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长链非编码RNA CBR3-AS1通过PI3K/AKT/mTOR/S6K通路介导白血病细胞对阿糖胞苷的耐药机制

Long non-coding RNA CBR3-AS1 mediates the mechanism of cytarabine resistance in leukemia cells through the PI3K/AKT/mTOR/S6K pathway

    摘要
  • 摘要:
    目的 探讨长链非编码RNA (lncRNA)CBR3-AS1通过PI3K/AKT/mTOR/S6K通路介导急性髓细胞白血病(AML)细胞对阿糖胞苷耐药的可能机制。
    方法 在2个AML细胞株K562和HL-60中建立耐药模型，分别为K562-R和HL-60-R。逆转录-实时定量聚合酶链反应(RT-qPCR)法检测lncRNA CBR3-AS1在K562-R和HL-60-R中的表达情况。在K562和HL-60细胞株中，利用质粒过表达lncRNA CBR3-AS1; 在K562-R和HL-60-R细胞株中，利用siRNA敲低lncRNA CBR3-AS1表达，并计算阿糖胞苷的半数抑制浓度(IC50)。蛋白印迹法检测PI3K/AKT/mTOR/S6K通路激活情况。
    结果 相较于K562和HL-60细胞株, K562-R和HL-60-R细胞株中lncRNA CBR3-AS1表达升高，差异有统计学意义(P < 0.05)。过表达lncRNA CBR3-AS1可提高阿糖胞苷IC50超过1 000 μmol/L(P < 0.05)。敲低lncRNA CBR3-AS1后，阿糖胞苷在耐药细胞株K562-R和HL-60-R中的IC50分别降低到21.27 μmol/L和12.10 μmol/L, 差异有统计学意义(P < 0.05)。过表达lncRNA CBR3-AS1显著提高磷酸肌醇3激酶(PI3K)、AKT、哺乳动物雷帕霉素靶蛋白(mTOR)、S6K蛋白的磷酸化水平(P < 0.05); 敲低lncRNA CBR3-AS1显著降低PI3K、AKT、mTOR、S6K蛋白的磷酸化水平(P < 0.05)。
    结论 lncRNA CBR3-AS1可能通过激活PI3K/AKT/mTOR/S6K通路介导AML细胞对阿糖胞苷的耐药。

     

    Abstract:
    Objective To investigate the possible mechanism of long non-coding RNA (lncRNA) CBR3-AS1 mediating cytarabine resistance in acute myeloid leukemia (AML) cells through PI3K/AKT/mTOR/S6K pathway.
    Methods Drug resistance models were established in two AML cell strains K562 and HL-60, named as K562-R and HL-60-R, respectively. Reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) was used to detect the expression of lncRNA CBR3-AS1 in K562-R and HL-60-R. LncRNA CBR3-AS1 was overexpressed by plasmids in the K562 and HL-60 cell strains; lncRNA CBR3-AS1 expression was knocked down by siRNA in K562-R and HL-60-R cell strains, and the 50% inhibitory concentration (IC50) of cytarabine was calculated. K562-R and HL-60-R cell lines, siRNA was used to knock down the expression of lncRNA CBR3-AS1. Western blot was used to detect the activation of PI3K/AKT/mTOR/S6K pathway.
    Results Compared with K562 and HL-60 cell strains, the expressions of lncRNA CBR3-AS1 in K562-R and HL-60-R cell strains were significantly increased (P < 0.05). Overexpression of lncRNA CBR3-AS1 increased the IC50 of cytarabine to over 1 000 μmol/L (P < 0.05). After knock down lncRNA CBR3-AS1, the IC50 of cytarabine in drug-resistant cell strains K562-R and HL-60-R were significantly reduced to 21.27 μmol/L and 12.10 μmol/L (P < 0.05), respectively. Overexpression of lncRNA CBR3-AS1 significantly increased the phosphorylation levels of PI3K, AKT, mammalian target of rapamycin (mTOR) and S6K proteins (P < 0.05). Knockdown lncRNA CBR3-AS1 significantly reduced the phosphorylation levels of PI3K, AKT, mTOR and S6K proteins (P < 0.05).
    Conclusion LncRNA CBR3-AS1 may mediate the resistance of AML cells to cytarabine by activating the PI3K/AKT/mTOR/S6K pathway.

     

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