Department of Hematology, Oslo University Hospital, Oslo, Norway.
Department of Hematology, the First Affiliated Hospital of Zhengzhou University, Zhengzhou, China.
J Thromb Haemost. 2016 Feb;14(2):387-96. doi: 10.1111/jth.13206. Epub 2016 Jan 30.
ESSENTIALS: A hypoxic microenvironment is a common feature of tumors that may influence activation of coagulation. MCF-7 and SK-BR-3 breast cancer cells and breast cancer tissue samples were used. The results showed transcriptional repression of tissue factor pathway inhibitor expression in hypoxia. Hypoxia-inducible factor 1α may be a target for the therapy of cancer-related coagulation and thrombosis.
Activation of coagulation is a common finding in patients with cancer, and is associated with an increased risk of venous thrombosis. As a hypoxic microenvironment is a common feature of solid tumors, we investigated the role of hypoxia in the regulation of tissue factor (TF) pathway inhibitor (TFPI) expression in breast cancer.
To explore the transcriptional regulation of TFPI by hypoxia-inducible factor (HIF)-1α in breast cancer cells and their correlation in breast cancer tissues.
MCF-7 and SK-BR-3 breast cancer cells were cultured in 1% oxygen or treated with cobalt chloride (CoCl2 ) to mimic hypoxia. Time-dependent and dose-dependent downregulation of TFPI mRNA (quantitative RT-PCR) and of free TFPI protein (ELISA) were observed in hypoxia. Western blotting showed parallel increases in the levels of HIF-1α protein and TF. HIF-1α inhibitor abolished or attenuated the hypoxia-induced downregulation of TFPI. Luciferase reporter assay showed that both hypoxia and HIF-1α overexpression caused strong repression of TFPI promoter activity. Subsequent chromatin immunoprecipitation and mutagenesis analysis demonstrated a functional hypoxia response element within the TFPI promoter, located at -1065 to -1060 relative to the transcriptional start point. In breast cancer tissue samples, gene expression analyses showed a positive correlation between the mRNA expression of TFPI and that of HIF-1α.
This study demonstrates that HIF-1α is involved in the transcriptional regulation of the TFPI gene, and suggests that a hypoxic microenvironment inside a breast tumor may induce a procoagulant state in breast cancer patients.
要点：缺氧微环境是肿瘤的常见特征，可能影响凝血的激活。使用 MCF-7 和 SK-BR-3 乳腺癌细胞和乳腺癌组织样本。结果表明，缺氧时组织因子途径抑制剂表达的转录抑制。缺氧诱导因子 1α 可能是癌症相关凝血和血栓形成治疗的靶点。
凝血激活是癌症患者的常见现象，与静脉血栓形成的风险增加有关。由于缺氧微环境是实体瘤的常见特征，我们研究了缺氧对乳腺癌中组织因子（TF）途径抑制剂（TFPI）表达的调节作用。
探讨缺氧诱导因子（HIF）-1α在乳腺癌细胞中对 TFPI 的转录调节及其在乳腺癌组织中的相关性。
将 MCF-7 和 SK-BR-3 乳腺癌细胞在 1%氧气或用钴氯化物（CoCl2）处理以模拟缺氧。在缺氧条件下，TFPI mRNA（定量 RT-PCR）和游离 TFPI 蛋白（ELISA）的时间依赖性和剂量依赖性下调。Western 印迹显示 HIF-1α 蛋白和 TF 的水平平行增加。HIF-1α 抑制剂消除或减弱了缺氧诱导的 TFPI 下调。荧光素酶报告基因检测显示，缺氧和 HIF-1α 过表达均可强烈抑制 TFPI 启动子活性。随后的染色质免疫沉淀和突变分析表明，TFPI 启动子内存在功能缺氧反应元件，位于转录起始点的-1065 到-1060 位。在乳腺癌组织样本中，基因表达分析显示 TFPI 和 HIF-1α 的 mRNA 表达之间存在正相关。
本研究表明 HIF-1α参与 TFPI 基因的转录调节，并提示乳腺癌肿瘤内的缺氧微环境可能在乳腺癌患者中诱导促凝状态。
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