Arsenic-stimulated liver sinusoidal capillarization in mice requires NADPH oxidase–generated superoxide
Adam C. Straub, … , Donna B. Stolz, Aaron Barchowsky
Adam C. Straub, … , Donna B. Stolz, Aaron Barchowsky
Published November 13, 2008
Citation Information: J Clin Invest. 2008;118(12):3980-3989. https://doi.org/10.1172/JCI35092.
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Research Article Hepatology

Arsenic-stimulated liver sinusoidal capillarization in mice requires NADPH oxidase–generated superoxide

Abstract

Environmental arsenic exposure, through drinking contaminated water, is a significant risk factor for developing vascular diseases and is associated with liver portal hypertension, vascular shunting, and portal fibrosis through unknown mechanisms. We found that the addition of low doses of arsenite to the drinking water of mice resulted in marked pathologic remodeling in liver sinusoidal endothelial cells (SECs), including SEC defenestration, capillarization, increased junctional PECAM-1 expression, protein nitration, and decreased liver clearance of modified albumin. Furthermore, the pathologic changes observed after in vivo exposure were recapitulated in isolated mouse SECs exposed to arsenic in culture. To investigate the role of NADPH oxidase–generated ROS in this remodeling, we examined the effect of arsenite in the drinking water of mice deficient for the p47 subunit of the NADPH oxidase and found that knockout mice were protected from arsenite-induced capillarization and protein nitration. Furthermore, ex vivo arsenic exposure increased SEC superoxide generation, and this effect was inhibited by addition of a Nox2 inhibitor and quenched by the cell-permeant superoxide scavenger. In addition, inhibiting either oxidant generation or Rac1-GTPase blocked ex vivo arsenic-stimulated SEC differentiation and dysfunction. Our data indicate that a Nox2-based oxidase is required for SEC capillarization and that it may play a central role in vessel remodeling following environmentally relevant arsenic exposures.

Authors

Adam C. Straub, Katherine A. Clark, Mark A. Ross, Ashwin G. Chandra, Song Li, Xiang Gao, Patrick J. Pagano, Donna B. Stolz, Aaron Barchowsky

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Figure 3

Arsenic inhibits SEC scavenging of acylated protein.

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Arsenic inhibits SEC scavenging of acylated protein. (A) Mice were injec...
(A) Mice were injected in the tail vein with 150 mg/kg FITC-labeled succinylated BSA (suc-BSA) or BSA (both green) in 200 μl saline. After 10 min, the mice were euthanized, and livers were excised, fixed in 4% paraformaldehyde, and sectioned for confocal microscopic analysis. Sections were stained with rhodamine-conjugated antibody to PECAM-1 (red) and DRAQ5 (blue). Images are representative of sections from 3 mice in each group. LV, large vessel; S, sinusoid vessel. Scale bar: 10 μm. (B) FITC-labeled acetylated LDL and biotin-labeled succinylated BSA (3 ml; 150 mg/ml saline) were infused over 3 min into the vena cavas of untreated mice and mice exposed to 100 ppb arsenic (As) for 2 wk. Livers were then excised, frozen in liquid N2, and sectioned, and total protein was extracted for assay of retained biotin label by immunoblotting, as described in Methods. Data are mean ± SD band density of biotin-labeled succinylated BSA normalized to β-actin (n = 3). NT, not treated. (C) Isolated SECs, incubated in the absence or presence of 1 mM tempol for 10 min, were left untreated or exposed to 2.5 μM arsenite for 24 h. Biotin-labeled succinylated BSA (20 μg/ml) was then added for 10 min, and after rinsing, total proteins were extracted for Western analysis (n = 3). Data are presented as in B. *P < 0.05, **P < 0.01 versus control; #P < 0.05 versus arsenic.