Protective Effect And Mechanism Of Tyrosol Combined With Lactobacillus Plantarum SC-5 On Ulcerative Colitis In Mice

Ulcerative colitis(UC)is a chronic,non-specific,recurrent,poor prognosis inflammatory bowel disease,clinical manifestations of diarrhea,hematochezia,abdominal pain and other symptoms.With the change of people’s dietary structure and the formation of new lifestyle,the incidence and prevalence of UC are on the rise,approaching three thousandths in the United States,five thousandths in Europe,and even more worldwide,so UC has become a serious public health problem worldwide[1].Although anti-inflammatory drugs,such as non-steroidal anti-inflammatory drugs(NSAIDs),can reduce the symptoms of UC,their long-term use is limited due to their severe hepatotoxicity and gastrointestinal adverse reactions [2,3].Therefore,it is crucial to explore alternative approaches for the treatment and prevention of UC.The pathogenesis of UC is complex,in which oxidative stress injury and intestinal flora disorder are important factors leading to the aggravation of UC.Studies have shown that the intake of specific natural antioxidant products can help regulate the intestinal microbiota and help treat and prevent colitis [4,5].The combination of different probiotics,or the combination of probiotics and other natural substances,is more effective in alleviating the UC process [6].Tyrosol(TY)is a phenolic compound derived from olive oil with anti-inflammatory,antioxidant,lipid-lowering,and anticancer properties [7].Lactobacilli can antagonize pathogenic bacteria and help to maintain the body’s immune response.Previous studies have found that Lactobacillus plantarum SC-5(S5)and TY have anti-inflammatory effects,and may contribute to the prevention or treatment of UC,but there is no systematic study on the mechanism of the combination of probiotics and natural antioxidant TY in the treatment of ulcerative colitis.Therefore,the purpose of this study is to investigate the protective effect of S5 combined with TY on UC and its related mechanisms,and to lay a theoretical foundation for further development of safe and natural drugs for UC prevention and treatment.S5 was used to interfere with 2.5% DSS-induced colitis model mice,and the disease activity index(DAI),colon length,and colonic histology were recorded and analyzed.The analysis showed that the combination of S5 and TY had more significant effects on alleviating the weight loss,colon length shortening,DAI score increased,inflammatory cell infiltration and intestinal crypt loss of colitis mice than single use.The pro-inflammatory cytokines(TNF-α,IL-1β,IL-6)in colonic tissue of mice were detected by Elisa,and the results showed that the combination of S5 and TY significantly reduced the pro-inflammatory cytokines in colonic tissue of mice.S5 and TY could also reduce the oxidative stress level(MPO)in UC mice,and the combination of S5 and TY could achieve better effect,basically restored to the same level as the normal group.We further detected the expression of NF-κB,MAPK and tight junction proteins ZO-1,Occludin,Claudin-3 in the colon of mice by Western blot and immunofluorescence assay.The results showed that the expression of ZO-1,Occludin,Claudin-3 and the number of goblet cells in colonic tissue of mice were significantly increased by the combination of S5 and TY.Meanwhile,our study further showed that the combination of S5 and TY inhibited the activation of NF-κB and MAPK by inhibiting the phosphorylation of NF-κB p65 subunit and MAPKp38,JNK and ERK1/2.Secondly,the intestinal microflora structure of mice was analyzed by using 16 s r RNA high-throughput sequencing technology.The analysis showed that the combination of S5 and TY had less effect on the structure of intestinal flora in DSSinduced ulcerative colitis mice than that alone,and the structure of intestinal flora in the combination of S5 and TY was similar to that in normal mice.By using Spearman correlation analysis,intestinal flora was closely related to colitis-related pathological parameters,in which beneficial bacteria such as Akkermansia,Faecalibaculum and Bifidobacterium were negatively correlated with DAI,pro-inflammatory cytokines and inflammatory cell infiltration.Desulfovibrio,Escherichia-Shigella and Erysipelatoclostridium were positively correlated with DAI,proinflammatory cytokines and inflammatory cell infiltration.This result suggests that the combination of S5 and TY may alleviate colitis-induced intestinal mucosal barrier damage by regulating intestinal flora.In order to verify the above hypothesis,we collected the feces of mice treated with S5 and TY and performed fecal bacterial transplantation(FMT)on UC mice.Subsequently,we detected the intestinal flora and inflammatory cytokines,inflammatory signaling pathways,tight junction proteins in the colon of mice.The results showed that FMT significantly alleviated the intestinal pathological tissue damage of DSS-induced ulcerative colitis mice,increased the number of goblet cells,inhibited the level of inflammation in mice,and increased the expression of intestinal tight junction proteins ZO-1,Claudin-3 and Occludin.FMT can also alleviate oxidative stress and inhibit NF-κB/MAPK-related inflammatory response stimulated by pathogenic bacteria.In addition,FMT significantly increased the abundance of the probiotic Akkermansia muciniphila in the gut of mice with DSS-induced colitis [10].These results suggest that the combination of S5 and TY can alleviate the intestinal barrier damage caused by DSS by regulating the intestinal flora structure.To sum up,this study shows that the combination of S5 and TY can enhance the intestinal physical barrier and improve the intestinal flora structure,and play a protective role against DSS-induced ulcerative colitis.This experiment provides experimental basis for the development of probiotics and natural antioxidant products,as well as public health,animal husbandry,disease drugs and other related fields.