Protective Effects Of Tea Polyphenols For Weaned Pigs Challenged With Oxidative Stress

Oxidative stress reduced growth performance and impaired immunity of piglets, further lead to depressed husbandry efficiency. The present researches were conducted to investigate whether tea polyphenol (TP) could relieve or eliminate oxidative stress suffered by piglets, and to research the underlying mechanisms from both in vitro and in vivo experiments, which may support evidence as well as new study direction for relieving oxidative stress that piglets had suffered. The main results were listed as following:Exp.1 Oxidative stress induced by H2O2 in piglets lymphocyte cultured in vitroThe present trial was conducted to study the effects of medium H2O2 levels on antioxidant enzyme activity and MDA contents of weaned piglets lymphocyte cultured in vitro, and to establish an oxidative stress lymphocyte model with H2O2 as stressor. In vitro, piglets lymphocytes in five treatments, with 6 replicates per treatment, were grown in media containing 0,200,400,600 and 800μM H2O2, respectively. Results showed that H2O2 dramatically decreased the activities of SOD and GSH-Px in piglets lymphocytes, meanwhile, significantly elevated lymphocytes MDA contents. When the concentration of H2O2 was 400μM, the lymphocytes cultured in vitro began to be challenged with apparent oxidative stress.Exp.2 Anti-oxidative stress of EGCG on piglets lymphocytes under oxidative stressIn this experiment, using the in vitro oxidative stress model established in experiment 1, the effects of EGCG on antioxidant enzyme activity and immune function of piglets lymphocytes under oxidative stress were determined. A 2x4 factorial design was conducted, with 6 replicates per treatment, lymphocytes under oxidative stress(400μM H2O2) or not(0μM H2O2) each was cultured in medium containing 0,20,40 and 80u.g/ml EGCG, respectively. The results showed that 400μM H2O2 significantly reduced the activities of SOD and GSH-Px in weaned piglets lymphocytes, improved lymphocytes MDA levels, lower the lymphocyte transformation rate, increased the medium concentrations of IL-2, TNF-aand IFN-y, whereas, decreased that of IL-4, promoted the mRNA expression of NF-kB, FOS and JUN in lymphocytes. Under normal circumstance, EGCG could elevate the activities of SOD and GSH-Px in piglets lymphocytes cultured in vitro, decrease lymphocytes MDA levels, improve the lymphocyte transformation rate, increase the medium concentrations of IL-2 and IL-4, whereas, lower that of TNF-αand IFN-γas well as the mRNA expression of NF-kB, FOS and JUN in lymphocytes. Under oxidative stress, EGCG could improve the activities of SOD and GSH-Px in piglets lymphocytes, down-regulate lymphocytes MDA levels, elevate the lymphocyte transformation rate, increased the medium concentrations of IL-2 and IL-4, whereas, inhibit that of TNF-αand IFN-γas well as the mRNA expression of NF-kB, FOS and JUN in lymphocytes.Those data demonstrated that 400μM H2O2 would induce oxidative stress, thus seriously impaired lymphocytes transformation rate, cytokine secretion and immune function. However, EGCG, by inhibiting the expression of NF-kB, FOS and JUN genes, could improve lymphocytes antioxidant enzyme activity, promote lymphocyte transformation and cytokine secretion, and therefore relieve the negative effects of oxidative stress on lymphocytes.Exp.3 Effects of TP on growth performance and immune function of piglets under oxidative stressTo gain insights into the effects of TP on growth performance, nutrients digestibility, serum redox state, immunity function and tissue structural integrity of piglets under oxidative stress, an oxidative stress model was established by intraperitoneally injecting with diquat of weaned piglets. Thirty-six 28-day-old Duroc×(Landrace×Yorkshire) piglets were randomly distributed into three groups with twelve piglets per group. Piglets in three treatments were fed with formulated diets containing 0,500 and 1000mg/kg TP, respectively. After 14d, half of piglets in each group were challenged with 10mg/kg diquat, the others were injected with the same volume of sterilized physiological saline. Digestion trial was carried out from 17 to 19d. Piglets were housed in single cage, and the whole experiment lasted for 21d.The results showed that diquat impaired growth performance of piglets, ADG (P＜0.01) and ADFI(P<0.01) reduced significantly and F/G (P<0.01) enhanced significantly. Nutrients (energy, crude protein and dry matter) digestibility was not