Molecular Oncology Laboratories, The Weatherall Institute of Molecular Medicine, The University of Oxford, John Radcliffe Hospital, OX3 9DS Oxford, UK.
Br J Cancer. 2013 Jul 9;109(1):50-9. doi: 10.1038/bjc.2013.240. Epub 2013 Jun 18.
Non-muscle invasive (NMI) bladder cancer is characterised by increased expression and activating mutations of FGFR3. We have previously investigated the role of microRNAs in bladder cancer and have shown that FGFR3 is a target of miR-100. In this study, we investigated the effects of hypoxia on miR-100 and FGFR3 expression, and the link between miR-100 and FGFR3 in hypoxia.
Bladder cancer cell lines were exposed to normoxic or hypoxic conditions and examined for the expression of FGFR3 by quantitative PCR (qPCR) and western blotting, and miR-100 by qPCR. The effect of FGFR3 and miR-100 on cell viability in two-dimensional (2-D) and three-dimensional (3-D) was examined by transfecting siRNA or mimic-100, respectively.
In NMI bladder cancer cell lines, FGFR3 expression was induced by hypoxia in a transcriptional and HIF-1α-dependent manner. Increased FGFR3 was also in part dependent on miR-100 levels, which decreased in hypoxia. Knockdown of FGFR3 led to a decrease in phosphorylation of the downstream kinases mitogen-activated protein kinase (MAPK) and protein kinase B (PKB), which was more pronounced under hypoxic conditions. Furthermore, transfection of mimic-100 also decreased phosphorylation of MAPK and PKB. Finally, knocking down FGFR3 profoundly decreased 2-D and 3-D cell growth, whereas introduction of mimic-100 decreased 3-D growth of cells.
Hypoxia, in part via suppression of miR-100, induces FGFR3 expression in bladder cancer, both of which have an important role in maintaining cell viability under conditions of stress.
非肌肉浸润性（NMI）膀胱癌的特征是 FGFR3 表达增加和激活突变。我们之前研究了 microRNAs 在膀胱癌中的作用，并表明 FGFR3 是 miR-100 的靶标。在这项研究中，我们研究了缺氧对 miR-100 和 FGFR3 表达的影响，以及 miR-100 和 FGFR3 在缺氧中的联系。
将膀胱癌细胞系暴露于常氧或缺氧条件下，并通过定量 PCR（qPCR）和 Western blot 检测 FGFR3 的表达，通过 qPCR 检测 miR-100 的表达。通过转染 siRNA 或 mimic-100，分别检测 FGFR3 和 miR-100 对二维（2-D）和三维（3-D）细胞活力的影响。
在 NMI 膀胱癌细胞系中，FGFR3 表达在转录和 HIF-1α 依赖性方式下被缺氧诱导。增加的 FGFR3 部分也依赖于 miR-100 水平，miR-100 在缺氧条件下降低。FGFR3 的敲低导致下游激酶丝裂原激活蛋白激酶（MAPK）和蛋白激酶 B（PKB）的磷酸化减少，在缺氧条件下更为明显。此外，转染 mimic-100 也降低了 MAPK 和 PKB 的磷酸化。最后，敲低 FGFR3 可显著降低 2-D 和 3-D 细胞生长，而转染 mimic-100 可降低细胞 3-D 生长。
缺氧部分通过抑制 miR-100 诱导膀胱癌中 FGFR3 的表达，这两者在应激条件下维持细胞活力中都具有重要作用。
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