Laboratory of Cell Biology, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka, 569-1094, Japan.
Laboratory of Cell Biology, Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara, Takatsuki, Osaka, 569-1094, Japan.
Biochem Biophys Res Commun. 2020 Jul 12;528(1):227-233. doi: 10.1016/j.bbrc.2020.05.020. Epub 2020 May 29.
We investigated the novel molecular mechanisms of the antitumor effect of berberine. In this study, two different human cell lines (breast cancer MCF7 cells and non-tumorigenic epithelial MCF12A cells) were treated with various concentrations of berberine. Treatment with 1 and 10 μM berberine inhibited proliferation with G/G cell cycle arrest in both cell lines, and treatment with 100 μM berberine triggered a marked level of cell death in MCF7 cells but not in MCF12A cells. Berberine increased the level of p53 protein and of its target p21 both time- and dose-dependently in MCF7 cells. At any concentration of berberine, immediate uptake (within 15 min) followed by predominantly mitochondrial accumulation were observed by confocal microscopy in both cell lines. At high concentrations (10 or 100 μM), accumulation in the nucleolus became prominent after the transition to the nucleoplasm, especially remarkable in MCF7 cells. Therefore, we evaluated the possibility of berberine-induced nucleolar stress and observed the disappearance of ribosomal protein (RP)L5 from the nucleolus and accumulation of p53 protein in the nucleus after treatment with 10 or 100 μM berberine in MCF7 cells. We also detected the accumulation of RPL5 and RPL11 in the nucleoplasm fraction where they bind to Mdm2. Moreover, downregulation of RPL5 inhibited berberine-driven induction of p53 and p21 and cell death in MCF7 cells. Whereas, in MCF12A cells, down-regulation of RPL5 had little effect on the growth inhibitory effect of high concentration of berberine. These results indicated that cell growth inhibition and cell death induced by higher doses (>10 μM) of berberine in MCF7 cells were due to the upregulation of p53 under the nucleolar stress response caused by a significant accumulation of berberine in the nucleoli.
我们研究了小檗碱抗肿瘤作用的新分子机制。在这项研究中，用不同浓度的小檗碱处理两种不同的人细胞系（乳腺癌 MCF7 细胞和非致瘤上皮 MCF12A 细胞）。用 1 和 10 μM 小檗碱处理，两种细胞系的细胞增殖均受到抑制，并出现 G1/G0 期细胞周期阻滞，用 100 μM 小檗碱处理，MCF7 细胞出现明显的细胞死亡，但 MCF12A 细胞没有。小檗碱在 MCF7 细胞中时间和剂量依赖性地增加了 p53 蛋白及其靶基因 p21 的水平。在任何浓度的小檗碱作用下，在两种细胞系中，通过共聚焦显微镜观察到小檗碱立即摄取（在 15 分钟内），随后主要在线粒体中积累。在高浓度（10 或 100 μM）时，在向核质转移后，核仁内的积累变得明显，在 MCF7 细胞中尤为明显。因此，我们评估了小檗碱诱导核仁应激的可能性，并观察到在 MCF7 细胞中用 10 或 100 μM 小檗碱处理后，核糖体蛋白（RP）L5 从核仁消失，p53 蛋白在核内积累。我们还检测到 RPL5 和 RPL11 在核质部分的积累，在核质部分它们与 Mdm2 结合。此外，下调 RPL5 抑制了 MCF7 细胞中小檗碱驱动的 p53 和 p21 的诱导和细胞死亡。然而，在 MCF12A 细胞中，下调 RPL5 对高浓度小檗碱的生长抑制作用影响不大。这些结果表明，在 MCF7 细胞中，高剂量（>10 μM）的小檗碱诱导的细胞生长抑制和细胞死亡是由于核仁中小檗碱的大量积累引起核仁应激反应导致 p53 的上调。
