| Early microbial colonization in suckling pigs has a profound effect on subsequent growth of piglets,and nutrients can effectively modulate the intestinal microbiota of piglets.Galacto-oligosaccharides(GOS),as a kind of prebiotic oligosaccharides with natural attributes,are not easily broken down in the small intestine due to the lack of corresponding digestive enzymes,and usually the vast majority of ingested GOS can directly enter the cecum and colon for fermentation and utilization by microorganisms.Studies have proved that GOS can modulate the intestinal microbiota in suckling piglets,and the modulation of intestinal microbiota is also beneficial to the intestinal barrier function.However,whether GOS can modulate the intestinal microbiota of suckling piglets in the intestinal inflammation injury model and relieve the intestinal inflammation and maintain the intestinal barrier function is worthy of our further exploration.As a strong inflammatory stimulant,lipopolysaccharides(LPS)are widely used in the construction of animal intestinal inflammatory injury models.Therefore,this experiment used lipopolysaccharides to challenge suckling piglets as the research object,aiming to study the effect of GOS on intestinal morphology,intestinal microbiota composition,inflammatory response and barrier function in LPS-challenged suckling piglets.The experimental design is to select 18 newborn suckling piglets with similar initial weights and divide them into three treatment groups,namely the control group(CON),the lipopolysaccharides challenge group(LPS-CON)and the GOS intervention group(LPS-GOS),6 piglets per group.From the first day after birth,piglets in the LPS-GOS group were fed with 1 g/kg body weight GOS solution every day,while the CON group and LPS-CON group were fed with the same amount of sterile saline.After 13 days of continuous feeding,the piglets in the LPS-CON group and the LPS-GOS group were intraperitoneally injected with an LPS solution of 80μg/kg body weight on the 14 th day.The piglets in the CON group were injected with an equal volume of sterile saline.Two hours after intraperitoneal injection of LPS solution or sterile saline,the piglets were slaughtered and sampled.This study was mainly divided into the following three parts:1.Effects of galacto-oligosaccharides on the morphology and antioxidant status in the hindgut of suckling piglets challenged with lipopolysaccharidesThis chapter aims to investigate the effects of galacto-oligosaccharides on morphological structure and antioxidant status in the cecum and colon of LPS challenged suckling piglets.The results showed that compared with the LPS challenge group,the colon length of the piglets in the GOS intervention group tended to increase(P = 0.089),and the ratio of colon length to body weight increased significantly(P <0.05).Observing the piglet’s cecum and colon HE stained sections,the results showed that the cecum and colon of the CON group were intact,while the LPS challenge group had obvious morphological injured,and the GOS intervention group had relatively integrated intestinal morphology.The antioxidant status of intestinal mucosa was evaluated,and the results showed that the intervention of GOS significantly decreased the concentration of MDA and the activity of ROS in cecum and colon of LPSchallenged suckling piglets(P < 0.05),and significantly increased the T-AOC level in cecum and colon mucosa(P < 0.05).And the activity of SOD in colonic mucosa tended to increase(P = 0.068).The result of the relative expression of apoptosis-related genes showed that the intervention of GOS significantly reduced the relative m RNA expression of caspase-3 in the cecal mucosa of LPS-challenged suckling piglets(P <0.05),and tended to decrease the relative m RNA expression of BAX(P = 0.065).In addition,the intervention of GOS significantly reduced the relative m RNA expression of caspase-3,caspase-9 and BAX in colonic mucosa,and significantly increased the relative m RNA expression of BCL2(P < 0.05).2.Effects of galacto-oligosaccharides on the intestinal microbiota and their metabolites in the hindgut of suckling piglets challenged with lipopolysaccharidesThis chapter aims to investigate the effects of galacto-oligosaccharides on the cecal and colonic microbiota and bacterial metabolites in LPS-challenged suckling piglets.The results showed that,in the cecal digesta,the intervention of GOS significantly increased the Shannon index and the relative abundance of Firmicutes in LPS-challenged suckling piglets(P < 0.05).At the genus level,compared with the CON group,LPS challenge significantly reduced the relative abundance of Ruminococcaceae＿NK4A214＿group and Prevotellaceae＿NK3B31＿group(P < 0.01),and tended to decrease the relative abundance of Lactobacillus(P = 0.054)and unclassified＿f＿＿Lachnospiraceae(P = 0.051).In addition,the intervention of GOS significantly increased the relative abundance of[Eubacterium]＿coprostanoligenes＿group,Phaseolarctobacterium,Prevotella＿2,Lachnospiraceae＿NK4A136＿group and