Dibutyryl-cAMP attenuates pulmonary fibrosis by blocking myofibroblast differentiation via PKA/CREB/CBP signaling in rats with silicosis

Our plain-English summary of Dibutyryl-cAMP attenuates pulmonary fibrosis by blocking myofibroblast differentiation via PKA/CREB/CBP signaling in rats with silicosis, by Y. Liu, H. Xu, Y. Geng, et al (Respiratory Research, 2017).

The study

• Authors: Y. Liu, H. Xu, Y. Geng, D. Xu, L. Zhang, Y. Yang, Z. Wei, B. Zhang, S. Li, X. Gao, R. Wang, X. Zhang, D. Brann, F. Yang
• Journal: Respiratory Research
• Published: 2017;18(1):38
• Cited by: 49 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Background: Myofibroblasts play a major role in the synthesis of extracellular matrix (ECM) and the stimulation of these cells is thought to play an important role in the development of silicosis. The present study was undertaken to investigate the anti-fibrotic effects of dibutyryl-cAMP (db-cAMP) on rats induced by silica.

Methods: A HOPE MED 8050 exposure control apparatus was used to create the silicosis model. Rats were randomly divided into 4 groups: 1)controls for 16 w; 2)silicosis for 16 w; 3)db-cAMP pre-treatment; 4) db-cAMP post-treatment. Rat pulmonary fibroblasts were cultured in vitro and divided into 4 groups as follows: 1) controls; 2) 10-7mol/L angiotensin II (Ang II); 3) Ang II +10-4 mol/L db-cAMP; and 4) Ang II + db-cAMP+ 10-6 mol/L H89. Hematoxylin-eosin (HE), Van Gieson staining and immunohistochemistry (IHC) were performed to observe the histomorphology of lung tissue. The levels of cAMP were detected by enzyme immunoassay. Double-labeling for α-SMA with Gαi3, protein kinase A (PKA), phosphorylated cAMP-response element-binding protein (p-CREB), and p-Smad2/3 was identified by immunofluorescence staining. Protein levels were detected by Western blot analysis. The interaction between CREB-binding protein (CBP) and Smad2/3 and p-CREB were measured by co-immunoprecipitation (Co-IP).

Results: Db-cAMP treatment reduced the number and size of silicosis nodules, inhibited myofibroblast differentiation, and extracellular matrix deposition in vitro and in vivo. In addition, db-cAMP regulated Gαs protein and inhibited expression of Gαi protein, which increased endogenous cAMP. Db-cAMP increased phosphorylated cAMP-response element-binding protein (p-CREB) via protein kinase A (PKA) signaling, and decreased nuclear p-Smad2/3 binding with CREB binding protein (CBP), which reduced activation of p-Smads in fibroblasts induced by Ang II.

Conclusions: This study showed an anti-silicotic effect of db-cAMP that was mediated via PKA/p-CREB/CBP signaling. Furthermore, the findings offer novel insight into the potential use of cAMP signaling for therapeutic strategies to treat silicosis.

In plain English

This laboratory study in rats and cultured rat lung cells concerned silicosis, lung scarring caused by crystalline silica dust, not asbestos. Male Wistar rats breathed silica dust for 3 hours a day at 2000 mg/m3 for 16 weeks. Some exposed rats received daily injections of dibutyryl-cAMP (db-cAMP), a laboratory form of a cell-signaling molecule. One group started it 48 hours before exposure and another after 4 weeks of exposure. Comparison groups received saline. Separately, lung fibroblasts (scar-forming cells) were stimulated with angiotensin II, with or without db-cAMP and a blocker called H89. Under the microscope, both db-cAMP schedules reduced the number and size of silicotic nodules and the build-up of collagen fibers. In lungs and in cells, db-cAMP reduced scar-related proteins and changed signaling proteins (PKA, phosphorylated CREB, phosphorylated Smad2/3). In cells, H89 reduced the effects of db-cAMP. The authors concluded that db-cAMP had an anti-silicotic effect working through PKA/p-CREB/CBP signaling. The article does not state how many rats were in each group, and it gives its results in figures rather than as numbers in the text.

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: Animal experiment (rats randomly divided into 4 groups) combined with experiments on cultured rat lung fibroblasts
• Who was studied: Male Wistar rats, 3 weeks of age at the start, in 4 groups: controls given saline for 16 weeks; silica-exposed for 16 weeks with saline; db-cAMP pre-treatment; db-cAMP post-treatment. Also rat lung fibroblasts grown in the laboratory. The number of rats per group is not stated.
• Exposure: Inhaled silica (SiO2, 5 um particles) from an exposure apparatus at a dust concentration of 2000 mg/m3, 3 hours per day, for 16 weeks. Treatment was db-cAMP, 10 mg/kg per day by injection under the skin. Pre-treatment started 48 hours before silica. Post-treatment started after 4 weeks of silica and continued for 12 weeks. Cultured fibroblasts were given angiotensin II (10 to the minus 7 mol/L), db-cAMP (10 to the minus 4 mol/L) and the PKA blocker H89 (10 to the minus 6 mol/L).
• Main results:
  • In silica-exposed rats, both db-cAMP pre-treatment and post-treatment reduced the number and size of silicotic nodules and the accumulation of collagen fibers on tissue staining
  • In rat lung tissue, fibronectin, collagen type I and alpha-smooth muscle actin were increased in the 16-week silica group compared with controls. Db-cAMP pre-treatment and post-treatment reduced these changes (Western blot, n = 6 independent experiments).
  • In fibroblasts stimulated with angiotensin II, db-cAMP reduced fibronectin, collagen type I and alpha-smooth muscle actin. Blocking PKA with H89 reduced this effect of db-cAMP.
  • In silicotic rat lung tissue, db-cAMP pre-treatment or post-treatment decreased the G proteins Gai2 and Gai3 and increased Gas and cAMP
  • Phosphorylated CREB was seen in cell nuclei of normal lung tissue, with no staining in silicotic nodules. PKA and phosphorylated CREB were reduced in the silicosis group, and db-cAMP pre-treatment or post-treatment increased both.
  • Db-cAMP inhibited the rise in phosphorylated Smad2/3 seen in silicotic rats and in angiotensin II-stimulated fibroblasts
  • In fibroblasts, db-cAMP increased the binding of phosphorylated CREB to CBP and reduced the binding of phosphorylated Smad2/3 to CBP (co-immunoprecipitation, n = 3 independent experiments)
• The authors' conclusion: The authors concluded that db-cAMP showed an anti-silicotic effect mediated through PKA/p-CREB/CBP signaling, by disrupting p-Smad2/3 binding with CBP and inhibiting myofibroblast differentiation

Topics

The National Library of Medicine indexes this study under subjects our site covers: Asbestosis.

Medical terms in this study, explained in our Lexicon: Pulmonary Fibrosis, Silicosis, Fibroblast, Immunohistochemistry, Deposition.

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