Department of Pathology, First Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510080, People's Republic of China.
Department of Pathology, First Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong 510080, People's Republic of China.
Biochem Biophys Res Commun. 2014 Oct 3;452(4):933-9. doi: 10.1016/j.bbrc.2014.09.009. Epub 2014 Sep 6.
Osteosarcoma is the most common primary malignant bone tumour in children and adolescents and is characterised by high malignant and metastatic potentials. However, the molecular mechanism underlying this invasiveness remains unclear. In this study, we determined that PD98059 and SP600125, the two mitogen-activated protein kinase (MAPK) family inhibitors, decreased the osteosarcoma cell U2OS-AEG-1 migration and invasion that was enhanced by astrocyte elevated gene-1 (AEG-1) in an in vitro wound-healing and Matrigel invasion assay independently of cell viability. These findings indicate that AEG-1 promoted osteosarcoma cell invasion is relevant to the MAPK pathways. The up-regulation of AEG-1 increased the levels of phosphor-c-Jun N-terminal kinase (JNK) and phosphor-c-Jun; however, there were no marked changes in the levels of phosphor-extracellular regulated kinase (ERK) 1/2 or phosphor-c-Fos due to the activation of AEG-1 in U2OS. SP600125 (a JNK inhibitor) decreased phosphor-c-Jun and MMP-2 in U2OS-AEG-1, while PD98059 (a ERK1/2 inhibitor) had no influence on the levels of phosphor-c-Jun or MMP-2 in U2OS-AEG-1. Further study revealed that the down-regulation of phosphor-c-Jun not only obviously decreased the MMP-2 protein level and the MMP-2 transcriptional activity that were up-regulated by AEG-1 in Western-blot and luciferase reporter assays, but also inhibited the migration and invasion abilities of the U2OS-AEG-1 cells, which suggests that AEG-1 mediated U2OS invasion at least partially via the JNK/c-Jun/MMP-2 pathway. Consistent with these observations, immunohistochemical (IHC) staining revealed that AEG-1 expression was associated with the protein levels of phosphor-c-Jun and MMP-2 in needle biopsy paraffin-embedded archival human osteosarcoma tissues. Taken together, our findings suggest that AEG-1 plays a crucial role in the aggressiveness of osteosarcoma via the JNK/c-Jun/MMP-2 pathway.
骨肉瘤是儿童和青少年中最常见的原发性恶性骨肿瘤，其特点是恶性程度和转移潜能高。然而，其侵袭性的分子机制尚不清楚。在这项研究中，我们发现，两种丝裂原活化蛋白激酶（MAPK）家族抑制剂 PD98059 和 SP600125，可降低星形细胞瘤上调基因-1（AEG-1）增强的骨肉瘤细胞 U2OS-AEG-1 迁移和侵袭能力，这在体外划痕和基质胶侵袭实验中是独立于细胞活力的。这些发现表明，AEG-1 促进骨肉瘤细胞侵袭与 MAPK 途径有关。AEG-1 的上调增加了磷酸化 c-Jun N-末端激酶（JNK）和磷酸化 c-Jun 的水平；然而，由于 AEG-1 在 U2OS 中的激活，ERK1/2 或磷酸化 c-Fos 的水平没有明显变化。SP600125（JNK 抑制剂）降低了 U2OS-AEG-1 中的磷酸化 c-Jun 和 MMP-2，而 PD98059（ERK1/2 抑制剂）对 U2OS-AEG-1 中磷酸化 c-Jun 或 MMP-2 的水平没有影响。进一步的研究表明，磷酸化 c-Jun 的下调不仅明显降低了 Western-blot 和荧光素酶报告基因分析中 AEG-1 上调的 MMP-2 蛋白水平和 MMP-2 转录活性，而且抑制了 U2OS-AEG-1 细胞的迁移和侵袭能力，这表明 AEG-1 通过 JNK/c-Jun/MMP-2 途径介导 U2OS 的侵袭。与这些观察结果一致，免疫组织化学（IHC）染色显示，AEG-1 的表达与骨肉瘤组织石蜡包埋活检存档组织中磷酸化 c-Jun 和 MMP-2 的蛋白水平相关。总之，我们的研究结果表明，AEG-1 通过 JNK/c-Jun/MMP-2 途径在骨肉瘤的侵袭性中发挥关键作用。
