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红景天苷通过调控miR-1343-3p/SOX18信号轴抑制胃癌细胞增殖

Salidroside Inhibits the Proliferation of Gastric Cancer Cells by Regulating the miR-1343-3p/SOX18 Signaling Axis

    摘要
  • 摘要:
    目的 研究红景天苷通过上调miR-1343-3p抑制胃癌细胞增殖的分子作用机制。
    方法 通过RNA数据库筛选与miR-1343-3p相关，且在红景天苷作用人胃癌细胞后表达水平显著变化的肿瘤增殖相关mRNA；基因比对及RNA结合蛋白免疫共沉淀分析miR-1343-3p与SOX18的关联性；免疫细胞化学法检测SOX18蛋白的定位；CCK-8检测红景天苷对人胃癌细胞（MGC-803、AGS）增殖的影响；将人胃癌细胞分为空白对照组和低、高剂量红景天苷组，实时荧光定量PCR（qPCR）检测miR-1343-3p和SOX18 mRNA的表达，蛋白质免疫印迹法检测SOX18蛋白表达；miR-1343-3p模拟物（miR-1343-3p mimic）、miR-1343-3p抑制剂（miR-1343-3p inhibitor）分别经LipofectamineTM 2000脂质体共转染胃癌细胞，qPCR检测miR-1343-3p和SOX18 mRNA的表达，蛋白质免疫印迹法检测SOX18蛋白表达。
    结果 生物信息学分析筛选获得miR-1343-3p下游mRNA为SOX18，基因比对验证了两者之间具有明确的结合位点，RNA结合蛋白免疫共沉淀验证了两者间存在靶向关系（P<0.05）；免疫细胞化学实验显示SOX18蛋白在核表达；CCK-8实验证明红景天苷明显抑制胃癌细胞的增殖且与作用时间和药物浓度呈现一定的依赖性；与空白对照组相比，红景天苷作用后胃癌细胞中SOX18 mRNA和蛋白表达均减少（P<0.05），miR-1343-3p表达增加（P<0.05）；与Control组相比，miR-1343-3p mimic组胃癌细胞中miR-1343-3p的表达升高、SOX18 mRNA和蛋白表达降低，miR-1343-3p inhibitor组胃癌细胞中miR-1343-3p的表达降低、SOX18 mRNA和蛋白表达升高（均P<0.05）。
    结论 红景天苷可能通过调控miR-1343-3p/SOX18信号轴抑制胃癌细胞增殖，这些调节因子有望成为胃癌新的潜在治疗靶点或生物标志物。

     

    Abstract:
    Objective  To investigate the molecular mechanism by which salidroside inhibits the proliferation of gastric cancer (GC) cells through upregulation of miR-1343-3p.
    Methods  RNA databases were used to screen for mRNAs associated with tumor proliferation and with miR-1343-3p, and exhibiting significant changes in their expression levels after salidroside treatment of human GC cells. Gene matching and immunoprecipitation of RNA-binding proteins were conducted to analyze the association between miR-1343-3p and SOX18. Immunocytochemistry was performed to determine the localization of SOX18 protein. The effect of salidroside on the proliferation of human GC cells (MGC-803 and AGS) was determined by CCK-8 assay. Human GC cells were divided into a blank control group and low- and high-dose salidroside groups. The expression of miR-1343-3p and SOX18 mRNA was measured by real-time quantitative fluorescence PCR (qPCR). The protein expression of SOX18 was measured by Western blot. GC cells were co-transfected with miR-1343-3p mimic and miR-1343-3p inhibitor, respectively, via LipofectamineTM 2000 liposomes. The expression of miR-1343-3p and SOX18 mRNA was measured by qPCR, and the protein expression of SOX18 was measured by Western blot.
    Results  Through bioinformatic analysis, SOX18 was identified as a downstream target of miR-1343-3p. Gene alignment confirmed the presence of specific binding sites between the two genes, and immunoprecipitation of RNA-binding proteins validated the targeting relationship between them (P < 0.05). Immunocytochemistry demonstrated the nuclear localization of SOX18 protein. CCK-8 assay findings demonstrated that salidroside significantly inhibited the proliferation of GC cells in a time- and dose-dependent manner. Compared with the blank control group, salidroside-treated GC cells showed decreased expression of both SOX18 mRNA and protein (P < 0.05) and an increased miR-1343-3p expression (P < 0.05). Compared with the control group, GC cells in the miR-1343-3p mimic group exhibited increased expression of miR-1343-3p and decreased expression of SOX18 mRNA and protein. In contrast, GC cells in the miR-1343-3p inhibitor group showed decreased expression of miR-1343-3p and increased expression of SOX18 mRNA and protein (all P < 0.05).
    Conclusion  Salidroside may inhibit the proliferation of GC cells by regulating the miR-1343-3p/SOX18 signaling axis and these regulators may present new potential therapeutic targets or biomarkers for gastric cancer.

     

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