Chen Wen-Liang, Barszczyk Andrew, Turlova Ekaterina, Deurloo Marielle, Liu Baosong, Yang Burton B, Rutka James T, Feng Zhong-Ping, Sun Hong-Shuo
Department of Surgery, University of Toronto, Toronto, Canada.
Department of Physiology, University of Toronto, Toronto, Canada.
Oncotarget. 2015 Jun 30;6(18):16321-40. doi: 10.18632/oncotarget.3872.
Glioblastomas are progressive brain tumors with devastating proliferative and invasive characteristics. Ion channels are the second largest target class for drug development. In this study, we investigated the effects of the TRPM7 inhibitor carvacrol on the viability, resistance to apoptosis, migration, and invasiveness of the human U87 glioblastoma cell line.The expression levels of TRPM7 mRNA and protein in U87 cells were detected by RT-PCR, western blotting and immunofluorescence. TRPM7 currents were recorded using whole-cell patch-clamp techniques. An MTT assay was used to assess cell viability and proliferation. Wound healing and transwell experiments were used to evaluate cell migration and invasion. Protein levels of p-Akt/t-Akt, p-ERK1/2/t-ERK1/2, cleaved caspase-3, MMP-2 and phosphorylated cofilin were also detected.TRPM7 mRNA and protein expression in U87 cells is higher than in normal human astrocytes. Whole-cell patch-clamp recording showed that carvacrol blocks recombinant TRPM7 current in HEK293 cells and endogenous TRPM7-like current in U87 cells. Carvacrol treatment reduced the viability, migration and invasion of U87 cells. Carvacrol also decreased MMP-2 protein expression and promoted the phosphorylation of cofilin. Furthermore, carvacrol inhibited the Ras/MEK/MAPK and PI3K/Akt signaling pathways.Therefore, carvacrol may have therapeutic potential for the treatment of glioblastomas through its inhibition of TRPM7 channels.
胶质母细胞瘤是具有毁灭性增殖和侵袭特性的进行性脑肿瘤。离子通道是药物开发的第二大靶点类别。在本研究中，我们研究了TRPM7抑制剂香芹酚对人U87胶质母细胞瘤细胞系的活力、抗凋亡能力、迁移和侵袭能力的影响。通过RT-PCR、蛋白质印迹法和免疫荧光检测U87细胞中TRPM7 mRNA和蛋白质的表达水平。使用全细胞膜片钳技术记录TRPM7电流。采用MTT法评估细胞活力和增殖。采用伤口愈合实验和Transwell实验评估细胞迁移和侵袭。还检测了p-Akt/t-Akt、p-ERK1/2/t-ERK1/2、裂解的caspase-3、MMP-2和磷酸化丝切蛋白的蛋白水平。U87细胞中TRPM7 mRNA和蛋白质的表达高于正常人星形胶质细胞。全细胞膜片钳记录显示，香芹酚可阻断HEK293细胞中的重组TRPM7电流以及U87细胞中的内源性TRPM7样电流。香芹酚处理降低了U87细胞的活力、迁移和侵袭能力。香芹酚还降低了MMP-2蛋白的表达并促进了丝切蛋白的磷酸化。此外，香芹酚抑制了Ras/MEK/MAPK和PI3K/Akt信号通路。因此，香芹酚可能通过抑制TRPM7通道而具有治疗胶质母细胞瘤的潜力。
