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Expression of Nitric Oxide-Transporting Aquaporin-1 Is Controlled by KLF2 and Marks Non-Activated Endothelium In Vivo.

Fontijn RD, Volger OL, van der Pouw-Kraan TC, Doddaballapur A, Leyen T, Baggen JM, Boon RA, Horrevoets AJ - PLoS ONE (2015)

Bottom Line: Chromosome immunoprecipitation (CHIP) confirms binding of KLF2 to the AQP1 promoter.We conclude that AQP1 expression is subject to KLF2-mediated positive regulation by atheroprotective shear stress and is downregulated under inflammatory conditions both in vitro and in vivo.Thus, endothelial expression of AQP1 characterizes the atheroprotected, non-inflamed vessel wall.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology and Immunology, VU Medical Center, Amsterdam, the Netherlands.

ABSTRACT
The flow-responsive transcription factor Krüppel-like factor 2 (KLF2) maintains an anti-coagulant, anti-inflammatory endothelium with sufficient nitric oxide (NO)-bioavailability. In this study, we aimed to explore, both in vitro and in human vascular tissue, expression of the NO-transporting transmembrane pore aquaporin-1 (AQP1) and its regulation by atheroprotective KLF2 and atherogenic inflammatory stimuli. In silico analysis of gene expression profiles from studies that assessed the effects of KLF2 overexpression in vitro and atherosclerosis in vivo on endothelial cells, identifies AQP1 as KLF2 downstream gene with elevated expression in the plaque-free vessel wall. Biomechanical and pharmaceutical induction of KLF2 in vitro is accompanied by induction of AQP1. Chromosome immunoprecipitation (CHIP) confirms binding of KLF2 to the AQP1 promoter. Inflammatory stimulation of endothelial cells leads to repression of AQP1 transcription, which is restrained by KLF2 overexpression. Immunohistochemistry reveals expression of aquaporin-1 in non-activated endothelium overlying macrophage-poor intimae, irrespective whether these intimae are characterized as being plaque-free or as containing advanced plaque. We conclude that AQP1 expression is subject to KLF2-mediated positive regulation by atheroprotective shear stress and is downregulated under inflammatory conditions both in vitro and in vivo. Thus, endothelial expression of AQP1 characterizes the atheroprotected, non-inflamed vessel wall. Our data provide support for a continuous role of KLF2 in stabilizing the vessel wall via co-temporal expression of eNOS and AQP1 both preceding and during the pathogenesis of atherosclerosis.

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Related in: MedlinePlus

AQP1 immunohistochemistry in vascular tissue specimen showing different stages of atherosclerosis.Overviews, showing immunohistochemistry for macrophages (HAM56, left column), are given for arteries without lesions (A, external iliac artery), with focal lesions of the initial stage (B, abdominal aorta, intimal xanthoma/fatty streak) or the advanced stage (C, common iliac artery, fibro-calcific plaque with signs of rupture). Rectangles within the HAM56 overview indicate the position of areas that are shown as magnification of serial sections stained for ICAM-1 (middle column) and AQP1 (right column). In addition, these sections/areas were further characterized with regard to their cellular composition by staining for macrophages (HAM56) and smooth muscle cells (anti α-actin) (S3–S5 Figs).
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pone.0145777.g005: AQP1 immunohistochemistry in vascular tissue specimen showing different stages of atherosclerosis.Overviews, showing immunohistochemistry for macrophages (HAM56, left column), are given for arteries without lesions (A, external iliac artery), with focal lesions of the initial stage (B, abdominal aorta, intimal xanthoma/fatty streak) or the advanced stage (C, common iliac artery, fibro-calcific plaque with signs of rupture). Rectangles within the HAM56 overview indicate the position of areas that are shown as magnification of serial sections stained for ICAM-1 (middle column) and AQP1 (right column). In addition, these sections/areas were further characterized with regard to their cellular composition by staining for macrophages (HAM56) and smooth muscle cells (anti α-actin) (S3–S5 Figs).

Mentions: Fig 5A shows a section of an external iliac artery without histological signs of atherosclerosis and negative for macrophage staining with HAM56 antibody. A homogenous AQP-1 immunostain was observed on endothelial cells lining the lumen. This observation is in agreement with microarray mRNA data for AQP1 that identify AQP1 as signature gene for the plaque-free situation (Fig 1A). Furthermore, AQP1 expression was abundant on endothelial cells lining capillaries within adventitial layers (Fig 5A).


