The Immunoprotective Effects And Mechanisms Of Polysaccharide Of Atractylodes Macrocephala Koidz On Goslings

Polysaccharide of Atractylodes macrocephala Koidz(PAMK)is polysaccharides extracted from the natural plant.It has been proved to have antibacterial,anti-inflammatory,anti-tumor,anti-oxidation and immune promoted functions,which can be developed as functional feed additives with no residues and no toxicity.However,there is still lack of comprehensive and systematic reports on the immunogenicity of PAMK.It is still unclear whether PAMK has immunoprotective effects on animals with immunosuppression or inflammatory pathological conditions.In addition,whether miRNA is involved in the immunoprotection of PAMK has not been reported.The study of immune regulation using goose is rare.In order to understand the protective effects and mechanisms of PAMK on immune function of goslings,the following five parts of the study were carried out.Part 1: Effects of PAMK on immune function of goslings0,100,200,400,600,800,1200,1600,and 2000 mg/kg PAMK were added into the feed respectively to select the optimal additive amount for goslings.The results showed that the suitable additive amount of PAMK was 400 mg/kg.In order to fully explore the impact of PAMK on the immune function of goslings,the related immune parameters of 400 mg/kg PAMK goslings were further tested.The results showed that PAMK can maintain the normal tissue morphology and ultrastructure of the immune organs of goslings,maintain serum levels of hemolysin,IgM,IgA,and IgG,and significantly increase serum levels of IL-4,IL-6,and IL-10 in goslings.The levels lower serum TGF-? and IFN-? levels.The above results revealed that PAMK improved the serum cytokine levels in goslings and promoted the immune function of goslings.Part 2: The protective effect of PAKM on immune function in immunosuppressed goslingsThis model of cyclophosphamide-induced immunosuppression in goslings was established.The goslings were injected with 20,40,80,and 100 mg/kg·BW cyclophosphamide respectively.The results showed that the suitable dose for cyclophosphamide-induced immune suppression was 40 mg/kg·BW.In order to explore whether PAMK can protect immune function of goslings under immunosuppression,cyclophosphamide was injected at the same time as the goslings were fed with PAMK to construct an immunosuppressive model.After comparative analysis found that PAMK alleviated the damage of the immune organ tissue morphology and ultramicrostructure made the immunity index to the level of healthy goslings.The above results show that the PAMK resisted the immune damage caused by cyclophosphamide,and it has a significant protective effect on the immune organs,humoral immunity and cellular immunity of goslings.Part 3: MiRNA sequencing to study the protected mechanism of PAMK on immunosuppressed goslingsThe results of the previous study indicate that PAMK had important protective effects on immunosuppressed goslings.This study intended to find out the miRNA immunoprotective mechanism of PAMK on immunosuppressed goslings.The thymus of the goslings of Control,PAMK,CTX and CTX+PAMK groups were selected for miRNA high-throughput sequencing.The protective mechanism of PAMK on immunosuppressed goslings was: novel_mir2 inhibited the expression of CTLA4,and then activated TCR signaling pathway,ultimately promoted the expression of transcription factors NFAT and AP-1.Part 4: Effectss of PAMK on intestinal mucosal immunity of LPS treated goslingsLPS treatment was used to establish a model of intestinal inflammation in goslings.Goslings were injected intraperitoneally with LPS injections of 0.5,1,2 mg/kg·BW.The results showed that the suitable dose of LPS to induce small intestine inflammation in goslings was 2 mg/kg·BW.To explore whether PAMK could protect the intestinal mucosal immunity of goslings induced by LPS,control,PAMK,LPS,PAMK and LPS co-treatment groups were used.The results showed that PAMK significantly alleviated the up-regulation of serum endotoxin,CRP,and proinflammatory cytokines caused by LPS,reduced tissue damage in the intestine,restore the normal number of IgA-secreting cells,and alleviated abnormal mRNA expression of TLR4,occludin,and ZO-1.The above results indicated that: PAMK significantly alleviated the symptoms of LPS-induced goose inflammation and protects intestinal mucosal immune function in terms of immune barriers and mechanical barriers.Part 5: Effects of PAMK on the intestinal microflora of goslings treated with LPSIn order to further study the mechanism of action of intestinal barrier in intestinal mucosal immunity in terms of biological barriers,the feces of control,PAMK,LPS,PAMK and LPS co-treatment groups were used for 16 S rDNA sequencing.The intestinal microbiota diversity and the changes of the microbial species in groups were analyzed.The results showed that the species abundance and diversity of intestinal microflora in the LPS group significantly increased,the number of Romboutia decreased significantly,and the composition of the intestinal microbiota was unbalanced.However,the species abundance and diversity of gut microbiota in the co-treatment group of PAMK and LPS were similar to those in the control group and the PAMK group,and there was no significant difference in the genus level.The above results show that: PAMK improved the imbalance of intestinal flora caused by LPS by maintaining the relative stability of Romboutsia.In summary,the results of this study show that PAMK may activate TCR signaling pathway by up-regulating expression of novel_mir2,targeting inhibition of CTLA4,and exerting immunoprotective effect on immunosuppressed goslings.At the same time,PAMK protected the intestinal mucosal immune function of enteritis goslings by improving the intestinal mechanical barrier,immune barrier and biological barrier function.The above research results fully reveal the immunoprotective effect and mechanism of PAMK on goslings,which provides an important basis for the further development of PAMK as a green natural gosling feed additive.