Yang Hua, Tian Shi-Ting, Wu Rui-Yun, Chen Yu, Mei Zhi-Nan, Wang Chao-Yuan, Yang Guang-Zhong
a Engineering Research Centre for the Protection and Utilization of Bioresource in Ethnic Area of Southern China, South-Central University for Nationalities , Wuhan 430074 , China.
J Asian Nat Prod Res. 2014 Oct;16(10):991-9. doi: 10.1080/10286020.2014.918961. Epub 2014 Jun 16.
Glycoborinine (GB), a natural carbazole alkaloid isolated from Glycosmis pentaphylla, has been shown to be a potential molecule against cancer cells. In this study, the cell-signaling pathway of its anti-tumor activity was investigated. MTT assay result showed that GB inhibited HepG2 cell proliferation in a dose- and time-dependent manner and 50% inhibiting concentration (IC50) of GB-induced cell death was 39.7 μM for a period of 48 h. GB-induced HepG2 apoptosis was confirmed by Hochest 33258 staining and PI staining. The level of reactive oxygen species (ROS) was measured with H2DCF-DA staining and the change of mitochondrial membrane potential (△Ψ(m)) was analyzed with tetrechloro-tetraethylbenzimidazolcarbocyanine iodide (JC-1) probe. Results showed that GB at 12.5, 25, and 50 μM promoted ROS production. GB induced HepG2 apoptosis through a mitochondrial apoptotic pathway, which was demonstrated by GB-induced increase in the ratio of Bax/Bcl-2, cytochrome C release, the ratio of cleaved caspase-3/procaspase-3, and the ratio of cleaved poly ADP-ribose polymerase (cleaved PARP)/poly ADP-ribose polymerase (PARP). To summarize, this study demonstrated that GB could induce HepG2 apoptosis through the mitochondrial-dependent pathway, which might provide a promising approach to cure liver cancer with GB.
甘柯宁碱（GB）是从五指毛桃中分离得到的一种天然咔唑生物碱，已被证明是一种潜在的抗癌细胞分子。在本研究中，对其抗肿瘤活性的细胞信号通路进行了研究。MTT 法检测结果显示，GB 以剂量和时间依赖性方式抑制 HepG2 细胞增殖，在 48 小时内，GB 诱导细胞死亡的 50%抑制浓度（IC50）为 39.7 μM。通过 Hoechst 33258 染色和 PI 染色证实了 GB 诱导的 HepG2 细胞凋亡。用 H2DCF-DA 染色测定活性氧（ROS）水平，用四氯四乙基苯并咪唑碳氰碘化物（JC-1）探针分析线粒体膜电位（△Ψ(m)）的变化。结果表明，12.5、25 和 50 μM 的 GB 促进了 ROS 的产生。GB 通过线粒体凋亡途径诱导 HepG2 细胞凋亡，这通过 GB 诱导 Bax/Bcl-2 比值增加、细胞色素 C 释放、裂解的半胱天冬酶-3/原半胱天冬酶-3 比值以及裂解的聚 ADP 核糖聚合酶（裂解的 PARP）/聚 ADP 核糖聚合酶（PARP）比值增加得到证明。综上所述，本研究表明 GB 可通过线粒体依赖途径诱导 HepG2 细胞凋亡，这可能为用 GB 治疗肝癌提供一种有前景的方法。
