A Study Of The Relationship Between Apoptotic Cell Clearance And Innate Immune Response In Caenorhabditis Elegans

The appropriate clearance of apoptotic cells is indispensable for apoptosis,which is crucial to maintain the development of multicellular organs and internal environmental homeostatic.If the apoptotic cells produced in the normal body can not be removed in time and effectively,the corpse and the harmful contents released by the apoptotic cells will lead to inflammation or autoimmune diseases(such as systemic lupus erythematosus and rheumatoid arthritis).and neurodegenerative diseases(such as Parkinson's disease and Alzheimer's disease).Apoptotic cell clearance is highly conserved in evolution,while it is extremely redundant and complex in mammals.To better explore the molecular mechanism of apoptotic cell clearance,people often rely on model organisms.C.elegans has been used as an excellent model organism for studying the mechanism of apoptotic cell clearance because of its own advantages.The invertebrate model C.elegans has also been widely used to study host-pathogen interactions due to the evolutionary conservation of the innate immune system.Similar to other invertebrates.C.elegans relies entirely on its innate immune system to defend against pathogens.Besides.P.aeruginosa PA14 and S.typhimurium SL1344 are commonly used to infect nematodes to study their innate immune response.Through the research on nematodes infected w ith various bacteria and other pathogens to help people understand the mechanism of innate immunity.In mammals,disorders of innate immunity are associated with inflammatory diseases.cancer,and autoimmune diseases.Previous studies have confirmed that the loss-of-function of DNase ?.a gene related to apoptotic cell clearance in Drosophila melanogaster,upregulated the expression of pathogen response genes and activated its innate immunity.Recent research found that activation of MPK-1 signaling pathway in C.elegans can facilitate the apoptosis of germ cells and that it depends at least in part on the phagocytic mechanism of apoptotic cells.However,little is known about the relationship between apoptotic cell clearance and innate immune in C.elegans.Based on this,this project explores the relationship between apoptotic cell clearance and innate immune response through the model organism of C.elegans,to provide a certain theoretical basis for revealing the pathogenesis of autoimmune diseases caused by apoptotic cell clearance disorders.Based on this research background,the following research results have been obtained on this subject:1.Apoptotic cell clearance defect mutant nematodes show resistance to P.aeruginosa PA14 and S.typhimurium SL1344.Previous studies have found C.elegans was more susceptible to produce antibacterial effect to resist pathogenic bacteria infection and its lifespan was extended when p38 MAPK-dependent innate immune response was activated.Therefore,lifespan analysis experiments were performed after apoptotic cell clearance defective mutant nematodes infected with pathogenic bacteria PA 14 and SL1344,respectively,and it was found that these defective mutant nematodes lifespan had been significantly extended than that of the wild-type nematodes N2.These results preliminarily indicate that the defect of apoptotic cell clearance may activate the innate immune response of C.elegans.2.The defect of apoptotic cell clearance can cause the upregulation of the expression of pathogen response genes in C.elegans.The expression of six pathogen response genes involved in innate immunity in apoptotic cell-deficient mutant nematodes were significantly increased,which were detected by qRT-PCR technology.nlp-29 is a pathogen response gene involved in innate immunity.The expression of GFP in HMZ009 transgenic nematode containing the pnlp-29?gfp reporter gene also increased significantly even without pathogenic bacterial infection after two apoptotic cell clearance related genes,ced-1 and ced-2,were silenced by using RNAi technology.This indicates that the pnlp-29?gfp reporter gene was activated.Our results further demonstrate that defect of apoptotic cell clearance can activate the innate immune response of C.elegans.3.The defect of apoptotic cell clearance causes the upregulation of PMK-1 and MPK-1 phosphorylation levels in the innate immunity of C.elegans.PMK-1 p38 MAPK and MPK-1/ERK MAPK are two major signaling pathways involved in the innate immunity of C.elegans.The levels of PMK-1 and MPK-1 phosphorylation in mutant nematodes involved in apoptotic cell clearance defect were significantly upregulated when compared to the control group,which were detected by Western blot.These indicate that PMK-1 and MPK-1 signaling pathways are activated.It is suggested that the defect of apoptotic cell clearance may be through the PMK-1 and MPK-1 signaling pathways to activate innate immune response of C.elegans.4.ced-1 and ced-2 participate in the innate immune response of C.elelegans against P.aeruginosa PA14 through MPK-1 signaling pathway.Lifespan analysis revealed that ced-1 and ced-2 participated in the innate immune response of C.elegans against P.aeruginosa PA14 through MPK-1 signaling pathway.Finally,the mechanism of apoptotic cell clearance regulating the innate immunity in C.elegans was explored by western blot and genetic epistasis analysis.We found that apoptotic cell clearance-related genes may directly or indirectly act on PMK-1 signaling pathway to regulate the innate immune response of C.elegans.In summary,our results show that the defect of apoptotic cell clearance can regulate the innate immune response of C.elegans through MPK-1 and PMK-1 signaling pathways,but the underlying mechanism remains to be further explored.