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. 2021 Jun 12;25(13):6032–6045. doi: 10.1111/jcmm.16460

FIGURE 1.

FIGURE 1

CXCL12‐induced miR‐126‐3p endogenous expression in vitro and ex vivo. (A) To study the effect of CXCL12 on egfl7‐miR‐126 promotor activity, Huh7 were co‐transfected with plasmid pGL3Basic‐miR‐126‐EGFL7‐Promoter (Addgene) and control pGL4.73 [hRluc/SV40] (Promega) for 24 hours. Then the cells were stimulated or not stimulated by CXCL12 for 24 h at 6 nmol.L‐1. Detection of luminescence was performed using Dual‐Glo® Luciferase Assay System (Promega). To study the effect of CXCL12 on miR‐126 expression level, HUVEC (B) or rat aortas (C) were stimulated or not stimulated by CXCL12 for 24 h at 6 nmol.L‐1. After total RNA extraction, miR‐126 level expression was analysed using qRT‐PCR with U6 snRNA as endogenous control. The results are expressed as mean ± SEM. Three independent experiments were performed for in vitro experiments and six for ex vivo experiments. **p <.01 vs Untreated cells; *p <.05 vs Untreated aortas. To analyse the up and down regulation of miR‐126‐3p and miR‐126‐5p, HUVEC were transfected with 20 nmol.L‐1 of premiR‐126‐3p, premiR‐126‐5p or inhibitors for 24 h. After total RNA extraction, miR‐126‐3p (D) and miR‐126‐5p (E) expression levels were analysed performing qRT‐PCR using U6 snRNA as endogenous control. The results are expressed with mean ± SEM. Three independent experiments were performed. **p <.01 vs SCL; *p <.05 vs SCL.