0.0001).The expression of p-MKK3 was mainly expressed in inflammatory cells of the skin tissue.Recent studies have shown that IL-1? is an important cytokine,which plays an important role in the process of skin abscess formation as well as inflammatory cells migration during S.aureus cutaneous infection.But the main resource of IL-1? depends on inflammsomes activation.In vitro,primary mice peritoneal macrophage culture cell were used as a model to investigate the role of MKK3 in the regulation of inflammsomes activation during S.aureus infection.The results of western blot have shown that the expression of Caspase-1,IL-1? and ASC protein was significantly increased in MKK3-/-mice macrophages,and this process mainly depends on p-JNK signals.To further reveal the role of MKK3 in the host's resistance to S.aureus skin infection.The colonization of S.aureus in mice skin tissue was analyzed by selecting a long time infection.Results showed that,after the 12 th and 14 th days of S.aureus infection,the bacterial colonization in MKK3-deficient mice skin tissue was significantly increased(p<0.05),compared with WT infection group.This indicates that MKK3-deficient was not beneficial for macrophages to remove pathogens in the late stage of S.aureus infection,which have an impact on the normal bactericidal function of macrophages.In conclusion,this study have shown that MKK3 kinase can inhibit the phosphorylation of JNK signals,that inhibits macrophage inflammsomes activation,thereby inhibit the strong inflammatory response caused by S.aureus infection,promoting the bactericidal function of macrophages,which plays a protective effect on hosts.