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. 2021 Jun 4:12:665994.
doi: 10.3389/fphys.2021.665994. eCollection 2021.

High Glucose Activates YAP Signaling to Promote Vascular Inflammation

Affiliations

High Glucose Activates YAP Signaling to Promote Vascular Inflammation

Jeremy Ortillon et al. Front Physiol. .

Abstract

Background and aims: The YAP/TAZ signaling is known to regulate endothelial activation and vascular inflammation in response to shear stress. Moreover, YAP/TAZ signaling plays a role in the progression of cancers and renal damage associated with diabetes. However, whether YAP/TAZ signaling is also implicated in diabetes-associated vascular complications is not known.

Methods: The effect of high glucose on YAP/TAZ signaling was firstly evaluated in vitro on endothelial cells cultured under static conditions or subjected to shear stress (either laminar or oscillatory flow). The impact of diabetes on YAP/TAZ signaling was additionally assessed in vivo in db/db mice.

Results: In vitro, we found that YAP was dephosphorylated/activated by high glucose in endothelial cells, thus leading to increased endothelial inflammation and monocyte attachment. Moreover, YAP was further activated when high glucose was combined to laminar flow conditions. YAP was also activated by oscillatory flow conditions but, in contrast, high glucose did not exert any additional effect. Interestingly, inhibition of YAP reduced endothelial inflammation and monocyte attachment. Finally, we found that YAP is also activated in the vascular wall of diabetic mice, where inflammatory markers are also increased.

Conclusion: With the current study we demonstrated that YAP signaling is activated by high glucose in endothelial cells in vitro and in the vasculature of diabetic mice, and we pinpointed YAP as a regulator of high glucose-mediated endothelial inflammation and monocyte attachment. YAP inhibition may represent a potential therapeutic opportunity to improve diabetes-associated vascular complications.

Keywords: YAP/TAZ; diabetes; endothelial cells; inflammation; vascular complications.

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Conflict of interest statement

All authors are Sanofi employees. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Effect of high glucose on YAP activation in endothelial cells. Representative protein expression blots and bar graphs of phospho-YAP (p-YAP; S127) and total YAP (A), phospho-TAZ (p-TAZ; S89) and total TAZ (B), CTGF and CYR61 (C) in HUVECs cultured under static conditions with different concentrations of glucose for 24 h. The control images of GAPDH are re-used for illustrative purposes. Data are presented as the mean ± SEM. N = 5/group for panels A and B; N = 7/group for panel C. *p < 0.05 between the indicated groups. Two-tailed Student’s t test was used for comparisons between the groups.
FIGURE 2
FIGURE 2
YAP mediates glucose-induced endothelial activation. Activity of TEAD-luciferase in TeloHAECs cultured under either normal or high glucose (A). Representative protein expression blots (B) and bar graphs showing the expression of VCAM-1 (C) and CTGF (D) in HUVECs cultured under static conditions with either normal or high glucose and additionally treated with siRNA for YAP/TAZ. Representative protein expression blots and bar graphs showing the expression of VCAM-1 and ICAM-1 (E) as well as their percentage (%) and cell surface expression (represented as mean of fluorescence – MFI) assessed by flow cytometry (F) in HUVECs cultured under static conditions with either normal or high glucose for 24 h and in the presence, or not, of the YAP/TEAD inhibitor K-975 at 200 nM. DMSO was used as control. Data are presented as the mean ± SEM. N = 3/group for panel A; N = 4/group for panel C; N = 6/group for panel D; N = 6–7/group for panel E; N = 5/group for panel F. p < 0.05 and ∗∗p < 0.01 between the indicated groups. Two-tailed Mann-Whitney test was used for comparisons between the groups.
FIGURE 3
FIGURE 3
Effects of high glucose and shear stress on YAP activation in endothelial cells. (A) Representative protein expression blots and bar graphs of phospho and total YAP, phospho and total TAZ in HUVECs cultured under either normal or high glucose and subjected to either laminar flow (LF, 12 dyn/cm2) or oscillatory flow (OF, 0.5 ± 6 dyn/cm2; 1 Hz) for 72 h. (B) Representative protein expression blots of VCAM-1 and CYR61 and bar graphs of VCAM-1 in HUVECs cultured under either normal or high glucose and subjected to LF or OF for 72 h. (C) Representative microphotographs and bar graphs showing THP-1 monocyte attachments to HUVECs cultured under either normal or high glucose, subjected to LF for 72 h and additionally treated with the YAP/TEAD inhibitor K-975 (200 nM). DMSO was used as control for K-975. (D) Representative microphotographs and bar graphs showing THP-1 monocyte attachments to HUVECs cultured under either normal or high glucose and subjected to either LF or OF for 72 h. THP-1 monocytes are GFP positive (green fluorescence). Nuclei are stained by DAPI (blue fluorescence). Data are presented as the mean ± SEM. N = 4–11/group for panel A; N = 6–15/group for panel B; N = 4–5/group for panel C and N = 5–6/group for panel D. p < 0.05 and ∗∗p < 0.01 between the indicated groups. Two-tailed Mann-Whitney test (A,C,D) or two-tailed Student’s t test (B) was used for comparisons between the groups.
FIGURE 4
FIGURE 4
YAP is activated in the vascular wall of diabetic mice. Representative microphotographs (×40 magnification) showing cross sections of either carotid arteries without cast (A) or cast-instrumented carotid arteries (B) isolated from db/ + or db/db mice and stained for p-YAP, total YAP and VCAM-1. Inserts at the lower left side show a detail of the arterial wall at higher magnification (×100), with the intima-media thickness indicated by an arrow. Bar graphs show the quantification of the intima-media area (C), and the expression of p-YAP (D), total YAP (E), the ratio p-YAP/YAP (F) and VCAM-1 (G). Data are presented as the mean ± SEM. N = 5–6/group. p < 0.05, ∗∗p < 0.01 between the indicated groups. Kruskal-Wallis test was used for comparisons.

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