| Weaning stress has an adverse effect on the growth performance and intestinal health of piglets, which can lead to severe outcomes such as diarrhea, slow-growing and reduction in resistance etc. Zinc Oxide(Zn O) is often excessively used in piglet industry as a mitigation measure although some problems like food safety and environmental pollution exist. As a kind of environmental protection feed additive, probiotics is much more used widely than before. This research was focused on protecting and improving intestinal mucosal barrier function in weaning piglets. Thus we investigated the effects and mechanism of the probiotics on relieving weaning stress and ameliorating intestinal health of piglets, which might provide a theoretical support for that probiotic could replace Zn O in piglets weaning period. The main results were listed as follows:Trial 1: Effects of Clostridium Butyricum and Lactobacillus salivarius ZJ614 on intestinal mucosal barrier function of actue diarrhea ratsForty-eight 21d-weanned wistar rats(weighing 57.45±6.23 g) were selected and randomly divided into four groups: Normal group(C1), model group(C2), Clostridium Butyricum-treated group(T1) and Lactobacillus salivarius ZJ614-treated group(T2). Each group had six replicates, each replicate had two rat. This experiment lasted for 9 days. C1, C2 groups were orally administered saline, and T1, T2 groups were administered probiotics accordingly. On the 8th day, every group except C1 was administered folium senna to cause acute diarrhea and then record diarrhea condition of rats. All rats were sacrificed and blood or intestinal samples were collected at the end of experiment. The results were showed as follows:1) All rats displayed diarrhea symptoms as long as administered folium senna. Loose stools rate, diarrhea index, serum endotoxin and D-lactic acid were significantly increased in C2 group compared with C1 group(P<0.01). The diarrhea index, serum endotoxin and D-lactic acid decreased significantly in T1 and T2 groups compared with C2 group(P<0.01).2) Compared with C2 group, T1 and T2 groups could significantly decrease the crypt depth(P<0.05),and significantly increase the ratio of villus height to crypt depth(v/c)(P<0.01).T1 group could improve the level of zonula occludens-1(ZO-1) and occludin m RNA expression significantly in comparison to the other three groups(P<0.05).3) C2 group significantly up-regulated the level of Interleukin-10(IL-10) and Interleukin-1β(IL-1β) m RNA expression compared with C1 group(P<0.01), and T1 group down-regulated the level of IL-10 m RNA expression compared with C2 group(P<0.01).In summary, Clostridium Butyricum or Lactobacillus salivarius ZJ614 can alleviate the actue diarrhea induced by senna on a certain degree, and also decrease the intestinal permeability, improve ileum morphology; furthermore, Clostridium Butyricum was better than Lactobacillus salivarius ZJ614 in the expression of tight junction protein and mucosal cytokine.Trial 2: Effects and its mechanism of Clostridium Butyricum on intestinal mucosal barrier function in weanling PigletsEighteen ‘Duroc ×Landrace ×Yorkshire’ crossbred piglets with an average body weight of(5.35±1.33) kg were weaned at 21 days of age and randomly allocated into three groups: Basal diet group(C group), basal diet + 500 mg/kg Clostridium Butyricum group(T group) and basal diet + 3 000 mg/kg Zn O group. Each group had six replicates with one piglet per replicate. This experiment lasted for 14 days. The results showed as follows:1) There was no difference in growth performance among three groups(P>0.05). Piglets in T group or Zn O group had lower diarrhea rate(P<0.05), lower crypt depth(P<0.05), higher ratio of villus height to crypt depth(v/c)(P<0.01) compared with C group. There was no difference in diarrhea rate, ileum morphology between T group and Zn O group(P>0.05). Compared with C group, piglets in Zn O group had higher jejunum villus height(P<0.05) and more ileum neutral acidic goblet cells(P<0.05). And there was a tendency that the number of total goblet cells in T group and Zn O group were increased(P<0.1).2) On the day 7 and 14, serum endotoxin and D-lactic acid were decreased significantly in T group or Zn O group compared with C group(P<0.05), and there was no difference between T group and Zn O group(P>0.05). T group or Zn O group had higher occludin m RNA expression in ileum and colon compared with C group(P<0.05). There was no difference in the zonula occludens-1(ZO-1) and occludin m RNA expression between T group and Zn O group(P>0.05). Compared with C group, Zn O or Clostridium Butyricum could up-regulate the protein expression of ZO-1 and occludin both in ileum and colon(P<0.05).3) Compared with C group, Zn O or Clostridium Butyricum could down-regulate the level of proinflammatory cytokine IL-1β in ileum and proinflammatory cytokine TNF-α m RNA expression in colon(P<0.05), up-regulate anti-inflammatory cytokine TGF-β1 m RNA expression in colon(P<0.05). T group and Zn O group had higher s Ig A content in ileum and colon mucosa in contrast with C group(P<0.05). There was no difference between T group and Zn O group in above indexes.4) Compared with C group, piglets in T group had higher gene expression of TLR2, My D88, NF-κB, TRAF6, and lower gene expression of TAK1 in ileum mucosa(P<0.05); There was no difference in the gene expression of NF-κB, TAK1, p38 MAPK between T group and Zn O group(P>0.05), but T group had higher TLR2, My D88, TRAF6 gene expression in comparison with Zn O group(P<0.05). In colon mucosa, Clostridium Butyricum elevated the gene expression of TLR2, NF-κB(P<0.05), and had a tendency to up-regulate My D88, TRAF6 expression compared with C group(P<0.1). There was no difference between T group and Zn O group in the gene expression of TLR2, My D88, NF-κB, TRAF6, TAK1, p38 MAPK in colon mucosa(P>0.05).5) In the ileum mucosa of piglets, the measured microflora mainly belonged to Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria. C group was greater than T group, and T group was greater than Zn O group in community richness and species evenness. Beta diversity analysis showed that, microflora in T group was relative concentration with less distance among the group samples than other groups. There was no difference in the phylum level among the three groups. But the addition of Zn O or Clostridium Butyricum elevated the proportion of Clostridiaceae, Streptomycetaceae; with the proportion of Lachnospiraceae, Bacteroidaceae, Nocardiaceae decreased(P<0.05). The microflora function prediction results showed that microflora in ileum intestinal of T group had potential value in cell motility, signal transduction, genetic translation, replication and repair, metabolism such as xenobolism biodegradation, carbohydrate metabolism, amino acid metabolism etc. And Zn O group mainly concentrated on metabolism such as xenobolism biodegradation and metabolism, lipid metabolism, metabolism of terpenoids and polyketides, amino acid metabolism etc.In conclusion, these results indicated that the diet supplemented with Clostridium Butyricum could decrease diarrhea rate, improve intestinal mucosal morphology, decrease intestinal permeability and up-regulate tight junction protein expression and improve mucosal immunity of weaning piglets. The effects of Clostridium Butyricum on improving intestinal barrier function was in comparable to high dose Zn O. And its mechanism might go through two pathways: on the one hand, it could activate mucosal immune function by regulating TLR/NF-κB signaling pathway; on the other hand, it could stabilize and improve ileum mucosal microflora, maintain intestinal microecological balance in weaning piglets. Above all, Clostridium Butyricum could replace high dose of Zn O to relieve weaning stress and keep normal performance in weaning piglets. |
