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完整後設資料紀錄
DC 欄位值語言
dc.contributor.advisor謝淑貞
dc.contributor.authorHsin-Yi Chiuen
dc.contributor.author邱欣怡zh_TW
dc.date.accessioned2021-06-16T23:53:36Z-
dc.date.available2012-07-27
dc.date.copyright2012-07-27
dc.date.issued2012
dc.date.submitted2012-07-19
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dc.identifier.urihttp://tdr.lib.ntu.edu.tw/jspui/handle/123456789/65608-
dc.description.abstract流行病學調查指出,過去二十年間,「異位性皮膚炎」的發生率及普及率在已開發國家大幅提升。已知此疾病的成因除了受基因調控外與西化的生活型態有密切關係,環境因子更是誘發此疾病的關鍵。研究顯示,體內正常的微生物菌叢在免疫調控與平衡上扮演重要的角色。由於衛生環境的改善、飲食習慣的改變以及抗生素廣泛使用等因素,大幅降低接觸外在微生物菌叢的機會,因此也提高了罹患免疫失衡疾病的機率。益生菌的免疫調節能力在動物與人體實驗上已被證實,但其調控的機轉仍待研究。本次報告內容是探討納豆菌(Bacillus subtilis var. natto)如何透過調控體內的免疫反應進而達到抑制臨床上犬異位性皮膚炎的症狀。本實驗室透過蛋白質體學的分析,發現犬隻在攝入108cfu/ml納豆菌(Bacillus subtilis var. natto),可誘發免疫細胞活化,進而抑制體內載脂蛋白A-I(Apolipoprotein A-I)的表現。藉由較低濃度的載脂蛋白A-I,可進一步維持或放大納豆菌所引發的促發炎現象,而達到抗過敏的效果。研究結果顯示納豆菌(Bacillus subtilis var. natto)可刺激免疫細胞活化進而影響載脂蛋白A-I的表現,並且分泌相關免疫調控蛋白來維持適當之免疫發炎反應以達到對急性期異位性皮膚炎之治療效果。zh_TW
dc.description.abstractAtopic dermatitis (AD) is one of the most common skin diseases that affects children and adults worldwide and its prevalence has been steadily increasing during the past decades. According to the hygiene hypothesis, the decreased exposure of microorganism results in altered T helper cells balance, increasing prevalence and severity of AD. Therefore, probiotics, with their important immune-regulatory potential, has been regarded as a novel strategy for the intervention of atopic disease. In this study, the probiotics, Bacillus subtilis var. natto, has been used for alleviating the symptoms of AD in clinical canine patients. After more than 3 weeks of treatment, the plasma samples were withdrawn from the dogs with significant clinical improvement to execute the proteomics analysis. After the statistical and comparative analysis, we have pinpointed that the down-regulation of apolipoprotein A-I (apoA-I), might get involved in the molecular mechanism mediated by Bacillus subtilis var. natto. Our studies have demonstrated that Bacillus subtilis var. natto could regulate the expression of apoA-I by its immunostimulatory activities and the down-regulation of apoA-I might play roles in the enhancement of its stimulatory effects. Furthermore, we also identified some immunoregulatory proteins, anexin-1, galectin-3 and thromospondin-1 might contribute to anti-AD effects supported by probiotics.en
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Previous issue date: 2012
en
dc.description.tableofcontentsCHAPTER 1 Literature Reviews 1
1.1 Atopic dermatitis 1
1.1-1 Introduction 1
1.1-2 Pathophysiology 6
1.1-3 Managements 9
1.2 Canine Atopic Dermatitis 11
1.2-1 Introduction 12
1.2-2 The similarity between canine AD and human AD 13
1.2-3 Management 16
1.3 Probiotics
1.3-1 Introduction 17
1.3-2 Bacillus subtilis var. natto 19
CHAPTER 2 Materials and Methods 20
2.1 Clinical canine AD experiment 20
2.1-1 Experiment design 20
2.1-2 Experimental material 20
2.1-2.1 Canine subjects recruitment 20
2.1-3 Experimental method 23
2.1-3.1 Probiotics treating protocol 23
2.1-3.2 Plasma specimen collection 23
2.2 Plasma proteomics analysis 24
2.2-1 Experiment design 24
2.2-2 Experimental material 24
2.2-2.1 Sample preparation and isolation 24
2.2-3 Experimental method 25
2.2-3.1 Quantitative proteomics analysis 25
2.2-3.2 Post-translational modification proteins analysis 28
