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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 王敏瑩(Man-Ying Wong) | |
| dc.contributor.author | Mei-Chih Sun | en |
| dc.contributor.author | 孫美芝 | zh_TW |
| dc.date.accessioned | 2021-06-13T06:40:55Z | - |
| dc.date.available | 2005-08-04 | |
| dc.date.copyright | 2005-08-04 | |
| dc.date.issued | 2005 | |
| dc.date.submitted | 2005-07-29 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/35100 | - |
| dc.description.abstract | 本實驗的目的是觀察牙周致病菌Porphyromonas gingivalis、Tannerella forsythensis〈舊名為Bacteroides forsythus〉、及其所萃取出的脂多醣內毒素，在體外對人類單核球衍生的樹突狀細胞，產生發炎前期細胞激素的影響。單核球衍生樹突狀細胞是從20個健康受試者之新鮮周邊血中分離並培養，分別以熱處理過的細菌、及不同濃度的脂多醣內毒素刺激24小時後，利用流式細胞儀觀察樹突狀細胞表型的改變，以ELISA方式測量分泌細胞激素產量、及反轉錄同步定量聚合酶連鎖反應偵測激素的基因表現，E. coli及其LPS在本實驗則為正向控制組。實驗結果顯示，體外經P. gingivalis、T. forsythensis及其LPS刺激樹突狀細胞，細胞的型態會改變、其中表面第1型及第2型MHC胜肽類複合物〈HLA-DR〉的表現及穩定度提昇，成熟化的記號〈CD83〉增加、共同刺激分子〈CD80、CD86〉的表現量也上昇。細胞激素測定方面，發現經細菌及其LPS刺激後，細胞激素的產量，與對照組相較，均顯著提昇，IL-10大於IL-12p40及IL-12p70，全菌的反應則較脂多醣內毒素明顯，皆具統計上的差異；樹突狀細胞受3種細菌的脂多醣內毒素刺激，分泌的細胞激素產量卻沒有明顯差別，IL-10具有劑量依存效應，但不具有統計上的意義；以E. coli刺激時，得到的IL-10、IL-12p70產量，顯著高於P. gingivalis和T. forsythensis，但在細胞激素分泌型式的比較上，沒有明顯的改變。本實驗所偵測到的IL-12，多為p40蛋白型式，IL-12p70的分泌量較低。細胞激素基因表現結果與ELISA結果類似，3種細菌刺激樹突狀細胞3小時後，IL-10 mRNA的表現最強，次為IL-12p40、IL-12p35最低，IL-12p35的mRNA，僅在以E. coli刺激時，有較顯著的提昇。由實驗結果得知，P. gingivalis及T. forsythensis感染寄主時，除了脂多醣內毒素外，其他細菌成份可能也在活化樹突狀細胞、及產生細胞激素的過程中參與反應。此外，牙周致病菌早期侵犯宿主時，可能因為較弱的免疫刺激能力、並能在初期免疫反應促使樹突狀細胞分泌高量抗發炎細胞激素，並誘導具有免疫抑制性質的調節性T淋巴球活化，使牙周感染區域形成慢性發炎的病灶，以避免遭到免疫細胞清除的命運。 | zh_TW |
| dc.description.abstract | The present study was to investigate the effect of periodontal pathogens Porphyromonas gingivalis, Tannerella forsythensis (Bacteroides forsythus) and their lipopolysaccharides (LPS) on the production of cytokines from human monocyte-derived dendritic cells (MDDC). MDDC were isolated from 20 healthy subjects and co-cultured with the heat treated whole bacteria or their LPS. The phenotypic changes were evaluated with flow cytometry, the cytokine profiles were determined by ELISA, and the mRNA expression of cytokine was analysis by real-time RT-PCR. E. coli and its LPS were used as positive control. The results showed that in vitro monocyte-derived DCs pulsed with P. gingivalis 33277, T. forsythensis 338 and their LPS would change the morphology and increase the surface expression and stability of MHC class I and class II-peptide complexes (HLA-DR), upregulated the maturation marker (CD83) and adhesion and co-stimulatory molecules(CD80, CD86). On cytokine detection, IL-10 production was higher than IL-12p40 and IL-12p70 in all groups. The response of whole bacteria was stronger than that of the LPS. Lipopolysaccharides of different bacteria induced similar IL-10 and IL-12 profiles from DCs with a dose dependent effect on IL-10. DCs pulsed with E. coli produced more IL-10 and IL-12 than P. gingivalis and T. forsythensis, without significant change of cytokine profile patterns. Most of IL-12 produced was in p40 form, rather than IL-12p70. IL-10 mRNA was highly expressed on DC after stimulation with the bacteria, but IL-12p35 mRNA level was elevated clearly only when DC pulsed with E. coli. The present study implicated that the components of P. gingivalis and T. forsythensis, other than LPS, maybe responsible for the interaction of the bacteria and the dendritic cells. The periodontal pathogens may induce the anti-inflammatory cytokine production to suppress the function of the dendritic cells in the innate immune response, and direct naïve T cells to regulatory T lymphocytes with immunosupressive properties. P. gingivalis and T. forsythensis with weaker immunostimulatory activity than E. coli may contribute to the induction of chronic inflammation at local lesion of periodontal infection. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-13T06:40:55Z (GMT). No. of bitstreams: 1 ntu-94-R91422001-1.pdf: 2771019 bytes, checksum: b0d0bf95f593b1cb0a0c40e7f7824b7e (MD5) Previous issue date: 2005 | en |
| dc.description.tableofcontents | 中文摘要 1
Abstract 3 文獻回顧 5 細菌在牙周病的致病角色 6 1.細菌與牙周致病因子相關性 6 引發組織破壞的機轉 7 2.牙周致病菌 8 Porphyromonas gingivalis抗原特性 10 Tannerella forsythensis抗原特性 13 宿主對牙周致病菌入侵的免疫反應 16 1. 初期免疫反應〈Innate immunity〉 16 牙齦組織的保護功能 16 牙周組織的初期免疫反應 17 2. 後繼免疫反應〈adaptive immunity〉 19 牙周組織的後繼免疫反應 19 3. T淋巴球控制後繼免疫反應的功能 20 牙周病免疫學對T淋巴球的研究 22 Th1與Th2免疫反應在牙周疾病進展的理論分歧 25 I. Seymour & Gemmel 牙周病進展模型假說的相關證據 26 II. Ebersole & Taubman牙周病進展模型假說的相關證據 28 牙周病對初期免疫反應引導T淋巴球分化的影響 31 樹突狀細胞與牙周疾病的相關性 33 1. 初期免疫反應中的樹突狀細胞 33 2. 樹突狀細胞調節T淋巴球反應機制 34 IL-12與引導初期及後繼免疫反應的功能 36 IL-10對抗原呈現細胞及免疫反應的影響 38 3. 牙周組織中的樹突狀細胞 40 在牙周組織的分佈位置 40 與牙周疾病的相關性 41 牙周致病菌對樹突狀細胞的影響 43 過去牙周致病菌與樹突狀細胞相關研究的缺失 44 實驗目的 46 實驗材料及方法 47 1. 細菌培養與脂多醣內毒素〈LPS〉的製備 47 2. 受試者樣本選取 48 3. 人類單核球衍生樹突狀細胞〈MDDCs〉的純化與培養 49 4. 細菌、脂多醣內毒素刺激單核球衍生樹突狀細胞產生細胞激素 50 5. 流式細胞儀測定樹突狀細胞成熟化表徵 51 6. IL-10、IL-12p40及IL-12p70細胞激素的測定 52 7. 