Sensitizing Effect of Berberine on Radiation-Induced Cytotoxicity in Human Liver Cancer Cells

黃蓮素對放射線毒殺人類肝癌細胞的增敏作用

田雨生(Yeu-Sheng Tyan) ； 劉文山(Wen-Shan Liu) ； 周芬碧(Fen-Pi Chou) ； 曾顯群(Hsien-Chun Tseng)

《放射治療與腫瘤學》 19卷1期 (2012/03) Pp. 63-72

https://doi.org/10.6316/TRO/201219(1)63

摘要

目的：研究黃蓮素對於放射線造成人類肝癌細胞的死亡是否有增敏作用。材料與方法：本研究使用人類肝癌細胞株Hep3B，比較以放射線或放射線合併黃蓮素對於Hep3B細胞死亡的影響，利用MTT assay分析細胞的存活，以DAPI stain計算細胞核內DNA濃染的數目，而flow cytometry分析定量細胞的凋亡，western blot分析與細胞凋亡相關蛋白的表現。結果：放射線主要誘導Hep3B細胞的凋亡，與放射劑量成正相關。比較單獨放射線照射10Gy與合併10Gy的放射線及10μM黃蓮素兩組細胞，流式細胞儀顯示細胞凋亡的比例自15.3%提升至47.2%。與細胞凋亡相關的蛋白如cytochrome c、Bax、p-JNK、cleaved caspase-3表現量增加，相反地，抗凋亡蛋白如Bcl-2與Akt的表現則降低。結論：上述結果證實黃蓮素能提升放射線毒殺Hep3B細胞的效果，其分子調控機轉推測可能是透過活化JNK與抑制Akt訊息傳導途徑，這部分有待進一步實驗證實。

關鍵字

黃蓮素 ； 細胞凋亡 ； 放射線敏感性

並列摘要

Purpose: To study the sensitizing effect of berberine on radiation-induced cytotoxicity in human liver cancer cells. Materials and Methods: Berberine, an isoquinoline alkaloid, was used in combination with radiotherapy to treat a human liver cancer cell line, Hep3B. The cells were irradiated with the indicated single dose of 10Gy with or without berberine (10μM) and cell viability was analyzed on MTT(3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyl- tetrazolium bromide) assay. DAPI(4',6-diamidino-2-phenylindole)staining was used to detect the apoptosis and acridine orange was used to detect presence of acidic organelles formation. The morphology of the cells was observed under fluorescence microscope and apoptosis was quantified by flow cytometry. The expressions of apoptosis-related proteins such as cytochrome c, cleaved caspase-3, Bax, p-JNK, Bcl-2 and Akt were analyzed on Western blot. Results: Radiation induced apoptotic cell death of Hep3B cells in a dose-dependent manner. The ratio of apoptotic cells increased from 15.3% in Hep3B treated with irradiation alone to 47.2% in Hep3B cells treated with irradiation in combination with berberine. Western blot analysis demonstrated increased expressions of apoptosis-related proteins, such as cytochrome c, Bax, p-JNK, and cleaved caspase-3, and decreased expressions of phosphorylated Akt and Bcl-2. Conclusion: Berberine enhances radiation-induced cytotoxicity in Hep3B cells most likely by activation of JNK and suppression of Akt.

並列關鍵字

Berberine ； Apoptosis ； Radiosensitivity