苦木酮通过下调 Nrf2、YAP 和 c-Myc 的表达抑制顺铂耐药膀胱癌细胞的生长和迁移。
Ailanthone inhibits cell growth and migration of cisplatin resistant bladder cancer cells through down-regulation of Nrf2, YAP, and c-Myc expression.
机构信息
Department of Clinical and Biological Science, University of Turin, Corso Raffaello 30, 10125 Torino, Italy.
Department of Clinical and Biological Science, University of Turin, Corso Raffaello 30, 10125 Torino, Italy.
出版信息
Phytomedicine. 2019 Mar 15;56:156-164. doi: 10.1016/j.phymed.2018.10.034. Epub 2018 Oct 29.
BACKGROUND
Ailanthone (Aila) is a natural active compound isolated from the Ailanthus altissima, which has been shown to possess an "in vitro" growth-inhibitory effect against several cancer cell lines. Advanced bladder cancer is a common disease characterized by a frequent onset of resistance to cisplatin-based therapy. The cisplatin (CDDP) resistance is accompanied by an increase in Nrf2 protein expression which contributes to conferring resistance. Recently, we demonstrated a cross-talk between Nrf2 and YAP. YAP has also been demonstrated to play an important role in chemoresistance of bladder cancer.
PURPOSE
We analyzed the antitumor effect of Aila in sensitive and CDDP-resistant bladder cancer cells and the molecular mechanisms involved in Aila activity.
STUDY DESIGN
Sensitive and CDDP-resistant 253J B-V and 253J bladder cancer cells, intrinsically CDDP-resistant T24 bladder cancer cells and HK-2 human renal cortex cells were used. Cells were treated with diverse concentrations of Aila and proliferation, cell cycle, apoptosis and gene expressions were determined.
METHODS
Aila toxicity and proliferation were determined by MTT and colony forming methods, respectively. Cell cycle was determined by cytofluorimetric analysis through PI staining method. Apoptosis was detected using Annexin V and PI double staining followed by quantitative flow cytometry. Expressions of Nrf2, Yap, c-Myc, and house-keeping genes were determined by western blot with specific antibodies. Cell migration was detected by wound healing and Boyden chamber analysis.
RESULTS
Aila inhibited the growth of sensitive and CDDP-resistant bladder cancer cells with the same effectiveness. On the contrary, the growth of HK-2 cells was only slightly reduced by Aila. Cell cycle analysis revealed an accumulation of Aila-treated bladder cancer cells in the G0/G1 phase. Interestingly, Aila strongly reduced Nrf2 expression in these cell lines. Moreover, Aila significantly reduced YAP, and c-Myc protein expression. The random and the oriented migration of bladder cancer cells were strongly inhibited by Aila treatment, in particular in CDDP-resistant cells.
CONCLUSION
Aila inhibited proliferation and invasiveness of bladder cancer cells. Its high effectiveness in CDDP resistant cells could be related to the inhibition of Nrf2, YAP, and c-Myc expressions. Aila could represent a new tool to treating CDDP-resistant bladder cancers.
背景
樗酮（Aila）是从臭椿中分离出的一种天然活性化合物，已被证明具有抑制几种癌细胞系生长的“体外”作用。晚期膀胱癌是一种常见疾病，其特点是经常对顺铂为基础的治疗产生耐药性。顺铂（CDDP）耐药伴随着 Nrf2 蛋白表达的增加，这有助于赋予耐药性。最近，我们证明了 Nrf2 和 YAP 之间存在串扰。YAP 也已被证明在膀胱癌的化疗耐药中发挥重要作用。
目的
分析樗酮在敏感和 CDDP 耐药膀胱癌细胞中的抗肿瘤作用及其作用机制。
研究设计
使用敏感和 CDDP 耐药的 253J B-V 和 253J 膀胱癌细胞、固有 CDDP 耐药的 T24 膀胱癌细胞和 HK-2 人肾皮质细胞。用不同浓度的 Aila 处理细胞，测定增殖、细胞周期、凋亡和基因表达。
方法
用 MTT 和集落形成方法分别测定 Aila 的毒性和增殖。用 PI 染色法通过细胞流式分析测定细胞周期。用 Annexin V 和 PI 双重染色法检测凋亡，并用定量流式细胞术进行检测。用特异性抗体通过 Western blot 测定 Nrf2、Yap、c-Myc 和管家基因的表达。用划痕愈合和 Boyden 室分析检测细胞迁移。
结果
Aila 以相同的效力抑制敏感和 CDDP 耐药膀胱癌细胞的生长。相反，Aila 对 HK-2 细胞的生长只有轻微的抑制作用。细胞周期分析显示 Aila 处理的膀胱癌细胞在 G0/G1 期积聚。有趣的是，Aila 强烈降低了这些细胞系中的 Nrf2 表达。此外，Aila 显著降低了 YAP 和 c-Myc 蛋白的表达。Aila 处理强烈抑制了膀胱癌细胞的随机和定向迁移，特别是在 CDDP 耐药细胞中。
结论
Aila 抑制了膀胱癌细胞的增殖和侵袭性。其在 CDDP 耐药细胞中的高疗效可能与 Nrf2、YAP 和 c-Myc 表达的抑制有关。Aila 可能成为治疗 CDDP 耐药膀胱癌的新工具。
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