过表达let-7f-5p通过靶向DARS2抑制MAPK信号通路调控膀胱癌细胞活性、侵袭和凋亡

    Overexpression of let-7f-5p regulates the viability, invasion and apoptosis of bladder cancer cells through targeting DARS2 and inhibiting MAPK signaling pathway

    • 摘要:
      目的: 研究let-7f-5p通过靶向线粒体天冬氨酰-tRNA合成酶2(DARS2)对膀胱癌细胞活性、侵袭和凋亡的影响及其作用机制。
      方法: 以人正常膀胱上皮细胞SV-HUC-1为对照,qRT-PCR检测膀胱癌细胞5637、T24中let-7f-5p和DARS2 mRNA的表达,Western blot检测DARS2蛋白表达。双荧光素酶报告实验检测let-7f-5p和DARS2之间靶向关系。在膀胱癌细胞T24中转染let-7f-5p mimic和DARS2过表达质粒,CCK-8检测细胞活性,transwell检测细胞侵袭,流式细胞检测细胞凋亡,Western blot检测MAPK通路蛋白表达。
      结果: 与SV-HUC-1组相比,膀胱癌5637和T24细胞组中let-7f-5p表达均降低,DARS2 mRNA和蛋白表达均升高(P < 0.05)。荧光素酶报告实验显示,let-7f-5p直接靶向作用于DARS2 3'-UTR。过表达let-7f-5p降低T24细胞活性和侵袭,促进细胞凋亡,并抑制p-p38 MAPK及p-ERK1/2蛋白表达(P < 0.05)。恢复DARS2表达后T24细胞活性和侵袭升高,细胞凋亡降低,p-p38 MAPK和p-ERK1/2蛋白表达提升(P < 0.05)。
      结论: Let-7f-5p通过靶向DARS2抑制MAPK信号通路降低膀胱癌细胞T24的细胞活性和侵袭,并促进细胞凋亡。

       

      Abstract:
      Objective To investigate the effects of let-7f-5p targeting DARS2 on the viability, invasion and apoptosis of bladder cancer (BC) cells and its underlying mechanism.
      Methods Normal human bladder epithelial cells SV-HUC-1 were used as control, the mRNA expression of let-7f-5p and DARS2 in BC cells 5637 and T24 were detected by qRT-PCR, and the protein expression of DARS2 was detected by western blot. Dual luciferase reporting assay was used to detect the targeting relationship between let-7f-5p and DARS2. Let-7f-5p mimic and DARS2 overexpression plasmid were transfected into bladder cancer cell T24. Cell viability was detected by CCK-8 method. Cell invasion was measured by transwell assay. Cell apoptosis was detected by flow cytometry. The expression of MAPK pathway proteins were detected by western blot.
      Results Compared with SV-HUC-1 group, the expression of let-7f-5p was significantly decreased, the mRNA and protein expression of DARS2 were significantly increased in bladder cancer cells 5637 and T24 (P < 0.05). Luciferase assay results showed that let-7f-5p directly targeted the 3'-UTR of DARS2. Overexpression of let-7f-5p significantly decreased the cell viability and invasion, promoted apoptosis, and inhibited the protein expression of p-p38 MAPK and p-ERK1/2 in T24 cells (P < 0.05). After DARS2 expression was restored, cell viability and invasion were significantly increased, cell apoptosis was decreased, and the protein expressions of p-p38 MAPK and p-ERK1/2 were significantly increased in T24 cells (P < 0.05).
      Conclusion Let-7f-5p reduces T24 cell viability and invasion, promotes cell apoptosis by targeting DARS2 and inhibiting MAPK signaling pathway.

       

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