Protective Effect And Mechanism Of Compound Chinese Herbal Medicine On IBDV-induced Immunosuppression In Green Foot Chicken

To study the protective effect and mechanism of two different concentrations of compound Chinese herbal medicine on IBDV-induced immunosuppression in green foot chicken.28-day-old green foot chicken without bursal vaccine were selected to infect IBDV strains by nose dropping and eye dropping after one week of adaptive feeding.They were randomly divided into 8 groups.The control group and the attack group were fed with basic diet.The experimental group A-1,A-2 and A-3 were fed with 0.5%,1.0%and 1.5%of Chinese herbal medicines in the basic diet after the attack.The experimental group B-1,B-2 and B-3 were fed with 0.5%,1.0%and 1.5%of Chinese herbal medicines in the basic diet after the attack,respectively.The experimental period was 7 days.On the 3rd and 7th day after infection,6 animals were randomly selected from each group for blood sampling.Histology,immunohistochemistry,real-time fluorescence quantitative PCR,Western-blot and ELISA were used to detect the indexes of immune organs in green foot chicken.The results are as follows:1.Effect of Compound Chinese Herbal Medicine on Serum Immunoglobulin,Antibody to IBDV and Cytokines of Immune Organs in green foot chicken Infected with IBDVCompound Chinese herbal preparations can significantly improve the effects of IBDV on immunoglobulin,IBDV antibody and cytokines of immune organs in green foot chicken.On the 3rd day after IBDV infection,the levels of IgA,IgG and IgM in the serum of the challenge group were significantly lower than those of the control group(P<0.05),and the level of IBDV antibody in the serum was significantly higher(P<0.05).The serum IgA and IgG levels in group B-1 were significantly higher than those in group B(P<0.05).Seven days after IBDV infection,compared with the control group,the serum IgA and IgG levels in the challenge group were significantly lower(P<0.05),and the serum IBDV antibody levels were significantly higher(P<0.05).The levels of IgA,IgG and IgM in serum and IBDV antibody in the test groups were higher than those in the control group,but there was no significant difference(P>0.05).Compared with the control group,the contents of IL-2 and INF-gamma in bursa of Fabricius,INF-gamma in thymus,IL-4 and INF-gamma in spleen were significantly decreased in the treatment group 3 days after IBDV infection(P<0.05).The contents of INF-gamma in bursa of Fabricius increased significantly in group A-1(P<0.05),IL-4 in bursa of Fabricius and thymus in group A-2(P<0.05),IL-4 in bursa of Fabricius and thymus in group B-1(P<0.05),and IL-4 in bursa of Fabricius and thymus in group B-3(P<0.05).At 7 days after IBDV infection,compared with the control group,the contents of IL-2 and INF-gamma in bursa of Fabricius and IL-2 in spleen were significantly decreased in the treatment group(P<0.05).The contents of INF-gamma in bursa of Fabricius increased significantly in group A-2(P<0.05);the contents of IL-2 in bursa of Fabricius and INF-gamma in spleen increased significantly in group A-3(P<0.05);the contents of INF-gamma in bursa of Fabricius and IL-4 in thymus increased significantly in group B-1(P<0.05);the contents of IL-2 in bursa of Fabricius and thymus increased significantly in group B-2(P<0.05);and the contents of IL-2 in bursa of Fabricius increased significantly in group B-3(P<0.05).2.Effect of Compound Chinese Herbal Medicine on the Development of Immune Organs in green foot chicken Infected with IBDVCompound Chinese herbal medicine can significantly reduce IBDV damage to immune organs of green foot chicken.On the 3rd day after IBDV infection,compared with the control group,the bursa index,spleen weight and index increased significantly(P<0.05),and the area of bursa lobule,thymus cortex and spleen nodule decreased significantly(P<0.05).Compared with the control group,the weight and index of bursa of Fabricius,spleen weight and index of A-1 group,spleen index of A-2 group decreased significantly(P<0.05),the area of bursa lobules and splenic nodules of A-2 and B-1 group increased significantly(P<0.05),and the thickness of thymic cortex of A-3 group increased significantly(P<0.05).On the 7th day