Figure 1
From: Shear stress induces endothelial-to-mesenchymal transition via the transcription factor Snail
Low shear stress induced mesenchymal genes via Snail. (a) HUVEC were exposed to low oscillatory (+/−4 dyn/cm2) or high (13 dyn/cm2) wall shear stress (WSS) using a parallel plate system. (b) PAEC were exposed to orbital flow to generate low (5 dyn/cm2) or high (15 dyn/cm2) wall shear stress. (a,b) After 72 h, levels of EndMT marker transcripts and VE-cadherin transcripts were quantified by qRT-PCR. The expression level at the low WSS site is presented relative to the expression at the high WSS site (normalised to 1; dotted line). Data were pooled from six independent experiments using cells from different donors and mean levels +/− SEM are shown. (c–e) HUVEC were exposed to orbital flow to generate low (5 dyn/cm2) or high (15 dyn/cm2) WSS for 72 h. (c) Expression of N-cadherin (green) and VE-cadherin (red) was determined by immunofluorescent staining and co-staining using DAPI (blue). Scale bar, 50 μm. The proportion of cells that expressed N-cadherin or VE-cadherin was measured. (d) The expression levels of N-cadherin (left) and VE-cadherin (right) were assessed by Western blotting using specific antibodies and anti-PDHX antibodies were used to control for total protein levels. Representative blots are shown. Bands were quantified by densitometry. (e) Expression of Snail (green) was determined by immunofluorescent staining and co-staining using DAPI (blue). Scale bar, 50 μm. Fluorescence intensity was quantified in multiple cells. (f) HUVEC were transfected with siRNA targeting Snail or with scrambled sequences and exposed to orbital flow for 72 h. Cells exposed to low WSS (5 dyn/cm2) were collected and transcript levels of Slug, N cadherin and α-SMA were quantified by qRT-PCR. (c–f) Data were pooled from three independent experiments using cells from different donors and mean levels +/− SEM are shown.
