香椿葉萃取液對非小細胞肺癌細胞株之抗癌功能研究

在我們過去的研究中，發現香椿葉水萃物（TSL-CE）有效抑制肺腺癌細胞A549的生長。而A549是非小細胞肺癌（NSCLC）中的一個類型，所以我們想了解香椿對人類肺腺癌細胞（H441）、肺鱗狀細胞癌細胞（H520）及肺大細胞癌細胞（H661）等三種非小細胞肺癌（NSCLC）的抗癌影響，結果顯示香椿樹各部份中以香椿葉最有抑癌效果，且NSCLC處理TSL-1(TSL-CE離心後之上清液)後比TSL-CE更有抑制效果，在對肺癌細胞增生之抗癌影響方面。以流式細胞儀分析發現，TSL-1處理三種細胞株的細胞週期都停滯在G1期，進一步以西方墨點法分析調控細胞週期的相關蛋白質分子表現，發現經TSL-1處理後，會致使Cyclin-D1在H441、H520及H661的表現量均下降，cdk4只在H441及H661的表現減少，p27在H520及H661的表現增加，而H661投予TSL-1後，在cylin D1、cdk4及p27表現量變化均較H441和H520明顯，在MTT assay測得IC50之濃度在H661亦是最低，所以TSL-1對NSCLC中的肺大癌細胞（H661）之抑癌效果是最好。在細胞死亡方式之抗癌影響方面，TSL-1處理後的肺癌細胞在細胞型態、Sub-G1的堆積、PARP被裂解情形、以及PI-Annexin-V雙染和TUNEL assay分析之結果，均證實TSL-1會造成三株細胞的凋亡。肺腺癌是NSCLC中最常見的類型，所以在動物試驗，選用H441（肺腺癌細胞）誘發裸鼠皮下腫瘤，分別以治療及預防二種實驗方式來探討，結果發現在治療上不論是灌食或腹腔注射均能有效抑制癌細胞生長；預防實驗中，先期灌食TSL-1亦使腫瘤細胞生長受到抑制而有預防效用。體內試驗之實驗組動物在血液生化值或組織器官上與對照組無差異。此項發現推測口服TSL-1可作為未來研發抗肺癌保健食品之重要學理依據。由本研究瞭解香椿在NSCLC抗癌能力上，以香椿葉抗癌效果最好，又以其成份TSL-1對於肺大細胞癌細胞（H661）有極佳的抑癌影響及誘發細胞凋亡；長時間餵食TSL-1對體內不會造成副作用，並可治療及預防癌細胞生長。因此TSL-1具有開發成為預防NSCLC之發生及治療用的保健食品。

關鍵字

非小細胞肺癌；保健食品；香椿

並列摘要

Our previous study showed that the crude extract from the leaves of Toona sinensis (TSL-CE) exerts potent anti-proliferative effect on A549 lung cancer cells. Since A549 is one of the Non-small-cell lung cancer (NSCLC) types, it is of interest to know whether TS also has anti-cancer effect on all three types of NSCLC. TSL-1, a fraction of TS crude extraction, is the supernatant parts of TSL-CE after centrifugation. The effects of TSL-1 on the proliferation of H441 cells (adenocarcinoma), H661 cells (large cell carcinoma) and H520 cells (squamous cell carcinoma) were investigated in this study. We tested the anti-cancer effect of TSL-1 on the proliferation and cell death. Our results showed that, the anti-proliferative effect of the TSL-1 is more potent than TSL-CE in all three cell types of NSCLC. Treatment of NSCLC cells with TSL-1 resulted in a G1-phase arrest by Flow cytometry. TSL-1 mediated G1 arrest was accompanied by the down-regulation of protein levels of cyclin-D1 in all three cell types of NSCLC, reduced cdk4 expression in H441 and H661, and the up-regulation of p27 in H520 and H661. We found that the decrease in cyclin D1 and cdk4 and the increase in p27 were most significant in H661 cells, compared to other two cell lines. Among the three cell lines, the IC50 for TSL-1 in H661 cells was the lowest, indicating the anti-cancer effect of TSL-1 is most potent in this cell line. TSL-1-induced apoptosis was confirmed by cell morphology, Sub-G1 peak accumulation, cleavage of PARP, PI-Annexin-V double staining and TUNEL assay in all three types of NSCLC. Adenocarcinoma of the lung cancer is now the most worldwide common subtype of NSCLC. Therefore, we performed in vivo experiments to evaluate the therapy and prevention effects of TSL-1 on H441 cells. Per os and intraperitoneal treatment of TSL-1 was used in therapy study, while only per os treatment of TSL-1 was used in prevention study. Our results revealed that treatment of TSL-1 inhibits H441 tumor growth in either therapy or prevention experiments. Whereas, TSL-1 did not show any side effects in these in vivo studies. Our results suggest that TSL-1 could be used as a health food for preventing and alleviating lung cancer proliferation and possess the potential to be developed as an anticancer drug.

並列關鍵字

NSCLC ； Toona sinensis ； health food

參考文獻

中文參考文獻

Google Scholar

Chang HC, Yeh CH, Huang MC, Hsu C, and Hsu HK: 香椿萃取液減少Bcl 2蛋白質表現而促進H520肺癌細胞自然凋亡。Health food society of Taiwan. 2001; Feb.30.

Google Scholar

Chen WI, Chen YR, Wang PH, Lue SI and Hsu HK: 椿葉萃取物對血小板凝集的作用。Health food society of Taiwan. 2002; Feb.70.

Google Scholar

Tsai LY, Huang CC, Hsiao JK, Lin SK, and Chang CS: Effects of Toona sinensis Roemor Concentrate on H2O2-induce Oxidative Damage in HL-60 and K562 Cell Lines. 中華民國醫事檢驗學會九十二年度學術發表會，2003/12/13-14，台北榮總致德樓.