| 徐慧英,刘晓冬,董永强,李文娜,李彩虹,刘兵,马淑梅.乏氧诱导因子1α在电离辐射诱导人乳腺癌细胞自噬性死亡中的作用[J].中华放射医学与防护杂志,2012,32(5):455-459 |
| 乏氧诱导因子1α在电离辐射诱导人乳腺癌细胞自噬性死亡中的作用 |
| Effects of hypoxia-inducible factor-1α on radiation-induced autophagic cell death in breast cancer cells |
| 投稿时间：2012-02-27 |
| DOI：10.3760/cma.j.issn.0254-5098.2012.05.002 |
| 中文关键词: 自噬 X射线 乏氧 乏氧诱导因子1α 辐射敏感性 |
| 英文关键词:Autophagy X-rays Hypoxia Hypoxia-inducible factor-1α Radiosensitivity |
| 基金项目:国家自然科学基金(30970682;30770649); 国家大学生创新项目(2010A72110) |
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| 中文摘要: |
| 目的 探讨乏氧诱导因子1α(HIF-1α)在辐射诱导人乳腺癌MCF-7细胞自噬性死亡中的作用。方法 将人乳腺癌MCF-7细胞分为对照组(常氧组,21%氧气)、照射组(8 Gy X射线)、乏氧组 (150 μmol/L CoCl2)、乏氧加照射组 (150 μmol/L CoCl2+8 Gy)。150 μmol/L CoCl2处理细胞模拟乏氧条件。Western blot法检测HIF-1α及MAPLC3蛋白表达;MDC及Hoechst染色方法分别用于检测细胞自噬和凋亡变化情况;克隆形成方法检测细胞辐射敏感性。构建携带人靶向HIF-1α-siRNA的反转录病毒载体,建立MCF-7 HIF-1α Ri沉默细胞模型,将细胞分为对照组(常氧组)、照射组(8 Gy)、乏氧组(CoCl2处理)、乏氧加照射组(CoCl2+8 Gy),检测细胞辐射敏感性、自噬及凋亡情况。结果 与对照组和照射组相比,HIF-1蛋白在乏氧组和乏氧加照射组表达明显增加,分别为0、0、1.00和1.89。与对照组相比,MAPLC3蛋白在照射组、乏氧组、乏氧加照射组的表达均明显上调,LC3II/LC3I比值分别为1.15、1.73、2.38和3.60。细胞辐射敏感性顺次降低,常氧+三甲基腺嘌呤(3MA)组>常氧组>乏氧+3MA组>乏氧组。成功构建HIF-1α沉默模型(pSUPER-HIF-1α Ri)与空载体对照模型(pSUPER),并检测了2种细胞的辐射敏感性。与常氧组相比,乏氧组MCF-7-pSUPER细胞存活分数显著增加(t=3.080、6.946、6.658、6.380,P<0.05),辐射敏感性降低。而MCF-7-pSUPER-HIF-1α Ri细胞,常氧与乏氧条件下细胞存活分数无明显改变。不同处理组(照射组、乏氧组和乏氧加照射组)的MCF-7-pSUPER-HIF-1α Ri细胞与MCF-7-pSUPER细胞相比较,自噬百分率分别降低21.1%、25.5%和15.5%(t=-4.635、-4.738、-6.354,P<0.05),但凋亡百分率差异无统计学意义。结论 在人乳腺癌MCF-7细胞中HIF-1α可增加辐射诱导的自噬,同时降低辐射敏感性,对细胞凋亡无影响。 |
| 英文摘要: |
| Objective To study the effects of hypoxia-inducible factor-1α (HIF-1α) on radiation-induced autophagic cell death in breast cancer cells.Methods MCF-7 cells were divided into four groups: control (normoxia, 21% Oxygen), irradiation (8 Gy X-rays), hypoxia (Cobalt chloride, CoCl2) and irradiation with hypoxia (CoCl2). 150 μmol/L CoCl2 was utilized to induce hypoxic conditions. Western blot was applied to detect the expression of HIF-1α and MAPLC3. MDC and Hoechst staining were used to detect autophagy and apoptosis. Radiosensitivity was detected by cloning formation. The short hairpin interfering RNA (shRNA) retroviral transduction particles targeting HIF-1α was transfected into MCF-7 cells to establish HIF-1α knockdown cells, then the radiosensibility, autophagy and apoptosis were detected. Results Compared with control group and irradiation group, the protein level of HIF-1 increased obviously in the normaxia, irradiation, hypoxia and irradiation with hypoxia groups,and the values were 0,0,1.00,1.89, respectively. The expression levels of MAPLC3 were markedly up-regulated in irradiation, hypoxia and irradiation with hypoxia groups as compared with control,and the ratios of LC3II/LC3I were 1.15, 1.73, 2.38 and 3.60, respectively. The radiosensitivity of MCF-7 cells decreased in the following order: normoxia with 3MA > normoxia > hypoxia with 3MA > hypoxia. HIF-1α knockdown cell (pSUPER-HIF-1α Ri) and vector control were constructed. After treatment with CoC12, survival fraction of MCF-7-pSUPER was significantly higher than that of control(t=3.080, 6.946, 6.658, 6.380, P<0.05), and radiosensitivity was down-regulated after irradiation, but there was no significant difference between normoxia and hypoxia in survival fraction of MCF-7-pSUPER-HIF-1α Ri. After treatment of irradiation or hypoxia, the autophagic fractions in MCF-7-pSUPER-HIF-1α Ri significantly decreased, reduced by 21.1%, 25.5%, 15.5%,respectively(t=4.635, 4.738, 6.354, P<0.05)as compared with MCF-7-pSUPER, but there was no change in apoptosis. Conclusions HIF-1α may increase radiation-induced autophagy and decrease radiosensitivity, but have no influence on apoptosis. |
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