| Objective:Containing methionine feed through to the establishing of homocysteine in the rat model of blood lipid index testing and intestinal flora the sequencing analysis of 16 s r DNA high variable area,explore high homocysteine levels,and the relationship between blood lipid changes and intestinal microecological bacteria,for clinical research on homocysteine blood disease pathogenesis and homocysteine and the relationship between intestinal microecological bacteria animal experimental basis,and to adjust the Angle of intestinal microecological bacteria from the treatment to prevent high homocysteine levels,provide theoretical basis.Methods:Twenty-two clean grade 8-week old male SD rats were selected and randomly divided into two groups,namely the normal control group and the high methionine group.Normal control group rats were fed with standard feed,and high methionine group rats were fed with standard feed added with 1.7% methionine.After the establishment of hyperhomocysteinemia SD rats model,blood samples were collected to detect common clinical biochemical indexes.Meanwhile,feces were collected to extract fecal DNA.16 S r DNA sequencing analysis technology was applied to analyze the Alpha diversity,Beta diversity and relative abundance of intestinal microflora in the two groups of rats.Results:1.Establishment of homocysteine rats modelAfter 4 weeks of methionine feeding,the homocysteine level of SD rats in the control group was 7.3±1.2μmol/L,and that of SD rats in the high methionine group was42.4±21.6μmol/L.The t test of the two groups of independent samples showed that P<0.05,indicating that the serum homocysteine level of the two groups of rats was significantly different,and the serum homocysteine level of the high methionine group of rats was significantly increased,suggesting that the model of hyperhomocysteinemia in SD rats had been established.2.Lipid metabolism of homocysteine ratsAfter 4 weeks of methionine feeding,serum lipid indexes of SD rats in the two groups were tested by t test of independent samples from the two groups,and there was no significant difference in serum triglyceride,cholesterol and LDL cholesterol levels between the two groups.There was a significant difference in serum HDL cholesterol between the two groups of SD rats,and the serum HDL cholesterol level in the high methionine group was 0.62± 0.09mmol/L,significantly lower than that in the control group.3.Changes of intestinal microflora in homocysteine rats3.1 Analysis of Alpha diversityCoverage analysis(sequencing depth index)showed that the two groups of sample libraries had the larger coverage,with the higher probability of sequence being detected in the samples and the lower probability of not being detected.The sequencing results can represent the real distribution of microorganisms in the samples.Simpson index indicated that hypermethionemia may reduce the diversity of microbial community.Chao1 index may showed that the hyperhomocysteinemia has no impact on the intestinal microflora abundance of single sample.3.2 Analysis of Beta diversityPrincipal component analysis(PCA),principal coordinate analysis(PCo A)and non-metric multidimensional analysis(NMDS)showed that the constitution of intestinal microflora in the two groups was significantly different.Moreover,Anosim analysis indicated that the composition difference of intestinal microflora between the two groups was greater than that inside each group.3.3 Analysis of relative abundanceLactobacillaceae has a high proportion of relative abundance in the two groups,and the relative abundance in the two groups is significantly different.The relative abundance of Lactobacillaceae in the high methionine group is 8.83%,which is lower than that in the control group,which is 19.34%.In addition,it was also found that the relative abundance of Christensenellaceae in the two groups was significantly different.The relative abundance of Christensenellaceae in the high-methionine group was 2.19%,which was higher than that in the control group which is 0.82%.The relative abundance of unidentified＿Clostridiales in the two groups was significantly different,1.40% in the high methionine group,which was higher than that in the control group which is0.94%.Lactobacillus has a high proportion of relative abundance in the two groups,and the relative abundance in the two groups is significantly different.The relative abundance in the high methionine group is 8.77%,which is lower than that in the control group,which is 19.34%.In addition,the relative abundance of Lactobacillus,Tyzzerella,unidentified＿Muribaculaceae,Odoribacter and Alloprevotella in the high-methionine group was significantly lower than that in the control group.The relative abundance of Negativibacillus,Butyrivibrio,unidentified＿Clostridiales,Dubosiella,Marvinbryantia,Parabacteroides,unidentified＿Christensenellaceae,Allobaculum,Turicibacter,Harryflintia,Pediococcus,unidentified＿Erysipelotrichaceae,Caproiciproducens,Enterococcus,Holdemania and Faecalitalea in the high methionine group was significantly higher than that in the control group.Conclusion:1.A rat model of high homocysteine was established after feeding SD rats with 1.7%methionine for 4 weeks.2.Hyperhomocysteinemia can cause changes in intestinal flora.3.There was a significant difference in the relative abundance of lactobacillus bacteria between the homocysteinemia group and the normal control group. |
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