探討肉桂醛抑制人類骨肉瘤轉移機制之研究

Study of the inhibitory effect of Cinnamaldehyde on invasion and migration of human Osteosarcoma cells.

10.6834/CSMU201900032

賴怡安

肉桂醛 ； 骨肉瘤 ； 侵襲 ； 轉移 ； Cinnamaldehyde ； Osteosarcoma ； invasion ； migration

中山醫學大學生化微生物免疫研究所學位論文

2019年

碩士

謝易修;陳霈霓

繁體中文

骨癌中最常見的是原發性骨肉瘤Osteosarcoma(OS)，好發於兒童期至青春期。惡性程度最高的通常位於膝關節，由於生長板的關係長的最快，也最容易轉移至肺臟及肝臟。目前癌症治療上，腫瘤移除手術搭配化學治療，能有效提升患者癒後存活率達68%，而轉移至肺臟的骨肉瘤至今癒後情況仍然不理想，並且會導致高死亡率。肉桂醛是一種自然中的有機化合物，主要存在肉桂、風信子、霍香和玫瑰中，常做為香料用於食品工業。先前已經有研究證實肉桂醛具有抗菌的作用，以及抑制人類黑色素瘤A375細胞的增殖、擴散和成長。但是對於抑制骨肉瘤侵襲和轉移的機制至今還沒有很多研究探討。首先我們利用人類骨肉瘤細胞143B和Saos-2做MTT assay 發現肉桂醛在一定的濃度範圍不會影響細胞存活率，而在相同的濃度範圍做Wound healing assay，發現肉桂醛顯著抑制細胞的移動能力。用gelatin zymography看matrix metalloproteinase (MMP-2)和用u-PA zymography觀察u-PA活性，隨著肉桂醛濃度的上升MMP-2和u-PA都有顯著的下降。利用Boyden chamber assay 觀察到細胞隨著肉桂醛濃度的上升invasion和migration顯著的被抑制。以Western blot 分析結果，主要明顯抑制p-FAK、p-MEK1/2蛋白表現量，以及核內NF-κB、Twist-1和snail-1。在近一步利用RT-PCR探討肉桂醛對細胞基因的影響，發現143B的u-PA的基因表現量降低了，而對Saos-2的u-PA和MMP-2的基因表現量也明顯降低了。最後使用已知FAK抑制劑(PF-573228)證實抑制劑增強肉桂醛抑制143B的invasion能力。

Osteosarcoma (OS) is the most common primary bone cancer with the highest level in malignancy. It frequently occurs at knees where containing growth plates with the fastest growing rate. The patients often die because of tumor progresses rapidly and making lung and liver metastases.Cinnamaldehyde is a natural organic compound found mainly in cinnamon, hyacinth, fragrant and rose, and is often used as a spice in the food industry.Previous studies have confirmed that cinnamaldehyde has an antibacterial effect and inhibits the proliferation, spread and growth of human melanoma A375 cells.However, there are still absence of studies on the mechanism of inhibiting the metastasis of osteosarcoma.In this study, we demonstrated that cinnamaldehydeelicited very low cytotoxicity and significantly inhibited cellularadhesion , motility,migration,and invasion in human osteosarcoma 143B and Saos-2 cells.Moreover, using RNAsequencing technology, we revealed that cinnamaldehyde repressedu-PA gene expression in 143B cells and u-PA, MMP-2 inSaos-2 cells.Moreover, cinnamaldehyde decreased the activation offocal adhesion kinase (FAK),respectively, in a dose-dependentmanner in 143B and Saos-2 cells and evident influence on the level and activation of p38, Akt, ERK1/2, steroid receptor coactivator. In furtherfunctional experiments, the use of FAK inhibitors(PF-573228)confirmed that the pharmaceutic inhibition of FAK augmentedthe cinnamaldehyde-mediated migrationsuppression of 143B cells.