KLF11 protects against abdominal aortic aneurysm through inhibition of endothelial cell dysfunction
Guizhen Zhao, Ziyi Chang, Yang Zhao, Yanhong Guo, Haocheng Lu, Wenying Liang, Oren Rom, Huilun Wang, Jinjian Sun, Tianqing Zhu, Yanbo Fan, Lin Chang, Bo Yang, Minerva T. Garcia-Barrio, Y. Eugene Chen, Jifeng Zhang
Guizhen Zhao, Ziyi Chang, Yang Zhao, Yanhong Guo, Haocheng Lu, Wenying Liang, Oren Rom, Huilun Wang, Jinjian Sun, Tianqing Zhu, Yanbo Fan, Lin Chang, Bo Yang, Minerva T. Garcia-Barrio, Y. Eugene Chen, Jifeng Zhang
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Research Article Vascular biology

KLF11 protects against abdominal aortic aneurysm through inhibition of endothelial cell dysfunction

Abstract

Abdominal aortic aneurysm (AAA) is a life-threatening degenerative vascular disease. Endothelial cell (EC) dysfunction is implicated in AAA. Our group recently demonstrated that Krüppel-like factor 11 (KLF11) plays an essential role in maintaining vascular homeostasis, at least partially through inhibition of EC inflammatory activation. However, the functions of endothelial KLF11 in AAA remain unknown. Here we found that endothelial KLF11 expression was reduced in the ECs from human aneurysms and was time dependently decreased in the aneurysmal endothelium from both elastase- and Pcsk9/AngII-induced AAA mouse models. KLF11 deficiency in ECs markedly aggravated AAA formation, whereas EC-selective KLF11 overexpression markedly inhibited AAA formation. Mechanistically, KLF11 not only inhibited the EC inflammatory response but also diminished MMP9 expression and activity and reduced NADPH oxidase 2–mediated production of reactive oxygen species in ECs. In addition, KLF11-deficient ECs induced smooth muscle cell dedifferentiation and apoptosis. Overall, we established endothelial KLF11 as a potentially novel factor protecting against AAA and a potential target for intervention in aortic aneurysms.

Authors

Guizhen Zhao, Ziyi Chang, Yang Zhao, Yanhong Guo, Haocheng Lu, Wenying Liang, Oren Rom, Huilun Wang, Jinjian Sun, Tianqing Zhu, Yanbo Fan, Lin Chang, Bo Yang, Minerva T. Garcia-Barrio, Y. Eugene Chen, Jifeng Zhang

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Figure 4

KLF11 inhibits MMP9 expression and activity in ECs.

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KLF11 inhibits MMP9 expression and activity in ECs. (A and B) Human aort...
(A and B) Human aortic ECs (HAECs) were infected with adenovirus carrying shRNA for KLF11 knockdown (Ad-shKLF11, 10 MOI, n = 4) or lacZ (Ad-shlacZ, n = 3) as control. After 48 hours, the total RNA was extracted for RNA sequencing. (A) The positive enrichment in the membrane protein proteolysis and metalloexopeptidase activity pathways is shown by gene set enrichment analysis plots (Ad-shKLF11 vs. Ad-shlacZ). NES, normalized enrichment score. (B) The differentially expressed genes (Ad-shKLF11 vs. Ad-shlacZ) are shown as volcano plots. Green dots, log2FoldChange < –0.5. Red dots, log2FoldChange > 0.5. Gray dots, –0.5 < log2FoldChange < 0.5. (C–F) HAECs were infected with Ad-shlacZ or Ad-shKLF11 or Ad-GFP or Ad-KLF11 (10 MOI). After 48 hours, they were treated with or without TNF-α (2 ng/mL) for 12 hours. (C and D) The mRNA levels of MMP2, MMP3, MMP9, MMP13, and MMP14 were determined by qPCR. (E and F) Representative gelatin zymography for the activity of MMP2 and MMP9 in the conditioned medium. Samples from E and F were run on 8% and 10% SDS-PAGE gels with 0.1% gelatin, respectively. (G) HAECs were infected with Ad-lacZ or Ad-flag-KLF11. After 48 hours, they were stimulated with TNF-α (2 ng/mL) for 4 hours. Chromatin immunoprecipitation (ChIP) assay was performed using an antibody against flag or IgG. (H) Luciferase reporter assay in HAECs transfected with an MMP9 promoter-driven luciferase reporter containing a KLF11 binding site and infected with Ad-lacZ or Ad-KLF11. After 48 hours, they were stimulated with TNF-α (2 ng/mL) for 4 hours. The luciferase activity was normalized against that of cotransfected Renilla. Data are presented as mean ± SEM. Two-way ANOVA followed by Holm-Sidak post hoc analysis (C, D, G, and H).