Protective Effects And Mechanism Of Sugar Cane Extract Against Gut Damage

The gut is one of the critical organs in animal production.Firstly,the absorption efficiency of nutrients in the intestine directly determines the amount of nutrients for animal growth.Secondly,the diversity and balance of intestinal flora maintain the health of the intestinal tract and inhibit the excessive proliferation of pathogenic bacteria.Finally,the gut serves as the largest immune organ in the body,which plays a role in regulating the intestinal barrier against pathogens.Its antioxidative and anti-inflammatory ability directly affect the integrity and the metabolic update rate of the gut,and eventually determines the health and the production effency of animals.With the extension of the research breadth and depth of animal nutrition,the research on functional extracts from plant sources has become a hot topic.This manuscript mainly focuses on the sugar cane extract(SCE)from sugar industry.Intestinal health is a major research field of nutritional science.Ulcerative colitis is one of the manifestations of unhealthy intestinal function,and its' incidence rate gradually increased.Meanwhile,it becomes a refractory case in our country.Research and development of functional feed from natural plant source has become a hot topic with a bright prospect.Firstly,this study analyzed the conventional and functional nutrients,such as amino acids,total polyphenols,stibene glucoside,rosemary acid,total flavonoid and the content of soybean isoflavaones of SCE.Secondly,the protective effects and mechanism of dietary SCE against diquat-induced gut damage in weaned piglet were studied.Thirdly,this study built the mice ulcerative colitis model through drinking water added with dextran sodium sulfate(DSS).Finally,this study focused on the protective effect and possible mechanism of adding 1%SCE against DSS-induced colonic inflammation in mice.The main results were as follows:1.Determination of the conventional and functional nutrients of SCEThis study measured the conventional and functional nutrients composition of SCE.The conventional nutrient composition of SCE consists of 8.27%water,9.08%ash,10.47%crude protein,3.02%crude fat,1.33%crude fiber and 67.83%nitrogen-free extracts.The amino acid contents(mg/g)of SCE were as following:Asp+Asn 9.876,Thr 2.727,Ser 4.091,Glu+Gln 30.675,Gly 3.079,Ala 3.209,Cys 0.746,Val 3.826,Met 0.207,Ile 2.882,Leu 5.717,Tyr 1.088,Phe 4.653,Lys 1.529,His 1.707,Arg 2.259,and Pro 4.187.The total polyphenol content is 0.587%.The rosemary acid content is 17.5 mg/kg.The wogonoside was not detected at the 0.2 mg/kg detection limit.The stibene glucoside was not detected at the 0.5 mg/kg detection limit.The soybean isoflavaones(daidzin,glycitin,genistin,daidzein,glycitein,genistein)was not detected at the 5 mg/kg detection limit.The above nutrients and functional ingredients in SCE laid a nutritional foundation for the subsequent study of its protective effects on intestinal oxidation and inflammatory damage.2.Effects of SCE against diquat-induced intestinal oxidative damage in weaned pigletTwenty-eight weaned piglets were randomly divided into four groups;the control group(Cont),Diqu group(Diqu),1%SCE supplementation group(1%SCE)and 2%SCE supplementation group(2%SCE).On d 21,piglets in the last three groups were intraperitoneally injected with 10 mg/kg of Diqu solution,and piglets in the Cont group were intraperitoneally injected with same amount of physical saline.All piglets were killed on d 24.The jejunum and ileum samples were separated to determine the changes of intestinal morphology.Results showed that 1%SCE supplementation increased average daily gain 12.94%and average daily feed intake 11.38%compared to the control group during the d21 before diaquat injection.The number of diarrhea piglets in SCE groups was significantly lower than that in the control group.During the 72 h after diaquat injection,2%SCE supplementation increased average daily gain and average daily feed intake(P<0.05)compared to the diaquat group.The recovery time of reintake of the 2%SCE group was about 15 hours earlier than that of the diquat group.Meanwhile,1%SCE supplementation significantly improved the depth of the crypt and the intestinal morphology after diaquat induced mucosa damage.Colonic length and morphological structure were improved by 2%SCE.3.Molecular mechanism of SCE against diquat-induced gut damage in weaned pigletThe experimental design and diet is the same with the above trial,also the same as the diquat challenge dose and slaughter time.The blood samples were taken to measure the GSH-PX,T-AOC,MDA,DAO and SOD,together with IgA,IgG and IgM.The jejunum and ileum tissue were taken for western blot annlysis of NF-?Bp65,Nrf2,ZO-1,Occludin and