Li Yang, Wei Zhenqing, Dong Bin, Lian Zhigang, Xu Yinghui
Department of Neurosurgery, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning 116011, P.R. China.
Int J Mol Med. 2016 Apr;37(4):998-1004. doi: 10.3892/ijmm.2016.2500. Epub 2016 Feb 23.
Phosphoglucose isomerase/autocrine motility factor (PGI/AMF) is secreted by tumors and influences tumor growth and metastasis. In order to investigate the effects of silencing PGI/AMF on the migration and the sphere forming abilities of human glioblastoma U87 cells, as well as on the side population cells (SPCs), PGI/AMF was silenced using siRNA. Western blot analysis and RT-qPCR were used to assess the expression of PGI/AMF, Akt and SRY (sex determining region Y)-box 2 (SOX2). Wound healing, migration and tumorsphere formation assays were performed to assess invasion and metastatic potential. The proportion of SPCs was determined using Hoechst 33342 dye and flow cytometric analysis. PGI/AMF silencing inhibited the wound healing capacity and migration ability of U87 cells by 52.6 and 80.4%, respectively, compared with the scrambled siRNA (both P<0.001). Silencing of PGI/AMF decreased the proportion of SPCs in the U87 cells by 80.9% (P<0.01). The silencing of PGI/AMF decreased the number and size of tumorspheres by 53.1 and 39.9%, respectively, compared with the scrambled siRNA (both P<0.01). The silencing of PGI/AMF decreased the levels of phosphorylated Akt (-71.9%, P<0.001) compared with the scrambled siRNA, as well as the levels of the stemness marker, SOX2 (-61.7%, P<0.01). Taken together, these findings suggest that PGI/AMF silencing decreases migration, tumorsphere formation as well as the proportion of SPCs in glioblastoma U87 cells. We suggest that the Akt pathway is involved, and our results provide a potential new target for the treatment of glioblastoma.
磷酸葡萄糖异构酶/自分泌运动因子(PGI/AMF)由肿瘤分泌,并影响肿瘤的生长和转移。为了研究沉默PGI/AMF对人胶质母细胞瘤U87细胞迁移和球形成能力以及对侧群细胞(SPCs)的影响,使用小干扰RNA(siRNA)沉默PGI/AMF。采用蛋白质免疫印迹分析和逆转录定量聚合酶链反应(RT-qPCR)评估PGI/AMF、蛋白激酶B(Akt)和Y染色体性别决定区高迁移率族蛋白盒2(SRY-box2,SOX2)的表达。进行伤口愈合、迁移和肿瘤球形成试验以评估侵袭和转移潜能。使用Hoechst 33342染料和流式细胞术分析确定SPCs的比例。与乱序siRNA相比,沉默PGI/AMF分别使U87细胞的伤口愈合能力和迁移能力降低了52.6%和80.4%(均P<0.001)。沉默PGI/AMF使U87细胞中SPCs的比例降低了80.9%(P<0.01)。与乱序siRNA相比,沉默PGI/AMF使肿瘤球的数量和大小分别减少了53.1%和39.9%(均P<0.01)。与乱序siRNA相比,沉默PGI/AMF使磷酸化Akt水平降低了71.9%(P<0.001),同时使干性标志物SOX2水平降低了61.7%(P<0.01)。综上所述,这些发现表明沉默PGI/AMF可降低胶质母细胞瘤U87细胞的迁移、肿瘤球形成以及SPCs的比例。我们认为Akt信号通路参与其中,我们的结果为胶质母细胞瘤的治疗提供了一个潜在的新靶点。