为探讨DNA去甲基化试剂5-氮杂-2′-脱氧胞苷(5-aza-2′-deoxycytidine,5-Aza-CdR)对三维(3D)培养模式下的肺腺癌细胞A549辐射敏感性的作用,开展了系列实验。使用不同浓度5-Aza-CdR处理单层(2D)A549细胞72 h后,MTT法检测其对A549细胞的增殖抑制作用。选取低浓度(2,5μmol/L)5-Aza-CdR预处理2D和3D培养的A549细胞72 h,X射线分别辐照1,2,4,6 Gy,检测微核形成率和克隆存活。实验结果显示,不同浓度的5-Aza-CdR 均能抑制2D的A549细胞增殖,且呈剂量依赖性。5μmol/L 药物预处理2D与3D细胞并联合辐照后诱导的细胞微核形成率均显著高于相应的对照组,并且细胞存活率显著降低。不过,较低浓度5-Aza-CdR(2μmol/L)预处理的3D培养A549细胞4,6 Gy辐照后微核数目较未用药处理组显著增加,克隆存活率较未用药组显著降低(P<0.05),而在2D培养A549细胞中未观测到上述现象。研究结果表明,5-Aza-CdR能抑制A549细胞增殖,3D培养A549细胞药物预处理更能增加其辐射敏感性。结果暗示,为减少对正常细胞的毒性作用,在临床放疗中,可低剂量使用5-Aza-CdR,实现肿瘤的有效靶向治疗。
5-Aza-CdR is a specific inhibitor of DNMTs which could suppress tumor growth by demethylation of genomic DNA. There have only few studies thus far concerning it as radiosensitizers in three-dimensional (3D) cells. The principal aim of this study is to evaluate the effects of 5-Aza-CdR on the radiosensitivity of A549 cells in monolayer (2D) and 3D cultures in an attempt to find out a new combination treatments with radiotherapy. The cell proliferation was detected by MTT assay after pretreated with different doses of 5-Aza-CdR for 72 h. A549 cells were treated with or without 5-Aza-CdR (2, 5 μmol/L) for 72 h before be exposed to X-rays of 1, 2, 4, 6 Gy, respectively. The DNA damage was evaluated by micronucleus assay and clonogenic assays. Pretreatment with 5-Aza-CdR inhibited the A549 cell proliferation significantly. More micronucleus were observed after irradiation in 3D cells pretreated with 2 and 5 μmol/L concentration of drug than those without treatment. The survival fractions of cells pretreated by both 2 and 5 μmol/L drug reduced significantly in 3D cultures after irradiation. These significances, however, were found in 2D cells pretreated by only 5 μmol/L drug. Our results suggest that 5-Aza-CdR can inhibit the A549 cells proliferation and apparently enhance the radiosensitivity of cells in 3D cultures. Using of the low dose 5-Aza-CdR in clinical radiotherapy may reduce side effects and enhance effectively the cancer target therapy.
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