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LncRNA TTTY15通过调控miR-520a-3p表达对高糖诱导大鼠心肌细胞增殖和凋亡的影响
作者:郝翠翠1  张青2 
单位:1. 聊城市第二人民医院 内分泌科, 山东 聊城 252600;
2. 聊城市第二人民医院 心内科, 山东 聊城 252600
关键词:TTTY15 miR-520a-3p 糖尿病 心肌细胞 增殖 凋亡 大鼠 
分类号:R-33;R363.1
出版年·卷·期(页码):2021·49·第一期(46-52)
摘要:

目的: 探讨长链非编码RNA TTTY15(LncRNA TTTY15)对高糖诱导大鼠心肌细胞H9c2增殖和凋亡的影响及作用机制。方法: 高糖诱导H9c2细胞后,实时荧光定量PCR(RT-qPCR)检测H9c2细胞中TTTY15和miR-520a-3p水平,四甲基噻唑蓝染色法(MTT)检测细胞增殖,流式细胞仪检测细胞凋亡,蛋白印迹(Western blotting)法检测细胞周期蛋白D1(cyclin D1)和活化的半胱天冬酶-3(C-caspase-3)蛋白水平。转染TTTY15小干扰RNA或miR-520a-3p模拟物至H9c2细胞,用上述相同方法观察下调TTTY15或上调miR-520a-3p对高糖诱导的H9c2细胞存活率、凋亡率以及cyclin D1和C-caspase-3蛋白表达的影响。双荧光素酶报告基因实验验证TTTY15与miR-520a-3p调控关系。结果:高糖促进了H9c2细胞中TTTY15表达、细胞凋亡率及C-caspase-3蛋白表达(P<0.05),降低了H9c2细胞存活率、miR-520a-3p表达及cyclin D1蛋白表达(P<0.05)。下调TTTY15或上调miR-520a-3p可提高高糖诱导的H9c2细胞存活率及cyclin D1蛋白水平(P<0.05),降低细胞凋亡率和C-caspase-3蛋白水平(P<0.05)。TTTY15在H9c2细胞中靶向负调控miR-520a-3p表达。下调miR-520a-3p可逆转下调TTTY15对H9c2细胞存活率、凋亡率及cyclin D1和C-caspase-3蛋白表达的影响。结论: TTTY15可能通过下调细胞中miR-520a-3p表达抑制高糖诱导的H9c2细胞增殖并促进细胞凋亡。

Objective: To investigate the effects of long non-coding RNA(LncRNA)TTTY15 on high glucose-induced rat cardiomyocyte H9c2 proliferation and apoptosis and its mechanism. Methods: After H9c2 cells were induced by high glucose, the levels of TTTY15 and miR-520a-3p in H9c2 cells were detected by real-time fluorescence quantitative PCR(RT-qPCR); cell proliferation was detected by MTT assay; cells apoptosis was detected by flow cytometry; the protein levels of cyclin D1 and activated caspase-3 (C-caspase-3) were detected by Western blotting. After TTTY15 small interfering RNA or miR-520a-3p mimic was transfected into H9c2 cells, the same methods as described above were used to observe the effects of down-regulating TTTY15 or up-regulating miR-520a-3p on high glucose-induced H9c2 cell survival rate, apoptosisrate and the protein expression levels of cyclin D1 and C-caspase-3. The dual luciferase reporter gene assay verified the regulatory relationship between TTTY15 and miR-520a-3p. Results: High glucose promoted the expression of TTTY15, apoptosis rate and the expression of C-caspase-3 protein in H9c2 cells (P<0.05), but reduced the expression of miR-520a-3p, survival rate and the expression of cyclin D1 protein (P<0.05). Down-regulating TTTY15 or up-regulating miR-520a-3p increased high glucose-induced H9c2 cell survival rate and the expression of cyclin D1 protein (P<0.05), while decreased apoptosis rate and the expression of C-caspase-3 protein (P<0.05). TTTY15 negatively regulated the expression of miR-520a-3p in H9c2 cells. Down-regulating miR-520a-3p reversed the effects of down-regulating TTTY15 on H9c2 cell survival rate, apoptosisrate and the expression levels of cyclin D1 and C-caspase-3 protein. Conclusion: TTTY15 inhibits high glucose-induced H9c2 cell proliferation and promotes apoptosis by down-regulating the expression of miR-520a-3p in cells.

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