| 摘要: | 樹突細胞是一專業性抗原呈獻細胞，也是扮演連接先天和後天免疫的關鍵角色。因此，在發展免疫調節作用劑的研究中，樹突細胞的活化與抑制是一個相當重要的篩選標的。在本研究中，我們選定傳統中藥材—金銀花及四物(當歸、川芎、芍藥、地黃)來測定其萃取物對樹突細胞的活化反應並尋找其活性成分。實驗結果發現，金銀花甲醇萃取物會增加樹突細胞之TNF-α及IL-6的表現，顯示金銀花甲醇萃取物具有活化樹突細胞的活性；進一步使用水和乙酸乙酯進行成分分離，分別測試兩分離層的萃取物對樹突細胞的調節活性，結果發現水層萃取物有刺激樹突細胞活化的活性；然而乙酸乙酯萃取物則具有抑制樹突細胞活化的活性。經HPLC分析發現，水層萃取物活性成分中綠原酸為其主要成分。進一步測試綠原酸對樹突細胞活化的活性反應，發現綠原酸的確具有刺激樹突細胞的活性。然而，部分實驗結果顯示綠原酸對樹突細胞並無活化效果，所以推測可能是綠原酸結構不穩定所致。因此，我們以加鹼處理方式模擬綠原酸被鹼水解，再測試綠原酸鹼性水解產物對促進樹突細胞活化的影響。實驗結果發現，加鹼處理之綠原酸對樹突細胞具有較強活化現象，然而綠原酸的水解產物：咖啡酸和奎寧酸，對樹突細胞並無抑制或活化反應。雖然未釐清綠原酸活性不穩定的原因，但我們意外發現pH增高能夠誘導樹突細胞活化。在四物的部分，分別測試四物湯以及當歸、川芎、芍藥、地黃的甲醇萃取物對樹突細胞的活性影響，實驗結果發現，四物湯、當歸和芍藥對樹突細胞有明顯的活化活性。然而，只有川芎對樹突細胞有明顯的抑制活性。進一步探討川芎中所具有抑制樹突細胞的活性成分，實驗結果發現川芎最主要的成分蒿本內酯對樹突細胞的活化則具有明顯的抑制活性。綜合以上實驗結果，我們發現中藥材具有調控免疫反應的活性以及活性成分，將提供中醫用藥以及免疫調節劑開發的重要參考資訊。
Dendritic cells (DCs) are professional antigen-presenting cells, and play an important role in connecting innate and adaptive immunity; therefore DCs are important target in developing immunomodulatory agents. In this study, we chose five traditional Chinese medicinal materials — Lonicerae Flos and Si Wu (Angelicae Sinensis Radix, Chuanxiong Rhizoma, Paeoniae Alba Radix, and Rehmanniae Radix) — to examine their effects on activity of DCs and to discover the active constituents therein. The results showed that methanol extract of Lonicerae Flos activated DCs via increasing production of TNF-α and IL-6. After further component separation with water and ethyl acetate, we examined immunomodulatory activity of the extracts in two layers on DCs, and found that water layer extract possessed an activity of stimulating DCs, whereas ethyl acetate layer extract had an activity of suppressing DCs. To determine the active constituent in water layer extract, we used HPLC and discovered that chlorogenic acid was the main component. Furthermore, we examined the effect of chlorogenic acid on DC activity, and ascertained its stimulating effect. However, some parts of our results did not show effect of chlorogenic acid on DC activity; thus we speculated that chlorogenic acid was unstable. Hence, we simulated chlorogenic acid hydrolysis by adding a base, and tested the influence of basic hydrolysis in promoting DC activation. The result revealed that base-added chlorogenic acid had stronger activating effect on DCs. However, hydrolyzed products of chlorogenic acid, caffeic acid and quinic acid, exhibited neither inhibitory nor stimulating effect on DCs. Although not clarifying why the activity of chlorogenic acid was unstable, we found higher pH inducing activation of DCs. In regard to Si Wu, we tested the influences in DC activity of Si Wu Tang and methanol extract of Angelicae Sinensis Radix, Chuanxiong Rhizoma, Paeoniae Alba Radix, and Rehmanniae Radix, respectively. The results showed that Si Wu Tang, Angelicae Sinensis Radix, and Paeoniae Alba Radix had obvious activating effects on DCs. However, only Chuanxiong Rhizoma possessed a significant inhibitory effect on DCs. Further investigation of the active constituent that had the inhibitory effect on DCs in Chuanxiong Rhizoma, we found that the main component, ligustilide, possesses an apparent inhibitory activity on DCs. Combining aforementioned results, we discovered that certain Chinese medicinal materials possess an immunomodulatory activity and immunomodulators. Such findings will offer important references for the use of Chinese medicinal materials and development of immunomodulatory agents. |
