Study On The Western-uyghur Medicine Treatment Of Asthma With Abnormal Savda Syndrome Based On Intestinal Flora And Metabolomics Research

Objectives: Analysis intestinal flora and metaboloties of asthma with Abnormal Savda syndrome(AASS) before and after Western-Uyghur medicine treatment and Healthy comtrols(HC), study the clinical efficacy and mechanisim of Western-Uyghur medicine treatment of AASS. Methods: 1) Totally 75 AASS were enrolled in this study and divided them to treated group and control group according to patient`s willing, 38 cases of treated group and 37 cases of control group. The treated group was treated with western-uyghur medication. The control group was treated with routine western medicine. Estimated and took notes the the symptom score, ACT score and AQLQ score of each group, compaired them before treatment and after treatment, and also compaired them in the two groups. 2) 38 AASS were enrolled in this study and given western-uyghur medication, and divided them to before treatment group and after treatment group, and 30 healthy volunteers were set up as controls. Collecting fresh stool samples from those cases, comparatively examined the structure of the intestinal flora in these asthma patients by using Illumina Miseq sequencing based on the 16 Sr RNA V3-V4 region of bacteria to provide an in-depth analysis of status-related differences of intestinal flora between different status according to two medicine systems within small and large population. 3) 32 asthma patients with abnormal Savda syndrome were enrolled in this study and given western-uyghur medication, and divided them to before treatment group and after treatment group, and 30 healthy volunteers were set up as controls. Collecting plasma samples from those cases, the plasma 1H NMR spectra were analyzed using the orthogonal projection to latent structure with discriminant analysis(OPLS-DA) method with unit variance scaling. The discriminative significance of the metabolites was determined using the Pearson’s product moment correlation coefficient. Results: 1) After 1 months of treatment, the clinical curative effect of two groups of were 94.8% and 91.9% respectively, the difference was not statistically significant(P>0.05); compared with before treatment, asthma symptoms score and ACT score of two groups were improved, but after 33 days treatment, day and night symptoms score and ACT score of two groups were not statistically significant(P>0.05). Compared with before treatment, AQLQ score of two groups were improved(P<0.05). After 33 days treatment, the treatment group was significantly higher than the control group in terms of activity limitation, psychological status, response to stimulus, and care for their own health, the difference was statistically significant(P<0.05). 2) 21489, 20771 and 21460 sequences per sample were obtained form the before treatment group, the after treatment group and the healthy control group(HC), binned into a mean of 154±48, 176±51 and 180±48 operational taxonomic units(OTUs) in the 3 group. The ace index in 3 groups were 202±51, 226±57, 232±51; the Chao index in 3 groups were 193±50, 221±61, 225±54; the shannon index in 3 groups were2.99±0.53, 3.14±0.59, 3.12±0.59, thsimpson index in 3 groups were 0.12±0.07, 0.11±0.11, 0.11±0.06. Before treatment, OTUs, ACE index and Chao index of AASS group were reduced, the difference of OTUs, ACE index and Chao index is statistically significant(P≤0.05), but the lowerness of Shannon index and Simpson index had no statistical significance(P>0.05). After treatment compared with before treatment, OTUs, ACE index and Chao index were all increased, and Shannon Weaver index increased, Simpson index decreased, which OTUs and Chao index differences have statistical significance(P≤0.05), ACE index, Shannon index and Simpson index difference had no statistical significance(P>0.05). Using rarefaction curve, Chao1 index, Shannon index and abundance Rank curve analysis showed the sample data of each group can be qualified, which can satisfy the following analysis, and the basic reached the ideal sequencing depth, which can cover all the bacterial species. The bacterial community was dominated by 12 phyla and 147 genus, which were identified as the difference between the three groups. At the phylum level, The intestinal bacterial community diversity between the three groups was different. The dominant phyla of HC were Bacteroidetes(46.18%), Firmicutes(41.49%), Proteobacteria(5.88%), Fusobacteria(1.22%), Verrucomicrobia(0.72%), Unclassified(0.61%). The dominant phyla of AASS before treatment were Firmicutes(49.16%), Bacteroidetes(40.14%), Actinobacteria(5.00%), Proteobacteria(4.59%), Fusobacteria(0.51%). The dominant phyla of AASS after treatment were Firmicutes(45.30%), Bacteroidetes(41.56%), Actinobacteria(5.60%), Proteobacteria(4.79%), Verrucomicrobia(1.08%), Unclassified(1.01%). At the phylum level, compaired with the HC, the relative abundance of Firmicutes and Actinobacteria were increased in the AASS before treatment group, and in which the relative abundance of Bacteroidetes, Proteobacteria and synergistetes were decreased.And the difference of synergistetes has signifant difference(P≤0.05). Compaired with the AASS before treatment, the relative abundance of Bacteroidetes, Actinobacteria, Candidate_division_TM7_norank, Cyanobacteria were increased in AASS after