Molecular Mechanisms Of Caspase 8 Promoting Innate Immunity Mediated By Porcine Epidemic Diarrhea Virus

Porcine epidemic diarrhea(PED)is an acute,highly fatal,contact infectious disease caused by the porcine epidemic diarrhea virus(PEDV),which can cause acute gastroenteritis in pigs,accompanied by watery diarrhea,vomiting,The dehydration and death of piglets and the mortality rate of piglets are as high as 100%,which brings huge economic losses to our country and the world.The current research on PEDV still needs to be in-depth.In the case of vaccine immunization,there will still be a popular trend.Therefore,it is necessary to study the mechanism of PEDV infection and immunity to provide new directions and theoretical technical support for the prevention,control and immunization of the pathogen.Apoptosis is a defense mechanism of the cell itself,which can prevent a wide range of infections by activating apoptosis when it is stimulated by pathogens.In cell apoptosis,Caspase-8(Caspase-8)plays an important role,and Caspase-8 can also participate in the regulation of immune-related pathways.When the cell undergoes apoptosis due to Caspase-8 cleavage,the activation of Caspase-8 can cleave the deubiquitinating enzyme Cylindromatosis(CYLD)and make it lose its deubiquitination function.Retinoic acid-induced gene protein regulated by CYLD I(Retinoic acid-inducible gene I,RIG-I)will smoothly ubiquitinate and activate the nuclear factor?B(NF-?B)pathway and the RIG-I-like receptor pathway.On the one hand,RIG-I ubiquitination plays an important role in the up-regulation of NF-?B phosphorylation.By up-regulating the phosphorylation of I?B kinase(Inhibitor of NF-?B kinase,IKK),it activates the I?B(NF-?B inhibitor alpha).The ubiquitination of I?B regulates the activity of NF-?B.On the other hand,it is also very important to regulate the phosphorylation of Interferon regulatory factor 3(IRF3)by up-regulating TANK binding kinase 1(TBK1).The phosphorylation of TBK1 activates IRF3,which can eventually produce type ? interferon,which activates the antiviral effect of cells.In this experiment,we firstly used cell morphology and flow cytometry to determine that PEDV can induce a deeper degree of exogenous apoptosis in porcine jejunal epithelial cells IPEC-DQ cells,and Western Blot technology was used to detect it.After PEDV infection,the expression of Caspase-related proteins in the apoptosis signaling pathway of IPEC-DQ cells indicates that PEDV can induce Caspase-8-mediated apoptosis in IPEC-DQ cells.Secondly,the cleavage of CYLD protein under the conditions of different infection concentrations and different infection time was tested to confirm that the cleavage of CYLD is caused when apoptosis occurs.At the same time,the cellular natural immune pathway proteins(RIG-I,IKK,I?B,p65,TBK1),IRF3)phosphorylation level,it was found that after PEDV infects IPEC-DQ cells,the cascade reaction eventually leads to the up-regulation of the phosphorylation level of key natural immune proteins NF-?B p65 and IRF3.In summary,PEDV infects IPEC-DQ cells and induces caspase-8-mediated exogenous apoptosis in the cells,activates the innate immune RIG-I-like receptor pathway and NF-?B pathway,and produces type ? interferon.The addition of the caspase-8 inhibitor Z-VAD blocked the activity of the NF-?B and RIG-I-like receptor pathways,providing a new idea for the prevention and treatment of PEDV.