HOXB7在电离辐射诱导的肺腺癌上皮细胞间质转化及转移中的作用和机制研究

Radiotherapy is a traditional treatment for cancer，but recent research have proved that ionizing irradiation can induce migration of esophageal cancer and other cancer cells，but the effect and mechanism of ionizing irradiation on lung adenocarcinoma cells are not clear. We found that HOXB7 is an important cancer-promoting molecule, and its expression was related to lymph nodes and distant metastasis of lung adenocarcinoma cells. We confirmed that the epithelial-mesenchymal transition and migration of lung adenocarcinoma can be induced by low dose ionizing irradiation，and this process was conducted by up-regulation of HOXB7 which was induced by γ–ray. Bioinformatics analysis revealed the expression of PIK3R1 was correlated with HOXB7, and PIK3R1 was a key gene for activation of PI3K/AKT signaling pathway. Moreover, the promoter region of PIK3R1 gene contain several binding sequence of HOXB7. Based on these studies, we propose to determine the effects of HOXB7 expression induced by ionizing irradiation, and explore the mechanisms of PIK3R1 expression regulated by HOXB7.Then we will access the role of HOXB7/PIK3R1/PI3K/AKT/Snail axis in ionizing irradiation-induced epithelial-mesenchymal transition and migration of lung adenocarcinoma cells. The results of this project can provide new targets for reversing metastasis of lung adenocarcinoma after radiotherapy.

放疗是传统的肿瘤治疗方法，但最新研究证实电离辐射可诱导食管癌等肿瘤发生转移，然而其对肺腺癌细胞转移的影响和机制尚不清楚。我们前期首次发现HOXB7是重要的促癌分子，与肺腺癌细胞淋巴结及远处转移密切相关，且HOXB7是细胞DNA损伤修复相关基因。本课题组预实验提示低剂量γ-射线可诱导肺腺癌细胞发生上皮间质转化并转移，而该效应与γ-射线上调HOXB7表达有关。WB证实HOXB7表达与PI3K/AKT信号通路关键激活因子PIK3R1表达呈正相关，生物信息学分析发现PIK3R1基因启动子区含HOXB7的结合序列。因此，本项目将进一步探明电离辐射对肺腺癌细胞HOXB7表达的影响，并深入探索HOXB7对PIK3R1表达的调控机制，进而明确HOXB7/PIK3R1/PI3K/AKT/Snail分子调控轴在电离辐射诱导的肺腺癌细胞上皮间质转化及转移中的调控作用。研究结果可为逆转肺腺癌放疗后转移提供新的思路

探索电离辐射对肺腺癌细胞中HOXB7基因表达的影响，以及HOXB7促进电离辐射诱导肺腺癌细胞上皮间质转化和迁移的分子机制。研究内容：1、检测电离辐射对肺腺癌中蛋白表达的影响2、在肺腺癌细胞下调HOXB7，以 X-射线辐照后第三天检测蛋白表达；3、在细胞下调PIK3R1，以 X-射线辐照后第三天后检测蛋白表达；4、分别用PI3K抑制剂或AKT抑制剂处理肺腺癌细胞，以 X-射线辐照第三天后检测蛋白表达情况；5、检测以上各组细胞的迁移能力的变化情况。结果：1、不同剂量的X-射线辐照，低剂量电离辐射可上调HOXB7、PIK3R1、Snail蛋白的表达，2Gy剂量的辐照时蛋白表达水平最高；用2Gy X-射线辐照发现辐照后第三天蛋白表达水平最高；电离辐射后肺腺癌细胞的迁移能力增强；降低HOXB7表达后，蛋白表达水平显著下调，上辐照后第三天HOXB7、PIK3R1、Snail蛋白表达水平最高；电离辐射后肺腺癌细胞的迁移能力增强；2.细胞中降低HOXB7表达水平后，PIK3R1、p-AKT、Snail表达下调，上皮表型标志分子表达上调，间质表型标志分子表达下调；肺腺癌细胞迁移能力降低。辐照后各蛋白的表达水平及细胞迁移能力与辐照前无明显变化。3、细胞中下调PIK3R1表达，肺腺癌HOXB7蛋白表达水平无明显变化，而p-AKT、Snail、Vimentin表达下调，E-cadherin表达上调；下调PIK3R1表达水平后，肺腺癌细胞迁移能力降低。辐照后HOXB7蛋白的表达水平上调，其余蛋白及细胞迁移能力较辐照前无明显变化。用PI3K抑制剂或AKT抑制剂处理肺腺癌A549细胞，利用Western blotting检测处理前后的肺腺癌A549细胞中各蛋白的表达，发现：抑制PI3K/AKT信号通路活性后，HOXB7及PIK3R1蛋白表达无明显变化，而Snail、E-cadherin表达上调，间质表型标志分子表达下调，细胞迁移能力降低；射线辐照抑制PI3K/AKT信号通路前后的肺腺癌细胞，检测各蛋白的表达，辐照后HOXB7及PIK3R1蛋白的表达水平提高，余蛋白的表达及细胞的迁移能力较辐照前无明显差异。结论：低剂量的电离辐射可上调肺腺癌中HOXB7的表达，进而活化PI3K/PI信号通路，最终促进肺腺癌A549细胞上皮间质转化和迁移。