Study On The Mitigation Action Of Chitosan On Oxidative Stress Of Weaned Piglets And The Underlying Mechanism

Weaning resulted in oxidative stress in piglets,leading to poor growth performance and health state.The objective of this experiment was to investigate the mitigation action of chitosan on oxidative stress of weaned piglets and the underlying mechanism in vivo and in vitro.The main results were listed as follows:Exp.1 Effects of chitosan on oxidative stress in weaned pigletsThe present experiment was conducted to investigate the effect of chitosan on the antioxidative function in weaned piglets challenged with diquat.A total of 60 weaned piglets（30 males and 30 females）were randomly divided into 5 treatments with 12 piglets each treatment.Piglets were fed with diets containing 0,250,500,1000 and 2000 mg/kg chitosan,respectively.After 14 days,half of piglets in each group were challenged with 10mg diquat per kg body weight.The others were injected with the same volume saline.The experiment lasted for 21 days.Body weight and feed intake were monitored.Blood samples were collected to determine antioxidative and immune parameters.The results showed that diquat induced oxidative stress and inflammatory responses by decreasing the activity of antioxidant and increasing pro-inflammatory cytokines.But dietary chitosan alleviated these negative effects induced by diquat and decreased serum concentrations of pro-inflammatory cytokines but increased activities of antioxidant enzymes and anti-inflammatory cytokines.These results suggested that chitosan attenuated the oxidative stress of piglets caused by diquat injection.Exp.2 Protective effect of chitosan against diquat-induced oxidative stress and the underlying mechanismThis experiment was conducted to investigate the underlying mechanism by which chitosan protected weaned piglets from oxidative stress induced by diquat.A total of 24weaned piglets（12 males and 12 females）were randomly divided into 2 treatments with 12piglets each treatment.Piglets were fed with diets containing 0 and 500 mg/kg chitosan,respectively.After 14 days,half of piglets in each group were challenged with 10 mg/kg diquat.The others were injected with the same volume saline.The experiment lasted for 21days.The results showed that dietary chitosan supplementation alleviated oxidative stress by improving growth performance,crude protein and calcium apparent digestibility,and increasing intestinal villus height and the ratio of villus height to crypt depth.Dietary chitosan increased the activities of SOD,CAT and GPx in serum,liver,spleen and small intestine.And chitosan up-regulated gene expression of CuZn-SOD,Mn-SOD,GPx4 in liver,CuZn-SOD and GPx4 in spleen,CAT,CuZn-SOD,Mn-SOD,GPx4 in duodenum,GPx4 in jejunum and CnZn-SOD,GPx4 in ileum.In addition,chitosan down-regulated the gene expression of p65 in liver as well as c-Fos in duodenum and jejunum.From these results,it was concluded that chitosan enhanced the antioxidative function by inhibiting NF-κB and AP-1 pathways in weaned piglets after diquat challenge.Exp.3 Protective effect of chitosan oligosaccharide against oxidative stress in lymphocytes of pigletsThe present experiment was conducted to investigate the effect of chitosan oligosaccharide on the antioxidative function in piglet’s lymphocytes challenged with H₂O₂.A 2×5 factorial design was conducted,with 6 replicates per group.One factor was oxidative stress treatment（Lymphocytes was challenged with H₂O₂ or not）.The other factor was chitosan oligosaccharide treatment（Culture medium contained 0,40,80,160and 320μg/mL chitosan oligosaccharide;respectively）.The results indicated that oxidative stress induced by H₂O₂ decreased the activities of SOD,CAT and GPx,but increased the level of MDA in lymphocytes.Both under normal and oxidative stress circumstance,chitosan oligosaccharide significantly increased activities of CAT and GPx,and reduced SOD activity and MDA concentration of lymphocytes.Exp.4 Mitigation action of chitosan oligosaccharide on H₂O₂-induced oxidative stress in lymphocytes through regulating NF-κB signaling pathwayThe present experiment was conducted to study the underlying mechanism of chitosan oligosaccharide protecting lymphocytes from oxidative stress.A 2×2×2 factorial design was conducted,including two oxidative stress treatments（with or without H₂O₂）,two chitosan oligosaccharide levels（0 or 160μg/mL chitosan oligosaccharide,the additive dose determined according to Exp.3）,and two PDTC levels（with or without PDTC）.The trial consists of 8 treatments with 6 replicates.The results showed that,under normal circumstance,chitosan oligosaccharide could reduce oxidative damage and elevate antioxidative function through increasing the activities of antioxidant enzymes and up-regulating GPx4 gene expression;under oxidative stress,chitosan oligosaccharide could increase CAT activity and up-regulate gene expression of Mn-SOD and GPx4through regulating NF-κB signaling pathway.Exp.5 Mitigation action of chitosan oligosaccharide on H₂O₂-induced oxidative stress in lymphocytes through regulating AP-1 signaling pathwayThe present experiment was conducted to study the underlying mechanism of chitosan oligosaccharide protecting lymphocytes from oxidative stress.A 2×2×2 factorial design was conducted,including two oxidative stress treatments（with or without H2O2）,two chitosan oligosaccharide levels（0 or 160μg/mL chitosan oligosaccharide,the additive dose determined according to Exp.3）,and two SP600125 levels（with or without SP600125）.The trial consists of 8 treatments with 6 replicates.The results showed that,under normal circumstance,chitosan oligosaccharide could increase activities of CAT and GPx and up-regulate gene expression of CAT and GPx4,but down-regulate gene expression of c-Jun and c-Fos in lymphocytes;under oxidative stress,chitosan oligosaccharide could increase CAT activity and down-regulate gene expression of c-Fos in lymphocytes.The results suggested that the protective mechanism of chitosan oligosaccharide on oxidative injury in lymphocytes might be attributed to the inhibition of AP-1 signaling pathway.