Potential Mechanism Of Flammulina Velutipes Polysaccharide In Preventing Inflammatory Bowel Diseases,structure Characterization And The Biological Effects Of Its Main Component

Background and objectiveInflammatory bowel disease(IBD)is a common and serious disease,and the incidence of IBD in the world is increasing each year.Traditional therapeutic drugs such as 5-aminosalicylic acid(5-ASA)and glucocorticoids have certain effects,but the side effects are large.In recent years,studies have found that plant polysaccharides and fungi polysaccharides have a better control effect on IBD.Flammulina velutipes polysaccharide(FVP)is the main active ingredient of Flammulina velutipes.Through pre-experiment,we have found that FVP can alleviate the symptoms of IBD induced by Dextran sulfate-sodium salt(DSS)in rats.The effects of FVP on preventing IBD,the effect mechanism,the structure of FVP main components and the activities of pure polysaccharides were studied in this researchMethods1.The rat model of DSS-induced IBD was established.Forty-eight male Sprague-Dawley rats were randomly divided into 6 groups(n=8),namely blank control group,IBD model,FVP low,medium and high dose group,and 5-ASA group.The control group was given normal saline for 14 days with normal drinking water;the model group was given normal saline for 14 days,the first 6 days given normal drinking water,the last 8 days given DSS drinking water(4.5%);the polysaccharide group and the positive drug group were orally administered FVP(50,100,200 mg/kg)and 5-ASA(200 mg/kg)for 14 days,normal drinking water for the first 6 days,DSS drinking water for the last 8 days,and animals were sacrificed on the 15th day.2.The effects of FVP on DSS-induced IBD in ras were evaluated.The pathological specimens were observed under the microscope for inflammation,and the colon tissue MPO was determined.3.The effect mechanism of FVP on DSS-induced IBD in rats was investigated.(1)determination of intestinal flora;(2)determination of short-chain fatty acids in the intestine;(3)detection of Occludin,ZO,TLR4,NF-κB expression in colonic tissues were done by RT-PCR and Werstern Blot.4.The main component of FVP was purified and its structure was identified.The crude polysaccharide of FVP was purified to obtain pure polysaccharide FVP1.The molecular weight,monosaccharide composition and glycosidic bond type of FVP 1 were analyzed.5.FVP1 activity study as follows(1)the effects of FVP1 on the production of NO,TNF-a,IL-6 and IL-12 in RAW264.7 cells;(2)the effects of FVP 1 on mitochondrial respiratory function in RAW264.7 cells.Results1.The epithelial damage was severe seen under the microscope,and the MPO activity was significantly higher than that of the control group.FVP relieved pathological damage of the colon and significantly reduced MPO activity.2.The ratio of Firmicutes to Bacteroides decreased(F/B),the alpha diversity index decreased,the abundance of pathogens such as Helicobacteraceae increased,and the abundances of beneficial bacteria such as Lachnospiraceae and Lactobacillales decreased.FVP up-regulated the F/B value and the alpha diversity index,which reduced the abundance of Helicobacteraceae and increased the abundance of Lachnospiraceae and Lactobacillales.3.The content of acetic acid,propionic acid and butyric acid in the model group was significantly lower than that in the control group.FVP increased the content of short-chain fatty acids,and the effect was significant in the medium and high dose groups.4.The expression of Occludin and ZO-1 was significantly lower than that of the control group.The expression of TLR4 and NF-κB was significantly higher than that of the control group.FVP had no significant effect on Occludin and ZO-1,but significantly decreased the expression of TLR4 and NF-κB.5.FVP1 has a molecular weight of 54.78 kDa and consists of mannose(7.74%),glucose(70.41%)and galactose(16.38%).The main glycosidic bond types of FVP1 are(1→6)-β-D-galactose,a-D-glucose,(1→3,6)-α-D-mannose.6.FVP1 promoted the secretion of NO,TNF-a,IL-6,and IL-12 in RAW264.7 cells;increased the maximum oxygen consumption rate of mitochondria in RAW264.7 cells.ConclusionsFVP has a preventive effect on DSS-induced colitis in rats,which can regulate intestinal flora,increase the production of short-chain fatty acids,and inhibit the expression of inflammatory factors in colonic mucosa.FVP 1 is a novel polysaccharide that has immunomodulatory and in vitro antiviral activity while improving mitochondrial respiratory function.