Department of Neurosurgery, Tianjin Medical University General Hospital, Tianjin, 300052, China.
Laboratory of Neuro-Oncology, Tianjin Neurological Institute, Tianjin, 300052, China.
J Exp Clin Cancer Res. 2019 Aug 5;38(1):339. doi: 10.1186/s13046-019-1319-4.
Glioma initiating cells (GICs), also known as glioma stem cells (GSCs), play an important role in the progression and recurrence of glioblastoma multiforme (GBM) due to their potential for self-renewal, multiple differentiation and tumor initiation. In the recent years, Notch1 has been found to be overexpressed in GICs. However, the regulatory mechanism of Notch1 in the self-renewal and invasion ability of GICs remains unclear. This study aims to explore the effect of Notch pathway on self-renewal and invasion of GICs and the underlying mechanisms.
Bioinformatic analysis and immunohistochemistry (IHC) were performed to evaluate the expression of Notch1 and Hes1 in GBM samples. Immunofluorescent (IF) staining was performed to observe the distribution of Notch1 and CXCR4 in GBM and GICs. Both pharmacological intervention and RNA interference were employed to investigate the role of Notch1 in GICs self-renewal, invasion and tumor growth in vitro or in vivo. The crosstalk effect of Notch1 and CXCL12/CXCR4 system on GIC self-renewal and invasion was explored by sphere formation assay, limiting dilution assay and Transwell assay. Western blots were used to verify the activation of Notch1/CXCR4/AKT pathway in self-renewal, invasion and tumor growth of GICs. Luciferase reporter assay was used to testify the potential binding site of Notch1 signaling and CXCR4. The orthotopic GICs implantations were established to analyze the role and the mechanism of Notch1 in glioma progression in vivo.
Notch1 signaling activity was elevated in GBM tissues. Notch1 and CXCR4 were both upregulated in GICs, compared to Notch1 positive glioma cells comprised a large proportion in the CD133+ glioma cell spheres, CXCR4 positive glioma cells which usually expressed Notch1 both and dispersed in the periphery of the sphere, only represent a small subset of CD133+ glioma cell spheres. Furthermore, downregulation of the Notch1 pathway by shRNA and MK0752 significantly inhibited the PI3K/AKT/mTOR signaling pathway via the decreased expression of CXCR4 in GICs, and weakened the self-renewal, invasion and tumor growth ability of GICs.
These findings suggest that the cross-talk between Notch1 signaling and CXCL12/CXCR4 system could contribute to the self-renewal and invasion of GICs, and this discovery could help drive the design of more effective therapies in Notch1-targeted treatment of GBMs.
神经胶质瘤起始细胞（GICs），也称为神经胶质瘤干细胞（GSCs），由于其自我更新、多向分化和肿瘤起始的能力，在多形性胶质母细胞瘤（GBM）的进展和复发中发挥重要作用。近年来，发现 Notch1 在 GICs 中过表达。然而，Notch1 通路在 GICs 自我更新和侵袭能力中的调控机制尚不清楚。本研究旨在探讨 Notch 通路对 GICs 自我更新和侵袭的影响及其潜在机制。
通过生物信息学分析和免疫组织化学（IHC）评估 GBM 样本中 Notch1 和 Hes1 的表达。免疫荧光（IF）染色观察 GBM 和 GICs 中 Notch1 和 CXCR4 的分布。通过药理学干预和 RNA 干扰研究 Notch1 在体外或体内 GICs 自我更新、侵袭和肿瘤生长中的作用。通过球体形成实验、有限稀释实验和 Transwell 实验探讨 Notch1 和 CXCL12/CXCR4 系统对 GIC 自我更新和侵袭的串扰作用。Western blot 验证 Notch1/CXCR4/AKT 通路在 GIC 自我更新、侵袭和肿瘤生长中的激活情况。荧光素酶报告实验验证 Notch1 信号通路与 CXCR4 的潜在结合位点。建立原位 GIC 植入模型分析 Notch1 在体内胶质瘤进展中的作用及其机制。
Notch1 信号活性在 GBM 组织中升高。与 Notch1 阳性胶质瘤细胞相比，GICs 中 Notch1 和 CXCR4 均上调，Notch1 阳性胶质瘤细胞在 CD133+胶质瘤细胞球中占很大比例，而 CXCR4 阳性胶质瘤细胞通常表达 Notch1 并分散在球体的外围，仅代表一小部分 CD133+胶质瘤细胞球。此外，shRNA 和 MK0752 下调 Notch1 通路通过降低 GICs 中 CXCR4 的表达，显著抑制了 PI3K/AKT/mTOR 信号通路，减弱了 GICs 的自我更新、侵袭和肿瘤生长能力。
这些发现表明 Notch1 信号与 CXCL12/CXCR4 系统的串扰可能有助于 GICs 的自我更新和侵袭，这一发现有助于推动 Notch1 靶向治疗 GBMs 的更有效治疗方法的设计。
