BJ-FA 對於人類血管內皮細胞的生長抑制作用及分子機轉
Molecular Mechanisms of BJ-FA-Induced anti-proliferation effect in human vascular endothelial cells
中文摘要
本篇研究計畫主要在探討化合物 (BJ-FA) 對人類臍靜脈內皮細胞 （Human umbilical vein endothelial cell，HUVEC）的生長抑制作用及其可能作用的分子機 轉。近年來，有關藉由抑制血管增生 (Angiogenesis) 進而抑制腫瘤生長的研究議 題，相當熱門。而在相關研究當中顯示，許多抑制血管增生的化合物不僅有抑制 腫瘤生長的作用且相對有著較低的副作用，因此尋找有效且容易被人體吸收的 化合物用以取代目前臨床對於癌症的治療藥物，似乎是比較妥當的方式。
本研究的發現，BJ-FA 能使人類臍靜脈內皮細胞的生長產生抑制作用，且其抑 制的效果和BJ-FA 的劑量成正相關性。而利用 3H-Thymidine incorporation 和 Flow cytometry 的實驗結果，我們發現 BJ-FA 會抑制血管內皮細胞的 DNA 合成，
並使細胞週期停滯在G0/G1。此外，利用 Western blot 的實驗分析，我們觀察到
與細胞週期停滯相關的蛋白p21，在血管內皮細胞的表現可受 BJ-FA 的刺激而
增加。我們進一步利用免疫沉澱法分析，發現BJ-FA 會使細胞內與 CDK2-cyclin complex 結合的 p21 有增加的現象。Kinase activity assay 發現 BJ-FA 會抑制 CDK2 的活性。利用微血管生成實驗，我們發現 BJ-FA 會抑制內皮細胞微小血管
的生成。因此初步推論BJ-FA 會干擾內皮細胞的細胞週期，進而達到抑制內皮
細胞生長的目的。其作用可能是透過增加抑制細胞週期進行的調控蛋白所引起。
因此，BJ-FA 或許具有潛力能成為抗血管增生的藥物。
英文摘要
The main purpose of this study is to investigate the antiproliferation effect of the compound, BJ-FA, on human umbilical vein endothelial cell (HUVEC), and its underlying molecular mechanisms. In these years, the topics about anti-angiogenesis have become an attractive theme for cancer therapy. It has been also demonstrated that lots of some chemical compounds not only have the utility of cancer therapy, but also can reduce the side effects of currently used drugs. Thus, to search for a
succedaneum much more effective and easily absorbed by human-body seems to be an appropriate approach to substitute the presently used clinical remedy against cancer.
Here we report that BJ-FA induced a dose-dependent inhibition of HUVEC growth.
[3H]-thymidine incorporation and flow cytometry analyses demostrated that the cell cycle was arrested in G0/G1 phase and decreased the DNA synthesis. Western blot analysis revealed that the cell cycle inhibitor protein, p21, was significantly increased in the BJ-FA treated HUVEC as compared with the control. Accordingly, we
hypothesize that BJ-FA may interrupt the progress of cell cycle of HUVEC and then inhibit the angiogenesis through increasing the levels of p21 protein which might inhibit the CDKs. Immuno-precipitation study showed that the CDK2-cyclin-linked p21 in HUVEC were significantly increased after BJ-FA treatment. Using matrix gel we further demonstrated that BJ-FA exerts the anti-capillary-like tube formation activity. Consequently, BJ-FA might have the potential to be an anti-angiogenesis drug.
