Chongqing productivity promotion center for the modernization of Chinese traditional medicine, School of Pharmaceutical Sciences, Southwest University, Chongqing, 400716, China.
School of Life Sciences, Southwest University, Chongqing, 400715, China; Chongqing Engineering Research Center for Pharmaceutical Process and Quality Control, Southwest University, Chongqing, 400716, China.
Phytomedicine. 2018 Sep 15;48:152-160. doi: 10.1016/j.phymed.2017.12.027. Epub 2017 Dec 26.
Colorectal cancer is the third leading cause of cancer-related deaths in the word. Coptisine (COP), an isoquinoline alkaloid derived from Coptis chinensis Franch, possesses a wide variety of pharmacological effects. However, its anti-proliferative effect on colon cancer is not fully elucidated. In the present study, we aimed to ascertain whether COP inhibits HCT-116 cell growth and to further explore the molecular mechanism in vitro and in vivo.
Cell viability was determined by MTT assay. Cell migration was detected using wound healing assay. Apoptosis, mitochondrial membrane potential (Δψ) and reactive oxygen species (ROS) was analysis via flow cytometry. Hoechst 33342 was used for morphology observation. The expression levels of proteins related to mitochondrial-mediated apoptotic pathway were detected by western blotting. In addition, the antitumor ability of COP was further measured in athymic nude mice.
COP significantly decreased cell viability and migration in HCT-116 cells. Flow cytometry and Hoechst 33342 analysis confirmed that COP suppressed cell proliferation by inducing apoptosis. COP decreased Δψ dose-dependently and induced intracellular ROS production time-dependently. Western blotting showed that COP activated mitochondrial-associated apoptosis by down-regulating Bcl-2, Bcl-XL, pro-caspase 3, XIAP level and up-regulating Bax, Bad, cytochrome c, Apaf-1, AIF and cleaved caspase-3 expression. In addition, COP also attenuated PI3K/Akt signaling pathway. In vivo study showed that 150 mg/kg COP significantly delayed the tumor development in BALB/c nude mice. Immunohistochemical analysis also confirmed the activated apoptosis in tumor tissue.
The results demonstrated that COP induces apoptosis in HCT-116 cells through PI3K/Akt and mitochondrial-associated apoptotic pathway. Our findings suggest that COP has potential to be a therapeutic candidate for colon cancer patients.
结直肠癌是全球癌症相关死亡的第三大原因。黄连碱（COP）是从黄连中提取的一种异喹啉生物碱，具有广泛的药理作用。然而，其对结肠癌的抗增殖作用尚未完全阐明。本研究旨在确定 COP 是否抑制 HCT-116 细胞生长，并进一步探讨其在体外和体内的分子机制。
采用 MTT 法测定细胞活力。划痕愈合实验检测细胞迁移。通过流式细胞术分析细胞凋亡、线粒体膜电位（Δψ）和活性氧（ROS）。Hoechst 33342 用于形态观察。Western blot 检测与线粒体介导的凋亡途径相关的蛋白表达水平。此外，还在裸鼠体内进一步测量了 COP 的抗肿瘤能力。
COP 显著降低 HCT-116 细胞的细胞活力和迁移。流式细胞术和 Hoechst 33342 分析证实，COP 通过诱导细胞凋亡抑制细胞增殖。COP 呈剂量依赖性降低 Δψ，并呈时间依赖性诱导细胞内 ROS 产生。Western blot 显示，COP 通过下调 Bcl-2、Bcl-XL、前半胱天冬酶 3、XIAP 水平和上调 Bax、Bad、细胞色素 c、凋亡蛋白酶激活因子 1、凋亡诱导因子和半胱天冬酶-3 的表达来激活线粒体相关凋亡。此外，COP 还抑制了 PI3K/Akt 信号通路。体内研究表明，150mg/kg COP 显著延缓了 BALB/c 裸鼠肿瘤的发展。免疫组织化学分析也证实了肿瘤组织中凋亡的激活。
研究结果表明，COP 通过 PI3K/Akt 和线粒体相关凋亡途径诱导 HCT-116 细胞凋亡。我们的研究结果表明，COP 有可能成为结肠癌患者的治疗候选药物。
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