| ObjectiveTo extract the most possible types of the targeted floras,by observing the gut microbiota distribution characters of SD rats fed with high-fat diet,and discussing the effect on gut microbiota intervened by Tangshen Capsule.Methods:1.Model building and division.90 six-week-old rats adaptably fed for 1 week were divided into normal control group(n=15)and high-fat diet group(n=75).Rats were fed with normal diet and high-fat one respectively.The feeding period was 8 weeks.On the foundation of plenty of former experimental results,evaluating whether MS model was successfully copied was on the basis of weight changes.After 8 weeks,rats fed with high-fat diet were randomly divided into high-fat diet group、berberine hydrochloride group,coptis decoction group,low dose of Tangshen capsule,high dose of Tangshen capsule.Each group remained the stable feeding.Anddrug intervention lasted for 4 weeks.2.Measure relative indicators of MS,weigh the weight of each group,calculate the weight change and the change rate,collect blood samples and measureblood glucose,blood fat,free fatty acid and fasting insulin,get epididymal fat to measure visceral fat mass,observe the weight change,the change rate and the change of MS relative indicators of rats after being intervened by berberine hydrochloride,coptis decoction andTangshen capsule.The differences between groups,measurement data were tested by the independent sample t.3.After 12 weeks,the rats were anesthetized and killed with intraperitoneal injection of 3%pentobarbital.Then the enterocoeliaswere opened,blood was collected from abdominalaortas,which was centrifuged to get serum and conserved to be used.The ceca of rats were intercepted from 2cm away from upper end and lower end respectively(The whole cecum was included and fresh feces was contained),then they were packed with silver paper and put into feces boxes of 50ml after being marked.And they were iced in drikold instantly and preserved to be used in-80 ℃.Six samples were randomly selectedin each group,and Huada Gene Limited Company was commissionedto detect 16S rDNA insamples(feces in ceca).Such software as flash/mothur were used to joint readl and read2,filter the data of low quality,exclude chimeric sequences,optimize,settle and analyze it after getting original data,and analyze the abundance and diversity of species in every sample.Moreover,OTU Venn diagram,PCA analysis diagram of OUT abundance,OTU Rank curve,species heatmap,phylogenetic tree diagram and so on were used to show.The flora differences in intestinal flora among six groups at the degree of Phylum,Class,Order,Family,Genus and Specieswere more analyzed,by using species profiling histogram to show.Results1.After 8-week high-fat diet,compared with normal matched group,the rats fed with high-fat diet weighed increasingly;the weight increment,growth rate advanced apparently.In week12,compared with normal matched group,high-fat diet group advanced more apparently in terms of the weight increment,growth rate and higher visceral fat mass,lower HDL-C,and higher LDL-C.Compared withhigh-fat diet group,high dose of Tangshen capsule group showed significant decrease of the weight increment and growth rate,and visceral fat mass decreased;HDL-C increased significantly,and LDL-C decreased significantly.Low dose of Tangshen capsulegroup had a decreasing trend of weight growth rate and an increasing one of HDL-C.Berberine hydrochloride group and coptis decoction groupshew a decreasing trend of weight increment and growth rate;visceral fat mass didn’t show no difference;and HDL-C and LDL-C didn’t show difference.2.Compared with high-fat diet group,the amount of OUT in normal matched group was increased significantly;there were significant differences between PC1 and PC2 dimension.OUT Rank curve shew a greater span and a gentler curve in normal matched group.The diversity of individual samples decreased significantly in high-fat diet group,and the diversity of Alpha and Beta decreased significantly.High-fat diet group shew a decreasing trend of Ackermann’s bacteria,an apparently decreasing trend of Butyric acid bacteria and Ross bellerella;an apparently increasing trend of Brausteria,Bacteroides eggerthiiand preval.Compared with high-fat diet group,the four groups of drug intervening therapy shew apparent differences in PC1 and PC2 dimension.The