【摘要】 目的:探讨miR-424/高迁移率蛋白A1基因（nigh mobility protein A1,HMGA1）分子轴对乳腺癌细胞放疗敏感性的影响及其分子机制。方法:收集2014年4月至2017年4月无锡市第四人民医院肿瘤放疗科经手术切除的50例乳腺癌患者的癌组织标本,用qPCR和Western blotting检测miR-424和HMGA1 m RNA与蛋白在乳腺癌放疗敏感患者和放疗抵抗患者癌组织中的表达水平。用不同辐射强度（0、2、4、6和8 Gy）的⁶⁰Coγ-射线处理人乳腺癌细胞株MDA-MB-468后,观察细胞miR-424和HMGA1的表达变化。向MDA-MB-468细胞中转染miR-424 mimic/inhibitor和pcDNA-HMGA1,采用平板克隆形成实验、MTT法、Transwell小室法和Annexin V-FITC/PI染色流式细胞术检测miR-424对辐射处理后乳腺癌细胞增殖、侵袭和凋亡的影响。用双荧光素酶报告基因验证miR-424与HMGA1的靶向关系。结果:与放疗抵抗的乳腺癌患者比较,miR-424在放疗敏感的乳腺癌患者癌组织中高表达（P<0.01）,HMGA1低表达（P<0.01）。与0、2和4 Gy处理组细胞比较,6和8 Gy的γ-射线处理后乳腺癌MDA-MB-468细胞凋亡率和miR-424的表达水平显著升高（均P<0.01）;细胞的侵袭能力和HMGA1的表达水平显著降低（均P<0.01）。双荧光素酶报告基因证实miR-424靶向作用HMGA1并下调其表达水平。miR-424通过靶向下调HMGA1显著抑制MDA-MB-468细胞的增殖、侵袭并促进细胞凋亡（均P<0.01）,进而上调MDA-MB-468细胞对放疗的敏感性。结论:miR-424/HMGA1分子轴可调控乳腺癌放疗敏感性,过表达miR-424可增强乳腺癌MDA-MB-468细胞对γ-射线放疗的敏感性。更多还原

【Abstract】 Objective: To explore the effect of miR-424/HMGA1（high mobility protein A1） axis on the radio-sensitivity of breast cancer cells and the possible mechanism. Methods: A total of 50 cases of breast cancer tissues from patients, who underwent surgical resection at the Department of Oncological Radiotherapy, Wuxi Fourth People’s Hospital from April 2014 to April 2017, were collected for this study. Real-time quantitative polymerase chain reaction（qPCR） and Western blotting were performed to evaluate the mRNA and protein expressions of miR-424 and HMGA1 in breast cancer tissues of radiation sensitive and insensitive patients. After being treated with different doses of ⁶⁰Co γ-ray radiation（0, 2, 4, 6 and 8 Gy）, the expression changes of miR-424 and HMGA1 in breast cancer MDA-MB-468 cells were observed. Subsequently, miR-424 mimic/inhibitor and pcDNA-HMGA1 were transfected into MDA-MB-468 cells, and the effect of miR-424 on cell proliferation, invasion and apoptosis of radiation-treated MDA-MB-468 cells were evaluated by colony formation assay, MTT assay, Transwell assay and Annexin V-FITC/PI double staining flow cytometry assay, respectively. Furthermore, dual luciferase reporter gene assay was used to verify whether HMGA1 was a target gene of miR-424. Results: The patients in radio-sensitive group exhibited higher miR-424 expression but lower HMGA1 expression than the patients in insensitive group（all P<0.01）. Compared with the cells treated with 0, 2 and 4 Gy radiation, the cells treated with 6 and 8 Gy radiation exhibited significantly higher apoptosis rate and miR-424 expression but lower HMGA1 expression and cell invasion（all P<0.01）. Moreover, luciferase reporter gene assay confirmed that miR-424 down-regulated HMGA1 expression. Mechanistically, miR-424 significantly inhibited cell proliferation, invasion and induced apoptosis of MDA-MB-468 cells（all P<0.01） via targeted down-regulating HMGA1, and further upregulated the radio-sensitivity of breast cancer cells. Conclusion: miR-424/HMGA1 axis regulates the radio-sensitivity of breast cancer,and over-expression of miR-424 may increase the sensitivity of MDA-MB-468 cells to γ-ray radiation therapy.更多还原