隋静, 张艳秋, 李成云, 付艳云, 梁戈玉. 微小RNA-27b对RAW264.7细胞生物学功能的作用及调控机制[J]. 环境与职业医学, 2016, 33(11): 1049-1054. DOI: 10.13213/j.cnki.jeom.2016.16287
引用本文: 隋静, 张艳秋, 李成云, 付艳云, 梁戈玉. 微小RNA-27b对RAW264.7细胞生物学功能的作用及调控机制[J]. 环境与职业医学, 2016, 33(11): 1049-1054. DOI: 10.13213/j.cnki.jeom.2016.16287
SUI Jing, ZHANG Yan-qiu, LI Cheng-yun, FU Yan-yun, LIANG Ge-yu. Biological Function of MicroRNA-27b on RAW264.7 Cells and Related Regulatory Mechanism[J]. Journal of Environmental and Occupational Medicine, 2016, 33(11): 1049-1054. DOI: 10.13213/j.cnki.jeom.2016.16287
Citation: SUI Jing, ZHANG Yan-qiu, LI Cheng-yun, FU Yan-yun, LIANG Ge-yu. Biological Function of MicroRNA-27b on RAW264.7 Cells and Related Regulatory Mechanism[J]. Journal of Environmental and Occupational Medicine, 2016, 33(11): 1049-1054. DOI: 10.13213/j.cnki.jeom.2016.16287

微小RNA-27b对RAW264.7细胞生物学功能的作用及调控机制

Biological Function of MicroRNA-27b on RAW264.7 Cells and Related Regulatory Mechanism

  • 摘要: 目的

    在分析纳米二氧化钛引起巨噬细胞微小RNA(miR)表达谱改变的基础上,选择与免疫通路相关的miR-27b,探讨miR-27b 对巨噬细胞RAW264.7 的生物学功能及其对靶基因磷脂酰肌醇-3- 激酶调节亚基3(Pik3r3)的调控作用。

    方法

    用Diana-mirPath 在线软件对miR-27b 进行生物学信息分析,并通过Targetscan 软件预测靶基因位点。通过转染使RAW264.7 细胞的miR-27b 表达上调,采用MTT、流式细胞技术观察转染后细胞增殖、细胞周期和细胞凋亡的变化,应用Western blot 检测靶基因的蛋白表达水平。

    结果

    Diana-mirPath 在线软件进行miR-27b 的信号通路及靶基因分析结果显示,miR-27b 参与的信号通路-ln P ≥2.99(P<0.05)的有23 个,其中得分最高的通路为T 细胞受体信号通路(T cell receptor signaling pathway),得分为16.89。靶基因预测结果显示Pik3r3 可能是其靶基因之一。转染miR-27b 模拟物后,转染组与阴性对照组相比较,光密度值无明显差异(P=0.725),细胞周期也无明显差异(G1、S 和G2 期分别为P=0.490,P=0.088,P=0.323),但细胞凋亡率明显增高(P=0.023)。此外,转染组Pik3r3蛋白表达量无明显差异(P >0.05)。

    结论

    miR-27b 可促进RAW264.7 细胞凋亡,其调控机制尚需进一步研究。

     

    Abstract: Objective

    Based on the analysis of microRNA (miR) profile of macrophages induced by titanium dioxide, to investigate biological functions of miR-27b which is associated with immune-related pathways on macrophages RAW264.7 and regulation function of miR-27b on phosphoinositide-3-kinase, regulatory subunit 3 (Pik3r3) gene.

    Method

    Diana-mirPath online software and Targetscan software were used to analyze the related pathways and putative targets of miR-27b. miR-27b mimics were transfected into RAW264.7 cells in order to upregulate the expression level of miR-27b. Then cell proliferation, cell cycle, and apoptosis were examined using MTT assay and flow cytometry method. The protein expression levels of the putative targets were detected by Western blot.

    Result

    The results of signaling pathway and target gene analyzed by Diana-mirPath online software showed that miR-27b participated in 23 pathways -ln P≥2.99 (P<0.05), and the pathway with the highest score (16.89) was T cell receptor signaling pathway. The putative target gene results showed that Pik3r3 might be one of the target genes of miR-27b. After transfection of miR-27b mimics, there was no significant difference in optical density (D) values and cell cycles between the transfected group and the negative control group (D: P=0.725; G1: P=0.490; S: P=0.088; G2: P=0.323, respectively). Apoptosis rate of macrophages in the miR-27b mimics transfected group was significantly higher than that in the negative control group (P=0.023). Besides, compared with the negative control group, the protein expression level of Pik3r3 in the transfected group showed no significant difference (P >0.05).

    Conclusion

    In the present study, miR-27b could promote the apoptosis of RAW264.7, and the regulation of miR-27b need more studies.

     

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