| Daily diet,environment and other external factors can cause the body to undergo oxidative stress,which is the main factor that causes various diseases such as diabetes,atherosclerosis,and inflammation.D-galactose is a common oxidative stress modeling reagent.This article mainly studies the effect of tea polysaccharide-tea polyphenol mixed functional components on experimental D-galactose-induced intestinal injury in mice,and explores its effect on the intestine The improvement effect and mechanism of injury.The materials used in the experiment contained about 57% tea polysaccharides,about20% tea polyphenols,and other non-functional components such as ash and moisture.By simulating the gastrointestinal digestion system in vitro,it is explored whether the tea polysaccharide-tea polyphenol mixed mixed functional ingredient(TPMFI)has an antioxidant effect,which provides basic data for later animal experiments.Since there is no antioxidant effect on other ingredients except tea polyphenols,in order to exclude the influence of the antioxidant effect of tea polyphenols on the results,a tea polyphenol group was set in the animal experiment,and the dose of tea polyphenol used in this group was the experimental material The same amount of tea polyphenols contained in the high-dose group.A model of intestinal oxidative stress was established by intraperitoneal injection of 10% D-galactose into mice for 56 days.After successful modeling,the mice were divided into normal group,model group,positive drug(200 mg/kg·bw reduced glutathione),tea polyphenol group(50mg/kg·bw),TPMFI low-dose group(40 mg/kg·bw),middle-dose group(100 mg/kg·bw),high-dose group(250 mg/kg·bw).All mice were given intragastric gavage for 45 days,once a day at the same time period,with a gavage volume of 0.1ml/10g·bw;normal group and model group were gavage with normal saline.Determine the relevant indicators,the experimental results are as follows:(1)In vitro experiment.After 2 hours of digestion of simulated gastric juice,compared with 5mg/ml TPMFI and 0.1mg/ml TPMFI,the DPPH·free radical scavenging rate,FRAP value,ORAC value,and-carotene fading inhibition rate increased by 63.1%,86.9%,30.1,respectively %,21.1%(p<0.05),the protein carbonyl content decreased by57.1%,and the brightness of supercoiled DNA bands increased.1mg/ml TPMFI was digested in simulated gastric juice for 2h and continued to be digested in simulated intestinal juice for 5h.Compared with gastric digestion for 2h,DPPH·free radical scavenging rate,FRAP value,ORAC value were reduced by 93.3%,78.9%(p<0.01),2.9 %(P>0.05),the fading inhibition rate of-carotene increased by 12.6%(p<0.05),the protein carbonyl content decreased by 15.2%(p<0.05),and the brightness of supercoiled DNA bands increased.It shows that under simulated gastrointestinal digestion conditions,TPMFI has the ability to scavenge free radicals and protect proteins and DNA from oxidative damage.(2)By measuring the content of relevant indicators of biological macromolecule oxidative damage in the jejunum tissue of experimental mice,it was found that the content of lipid oxidation products,protein oxidation products,and DNA oxidation products in mice with D-galactose-induced intestinal damage increased significantly(p< 0.05),the DNA oxidative repair index content was significantly reduced(p<0.05).After intervention with TPMFI,compared with the model group,the contents of MDA,4-HNE,8-iso-PGF2α,AOPPs,Dityr,PCO,and 8-OHd G in the jejunum tissue of the high-dose TPMFI group were reduced by 83.6%、 71.8%、62.2%、62.4%、78.5%、75.2%、63.6%(p<0.05),the 5-hm C content increased by 410%(p<0.05),all returned to the normal control group level,and compared with the tea polyphenol group There is a significant difference(p<0.05).It shows that in addition to tea polyphenols,other functional components mainly composed of tea polysaccharides can also repair the oxidative damage of the intestinal biological macromolecules of experimental mice.(3)By measuring the content of antioxidant-related indicators in the jejunum tissue of experimental mice and the m RNA expression of TLR4/p38/Nrf2 signaling pathway-related factors,it is found that the intestinal oxidative stress induced by D-galactose leads to the antioxidant capacity of mice Decrease,the m RNA expression of the relevant indicators of the antioxidant signal pathway is abnormal.After intervention with TPMFI,compared with the model group,high-dose TPMFI increased the T-AOC content,GR,and HO-1 activity in jejunum tissue by 220%,280%,and 170%,respectively(p<0.05),and reduced ROS content 72.8%(p<0.05);TLR4 and p38 m RNA expressions were reduced by 39.1%,82.4%(p<0.05),and Nrf2,GR,HO-1 m RNA expressions were increased by 83.6%,830%,96.3%,respectively(P<0.05),all recovered to the level of the normal control group,and there was a significant difference between the tea polyphenol group and the tea polyphenol group(p<0.05).It shows that TPMFI has the effect of restoring the antioxidant capacity of the jejunum tissue,and can improve the oxidative damage of the intestine by regulating the m RNA expression of related factors in the TLR4/p38/Nrf2 signaling pathway.(4)By measuring the contents of serum midgut mechanical barrier-related indicators and chemical barrier-related indicators in jejunum tissue,as well as MLCK pathway related factors and MUC2 m RNA expression in jejunum tissue,it was found that the small intestinal villi of model mice were severely broken and the intestinal epithelial cells were tightly connected.Destroyed;serum midgut injury-related indicators increased significantly(p<0.05),MLCK pathway related factors and MUC2 m RNA expression were abnormal.After intervention with TPMFI,compared with the model group,high-dose TPMFI restored the tight junctions of small intestinal villi and intestinal epithelial cells in mice;the contents of LPS,DAO,D-LAC,and i FABP were reduced by 37.7%,43.3%,50.6%,and 33.7%,respectively(P<0.05),the contents of MUC1 and MUC2 increased by220% and 210%,respectively;the expression of MLCK m RNA decreased by 76.0%(p<0.05),and the expression of ZO-1,claudin-1 and MUC2 m RNA increased respectively130%,350%,230%(p<0.05),all recovered to the level of the normal control group,and there was a significant difference between the tea polyphenol group(p<0.05).It shows that TPMFI can repair the intestinal mechanical and chemical barrier damage caused by oxidation by regulating the expression of MLCK pathway related factors and MUC2 m RNA.(5)By measuring the intestinal immune barrier-related indicators in the jejunum tissue and the m RNA expression of TLR4/Nf-kb pathway-related factors,as well as detecting the intestinal biological barrier by 16 s r RNA high-throughput sequencing,it was found that the content of the immune barrier-related indicators in model mice And TLR4/Nf-kb pathway related factors m RNA expression is abnormal.After TPMFI intervention,compared with the model group,the high-dose group TPMFI reduced the TNF-α and MPO contents in the jejunum by 66.9% and 54.7%,respectively(p<0.05),and the IL-10 content increased by 77.2%(p <0.05),the expression of Nf-kb and TNF-αm RNA were reduced by 47.6% and 74.2% respectively(p<0.05),and there were significant differences with the tea polyphenol group(p<0.05).It shows that TPMFI can improve the intestinal immune barrier damage caused by oxidation by regulating the TLR4/Nf-kb pathway related factors.Compared with the normal control group,the abundance,diversity,and ratio of the intestinal flora of the model group were significantly different(p<0.05).Different doses of T PMFI can improve this difference to a certain extent but not all the return to the normal group level indicates that TPMFI can regulate the types,abundance and uniformity of microorganisms in the intestinal tract,thereby protecting the intestinal biological barrier to perform its normal function. |
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