diversity of Alpha differed significantly,showing normal matched group>high dose of Tangshen capsule group>Low dose of Tangshen capsule group>coptis decoction group>high-fat diet group>berberine hydrochloride group.The diversity of Beta differed significantly,and the most was high dose of Tangshen capsule group,closest to normal matched group.Compared with high-fat diet group,OUT Rank curve inhigh and low dose of Tangshen capsule,coptis decoction group shew a greater span and a gentler slope;berberine hydrochloride group shew a decreasing curve span and slope.Compared with high-fat dietgroup,in high and low dose of Tangshen capsulegroup:two groups of Tangshen capsule shew a significant decrease of Bacteroidetes,in high dose group,it shew more apparent.Firmicutes shew an increasing trend in grouphigh and low dose of Tangshen capsule,coptis decoction group,while shew a decreasing one in berberine hydrochloride group.Alpha Proteobacteria,Ackerman’s genus,butyric acid bacillus and coccus,Przewalski’s habitat Ross Berry’s bacteria shew a significant difference.Ackerman’s bacteria repairing intestinal barrier were significantly gathered in high and low dose of Tangshen capsulegroup,and Bacteroides,Staphylococcus aureus,butyric acid,rumen bacteria,Lactobacillus,bacillus,koala hair screw Ross Berry’s bacteria increased.The diversity and abundance of segmentedfilamentousbacteria,Clostridiumleptum,Prevotella and so onwere reduced.As for efficacy,high dosegroupwas much more apparent than low dosegroup.Coptis decoction group mainly increased Ackerman’s bacteria,butyric acid coccus and other bacteria,and the efficacy was lower than high and low dose of Tangshen capsulegroup.Berberine hydrochloride group mainly increased Ackerman’s bacteria.Conclusion1.Tangshen capsule of both high and low dose can significantly improve part of relative indicators to MS of the SD rats intervened with high-fat diet,and that of high dose shew better curative effect,indicating there’s a positive correlation between curative effect and the dose;With a more significant curative effect than berberine of higher dose in Berberine hydrochloride group and coptis of higher dose in Coptis decoction,it indicated the curative effect of Tangshen capsule was achieved through more targets and more methods.2.The intestinal microflora of SD rats model intervened by high-fat diet in this research changed significantly,and its characteristic was the significant reduce of the sample abundance and diversity;Firmicutes decreased significantly,Mollicutes and Elusimicrobia decreased significantly,and Bacteroidetes shew an increasing trend.Ackerman’s bacteria of the rats showed a decreasing trend,suggesting a failing ability of repairing intestinal barrier;beneficial bacteria shew an obviously decreasing trend,such as butyric acid bacteria and Ross berry strains;such opportunistic pathogens as Brausteria,Bacteroides eggerthii,Prevotella faecalis and coprococcus increased significantly.3.Tangshen capsule significantly increased the abundance and diversity of the intestinal microflora,meanwhile promoted repairing the recovery of intestinal barrier,increased the abundance of beneficial bacteria,reduced the kind and abundance of opportunistic bacteria.It intensively gathered Ackerman’s bacteria repairing the recovery of intestinal barrier,increased the abundance of such beneficial bacteria as Bacteroides,butyric acid bacteria,Ruminococcus,Lactobacillus,Lachnospira,Phascolarcto bacterium,and Ross bellerella,and decreased the kind and abundance of opportunistic bacteria,such as segmented filamentous bacteria,Clostridium leptum,and Pevotellaspecies.Compared with higher dose of berberine hydrochlorideand coptisdecoction with simplecoptis,after being intervened by Tangshen capsule,the structure of intestinal microflora shew a more apparent trend of increasing beneficial bacteria and reducing opportunistic bacteria and Ackermann’s bacteria majoring,in intestinal repairing gathered more apparently.Considering that the mode of action is related to coptis in compounds TCM or berberine hydrochloride,but more apparent and more efficient mode of action shows compounds TCM Tangshen capsule’s characters of more ingredients,more approaches and targets,also shows the efficiency,high efficiency,integrity and dominance. |
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