Analysis Of Characteristics Of Intestinal Microflora In Down's Syndrome Patients And Research On Regulation

Down's syndrome(Trisomy 21,DS)is a genetic disorder,which is usually associated with physical and mental defects.The incidence rate of Down's syndrome is0.1447% in neonates.With the increase of the age of pregnant women,the incidence rate of the disease is significantly increased.So far,very little is known about whether the symptoms are related to the intestinal flora of DS patients.In this study,the intestinal microflora compositions were measured between DS patients and healthy people.The effects of fecal and blood metabolites on DS patients were analyzed using metabonomics.Finally,the characteristic intestinal flora was reconstructed in sterile mice through flora transplantation.The data are displayed as follows:The V3-V4 region of the 16 S r RNA gene in 39 fecal samples(17 samples from DS patients and 22 samples from healthy people)were analyzed using Illumina miseq high-throughput sequencing technology and multivariate statistical method.Compared with the HC(healthy people)group,the relative abundance of Bacteroides,Bacteroidaceae and Lachnospiracea?incertae were significantly decreased in the DS group,whereas the relative abundance of Prevotella,Bacteroides,Faecalibacterium,Alloprevotella,Megasphaera and Dialister were significantly increased.We speculate that the difference of intestinal flora is related to the inflammation caused by the overexpression of SOD(Superoxide dismutase)gene on chromosome 21 in DS patientsAccording to the results of 16 S r RNA sequencing,we sequenced the metagenome of8 DS patients and 4 healthy people.Compared with the HC group,the relative abundance of Prevotella copri CAG164,Prevotella copri,Prevotella sp CAG386,Oscillibacter sp ER4 were significantly increased in the DS group.Based on GC-TOF-MS,the metabolic profiles of feces in HC and DS group were analyzed.Thirty-five different metabolites of feces were screened by multivariate statistical analyses(Twenty-one metabolites increased significantly and fourteen metabolites decreased significantly in DS patients).It was found that several inflammatory markers in the intestinal tract of DS patients were significantly up-regulated,among which methionine sulfoxide was up-regulated by 4.98 times and3-Hydroxybutyric acid was up-regulated by 3.36 times.Multiple up-regulated metabolites could aggravate the development of inflammation,increase intestinal membrane permeability and destroy energy metabolism.These key metabolites were significantly correlated with Prevotella sp CAG5226,Prevotella sp CAG520,Prevotella sp CAG386 and oscillibacter sp Er4.Significant difference of the metabolites are mainly mapped on multiple amino acid metabolic pathways,involving multiple neurotrophin-related metabolic pathways.It is speculated that these bacteria may affect the neural phenotype by destroying energy metabolism,promoting intestinal inflammation and increasing intestinal membrane permeability..Based on GC-TOF-MS technology,the metabolic spectrum of blood samples was analyzed in the HC and DS group.Twenty different metabolites were screened from blood by multivariate statistical analysis(Twelve metabolites increased significantly in DS patients and eight metabolites decreased significantly).Among the significant difference metabolites,we found that the metabolites associated with TCA cycle,such as Pyruvic acid,L-Malic acid,Fumaric acid and Oxalic acid,are accumulated significantly,especially the Fumaric acid was up-regulated by 5.6 times,indicating that the pathway of energy metabolism in the DS patients may be blocked.These metabolites were positively correlated with Prevotella sp CAG5226,Prevotella sp CAG520,Prevotella sp CAG386 and oscillibacter sp Er4,which confirmed our previous hypothesis that these kinds of bacteria destroyed the body's energy metabolism.The data of blood cytokine analyses showed that the concentrations of GM-CSF,IL-1?,IL-17 A,G-CSF,IL-8,MCP-1,IL-6,TNF-?,Fractalkine,IL-7,MIP-1?,MIP-1?,IL-1?,Anglogenin and Eotaxinin had a rising trend compared with the healthy people,and MIG and IL-10 were increased significantly,which further indicates that the inflammation in DS patients is enhanced..The effects of feces from DS patients and different strains on the behavior of mice were evaluated by the studies using sterile mouse model.The results showed that the percentage of sucrose solution intake,the total movement distance and the time in center were significantly reduced,whereas the immobility time was prolonged in the forced swimming test.It indicated that the characteristic bacteria of DS patients could induce the mice to have the phenotypes of depression and decreased exercise desire.These results indicate that these bacteria can affect the neurological functionss.Furthermore,the results showed that the levels of corticosterone,LPS and C-reactive protein(CRP)were significantly increased in the feces and different strains of DS patients.It is speculated that the flora may affect the neurophenotype by causing intestinal inflammation.In summary,there are significant differences in the intestinal flora,fecal metabolites and blood metabolites between the DS patients and healthy people,and the correlation between the fecal metabolites and blood metabolites of DS patients and the characteristic intestinal flora is obvious.We speculate that the overexpression of SOD1 gene on chromosome 21 of DS patients causes oxidative stress,raises intestinal reactive oxygen level,increasesintestinal strains Prevotella sp.and Oscillibacter sp.,induces inflammatory response and intestinal leakage,causes abnormal energy metabolism,and aggravates the development of neurophenotype.This provides a theoretical basis for the research of disease progression of DS patients and the improvement of symptoms by fecal bacteria transplantation in the future.