Yanarojana Mongkol, Nararatwanchai Thamthiwat, Thairat Sarut, Tancharoen Salunya
School of Anti-Aging and Regenerative Medicine, Mae Fah Luang University, Bangkok, Thailand.
Department of Pharmacology, Faculty of Dentistry, Mahidol University, Bangkok, Thailand.
Anticancer Res. 2017 Dec;37(12):6619-6628. doi: 10.21873/anticanres.12119.
BACKGROUND/AIM: To analyze the apoptotic effect of Houttuynia cordata Thunb (HCT) extract on human melanoma A375 cells and its underlying mechanisms.
The effects of HCT on cell death were determined using the MTT assay. Hoechst 33342 staining was conducted to confirm the detection of cell apoptosis. Caspase-3 and caspase-8 mRNA and cleaved protein levels were investigated by RT-PCR and western blotting, respectively. The release of high mobility group box 1 (HMGB1) and phosphorylation of mitogen-activated protein kinase (MAPK) were determined by ELISA.
Caspase-3 and caspase-8 specific inhibitors suppressed HCT-induced cell death. HCT increased caspase-3 and caspase-8 mRNA, protein levels, and caspase activities in a concentration- and time-dependent manner. HCT induced MAPK phosphorylation in a time-dependent fashion. Pretreatment of cells with a selective inhibitor of p38 MAPK reduced apoptosis and reversed the levels of HMGB1 release in response to HCT treatment.
HCT induces A375 programmed cell death by activating the caspase-dependent pathway and by p38 phosphorylation associated with HMGB1 reduction.
背景/目的：分析鱼腥草提取物对人黑色素瘤A375细胞的凋亡作用及其潜在机制。
采用MTT法测定鱼腥草对细胞死亡的影响。进行Hoechst 33342染色以确认细胞凋亡检测。分别通过RT-PCR和蛋白质印迹法研究半胱天冬酶-3和半胱天冬酶-8的mRNA及裂解蛋白水平。通过酶联免疫吸附测定法测定高迁移率族蛋白B1（HMGB1）的释放和丝裂原活化蛋白激酶（MAPK）的磷酸化。
半胱天冬酶-3和半胱天冬酶-8特异性抑制剂抑制了鱼腥草诱导的细胞死亡。鱼腥草以浓度和时间依赖性方式增加半胱天冬酶-3和半胱天冬酶-8的mRNA、蛋白水平及半胱天冬酶活性。鱼腥草以时间依赖性方式诱导MAPK磷酸化。用p38 MAPK选择性抑制剂预处理细胞可减少细胞凋亡，并逆转鱼腥草处理后HMGB1的释放水平。
鱼腥草通过激活半胱天冬酶依赖性途径以及与HMGB1减少相关的p38磷酸化诱导A375细胞程序性死亡。