反轉錄─同步定量聚合酶連鎖反應〈RT-real time PCR〉 53 單核球衍生樹突狀細胞mRNA的分離 53 mRNA反轉錄反應〈Reverse transcriptase reaction〉 54 同步定量聚合酶連鎖反應〈Real time-PCR〉 55 8. 統計分析 57 實驗結果(Results) 58 1.單核球衍生樹突狀細胞激活後的型態改變 58 2.流式細胞儀的訊號表現 59 純化CD14+單核球的表型 59 未成熟樹突狀細胞表面的訊號表現 59 成熟樹突狀細胞表面的訊號表現 60 3. ELISA偵測細胞激素濃度的結果 63 IL-10的濃度變化 64 IL-12的濃度變化 65 樹突狀細胞分泌IL-10與IL-12p70的比例 67 4.反轉錄同步定量聚合酶連鎖反應的結果 68 β-actin mRNA的表現 68 Realtime-PCR定量的結果 68 標定基因電泳的結果 70 討論(Discussion) 71 抗原型式與細胞激素產量的差異 72 牙周致病菌刺激樹突狀細胞分泌的早期激素 74 與過去研究相較樹突狀細胞對T淋巴球影響 78 IL-12 80 體外培養樹突狀細胞及組織學 82 結論(Conclusion) 87 圖表 88 參考書目〈References〉 114 圖表目錄 圖1：CD14+單核球的型態 89 圖2：〈100X〉單核球衍生樹突狀細胞型態改變 90 圖3：〈400X〉單核球衍生樹突狀細胞型態改變 91 圖4：〈1000X〉單核球衍生樹突狀細胞型態改變 92 圖5：單核球CD14+的表現 93 圖6：培養6天之單核球表現CD14的情形 93 圖7：未成熟樹突狀細胞的表型： 94 圖8：牙周致病菌刺激DC表型的改變 95 圖9：細菌活化CD83表現的改變 96 圖10：LPS活化CD83表現的改變 97 圖11：細菌活化CD80表現的改變 98 圖12：LPS活化CD80表現的改變 99 圖13：細菌活化CD86表現的改變 100 圖14：LPS活化CD86表現的改變 101 圖15：細菌活化HLA-DR表現的改變 102 圖16：LPS活化HLA-DR表現的改變 103 TABLE I.：牙周致病菌活化樹突狀細胞分泌激素的產量 104 圖17：樹突狀細胞激素型式的比較 104 圖18：細菌和LPS種類對 IL-10產量的影響 105 圖19：抗原型式對IL-10分泌的影響 105 圖20：細菌和LPS種類對IL-12p40產量的影響 106 圖21：抗原型式對IL-12p40分泌的影響 106 圖22：細菌和LPS種類對IL-12p70產量的影響 107 圖23：抗原型式對IL-12p70分泌的影響。 107 圖24：樹突狀細胞分泌IL-12p40與p70的比例 108 圖25：樹突狀細胞IL-10與IL-12p70產量的比例………………………………108 圖26：樹突狀細胞Β-actin的表現 109 圖27：cDNA濃度與ΔCT 109 TABLE II：Validation Test 110 圖28：Validation Test (GAPDH與IL-10、IL-12p40、p35) 110 圖29：樹突狀細胞GAPDH的表現 111 TABLE III：樹突狀細胞IL-10, IL-12p40 & p35的mRNA表現 111 圖30：樹突狀細胞IL-10、IL-12p40、及IL-12p35基因表現倍數差異 112 圖31：GAPDH電泳結果 112 圖32：IL-10 mRNA電泳的結果 113 圖33：IL-12p40 mRNA電泳的結果 113 圖3：IL-12p35 mRNA電泳的結果 113 | |
| dc.language.iso | zh-TW | |
| dc.title | 牙周致病菌Porphyromonas gingivalis、Tannerella forsythensis
對人類單核球衍生樹突狀細胞產生細胞激素的影響 | zh_TW |
| dc.title | Effects of periodontal pathogens Porphyromonas gingivalis and Tannerella forsythensis on cytokine production of human monocyte-derived dendritic cells. | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 93-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 江伯倫(Bor-Luen Chiang) | |
| dc.contributor.oralexamcommittee | 侯連團(Lein-Tuan Hou),傅鍔 | |
| dc.subject.keyword | 樹突狀細胞,脂多醣內毒素,細胞激素,T淋巴球,IL-10,IL-12, | zh_TW |
| dc.subject.keyword | Porphyromonas gingivalis,Tannerella forsythensis,dendritic cell,LPS,cytokine,T cell,IL-10,IL-12, | en |
| dc.relation.page | 136 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2005-08-01 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 臨床牙醫學研究所 | zh_TW |
| 顯示於系所單位： | 臨床牙醫學研究所 | |
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