2.2-3.3 Mass-spectrometry and database search 28
2.2-3.4 Comparative analysis and pathway mapping 28
2.3 In Vitro cell model system 30
2.3-1 Experiment design 30
2.3-2 Experimental material 30
2.3-2.1 Preparation of heat-killed microorganism 30
2.3-2.2 RAW264.7 macrophages cells culture 31
2.3-2.3 Preparation of conditioned media for nitric oxide assay 32
2.3-2.4 Preparation of serum-free conditioned media 32
2.3-3 Experimental method 33
2.3-3.1 Luciferase Assay 33
2.3-3.2 Nitric oxide (NO) Assay 34
2.3-3.3 TNF-α ELISA assay 35
2.3-3.4 Preparation of conditioned media for secretome analysis 36
CHAPTER 3 Results and Discussion 38
3.1Clinical canine AD dogs experiment 38
3.1-1 Clinical evaluation and case selection 38
3.1-2 Isolation of up-regulated proteins after probiotics treatment 39
3.1-3 Change of proteins expression profiles after probiotics treatment 40
3.1-4 PTMs proteins analysis and mapping 41
3.1-5 Hypothesized mechanisms 42
3.2 Validation of hypothesized mechanism in cell culture system 44
3.2-1 Murine macrophage cell model establishment 45
3.2-2 Pro-inflammatory response of probiotics 47
3.2-3 Analysis the cells immune responses under serum free condition 49
3.2-4 Proteomics analysis of conditioned medium 52
3.2-5 Identification of immunoregulatory proteins 53
CHAPTER 4 Conclusion 57
CHAPTER 5 Discussions 57
CHAPTER 6 References 62
Appendix
Figure 1. The pathogenesis of atopic dermatitis
(Chen et al., 2004) 8
Figure 2. General approaches of quantitative proteomics (Zhu et al., 2009) 27
Figure 3. Comparative analysis of proteins expression in 5 canine AD patients 29
Figure 4. Quantification analysis of proteins expression in 5 canine AD patients 40
Figure 5. Relative COX-2 gene expression profile 48
Figure 6. Nitrite (NO) induction profile 48
Figure 7. Relative iNOS gene expression profile under serum free condition 50
Figure 8. NO induction profile under serum free condition 51
Figure 9. TNF-α secretion profile under serum free condition 51
Figure 10. Quantitative analysis of B. subtilis var. natto-treated conditioned medium 52
Table 1. Clinical features of atopic dermatitis
(Leung et al., 2007) 3
Table 2. Similarities between canine AD and human AD (Marsella et al., 2009) 12
Table 3. The table of CADESI-03 scoring scheme
(Olivry et al., 2007) 22
Table 4. The clinical conditions of atopic dogs after probiotics treatment 38
Table 5. Ten differentially expressed protein 41
Table 6. Differentially expression of inflammation-associated proteins in conditioned medium 56
dc.language.isoen
dc.subject納豆菌zh_TW
dc.subject載脂蛋白A-Izh_TW
dc.subject免疫調控蛋白zh_TW
dc.subject異位性皮膚炎zh_TW
dc.subject蛋白質體學zh_TW
dc.subjectBacillus subtilis var. nattoen
dc.subjectapolipoprotein A-Ien
dc.subjectproteomicsen
dc.subjectatopic dermatitisen
dc.subjectimmunoregulatory proteinsen
dc.title益生菌在犬異位性皮膚炎之機制探討zh_TW
dc.titleThe Mechanism Study of Probiotics on Canine Atopic Dermatitisen
dc.typeThesis
dc.date.schoolyear100-2
dc.description.degree碩士
dc.contributor.oralexamcommittee廖辰中,羅翊禎,陳念榮
dc.subject.keyword異位性皮膚炎,納豆菌,蛋白質體學,載脂蛋白A-I,免疫調控蛋白,zh_TW
dc.subject.keywordatopic dermatitis,Bacillus subtilis var. natto,proteomics,apolipoprotein A-I,immunoregulatory proteins,en
dc.relation.page77
dc.rights.note有償授權
dc.date.accepted2012-07-19
dc.contributor.author-college生物資源暨農學院zh_TW
dc.contributor.author-dept食品科技研究所zh_TW
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