Aloperine induces apoptosis and G2/M cell cycle arrest in hepatocellular carcinoma cells through the PI3K/Akt signaling pathway
Aloperine induces apoptosis and G2/M cell cycle arrest in hepatocellular carcinoma cells through the PI3K/Akt signaling pathway
复制标题
Aloperine 通过 PI3K/Akt 信号通路诱导肝细胞癌细胞凋亡和 G2/M 细胞周期阻滞
DOI：
10.1016/j.phymed.2019.152843
复制
发表时间：
2019-08-01
期刊：
影响因子：
7.9
通讯作者：
Yu, Zhi-Ling
中科院分区：
文献类型：
--
作者：
Liu, Jun-Shan;Huo, Chu-Ying;Yu, Zhi-Ling
背景：肝细胞癌（HCC）在全世界癌症相关死亡的最常见原因中排名第三。目前，肝癌的化疗仍显不足。在从传统中药中寻找有效的抗肝癌药物的过程中，我们发现苦豆碱（aloperine，ALO）是一种从苦豆子（Sophora alopecuroides L.）发挥抗HCC活性。然而，ALO对肝癌的作用研究较少，其机制尚不清楚。目的：本研究旨在评价ALO的抗肝癌活性，并探讨其作用机制。方法：采用MTT法和集落形成实验观察ALO对人肝癌细胞株Hep 3B和Huh 7增殖的抑制作用。Hoechst 33258染色观察ALO作用后细胞形态学变化。流式细胞仪分析细胞凋亡诱导、线粒体膜电位的崩溃和细胞周期的分布。Western blotting检测细胞凋亡和细胞周期阻滞相关蛋白以及PI 3 K/Akt信号通路中关键蛋白的表达水平。采用小干扰RNA（siRNA）转染方法研究Akt在ALO诱导的细胞凋亡和细胞周期阻滞中的作用。结果：ALO对肝癌细胞株Hep 3B和Huh 7的增殖有明显的抑制作用。ALO可诱导肝癌细胞凋亡，并伴有线粒体电位下降、细胞色素c释放、caspase-9、caspase-3和PARP裂解增加。ALO通过下调cdc 25 C、cdc 2和cyclin B1的表达，诱导G2/M期细胞阻滞。此外，ALO通过降低p110 α、p85、Akt和p-Akt的表达水平来抑制PI 3 K/Akt信号通路的激活（（）（Ser 473）（））。进一步的研究表明，通过siRNA抑制Akt增强ALO介导的肝癌细胞凋亡和G2/M期细胞阻滞。结论：ALO通过抑制PI 3 K/Akt信号通路诱导肝癌细胞凋亡，并使肝癌细胞G2/M期阻滞。本研究为ALO作为一种潜在的肝癌化疗药物提供了理论依据。
Background: Hepatocellular carcinoma (HCC) ranks third among the most common causes of cancer-related deaths worldwide. The chemotherapy for HCC is still insufficient, so far. In searching for effective anti-HCC agents from traditional Chinese medicine, we discovered that aloperine (ALO), a quinolizidine alkaloid from Sophora alopecuroides L., exerts anti-HCC activities. However, the effects of ALO on HCC have been rarely studied, and its underlying mechanisms remain unknown.Purpose: This study aims to evaluate the anti-HCC activities of ALO and explore its underlying mechanisms. Methods: MTT assay and colony formation assay were used to investigate the anti-proliferative effects of ALO on human HCC Hep3B and Huh7 cells. Hoechst 33258 staining was used to observe the morphological changes of cells after ALO treatment. Flow cytometry was used to analyze apoptosis induction, the collapse of the mitochondrial membrane potential and cell cycle distribution. Western blotting was used to examine the expression levels of proteins associated with apoptosis and cell cycle arrest, and key proteins in the PI3K/Akt signaling pathway. Small interfering RNA (siRNA) transfection was used to investigate the role of Akt in ALO-induced apoptosis and cell cycle arrest. Zebrafish tumor model was used to evaluate the anti-HCC effects of ALO in vivo.Results: ALO inhibited the proliferation of Hep3B and Huh7 cells. ALO induced apoptosis in HCC cells, which was accompanied by the loss of mitochondrial potential, the release of cytochrome c into cytosol, as well as the increased cleavages of caspase-9, caspase-3 and PARP. Moreover, ALO induced G2/M cell cycle arrest by downregulating the expression levels of cdc25C, cdc2 and cyclin B1. In addition, ALO inhibited activation of the PI3K/Akt signaling pathway by decreasing the expression levels of p110 alpha, p85, Akt and p-Akt (()(Ser473)()). Further study showed that inhibition of Akt by siRNA augmented ALO-mediated apoptosis and G2/M cell cycle arrest in HCC cells. Critically, ALO inhibited the growth of Huh7 cells in vivo.Conclusion: We first demonstrated that ALO induced apoptosis and G2/M cell cycle arrest in HCC cells through inhibition of the PI3K/Akt signaling pathway. This study provides a rationale for ALO as a potential chemotherapeutic agent for HCC.