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  

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

作　　者：郑宝莲 何嘉轩 梁珮琪 李丹 刘小川 张静莹 ZHENG Bao-Lian;HE Jia-Xuan;LIANG Pei-Qi;LI Dan;LIU Xiao-Chuan;ZHANG Jing-Ying(Key Laboratory of 3D Printing for Stomatology,the First Dongguan Affiliated Hospital,Guangdong Medical University,Dongguan 523710,Guangdong,China)

机构地区：[1]广东医科大学附属东莞第一医院,口腔医学3D打印技术重点实验室,广东东莞523710

出　　处：《中国生物化学与分子生物学报》2022年第3期308-319,共12页Chinese Journal of Biochemistry and Molecular Biology

基　　金：广东省基础与应用基础研究基金联合基金(No.2020B1515120001);广东省普通高校重点领域(No.2020ZDZX2013);“攀登计划”广东大学生科技创新培育专项资金项目(No.Pdjh2021b2280);广东医科大学学科建设项目(No.4SG21019G)资助。

摘　　要：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。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.

关 键 词：氧化应激 hsa_circ_0087354 hsa-miR-199-3p 核因子E2相关因子2 溶质载体家族7成员11(SLC7A11) 

分 类 号：Q25[生物学—细胞生物学] Q26