Liang Hui, Dong Ziyan, Liu Jiang-Feng, Chuang Wei, Gao Li-Zhen, Ren Yu-Guo
Department of Hematology, Qingdao Women and Children's Hospital, Qingdao, China.
Department of Ophthalmology and Otorhinolaryngology, People's Hospital of Lanshan District, Rizhao, China.
Histol Histopathol. 2017 Sep;32(9):899-907. doi: 10.14670/HH-11-837. Epub 2016 Oct 27.
Recent studies have shown that high miR-155 expression was associated with poor prognosis in patients with acute myelogeneous leukemia (AML). Furthermore, targeting miR-155 results in monocytic differentiation and apoptosis. However, the exact role and mechanisms of miR-155 in human AML remains speculative. HL-60 cells were treated with anti-miR-155 for 72 h. Cell growth and apoptosis in vitro were detected by MTT, BrdU proliferation, colony formation and flow cytometry assay. The effect of anti-miR-155 on growth of HL-60 cells was also evaluated in a leukemia mouse model. Slug cDNA and PUMA siRNA trannsfection was used to assess the signal pathway. Different protein expression was detected by western blot assay and quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) assay. The results shown that targeting miR-155 resulted in a 24-fold decrease of miR-155 expression compared to negative control in the HL-60 cells. Targeting miR-155 significantly downregulated Slug and upregulated PUMA expression, and decreased HL-60 cell growth by 70% , impaired colony formation by approximately 60%, and increased HL-60 cell apoptosis by 45%. Targeting PUMA reversed miR-155 sliencing-induced proliferation and apoptosis of HL-60 cells. Restoration of Slug decreased PUMA expression. In murine engraftment models of HL-60 cells, we showed that targeting miR-155 was able to reduce tumor growth. This was accompanied with decreased Slug expression and increased PUMA expression in these tumors. Collectively, our findings strongly suggest targeting miR-155 exhibited in vivo and in vitro antileukemic activities in AML through a novel mechanism resulting in inhibition of Slug expression and increase of PUMA expression.
近期研究表明，急性髓系白血病（AML）患者中miR-155高表达与预后不良相关。此外，靶向miR-155可导致单核细胞分化和凋亡。然而，miR-155在人类AML中的确切作用和机制仍具有推测性。用抗miR-155处理HL-60细胞72小时。通过MTT、BrdU增殖、集落形成和流式细胞术检测体外细胞生长和凋亡。在白血病小鼠模型中也评估了抗miR-155对HL-60细胞生长的影响。使用Slug cDNA和PUMA siRNA转染来评估信号通路。通过蛋白质印迹分析和定量逆转录聚合酶链反应（qRT-PCR）检测不同的蛋白质表达。结果显示，与HL-60细胞中的阴性对照相比，靶向miR-155导致miR-155表达降低24倍。靶向miR-155显著下调Slug并上调PUMA表达，使HL-60细胞生长减少70%，集落形成受损约60%，并使HL-60细胞凋亡增加45%。靶向PUMA可逆转miR-155沉默诱导的HL-60细胞增殖和凋亡。恢复Slug可降低PUMA表达。在HL-60细胞的小鼠移植模型中，我们表明靶向miR-155能够减少肿瘤生长。这伴随着这些肿瘤中Slug表达降低和PUMA表达增加。总体而言，我们的研究结果强烈表明，靶向miR-155通过一种新机制在体内和体外对AML表现出抗白血病活性，该机制导致Slug表达受到抑制和PUMA表达增加。