after IBDV infection,compared with the control group,the weight and index of bursa of Fabricius decreased significantly(P<0.05),while the weight and index of spleen increased significantly(P<0.05),the area of bursa of Fabricius decreased significantly(P<0.05),and the area of splenic nodule and lymphatic sheath around splenic artery increased significantly(P<0.05).The spleen index of group A-1 was significantly lower than that of group A-1(P<0.05),bursa weight and index of group A-2 were significantly higher(P<0.05),bursa index of group A-3 and B-1 was significantly higher(P<0.05),spleen index of group A-3,B-1 and B-2 was significantly lower(P<0.05),bursa lobule area of group A-2 and B-1 was significantly higher(P<0.05),thymus cortex of group B-1 was significantly higher than that of Attack group(P<0.05)。2.Effect of Compound Chinese Herbal Medicine on the Development of Immune Organs in green foot chicken Infected with IBDVCompound Chinese herbal medicine can significantly reduce IBDV damage to immune organs of green foot chicken.On the 3rd day after IBDV infection,compared with the control group,the bursa index,spleen weight and index increased significantly(P<0.05),and the area of bursa lobule,thymus cortex and spleen nodule decreased significantly(P<0.05).Compared with the control group,the weight and index of bursa of Fabricius,spleen weight and index of A-1 group,spleen index of A-2 group decreased significantly(P<0.05),the area of bursa lobules and splenic nodules of A-2 and B-1 group increased significantly(P<0.05),and the thickness of thymic cortex of A-3 group increased significantly(P<0.05).On the 7th day after IBDV infection,compared with the control group,the weight and index of bursa of Fabricius decreased significantly(P<0.05),while the weight and index of spleen increased significantly(P<0.05),the area of bursa of Fabricius decreased significantly(P<0.05),and the area of splenic nodule and lymphatic sheath around splenic artery increased significantly(P<0.05).The spleen index of group A-1 was significantly lower than that of group A-1(P<0.05),bursa weight and index of group A-2 were significantly higher(P<0.05),bursa index of group A-3 and B-1 was significantly higher(P<0.05),spleen index of group A-3,B-1 and B-2 was significantly lower(P<0.05),bursa lobule area of group A-2 and B-1 was significantly higher(P<0.05),thymus cortex of group B-1 was significantly higher than that of Attack group(P<0.05)。3.Effect of Compound Chinese Herbal Medicine on Antioxidant Function of Immune Organs in green foot chicken Infected with IBDVCompound Chinese herbal medicine can significantly improve the anti-oxidative effect of IBDV on immune organs of green foot chicken.Three days after IBDV infection,compared with the control group,T-AOC activity,T-SOD activity and GSH-Px content in bursa of Fabricius,T-AOC activity in thymus and GSH-Px content decreased significantly(P<0.05),while MDA content in bursa of Fabricius,thymus and spleen increased significantly(P<0.05);MDA content in bursa of A-1 group decreased significantly(P<0.05);MDA content in bursa of thymus and bursa of A-2 group decreased significantly(P<0.05).T-SOD activity in bursa of Fabricius,GSH-Px content in thymus,T-SOD activity in spleen and GSH-Px content were significantly increased(P<0.05);T-AOC activity in thymus was significantly increased in group A-3(P<0.05);MDA content in bursa of Fabricius,thymus and spleen was significantly decreased in group B-1(P<0.05);T-AOC activity in bursa and thymus,T-SOD activity and GSH-Px content in spleen were significantly increased in group B-3(P<0.05).The activity increased significantly(P<0.05).Compared with the control group,the MDA content in bursa of Fabricius and thymus increased significantly(P<0.05),the T-AOC activity and GSH-Px content in bursa of Fabricius and the GSH-Px content in thymus and spleen decreased significantly(P<0.05),the GSH-Px content in bursa of Fabricius in group A-1 increased significantly(P<0.05),the MDA content in bursa of Fabricius and thymus decreased significantly(P<0.05),and the MDA content in group A-2 decreased significantly(P<0.05).T-SOD activity,GSH-Px content in thymus,T-SOD activity in spleen and GSH-Px content in thymus increased significantly(P<0.05);MDA