Wei Dian-Fang, Tang Ming-Ke, Liu Yang, Zhang Chao-Yue, Qin Li-Juan
School of Basic Medical Sciences, North China University of Science and Technology, Tangshan 063000, China.
Hebei Key Laboratory for Chronic Diseases, Tangshan 063000, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2019 Mar;50(2):188-192.
To study the relationship between hypoxia and the hypoxia inducible factor-1α (HIF-1α) from lung cancer cells, to reveal the possible mechanism of brain metastases of lung cancer.
The hypoxia model of A549 lung cancer cells was established. After hypoxia culture of A549 cells for 0.5, 2, 4, 8, 12 and 24 h (normal oxygen culture at the same time point was set as the control group), the mass concentration of HIF-1α in A549 lung cancer cell culture medium were determined by ELISA. Transwell chamber was used to construct an blood brain barrier model, was treated with A549 lung cancer cell culture medium after different time points of hypoxia, Tran endothelial resistance (TER) change of blood-brain barrier model in instrument, to reflect the changes of blood-brain barrier permeability ; A549 lung cancer cells in the culture medium were counted under Transwell room. A549 lung cancer cells with hypoxia at different time points injected into Wistar rats via tail vein, Western blot method was used to menstruate expression of tight junction associated protein Claudin-5 in the brain tissues, Evans blue to detect the change of blood brain barrier permeability in rats.
Compared with the control group, the HIF-1α mass concentration in the cell culture solution of A549 increased, the blood-brain barrier model TER decreased, and the cell number of A549 that passed through transwell into the lower chamber increased (all <0.05) after hypoxia 2 h, the above effect was most obvious when hypoxia 8 h (all <0.01). After hypoxia 24 h, it was restored to the control group level. In the experiment of rats, compared with the control group, the mass percent of Evans blue in rat brain tissues increased after A549 cell culture solution with hypoxia 2 h was injected via caudal vein, meaning increased the permeability of rat blood brain barrier, while the expression of Claudin-5 protein in rat brain tissues decreased (all <0.05). The effect was most obvious when A549 cell culture solution with hypoxia 8 h was injected into rat tail vein (<0.01 ). Ejectionof hypoxia 24 h A549 cell culture solution yielded the same effects as those in the control group.
Hypoxia can induce the increase of HIF-1α in lung cancer cells. The increase of HIF-1α results in the decrease of Claudin-5 expression and increase of blood-brain barrier permeability, leading to lung cancer cells metastasis into the brain.
研究缺氧与肺癌细胞中缺氧诱导因子-1α（HIF-1α）的关系，揭示肺癌脑转移的可能机制。
建立A549肺癌细胞缺氧模型。将A549细胞进行缺氧培养0.5、2、4、8、12和24 h（同时设常氧培养的同一时间点为对照组），采用ELISA法测定A549肺癌细胞培养液中HIF-1α的质量浓度。用Transwell小室构建血脑屏障模型，在不同缺氧时间点后用A549肺癌细胞培养液处理，通过仪器检测血脑屏障模型的跨内皮电阻（TER）变化，以反映血脑屏障通透性的改变；在Transwell小室内计数培养液中的A549肺癌细胞。将不同时间点缺氧的A549肺癌细胞经尾静脉注入Wistar大鼠，采用Western blot法检测脑组织中紧密连接相关蛋白Claudin-5的表达，用伊文思蓝检测大鼠血脑屏障通透性的变化。
与对照组相比，缺氧2 h后A549细胞培养液中HIF-1α质量浓度升高，血脑屏障模型TER降低，穿过Transwell进入下室的A549细胞数增加（均P<0.05），缺氧8 h时上述效应最明显（均P<0.01）。缺氧24 h后恢复至对照组水平。在大鼠实验中，与对照组相比，经尾静脉注射缺氧2 h的A549细胞培养液后，大鼠脑组织中伊文思蓝质量百分比增加，即大鼠血脑屏障通透性增加，同时大鼠脑组织中Claudin-5蛋白表达降低（均P<0.05）。经尾静脉注射缺氧8 h的A549细胞培养液时效应最明显（P<0.01）。注射缺氧24 h的A549细胞培养液产生的效应与对照组相同。
缺氧可诱导肺癌细胞中HIF-1α升高。HIF-1α升高导致Claudin-5表达降低和血脑屏障通透性增加，从而致使肺癌细胞转移至脑内。
