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Receptor-activated Smad localisation in Bleomycin-induced pulmonary fibrosis
  1. Hiroyuki Higashiyama,
  2. Daisuke Yoshimoto,
  3. Yuji Okamoto,
  4. Hideo Kikkawa,
  5. Satoshi Asano,
  6. Mine Kinoshita
  1. Department of Pharmacology, Tsukuba Research Laboratories, GlaxoSmithKline, Tsukuba, Ibaraki, Japan
  1. Correspondence to:
 Dr S Asano
 Department of Pharmacology, Tsukuba Research Laboratories, GlaxoSmithKline, 43 Wadai, Tsukuba, Ibaraki 300-4247, Japan;satoshi.asano{at}


Background: Recent advances in fibrosis biology have identified transforming growth factor (TGF)-β type I receptor-mediated activation of Smads as playing a central part in the development of fibrosis. However, to date, there have been few studies that examined the localisation and distribution of receptor-activated Smads protein (R-Smads: Smad2 and 3) during the fibrosis progression.

Aims: To histopathologically assess the time-course change of the localisation and distribution of the Smads protein in pulmonary fibrosis.

Methods: Pulmonary fibrosis was induced by intranasal injection of bleomycin (0.3 U/mouse). Lungs were isolated 2, 5, 7, 9 and 14 days after bleomycin treatment. Histological changes in the lungs were evaluated by haematoxylin-eosin stain or Masson’s trichrome stain, and scored. TGF-β1, Smad3 and phosphorylated Smad2 localisations in lung tissues were determined by immunohistochemistry.

Results: The bleomycin treatment led to considerable pulmonary fibrotic changes accompanied by marked increase in TGF-β1 expression in infiltrating macrophages. With the progression in fibrosis (day 7–14), marked increases in Smad3-positive and pSmad2-positive cells were observed. There were intense Smad3-positive and pSmad2-positive signals localised to the nuclei of the infiltrating macrophages and to type II epithelial cells, and less intense signals in fibroblasts and hyperplastic alveolar/bronchiolar epithelial cells.

Conclusions: The time-course data of TGF-β1 and R-Smads indicate that progressive enhancement of TGF-β1 signalling via R-Smad is activated in the process of fibrosis progression.

  • ECM, extracellular matrix
  • TGF, transforming growth factor

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  • Published Online First 2 June 2006

  • Competing interests: None.