Modulation of the Nrf2 signalling pathway in Hct116 colon carcinoma cells by baicalein and its methylated derivative negletein
Modulation of the Nrf2 signalling pathway in Hct116 colon carcinoma cells by baicalein and its methylated derivative negletein
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DOI：
10.3109/13880209.2015.1104703
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发表时间：
2016-09-01
影响因子：
3.8
通讯作者：
Waetjen, Wim
中科院分区：
文献类型：
--
作者：
Havermann, Susannah;Chovolou, Yvonni;Waetjen, Wim
背景：黄芩素是黄芩（唇形科）提取物中的主要化合物，在中药中用于治疗炎症和胃肠道疾病。该黄酮类化合物是Nrf2信号通路的激活剂，但其分子机制尚不清楚。目的：通过蛋白酶体活性、自由基清除活性分析以及与黄芩素衍生物的比较，研究黄芩素介导的Hct116细胞中Nrf2激活的分子模式。材料与方法：自由基清除活性（TEAC、DCF）高达25M，细胞毒性在 Hct116 人结肠癌细胞中，分析了浓度高达 40 或 50M 的化合物的浓度高达 100M（MTT 测定，48 小时）的蛋白酶体活性和 Nrf2 激活（荧光素酶测定、泛素化、蛋白质印迹、Ser40 磷酸化；孵育 1 或 4 小时）。结果：Nrf2 的泛素化、蛋白酶体活性和 Nrf2 活性没有变化。可检测到 NRF2 基因的转录。黄芩素降低 Nrf2 的磷酸化（IC(50-) 值约为 20M），表明黄酮类化合物对蛋白激酶具有抑制作用。由于黄芩素对 Nrf2 途径的激活也可能是由于该化合物的氧化还原活性，因此我们研究了甲基化黄芩素衍生物 Oroxylin A、negeletein 和黄芩素三甲醚的作用。 Oroxylin A 和 negletein 显示出相当的氧化还原活性电位，但只有 negletein (50M，4h) 能够激活 Nrf2。 结论：该结果证实了黄芩素（黄芩提取物中的一种成分）的假设，可引起 Nrf2 的激活，而与细胞氧化还原电位的调节无关。
Context: Baicalein is a major compound in extracts derived from Scutellaria baicalensis Georgi (Lamiaceae) which are used in the Traditional Chinese Medicine for the treatment of inflammatory and gastrointestinal diseases. This flavonoid is an activator of the Nrf2 signalling pathway but the molecular mechanism is not clearly established.Objective: We investigated the molecular mode of baicalein-mediated Nrf2-activation in Hct116 cells by the analysis of proteasomal activity, radical-scavenging activity and the comparison with baicalein derivatives.Materials and methods: The radical-scavenging activity (TEAC, DCF) up to 25M, cytotoxicity (MTT assay, 48h) up to 100M, proteasomal activity and the Nrf2-activation (luciferase assay, ubiquitinylation, western blot, Ser40-phosphorylation; incubation for 1 or 4h) by concentrations up to 40 or 50M of the compounds were analysed in Hct116 human colon carcinoma cells.Results: No change in the ubiquitinylation of Nrf2, proteasomal activity and transcription of the NRF2 gene were detectable. Baicalein decreased the phosphorylation of Nrf2 (IC(50-)value approximately 20M) suggesting an inhibitory effect of the flavonoid on protein kinases. Since the activation of the Nrf2 pathway by baicalein might be also due to redox-activity of the compound, we investigated the effects of methylated baicalein derivatives oroxylin A, negeletein and baicaleintrimethylether. Oroxylin A and negletein showed a comparable redox-active potential, but only negletein (50M, 4h) was able to activate Nrf2.Conclusion: This result confirms the hypothesis that baicalein, a component of extracts derived from Baical Skullcap, causes an activation of Nrf2 independent of a modulation of the cellular redox potential.