Hsa_circ_0087354通过hsa-miR-199-3p/SLC7A11调节MG-63细胞增殖及氧化还原状态

环状RNA(circular RNAs, circRNAs)是一类新型非编码RNA。已有研究表明,其在细胞氧化还原反应中发挥重要作用。在本文前期研究中,发现通过real-time PCR检测,hsa_circ_0087354与细胞的氧化还原状态密切相关。过表达hsa_circ_0087354后,活性氧1(reactive oxygen species1,ROS1)基因表达显著下降(P＜0.01),超氧化物歧化酶1(surperoxide dismutase1,SOD1)表达显著升高(P＜0.05);细胞内SOD和谷胱甘肽过氧化物酶(glutathione peroxidase,GPx)活性以及谷胱甘肽(glutathione,GSH)浓度显著升高(P＜0.01),细胞增殖能力增强(P＜0.05)。生物信息学分析预测,hsa-miR-199-3p与hsa_circ_0087354和溶质载体家族7成员11(solute carrier family 7 member 11,SLC7A11)存在结合位点,可能存在靶向调控关系。双荧光素酶报告基因结果证实了hsa-miR-199-3p与hsa_circ_0087354和SLC7A11之间的靶向调控关系。构建过表达hsa_circ_0087354质粒和ctrl质粒,合成hsa-miR-199a-3p、hsa-miR-199b-3p 和hsa-miR-NC mimics。通过Real-time PCR分析发现,转染hsa_circ_0087354后,hsa-miR-199-3p表达显著降低(P＜0.01),SLC7A11表达显著升高(P＜0.05)。转染hsa-miR-199-3p后,SLC7A11基因表达显著下降(P＜0.001),细胞内SOD和GPx活性以及GSH浓度显著降低(P＜0.01),细胞增殖能力下降(P＜0.05)。研究结果表明,hsa_circ_0087354通过吸附hsa-miR-199-3p,增强SLC7A11表达,促进氧化应激MG-63细胞增殖,降低氧化应激水平。

Hsa_circ_0087354 Regulates Proliferation and Redox State via hsa-miR- 199-3p/SLC7A11 in MG-63 Cells

Circular RNAs (circRNAs),a kind of novel non-coding RNAs, have been shown to play an important role in cellular redox reactions. In the previous study, we found that hsa_circ_0087354 was closely related to the cellular redox state by real-time PCR. After overexpression of hsa_circ_0087354, the relative expression of ROS1 were decreased significantly (P<0.01), while the levels of SOD1 were increased significantly (P<0.05). The activities of SOD and Gpx as well as GSH concentration were significantly increased (P<0.01), and cell proliferation was promoted in cells (P<0.05). Bioinformatics analysis predicted that there were binding sites between hsa-miR-199-3p and hsa_circ_0087354 as well as solute carrier family 7 member 11 (SLC7A11), which might have a targeted regulatory relationship. Dual luciferase reporter assay confirmed the targeted regulatory relationship between hsa-miR-199-3p with hsa_circ_0087354, and SLC7A11. Overexpressed hsa_circ_0087354 plasmid and ctrl plasmid were constructed, hsa-miR-199a-3p, hsa-miR-199b-3p and hsa-miR-NC mimics were synthesized. Real-time PCR analysis showed that the relative expression of hsa-miR-199-3p was observably decreased (P<0.01), while the relative expression of SLC7A11 in cells was dramatically increased after transfection of has_circ_0087354 plasmid (P<0.05). After transfection with hsa-miR-199-3p, the relative expressions of SLC7A11 were markedly decreased (P<0.001). The activities of SOD and Gpx, GSH concentration (P<0.01), and cell proliferation rate (P<0.05) were significantly reduced. In conclusion, hsa_circ_0087354 could enhance the expression of SLC7A11, promote the proliferation of cells and reduce the oxidative stress by adsorption of hsa-miR-199-3p.