Pirfenidone inhibits TGF-β1-induced over-expression of collagen type I and heat shock protein 47 in A549 cells

doi:10.1186/1471-2466-12-24

BMC Pulm Med. 2012; 12: 24.

Published online 2012 June 13. doi: 10.1186/1471-2466-12-24

PMCID: PMC3403980

Pirfenidone inhibits TGF-β1-induced over-expression of collagen type I and heat shock protein 47 in A549 cells

Keiko Hisatomi,¹ Hiroshi Mukae,² Noriho Sakamoto,¹ Yuji Ishimatsu,¹ Tomoyuki Kakugawa,¹ Shintaro Hara,¹ Hanako Fujita,¹ Seiko Nakamichi,³ Hisashi Oku,⁴ Yoshishige Urata,⁵ Hiroshi Kubota,⁶ Kazuhiro Nagata,⁶ and Shigeru Kohno¹

¹Second Department of Internal Medicine, Nagasaki University School of Medicine, 1-7-1 Sakamoto, Nagasaki, 852-8501, Japan

²Department of Respiratory Medicine, University of Occupational and Environmental Health, Kitakyushu, Japan

³Department of General Medicine, Nagasaki University Hospital, Nagasaki, Japan

⁴Discovery Research Laboratories, Shionogi & Co. Ltd, Osaka, Japan

⁵Department of Stem Cell Biology, Nagasaki University Graduate of Biomedical Science, Nagasaki, Japan

⁶Department of Molecular and Cellular Biology, Institute for Frontier Medical Science, Kyoto University, Kyoto, Japan

Corresponding author.

Keiko Hisatomi: keiko_hisatomi/at/yahoo.co.jp; Hiroshi Mukae: hmukae/at/med.uoeh-u.ac.jp; Noriho Sakamoto: nsakamot/at/nagasaki-u.ac.jp; Yuji Ishimatsu: yuji-i/at/nagasaki-u.ac.jp; Tomoyuki Kakugawa: kakugawa/at/nagasaki-u.ac.jp; Shintaro Hara: hara_shin123/at/me.com; Hanako Fujita: hanakofu0925/at/yahoo.co.jp; Seiko Nakamichi: seiko-n/at/nagasaki-u.ac.jp; Hisashi Oku: hisashi.oku/at/shionogi.co.jp; Yoshishige Urata: urata/at/net.nagasaki-u.ac.jp; Hiroshi Kubota: hkubota/at/ipc.akita-u.ac.jp; Kazuhiro Nagata: nagata/at/frontier.kyoto-u.ac.jp; Shigeru Kohno: s-kohno/at/nagasaki-u.ac.jp

Received December 20, 2011; Accepted June 13, 2012.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background

Pirfenidone is a novel anti-fibrotic and anti-inflammatory agent that inhibits the progression of fibrosis in animal models and in patients with idiopathic pulmonary fibrosis (IPF). We previously showed that pirfenidone inhibits the over-expression of collagen type I and of heat shock protein (HSP) 47, a collagen-specific molecular chaperone, in human lung fibroblasts stimulated with transforming growth factor (TGF)-β1 in vitro. The increased numbers of HSP47-positive type II pneumocytes as well as fibroblasts were also diminished by pirfenidone in an animal model of pulmonary fibrosis induced by bleomycin. The present study evaluates the effects of pirfenidone on collagen type I and HSP47 expression in the human alveolar epithelial cell line, A549 cells in vitro.

Methods

The expression of collagen type I, HSP47 and E-cadherin mRNAs in A549 cells stimulated with TGF-β1 was evaluated by Northern blotting or real-time PCR. The expression of collagen type I, HSP47 and fibronectin proteins was assessed by immunocytochemical staining.

Results

TGF-β1 stimulated collagen type I and HSP47 mRNA and protein expression in A549 cells, and pirfenidone significantly inhibited this process. Pirfenidone also inhibited over-expression of the fibroblast phenotypic marker fibronectin in A549 cells induced by TGF-β1.

Conclusion

We concluded that the anti-fibrotic effects of pirfenidone might be mediated not only through the direct inhibition of collagen type I expression but also through the inhibition of HSP47 expression in alveolar epithelial cells, which results in reduced collagen synthesis in lung fibrosis. Furthermore, pirfenidone might partially inhibit the epithelial-mesenchymal transition.

Keywords: Pneumocyte, Interstitial pneumonia, Epithelial cell, Epithelial mesenchymal transition, Pulmonary fibrosis

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