大黃酸誘導乳癌細胞凋亡之探討

Rhein induces apoptosis in human breast cancer cells

宋明璋(Ming Zhang Song)

指導教授 ： 張清堯

亞洲大學/健康學院/生物科技學系碩士班/碩士(2008年)

摘要

摘要
中國傳統草藥有許多具有抗腫瘤的功能藥物，如大黃酸(Rhein)。大黃酸是大黃根莖中的一種蒽醌類，已有文獻報導抑制人類肝癌、神經膠質瘤與骨髓癌細胞生長。乳癌為臺灣女性癌症首位，每年約有兩千人罹患，有一半以上病患會因此病而死亡。臨床上有些乳癌患者HER2過度表現患者難治療，癒後較差。本論文研究目的為從25種中草藥指標成分中與大黃酸（Rhein）比較證明出可同時抗大量表現HER2乳癌細胞(MCF-7/HER2)及低度表現HER2乳癌細胞(MCF-7/WT)生長的藥物大黃酸，並且探討其抗腫瘤的作用機轉。大黃酸(Rhein)對 MCF-7/WT細胞的百分之五十藥物抑制濃度(IC50)為83.66 μg/ml ，及對 MCF-7/ HER2細胞的IC50為47.67μg/ml，為26種中草藥指標成分中最顯著的抗乳癌的藥物。藉由流式細胞儀分析細胞週期(cell cycle)，在此實驗發現大黃酸對MCF-7/ HER2細胞和 MCF-7/WT細胞具有抑制細胞增生作且誘導細胞凋亡的發生，尤其對MCF-7/ HER2細胞誘導細胞凋亡程度比MCF-7/WT細胞更顯著。由西方墨點法(Western blotting)來看大黃酸對MCF-7/ HER2細胞和 MCF-7/WT細胞活化Caspase-3與Caspase-9。另外，進一步探討細胞凋亡機轉，以流式細胞儀分析發現大黃酸誘導細胞粒線體膜電位的下降及促進ROS的產生。且用細胞內訊號傳遞路徑分析了解大黃酸活化NF-κB與 p53訊號傳遞。由本論文研究成果證明大黃酸會促進ROS產生與粒線體膜電位下降，活化NF-κB與 p53訊號傳遞，進而誘發細胞凋亡，抑制人類乳癌細胞的細胞增生。因此，未來大黃酸可進一步評估作為抗人類乳癌藥物的可能性。

關鍵字

大黃酸 ； 乳癌 ； 細胞凋亡

並列摘要

Abstract
Chinese traditional herbs, such as rhubarb, have been demonstrated anti-cancer function. Rhein ，4, 5-dihydroxyanthraquinone-2-carboxylic acid，is an anthraquinone compound that is enriched in rhubarb with anti-liver cancer, anti-glioma and anti-myeloma activities in vitro and in vivo assays. Breast cancer occupies the first position among female cancer deaths in Taiwan. The patients whose breast cancer cells over-express the HER2 protein are likely resistant to a variety of chemotherapy agents. In this study, 26 Chinese traditional herb compounds were selected to test the activities and mechanism of anti-breast cancer in vitro assays of MCF-7/WT cells expressing the low level of HER2 and MCF-7/HER2 cells expressing the high level of HER2. MTT assay indicated that rhein showed the best ability to inhibit the growth of both MCF-7/WT cells and MCF-7/HER2 cells among these Chinese traditional herb compounds. The 50% inhibitory concentration (IC50) value of rhein in MCF-7/WT cells was 83.66 μg/ml, whereas the IC50 value of rhein in MCF-7/ HER2 cells was 47.67 μg/ml. Flow cytometry showed that rhein induces dose-dependent apoptosis. Furthermore, Western blotting demonstrated caspase 3 and caspase 9 dependent apoptosis induced by rhein. Flow cytometry assays showed that rhein significantly increased in the production of reactive oxygen species (ROS) and decreased in the mitochondrial membrane potential (MMP) using. Moreover, in vivo signaling pathway assay revealed that NF-κB and p53, but not AP1 were activated in rhein-treated MCF-7/WT cells and MCF-7/HER2 cells, being associated with ROS-mediated apoptosis. Therefore, rhein could be useful for the clinical application of treatment for patients with breast cancer with low and high levels of HER2 expression.

並列關鍵字

Rhein ； HER2 ； cell cycle

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