affected. Serum SOD, GSH-Px, CAT and T-AOC activity were down-regulated by diquate injection, whereas, serum MDA level was enhanced. Diquate injection impressed humoral immunity, decreased serum IgG, IgA and IgM contents, moreover, diquate injection reduced serum CD4+level, elevated that of CD8+, decrease CD4+/CD8+ratios, but did not change serum CD3+level. Diquate injection increased serum IL-2, IFN-yand TNF-aconcentrations, lower that of IL-4, and impaired tissue structure of liver, spleen and lymphaden.Under normal circumstance, growth performance and nutrients digestibility of piglets, serum CAT, T-AOC, MDA, IgG, IgA, IgM, CD3+, CD8+, IFN-yand TNF-alevels and tissues structural integrity were not affected by TP-supplementation. However, TP elevated serum SOD and GSH-Px activities, serum CD4+, IL-2 and IL-4 levels as well as CD4+/CD8+ratios.Under oxidative stress, TP could improve growth performance of piglets, but had no significantly effect on nutrients (energy, crude protein and dry matter) digestibility. Feeding TP-supplemented diets increased serum SOD, GSH-Px and CAT activities and T-AOC, reduced serum MDA concentration, thus relieve oxidative stress. TP played an important role in both cellular immunity and humoral immunity by affecting serum antibodies and cytokine levels. The tissue injuries caused by oxidative stress could also be alleviated by TP.Those data indicated that piglets challenged with 10mg/kg BW diquate showed poor growth performance, lower serum antioxidant enzyme activity and higher serum MDA level, which suggested an oxidative stress. Additionally, diquate injection reduced serum immunoglobin contents, changed lymphocyte subsets, upset cytokine secretion and did harm to tissue structure integrity of liver, spleen and lymphaden, finally impaired cellular immunity and humoral immunity. Under normal condition, TP had no dramatically effect on growth performance, apparent nutrient digestibility, serum immunoglobin contents and tissue structure, whereas, long term intake of TP resulted in better antioxidant ability and immunity of piglets by elevating serum antioxidant enzyme activity, serum CD4+contents and CD4+/CD8+ratios, and by promoting secretion of IL-2 and IL-4. Moreover, feeding TP-supplemented diet could help relieve the negative effects of oxidative stress on growth performance, immunity and tissue structural integrity of piglets in specified extent.Exp.4 Protecting effects of TP on piglets from oxidative stressThis present trial was conducted to investigate the underlying mechanism by which TP protecting growth performance and immunity of piglets from oxidative stress. The effects of TP on the excretion of corticotrophin (ACTH) and glucocorticoids (GC) and the mRNA expression of NF-kB, FOS and JUN in piglets under oxidative stress were investigated. The materials and procedure in this experiment was the same as experiment 3. The results showed that serum ACTH and GC concentrations of piglets were increased under oxidative stress induced by diquat. TP had no significant effect on serum ACTH and GC concentrations of piglets under normal condition, however, intake of TP could reduce serum ACTH and GC secretion and toward to normal levels. The mRNA expression of NF-kB, JUN and FOS of liver, spleen and lymphaden in piglets were higher under oxidative stress, and feeding TP-supplemented diets could recover the detrimental expression of NF-kB, JUN and FOS.Those data indicated that inhibiting the synthesis and secretion of ACTH and GC in HPA as well as decreasing the expression of NF-kB, JUN and FOS in liver, spleen and lymphaden were the underlying mechanisms by which TP protected growth performance, immunity and tissue structural integrity of piglets from oxidative stress.ConclusionsThese results demonstrated that oxidative stress reduce growth performance, serum antioxidant enzyme activity and immunoglobin contents, moreover, upset lymphocyte subsets and cytokine secretion and finally impair immunity and tissue structure integrity of piglets. A long term intake of TP could relieve growth depression, improve anti-oxidant ability, circle immunoglobin contents and lymphocyte subsets differentiation of piglets, also promote some cytokine excretion, finally relieve the detrimental effects caused by oxidative stress. TP supplementation had beneficial effects in piglets induced by oxidative stress through regulating ACTH and GC section in HPA endocrine and decreasing the expression of NF-kB, JUN and FOS in liver, spleen and lymphaden.