- 學位論文
桂花粗萃物與其相關成分之免疫調節能力分析
The study of immunomodulatory effect of Osmanthus fragrans extract and its related compounds
摘要
桂花(Osmanthus fragrans flower)，是東方特有的藥草植物，根據中國醫書的記載，桂花除了有特殊香氣可作為食品外，枝葉、種子、根亦可供藥用。文獻指出，桂花中含有許多具有抗氧化與清除自由基能力之抗氧化性物質，分別為類黃酮類之rutin及酚酸類之verbascoside。Verbascoside亦是桂花純化物質中含量較高的化合物，但其是否具有抗發炎之免疫調節能力仍值得關注。目前經由動物實驗證實桂花粗萃物具有抗氧化與改善氣喘發炎的能力，而更深入的機轉探討並未進行。故本論文的研究著重在細胞層面的探討，而在未來開發相關桂花產品也可當作科學依據。實驗設計分為下列幾個部分：(1)利用人類腸道上皮細胞株與T細胞株模擬桂花粗萃物、rutin及verbascoside接觸腸道後之毒性反應，並藉由細胞激素IL-8、IL-2的表現情況來評估是否具有抗發炎能力。接續建立共培養系統觀察桂花粗萃成分是否經由影響腸道上皮細胞而更進一步影響免疫細胞。(2)探討桂花粗萃及其主要成份是否具有抗氧化能力及是否可改善處在氧化壓力下腸道上皮細胞分泌細胞激素IL-8的能力。(3)最後利用小鼠樹突細胞模擬觀察桂花粗萃物對於樹突細胞成熟發展之影響。本論文的結果顯示(1)桂花粗萃物與其主要成分均無細胞毒性的產生，細胞激素IL-2、IL-8亦有被抑制表現。此現象以桂花粗萃物能力最佳。(2)在氧化壓力情況下，桂花粗萃及其主要成份具有抑制腸道上皮細胞分泌IL-8的能力，(3)桂花粗萃物與其主要成分直接接觸樹突細胞後並未刺激樹突細胞活化成熟，甚至在脂多醣及卵蛋白的作用下，桂花粗萃物仍可抑制樹突細胞成熟，但其細胞激素分泌量並未有差異。由以上結果可得知桂花粗萃物確實具有免疫調節能力，但其影響途徑並不一定是經由抗氧化(清除氫氧自由基)所導致，需要更進一步測試訊息傳遞物質表現情況如NF-κB來判斷。
並列摘要
Osmanthus fragrans flower is one popular herb in the East. According to Chinese medical book, it recorded that O. fragrans leaves, seeds, roots all have medicinal benefit. O. fragrans contains many antioxidant compounds that have antioxidant and free radical scavenging capacity. Rutin(which belongs to Flavonoid) and verbascoside (which belongs to Polyphenol) are the main purified compounds of O. fragrans, but the capacity of anti-inflammatory and immunomodulatory is still unclear. In asthma animal model, extract of O. fragrans (OFE) has been confirmed that it can alleviate the airway inflammation and induced anti-oxidant status in lungs, but the mechanism was not discussed in detail. This study is focused on in vitro cell experiments and hopes these experiment results can provide the therapy application in the future. Experimental design was divided into the following sections:(1) Using human intestinal epithelial cell line (HT-29 cells) and T cell line (Jurkat cells) to evaluate the cell viability (using MTT assay) and secretion ability of IL-8, IL-2 (using ELISA assay) after treated with OFE and its related compounds. Furthermore, we also set up the in vitro co-culture system to investigate whether the supernatant of OFE treated intestinal epithelial cells can influence the Jurkat T cells. (2) Evaluate the anti-inflammatory ability of OFE on the IL-8 secretion of HT-29 cells under the H2O2 oxidative stress. (3) Finally, we observed the influence to OFE for maturation of dendritic cells. Results show that (1) OFE and its major compounds had no cytotoxicity, moreover, OFE have stronger ability to suppress the IL-2 secretion of Jurkat cells and IL-8 secretion of HT-29 cells. (2) OFE and its major compounds inhibited the secretion of IL-8 on intestinal epithelial cells under oxidative stress. (3) These compounds could not promote the maturation of dendritic cells, furthermore, OFE can suppress the maturation of lipopolysaccharide or ovalbumin treated dendritic cells. However, cytokines expression by DCs was not influenced. By the above results show that OFE does have the effect of immunomodulatory capabilities, but the mechanism did not only through anti-oxidant activity of OFE. The signal transduction pathways of inflammation such as NF-κB influenced by OFE still need further investigation.