Adjuvant Activity And Application Of Antimicrobial Peptide BSN-37

With the development of large-scale and industrialized livestock breeding,some infectious diseases will occur in the breeding process.With the increasingly strict policy situation of banning and substituting antibiotics,from an economic point of view,vaccination with inactivated vaccines has gradually become an effective means of current prevention.Inactivated vaccines are safe and widely used,but they have defects in immune protection period and immunogenicity,and more and more attention has been paid to continuously seeking green safe,non-toxic or low-toxic,efficient adjuvants.Antimicrobial peptides（AMPs）,as peptides derived from animals,plants and microorganisms,have the advantages of small molecular weight,wide antibacterial spectrum,good thermal stability,difficult to produce drug resistance,and enhanced immunity etc.With the advancement of research,adjuvant activity of antimicrobial peptides has received much attention.The purpose of this study was to investigate the regulatory effect of antimicrobial peptide BSN-37 on immune function in vivo and in vitro and its adjuvant activity on trivalent inactivated piglet Escherichia coli vaccine,in order to fully utilize and develop antimicrobial peptides resources,and provide an important basis for seeking safe and effective adjuvants for new veterinary vaccine.1.Different doses of antimicrobial peptide BSN-37 were added to stimulate BMDC and RAW264.7 in vitro,and CCK-8 was used to detect cell proliferation after BSN-37 stimulation;Western blot was detected using costimulatory molecules CD80,CD40,MHC I,and MHC II expression;ELISA was used to detect the secretion levels of cytokines IL-2,IL-4,IFN-γ and IL-10 in the cell culture supernatant.The results showed that after the antimicrobial peptide BSN-37 treated BMDC and RAW264.7 cells,there was no significant change in cell morphology and no cytotoxicity.In vitro,it promoted the expression of costimulatory molecules CD80,CD40,MHC I and MHC II,and promoted the Th1-type cytokine IL-2 and IFN-γ,Th2-type cytokines IL-10 and IL-4.The results show that antimicrobial peptide BSN-37 has no cytotoxicity,can promote cellular immune response,and has immune adjuvant activity.2.Babl/c mice were immunized with different doses of antimicrobial peptide BSN-37 combined with OVA model antigen,CCK-8 was used to detect the effect of BSN-37 on OVA-induced splenocyte proliferation stimulation index;ELISA was used to detect the cells in the supernatant of splenocyte culture Factors IL-2,IL-4,IFN-γ,IL-10 secretion levels and serum Ig G,Ig G1,Ig G2 a antibody content.The results showed that antimicrobial peptide BSN-37 had no effect on the growth of mice and had good safety;The secretion levels of cytokines（including IL-2,IL-10,IL-4 and IFN-γ）and serum levels of Ig G,Ig G1,The content of Ig G2 a antibody was significantly increased;The proliferation and activation ability of mouse spleen lymphocytes were enhanced.Thus,the antimicrobial peptide BSN-37 induced a higher level of immune response in the body.3.Using Escherichia coli C83549,C83917,and C83644 as alternative vaccine strains,a trivalent inactivated vaccine for piglet Escherichia coli was prepared.Different doses of antimicrobial peptide BSN-37 combined with trivalent inactivated vaccine were subcutaneously injected to immunize mice,and the optimal dose of antimicrobial peptide BSN-37 was determined to be 25 μg.With an increase in the content of specific Ig G,Ig G1,and Ig G2 a antibodies in the serum,and the proliferation ability of spleen lymphocytes increased.It shows that antimicrobial peptide BSN-37 can significantly enhance the immune response of mice,which proves adjuvant activity of the antimicrobial peptide BSN-37.The optimal dose of antimicrobial peptide BSN-37 combined with different doses of trivalent inactivated vaccine subcutaneously to immunize mice,and the optimal compatible dose of trivalent inactivated vaccine was determined to be 0.3 m L.The content of Ig G,Ig G1,Ig G2 a antibodies and cytokines IL-2,IL-10,IL-4,IFN-γ in serum were increased,promoting the proliferation ability of splenocyte lymphocytes.It shows that antimicrobial peptide BSN-37 can significantly improve the immune function of mice,confirms the adjuvant activity of antimicrobial peptide BSN-37,which provides a basis for the development of antimicrobial peptide BSN-37 as a safe and efficient new immune adjuvant.