Faecalibacterium(P < 0.05)in the caecal digesta of LPS-challenged suckling piglets.In the colonic digesta microbiota,no significant differences were found in the three groups at the phylum level.At the genus level,compared with the CON group,LPS challenge tended to reduce the relative abundance of Lactobacillus(P = 0.059)and Veillonella(P = 0.064).In addition,compared with the LPS challenge group,the GOS intervention significantly increased the relative abundance of Blautia,Ruminococcaceae＿UCG＿005,Ruminococcus＿2,Oscillibacter,Lachnospiraceae＿NK4A136＿group,Lachnospiraceae＿FCS020＿group,Ruminiclostridium＿9 and Coprococcus＿3(P < 0.05).In the colonic mucosa,compared with the CON group,LPS challenge increased the relative abundance of Actinobacteria(P < 0.05)and tended to decrease the relative abundance of Firmicutes(P = 0.054).At the genus level,LPS challenge significantly increased the relative abundance of Alloprevotella,Campylobacter,and Helicobacter(P < 0.05),and tended to increase the relative abundance of Alipipes(P = 0.085).In addition,LPS challenge significantly reduced the relative abundance of norank＿f＿＿Muribaculaceae,Prevotellaceae＿NK3B31＿group and Ruminococcaceae＿NK4A214＿group(P < 0.05),and tended to decrease the relative abundance of Lactobacillus(P = 0.097)and Romboutsia(P = 0.083).The GOS intervention significantly increased the relative abundance of norank＿f＿＿Muribaculaceae and Romboutsia in the colonic mucosa of LPS-challenged suckling piglets(P < 0.05),and tended to increase the relative abundance of Lactobacillus(P = 0.058);in addition,it significantly reduced the relative abundance of Alloprevotella,Alitipes(P < 0.05),Campylobacter and Helicobacter(P< 0.01).In terms of microbial metabolites,the intervention of GOS significantly increased the concentration of acetate,propionate(P < 0.05),butyrate and total SCFAs(P < 0.01)in the cecal digesta of LPS-challenged suckling piglets.In addition,it significantly increases the concentration of acetate,butyrate,total SCFAs and lactate in the colonic digesta(P < 0.05).3.Effect of galacto-oligosaccharide on inflammation and barrier function in the hindgut of suckling piglets challenged with lipopolysaccharidesThis chapter aims to investigate the effects of GOS on the inflammation and barrier function of the cecum and colon in LPS-challenged suckling piglets.The results showed that compared with the CON group,LPS challenge significantly increased the concentration of IL-1β,IL-6,IL-8 and TNF-α in the mucosa of the cecum and colon(P< 0.05).Compared with the LPS challenge group,the concentrations of IL-1β,IL-6 and TNF-α in the cecum and colon mucosa of the piglets in the GOS intervention group were significantly reduced(P < 0.05).Further analysis of the expression of inflammatory pathway related factors in cecal mucosa showed that LPS challenge increased the relative m RNA expression of CD14(P < 0.05),TLR4(P < 0.05),My D88(P = 0.078)and NF-κB p65(P < 0.05).Compared with LPS challenge group,GOS intervention reduced the relative m RNA expression of CD14(P = 0.057),TLR4(P <0.01)and NF-κB p65(P = 0.088).In colonic mucosa,LPS challenge significantly increased the concentration of CD14(P < 0.01)and the relative m RNA expression of TLR4,My D88,and NF-κB p65(P < 0.05).Compared with the LPS challenge group,the concentration of CD14 and the relative m RNA expression of TLR4 and My D88 in the GOS intervention group were significantly reduced(P < 0.05),and the relative m RNA expression of NF-κB p65 tended to decrease(P = 0.062).In terms of intestinal barrier function,compared with the CON group,LPS challenge significantly increased the relative m RNA expression of MUC2 in the cecal mucosa,and significantly increased the relative m RNA expression of MUC1 and MUC2 in the colonic mucosa(P< 0.05).Compared with the LPS challenge group,the GOS intervention significantly reduced the relative m RNA expression of MUC2 in the colonic mucosa of piglets(P <0.05),and tended to decrease the relative m RNA expression of MUC1(P = 0.080).In addition,compared with the CON group,LPS challenge significantly reduced the protein expression of Claudin-1 and ZO-1 in the cecal and colonic mucosa(P < 0.05),and tended to decrease the protein expression of Occludin in the cecal mucosa(P =0.088).Compared with the LPS challenge group,the GOS intervention significantly increased the protein expression of Claudin-1 in the cecal and colonic mucosa and the protein expression of ZO-1 in the colonic mucosa(P <0.05).In summary,GOS enhanced the antioxidant capacity in the hindgut of LPSchallenged suckling piglets,alleviated the morphological injured of the hindgut,and maintained the morphological integrity.GOS modulated the microbial composition in the hindgut of LPS-challenged suckling piglets,increases the concentration of shortchain fatty acids in the hindgut.GOS alleviated the intestinal inflammatory response and maintained the intestinal epithelial barrier function of of LPS-challenged suckling piglets. |
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