Department of Pediatrics, Jining No. 1 People's Hospital, Jining, Shandong, China.
J Cell Biochem. 2018 Dec;119(12):9866-9877. doi: 10.1002/jcb.27307. Epub 2018 Aug 28.
Medulloblastoma is a primitive neuroectodermal-derived brain tumor and the most common malignant brain tumor in children. Triptolide (TPL) is the major active component extracted from Tripterygium wilfordii Hook F. This study aimed to explore the effects of TPL on medulloblastoma cell proliferation, migration, and apoptosis, as well as the underlying possible molecular mechanism. Viability, proliferation, and apoptosis of Daoy cells were measured using cell counting kit-8 assay, 5-bromo-2'-deoxyuridine incorporation assay, and Guava Nexin assay, respectively. Cell migration was detected using two-chamber transwell assay and wound healing assay. Quantitative reverse transcription polymerase chain reaction (qRT-PCR) was performed to determine the relative expression of microRNA-138 (miR-138) in Daoy cells. Cell transfection was used to change the expression of miR-138 in cells. Western blot analysis was used to analyze the expression of key factors involved in cell apoptosis, cell migration, the phosphatidylinositol 3-kinase (PI3K)/protein kinase 3 (AKT) pathway, and the Notch pathway in Daoy cells. We found that TPL significantly inhibited the viability, proliferation, and migration of Daoy cells but promoted Daoy cell apoptosis. The expression levels of matrix metalloproteinases (MMP)-2 and MMP-9 after TPL treatment were decreased. The expression of miR-138 in Daoy cells after TPL treatment was increased. Suppression of miR-138 obviously reversed the TPL-induced Daoy cell proliferation, migration inhibition, and cell apoptosis enhancement, as well as the inactivation of the PI3K/AKT and Notch pathways. Cyclin-dependent kinase 6 (CDK6) was a direct target gene of miR-138, which might be involved in the antitumor effects of TPL on Daoy cells. In conclusion, our study verified that TPL exerted anticancer effects on medulloblastoma cells possibly via upregulating miR-138 and inactivating the PI3K/AKT and Notch pathways.
髓母细胞瘤是一种原始神经外胚层来源的脑肿瘤，也是儿童中最常见的恶性脑肿瘤。雷公藤红素（TPL）是从雷公藤中提取的主要活性成分。本研究旨在探讨 TPL 对髓母细胞瘤细胞增殖、迁移和凋亡的影响及其潜在的分子机制。使用细胞计数试剂盒-8 测定法、5-溴-2'-脱氧尿苷掺入测定法和 Guava Nexin 测定法分别测定 Daoy 细胞的活力、增殖和凋亡。使用双室 Transwell 测定法和划痕愈合测定法检测细胞迁移。定量逆转录聚合酶链反应（qRT-PCR）用于测定 Daoy 细胞中 microRNA-138（miR-138）的相对表达。细胞转染用于改变细胞中 miR-138 的表达。Western blot 分析用于分析 Daoy 细胞中细胞凋亡、细胞迁移、磷脂酰肌醇 3-激酶（PI3K）/蛋白激酶 B（AKT）途径和 Notch 途径中涉及的关键因子的表达。我们发现 TPL 显著抑制 Daoy 细胞的活力、增殖和迁移，但促进 Daoy 细胞凋亡。TPL 处理后基质金属蛋白酶（MMP）-2 和 MMP-9 的表达水平降低。TPL 处理后 Daoy 细胞中 miR-138 的表达增加。抑制 miR-138 明显逆转了 TPL 诱导的 Daoy 细胞增殖、迁移抑制和细胞凋亡增强，以及 PI3K/AKT 和 Notch 途径失活。细胞周期蛋白依赖性激酶 6（CDK6）是 miR-138 的直接靶基因，可能参与了 TPL 对 Daoy 细胞的抗癌作用。总之，本研究证实 TPL 通过上调 miR-138 并使 PI3K/AKT 和 Notch 途径失活，对髓母细胞瘤细胞发挥抗癌作用。
