circ_UBE2D2/miR-144-3p调控宫颈癌SiHa细胞增殖、凋亡、迁移及侵袭的机制研究

Study on the Mechanism of circ_UBE2D2/miR-144-3p Regulating the Proliferation,Apoptosis,Migration and Invasion of Cervical Cancer SiHa Cells

该研究探讨circ_UBE2D2/miR-144-3p分子轴对宫颈癌SiHa细胞增殖、凋亡、迁移、侵袭的影响及其可能作用机制。体外培养人宫颈癌细胞SiHa,并将各蛋白基因分别转染至其中,再将si-circ_UBE2D2和anti-miR-NC、si-circ_UBE2D2和anti-miR-144-3p分别共转染至SiHa细胞;应用双荧光素酶实验检测miR-144-3p过表达对野生型载体WT-circ_UBE2D2与突变型载体MUT-circ_UBE2D2荧光素酶活性的影响;检测各组细胞相关生物学行为活性;Western blot检测相关蛋白表达量。上调miR-144-3p表达对WT-circ_UBE2D2的荧光素酶活性降低具有明显效用(P<05),而未能影响MUT-circ_UBE2D2的荧光素酶活性;分别与转染si-NC或miR-NC相比,转染si-circ_UBE2D2或转染miR-144-3p mimics后细胞增殖抑制率、细胞凋亡率和Cleaved-caspase3、Bax蛋白水平升高(P<0.05),细胞克隆及迁移活性均显著降低(P<0.05), Bcl-2蛋白呈明显低表达(P<0.05);与共转染si-circ_UBE2D2和anti-miR-NC相比,共转染sicirc_UBE2D2和anti-miR-144-3p后,细胞增殖抑制率、细胞凋亡率和Cleaved-caspase3、Bax蛋白水平降低(P<0.05),细胞克隆、迁移等活性均明显升高(P<0.05), Bcl-2蛋白呈明显高表达(P<0.05)。干扰circ_UBE2D2表达可通过上调miR-144-3p表达而抑制宫颈癌细胞增殖、克隆形成、迁移及侵袭,并可诱导细胞凋亡。

The aim of this work is to explore the influence and its possible mechanism of circ_UBE2D2/miR-144-3p molecular axis on the proliferation,apoptosis,migration and invasion of cervical cancer SiHa cells.si-NC,si-circ_UBE2D2,miR-NC,miR-144-3p mimics were transfected into SiHa cells.si-circ BE2D2 and antimiR-NC,or si-circ_UBE2D2 and anti-miR-144-3p were co-transfected into SiHa cells.The dual luciferase reporter experiment was used to detect the effect of miR-144-3p overexpression on the luciferase activity of wild-type vector WT-circ_UBE2D2 or mutant vector MUT-circ_UBE2D2.MTT assay,plate colony formation assay,flow cytometry,and Transwell assay were used to detect the proliferation,colony formation,apoptosis,migration and invasion of SiHa cells,respectively.Western blot was used to detect the protein expression of Cleaved-caspase3,Bax,and Bcl-2.Overexpression of miR-144-3p could reduce the luciferase activity of WT-circ_UBE2D2(P<0.05)but not that of MUT-circ_UBE2D2.Compared with the si-NC or miR-NC group,the cell proliferation inhibition rate,apoptosis rate and the protein levels of Cleaved-caspase3,Bax were increased in the si-circ_UBE2D2 or miR-144-3p mimics group(P<0.05),while the number of cell clone formation,migration and invasion cells,and the protein level of Bcl-2 were decreased(P<0.05).Compared with the si-circ_UBE2D2+anti-miR-NC group,the cell proliferation inhibition rate,apoptosis rate and the protein levels of Cleaved-caspase3,Bax were reduced(P<0.05)in the si-circ_UBE2D2+anti-miR-144-3p group,while the number of cell clone formation,migration and invasion cells and Bcl-2 protein level were increased(P<0.05).circ BE2D2 interference could inhibit the proliferation,clone formation,migration and invasion,induce cell apoptosis of cervical cancer cells by up-regulating miR-144-3p.