The Effects And Mechanisms Of GPR120 Protecting Against Clostridium Perfringens Infection

Clostridium perfringens is an important zoonosis pathogen,which seriously endangers the development of animal husbandry and human health.It is one of the important public health problems in the world.Due to the emergence of monoresistant and multidrug-resistant strains,the expansion of drug resistance spectrum and the mixed infection of multiple serotypes,new prevention and treatment methods are urgently needed.As a membrane protein receptor,G-protein coupled receptor 120（GPR120）has a typical seven transmembrane structure,which can activate intracellular G protein and signal transduction molecules,and then induce biological effects.At present,the reports on the biological function of GPR120 mainly focus on the function of GPR120ω-Fatty acid receptors are involved in the regulation of obesity,type 2 diabetes,fatty liver and some self-inflammatory diseases.Although the function of GPR120 has been known,little is known about its function and mechanism in anti infective immune response.The infectivity and pathogenicity of pathogens largely depend on the interaction between pathogens and host immune system,so it is difficult to effectively control pathogens only from the perspective of pathogens.Innate immunity is the first line of defense of host against pathogen infection.It is beneficial to the prevention and control of diseases to study the anti infection immune response of the body and find the intervention target.This study investigated the role of GPR120 in host defense against Clostridium perfringens infection.WT and GPR120^-/-mice were used to establish the model of Clostridium perfringens muscle infection（gas gangrene）.Firstly,whether GPR120 affects the susceptibility of host to Clostridium perfringens was analyzed.The results showed that compared with WT mice,GPR120^-/-mice were more susceptible to Clostridium perfringens gas gangrene,mainly manifested by more bacteria loading in muscle tissue,more severe pathological damage and recruitment of a large number of neutrophils and macrophages.In addition,Clostridium perfringens infection significantly promoted the expression of IL-1β,TNF-αand KC in the muscle tissue of WT mice.However,compared with WT mice,the expression level of IL-1βin the muscle tissue of GPR120^-/-mice was significantly decreased,the expression level of TNF-αwas not significantly different,and the expression level of KC was significantly increased.These results suggest that GPR120 is involved in the host immune defense against Clostridium perfringens infection,which can inhibit the proliferation of Clostridium perfringens,reduce the histopathological damage,and play an indispensable protective role in the host defense against Clostridium perfringens infection.Since GPR120 is mainly expressed in the inflammatory cells in the muscle tissue of mice infected with Clostridium perfringens,it is also found that GPR120 is expressed in macrophages,and Clostridium perfringens can significantly up regulate the expression of GPR120.Therefore,mouse macrophages are the main cells for in vitro study to clarify the molecular mechanism of GPR120limiting Clostridium perfringens infection.Based on the results of in vivo study,compared with WT mice,the expression level of inflammatory cytokine IL-1βin muscle tissue of GPR120^-/-mice was significantly decreased after Clostridium perfringens infection,but there was no significant difference in the expression level of non-inflammatory cytokine TNF-α.At the same time,recent studies have shown that NLRP3 inflammasome plays an important role in host defense against pathogen infection.Therefore,we analyzed whether GPR120 mediated anti infection protection depends on NLRP3inflammasome signal.The results showed that GPR120 deletion significantly inhibited K⁺dependent NLRP3 inflammasome signal activation induced by Clostridium perfringens.At the same time,we used WT and NLRP3^-/-mice to construct Clostridium perfringens gas gangrene model,and compared the anti infective effects of GPR120 and NLRP3.The results showed that,similar to GPR120^-/-mice,NLRP3^-/-mice were more susceptible to Clostridium perfringens gas gangrene than WT mice.The main manifestations were more severe pathological injury of muscle tissue,more bacteria loading,a large number of inflammatory cell infiltration and significantly decreased expression of IL-1β.There was no significant difference in the expression of TNF-α,but KC expression was significantly increased.In addition,GPR120 and NLRP3 deletion inhibited the killing ability of macrophages against Clostridium perfringens,and showed convergence effect.These results suggest that the anti infective protective effect mediated by GPR120 depends on NLRP3 inflammasome.It is known that the activation of NLRP3 inflammasome requires two signals:signal stage I mainly activates NF-κB signaling pathway through IL-1r1,TLRs,TNFR1,TNFR2 and other ligands to promote the expression of NLRP3,pro-Caspase-1 and pro-IL-1β;signal stage II mainly involves the assembly of NLRP3 inflammasome,which mediates cell apoptosis and cleaves and secretes bioactive IL-1βand IL-18 to the outside of cells.The results showed that GPR120 deletion did not affect the expression of NLRP3,pro-IL-1β,GSDMD-FL and ASC,but significantly inhibited the production of ROS,the formation of ASC spots and the activation of K⁺dependent NLRP3inflammasome.These results suggest that the regulation of NLRP3 inflammasome signal by GPR120does not depend on the regulation of the first signal,but on the regulation of the second signal,and the specific mechanism needs to be further verified.In conclusion,GPR120 is involved in host immune defense against Clostridium perfringens infection,and can inhibit the proliferation and histopathological damage of Clostridium perfringens.Moreover,GPR120 mediated anti infection protection depends on NLRP3 inflammasome.We found a new intervention target for Clostridium perfringens disease,which provides a basis for further development of GPR120 signaling for the prevention and treatment of animal and human Clostridium perfringens disease objective to provide scientific basis for drug and vaccine development and clinical medication.