Oxidative stress alters syndecan-1 distribution in lungs with pulmonary fibrosis

Our plain-English summary of Oxidative stress alters syndecan-1 distribution in lungs with pulmonary fibrosis, by C.R. Kliment, J.M. Englert, B.R. Gochuico, et al (The Journal of Biological Chemistry, 2009).

The study

• Authors: C.R. Kliment, J.M. Englert, B.R. Gochuico, G. Yu, N. Kaminski, I. Rosas, T.D. Oury
• Journal: The Journal of Biological Chemistry
• Published: 2009;284(6):3537-45
• Cited by: 107 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Idiopathic pulmonary fibrosis (IPF) is an interstitial lung disease characterized by severe, progressive fibrosis. Roles for inflammation and oxidative stress have recently been demonstrated, but despite advances in understanding the pathogenesis, there are still no effective therapies for IPF. This study investigates how extracellular superoxide dismutase (EC-SOD), a syndecan-binding antioxidant enzyme, inhibits inflammation and lung fibrosis. We hypothesize that EC-SOD protects the lung from oxidant damage by preventing syndecan fragmentation/shedding. Wild-type or EC-SOD-null mice were exposed to an intratracheal instillation of asbestos or bleomycin. Western blot was used to detect syndecans in the bronchoalveolar lavage fluid and lung. Human lung samples (normal and IPF) were also analyzed. Immunohistochemistry for syndecan-1 and EC-SOD was performed on human and mouse lungs. In vitro, alveolar epithelial cells were exposed to oxidative stress and EC-SOD. Cell supernatants were analyzed for shed syndecan-1 by Western blot. Syndecan-1 ectodomain was assessed in wound healing and neutrophil chemotaxis. Increases in human syndecan-1 are detected in lung homogenates and lavage fluid of IPF lungs. Syndecan-1 is also significantly elevated in the lavage fluid of EC-SOD-null mice after asbestos and bleomycin exposure. On IHC, syndecan-1 staining increases within fibrotic areas of human and mouse lungs. In vitro, EC-SOD inhibits oxidant-induced loss of syndecan-1 from A549 cells. Shed and exogenous syndecan-1 ectodomain induce neutrophil chemotaxis, inhibit alveolar epithelial wound healing, and promote fibrogenesis. Oxidative shedding of syndecan-1 is an underlying cause of neutrophil chemotaxis and aberrant wound healing that may contribute to pulmonary fibrosis.

