Huang Jian-Lan, Liang Li, Xie Pei-En, Sun Wei-Liang, Wang Li, Cai Zheng-Wen
Department of Medical Oncology, The Second Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi Zhuang Autonomous Region 530007, P.R. China.
Exp Ther Med. 2023 Sep 1;26(4):484. doi: 10.3892/etm.2023.12183. eCollection 2023 Oct.
Cucurbitacin B (CuB) is a member of the cucurbitacin family, which has shown potent anticancer pharmacological activity. Prolonged or severe endoplasmic reticulum stress (ERS) induces apoptosis; therefore, the present study investigated whether CuB may activate the ERS pathway to induce apoptosis. HT-29 and SW620 colorectal cancer (CRC) cells were treated with a range of concentrations of CuB for 48 h, and the viability and proliferation of cells were determined using Cell Counting Kit 8 (CCK8) and colony formation assays. Subsequently, the appropriate CuB concentration (5 µM) was selected for treatment of CRC cells for 48 h. Western blot analysis was used to measure the expression levels of ERS-related proteins, flow cytometry was used to evaluate apoptosis, the dichlorodihydrofluorescein diacetate fluorescent probe was used to detect reactive oxygen species (ROS) production, and the relationship between ROS and ERS was determined by western blot analysis. Furthermore, flow cytometry was used to evaluate apoptosis after treatment with the ERS inhibitor 4-phenylbutyric acid, the ROS inhibitor N-acetylcysteine and following knockdown of CHOP expression. In addition, western blot analysis was performed to measure Bax and Bcl2 protein expression levels, and a CCK8 assay was performed to evaluate the viability of cells following knockdown of CHOP. Notably, CuB treatment increased apoptosis and inhibited cell proliferation in CRC cell lines, and these effects were mediated by ROS and ROS-regulated activation of the PERK and XBP1 ERS pathways. In conclusion, CuB may induce apoptosis in HT-29 and SW620 CRC cells via ROS and ERS.
葫芦素B（CuB）是葫芦素家族的一员，已显示出强大的抗癌药理活性。长期或严重的内质网应激（ERS）会诱导细胞凋亡；因此，本研究调查了CuB是否可能激活ERS途径以诱导细胞凋亡。用一系列浓度的CuB处理HT-29和SW620结肠直肠癌（CRC）细胞48小时，并使用细胞计数试剂盒8（CCK8）和集落形成试验测定细胞的活力和增殖。随后，选择合适的CuB浓度（5μM）处理CRC细胞48小时。采用蛋白质免疫印迹分析来检测ERS相关蛋白的表达水平，流式细胞术用于评估细胞凋亡，二氯二氢荧光素二乙酸酯荧光探针用于检测活性氧（ROS）的产生，并且通过蛋白质免疫印迹分析确定ROS与ERS之间的关系。此外，流式细胞术用于评估在用ERS抑制剂4-苯基丁酸、ROS抑制剂N-乙酰半胱氨酸处理后以及CHOP表达敲低后的细胞凋亡情况。另外，进行蛋白质免疫印迹分析以检测Bax和Bcl2蛋白的表达水平，并进行CCK8试验以评估CHOP敲低后细胞的活力。值得注意的是，CuB处理增加了CRC细胞系中的细胞凋亡并抑制了细胞增殖，并且这些作用是由ROS以及ROS调节的PERK和XBP1 ERS途径的激活介导的。总之，CuB可能通过ROS和ERS诱导HT-29和SW620 CRC细胞凋亡。
