Transforming growth factor β1-induced collagen production in myofibroblasts is mediated by reactive oxygen species derived from NADPH oxidase 4

Intestinal fibrosis with stricture formation is a severe complication of Crohn's disease (CD). Though new therapeutic targets to enable the prevention or treatment of intestinal fibrosis are needed, markers of this condition and the basic mechanisms responsible have not been established. NADPH...

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Published in:Biochemical and biophysical research communications 2018-11, Vol.506 (3), p.557-562
Main Authors: Hotta, Yuma, Uchiyama, Kazuhiko, Takagi, Tomohisa, Kashiwagi, Saori, Nakano, Takahiro, Mukai, Rieko, Toyokawa, Yuki, Yasuda, Tomoyo, Ueda, Tomohiro, Suyama, Yosuke, Murakami, Takaaki, Tanaka, Makoto, Majima, Atsushi, Doi, Toshifumi, Hirai, Yasuko, Mizushima, Katsura, Morita, Mayuko, Higashimura, Yasuki, Inoue, Ken, Fukui, Akifumi, Okayama, Tetsuya, Katada, Kazuhiro, Kamada, Kazuhiro, Handa, Osamu, Ishikawa, Takeshi, Naito, Yuji, Itoh, Yoshito
Format: Article
Language:English
Subjects:
60 APPLIED LIFE SCIENCES
Animals
Cell Line
COLLAGEN
Collagen - biosynthesis
Crohn's disease
FIBROSIS
GROWTH FACTORS
INFLAMMATION
Inflammatory bowel disease
LUNGS
Mice
Myofibroblasts - drug effects
Myofibroblasts - metabolism
NADPH oxidase 4
NADPH Oxidase 4 - genetics
NADPH Oxidase 4 - metabolism
OXIDASES
Reactive oxygen species
Reactive Oxygen Species - metabolism
RNA
RNA, Messenger - genetics
RNA, Messenger - metabolism
Transforming Growth Factor beta1 - pharmacology
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Summary:Intestinal fibrosis with stricture formation is a severe complication of Crohn's disease (CD). Though new therapeutic targets to enable the prevention or treatment of intestinal fibrosis are needed, markers of this condition and the basic mechanisms responsible have not been established. NADPH oxidase (NOX) 4 has already been reported to play a key role in models of fibrogenesis, including that of the lung. However, its importance in intestinal fibrogenesis remains unclear. In this study, we examined the role of NOX4 in collagen production by intestinal myofibroblasts stimulated with transforming growth factor (TGF)-β1. Using LmcMF cells, an intestinal subepithelial myofibroblast (ISEMF) line, we first examined the induction of collagen production by TGF-β1. Subsequently, we investigated the role of NOX4 in TGF-β1-induced collagen I production in these cells using SB525334 (an SMAD2/3 inhibitor), diphenyleneiodonium (an NOX inhibitor), and Nox4 small interfering RNA (siRNA). Production of collagen was assessed with Sirius red staining, and Nox4 expression was measured by quantitative real-time PCR. Reactive oxygen species (ROS) production was determined using DCFDA and fluorescent microscopy. We observed that TGF-β1 induced collagen production via NOX4 activation and ROS generation in LmcMF cells. Nox4 siRNA and inhibitors of TGF-β1 receptor and NOX significantly reduced TGF-β1-induced ROS and collagen production. Thus, in the present study, we revealed that collagen production in ISEMFs is induced via an NOX4-dependent pathway. This work supports a function for NOX4 in intestinal fibrogenesis and identifies it as a potential therapeutic target in recalcitrant fibrotic disorders of CD patients. •Intestinal fibrosis with stricture is severe complication of Crohn's disease.•TGF-β1-induced collagen production in intestinal myofibroblast is dependent on NOX4.•NOX inhibitor significantly diminished TGF-β1-induced ROS and collagen production.•NOX4 may constitute a therapeutic target in fibrotic disorders among Crohn's disease.
ISSN:0006-291X
1090-2104
DOI:10.1016/j.bbrc.2018.10.116