Immune Response Of B Cells Induced By Porcine Epidemic Diarrhea Virus

Porcine epidemic diarrhea(PED)is an acute,highly contacted contagious intestinal infectious disease of swine,caused by porcine epidemic diarrhea virus(PEDV).Diarrhea,vomiting,acute enteritis is major characteristic of PED.PEDV is one of the major causes of diarrhea and death in neonnate piglets.Mature primary B cells recognize antigens,activate,proliferate,and differentiate into plasma cells,synthesize and secrete antibodies to clear pathogens.At present,the mechanism of B cell immune response induced by PEDV is still unclear.This study explored the immune response of B cells from pigs in different ages to PEDV virulent and attenuated strains in vitro in order to establish the foundation for clarifying the immune mechanism of PEDV infection.In order to clarify the activation of PEDV on B cells,the anterior vena cava blood of neonate piglets,weaned pigs and adult sows were collected separately,and CD21~+B cells were sorted from peripheral blood by density gradient centrifugation combined with immunomagnetic beads sorting cell method.PEDV virulent strain QY2016 and attenuated strain CV777 were inoculated on B cells of neonate piglets,weaned pigs and adult sows in vitro.Expression levels of B cell surface molecules MHC II,CD80,CD54,CD27 and mIgM were measured by Flow cytometry at 24 h after infection.The virulent strains QY2016 and CV777 could not induce the increase of MHC II~+,CD80~+and mIgM~+B cells in pig B cells at each growth stage,but it could cause the content of CD54~+and CD27~+B cells in B cells of neonatal piglets increase significantly.The content of B cells expressing MHC II~+,CD80~+,mIgM~+and mIgM~+CD27~+in B cells inoculated with virulent strain QY2016 was significantly lower than that of B cells inoculated with attenuated strain CV777.ELISA was used to detect the expression levels of IL-1,IL-22,IL-4,IL-6,IL-10,IFN-?,GM-CSF,M-CSF,IFN-?and TGF-?in the supernatant of B cells from pigs in different ages at 0 h,24 h and 48 h after viral inoculation.Both QY2016 strain and CV777strain could cause IFN-?and GM-CSF secretion from B cells of neonatal piglets,IL-1,IFN-?and IL-4 secretion from B cells from weaned pigs,and TGF-?secretion from sows,whereas could not cause secretion of IL-6 and IL-10 from B cells at each growth stage pigs.The secretion of other cytokines differed between PEDV virulent and attenuated strains and pigs in different ages,but B cells secreted more cytokines in weaned pigs than those in neonate piglets and adult sows.The levels of IgM,IgA and IgG in the supernatant of B cells were determined by ELISA at 0 d,4 d and 7 d after virus infection.The virulent strain QY2016 was only able to induce the secretion of IgM and IgG from the B cells of the weaned pigs,while the attenuated strain CV777 was able to induce secretion of IgM and IgA from the weaned B cells,and the secretion of IgA and IgG from the sow B cells.Both virulent strain and attenuated strain could not induce the secretion of IgM,IgA and IgG from B cells of neonatal piglets.The above results indicate that B cell surface molecule expression,cytokines and antibody responses induced by PEDV are closely related to the ages of pig and the virulence of PEDV strains.Compared with the attenuated strain,the responses of B cells to virulent strains are weak,and the responses of neonatal piglets and sow B cells to PEDV were lower than those of weaned pigs.