Inhibition of TPA-induced protein kinase C and transcription activator protein-1 binding activities by theaflavin-3,3′-digallate from black tea in NIH3T3 cells
Inhibition of TPA-induced protein kinase C and transcription activator protein-1 binding activities by theaflavin-3,3′-digallate from black tea in NIH3T3 cells
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DOI：
10.1021/jf981099k
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发表时间：
1999-04-01
影响因子：
6.1
通讯作者：
Lin, JK
中科院分区：
文献类型：
--
作者：
Chen, YC;Liang, YC;Lin, JK
茶是世界上最受欢迎的饮料之一。几份报告表明，绿色茶和红茶都能够抑制动物模型中的肿瘤细胞增殖。本研究以茶黄素（TF-1）、茶黄素-3-没食子酸酯（TF-2a）、茶黄素-3 '-没食子酸酯（TF-2b）的混合物（TF-2）、茶黄素-3，3'-二没食子酸酯（TF-3）、茶红素（TR）、和主要的绿色茶多酚（-）-表没食子儿茶素-3-没食子酸酯（EGCG）对12-O-十四烷酰佛波醇13-乙酸酯（TPA）诱导的蛋白激酶C（PKC）和转录激活蛋白-1（AP-1）的影响在NIH 3 T3细胞中的结合活性。在用部分纯化的制备物分析PKC活性时，TPA（100 ng/mL）处理能够将膜相关的PKC活性提高约3倍，并且用TF-3（20 μ M）和EGCG（20 μ M）处理分别显示出对TPA诱导的PKC活性的94.5%和9.4%的抑制。在TPA处理的NIH 3 T3细胞中检测到PKC α蛋白从胞浆到膜的易位，TF-3能够阻断其易位。利用髓鞘碱性蛋白（MBP）作为PKC特异性底物进行体外激酶活性测定，结果发现TPA处理后MBP蛋白磷酸化程度增加，TF-3对其磷酸化有最强的抑制作用，而EGCG的抑制作用较弱。采用电泳迁移率变动法（electrophoretic mobility shift assay，EBT）和北方印迹法（northern blot）分析了茶多酚对AP-1结合活性的抑制作用，并采用western blot分析了茶多酚对c-Jun基因表达的抑制作用。本研究结果为了解茶多酚对TPA促肿瘤作用的抑制作用提供了新的分子基础。
Tea is one of the most popular beverages in the world. Several reports have shown that both green tea and black tea were able to inhibit tumor cell proliferation in animal models. In this study, we investigated the inhibitory effects of black tea polyphenols including theaflavin (TF-1), the mixture (TF-2) of theaflavin-3-gallate (TF-2a), and theaflavin-3'-gallate (TF-2b), theaflavin-3,3'-digallate (TF-3), thearubigin (TR), and a major green tea polyphenol (-)-epigallocatechin-3-gallate (EGCG) on 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced protein kinase C (PKC) and transcription activator protein-1 (AP-1) binding activities in NIH3T3 cells. On analysis of PKC activity with partial purified preparation, TPA (100 ng/mL) treatment was able to elevate membrane-associated PKC activity similar to 3-fold, and treatment with TF-3 (20 mu M) and EGCG (20 mu M) showed 94.5% and 9.4%; suppression on TPA-induced PKC activity, respectively. Translocation of PKC alpha protein from cytosol to membrane was detected in TPA-treated NIH3T3 cells, and TF-3 was able to block its translocation. By in vitro kinase assay using myelin basic protein (MBP) as a PKC-specific substrate, we found that TPA treatment was able to increase PKC kinase activity by detection of phosphorylated MBP protein and TF-3 showed strongest inhibitory effect on its phosphorylation while EGCG was shown to be less effective. We also analyzed the AP-1 binding activity by electrophoretic mobility shift assay and c-Jun gene expression by northern blot and western blot, the results showed that TF-3 is the most potent inhibitor on TPA-induced AP-1 binding activity and c-Jun gene expression among these five tea polyphenols. Our results might provide new molecular basis for understanding the inhibitory effects of tea polyphenols on TPA-mediated tumor promotion.