趨化激素（chemokines）在氣喘的發病機制中扮演著重要的角色。芝麻素（sesamin）是一種從芝麻種子中純化出來的植物雌激素，在一些研究中發現與抗發炎有關，但芝麻素對氣喘相關的趨化激素是否有影響仍屬未知，本研究澄清芝麻素對人類單核球細胞表現干擾素γ誘導蛋白10（interferon-γ-inducible protein-10，IP-10/CXCL10）、巨噬細胞衍生趨化激素（macrophage-derived chemokine，MDC/CCL22）、生長相關癌基因α（growth-related oncogene-α，GRO-α/ CXCL1）和腫瘤壞死因子α（tumor necrosis factor-α，TNF-α）的影響。首先以芝麻素對細胞進行前處理，之後再以細菌脂多醣（lipopolysaccharide，LPS）刺激，利用酵素免疫分析法（ELISA）測定IP-10、MDC、GRO-α、TNF-α的濃度。為確認其調控機制，進行細胞表面受體之拮抗劑及細胞內路徑抑制劑、西方墨點分析（Western blot）和染色質免疫沉澱（chromatin immunoprecipitation，ChIP）等研究。結果顯示芝麻素抑制了MDC的表現，但對IP-10，GRO-α和TNF-α的表現沒有影響。給予雌激素受體（estrogen receptor，ER）拮抗劑和過氧化物酶體增殖物激活受體（peroxisomal proliferators-activated receptor，PPAR）-α拮抗劑會使被抑制的MDC表現回復。芝麻素可抑制細胞絲裂原活化蛋白激酶（mitogen-activated protein kinase，MAPK）中p38和核因子κB（nuclear factor κB，NFκB）中p65的磷酸化，亦可抑制組蛋白H3/H4在MDC啟動子區域的乙烯化。芝麻素可藉由通過ER/ PPAR-α，MAPK-p38和NFκB-p65的路徑抑制MDC的表現和表觀基因調控而抑制氣喘的發病。

Certain chemokines play important roles in asthma. Sesamin has anti-inflammatory effects. Whether sesamin modulates asthma-related chemokines is unknown. We investigated the effects of sesamin on interferon-γ-inducible protein-10 (IP-10/CXCL10), macrophage-derived chemokine (MDC/CCL22), growth-related oncogene-α (GRO-α/CXCL1) and tumor necrosis factor (TNF)-α expression in human monocytes. Cells were pretreated with sesamin and then stimulated with lipopolysaccharide (LPS). IP-10, MDC, GRO-α and TNF-α were measured by ELISA. Mechanisms were investigated by receptor antagonists, pathway inhibitors, Western blotting and chromatin immunoprecipitation. Sesamin suppressed the expression of MDC but had no effects on IP-10, GRO-α and TNF-α expression. The suppressive effect was reversed by the estrogen receptor (ER) and peroxisomal proliferators-activated receptor (PPAR)-α antagonists. Sesamin suppressed phosphorylation of mitogen-activated protein kinase (MAPK)-p38 and nuclear factor κB (NFκB)-p65. Sesamin suppressed histone H3/H4 acetylation in MDC promoter region. Sesamin may suppress asthmatic inflammation by suppressing MDC expression via the ER/PPAR-α, the MAPK-p38 pathway and the NFκB-p65 pathway and the epigenetic regulation.

論文審定書……....…………………………………………….…...…...…i
致謝……………………………………..………..........…………...……..ii
中文摘要……………………………………………………….…....……iv
英文摘要……………………………………..……………………......…..v
第 一 章 前言…………………………………………………..……....1
1.1 簡介………………………………………………………......…..1
1.2 單核球細胞（monocyte）…………………………………..…..2
1.3 芝麻素（sesamin）……………………………………..……….4
1.4 MAPKs及NFκB細胞訊息傳遞路徑……………….…………..6
1.5 組蛋白修飾（histone modification）……….…………………..7
第 二 章 研究目的………………………………..……………………9
第 三 章 材料與方法………………………………………..………..11
3.1 細胞培養...…………………………………………....................11
3.2 芝麻素的來源及配製…………………………….......................13
3.3 細胞存活率（Cell viability assay）……………........................14
3.4 酵素免疫分析法（Enzyme-linked immunosorbent assay，ELISA）…………………………………………………………..14
3.5 西方墨點分析法（Western blotting）……………………..….....15
3.6 染色質免疫沉澱法（Chromatin immunoprecipitation assay，ChIP）.................................................................................................................17
3.7 聚合酶鏈鎖反應（polymerase chain reactio，PCR）.....................18
3.8 數據統計分析................................................................................19
第 四 章 結果……………..…………………………………………..20
4.1 芝麻素會抑制LPS刺激後THP-1和human primary monocytes 表現MDC..................................................................................20
4.2 芝麻素對THP-1沒有細胞毒殺作用……………………..….21
4.3 芝麻素是經由ER和PPAR-α表面受體而抑制LPS誘導表現的MDC……………..…………………………………….…...22
4.4 MAPK抑制劑和NFκB抑制劑皆可抑制LPS誘導產生的MDC…………………………………………………………...22
4.5 芝麻素抑制LPS誘導人類單核球細胞表現的MDC是經由MAPK-p38和NFκB-p65訊息傳遞路徑………………...…...23
4.6 芝麻素經由抑制NFκB相關的乙醯轉移酶CBP抑制histone acetylation而使LPS誘導表現的MDC被抑制…………….24
第 五 章 討論………………………………………..……………..…26
第 六 章 結論……………………………………………..…………..32

參考文獻……………………………………………………...………….42
附錄………………………………………………………..……………..51

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