siRNA-mediated downregulation of MMP-9 and uPAR in combination with radiation induces G2/M cell-cycle arrest in Medulloblastoma.
siRNA-mediated downregulation of MMP-9 and uPAR in combination with radiation induces G2/M cell-cycle arrest in Medulloblastoma.
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DOI：
10.1158/1541-7786.mcr-10-0399
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发表时间：
2011-01
期刊：
影响因子：
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通讯作者：
Rao JS
中科院分区：
文献类型：
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作者：
Nagaraju GP;Nalla AK;Gupta R;Mohanam S;Gujrati M;Dinh DH;Rao JS
我们之前的工作和其他研究人员的工作强烈表明 MMP-9 和 uPAR 的上调与肿瘤血管生成和转移之间存在关系。在本研究中，我们评估了 MMP-9 和 uPAR 在髓母细胞瘤癌细胞对电离辐射 (IR) 抵抗中的作用，并测试了 siRNA 单独或联合 (pUM) 针对 MMP-9 (pM) 和 uPAR (pU) 的抗肿瘤功效。进行细胞增殖（BrdU 测定）、细胞凋亡（用于 DNA 片段化的原位 TUNEL）和细胞周期 (FACS) 分析，以确定 siRNA 单独或与 IR 组合对髓母细胞瘤细胞中 G2/M 细胞周期停滞的影响。 IR 上调髓母细胞瘤细胞中 MMP-9 和 uPAR 的表达； pM、pU 和 pUM 与 IR 联合有效降低 MMP-9 和 uPAR 表达，从而导致髓母细胞瘤细胞的放射敏感性增加。 siRNA 治疗（pM、pU 和 pUM）还促进 IR 诱导的细胞凋亡，并增强细胞周期进展过程中 IR 诱导的 G2/M 期停滞。 IR 通过抑制 pCdc2 和细胞周期蛋白 B 调节的信号通路（涉及 p53、p21/WAF1 和 Chk2 基因表达）诱导 G2/M 细胞周期停滞，而 siRNA（pM、pU 和 pUM）单独或与 IR 组合通过抑制 pCdc2 和细胞周期蛋白 B1 调节的信号通路（涉及 Chk1 和 Cdc25A 基因表达）介导 G2/M 细胞周期停滞。综上所述，我们的数据表明，MMP-9 和 uPAR 的下调会诱导 Chk1 介导的 G2/M 细胞周期停滞，而单独 IR 引起的破坏则依赖于 p53 和 Chk2 介导的 G2/M 细胞周期停滞。
Our previous work and that of other investigators strongly suggest a relationship between the upregulation of MMP-9 and uPAR and tumor angiogenesis and metastasis. In this study, we evaluated the role of MMP-9 and uPAR in medulloblastoma cancer cell resistance to ionizing irradiation (IR) and tested the anti-tumor efficacy of siRNA against MMP-9 (pM) and uPAR (pU) either alone or in combination (pUM). Cell proliferation (BrdU assay), apoptosis (in situ TUNEL for DNA fragmentation), and cell cycle (FACS) analyses were carried out to determine the effect of siRNA either alone or in combination with IR on G2/M cell cycle arrest in medulloblastoma cells. IR upregulated MMP-9 and uPAR expression in medulloblastoma cells; pM, pU, and pUM in combination with IR effectively reduced both MMP-9 and uPAR expression, thereby leading to increased radiosensitivity of medulloblastoma cells. siRNA treatments (pM, pU, and pUM) also promoted IR-induced apoptosis and enhanced IR-induced G2/M arrest during cell cycle progression. While IR induces G2/M cell cycle arrest through inhibition of the pCdc2 and cyclin B-regulated signaling pathways involving p53, p21/WAF1, and Chk2 gene expression, siRNA (pM, pU, and pUM) alone or in combination with IR induced G2/M arrest mediated through inhibition of the pCdc2 and cyclin B1-regulated signaling pathways involving Chk1 and Cdc25A gene expression. Taken together, our data suggest that downregulation of MMP-9, and uPAR induces Chk1-mediated G2/M cell cycle arrest, whereas the disruption caused by IR alone is dependent on p53- and Chk2-mediated G2/M cell cycle arrest.