| Abstract: | 由衛生署公佈之台灣十大死亡原因顯示，癌症仍為十大死因之首，因此，如何預防或延緩癌症的發生為刻不容緩的事。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。目前許多研究趨向以天然物成份或複方來達到抑制癌細胞的增生及惡化，例如Tea polyphenolic extracts, curcuminoid extracts及broccoli extracts （sulforaphane）已被廣泛的應用為化學預防物質（chemopreventive agent）。癌症預防已為一世界潮流，因此，尋找化學預防物質為一重要工作。本論文主要以下列兩種天然物為研究材料，戊乙醯去羥梔子[penta-acetyl geniposide，(Ac)5GP]以及洛神花花青素（Hibiscus anthocyanins，HAs），探討此兩種天然物對癌細胞之作用及抗癌之分子機制。近年來，有許多研究指出一些抗癌藥物可藉由引起癌細胞的程序式死亡或改變癌細胞的生長週期，來達到抑制癌細胞的增生與惡化。戊乙醯去羥梔子是從山梔子（Gardenia fructus）萃取出來的一種醣（glycoside），再經乙醯化（acetylation）而得到的物質。本論文研究發現，戊乙醯去羥梔子處理C6細胞後，觀察到細胞發生死亡現象，且p53、c-Myc及Bax蛋白表現增加，導致細胞走向細胞凋亡。以戊乙醯去羥梔子分別處理C6細胞0至24小時後，同樣也發現p53蛋白表現量增加，使下游的p21活化，進一步使cyclin D1/cdk 4複合體減少，此時，phospho-Rb蛋白的表現受抑制，相對的Rb/E2F複合體增加，最後導致細胞週期停滯在G0/G1期。因此，推斷戊乙醯去羥梔子會引起C6神經膠瘤細胞的細胞週期停滯，且造成C6細胞凋謝死亡。另外，花青素廣存於蔬菜、水果等植物中，已有許多報告證明花青素具抗氧化活性，降低血脂肪及防癌等作用。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。結果顯示，洛神花花青素會使磷酸化的p38及c-Jun表現量上升以及tBid、Fas、FasL、cytochrome c等蛋白表現增加而誘導HL-60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N-Nitroso-N-Methylurea（NMU）誘發Sprague-Dawley大白鼠血癌。結果發現，洛神花花青素可以抑制白血球對肝臟及脾臟浸潤，並且進一步抑制大鼠罹患血癌。從以上的結果，我們證實戊乙醯去羥梔子及洛神花花青素可以預防及治療癌症並說明其作用機轉。希望本實驗室所合成的戊乙醯去羥梔子及洛神花花青素在釐清其作用機轉後發展成一具廣泛性、無毒害，且在人體治療上適用於各種癌症之抗癌藥。
Department of Health of Executive Yuan announced that amonth the Taiwanese ten major death causes of Taiwanese, cancer is still listed as the first. Hence, it is a very important to prevent or delay the emergence of the cancer. The development of cancer is complicated and multifactoral. Although some of the mechanisms of carcinogenesis are well studied, the efficiency of treatment is still limited and the complication is severe. A lot of studies tend to suppress the proliferation and deterioration of cancer cells by using natural compounds or medicine composed of two or more ingredients, such as tea polyphenolic extracts, curcuminoid extracts and broccoli extracts (sulforaphane) which have already been extensively applied as chemopreventive agents. Prevention of cancer development is the trend, and a vital part of it is to look for chemopreventive agents. This thesis focued on two natural compounds, penta-acetyl geniposide, [(Ac)5GP], and Hibiscus anthocyanins (HAs) to explore the molecular mechanisms of their inhibitory effect on cancer cells. In recent years, many researches showed that some anticancer drugs could induce the death or influence the cell cycle of cancer cells, resulted in suppressing the proliferation and deterioration of cancer cells. (Ac)5GP is the acetylated form of glycoside that was extracted from Gardenia fructus. This thesis discovered that (Ac)5GP could induce C6 glioma cell death and increase the expression of some proteins, such as p53, c-Myc and Bax, which led C6 glioma cell to apoptosis. C6 cell treated with (Ac)5GP for 0 to 24 hours revealed an increased expression of p53 protein that, in turn, activated its downstream protein (p21), which consequently reduced cyclin D1/cdk 4 complex, suppressed the level of phospho-Rb, and increased the relative level of Rb/E2F complex. These events caused cell cycle arrest in G0/G1 phase. It was demonstrated that (Ac)5GP could induce cell cycle arrest and apoptosis of C6 glioma cells. In addition, anthocyanins exist widely stored in plants (i.e. vegetables, fruit, etc.). Several reports have proved that anthocyanins have anti-oxidant activity, anti-cancer function, and reduce blood cholesterol. 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. The results showed that HAs not only induced phosphorylation of p38 and c-Jun proteins but also elevated expression of tBid, Fas, FasL, cytochrome c proteins, leading to HL-60 cells death. In the second part, we conducted animal experiment to investigate the function of HAs to prevent or delay the development of cancer. The animal model of leukemia was carried out by injecting N-Nitroso-N-Methylurea (NMU) to Sprague-Dawley rats. We found that HAs suppressed the leukocytes to invade the liver and spleen, and inhibited NMU-induced rat leukemia. The above mentioned resultes demonstrated the mechanisms via which (Ac)5GP and HAs prevent and delay the development of cancer (leukemia). We expect that (Ac)5GP and HAs could be developed as the anticancer drugs with the advantage of widely used, non-toxic and suitable for various kinds of treatments of human cancers. |