content in bursa of Fabricius decreased significantly in group A-3(P<0.05),GSH-Px content in thymus increased significantly(P<0.05);MDA content in bursa of Fabricius and thymus decreased significantly in group B-1(P<0.05),GSH-Px content in bursa of Fabricius and T-SOD activity in thymus increased significantly(P<0.05).The activity of T-SOD and the content of GSH-Px in spleen of group B-3 increased significantly(P<0.05).4.Effect of Compound Chinese Herbal Medicine on Apoptosis of Immune Organs in green foot chicken Infected with IBDVCompound Chinese herbal medicine can promote the apoptosis of immune organ cells of broiler chickens infected with IBDV.At 3 days after IBDV infection,compared with the control group,the expression of Caspase-3 in bursa of Fabricius and thymus and the number of Caspase-3 positive cells were significantly decreased in the challenge group(P<0.05),and the number of Caspase-3 positive cells in spleen was significantly decreased in the challenge group(P<0.05).The expression of Caspase-3 in bursa of Fabricius and the number of Caspase-3 positive cells in thymus in group A-1 were significantly higher than those in group A-1(P<0.05);the expression of Caspase-3 in bursa of Fabricius,Caspase-3 in spleen and Caspase-3 positive cells in group A-2 were significantly higher than those in group B-1(P<0.05);the expression of Caspase-3 in bursa of Fabricius and the number of positive cells,Caspase-3 in thymus and protein in group B-1 were significantly higher than those in group B-1(P<0.05),The number of positive cells increased significantly(P<0.05),and the number of Caspase-3 positive cells in B-3 thymus increased significantly(P<0.05).7 days after IBDV infection,compared with the control group,Caspase-3 protein expression and expression in bursa of Fabricius,Caspase-3 positive cells and thymic Caspase-3 positive cells were significantly decreased in the challenge group(P<0.05);Caspase-3 protein expression and positive cells in bursa of Fabricius in group A-2 were significantly increased(P<0.05);Caspase-3 positive cells in thymus in group B-2 were significantly increased(P<0.05);Caspase-3 positive cells in group B-3 were significantly increased(P<0.05).B-3.The expression of 3 mRNA was significantly decreased(P<0.05).5.Effect of Compound Chinese Herbal Medicine on TLR3/NFκB Signaling Pathway in Immune Organs of green foot chicken Infected with IBDVThe protective effect of compound Chinese herbal medicine on immune organs of green foot chicken infected with IBDV is related to TLR3/NFκB signaling pathway.After 3 days of IBDV infection,the expression of TLR3/κB in bursa of Fabricius and spleen of the challenge group was significantly higher than that of the control group(P<0.05).The expression of TLR3 and NFκB in bursa of Fabricius in group A-1 was significantly lower than that in group A-1(P<0.05);the expression of TLR3/NFκB in bursa of Fabricius in group A-2 was significantly lower than that in group B-1(P<0.05);the expression of TLR3 and TLR3/NFκB in bursa of Fabricius in group B-1 was significantly lower than that in group B-1(P<0.05).After 7 days of IBDV infection,compared with the control group,the expression of TLR3 in bursa of Fabricius and spleen increased significantly(P<0.05)and the expression of TLR3/NFκB in thymus increased significantly(P<0.05).The expression of TLR3 protein in spleen of group A-1 was significantly lower than that of group A(P<0.05).Conclusion:IBDV can obviously destroy the blood index,immune organ quality and index,tissue structure,antioxidant and immune function of green foot chicken,leading to immune suppression.Adding compound Chinese herbal medicines to the feed after challenge can significantly improve the injury of IBDV to the body,of which the best concentration is 1.0%A and 0.5%B.The results also showed that IBDV infection was related to TLR3/NFκB,and the protective effect of Chinese herbal medicine on IBDV green foot chicken was also related to TLR3/NFκB.Therefore,this compound Chinese herbal medicine can be used as a feed additive to provide theoretical reference for the application of Chinese herbal medicine in the prevention and treatment of chicken infectious bursal disease and other livestock and poultry related diseases.