Sal-miR-58通过HAX1介导胶质瘤细胞放射敏感性的作用及机制研究  

Effect and mechanism of the role of Sal-miR-58 in mediating radiosensitivity of glioma U251 cells via HAX1

作　　者：王国辉 葛鸿瑶 杜振宇 杨高山 Wang Guohui;Ge Hongyao;Du Zhenyu;Yang Gaoshan(Department of Radiotherapy,Tianjin First Center Hospital,Tianjin 300000,China;Department of Biochemistry and Biology,College of Basic Medicine,Hebei University of Chinese Medicine,Shijiazhuang 050200,China)

机构地区：[1]天津市第一中心医院放疗科,天津300000 [2]河北中医药大学基础医学院生物化学与生物学教研室,石家庄050200

出　　处：《中华放射肿瘤学杂志》2024年第8期746-752,共7页Chinese Journal of Radiation Oncology

基　　金：河北省自然科学基金(H2021423069);河北省高等学校科学技术研究项目(BJ2021031);河北中医学院博士科研基金项目(BSZ2021003)。

摘　　要：目的探讨丹参来源的Sal-miR-58对人胶质瘤细胞U251放射敏感性的影响及其分子机制。方法分别采用体外合成的不同浓度的miR-Ctl和Sal-miR-58模拟物处理胶质瘤U251细胞,再给予X线照射。MTT检测Sal-miR-58对U251细胞活力的影响。Hoechst33342/碘化丙啶(PI)双染检测Sal-miR-58对照射后胶质瘤U251细胞凋亡的影响。2'',7''-二氯二氢荧光素二乙酸酯(DCFH-DA)探针检测Sal-miR-58联合照射对肿瘤细胞活性氧含量的影响。克隆形成实验检测Sal-miR-58联合照射对U251细胞放射敏感性的影响。蛋白质印迹法检测Sal-miR-58调控抗凋亡蛋白造血细胞特异性蛋白1-相关蛋白X-l(HAX1)及凋亡标志蛋白B细胞淋巴瘤2(Bcl-2)、活化的胱天蛋白酶(Cleaved-Caspase)9、Cleaved-Caspase 3的表达情况。多组间比较行单因素方差分析,两组间比较采用独立样本t检验。结果Sal-miR-58能加重照射后U251细胞增殖的抑制(P<0.05)。Sal-miR-58促进U251细胞凋亡,并且增加照射后U251细胞活性氧的产生(P<0.05)。克隆形成实验显示,Sal-miR-58增加U251细胞放射敏感性,放射增敏比为1.43。蛋白质印迹法结果显示,Sal-miR-58抑制放射诱导的U251细胞HAX1表达,且Sal-miR-58能通过阻抑HAX1表达进而抑制Bcl-2表达,促进Caspase 9及Caspase 3的活化。结论Sal-miR-58具有增强U251细胞放射敏感性的作用,其可能机制是Sal-miR-58通过下调射线诱导的HAX1表达,促进肿瘤细胞的凋亡。ObjectiveTo investigate the effect of salvia miltiorrhiza-derived Sal-miR-58 on the radiosensitivity of glioma U251 cells and its possible mechanism.MethodsGlioma U251 cells were treated with different concentrations of miR-Ctl or Sal-miR-58 mimic,and subsequently treated with radiation to establish the radiotherapy model in vitro.The effect of Sal-miR-58 upon U251 cell viability was assessed by MTT assay.The effect of Sal-miR-58 on apoptosis of glioma U251 cells was evaluated by Hoechst33342/propidium iodide(PI)staining.The changes of reactive oxygen species(ROS)content in U251 cells were detected by 2',7'-dichlorodihydrofluorescein diacetate(DCFH-DA)fluorescent probe.The effect of Sal-miR-58 combined irradiation on the radiosensitivity of U251 cells was detected by clone formation assay.The expression levels of HCLS1-related protein X-1(HAX1),apoptosis marker proteins B cell lymphoma 2(Bcl-2),cleaved-cysteine-containing aspartate-specific protease(Cleaved-Caspase)9 and Cleaved-Caspase 3 were detected by Western blot.Multi-group comparison was conducted by one-way ANOVA.Two-group comparison was performed by independent sample t-test.ResultsSal-miR-58 could exacerbate the inhibition of U251 cell proliferation after irradiation(P<0.05).Sal-miR-58 could promote the apoptosis of U251 cells and increase the production of ROS in U251 cells after radiation(P<0.05).Clone formation assay showed that Sal-miR-58 increased the radiosensitivity of U251 cells,with a radiosensitization ratio of 1.43.Western blot showed that Sal-miR-58 inhibited the expression of HAX1 in U251 cells.Sal-miR-58 could inhibit the expression of Bcl-2 and increase the activation of Caspase 3 and Caspase 9 by reducing the expression of HAX1.ConclusionsSal-miR-58 enhances the radiosensitivity of U251 cells.The possible mechanism is that Sal-miR-58 inhibits the expression of HAX1 induced by radiation and accelerates the apoptosis process of tumor cells.

关 键 词：微RNAs 丹参来源 HAX1基因 放射敏感性 人胶质瘤 

分 类 号：R739.4[医药卫生—肿瘤]