Expression of Nitric Oxide-Transporting Aquaporin-1 Is Controlled by KLF2 and Marks Non-Activated Endothelium In Vivo.

Fontijn RD, Volger OL, van der Pouw-Kraan TC, Doddaballapur A, Leyen T, Baggen JM, Boon RA, Horrevoets AJ - PLoS ONE (2015)

AQP1 immunohistochemistry in vascular tissue specimen showing different stages of atherosclerosis.Overviews, showing immunohistochemistry for macrophages (HAM56, left column), are given for arteries without lesions (A, external iliac artery), with focal lesions of the initial stage (B, abdominal aorta, intimal xanthoma/fatty streak) or the advanced stage (C, common iliac artery, fibro-calcific plaque with signs of rupture). Rectangles within the HAM56 overview indicate the position of areas that are shown as magnification of serial sections stained for ICAM-1 (middle column) and AQP1 (right column). In addition, these sections/areas were further characterized with regard to their cellular composition by staining for macrophages (HAM56) and smooth muscle cells (anti α-actin) (S3–S5 Figs).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4696733&req=5

pone.0145777.g005: AQP1 immunohistochemistry in vascular tissue specimen showing different stages of atherosclerosis.Overviews, showing immunohistochemistry for macrophages (HAM56, left column), are given for arteries without lesions (A, external iliac artery), with focal lesions of the initial stage (B, abdominal aorta, intimal xanthoma/fatty streak) or the advanced stage (C, common iliac artery, fibro-calcific plaque with signs of rupture). Rectangles within the HAM56 overview indicate the position of areas that are shown as magnification of serial sections stained for ICAM-1 (middle column) and AQP1 (right column). In addition, these sections/areas were further characterized with regard to their cellular composition by staining for macrophages (HAM56) and smooth muscle cells (anti α-actin) (S3–S5 Figs).
Mentions: Fig 5A shows a section of an external iliac artery without histological signs of atherosclerosis and negative for macrophage staining with HAM56 antibody. A homogenous AQP-1 immunostain was observed on endothelial cells lining the lumen. This observation is in agreement with microarray mRNA data for AQP1 that identify AQP1 as signature gene for the plaque-free situation (Fig 1A). Furthermore, AQP1 expression was abundant on endothelial cells lining capillaries within adventitial layers (Fig 5A).

Bottom Line: Chromosome immunoprecipitation (CHIP) confirms binding of KLF2 to the AQP1 promoter.We conclude that AQP1 expression is subject to KLF2-mediated positive regulation by atheroprotective shear stress and is downregulated under inflammatory conditions both in vitro and in vivo.Thus, endothelial expression of AQP1 characterizes the atheroprotected, non-inflamed vessel wall.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Cell Biology and Immunology, VU Medical Center, Amsterdam, the Netherlands.

ABSTRACT
The flow-responsive transcription factor Krüppel-like factor 2 (KLF2) maintains an anti-coagulant, anti-inflammatory endothelium with sufficient nitric oxide (NO)-bioavailability. In this study, we aimed to explore, both in vitro and in human vascular tissue, expression of the NO-transporting transmembrane pore aquaporin-1 (AQP1) and its regulation by atheroprotective KLF2 and atherogenic inflammatory stimuli. In silico analysis of gene expression profiles from studies that assessed the effects of KLF2 overexpression in vitro and atherosclerosis in vivo on endothelial cells, identifies AQP1 as KLF2 downstream gene with elevated expression in the plaque-free vessel wall. Biomechanical and pharmaceutical induction of KLF2 in vitro is accompanied by induction of AQP1. Chromosome immunoprecipitation (CHIP) confirms binding of KLF2 to the AQP1 promoter. Inflammatory stimulation of endothelial cells leads to repression of AQP1 transcription, which is restrained by KLF2 overexpression. Immunohistochemistry reveals expression of aquaporin-1 in non-activated endothelium overlying macrophage-poor intimae, irrespective whether these intimae are characterized as being plaque-free or as containing advanced plaque. We conclude that AQP1 expression is subject to KLF2-mediated positive regulation by atheroprotective shear stress and is downregulated under inflammatory conditions both in vitro and in vivo. Thus, endothelial expression of AQP1 characterizes the atheroprotected, non-inflamed vessel wall. Our data provide support for a continuous role of KLF2 in stabilizing the vessel wall via co-temporal expression of eNOS and AQP1 both preceding and during the pathogenesis of atherosclerosis.

Show MeSH
Related in: MedlinePlus