洛神花青素在人類血癌細胞株（MOLT-4）和利用小鼠血癌細胞株（WEHI-3）誘發血癌模式的細胞生長與凋亡之探討

癌症的形成是透過許多複雜且多重的過程，其中形成的原因雖然被了解，但治療的效果仍然有限，且亦對人體造成極大的傷害。因此，尋找化學預防劑，是一個非常重要的事情。花青素是一種廣泛存在於蔬果植物中的天然色素。內含有豐富的類黃酮、花青素、原兒茶酸等天然抗氧化成分外，在過去許多的研究中，結果證實洛神花中的花青素成分具有抗氧化活性、護肝及抗癌的功效。本實驗探討洛神花花青素（Hibiscus anthocyanins，HAs）是否對於人類血癌細胞株(MOLT4)和小鼠血癌細胞株(WEHI-3)會誘發抗癌活性及其探討分子機轉。由於洛神花在國內容易栽培，產率高，而洛神花花青素也容易分離，因此，本研究擬利用台灣特有之洛神花所分離之花青素，進行體外（in vitro）及體內（in vivo）實驗，首先 以MTT分析HAs對MOLT4細胞之生長作用，發現在處理24小時下，其 IC50為 0.2mg/ml ，再藉由流式細胞儀分析結果發現洛神花花青素能夠讓細胞週期大量在SubG1期，使細胞走向凋亡,也藉由流式細胞儀分析測得經洛神花花青素作用後的人類血癌細胞(MOLT4)使粒線體膜電位下降之細胞凋亡，由以上實驗再經由西方墨點法指出洛神花花青素能使癌細胞中促凋亡蛋白 Bid，Bax，Bad 及p-Jnk表現量上升而誘導MOLT4細胞死亡。接著，利用動物實驗探討洛神花花青素預防或延緩癌症之作用，動物模式為使用小鼠血癌細胞(WEHI-3)以腹腔注射方式誘發小白鼠產生血癌，並處理不同濃度花青素兩週，根據血液分析、生化分析、H&E 染色、劉氏染色等實驗方法，洛神花花青素可以抑制癌細胞浸潤肝臟及脾臟，進一步抑制小鼠血癌惡化，從以上的結果，我們證實洛神花花青素可以治療癌症並說明其作用機轉。

The development of cancer is complicated and multifactoral. Ahough some of the mechanisms of carcinogenesis are well studied. Anthocyanin is a widely present in fruits and vegetables. It contains highly valuable nutrients, such as protocatechuic acid, flavonoids, and anthocyanins. Some previous reports demonstrated that H. sabdariffa anthocyanins (HAs) were anti-oxidant, liver-protective and anti-tumorigenic. We examined the effect of HAs on human leukemia MOLT4 and mouse WEHI-3 cell. Hibiscus sabdariffa L. easily cultured in Taiwan has high production rate of anthocyanins. Besides, HAs are easy to extract. Therefore, it is an easily available material for research here. This research carried out in vitro and in vivo tests with HAs. MTT was used to analyze the growth- inhibiting effect of HAs. The IC50 was 0.2 mg/ml after 24-hour-treatment. Therefore，the result from flow cytomertic analysis indicated that HAs cell cycle in SubG1 and induced apoptosis in examined cell line. Flow cytomertic also used for analysis the levels of decreased the JC1 stain level. Western blot show that HAs treatment gradually increased the levels of the pro-apoptotic protein Bid,Bax,Bad and p-Jnk. Animal model to use the mouse leukemia cells (WEHI-3) induced by intraperitoneal injection of mice leukemia, and tube feeding different concentrations of HAs two weeks, Based on the data of blood analysis, biochemical analysis, H&E stain, Liu’s stain. we further demonstrate that HAs inhibits by WEHI3 reduces invasion of liver and spleen. Further deterioration of inhibition of mouse leukemia. The above mentioned resultes demonstrated the mechanisms HAs prevent and delay the development of cancer (leukemia).