Effect of miR-499-5p/SOX6 axis on atrial fibrosis in rats with atrial fibrillation

Figures

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  Figure 1. miR-499-5p was underexpressed in myocardial tissues of AF rats and inversely correlated with SOX6. Rats were injected daily with Ach–CaCl2 mixture via tail veins for 7 days. (a) ECG analysis; (b) RT-qPCR determined the levels of miR-499-5p and SOX6 in rat myocardial tissues; (c) Western blotting determined the SOX6 protein levels in rat myocardial tissues. N = 6. An independent sample t-test was used for comparisons between groups in panels b and c. **p < 0.01.

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  Figure 2. miR-499-5p alleviated AF in rats by inhibiting atrial fibrosis. Rats were injected with Lv-miR-499-5p/Lv-NC plasmids via tail veins, followed by AF modeling 14 days later. (a) RT-qPCR measured miR-499-5p level in rat myocardial tissues; (b) ECG analysis; (c) Masson staining evaluated atrial fibrosis in rats; (d) Western blotting measured the protein levels of atrial fibrosis markers collage I, α-SMA, and TGFβ1; N = 6. An independent sample t-test was used for comparisons between two groups in panel (a), and one-way ANOVA was used for comparisons among multiple groups in panels (b)/(c)/(d), followed by Tukey’s test. *p < 0.05, **p < 0.01.

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  Figure 3. miR-499-5p attenuated atrial fibrosis by targeting SOX6 in AF rats. (a) TargerScan7.2 database predicted the targeted binding sites of miR-499-5p and SOX6; (b) the dual-luciferase assay; (c) RT-qPCR determined SOX6 mRNA level in rat myocardial tissues; (d) Western blotting determined SOX6 protein level in rat myocardial tissues; (e) Masson staining evaluated atrial fibrosis in rats; (f) Western blotting determined collage I, α-SMA, and TGFβ1 protein levels in rat myocardial tissues. N = 6. An independent sample t-test was conducted for comparisons between groups in panels (b)/(e)/(f), and one-way ANOVA was used for multiple groups in panels c/d, followed by Tukey’s test. *p < 0.05, **p < 0.01.

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  Figure 4. AF upregulated p21 to trigger cell cycle arrest, senescence, and apoptosis in rat cardiomyocytes. (a) Western blotting determined p21, CDC25, and Cyclin B1 protein levels in myocardial tissues; (b) immunohistochemistry assay determined SA-β-gal and γ-H2AX levels in myocardial tissues; (c) TUNEL staining evaluated cell apoptosis. N = 6. An independent sample t-test was used for comparisons between two groups. **p < 0.01.

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  Figure 5. SOX6 silencing downregulated p21 and mitigated cell cycle arrest, senescence, and apoptosis in rat cardiomyocytes. Rats were injected with si-SOX6 plasmids via tail veins on day 14 before AF modeling. (a) RT-qPCR measured SOX6 mRNA level in rat myocardial tissues; (b) Western blotting measured SOX6, p21, CDC25, and Cyclin B1 protein levels in rat myocardial tissues; (c) immunohistochemistry assay determined SA-β-gal and γ-H2AX protein levels; (d) TUNEL staining assessed cell apoptosis. N = 6. An independent sample t-test was conducted for comparisons between two groups. **p < 0.01.