Park Jun Chul, Chang In Bok, Ahn Jun Hyong, Kim Ji Hee, Song Joon Ho, Moon Seung Myung, Park Young-Han
Department of Neurosurgery, Hallym University Sacred Heart Hospital, Anyang, Korea.
Department of Neurosurgery, Dongtan Sacred Heart Hospital, Hwaseong, Korea.
J Korean Neurosurg Soc. 2018 Jul;61(4):441-449. doi: 10.3340/jkns.2017.0219. Epub 2018 Jul 1.
Notch receptors are heterodimeric transmembrane proteins that regulate cell fate, such as differentiation, proliferation, and apoptosis. Dysregulated Notch pathway signaling has been observed in glioblastomas, as well as in other human malignancies. Nerve growth factor (NGF) is essential for cell growth and differentiation in the nervous system. Recent reports suggest that NGF stimulates glioblastoma proliferation. However, the relationship between NGF and Notch1 in glioblastomas remains unknown. Therefore, we investigated expression of Notch1 in a glioblastoma cell line (U87-MG), and examined the relationship between NGF and Notch1 signaling.
We evaluated expression of Notch1 in human glioblastomas and normal brain tissues by immunohistochemical staining. The effect of NGF on glioblastoma cell line (U87-MG) was evaluated by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. To evaluate the relationship between NGF and Notch1 signaling, Notch1 and Hes1 expression were evaluated by reverse transcription polymerase chain reaction (RT-PCR) and Western blot analysis, respectively. To confirm the effects of NGF on Notch1 signaling, Notch1 and Hes1 small interfering RNAs (siRNAs) were used.
In immunohistochemistry, Notch1 expression was higher in glioblastoma than in normal brain tissue. MTT assay showed that NGF stimulates U87-MG cells in a dose-dependent manner. RT-PCR and Western blot analysis demonstrated that Notch1 and Hes1 expression were increased by NGF in a dose-dependent manner. After transfection with Notch1 and Hes1 siRNAs, there was no significant difference between controls and 100 nM NGF-β, which means that U87-MG cell proliferation was suppressed by Notch1 and Hes1 siRNAs.
These results indicate that NGF stimulates glioblastoma cell proliferation via Notch1 signaling through Hes 1.
Notch受体是异二聚体跨膜蛋白，可调节细胞命运，如分化、增殖和凋亡。在胶质母细胞瘤以及其他人类恶性肿瘤中均观察到Notch信号通路失调。神经生长因子（NGF）对神经系统中的细胞生长和分化至关重要。最近的报告表明，NGF可刺激胶质母细胞瘤增殖。然而，NGF与胶质母细胞瘤中Notch1之间的关系仍不清楚。因此，我们研究了胶质母细胞瘤细胞系（U87-MG）中Notch1的表达，并检测了NGF与Notch1信号传导之间的关系。
我们通过免疫组织化学染色评估人胶质母细胞瘤和正常脑组织中Notch1的表达。通过3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）试验评估NGF对胶质母细胞瘤细胞系（U87-MG）的影响。为了评估NGF与Notch1信号传导之间的关系，分别通过逆转录聚合酶链反应（RT-PCR）和蛋白质免疫印迹分析评估Notch1和Hes1的表达。为了证实NGF对Notch1信号传导的影响，使用了Notch1和Hes1小干扰RNA（siRNA）。
免疫组织化学显示，胶质母细胞瘤中Notch1的表达高于正常脑组织。MTT试验表明，NGF以剂量依赖性方式刺激U87-MG细胞。RT-PCR和蛋白质免疫印迹分析表明，NGF以剂量依赖性方式增加Notch1和Hes1的表达。用Notch1和Hes1 siRNA转染后，对照组与100 nM NGF-β之间无显著差异，这意味着Notch1和Hes1 siRNA抑制了U87-MG细胞增殖。
这些结果表明，NGF通过Hes 1经由Notch1信号传导刺激胶质母细胞瘤细胞增殖。
