Department of Neurosurgery, Nanfang Hospital, Southern Medical University, Guangzhou, China.
Nanfang Neurology Research Institution, Nanfang Hospital, Southern Medical University, Guangzhou, China.
Theranostics. 2019 Mar 16;9(7):1909-1922. doi: 10.7150/thno.30578. eCollection 2019.
Glioblastoma (GBM) is the most common and aggressive brain tumor, characterized by its propensity to invade the surrounding brain parenchyma. The effect of extracellular high-mobility group box 1 (HMGB1) protein on glioblastoma (GBM) progression is still controversial. p62 is overexpressed in glioma cells, and has been associated with the malignant features and poor prognosis of GBM patients. Hence, this study aimed to clarify the role of p62 in HMGB1-induced epithelial-mesenchymal transition (EMT) of GBM both and . Immunoblotting, immunofluorescence and qRT-PCR were performed to evaluate EMT progression in both human GBM cell line and primary GBM cells. Transwell and wound healing assays were used to assess the invasion and migration of GBM cells. shRNA technique was used to investigate the role of p62 in HMGB1-induced EMT both and orthotopic tumor model. Co-immunoprecipitation assay was used to reveal the interaction between p62 and GSK-3β (glycogen synthase kinase 3 beta). Immunohistochemistry was performed to detect the expression levels of proteins in human GBM tissues. In this study, GBM cells treated with recombinant human HMGB1 (rhHMGB1) underwent spontaneous EMT through GSK-3β/Snail signaling pathway. In addition, our study revealed that rhHMGB1-induced EMT of GBM cells was accompanied by p62 overexpression, which was mediated by the activation of TLR4-p38-Nrf2 signaling pathway. Moreover, the results demonstrated that p62 knockdown impaired rhHMGB1-induced EMT both and . Subsequent mechanistic investigations showed that p62 served as a shuttling factor for the interaction of GSK-3β with proteasome, and ultimately activated GSK-3β/Snail signaling pathway by augmenting the degradation of GSK-3β. Furthermore, immunohistochemistry analysis revealed a significant inverse correlation between p62 and GSK-3β, and a combination of the both might serve as a more powerful predictor of poor survival in GBM patients. This study suggests that p62 is an effector for HMGB1-induced EMT, and may represent a novel therapeutic target in GBM.
胶质母细胞瘤（GBM）是最常见和侵袭性最强的脑肿瘤，其特征是倾向于侵犯周围的脑实质。细胞外高迁移率族蛋白 B1（HMGB1）蛋白对胶质母细胞瘤（GBM）进展的影响仍存在争议。p62 在神经胶质瘤细胞中过表达，并与 GBM 患者的恶性特征和不良预后相关。因此，本研究旨在阐明 p62 在 HMGB1 诱导的 GBM 上皮-间充质转化（EMT）中的作用。通过免疫印迹、免疫荧光和 qRT-PCR 评估了人 GBM 细胞系和原代 GBM 细胞的 EMT 进展。Transwell 和划痕愈合实验用于评估 GBM 细胞的侵袭和迁移。shRNA 技术用于研究 p62 在 HMGB1 诱导的 EMT 中的作用，包括体外和原位肿瘤模型。免疫共沉淀实验用于揭示 p62 与 GSK-3β（糖原合成酶激酶 3β）之间的相互作用。免疫组织化学用于检测人 GBM 组织中蛋白质的表达水平。
在这项研究中，用重组人 HMGB1（rhHMGB1）处理的 GBM 细胞通过 GSK-3β/Snail 信号通路自发发生 EMT。此外，我们的研究表明，rhHMGB1 诱导的 GBM 细胞 EMT 伴随着 p62 的过表达，这是通过 TLR4-p38-Nrf2 信号通路的激活介导的。此外，研究结果表明，p62 敲低可削弱 rhHMGB1 诱导的 EMT 作用，包括体外和原位肿瘤模型。随后的机制研究表明，p62 作为 GSK-3β 与蛋白酶体相互作用的穿梭因子，通过增强 GSK-3β 的降解，最终激活 GSK-3β/Snail 信号通路。此外，免疫组织化学分析显示 p62 与 GSK-3β 呈显著负相关，两者的结合可能成为 GBM 患者预后不良的更有力预测指标。
这项研究表明，p62 是 HMGB1 诱导的 EMT 的效应物，可能成为 GBM 的一个新的治疗靶点。
