Blocking Effect Of Early Pregnancy Factor On Red Meat Neu5Gc-induced Antibody Production In CMAH^-/- Mice

The meat of mammals such as cows,sheep,and pigs are often referred to as red meat and is an essential animal source food in our daily lives.Different eating habits make red meat intake associated with various human diseases,such as cancer,cardiovascular disease,type II diabetes,hyperlipidemia and hypertension.There is a large amount of evidence to confirm that prolonged consumption of red meat and its products may increase the risk of developing tumors and cardiovascular diseases in the human body.In recent years,studies have found that there is a potential carcinogen in red meat,N-hydroxyacetylneuraminic acid(Neu5Gc).As a special antigen,it can be modified on the surface of human cells through various ways to activate the immune system and produce antibodies.The sustained interaction between this antibody and antigen triggers chronic inflammatory reactions,further leading to the occurrence and development of tumors and cancer.From this,it can be seen that in the process of Neu5Gc inducing chronic inflammation,the production of antibodies is the key point that triggers the disease,and the production of antibodies depends on the auxiliary effect of T cells.If T cell activity is inhibited,it will have a certain impact on the activation of B cells and the production of antibodies.If the key pathogenic link of blocking antibody production can be achieved,reducing the chronic inflammation caused by the continuous action of antigens and antibodies,It is expected to solve the meat safety problem caused by red meat Neu5Gc.It has been found that early pregnancy factor(EPF)can reduce the expression and secretion of CD4~+T cell adhesion molecules,down regulate the activity of T cells,and prevent the fetus from being cleared as a foreign body in the mother.It is considered to be a mild immunosuppressive factor.It was first discovered by Australian researcher Morton in the serum of pregnant mice and is currently one of the earliest biochemical indicators to confirm maternal pregnancy.Currently,it has been widely used for early pregnancy diagnosis,fetal testing,and the treatment of autoimmune diseases.Given the unique immunosuppressive function of EPF and the mutual activation relationship between T/B cells in antibody generation,this study aims to explore whether EPF can inhibit Th cell activation and hinder B cell antibody synthesis,thereby reducing chronic inflammation caused by the interaction between antibodies and Neu5Gc.This study first cloned the nucleic acid sequence encoding the EPF protein,induced the expression of the sumo EPF recombinant protein(srEPF)and the EPF(10 k D)protein that removed the inhibitory region through a prokaryotic expression system,and purified the EPF antibody;And verify its activity at the cellular level through rosette inhibition test;To investigate the effect of EPF on Neu5Gc induced chronic inflammation in mice with CMAH gene deletion as a simulated human model animal.The results showed that srEPF and EPF(10 k D)proteins were successfully expressed and purified,and high purity and high specificity of EPF antibodies were obtained.EPF exerted a better immunosuppressive effect by inhibiting the formation rate of rosette and its binding form in vitro.The animal model mediated by Neu5Gc in red meat was established,and EPF intervention could alleviate the slow weight gain and shortening of colon length in mice.ELISA results showed that injection of EPF significantly reduced the level of anti-Neu5Gc antibody,the contents of inflammatory factors IL-6,IL-1β,TNF-αand MPO activity in vivo.HE staining showed that the pathological changes of liver and intestinal tissues were alleviated after EPF treatment.Immunohistochemical observation of CD4 cells in mice showed that the number of CD4 cells in EPF group was significantly lower than that in inflammation promotion group,and fluorescence quantitative results showed that the number of B cell activation molecules CD80 and CD86 molecules in mice after EPF treatment was decreased,indicating that EPF may inhibit CD4 lymphocytes.Intervention of B cell activation signal transmission,and then affect the production of anti-Neu5Gc antibody in vivo,reduce the body inflammation caused by red meat Neu5Gc.In summary,the EPF antigen prepared in this experiment has an inhibitory effect on the production of antibodies caused by red meat Neu5Gc,which can reduce the level of inflammation in mice and alleviate the inflammation caused by Neu5Gc.This provides a new research approach and effective solution for exploring and solving the safety hazards caused by long-term consumption of red meat.