Department of Urology, San Francisco Veterans Affairs Medical Center, San Francisco, CA, USA.
Int J Cancer. 2011 Apr 15;128(8):1793-803. doi: 10.1002/ijc.25507.
The functional significance of Wnt antagonist DKK1 has not been investigated in renal cell carcinoma (RCC). Therefore, we hypothesized that DKK1 may be a tumor suppressor gene and is epigenetically silenced, thus decreased DKK1 may cause progression of RCC. To assess the function of DKK1, we established stable DKK1 transfected cells and monitored them regarding cell viability, colony formation, apoptosis, cell cycle, and invasive capability. RCC cell lines had decreased levels of DKK1, which were increased after treatment with 5-Aza-2'-deoxycytidine and trichostatin A. In chromatin immunoprecipitation assay, the level of dimethyl H3K9 and trimethyl H3K27 was decreased after 5-Aza-2'-deoxycytidine/trichostatin A treatment in RCC cell lines. Increased methylation was also associated with higher pathological stages in primary RCC tissues. T-cell factor/lymphoid enhancer factor activity and nuclear beta-catenin expression were not changed in DKK1 transfectants. Also the expression of cyclinD1 and c-Myc was not changed in DKK1 transfectants. These results suggest that DKK1 may not be involved in the beta-catenin dependent pathway. We also evaluated the expression of various related genes. Cleaved caspase3, p53, p21 and puma expression were significantly upregulated in the DKK1 transfected cells. The population of apoptotic cells was increased in stable DKK1 cells and tumor growth suppression was also observed in nude mice with DKK1 transfected cells. In conclusion, this is the first report to show that DKK1 expression is epigenetically silenced in kidney cancer and its reexpression induces apoptosis and cell cycle arrest in RCC.
DKK1 在肾细胞癌 (RCC) 中的功能意义尚未得到研究。因此，我们假设 DKK1 可能是一种肿瘤抑制基因，并被表观遗传沉默，因此 DKK1 的减少可能导致 RCC 的进展。为了评估 DKK1 的功能，我们建立了稳定转染 DKK1 的细胞，并监测它们的细胞活力、集落形成、细胞凋亡、细胞周期和侵袭能力。RCC 细胞系中 DKK1 的水平降低，经 5-Aza-2'-脱氧胞苷和曲古抑菌素 A 处理后增加。在染色质免疫沉淀测定中，在 RCC 细胞系中用 5-Aza-2'-脱氧胞苷/曲古抑菌素 A 处理后，二甲基 H3K9 和三甲基 H3K27 的水平降低。甲基化增加也与原发性 RCC 组织中的较高病理分期相关。在 DKK1 转染子中，T 细胞因子/淋巴增强因子活性和核 β-连环蛋白表达没有改变。在 DKK1 转染子中，cyclinD1 和 c-Myc 的表达也没有改变。这些结果表明 DKK1 可能不参与β-连环蛋白依赖途径。我们还评估了各种相关基因的表达。在 DKK1 转染细胞中，cleaved caspase3、p53、p21 和 puma 的表达显著上调。在稳定转染 DKK1 的细胞中，凋亡细胞的比例增加，并且在裸鼠中用 DKK1 转染的细胞也观察到肿瘤生长抑制。总之，这是第一项表明 DKK1 表达在肾癌中被表观遗传沉默，其重新表达诱导 RCC 细胞凋亡和细胞周期停滞的报告。
