Department of Gynecology, Cancer Hospital of China Medical University, Liaoning Cancer Hospital and Institute, Shenyang, Liaoning 110042, P.R. China.
Int J Mol Med. 2018 Mar;41(3):1349-1356. doi: 10.3892/ijmm.2017.3342. Epub 2017 Dec 22.
B-Raf proto-oncogene, serine/threonine kinase (BRAF) has previously been identified as a candidate target gene in endometriosis. Wild-type and mutated BRAF serve important roles in different diseases. The aim of the present study was to explore BRAF mutation, the mRNA and protein expression of wild-type BRAF (wtBRAF) in endometriosis, and the association between the expression levels of wtBRAF and the predicted transcription factor cAMP responsive element binding protein 1 (CREB1). In the present study, BRAF mutation was detected using Sanger sequencing among 30 ectopic and matched eutopic endometrium samples of patients with endometriosis as well as 25 normal endometrium samples, and no BRAF mutation was detected in exons 11 or 15. A region of ~2,000 bp upstream of the BRAF gene was then screened using NCBI and UCSC databases, and CREB1 was identified as a potential transcription factor of BRAF by analysis with the JASPAR and the TRANSFAC databases. Quantitative polymerase chain reaction was used to analysis the mRNA expression levels of wtBRAF and CREB1, and the corresponding protein expression levels were evaluated using immunohistochemistry and western blot analysis. The results revealed that the mRNA and protein expression levels of wtBRAF and CREB1 were significantly upregulated in the eutopic endometrial tissues of patients with endometriosis compared with normal endometrial tissues (P<0.05) and no significant difference in wtBRAF and CREB1 levels was detected between the ectopic and eutopic endometrium (P>0.05). In addition, correlation analysis revealed that the protein expression of CREB1 was positively correlated with the transcript level and protein expression of wtBRAF. It is reasonable to speculate that CREB1 may activate the transcription of wtBRAF through directly binding to its promoter, increasing BRAF expression and regulating the cell proliferation, migration and invasion of endometriosis.
B-Raf 原癌基因丝氨酸/苏氨酸激酶（BRAF）先前已被确定为子宫内膜异位症的候选靶基因。野生型和突变型 BRAF 在不同疾病中发挥重要作用。本研究旨在探讨子宫内膜异位症中 BRAF 突变、野生型 BRAF（wtBRAF）的 mRNA 和蛋白表达，以及 wtBRAF 表达水平与预测转录因子 cAMP 反应元件结合蛋白 1（CREB1）之间的关系。在本研究中，通过对 30 例子宫内膜异位症患者的异位和配对在位内膜样本以及 25 例正常内膜样本进行 Sanger 测序，检测到 BRAF 突变，在外显子 11 或 15 中未检测到 BRAF 突变。然后使用 NCBI 和 UCSC 数据库筛选 BRAF 基因上游约 2000bp 的区域，通过 JASPAR 和 TRANSFAC 数据库分析鉴定出 CREB1 是 BRAF 的潜在转录因子。采用定量聚合酶链反应分析 wtBRAF 和 CREB1 的 mRNA 表达水平，采用免疫组织化学和 Western blot 分析评估相应的蛋白表达水平。结果显示，与正常子宫内膜组织相比，子宫内膜异位症患者的在位子宫内膜组织中 wtBRAF 和 CREB1 的 mRNA 和蛋白表达水平均显著上调（P<0.05），异位和在位子宫内膜组织中 wtBRAF 和 CREB1 水平无显著差异（P>0.05）。此外，相关性分析显示 CREB1 蛋白表达与 wtBRAF 的转录水平和蛋白表达呈正相关。有理由推测 CREB1 可能通过直接结合其启动子激活 wtBRAF 的转录，增加 BRAF 表达，调节子宫内膜异位症中细胞的增殖、迁移和侵袭。
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