장내 미생물 네트워크 조절을 통한 궤양성 대장염 완화 : 동물 모델과 인간 연구의 통찰 = Modulating Gut Microbiota Networks to Mitigate Ulcerative Colitis : Insights from Animal Models and Human Studies

목차 (Table of Contents)

• ABSTRACT . ⅰ
• Ⅰ. Introduction . 1
• Ⅱ. Research Background . 7
• 1. UC . 7
• 2. Cognitive impairment induced by UC-8 . 8
• 3. Association of UC with other diseases 10
• 4. Metagenomics and meta-analysis 14
• 5. Microbial metabolic network reconstruction and protein homology analysis 17
• 6. Ruminococcus gnavus (R. gnavus) . 20
• 7. Bacteroides uniformis (B. uniformis) . 22
• 8. Anaerostipes hadrus (A. hadrus) 23
• 9. C. difficile . 24
• 10. Enterotypes 27
• 11. Bile Acids 29
• 12. SCFAs . 31
• 13. Inulin (Inu) 35
• 14. GOS . 36
• Ⅲ.Experimental Methods 38
• 1. Meta-analysis of UC 38
• a. Data collection and preprocessing . 38
• b. Inclusion criteria 38
• c. Metagenomic data analysis and downstream analysis 46
• d. Deep Neural Network (DNN) and SHapley Additive exPlanations (shap) 44
• e. Network analysis and prediction of gut microbiome functions 46
• 2. Microbial experiments . 47
• a. Bacterial strains and media preparation 47
• b. Polysaccharide substrate, carbon source utilization, and metabolite prediction . 48
• c. Carbohydrate utilization and bacterial growth assessment. 50
• d. Bacterial co-culture. 51
• (1) Co-culture of B. uniformis and A. hadrus. 52
• (2) Co-culture with C. difficile. 52
• e. Bacterial DNA, RNA extraction and quantitative PCR 53
• f. SCFA and bile acid detection in culture supernatant 55
• 3. Cell experiments 58
• a. Cell culture and co-incubation with culture supernatant 58
• b. Caco-2 cell mRNA expression. 59
• c. Caco-2 cell western blot . 60
• d. Caco-2 cell ZO-1 immunofluorescence staining . 62
• e. Caco-2 cell toluidine blue staining 63
• 4. Animal experiments . 63
• a. Chronic colitis mouse model induced by dextran sodium sulfate (DSS) 63
• (1) Animal management 63
• (2) Animal grouping, diet composition, and microbial feeding . 64
• (3) Chronic colitis and CDI induction and evaluation . 67
• b. Cognitive ability experiments 68
• (1) Y-maze test 68
• (2) Novel object recognition test (ORT). 69
• c. Gut barrier permeability assay 70
• d. Histopathological analysis . 71
• e. Metabolomic analysis. 75
• (1) SCFA gas chromatography analysis . 75
• (2) Bile acid high-performance liquid chromatography analysis 76
• f. Molecular biological analysis 76
• (1) Protein western blot . 76
• (2) Real-time quantitative PCR 77
• g. Metagenomic sequencing and analysis 77
• (1) Fecal DNA extraction and 16S rRNA amplification 77
• (2) Metagenomic analysis. 79
• 5. Statistical analysis. 80
• Ⅳ. Experimental results . 81
• 1. Meta-analysis of UC 81
• a. Study selection. 81
• b. Enterotype and biodiversity analysis. 86
• c. DNN classification model construction and performance 93
• d. Shap interpreter and network analysis in the total cohort. 96
• e. Shap interpreter and network analysis in the ET-L cohort. 103
• f. Shap interpreter, network analysis, and picrust functional prediction in the ET-B cohort. 109
• g. ET-C enterotype network analysis 118
• 2. Meta-analysis of UC 124
• a. Analysis of carbohydrate utilization by microorganisms 125
• b. Results of B. uniformis and A. hadrus co-culture . 133
• c. Inhibitory effects of B. uniformis and A. hadrus on C. difficile 141
• d. Analysis of microbial bile acid metabolism using protein homology . 146
• 3. Results of cell experiments 150
• a. RT-PCR results 152
• b. Western blot results 157
• c. Trouidin staining 160
• 4. Results of animal experiments . 163
• a. Chronic Colitis Induced by Low-Molecular-Weight DSS and CDI Exacerbates Colitis Symptoms 166
• b. Mild Colitis Induced by DSS Affects Cognitive Function 173
• c. PCR and western blot results. 176
• d. Intestinal permeability detection . 192
• e. Histopathological detection 194
• (1) Brain tissue staining and evaluation . 194
• (2) Colon tissue staining and evaluation 197
• f. Chromatographic analysis of SCFAs and BAs in mouse colon feces 207
• g. Gut microbiome analysis 216
• Ⅴ. Discussion . 234
• Ⅵ. Conclusion . 262
• Reference 266
• Abbreviation . 291