Regulation Mechanism Of Lactobacillus Amylovour On IκB/NF-κB Pathway In Porcine Rotavirus Infection

Probiotics are a group of living microorganisms that are beneficial to the body and can be stably colonized in the intestine.Relevant research shows that probiotics have antiviral and regulating body immunity.Rotavirus(RV)mainly causes viral gastroenteritis,which is an important cause of diarrhea and even death in infants and young animals,and it poses a huge threat to the healthy growth of infants and young children and the development of the breeding industry.The main function of the non-structural protein NSP1 encoded by RV is to help the virus escape the innate immunity of the body in the early stage of RV infection.At present,studies have pointed out that during RV infection,NSP1 can phosphorylate CKII and then compete with IкB to bind β-Trc P to inhibit the activation of NF-кB to avoid the innate immunity of the host.Lactobacillus amylovorus is one of the probiotic lactic acid bacteria.It is known that Lactobacillus amylovorus can effectively reduce RV infection.Both its live bacteria and metabolites can prevent the adhesion and replication of RV in intestinal tissue cells in different directions and different degrees.However,there is no relevant report on whether Lactobacillus amylovorus and its secreted supernatant play a role through the IкB / NF-кB pathway in rotavirus infection,and its impact on NSP1 is also unclear.These all need further study.Accordingly,in this study,Lactobacillus amylovorus and its secreted supernatant were used in the in vivo and in vitro experiments,and the normal group and the RV infection group were set as controls.Mainly explore the regulation of RV NSP1 and IкB / NF-kB under the preventive and therapeutic interventions of Lactobacillus amylovorus and its secreted supernatant.Initially probe the effect of Lactobacillus amylovorus and its secreted supernatant on the production and efficacy of CKII.The experimental design completed the analysis of the anti-RV ability of Lactobacillus amylovorus and its secreted supernatant in vivo and in vitro;the expression,identification and immunogenicity determination of recombinant porcine rotavirus NSP1 protein.The changes of m RNA and protein levels of NSP1,IкB and NF-кB under the preventive and therapeutic intervention of Lactobacillus amylovorus and its secreted supernatant can regulate the m RNA level of CKII.The results of the study showed that in the in vitro test,the content of NSP1 in the Lactobacillus amylovorus Secrete supernatant before inoculation group(S.BI group)was significantly lower than that in the RV group(P<0.05).The level of NSP1 in the Lactobacillus amylovorus Secrete supernatant after inoculation group(S.AI group)was significantly lower than that in the RV group(P<0.05),and there was no significant difference in protein level compared with the RV group(P>0.05).The trends of m RNA levels of IкB and NF-кB were similar.The NF-kB mRNA of S.BI group was significantly different from that of RV group(P <0.05),and it was in a continuously stable up-regulation state.At the protein level,there was a large amount of p-IкB accumulation in the RV group,but the expression of p-NF-кB was very small.The expression level of p-NF-кB in the S.BI group was significantly higher than that in the RV group(P<0.05).In the in vivo test,the Lactobacillus amylovorus before inoculation group(BI group)can significantly increase the level of NF-кB m RNA(P<0.05)within three days of RV infection.The Lactobacillus amylovorus after inoculation group(AI group)is in sharp contrast.The IкB m RNA levels of the RV group,BI group and AI group increased significantly after 1 day of infection with RV and then showed a downward trend(P<0.05),and there was no significant difference on the 7th day compared with the normal group(P>0.05).The differences between the groups are relatively small.Both in vivo and in vitro experiments indicated that Lactobacillus amylovorus and its secreted supernatant could effectively inhibit the up-regulation of CKII m RNA level,which was significantly different from that of RV group(P<0.05).TBB(CKII inhibitor)was used to inhibit CKII in RV group.The results showed that p-NSP1 protein level was not expressed.Compared with the S.BI group,this result corroborates that Lactobacillus amylovorus and its secreted supernatant can reduce the expression of CKII and the phosphorylation level of NSP1,and ensure the effective activation of NF-кB pathway.In summary,Lactobacillus amylovorus and its secreted supernatant are effective in inhibiting RV infection of intestinal tissue cells.They can inhibit the high expression of NSP1,produce biological activity by reducing the content of CKII,inhibit NSP1 phosphorylation,and effectively activate the IкB / NF-кB pathway to prevent RV from evading innate immunity through NSP1.The research provides a certain reference and basis for that the role of probiotics and its secreted products in regulating the body’s innate immune pathways,preventing and reducing viral infections.