研究甘草查爾酮A抑制人類肝癌細胞侵襲之分子機制

人類肝細胞癌是全世界最普遍的惡性腫瘤之一，尤其在亞洲和非洲的比例最高，平均約有百分之八十的癌症病人因癌細胞之轉移而致命，故若能有效抑制癌細胞的轉移及入侵，將可大幅減低癌症的死亡率。因此發展新穎天然藥物來當作癌症預防或是治療藥物是目前最重要的課題。甘草查爾酮A ( Licochalcone A )是甘草成分中最具有活性的化合物，本身也是estrogenic且屬於黃酮類分子。目前根據研究報導指出甘草查爾酮A具有抗腫瘤和抗發炎之功能。但是，目前甘草查爾酮A對於人類肝癌細胞的抗癌作用機制，至今仍然未知。本研究以MTT方式證實甘草查爾酮A不會降低六株人類肝癌細胞株(HA22T/VGH、 SK-Hep-1、Huh-7、PLC/PRF/5、Hep3B 和 HepG2)及正常人類肝細胞(Chang liver) 的細胞存活率。腫癌細胞的侵襲和轉移是惡性腫瘤的重要特徵之一，也是在臨床治療上最一致的現象。因此本研究以高度侵襲肝癌細胞 HA22T/VGH 和 SK-Hep-1為模式來探討甘草查爾酮A抑制人類肝癌細胞移動和侵襲能力。採用細胞傷口癒合、細胞移動和侵襲實驗證實甘草查爾酮A 抑制HA22T/VGH 和SK-Hep-1 細胞的移動和侵襲能力。利用西方墨點法、casein zymography方法、免疫螢光法和RT-PCR方式證實甘草查爾酮A抑制人類肝癌細胞侵襲是透過抑制 u-PA 蛋白和mRNA表現。同時也證實甘草查爾酮A會專一性抑制JNK1/2磷酸化；同時加入JNK1/2抑制劑(SP600125)會加強甘草查爾酮A抑制u-PA蛋白和mRNA表現。我們利用轉錄因子結合區的軟體尋找到u-PA 啟動子的轉錄因子(AP-1、ATF-2和CREB-1)，並以Luciferase 方法和西方墨點證實甘草查爾酮A會抑制u-PA 啟動子的活性，同時也會抑制u-PA啟動子上的CREB-1蛋白表現，不影響AP-1和ATF-2蛋白表現。此研究結果說明甘草查爾酮A可能在作抗肝癌藥物和抗侵襲藥物的發展上深具潛力。

關鍵字

甘草查爾酮A ；轉移；侵襲；人類肝癌細胞；尿激酶-型血漿酶原催化劑

並列摘要

Hepatocellular carcinoma (HCC) is one of the most common malignant tumors in the world, and especially prevalent in parts of Asia and Africa. The metastasis (invasion) of cancer cells leads 80% patients died, and inhibition of cancer cells metastasis will markly decrease the death rate. Therefore, development of novel nature drug or drug therapy was important topic. Licochalcone A is the main active compound of the licorice species Glycyrrhiza inflate, and is an estrogenic flavonoid with anti-metastatic, anti-cacinogenic, anti-inflammatory, anti-angiogenic, anti-oxidant and apoptosis effects. However, Licochalcone A effect molecular mechanism of human hepatocellular carcinoma is presently unknown. In our studies found that the cell growth inhibitions of Licochalcone A on six HCC cancer cells (HA22T/VGH、 SK-Hep-1、Huh-7、PLC/PRF/5、Hep3B and HepG2) and normal liver cells (Chang liver) were not affecting the cell growth by MTT assay. We found that the Licochalcone A inhibited the migrate and invasive on HA22T/VGH and SK-Hep-1 cells were observed by wound healing assay, invasion assay and migration assay after treatment for 24 h. Licochalcone A inhibited the HCC cell migration and invasion through suppression of u-PA expression by western blots, casein zymography, immunofluorescence assay and RT-PCR assay. Furthermore, Licochalcone A specifically inhibited phosphorylation of JNK1/2, and inhibition of JNK1/2 by JNK1/2 inhibitor (SP600125) significantly enhanced the Licochalcone A -inhibited u-PA activity, protein and mRNA expression. We used the software of transcription factor binding sites to find that transcription factors of u-PA promoter (AP-1、ATF-2 and CREB-1), and we also suggested that Licochalcone A inhibited u-PA promoter transcription activity in a dose-dependent by luciferase assay, moreover it also inhibit the expression of CREB-1, not effect AP-1 and ATF-2. Therefore, Licochalcone A is a promising candidate for further development as an anti-tumor or anti-invasive agent against HCC progression..

並列關鍵字

Licochalcone A ； Migration ； Invasion ； Human hepatocellular carcinoma cells ； urokinase-type plasminogen activator

參考文獻

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研究甘草查爾酮A抑制人類肝癌細胞侵襲之分子機制

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