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刘念, 张红, 薛中柱. miRNA-199a-5p对骨肉瘤细胞增殖及自噬的影响[J]. 肿瘤防治研究, 2017, 44(3): 197-201. DOI: 10.3971/j.issn.1000-8578.2017.03.009
引用本文: 刘念, 张红, 薛中柱. miRNA-199a-5p对骨肉瘤细胞增殖及自噬的影响[J]. 肿瘤防治研究, 2017, 44(3): 197-201. DOI: 10.3971/j.issn.1000-8578.2017.03.009
LIU Nian, ZHANG Hong, XUE Zhongzhu. Effect of miRNA-199a-5p on Cell Proliferation and Autophagy in Osteosarcoma[J]. Cancer Research on Prevention and Treatment, 2017, 44(3): 197-201. DOI: 10.3971/j.issn.1000-8578.2017.03.009
Citation: LIU Nian, ZHANG Hong, XUE Zhongzhu. Effect of miRNA-199a-5p on Cell Proliferation and Autophagy in Osteosarcoma[J]. Cancer Research on Prevention and Treatment, 2017, 44(3): 197-201. DOI: 10.3971/j.issn.1000-8578.2017.03.009

miRNA-199a-5p对骨肉瘤细胞增殖及自噬的影响

Effect of miRNA-199a-5p on Cell Proliferation and Autophagy in Osteosarcoma

  • 摘要:
    目的 探讨miRNA-199a-5p对人骨肉瘤细胞株U2OS增殖及自噬的影响及其可能的作用机制。
    方法 实时定量PCR法检测32例骨肉瘤患者的病理组织标本及配对癌旁正常组织中miRNA-199a-5p的表达水平。将miRNA-199a-5p mimics转染入U2OS细胞,使其过表达miRNA-199a-5p,采用CCK-8法和流式细胞仪检测细胞增殖及周期的变化;Western blot检测周期相关蛋白(cyclinD1、P27、pRb)及自噬相关蛋白(Beclin 1、LC3Ⅱ/Ⅰ)表达水平。
    结果 miRNA-199a-5p在骨肉瘤组织及癌细胞U2OS中低表达,且表达水平与肿块大小、远距转移、Enneking临床分期及病理分级密切相关。过表达miRNA-199a-5p后,U2OS细胞的增殖能力显著降低(P < 0.05),同时cyclinD1、pRb表达水平显著下降,而P27、LC3Ⅱ/Ⅰ、Beclin 1的表达水平显著增加(P < 0.05)。
    结论 miRNA-199a-5p能够抑制人骨肉瘤细胞株U2OS的增殖并激活细胞自噬;其作用机制可能与下调cyclinD1、上调P27、抑制pRb蛋白的磷酸化、造成细胞周期G1/S阻滞有关。提示miRNA-199a-5p可以作为骨肉瘤临床治疗的潜在作用位点。

     

    Abstract:
    Objective To investigate the effect of miRNA-199a-5p on cell proliferation and autophagy in osteosarcoma cell line U2OS and its possible mechanism.
    Methods The expression of miRNA-199a-5p were detected by qRT-PCR after transfection with the miRNA-199a-5p mimics in U2OS cells in paraffin section and paired adjacent normal tissue from 32 patients with osteosarcoma. The proliferation were measured by CCK-8 assay and flow cytometry assay. The expression of cell cycle related protein (cyclinD1, P27, pRb) and autography related protein (Beclin 1, LC3Ⅱ/Ⅰ) were measured by Western blot assay.
    Results MiRNA-199a-5p was lowly expressed in both osteosarcoma tissues and U2OS cells (P < 0.05). The expression level of miRNA-199a-5p was correlated with the tumor size, distant metastasis, Enneking stage and pathological grade stage (P < 0.05). Overexpression of miRNA-199a-5p suppressed U2OS cell proliferation (P < 0.05), and downregulated the expression of cyclinD1 and pRb, while upregulated the expression of P27, Beclin 1 and LC3Ⅱ/Ⅰ(P < 0.05).
    Conclusion miRNA-199a-5p suppresses cell proliferation but induces cell autophagy. And its mechanism may be related to thedownregulation of cyclinD1 and upregulation of P27 protein expression, then supressed the phosphorylation of pRb, resulting in G1/S arrest in human osteosarcoma cell line U2OS. It suggests that miRNA-199a-5p may be used as a potential new target for targeted therapy of osteosarcoma.

     

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