Department of Neurosurgery, The First Affiliated Hospital of Xinxiang Medical University, Xinxiang, Henan, China.
Mol Cell Biochem. 2013 May;377(1-2):87-96. doi: 10.1007/s11010-013-1573-2. Epub 2013 Jan 29.
Tumor necrosis factor receptor-associated factor 6 (TRAF6), which plays an important role in inflammation and immune response, is an essential adaptor protein for the NF-κB (nuclear factor κB) signaling pathway. Recent studies have shown that TRAF6 played an important role in tumorigenesis and invasion by suppressing NF-κB activation. However, up to now, the biologic role of TRAF6 in glioma has still remained unknown. To address the expression of TRAF6 in glioma cells, four glioma cell lines (U251, U-87MG, LN-18, and U373) and a non-cancerous human glial cell line SVG p12 were used to explore the protein expression of TRAF6 by Western blot. Our results indicated that TRAF6 expression was upregulated in human glioma cell lines, especially in metastatic cell lines. To investigate the role of TRAF6 in cell proliferation, apoptosis, invasion, and migration of glioma, we generated human glioma U-87MG cell lines in which TRAF6 was either overexpressed or depleted. Subsequently, the effects of TRAF6 on cell viability, cell cycle distribution, apoptosis, invasion, and migration in U-87MG cells were determined with 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl tetrazolium bromide (MTT) assay, flow cytometry analysis, transwell invasion assay, and wound-healing assay. The results showed that knockdown of TRAF6 could decrease cell viability, suppress cell proliferation, invasion and migration, and promote cell apoptosis, whereas overexpression of TRAF6 displayed the opposite effects. In addition, the effects of TRAF6 on the expression of phosphor-NF-κB (p-p65), cyclin D1, caspase 3, and MMP-9 were also probed. Knockdown of TRAF6 could lower the expression of p-p65, cyclin D1, and MMP-9, and raise the expression of caspase 3. All these results suggested that TRAF6 might be involved in the potentiation of growth, proliferation, invasion, and migration of U-87MG cell, as well as inhibition of apoptosis of U-87MG cell by abrogating activation of NF-κB.
肿瘤坏死因子受体相关因子 6（TRAF6）在炎症和免疫反应中发挥重要作用，是 NF-κB（核因子 κB）信号通路的重要衔接蛋白。最近的研究表明，TRAF6 通过抑制 NF-κB 的激活在肿瘤发生和侵袭中发挥重要作用。然而，到目前为止，TRAF6 在神经胶质瘤中的生物学作用仍不清楚。为了研究 TRAF6 在神经胶质瘤细胞中的表达，我们使用了四种神经胶质瘤细胞系（U251、U-87MG、LN-18 和 U373）和一种非癌性人神经胶质细胞系 SVG p12 来通过 Western blot 探索 TRAF6 的蛋白表达。我们的结果表明，TRAF6 在人神经胶质瘤细胞系中表达上调，尤其是在转移性细胞系中。为了研究 TRAF6 在神经胶质瘤细胞增殖、凋亡、侵袭和迁移中的作用，我们构建了 TRAF6 过表达或缺失的人神经胶质瘤 U-87MG 细胞系。随后，通过 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）测定法、流式细胞术分析、Transwell 侵袭测定法和划痕愈合测定法来确定 TRAF6 对 U-87MG 细胞活力、细胞周期分布、凋亡、侵袭和迁移的影响。结果表明，TRAF6 敲低可降低细胞活力，抑制细胞增殖、侵袭和迁移，并促进细胞凋亡，而 TRAF6 过表达则呈现相反的效果。此外，还研究了 TRAF6 对磷酸化 NF-κB（p-p65）、细胞周期蛋白 D1、半胱天冬酶 3 和基质金属蛋白酶 9（MMP-9）表达的影响。TRAF6 敲低可降低 p-p65、细胞周期蛋白 D1 和 MMP-9 的表达，提高半胱天冬酶 3 的表达。所有这些结果表明，TRAF6 可能参与了 U-87MG 细胞生长、增殖、侵袭和迁移的增强，以及通过 NF-κB 激活的抑制促进了 U-87MG 细胞的凋亡。
