The unique expression of non-coding microRNAs in radioresistant fraction of acute promyelocytic leukemia, HL60 cell. The unique expression of non-coding microRNAs in radioresistant fraction of acute promyelocytic leukemia, HL60 cell

A one of general treatments of patient with acute promyelocytic leukemia (APL) is performed a high dose rate of total-body irradiation therapy. However, the appearance of radioresistant cell that is rarely expressed induces a harmful outcome even performing radiotherapy. We have been established a radioresistant APL cell model (Resistant type), and some characteristics was clarified. But little is known as to the micro RNAs (miRs) expression profile in this cell. In the present study, we analyzed on the expression profile of miRs included in small RNAs using miRNA microarray, The focused miRs that is enabled to investigate by normalization was 1187 molecules. Among these miRs, 27 miRNAs (10 up-regulated miRs and 17 down-regulated miRs) in Res-HL60 showed a p-value 1.5 or Fold change < 0.66 in comparison to Wt-HL60. In addition, five miRs were reproducible in RT-qPCR (miR-146a-5p, miR-30c-1-3p, miR-671-5p, miR-610, miR-3675-5p). miRNA-target gene regulation networks were looked for the five miRNAs using OmicsNet 2.0. Interestingly, the five miRNAs were Superoxide Dismutase 2 (SOD2) mRNA. These results were suggested that radioresistant leukemia cell acquires antioxidant system through from specific miRs endogenously. Overall design: Human acute promyelocytic leukemia cell line HL60 was prepared after frequent irradiation (4Gy X-irradiation ×4 fractions per 4 weeks) with confirmed radiation-resistance characteristics. The extracted total RNA in wild type cell and radiation-resistant cell were performed micro RNA microarray and calculate the expression of micro RNAs.

急性早幼粒细胞白血病（acute promyelocytic leukemia, APL）患者的常规治疗方案之一为高剂量率全身照射治疗。然而，极少数表达的辐射抗性细胞会导致即便实施放疗，仍出现不良预后。本团队已成功构建辐射抗性APL细胞模型（抗性株）并明确了其部分生物学特征，但该细胞中微小RNA（micro RNAs, miRs）的表达谱仍有待深入研究。
本研究通过miRNA微阵列（miRNA microarray）分析小RNA组分中miRs的表达谱，经标准化处理后可有效纳入分析的miRs共计1187个。相较于野生型HL60细胞（Wt-HL60），辐射抗性HL60细胞（Res-HL60）中有27个miRNAs呈现显著差异表达，其中10个上调、17个下调，且其p值符合1.5阈值、变化倍数<0.66。此外，通过实时定量聚合酶链反应（RT-qPCR）验证了5个miRs的表达差异，分别为miR-146a-5p、miR-30c-1-3p、miR-671-5p、miR-610及miR-3675-5p。
本研究借助OmicsNet 2.0平台对上述5个miRs进行靶基因调控网络预测。值得注意的是，其中5个miRs的靶基因涵盖超氧化物歧化酶2（Superoxide Dismutase 2, SOD2）的mRNA。上述结果提示，辐射抗性白血病细胞可通过内源性特异性miRs获得抗氧化系统。
实验整体设计：将人急性早幼粒细胞白血病细胞系HL60经反复辐照构建辐射抗性模型：每周给予4次、每次4Gy的X射线照射，持续4周，并验证其辐射抗性特征。分别提取野生型细胞与辐射抗性细胞的总RNA，进行miRNA微阵列检测并计算miRNAs的表达水平。