circ_0001461靶向抑制miR-30a-5p对骨肉瘤细胞增殖和凋亡的影响

摘要 目的：探讨circ_0001461对骨肉瘤细胞增殖和凋亡的影响及调控机制。方法：采用实时荧光定量聚合酶反应（qRT-PCR）检测检测circ_0001461在骨肉瘤组织和细胞中的表达水平。在U2OS和HOS细胞中转染sh-NC和sh-circ_0001461后，采用CCK8检测细胞增殖情况，流式细胞术检测细胞凋亡情况，qRT-PCR检测增殖相关分子Ki-67 mRNA的表达水平，Western Blot检测凋亡相关分子Cleaved-caspase-3蛋白的表达水平。采用双荧光素酶报告基因检测circ_0001461和miR-30a-5p的结合情况。结果：circ_0001461在骨肉瘤组织中的表达水平明显高于癌旁正常组织（P<0.05），circ_0001461在骨肉瘤细胞U2OS和HOS中的表达水平均明显高于成骨细胞NHOst（P<0.05）。低表达circ_0001461能够抑制骨肉瘤细胞U2OS和HOS的增殖和增殖相关分子Ki-67的表达（P<0.05）；促进骨肉瘤细胞U2OS和HOS的凋亡和凋亡相关分子Cleaved-caspase-3蛋白的表达（P<0.05）。双荧光素酶结果显示circ_0001461能够靶向结合miR-30a-5p。低表达circ_0001461能够促进miR-30a-5p的表达（P<0.05），circ_0001461和miR-30a-5p在骨肉瘤组织中的表达呈负相关（P<0.05）。在U2OS细胞中共转染sh-circ_0001461和miR-30a-5p mimics后能够进一步加强单独转染sh-circ_0001461对U2OS细胞增殖和凋亡的影响（P<0.05）；在HOS细胞中共转染sh-circ_0001461和miR-30a-5p inhibitors后能够逆转单独转染sh-circ_0001461对U2OS细胞增殖和凋亡的影响（P>0.05）。结论：circ_0001461在骨肉瘤组织和细胞中明显高表达，低表达circ_0001461能够靶向促进miR-30a-5p的表达进而抑制骨肉瘤细胞增殖和促进细胞凋亡。

The Effect of Circ_0001461 Targeting Inhibition of miR-30a-5p on the Proliferation and Apoptosis of Osteosarcoma Cells

ABSTRACT Objective: To investigate the effect of circ_0001461 on the proliferation and apoptosis of osteosarcoma cells and its regulatory mechanism. Methods: Real-time fluorescent quantitative polymerase reaction(qRT-PCR) was used to detect the expression level of circ_0001461 in osteosarcoma tissues and cells. After transfection of sh-NC and sh-circ_0001461 in U2OS and HOS cells, CCK8 was used to detect cell proliferation, flow cytometry was used to detect cell apoptosis, qRT-PCR was used to detect the expression level of proliferation-related molecule Ki-67 mRNA, Western Blot was used to detect the expression level of apoptosis-related molecule Cleaved-caspase-3 protein. The dual luciferase reporter gene was used to detect the binding of circ_0001461 and miR-30a-5p. Results: The expression level of circ_0001461 in osteosarcoma tissue was significantly higher than that in normal adjacent tissues(P<0.05), and the expression level of circ_0001461 in osteosarcoma cells U2OS and HOS was significantly higher than that in osteoblast NHOst (P<0.05). Low expression of circ_0001461 could inhibit the proliferation of osteosarcoma cells U2OS and HOS and the expression of proliferation-related molecule Ki-67(P<0.05); promote the apoptosis of osteosarcoma cells U2OS and HOS and the apoptosis-related molecule Cleaved-caspase-3 protein Expression(P<0.05). The results of dual luciferase showed that circ_0001461 could target miR-30a-5p. Low expression of circ_0001461 could promote the expression of miR-30a-5p(P<0.05), and the expression of circ_0001461 and miR-30a-5p in osteosarcoma tissues was negatively correlated (P<0.05). Co-transfection of sh-circ_0001461 and miR-30a-5p mimics in U2OS cells could further enhance the effect of single transfection of sh-circ_0001461 on the proliferation and apoptosis of U2OS cells (P<0.05); co-transfection of sh-circ_0001461 and miR-30a-5p inhibitors in HOS cells could reverse the effect of single transfection sh-circ_0001461 on the proliferation and apoptosis of U2OS cells (P>0.05). Conclusion: circ_0001461 was highly expressed in osteosarcoma tissues and cells. The low expression of circ_0001461 could target the expression of miR-30a-5p to inhibit the proliferation of osteosarcoma cells and promote cell apoptosis.