Diallyltrisulfide 經由活性氧化物誘導人類口腔癌細胞凋亡

Abstract

流行病學研究指出，飲食中攝取青蔥屬（Allium）的蔬菜，例如：洋蔥、大蒜等，可以降低肺癌、結腸癌、攝護腺癌和乳癌等多種癌症發生的危險性。青蔥屬蔬菜在加工過程中會產生有機硫化合物（organosulfur compounds, OSCs），使這類蔬菜具有抗癌效果。 化學物質誘發癌症實驗動物研究發現，青蔥屬蔬菜產生的OSCs成分中，以diallyl sulfide (DAS) ，diallyl disulfide (DADS) ，和diallyl trisulfide (DATS) 較具有抗癌保護的效果，但其作用機轉尚未明瞭。 近年研究指出， DAS, DADS 和DATS可有效的抑制皮膚癌、小腸癌、乳癌和結腸癌等等之癌細胞的生長並促使細胞凋亡。 先前本實驗室研究發現DATS可引起人類口腔鱗狀細胞癌細胞株Ca9-22與SAS細胞凋亡。 DATS促使癌細胞凋亡的效果明顯比DAS及DADS好，但DATS對於口腔癌細胞的作用機轉尚未完全明瞭。 本研究發現以DATS分別處理口腔癌細胞株SAS及Ca922後，會產生Reactive oxygen species （ROS） 以及造成JNK的活化，加入了JNK 的抑制劑 (SP600125) 及 ROS的抑制劑 N-acetylcystine (NAC)可抑制由DATS所引起的細胞凋亡。 顯示DATS 可經由 ROS 產生及JNK的活化引起細胞凋亡。 西方墨點法分析顯示DATS可使得FADD大量表現、Bcl-2表現量降低、caspase-8及caspase-9活化以及PARP的水解。另外我們也發現細胞在加入DATS之後Caspase 4有明顯的受到活化，可見得DATS也可以經由導致ER stress而走向細胞凋亡。 總結本研究中結果發現，DATS造成口腔癌細胞Ca9-22走向細胞凋亡主要的機制為細胞內產生ROS而導致JNK的磷酸化以及下游的FADD的大量表現、Bcl-2表現量降低、caspase-8、caspase-9和caspase-4的活化以及PARP的水解。對於DATS是否可當作口腔癌抗癌藥物則須先以動物實驗做更深入的評估其抗癌的功效。

Epidemiological studies support the premise that dietary intake of Allium vegetables (e.g., garlic, onions and so forth) may lower the risk of varies types of cancer. Garlic-derived compounds have been shown to offer protection against oral tumor in animal models induced by carcinogens. Garlic-derived organosulfur compounds (OSCs) including diallyl sulfide (DAS), diallyl disulfide (DADS) and diallyl trisulfide (DATS) have been shown to suppress proliferation and induce apoptosis in various cancer cell lines including skin, intestinal, breast and colon cancer cells. Previously, we have shown that DATS can induce apoptosis in oral cancer SAS and Ca9-22 cell lines. DATS was more effective than DAS and DADS for induction of apoptosis. However, the effects of DATS on oral cancer cells are not fully understood. In this study, we found treatment of SAS and Ca9-22 cells with DATS induced reactive oxygen species (ROS) production as detected by DCF fluorescence. Pretreatment of cells with NAC reduced the DATS-induced apoptosis. We also found that DATS-PDT was able to induced JNK activation and pretreatment of cells with SP600125 (JNK inhibitor) inhibited DATS -induced PARP cleavage. Western blot analysis showed DATS treatment induced FADD overexpression, Bcl-2 down-regulation, caspase-8, caspase-9 activation and PARP cleavage. We also observed that treatment of Ca9-22 cells with DATS activated of caspase-4, which indicated involvement of endoplasmic reticulum (ER) stress in apoptosis. Taken together, our study showed that DATS can induce apoptosis in oral cancer cells via ROS production, JNK phosphorylation, activating intrinsic- and extrinsic-apoptosis pathway, and caspase-4 activation.