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. 2020 Apr 24;11(4):284. doi: 10.1038/s41419-020-2447-7

Fig. 2. Prediction and validation of circGCN1L1-binding miRNAs.

Fig. 2

a Circular RNA Interactome, StarBase, and RegRNA2.0 databases were used to predict the interacting miRNAs of circGCN1L1 and its potential binding sites. b RT-qPCR detection of circGCN1L1 and miR-330-3p expression in synovial tissues and synoviocytes. N = 15 (five different samples in each group for three independent experiments). *p < 0.05. c HEK-293T cells were transfected with circGCN1L1 reporter or circGCN1L1 MT reporter and miR-330-3p or NC, followed by luciferase activity detection. Relative luciferase activity is demonstrated in the histogram. N = 3 (three replicates). *p < 0.05. d Synoviocytes from the control patient were transfected with 50 nM circGCN1L1 or shcircGCN1L1 plasmid. After 48 h, the expression levels of circGCN1L1 and TNF-α were measured by RT-qPCR and normalized to the β-actin level. The expression of miR-330-3p was measured by RT-qPCR and normalized to U6 expression. N = 6 (two different samples for three independent experiments). *p < 0.05. e Synoviocytes from the control patient were transfected with 50 nM circGCN1L1 plasmid and/or miR-330-3p mimics. After 48 h, the expression levels of circGCN1L1, TNF-α, MMP3, and MMP13 were measured by RT-qPCR and normalized to the β-actin level. The expression of miR-330-3p was normalized to U6 expression. N = 6 (two different samples for three replicates). *p < 0.05. Data are presented as mean ± S.D. Two-tailed t-test (b, d) and one-way ANOVA with Bonferroni test (c, e) were performed. TNF tumor necrosis factor, NC normal control, RT-qPCR real-time quantitative polymerase chain reaction.