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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Ang II induces biological effects in human HSCs in a redox-sensitive man...
Ang II induces biological effects in human HSCs in a redox-sensitive manner. (a) Ang II (10–8 M) increases DNA synthesis. This effect was prevented by losartan (10–7 M), the ERK inhibitor PD98059 (5.10–6 M), the PI3K inhibitor LY294002 (2.10–6 M), NAC (10–5 M), and DPI (10–6 M), but not by the AT2 antagonist PD123319 (10–7 M). Results are expressed as mean ± SD of three independent experiments. *P < 0.01 vs. buffer; ‡P < 0.01 vs. buffer + Ang II. (b) Ang II increased the number of HSCs migrating through a modified Boyden chamber. Pretreatment with losartan, LY294002, NAC, and DPI inhibited this effect. Results are the mean ± SD from three independent experiments. *P < 0.01 vs. buffer; ‡P < 0.05 vs. buffer + Ang II. (c) In vitro wound-healing assay. Ang II induced migration of cells into the wound. This effect was inhibited by losartan and DPI. Images are representative of three independent experiments. (d) Ang II induces the secretion of proinflammatory chemokines. Ang II increased secretion of IL-8 (white bars) and MCP-1 (black bars). The base-line levels of MCP-1 and IL-8 were 545 ± 43 pg/ml and 119 ± 23 pg/ml, respectively. Pretreatment with losartan, NAC, and DPI attenuated this effect. Incubation with PD98059, LY294002, the p38 MAPK inhibitor SB203580 (10–6 M), and the JNK inhibitor SP600125 (20.10–6 M) attenuated cytokine secretion. Results are the mean ± SD from six independent experiments. *P < 0.05 vs. buffer; ‡P < 0.05 vs. buffer + Ang II.