摘要
目的探讨miR-21对肺癌干细胞增殖、侵袭能力,以及对化疗药物敏感性的影响。方法采用流式细胞边缘群分选技术从人肺癌细胞系A549中分离出肺癌干细胞。应用脂质体分别介导成熟miR-21的阻遏物(inhibitors)和模拟物(mimics)来抑制和增强肺癌干细胞miR-21的表达;实时荧光定量PCR检测肺癌干细胞中miR-21表达水平的变化;噻唑蓝(MTT)法检测转染干预前后肺癌干细胞的增殖情况和对化疗药物的敏感性;Transwell小室检测肺癌干细胞迁移能力。结果抑制组肺癌干细胞miR-21表达水平明显低于非转染组和阴性对照组,而增强组肺癌干细胞miR-21表达水平则显著高于非转染组和阴性对照组(P<0.05);抑制组肺癌干细胞转染后各时间点增殖率较非转染组和阴性对照组明显降低,而增强组肺癌干细胞转染后各时间点增殖率较非转染组和阴性对照组显著升高(P<0.05);Transwell实验显示抑制组转染后48 h穿过人工基底膜的细胞数为(29.55±5.48)个,明显少于非转染组和阴性对照组,而增强组转染后48 h穿过人工基底膜的细胞数为(74.47±8.63)个,明显多于非转染组和阴性对照组(P<0.05);顺铂(DDP)和吉西他滨(GEM)对抑制组转染后48 h肺癌干细胞的IC50值明显低于非转染组和阴性对照组(P<0.05);DDP和GEM对增强组转染后48 h肺癌干细胞的IC50值则明显高于非转染组和阴性对照组(P<0.05)。结论下调miR-21的表达可以抑制肺癌干细胞的增殖和侵袭能力,增强其对化疗药物的敏感性。
Objective To investigatet the effect of miR-21 on the capacity of proliferation,invasion and chemo-sensitivity in lung cancer stem cells. Methods Lung cancer stem cells were isolated by fluorescence activated cell sorting （FACS） from human lung adenocarcinoma cell line A549.The miR-21 inhibitors and mimics were transfected into Lung cancer stem cells by liposome.The expression levels of miR-21 in Lung cancer stem cells was measured by reverse transcription polymerase chain reaction （RT-PCR）.M3T method was used to evaluate the capability of cell proliferation and chemoresistance.The invasive capability of cells was evaluated by using transwell chamber model. Results The expression of miR-21 in inhibition group was obviously lower than non-transfection group and negative control group, and in enhanced group it was obviously higher than non-transfection group and negative control group （P〈0.05）.The proliferation rate at each time point in inhibition group was down-regulated compared with non-transfection group and negative control group, and in enhanced group it was up-regulated compared with non-transfeetion group and negative control group （P 〈 0.05 ）. Transwell chamber assay demonstrated the cell penetrating number was 29.55 ±5.48 in inhibition group at 48 h post-transfection, which was obviously less than non-transfection group and negative control group （P〈0.05）.In enhanced group, the cell penetrating number was 74.47 ±8.63 ,which was obviously more than non-transfection group and negative control group （P〈0.05） .The half inhibition concentration （IC50） of Cisplatin （DDP） and Gemeitabine （GEM） in inhibition group was obviously lower than non-transfection group and negative control group.And in enhanced group it was obviously higher than non-transfection group and negative control group （P〈0.05）. Conclusion Down-regulation of miR-21 expression in lung cancer stem ceils would inhibit their ability of proliferation and invasion, enhance the sensitivity to chemotherapy.
出处
《中南医学科学杂志》
CAS
2016年第6期643-647,共5页
Medical Science Journal of Central South China
