吳茱萸鹼誘導人類食道癌細胞死亡之作用 機轉

吳茱萸鹼誘導人類食道癌細胞死亡之作用 機轉

 根據行政院衛生署民國九十五年公佈台灣地區主要癌症死亡原因，食道癌位居第九位，居男性癌症 死因的第六位，五年的存活率低於 10% 。食道癌早期常無明顯症狀，因此大部分病患都無法於早 期獲得診斷，大約只有 30% 之病患在診斷之時，有機會接受手術之治療。所以發展新一代藥劑單 獨使用或併用他藥來治療食道癌，有即刻與強烈的需要。

吳茱萸中的成分吳茱萸鹼 (evodiamine) 是多功能物質，具有降血壓、抗心律不整、刺激內皮細胞和 抑制巨噬細胞釋放一氧化氮之作用。抗腫瘤的作用包括誘導細胞凋亡、抑制血管生成的作用及抑制 腫瘤細胞的侵入和轉移等等。

本實驗首次將吳茱萸鹼作用於人類食道癌細胞株 CE 81T/VGH 和 TE2 。研究結果顯示：隨著時間 及劑量之增加，吳茱萸鹼對人類食道癌細胞株 CE 81T/VGH 和 TE2 的生長抑制亦隨之增加，在 72 小時可以高達 80% 。在加藥處理後皆可以見到 CE81T/VGH 及 TE2 細胞株具有有絲分裂風暴之形 態學特徵，所以我們推測造成 CE 81T/VGH 和 TE2 細胞株死亡的方式主要為有絲分裂風暴。在免 疫螢光染色下， CE 81T/VGH 及 TE2 細胞株在藥物處理後 24 小時皆可以見到因中心體數目增加 而形成的多端紡錘體細胞。造成 CE 81T/VGH 及 TE2 細胞株中心體異常增多的原因可能為 Aurora A 的過度表現。與有絲分裂有關的調節蛋白如 MAD1 、 Bub R1 、 Aurora B 及 Survivin 亦隨著吳 茱萸鹼作用時間的增長而有不同的表現，所以我們推測紡綞體檢查點蛋白的異常表現是吳茱萸鹼造 成 CE81T/VGH 及 TE2 細胞株紡綞體檢查點缺失或異常的主要原因。以流式細胞儀分析發現 CE 81 T/VGH 及 TE2 細胞株在藥物處理後 24 小時細胞週期停滯在 G2/M 。以西方墨點法分析發現 CE 81 T/VGH 和 TE2 細胞株的細胞週期調節蛋白 cyclin B1 及 cdk1 在第一天皆有不正常的表現。

吳茱萸鹼在極低濃度下就有很高的細胞生長抑制的效果，所以具有對抗食道癌細胞之作用，因此有 可能作為食道癌的化學治療藥劑。而且吳茱萸鹼可以讓食道癌細胞週期停滯在 G2/M ，具有輻射致 敏劑的潛力。因此茱萸鹼有可能成為新一代治療食道癌的治療藥劑。但是吳茱萸鹼是否可以直接用 於治療食道癌病患，則需進一步的研究發展。

The effect and mechanism of evodiamine on induci ng cell

death of human esophageal cancer cells



Malignant tumor is one of the most common causes of death worldwide and cancer-related mortality is expected to increase c onsiderably. Esophageal cancer, usually is unresectable while diagnosis, has dismal prognosis. Despite aggressive treatment, t he overall 5-year survival rate is less than 10%. Clearly, development of new effective therapy in this malignancy remains the critically important issue in clinical practice.

Evodiamine is a naturally occurring bioactive ingredient of Evodia rutaecarpa (Juss.) Benth. Previous studies on evodiamine s howed several biological functions including anti-hypertension, anti-arrhythmia, stimulation on endothelial cells and inhibitio n on nitric oxide release from macrophages. It has also been demonstrated possessing anti-tumor effects on inducing apoptosis as well as inhibiting tubulin polymerization, angiogenesis, tumor invasion, and metastasis.

This study is the first to examine the anti-tumor effect of evodiamine against human esophageal carcinoma cell lines CE 81T/

VGH and TE2. The results show that evodiamine inhibited the growth of these two cell lines, up to 80% inhibition at 72 hours , in a dose- and time-dependent manner. Treatment with evodiamine arrested cell cycle at G2/M phase and induced morpholo gical changes characteristic of mitotic catastrophe in both cell lines. By immunofluorescence staining, centrosome amplificati on and multipolar spindle were observed. To link these mitosis-related cellular events, the expression of proteins involving ch k-cdk1-cyclin B mitotic pathway and spindle checkpoints was assessed simultaneously. Upon evodiamine treatment, cdk1 and cyclin B1 were up-regulated accompanied by a moderate increase in phosphorylation of histone 3, validating an arrest at G2/

M. The molecules involving centrosome maturation and spindle checkpoint function, such as Aurora A, Mad1, BubR1, Auror a B and Survivin were up-regulated by evodiamine with a similar time to plateau over 4 – 24 hours followed by degradation.

In conclusion, evodiamine possesses growth inhibitory activity against human esophageal cancer cells. The major mode of cel

l death induced by evodiamine is mitotic catastrophe accompanied by G2/M arrest and centrosome amplification. The putative

mechanisms of action might be those mediating centrosome maturation, amplification and spindle checkpoint function. It indi

cates that evodiamine might be a potent and novel therapeutic agent against esophageal cancer and may have a role in radiatio

n sensitization. Further in vivo studies including anti-tumor effect and toxicity are warranted.