Department of Neurosurgery, Beijing Tiantan Hospital, Capital Medical University, Beijing 100050, P.R. China.
Core Laboratory for Clinical Medical Research, Beijing Tiantan Hospital, Capital Medical University, Beijing 100050, P.R. China.
Oncol Rep. 2018 Mar;39(3):1441-1453. doi: 10.3892/or.2018.6208. Epub 2018 Jan 10.
The blood‑brain barrier (BBB) constitutes an efficient organization of tight junctions that limits the delivery of tumor to the brain. The principal tight junction protein in BBB is claudin‑5 (CLDN5), but its mechanism of action remains largely unknown. Long non‑coding RNAs (lncRNAs) are aberrantly expressed in many cancers, some lncRNAs play key roles in regulating BBB permeability and are involved in tumor brain metastasis. In particular, lncRNAs can function as competing endogenous RNAs (ceRNAs). Herein, we investigated whether ceRNA dysregulation is associated with alterations of the level of CLDN5 in human brain vascular endothelial hCMEC/D3 cells. The Affymetrix Human Transcriptome Array 2.0 and Affymetrix GeneChip miRNA 4.0 Array were used to detect the expression levels of 2,578 miRNAs, 22,829 lncRNAs, and 44,699 mRNAs in pLL3.7‑CLDN5‑transfected and pLL3.7 control hCMEC/D3 cells. The distinctly expressed miRNAs, lncRNAs, and mRNAs were subjected to construction of miRNA‑lncRNA‑mRNA interaction network. A total of 41 miRNAs, 954 lncRNAs, and 222 mRNAs were found to be differentially expressed between the CLDN5‑overexpressing and control group. 148 lncRNA acting as ceRNAs were identified based on the miRNA‑lncRNA‑mRNA interaction network. The function of differential mRNA in the network was determined by GO and pathway analysis. The potential roles of the 27 ceRNAs were revealed, the possible biology functions of these regulatory ceRNAs mainly included tight junction, focal adhesion, cell‑cell adhesion, cell growth and apoptosis. The identified sets of miRNAs, lncRNAs and mRNAs specific to CLDN5‑overexpressing hCMEC/D3 cells were verified by quantitative real‑time RT‑PCR experiment. Our study predicts the biological functions of a multitude of ceRNAs associated with the alteration of CLDN5 in brain vascular endothelial cells. Our data suggest that these dysregulated ceRNAs, in conjunction with the high CLDN5 levels, could serve as useful targets of prevention of brain metastasis formation. Further studies are warranted to determine the role of these ceRNAs in facilitating the function of CLDN5 in brain‑tumor barrier.
血脑屏障（BBB）构成了紧密连接的有效组织，限制了肿瘤向大脑的输送。BBB 的主要紧密连接蛋白是 Claudin-5（CLDN5），但其作用机制在很大程度上仍不清楚。长链非编码 RNA（lncRNA）在许多癌症中表达异常，一些 lncRNA 在调节 BBB 通透性方面发挥着关键作用，并参与肿瘤脑转移。特别是，lncRNA 可以作为竞争性内源 RNA（ceRNA）发挥作用。在此，我们研究了 ceRNA 失调是否与人类脑血管内皮 hCMEC/D3 细胞中 CLDN5 水平的改变有关。使用 Affymetrix Human Transcriptome Array 2.0 和 Affymetrix GeneChip miRNA 4.0 Array 检测了 pLL3.7-CLDN5 转染和 pLL3.7 对照 hCMEC/D3 细胞中 2578 个 miRNA、22829 个 lncRNA 和 44699 个 mRNA 的表达水平。对明显表达的 miRNA、lncRNA 和 mRNA 进行了 miRNA-lncRNA-mRNA 相互作用网络的构建。在 CLDN5 过表达组和对照组之间共发现 41 个 miRNA、954 个 lncRNA 和 222 个 mRNA 差异表达。基于 miRNA-lncRNA-mRNA 相互作用网络，共鉴定出 148 个作为 ceRNA 的 lncRNA。通过 GO 和通路分析确定网络中差异 mRNA 的功能。揭示了 27 个 ceRNA 的潜在作用，这些调节 ceRNA 的可能生物学功能主要包括紧密连接、焦点黏附、细胞-细胞黏附、细胞生长和凋亡。通过定量实时 RT-PCR 实验验证了 CLDN5 过表达 hCMEC/D3 细胞中特定的 miRNA、lncRNA 和 mRNA 集合。本研究预测了与脑血管内皮细胞中 CLDN5 改变相关的大量 ceRNA 的生物学功能。我们的数据表明，这些失调的 ceRNA 与高 CLDN5 水平一起，可能成为预防脑转移形成的有用靶点。进一步的研究需要确定这些 ceRNA 在促进 CLDN5 在血脑屏障中的功能中的作用。