treatment group, and in which the relative abundance of Firmicutes were decreased, and the difference of Candidate_division_TM7_ norank, Cyanobacteria has signifant difference(P≤0.05). After western-uyghur medicine treatment, the intestinal flora structure of AASS was similar with HC. At the genus level, The intestinal bacterial community diversity between the three groups was different. The dominant genus of HC were bacteroides(30.24%), faecalibacterium(3.64%), bifidobacterium(2.94%), C coli Shigella(2.44%), Alistipes(2.18%), blautia(1.24%), dialister(1.23%), dorea(1.23%), coprococcus(1.05%), barnesiella(0.51%), Eryspelotrichaceae_incertae_Sedis(0.46%). The dominant genus of of AASS before treatment were bacteroides(21.05%), bifidobacterium(4.56%), faecalibacterium(4.44%), C coli Shigella(2.68%), dialister(2.64%), Blautia(2.34%), coprococcus(1.51%), Catenibacterium(1.46), Alistipes(1.18%), dorea(0.85%), Eryspelotrichaceae_incertae_ Sedis(0.68%). The dominant genus of AASS after treatment were bacteroides(25.50%), enterococcus(2.20%), bifidobacterium(4.69%), faecalibacterium(4.67%), dialister(3.39%), Alistipes(1.77%), blautia(1.75%), C coli Shigella(1.68%), coprococcus(1.18%), akkermansia(1.08%), Eryspelotrichaceae_incertae_Sedis(0.79%), dorea(0.72%), barnesiella(0.44%). At the genus level, compaired with the HC, the relative abundance of bacteroides, Alistipes, dorea, bilophila, burkholderia and butyricimonas were decreased, and in which the relative abundance of catenabacterium, bifidobacterium, faecalibacterium, C coli Shigella, dialister, blautia, coprococcus and Eryspelotrichaceae_ incertae_Sedis were increased, and the difference of bilophila, burkholderia and butyricimonas has signifant difference(P≤0.05). Compaired with the AASS before treatment, the relative abundance of enterococcus, akkermansia, bacteroides, bifidobacterium, faecalibacterium, dialister, Alistipes, Eryspelotrichaceae_incertae_Sedis, barnesiella, acinetobacterium, chloroplast_norank, comamonas, flavonifracter and burkholderia were increased, and in which the relative abundance ofblautia, C coli Shigella, coprococcus and dorea were decreased, and the difference of acinetobacterium, chloroplast_norank, comamonas and burkholderia has signifant difference(P≤0.05). The bacterial microbiota composition of the three groups could not be distinguished by PCA, NMDS and PCo A, the Unifra diversity of NMDS and Beta diversity analysis. The composition of the intestinal flora in the three groups was different from that of the Heatmap, LEf SE, ANOSIM analysis. 3) Compared with the healthy controls, the asthma patients with abnormal Savda syndrome before treatment had uniformly correlative low levels of leucine, lactic acid, 1-methy histidine, lipidtemns such as low-density lipoprotein(LDL)(P<0.05), but had significantly high levels of valine, alanine, acetic acid, citric acid, glycine, tyrosine, α-glucose, β-glucose, histidine(P<0.05). Compared with the before treatment group, the after treatment group patients had low levels of acetic acid, citric acid, tyrosine, 1-methy histidine, α-glucose, β-glucose(P < 0.05), but had significantly high levels of isoleucine, lactic acid, glycoprotein, histidine, lipidtemns such as low-density lipoprotein(LDL)(P<0.05). Conclusion: 1) Western-uygur medication can improve patients day and night symptom control, ACT, AQLQ and clinical efficacy, especially improve patients quality of life including the activities of limited, psychological conditions, the reflection of the stimulus, for their own health care. And then improves the asthma patients mental health and emotional well-being. 2) At the phylum and genus level, the structure and diversity of intestinal flora of AASS was altered, the number of OTUs, the relative abundance and diversity were all decreased. The intestinal flora has relationship with AASS. 3) Western-uygur medication can change the structure and diversity of intestinal flora of AASS at the phylum and genus level, it increase the number of OTUs and the relative abundance and diversity of AASS. Western-uygur medication would reduce the harmful bacteria increase the beneficial bacteria,correct the structure of intestinal flora imbalance, maintain the ecological balance of micro organism, and adjust the immune function, and then play a role in the treatment of asthma and improves the asthma patients mental health and emotional condition. This is consistent with the previous study of the neural endocrine immune network mechanism of asthma.It provides a thought and foundation for the study of the microbial-gut-brain axis and the gut-brain-endocrain axis of the AASS. Regulation of intestinal flora may be one of the therapeutic mechanism of Western-uygur medication of AASS. 4) The metabolites in the AASS were significantly different from those in the healthy’s, many metabolimic pathways such as histidine-histamine conversion, TCA cycle, energy metabolism, fatty acid metabolism and amino acid metabolism were altered, suggesting that the changed metabolimic pathways have the close relation to AASS by inducing inflammation of the airway. Western-uyghur medication has significant regulatory effects on plasma metabolites of AASS, it markedly regulates the contents of final metabolites. Readjusting the altered metabolimic pathways may be another one of the therapeutic mechanism of Western-uygur medication of AASS.