Claudinl.The colonic digesta were taken for microbial diversity analysis.Results showed that SCE supplementation increased the concentration of IgM and GSH-PX in the blood(P<0.05).The expression level of NF-?Bp65 in jejunum of piglets in 1%SCE group was significantly lower than that of control group(P<0.05).Adding 2%SCE significantly increased the expression level of Nrf2 protein in jejunum of piglets(P<0.05).The expression level of occludin in jejunum of piglets in 1%SCE group was significantly higher than that of control group(P<0.05).Concurrently,a significant discrepancy in colonic microbiota existed among four groups.At the phylum level,when compared to the Diqu group,2%SCE supplementation decreased the relative abundance of Spirochaetae,Euryarchaeotaand Verrucomicrobia(P<0.05).At the genus level,increases in relative abundances of Lactobacillus and Ruminococcus_gauvreauii group,while decreases in relative abundances of Clostridium_sensu_stricto₁,Klebsiella and Sharpea were observed in the SCE supplementation group.In conclusion,dietary supplementation with SCE might alleviate the gut oxidative stress induced by Diqu,through promoting colonization of beneficial bacteria,and inhibiting colonization of harmful pathogens in the colon of weaned piglets.4.Appropriate dose and exposure time of dextran sulfate sodium to induce the ulcerative colitis in miceForty female ICR mice(26.63±0.19 g)were divided into four groups(a control group,3%,4%and 5%dextran sodium sulfate(DSS)challenged group)to build acute ulcerative colitis.The method is that each group continuously drank up deionized water,3%,4%,5%DSS water for 7 d.Taken the body weight change,colonic length and histological structure together,3%,4%,5%DSS aqueous solution all could induce ulcerative colitis.Severity of the three groups was as following:3%group was partial light,4%group was moderate,5%group was partial serious and even some of the mice had the risk of death.Therefore,we chose the 4%DSS aqueous solution for 7 d to build female ICR mice UC model for further research on the protective effect and possible mechanism of SCE.5.Protective effect and possible mechanism of SCE against DSS-induced colonic inflammation in miceForty female ICR mice(26.63 ± 0.19 g)were randomly divided into four groups.The groups were arranged as a 2 × 2 factorial design,which included 2 SCE levels(0%,1%)and 2 levels of DSS(0%,4%).0%SCE-DSS(+),.Mice in 0%SEC groups were fed basal diet and the other mice received 1%SCE supplemented in basic diet from 6-week to 8-week for 14 d.At 7-week-old,mice in the 0%SCE-DSS(+)group and the 1%SCE-DSS(+)group were also given a 4%DSS solution for 7 d(d 8 to d 14 of the experiment)via drinking water to induce the colonic inflammation.The results showed that SCE played a protective effect of ulcerative colitis in mice by increasing daily weight gain,stabilizing spleen weight,increasing colonic length and improving colonic histological structure.A possible mechanism underlying the protective of SCE against the DSS-induced UC consists of four aspects.To begin with,SCE could significantly increase mice's serum Ig levels to enhance immunity.Moreover,SCE could strengthen the antioxidative capacity(GSH-PX,MDA and T-AOC)via increasing the antioxidative gene GPX1 and GPX4 expression.Once more,SCE could decrease the proinflammatory IL-17 expression,and further inhibit the p65 signal pathway,which depress the transcription of downstream proinflammatory genes(TNF-? and IL-1?).In the end,SCE could enhance the intestinal mucosal mechanical barrier by up-regulating the tight junction-related proteins(ZO-1,Occludin and Claudin1).To sum up,this study comes to the following conclusions:1.The sugar cane extract had a high nutritional value and strong biological activity,which layed a nutritional foundation for the follow-up study on its effect and mechanism of intestinal damage.2.Dietary supplementation with SCE might protect the gut oxidative damage induced by Diqu,through increasing serum immunoglobulin content,maintaining the small intestine and colon mucosa barrier structure,promoting colonization of beneficial bacteria,and inhibiting colonization of harmful pathogens in the colon of weaned piglets.3.Drinking water with DSS could induce ulcerative colitis in female ICR mice,preferably at a concentration of 4%for 7 d.Dietary supplementation with SCE could alleviate mice colitis by increasing daily weight gain,increasing colon length and improving colon tissue structure.The mechanism of alleviation was mainly achieved by increasing serum IgA concentration,enhancing the expression level of antioxidant genes GPX1 and GPX4,blocking the activation of NF-?B signaling pathway involved in nuclear protein p65,and increasing the expression levels of cell tight junction proteins.