Department of Biochemistry and Molecular Biology, School of Medicine, Medical Research Center for Bioreaction to Reactive Oxygen Species, Biomedical Science Institute, Kyung Hee University, Seoul, 130-701, Republic of Korea.
Apoptosis. 2014 Apr;19(4):682-97. doi: 10.1007/s10495-013-0955-y.
Licochalcone A (LicA), an estrogenic flavonoid, induces apoptosis in multiple types of cancer cells. In this study, the molecular mechanisms underlying the anti-cancer effects of LicA were investigated in HepG2 human hepatocellular carcinoma cells. LicA induced apoptotic cell death, activation of caspase-4, -9, and -3, and expression of endoplasmic reticulum (ER) stress-associated proteins, including C/EBP homologous protein (CHOP). Inhibition of ER stress by CHOP knockdown or treatment with the ER stress inhibitors, salubrinal and 4-phenylbutyric acid, reduced LicA-induced cell death. LicA also induced reactive oxygen species (ROS) accumulation and the anti-oxidant N-acetylcysteine reduced LicA-induced cell death and CHOP expression. In addition, LicA increased the levels of cytosolic Ca(2+), which was blocked by 2-aminoethoxydiphenyl borate (an antagonist of inositol 1,4,5-trisphosphate receptor) and BAPTA-AM (an intracellular Ca(2+) chelator). 2-Aminoethoxydiphenyl borate and BAPTA-AM inhibited LicA-induced cell death. Interestingly, LicA induced phosphorylation of phospholipase Cγ1 (PLCγ1) and inhibition of PLCγ1 reduced cell death and ER stress. Moreover, the multi-targeted receptor tyrosine kinase inhibitors, sorafenib and sunitinib, reduced LicA-induced cell death, ER stress, and cytosolic Ca(2+) and ROS accumulation. Finally, LicA induced phosphorylation of vascular endothelial growth factor receptor 2 (VEGFR2) and c-Met receptor and inhibition of both receptors by co-transfection with VEGFR2 and c-Met siRNAs reversed LicA-induced cell death, Ca(2+) increase, and CHOP expression. Taken together, these findings suggest that induction of ER stress via a PLCγ1-, Ca(2+)-, and ROS-dependent pathway may be an important mechanism by which LicA induces apoptosis in HepG2 hepatocellular carcinoma cells.
甘草查尔酮 A（LicA）是一种具有雌激素样作用的黄酮类化合物，可诱导多种类型的癌细胞凋亡。在这项研究中，我们研究了 LicA 在 HepG2 人肝癌细胞中的抗癌作用的分子机制。LicA 诱导细胞凋亡，激活半胱天冬酶-4、-9 和 -3，并表达内质网（ER）应激相关蛋白，包括 C/EBP 同源蛋白（CHOP）。通过 CHOP 敲低或用 ER 应激抑制剂 salubrinal 和 4-苯基丁酸处理抑制 ER 应激，可减少 LicA 诱导的细胞死亡。LicA 还诱导活性氧（ROS）积累，抗氧化剂 N-乙酰半胱氨酸可减少 LicA 诱导的细胞死亡和 CHOP 表达。此外，LicA 增加了细胞质 Ca（2+）水平，而 2-氨基乙氧基二苯硼酸盐（三磷酸肌醇受体拮抗剂）和 BAPTA-AM（细胞内 Ca（2+）螯合剂）可阻断该作用。2-氨基乙氧基二苯硼酸盐和 BAPTA-AM 抑制 LicA 诱导的细胞死亡。有趣的是，LicA 诱导磷脂酶 Cγ1（PLCγ1）磷酸化，抑制 PLCγ1 可减少细胞死亡和 ER 应激。此外，多靶点受体酪氨酸激酶抑制剂索拉非尼和舒尼替尼可减少 LicA 诱导的细胞死亡、ER 应激以及细胞质 Ca（2+）和 ROS 积累。最后，LicA 诱导血管内皮生长因子受体 2（VEGFR2）和 c-Met 受体磷酸化，并用 VEGFR2 和 c-Met siRNA 共转染逆转 LicA 诱导的细胞死亡、Ca（2+）增加和 CHOP 表达。总之，这些发现表明，通过 PLCγ1、Ca（2+）和 ROS 依赖性途径诱导 ER 应激可能是 LicA 诱导 HepG2 肝癌细胞凋亡的重要机制。
