NADPH oxidase signal transduces angiotensin II in hepatic stellate cells and is critical in hepatic fibrosis
Ramón Bataller, … , John J. Lemasters, David A. Brenner
Ramón Bataller, … , John J. Lemasters, David A. Brenner
Published November 1, 2003
Citation Information: J Clin Invest. 2003;112(9):1383-1394. https://doi.org/10.1172/JCI18212.
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Article Hepatology

NADPH oxidase signal transduces angiotensin II in hepatic stellate cells and is critical in hepatic fibrosis

Abstract

Angiotensin II (Ang II) is a pro-oxidant and fibrogenic cytokine. We investigated the role of NADPH oxidase in Ang II–induced effects in hepatic stellate cells (HSCs), a fibrogenic cell type. Human HSCs express mRNAs of key components of nonphagocytic NADPH oxidase. Ang II phosphorylated p47phox, a regulatory subunit of NADPH oxidase, and induced reactive oxygen species formation via NADPH oxidase activity. Ang II phosphorylated AKT and MAPKs and increased AP-1 DNA binding in a redox-sensitive manner. Ang II stimulated DNA synthesis, cell migration, procollagen α1(I) mRNA expression, and secretion of TGF-β1 and inflammatory cytokines. These effects were attenuated by N-acetylcysteine and diphenylene iodonium, an NADPH oxidase inhibitor. Moreover, Ang II induced upregulation of genes potentially involved in hepatic wound-healing response in a redox-sensitive manner, as assessed by microarray analysis. HSCs isolated from p47phox–/– mice displayed a blunted response to Ang II compared with WT cells. We also assessed the role of NADPH oxidase in experimental liver fibrosis. After bile duct ligation, p47phox–/– mice showed attenuated liver injury and fibrosis compared with WT counterparts. Moreover, expression of smooth muscle α-actin and expression of TGF-β1 were reduced in p47phox–/– mice. Thus, NADPH oxidase mediates the actions of Ang II on HSCs and plays a critical role in liver fibrogenesis.

Authors

Ramón Bataller, Robert F. Schwabe, Youkyung H. Choi, Liu Yang, Yong Han Paik, Jeffrey Lindquist, Ting Qian, Robert Schoonhoven, Curt H. Hagedorn, John J. Lemasters, David A. Brenner

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Figure 5

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Effects of Ang II in WT and p47phox–/– mouse HSCs. HSCs were isolated fr...
Effects of Ang II in WT and p47phox–/– mouse HSCs. HSCs were isolated from C57BL/6 WT and p47phox–/– mice. Cells were studied after 14 days in culture (activated phenotype). (a) ROS production was estimated in DCFDA-loaded mouse HSCs by a fluorometer at 485/535 nm. Ang II (10–8 M) increased intracellular ROS in WT HSCs (open triangles), while the increase in ROS was attenuated in p47phox–/– HSCs (filled circles). Cells exposed to buffer did not show any increase in cell fluorescence (WT, filled triangles; p47phox–/–, open circles). (b and c) Ang II (10–8 M) increases DNA synthesis (b) and cell migration (c) in WT mouse HSCs, as assessed by 3H-thymidine uptake and an in vitro migration assay in a modified Boyden chamber, respectively. These effects were markedly attenuated in p47phox–/– HSCs. *P < 0.05, WT buffer vs. WTAng II. ‡P < 0.05, WT Ang II vs. KO. (d) Effect of Ang II on phosphorylation of ERK and c-Jun, as assessed by Western blot analysis. Ang II (10–8 M) stimulates ERK and c-Jun phosphorylation in WT mouse HSCs, while both effects were markedly blunted in p47phox–/– HSCs. Results are representative of three independent experiments.