In plain English

This laboratory study examined how oxidative stress affects syndecan-1, a lung cell-surface protein, in pulmonary fibrosis (lung scarring). The authors caused lung injury in normal mice and in mice lacking the antioxidant enzyme EC-SOD. They used crocidolite asbestos (0.1 mg into the windpipe), inert titanium dioxide as a control, or bleomycin. In normal mice, syndecan-1 shed into lung fluid rose 1, 14 and 28 days after asbestos. Levels were further increased in mice lacking EC-SOD at days 1 and 28, and were significantly higher 7 days after bleomycin. In mice lacking EC-SOD, giving EC-SOD with asbestos lowered neutrophils (a type of white blood cell) and shed syndecan-1 in lung fluid after 24 hours. Lung fluid and tissue from patients with idiopathic pulmonary fibrosis had significantly more syndecan-1 than samples from control subjects. In cell experiments, oxidants raised shed syndecan-1 from 9.2 to 42.3 ng/ml; EC-SOD reduced this shedding. The shed protein attracted neutrophils. A lab-made version also attracted neutrophils, reduced healing of scratched lung-cell layers and increased lung fibroblast growth. The authors concluded that shed syndecan-1 contributes to pulmonary fibrosis.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory study combining animal experiments (mice given asbestos or bleomycin into the lungs), analysis of human lung fluid and tissue samples, and cell-culture experiments
• Who was studied: Normal (wild-type C57BL/6) mice and mice lacking EC-SOD. Lung lavage fluid and lung tissue from patients with idiopathic pulmonary fibrosis (IPF) and from people with normal lungs, obtained at the NIH Clinical Center (Bethesda, MD) or the University of Pittsburgh. Cultured cells: human A549 alveolar epithelial cells, LL47 human lung fibroblasts, primary mouse alveolar epithelial cells, and human neutrophils. Figure legends give group sizes of about 4 to 5.
• Exposure: Mice received 0.1 mg crocidolite asbestos or titanium dioxide (inert control particle) into the trachea and were studied at 1, 14 and 28 days. Other mice received bleomycin and were studied at 7 days. Cells were exposed to laboratory-generated reactive oxygen species (oxidants), with or without EC-SOD, and to a lab-made syndecan-1 ectodomain protein.
• Main results:
  • In wild-type mice, shed syndecan-1 in lung lavage fluid increased at 1, 14 and 28 days after asbestos.
  • In mice lacking EC-SOD, shed syndecan-1 after asbestos was significantly further increased at days 1 and 28.
  • Seven days after bleomycin, shed syndecan-1 in lavage fluid was significantly higher in mice lacking EC-SOD than in wild-type mice.
  • In mice lacking EC-SOD, 24 hours after co-treatment with asbestos and 50 units of EC-SOD, lavage fluid showed a significant decrease in neutrophils and a decrease in shed syndecan-1.
  • In patients with IPF, syndecan-1 was significantly increased in lavage fluid and lung tissue compared with control subjects.
  • On tissue staining, fibrotic areas of IPF lungs showed increased syndecan-1 and decreased interstitial EC-SOD.
  • In wild-type mice 14 days after asbestos, EC-SOD gene expression was 76.32% of the level in titanium dioxide-treated lungs (set at 100%).
  • In A549 cells, EC-SOD significantly protected against oxidant-induced syndecan-1 shedding. Shed syndecan-1 measured by ELISA was 9.2 ng/ml after medium versus 42.3 ng/ml after oxidant treatment.
  • Fluid from oxidant-treated A549 cells attracted neutrophils: migration index 1.70 versus 1.00 for the control solution. Knocking down syndecan-1 in the cells reduced the index to 1.12.
  • The lab-made syndecan-1 ectodomain alone attracted human neutrophils: migration index 4.42 versus 1.0 for the control solution.
  • In primary mouse alveolar epithelial cells, wound healing after 20 hours was 65.0% with medium alone versus 51.6% with the lab-made syndecan-1 ectodomain.
  • In A549 cells, knocking down syndecan-1 reduced wound healing from 80.1% (control siRNA) to 49.3%.
  • In lung fibroblasts, the lab-made syndecan-1 ectodomain significantly increased proliferation after 24 hours.
  • Lung fibroblasts treated for 48 hours released more TGF-beta1: 0.80 ng/ml with the ectodomain versus 0.34 ng/ml with medium.
• The authors' conclusion: The authors concluded that shed syndecan-1 contributes to the development of pulmonary fibrosis by attracting neutrophils, impairing repair of the lung lining after injury, and promoting fibrosis. They also concluded that EC-SOD helps prevent oxidant-induced shedding of syndecan-1.
• Limitations the authors state:
  • The authors state that the mechanism by which shed syndecan-1 attracts neutrophils is not known.
  • Knocking down syndecan-1 only partly blocked neutrophil attraction; the authors note that other factors also contribute.
  • The lab-made ectodomain used in cell experiments lacked the sugar (glycosaminoglycan) side chains, so those experiments addressed only the protein itself.

Topics

Medical terms in this study, explained in our Lexicon: Oxidative Stress, Pulmonary Fibrosis, Idiopathic, Interstitial Lung Disease, Fibrosis, Inflammation, Superoxide, Bronchoalveolar Lavage (BAL).

Related studies

• Extracellular superoxide dismutase inhibits inflammation by preventing oxidative fragmentation of hyaluronan — F. Gao, J.R. Koenitzer, et al. (2008)
• Syndecan-1 alters heparan sulfate composition and signaling pathways in malignant mesothelioma — G. Heidari-Hamedani, R.R. Vivès, et al. (2015)
• Paradoxical function for the receptor for advanced glycation end products in mouse models of pulmonary fibrosis — J.M. Englert, C.R. Kliment, et al. (2011)
• Attenuation of oxidative stress and chromosomal aberrations in cultured macrophages and pulmonary cells following self-sustained high temperature synthesis of asbestos — S. La Maestra, R.T. Micale, et al. (2020)
• Patterns of 8-hydroxydeoxyguanosine formation in DNA and indications of oxidative stress in rat and human pleural mesothelial cells after exposure to crocidolite asbestos — H. Fung, Y.W. Kow, et al. (1997)
• Mitochondrial catalase overexpressed transgenic mice are protected against lung fibrosis in part via preventing alveolar epithelial cell mitochondrial DNA damage — S.J. Kim, P. Cheresh, et al. (2016)

Browse more research in The Journal of Biological Chemistry, or return to medical research.