Long Noncoding RNA SOX2-OT Aggravates Doxorubicin-Induced Apoptosis of Cardiomyocyte by Targeting miR-942-5p/DP5

Figures

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  Figure 1. DOX treatment inhibited cell viability, promoted apoptosis, and up-regulated the expression of SOX2-OT in cardiomyocytes. (A and C and E) Cardiomyocytes were stimulated with different concentrations of DOX (0.1, 1, 5, 10 μM) for 24 h. (A) Cell viability were detected by CCK-8 assay. (C) Apoptotic cells were detected using a flow cytometry. (E) The expression of SOX2-OT was detected by RT-qPCR. (B and D and F) Cardiomyocytes were stimulated with 5 μM DOX for 12, 24, 48, and 72 h. (B) Cell viability was detected by CCK-8 assay. (D) Apoptotic cells were detected using a flow cytometry. (F) The expression of SOX2-OT was detected by RT-qPCR. *p < 0.05, **p < 0.01, ***p < 0.001 vs the control group.

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  Figure 2. SOX2-OT acted as a sponge for miR-942-5p to exert biological effects. (A) Putative targeting sites for SOX2-OT and miR-942-5p. (B) Analysis of luciferase activity in HEK293 cells co-transfected with miR-942-5p mimics and SOX2-OT-WT or SOX2-OT-Mut vectors. **p < 0.01 vs SOX2-OT-MUT. (C) mRNA expression level of miR-942-5p in primary cardiomyocytes knocked down or overexpressed by SOX2-OT. **p < 0.01 vs siNC or NC. (D) Cardiomyocytes were stimulated with different concentrations of DOX (0.1, 1, 5, 10 μM) for 24 h. The expression of miR-942-5p was detected by RT-qPCR. (E) Cardiomyocytes were stimulated with 5 μM DOX for 12, 24, 48, and 72 h. *p < 0.05, **p < 0.01, ***p < 0.001 vs the control group.

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  Figure 3. SOX2-OT upregulated the expression of DP5 by sponging miR-942-5p. (A) Putative binding sites for miR-942-5p and DP5 3ʹ-UTR. (B) Analysis of luciferase activity in HEK293 cells co-transfected with miR-942-5p mimic and DP5 3ʹ-UTR-WT or DP5 3ʹ-UTR-Mut vector. **p < 0.01 vs DP5-WT. (C) DP5 mRNA expression levels in primary cardiomyocytes co-transfected with miR-942-5p mimic and SOX2-OT. **p < 0.01 vs miR-NC; ##p < 0.01 vs miR-NC+SOX2-OT. (D) Cardiomyocytes were stimulated with different concentrations of DOX (0.1, 1, 5, 10 μM) for 24 h. The expression levels of DP5 were detected by RT-qPCR. (E) Cardiomyocytes were stimulated with 5 μM DOX for 12, 24, 48, and 72 h. *p < 0.05, **p < 0.01, ***p < 0.001 vs the control group.

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  Figure 4. Biological role of SOX2-OT in DOX-induced cardiomyocyte apoptosis. Cells transfected with si-NC, si-SOX2-OT, miR-NC, miR-942-5p mimic or si-DP5 were treated with DOX for 24 h. Apoptosis rate of primary cardiomyocytes transfected with (A and C) si-NC, si-SOX2-OT and miR-NC, miR-942-5p mimic or si-DP5. (B and D) Determination of caspase-3 activity in si-NC, si-SOX2-OT and miR-NC, miR-942-5p mimic or si-DP5 transfected primary cardiomyocytes. ***p<0.001 vs control; ##p<0.01 vs si-NC; &p<0.05 vs si-SOX2-OT.

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  Figure 5. Effect of overexpression of SOX2-OT on myocardial function and apoptosis. (A) The expression levels of SOX2-OT, miR-942-5p and DP5. (B) Protein expression levels of DP5 in rats. Quantitative analysis of LVEF (C), and LVFS (D) in rats. (E) Apoptosis of cardiomyocytes was measured by the TUNEL method. **p < 0.01 and ***p < 0.001 vs Control; #p < 0.05, ##p < 0.01, and ###p < 0.001 vs si-NC; &p < 0.05 vs si-SOX2-OT.