The Study Of The Effects Of Yupingfeng Polysaccharide Liposomes On Immune System Of Chickens

It was demonstrated that Yupingfeng polysaccharides(YPF-P)enhanced the immune-modulating activity significantly.The preparation of YPF-P has been performed and the components of YPF-P were separated and identified by our research group in the preliminary experiment.In order to explore the medicinal value and expand the range of clinical application,a novel formulation of YPF-P was prepared by reverse evaporation method,which was named Yupingfeng polysaccharide Liposomes(YPF-PL).The previous results show that YPF-PL was high encapsulation rate and superior slow-release effect.Combining in vivo experiments with in vitro cell experiments,the effects of YPF-PL and YPF-P on the immune system of target animal(chickens)were compared by tissue sections,immunohistochemistry,immunofluorescence,flow cytometry and ELISA.The effects of YPF-PL on the central immune organs,peripheral immune organs and mucus-related lymphocyte tissues of chicks were discussed.The immunomodulatory effects of the YPF-PL were studied by the morphological structure and immune function.180 13-days-old chickens were randomly divided into 6 groups,including negative group(B group),Immune group(I group),YPF-P group(YPF-P group200mg/Kg),Low,Medium and High dose YPF-PL groups(YPF-PL_Lgroup100mg/Kg;YPF-PL_Mgroup 200mg/Kg;YPF-PL_Hgroup 400mg/Kg).The drug was administered twice continuous,each for 7 days.The chickens were immunized with H5N1 avian influenza inactivated vaccine during the period of time;10 chickens of each group were execute randomly on 42d.The central immune organ(bursa of Fabricius),peripheral immune organs(spleen)and mucosal associated lymphoid tissue(lung and jejunal structures)were sampled.Then the conventional paraffin sections,H&E staining and image software were applied to analyze the change of the organization structure.The expression of IgA~+,IgM~+,IgG~+B lymphocytes and activepentapeptides(Bursopentine,BP5)of bursa were detected by immunohistochemistry and immunofluorescence staining.Spleen lymphocyte suspension was isolated and prepared in vitro,the proliferative activity and the transformation capacity with Con A were determined by MTT assay.Besides,the changes in the proportion of Bu-1~+B cells,CD3~+,CD4~+and CD8~+T lymphocyte subsets in spleen lymphocytes were assayed by Flow cytometry.Finally,secretory SIgA of the lung and jejunum were detected by ELISA,and the change of the number of GC,IEL and LPL in jejunum were determined by image analysis.The results showed that the effects of YPF-PL on the tissue structure of the immune system in chickens were exhibited as following:1.the ratio of the medulla to the cortex in the bursa of farrar and the volume of lymphatic nodules(follicles)and lobules(plica)of bursa were increased significantly;2.the longitudinal section of the white pulp of the spleen was widen,and the number of white and red medulla lymphocytes were increased obviously;3.the lung tissue was stained deeply;the lymphocytes of the lung was diffusedly increased;the wall full of many lymphocytes of the lung was obviously thickened;broncho-associated lymphoid tissues(BALT)increased in size;the phenomenon of lymphocyte adhesion and migration in the inner wall of blood vessels increased,which promoted the homing of lymphocytes in lung tissues.4.the growth of jejunal villi was significantly promote;the V/C ratio was improved,the structural integrity of the small intestine was protected,and the jejunal mucosal mechanical immune barrier was strengthened.The results also showed that the effects of YPF-PL on immune system function in chickens were exhibited as following:1.the expression rates of IgM~+,IgA~+and IgG~+B cells in bursa and the expression intensity of BP5 were increased significantly;2.The proliferation of lymphocytes and lymphocyte proliferation and transformation in the chicken spleen were stimulated.Meanwhile,the proportion of Bu-1~+B lymphocytes,CD3~+and CD4~+T lymphocyte subsets and the CD4~+/CD8~+ratio in chicken spleen were increased significantly.3.the number of GC,IEL and LPL cells in jejunum were increased significantly and the secretion of SIgA in jejunum and pulmonary mucosa were promoted markedly.In summary,this study demonstrated that YPF-PL has a significant positive regulation effect on the structure and function of immune system in chickens.