臺灣博碩士論文加值系統

Psoralidin是由中藥補骨脂(Psoralea corylifolia L.)所萃取而來的天然物。psoralidin曾被報導過，於小鼠腹腔分離出的巨噬細胞能抑制LPS 誘導之NO生成，但抑制NO的作用機轉還未釐清，因此，本論文利用LPS刺激RAW264.7巨噬細胞株活化TLR4的訊息傳導，結果顯示，psoralidin於濃度10、 20 和30 uM能濃度相關的抑制LPS誘導之NO生成，並干擾NF-kB/IkB/IKK和PI3K/Akt路徑活化，而不影響MAPK路徑，psoralidin亦能抑制LPS誘導之TAK1磷酸化和IRAK1活化。Syk已知參與TLR4的上游訊息傳導之調控，但詳細機轉不明，本論文利用syk 抑制劑(piceatannol)處理細胞可知，piceatannol能抑制LPS誘導之NO生成、NF-kB和PI3K/Akt路徑及IRAK1活化，但不影響IKK/IkB路徑。這些結果顯示psoralidin和Syk兩者抑制NO生合成的機制有部分相同。然而，由細胞處理IRAK1的siRNA後發現與已知機轉不同的是，抑制IRAK1活化並不影響LPS活化的IKK/IkB 路徑。因此，psoralidin抗發炎的藥理機轉，也許是分別抑制了LPS誘導之IRAK1活化、PI3K/Akt路徑和IKK/IkB路徑後，進而影響NF-kB活化，最後抑制NO生合成。

Psoralidin is a natural product isolated from Psoralea corylifolia L., a herbal plant that is used in traditional Chinese medicine. The present results showed that this compound concentration (10, 20 and 30 uM)-dependently inhibited LPS-induced NO production in RAW264.7 macrophages via interfering with the activation of NF-kB/IkB/IKK and PI3K/Akt pathways,but didn’t affect MAPKs pathways activations. Furthermore, psoralidin inhibited LPS-induced IRAK1 activation and TAK1 phosphorylation. Syk is an upstream regulator of TLR4 signaling. However, the detail mechanism of Syk regulated TLR4 signaling is unclear. In this study, syk inhibitor piceatannol inhibitsed LPS-induced NO production, NF-kB, PI3K/Akt pathway and IRAK1 activations, but didn’t affect IKK/IkB pathway activation. It appears some similar action mechanisms between psoralidin and piceatannol. Transfection of siRNA targeting IRAK1, didn’t affect LPS-induced IKK/IkB pathway. Therefore, the anti-inflammatory mechanisms of psoralidin might inhibit LPS-induced IRAK1, PI3K/Akt pathway and IKK/I�菣B pathway independently and subsequently suppressing NF-k�羠 activation and NO production in RAW264.7 macrophages.

中文摘要..................................................1
英文摘要..................................... ............2
背景介紹..................................................3
壹、發炎反應(inflammation)................................3
貳、一氧化氮及一氧化氮合成酶..............................5
參、巨噬細胞的辨識系統: 類鐸受體(Toll-like receptors).....7
肆、以LPS刺激巨噬細胞所活化的NO訊息傳遞路徑...............11
伍、NF-kB/IKK 路徑........................................11
陸、絲裂原活化蛋白激酶(Mitogen-activated protein kinases, MAPKs)....................................................14
柒、PI3K/Akt 路徑.........................................15
捌、Non-receptor PTKs.....................................17
玖、Psoralidin............................................20
研究目的..................................................23
實驗材料與方法............................................24
壹、實驗藥品及材料來源....................................24
貳、緩衝液配製............................................26
参、實驗方法..............................................27
研究結果..................................................36
討論......................................................45
NO生成含量測定及psoralidin對NO生成的影響..................45
Psoralidin抑制iNOS mRNA及iNOS蛋白質含量...................45
Psoralidin抑制LPS誘導之TAK1活化卻不影響MAPKs..............46
Psoralidin抑制LPS誘導之IRAK1活化..........................46
Psoralidin影響的機轉層面..................................47
參考文獻..................................................49
圖表......................................................56

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