| The diarrhea of weaned piglet is an urgent problem to be solved in the production of piglets. The dysfunction of intestinal mucosal barrier resulted from the various factors is the pathological characteristics and internal cause of diarrhea in weaned piglets. Therefore, the improvement of the intestinal health and the reduction of the diarrhea rate of weaned piglets are of great importance for increasing the production efficiency and level of the weaned piglets. The objective of the present study is to investigate the effects of Actigen, a second-generation mannan rich fraction, in antibiotics-free diet on the growth performance and intestinal barrier function in weaned piglets. One hundred and fifty30-day-old piglets(Duroc × Landrace × Yorkshire) were randomly divided into three groups, with 5 replications in each group and 10 piglets in a replication. The piglets in the control group were fed with the basal diet without antibiotics. The piglets in the Antibiotics group were fed with the basal diet containing100 mg/kg antibiotics, and the piglets in the Actigen group were fed with the basal diet supplemented with 800 mg/kg Actigen. The piglets were left to acclimate one week before the experimental and the formal experiment lasted for 28 days. The weaned piglets were freely accessed to the feed and water during the experiment. The feed intake and the diarrhea were recorded daily. At the end of the experiment, the piglets were weighted and 10 piglets were randomly selected in each group to be sacri?ced for sampling. The blood, intestinal contents, mucosa and intestinal tissue samples of weaned piglets were collected for further examination. The serum levels of protein, glucose and lipid metabolism and nitrogen metabolism related indicators, as well as the disaccharidases(sucrase, lactase, maltase) activity in ileal contents of weaned piglets were detected by biochemical kits. The level of serum endotoxin, D-lactic acid, diamineoxidase, inflammatory factor content and s Ig A in ileum mucous of weaned piglets were detected by using ELISA and RIA method. The HE staining was used to observe the intestinal morphology of weaned piglets and the PAS staining was applied to determine mucus secretion the intestine of weaned piglets. Western blot was used to detect the expression of tight junction protein(ZO-1, Occludin, Claudin-1) and inflammatory pathway related proteins(TLR4, My D88, p-IKK, p-JNK) in weaned piglets. The expression level of intestinal mucous inflammatory factor m RNA and ileum contents microflora 16 S r DNA on weaned piglets were tested by Real-time quantitative PCR.The research results are showed as follows:1. Effect of Actigen on the growth performance and diarrhea rate of weaned piglets.Compared with the control group, Actigen and antibiotics were able to achieve a significant reduction in the rate of diarrhea in weaned piglets.2. Effect of Actigen on intestinal physical barrier of weaned piglets. Compared with the control group, the serum levels of endotoxin were significantly decreased by Actigen and Antibiotics(p < 0.05). In addition, antibiotics significantly increased the level of D-lactic acid in the serum of weaned piglets(p<0.05). HE staining results showed that Actigen significantly increased the villus height, reduced the crypt depth, and elevated the ratio of the height of the villus and crypt depth of small intestinal(p<0.05). Western blot results showed that compared with the control group and Antibiotics group, Actigen significantly promoted the expression of tight junction protein(ZO-1, occludin) in the mucosa of duodenal and jejunum, and stimulated the expression of occludin in Ileum mucosa of weaned piglet(p <0.05). The results showed that Actigen could increase the intestinal physical barrier function of weaned piglets.3. Effects of Actigen on intestinal chemical barrier in weaned piglets. Compared with the control group, Actigen had the trend(p = 0.053) to decrease the p H value of duodenal contents in weaned piglets. PAS staining results showed that Actigen could significantly increase the number of goblet cells in jejunum mucosa of weaned piglets, and promote the secretion of intestinal mucus. In addition, Actigen also resulted a significant increase of the lactase activity in ileum content(p<0.05). Compared with the antibiotics group, Actigen could significantly increase the activity of invertase and maltase in the ileum contents(p<0.05). These results indicated that Actigen could regulate intestinal contents p H value and digestive enzyme activity so as to promote intestinal digestion and absorption of nutrients.4. Effects of Actigen on intestinal microbial barrier in weaned piglets. The results of microbial flora of the ileum contents in weaned piglets showed that compared with the control group, Actigen significantly increased the total number of intestinal bacteria and Escherichia coli(p <0.05). Meanwhile, Antibiotics significantly inhibited the number of Salmonella and lactobacilli and had thetrend(p=0.079) to increasethe number of bifidobacteria.5. Effect of Actigen on immune barrier of weaned piglets. The serum levels of inflammatory factors and the m RNA expression of inflammatory cytokine in ileum mucosa were detected. The results showed that Actigen and Antibiotics significantly reduced the serum levels of inflammatory cytokines of weaned piglets(p<0.05), with higher reduction in the Actigen group. Meanwhile, Actigen significantly increased the serum content of antiinflammatory factor IL-10 of weaned piglets(p<0.05). In accord, compared with control group, Actigen could significantly increase the m RNA expression of anti-inflammatory factor IL-10 and IL-4 in weaned piglets(p<0.05).The results of the expression of TLR signaling pathway in Ileum mucosa of weaned piglets demonstrated that compared with control group, Antibiotics significantly increased the expression of TLR4, My D88 and p-IKK(p < 0.05). Compared with the Antibiotics group, the expression of TLR4, My D88, p-IKK in the ileum mucosa of weaning piglets was significantly decreased by Actigen(p<0.05).In conclusion, Actigen was able to reduce the intestinal permeability of weaned piglets by increasing the expression of intestinal tight junction protein and mucin, improving the integrity of intestinal structure. In addition, Actigen could also increasethe activity of disaccharidase in ileum contents, decrease the level of pro-inflammatory factors in serum and reduce the expression of inflammatory pathway protein in the ileum mucous. The results showed that Actigen could improve the intestinal health, which provides a scientific basis for the application of Actigen as an alternative to antibiotics in the pig production. |
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