Figure 1: miR-712 is a flow-sensitive miRNA upregulated by d-flow in vitro and in vivo. | Nature Communications

Figure 1: miR-712 is a flow-sensitive miRNA upregulated by d-flow in vitro and in vivo.

From: The atypical mechanosensitive microRNA-712 derived from pre-ribosomal RNA induces endothelial inflammation and atherosclerosis

Figure 1

(a) The schema shows naturally occurring d-flow (lesser curvature, LC) and s-flow regions (greater curvature, GC) in the aortic arch. Also shown is the surgically induced d-flow in partial carotid ligation model in which three of the four caudal branches of the left common carotid artery (LCA) are ligated, while the contralateral right common carotid artery (RCA) remains untouched as an internal control. (b) Endothelial-enriched total RNAs obtained from intima of mouse (C57BL/6) left carotid (flow-disturbed LCA) and right carotid (contralateral control, RCA) at 48 h post ligation were analysed by gene array (Illumina BeadChip). Hierarchical clustering analyses of mechanosensitive miRNAs found in LCA endothelium compared with that of RCA are shown as heat maps. The colour represents the expression level of the gene. Red represents high expression, while green represents low expression. The expression levels are continuously mapped on the colour scale provided at the top of the figure. Each column represents a single sample pooled from three different LCAs or RCAs, and each row represents a single miRNA probe (n=3). (c–f) Validation of miRNA microarray results by qPCR. Quantitative PCR (qPCR), using additional independent RNA samples, was used to validate the above miRNA array data. Ten miRNAs (five up-, five downregulated miRNAs at 48 h post ligation) were selected based on fold change by flow. The qPCR study validated the microarray results for (c) five upregulated (miR-712, -330*, -699, -223 and 770-5p) and (d) five downregulated (miR-195, -30c, -29b, -26b and let-7d) miRNAs at the 48 h time point (n=5 each, data shown as mean±s.e.m.; *P<0.05 as determined by paired t-test). To further validate whether the mechanosensitive miRNAs that were identified in vivo responded specifically to shear stress, we tested expression of these miRNAs in vitro using iMAECs that were subjected to LS or OS, mimicking s-flow and d-flow in vitro, respectively14. Among the nine different miRNAs examined, seven miRNAs were differentially expressed under OS (n=6 each, data shown as mean±s.e.m.; *P<0.05 as determined by paired t-test) (e). These results showed that miR-712 was the most consistently and robustly upregulated miRNA both in vivo and under d-flow conditions in vitro.

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