木犀草素對 TGF-β1 與 MMPs 基因表現及對人類肺癌細胞侵襲能力之影響

The effect of luteolin on TGF-β1-mediated MMPs gene expression and cell invasion in human lung cancer cells

林伯儒(Po-Ru Lin)

指導教授 ： 吳立真

共同指導教授 ： 徐士蘭

國立暨南國際大學/科技學院/應用化學系/碩士(2012年)

https://doi.org/10.6837/NCNU.2012.00192

摘要

在肺癌組織中，Transforming growth factor beta ( TGF-β1 ) 因不斷刺激與發炎而發生突變，促使肺癌細胞侵襲 ( invasion ) 能力增加，並且高量表現，為腫瘤轉移過程中重要的促進分子。肺癌細胞轉移時，型態會改變，以利爬行，亦會分泌蛋白水解酵素( matrix metalloproteinases, MMPs )，分解胞外基質，侵入並擴散至周圍組織，造成肺癌轉移惡化的症狀，嚴重危及患者的生命。木犀草素
( 3’,4’,5’,7’-tetrahydroxyflavone )，是一個天然黃酮類化合物，存在於許多蔬菜、水果和中草藥植物中，是中藥金銀花的指標成分之一，其有抗發炎、抗氧化、抗增生、抗病毒、抗過敏及抗癌的功能。但其對於癌細胞轉移能力，尤其是 TGF-β1 誘發的癌細胞的侵襲能力之影響，無任何文獻研究討論。本論文研究利用高度轉
移能力的人類肺腺癌細胞株 A549，探討木犀草素對於癌細胞侵襲能力之分子機轉。由 Transwell 實驗結果顯示，前處理木犀草素，能有效抑制 TGF-β1 促進 A549 細胞之侵襲能力。由 gelatin zymography 和 RT-PCR 或是 real-time PCR 的實驗結果發現，木犀草素能阻斷 TGF-β1 活化的 MMP-2 和 MMP-9 之基因轉錄和酵素活性。進一步探討其分子作用機轉，TGF-β1能刺激活化 PI3K/AKT
和 NFκB 路徑，促進 MMP-2 和 MMP-9 表現，而木犀草素可抑制 TGF-β1誘發 AKT/ NFκB 活化路徑，而降低 MMP-2 和 MMP-9 之基因表現及活性，達到抑制轉移之效果。

關鍵字

TGF-β1 ； 侵襲 ； MMPs ； 木犀草素 ； A549 細胞株

並列摘要

TGF-β1 is a potent inducer of invasion which plays an important role in lung cancer metastasis. During metastatic process, lung cancer cells increase cell mobility and especially increase of matrix metalloproteinases (MMPs) activity which can destruct extracellular matrix structure to promote cell invasion. Luteolin, 3’,4’,5’,7’-tetrahydroxyflavone, a natural flavonoid existing in many types of plants including fruits, vegetables, and medicinal herbs, has potent anti-oxidative, anti-inflammatory, anti-proliferative, anti-metastasis, and anti-cancer properties both in vitro and in vivo. However, the molecular mechanisms of luteolin-mediated anti-metastatic activity remains largely unclear. To examine whether lutoelin might suppressed cancer cell invasion induced by TGF-β1, human lung adenocarcinoma A549 cell line was used. Our results showed that luteolin significantly attenuated TGF-β1-induced A549 cell invasion. This event was accompanied by the down-regulation of MMP-2 and -9 function and gene expression. Additionally, incubation of A549 cells with TGF-β1 resulted in activation of phosphorylated Akt as evidenced by increased Akt-Ser473, this event was significantly inhibited by luteolin. Moreover, luteolin blocked TGF-β1-mediated IκB degradation and NFκB activation. Interestingly, TGF-β1-stimulated cell invasion and MMPs upregulation was blocked by inhibitors of PI3K/Akt and NFκB. These data imply that the activation of Akt and NFκB pathway was necessary for TGF-β1-stimulated cell invasion and MMP upregulation. Overall, our findings indicate that TGF-β1 may activate NFκB by Akt that upregulates MMPs gene expression, and consequently induced cell invasion, however, this event can significantly block by luteolin, suggesting that luteolin inhibit TGF-β1-stimulated cell invasion and MMPs upregulation through the inhibiting Akt-NFκB signaling pathway.

並列關鍵字

TGF-β1 ； invasion ； MMPs ； luteolin ； A549 cell line

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