Advertisement

Histochemistry and Cell Biology

, Volume 121, Issue 2, pp 131–140 | Cite as

Early signs of lung fibrosis after in vitro treatment of rat lung slices with CdCl2 and TGF-β1

  • M. KasperEmail author
  • D. Seidel
  • L. Knels
  • N. Morishima
  • A. Neisser
  • S. Bramke
  • R. Koslowski
Original Paper

Abstract

Precision-cut rat lung slices have been employed in combination with an extensive immunohistochemistry of paraffin-embedded slices for monitoring of early pathohistological changes after exposure to CdCl2/TGF-β1. Three days of CdCl2 exposure in combination with TGF-β1 seem to be sufficient to induce lung injury with alterations similar to changes observed in early lung fibrogenesis: (1) extracellular matrix accumulation and myofibroblast transdifferentiation (Sirius red staining, collagen type IV, α-smooth muscle actin), (2) type I cell injury with loss of type I cell antigens (T1α antigen, aquaporin-5, RAGE), (3) increased apoptosis of pulmonary cells (active caspase-3, vimentin cleavage product V1 of caspase-9), and (4) activation of microvascular endothelial cells (podocalyxin, caveolin-1). Western blot analysis confirmed the increasing amount of α-smooth muscle actin, the loss of T1α antigen, and the increase in caveolin-1 immunoreactivity. The explant culture using CdCl2/TGF-β1 provides a suitable tool for the study of other factors involved in pulmonary pathology including transcription factors, cytokines, and other metabolites involved in early stages of fibrogenesis.

Keywords

Slice culture Rat lung Cadmium chloride Fibrosis Aquaporin-5 

Notes

Acknowledgements

The authors thank Dr. Stefan Uhlig, Head Division Pulmonary Pharmacology, Research Center Borstel, Germany, and Dr. Jean-Paul Morin, Faculte de Médecine Pharmacie, Universite de Rouen for introducing the slice culture techniques and all advice. The authors also thank Mrs. Petra Peche for excellent technical assistance. The financial support by elbionAG Dresden is acknowledged.

References

  1. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefPubMedGoogle Scholar
  2. Damiano VV, Cherian PV, Frankel FR, Steeger JR, Sohn M, Oppenheim D, Weinbaum G (1990) Intraluminal fibrosis induced unilaterally by lobar instillation of CdCl2 into the rat lung. Am J Pathol 137:883–894PubMedGoogle Scholar
  3. de Kanter R, Monshouwer M, Meijer DK, Groothuis GM (2002) Precision-cut organ slices as a tool to study toxicity and metabolism of xenobiotics with special reference to non-hepatic tissues. Curr Drug Metab 3:39–59PubMedGoogle Scholar
  4. Drab M, Verkade P, Elger M, Kasper M, Lohn M, Lauterbach B, Menne J, Lindschau C, Mende F, Luft FC, Schedl A, Haller H, Kurzchalia TV (2001) Loss of caveolae, vascular dysfunction, and pulmonary defects in caveolin-1 gene-disrupted mice. Science 293:2449–2452PubMedGoogle Scholar
  5. Driscoll KE, Maurer JK, Poynter J, Higgins J, Asquith T, Miller NS (1992) Stimulation of rat alveolar macrophage fibronectin release in a cadmium chloride model of lung injury and fibrosis. Toxicol Appl Pharmacol 116:30–37PubMedGoogle Scholar
  6. Fehrenbach H, Kasper M, Tschernig T, Shearman MS, Schuh D, Muller M (1998) Receptor for advanced glycation end products (RAGE) exhibits highly differential cellular and subcellular localisation in rat and human lung. Cell Mol Biol (Noisy-le-grand) 44:1147–1157Google Scholar
  7. Fisher GL, Placke ME (1987) In vitro models of lung toxicity. Toxicology 47:71–93CrossRefPubMedGoogle Scholar
  8. Hoyt DG, Lazo JS (1993) NAD depletion after in vitro exposure of murine lung slices to bleomycin. Biochem Pharmacol 46:1819–1824Google Scholar
  9. Huttunen HJ, Kuja-Panula J, Rauvala H (2002) Receptor for advanced glycation end products (RAGE) signaling induces CREB-dependent chromogranin expression during neuronal differentiation. J Biol Chem 277:38635–38646CrossRefPubMedGoogle Scholar
  10. Kapanci Y, Ribaux C, Chaponnier C, Gabbiani G (1992) Cytoskeletal features of alveolar myofibroblasts and pericytes in normal human and rat lung. J Histochem Cytochem 40:1955–1963PubMedGoogle Scholar
  11. Kasper M, Reimann T, Hempel U, Wenzel KW, Bierhaus A, Schuh D, Dimmer V, Haroske G, Muller M (1998) Loss of caveolin expression in type I pneumocytes as an indicator of subcellular alterations during lung fibrogenesis. Histochem Cell Biol 109:41–48PubMedGoogle Scholar
  12. Kelly M, Kolb M, Bonniaud P, Gauldie J (2003) A re-evaluation of fibrogenic cytokines in lung fibrosis. Curr Pharm Des 9:39–49PubMedGoogle Scholar
  13. Kinnard WV, Tuder R, Papst P, Fisher JH (1994) Regulation of alveolar type II cell differentiation and proliferation in adult rat lung explants. Am J Respir Cell Mol Biol 11:416–425PubMedGoogle Scholar
  14. Klein JM, McCarthy TA, Dagle JM, Snyder JM (2002) Antisense inhibition of surfactant protein A decreases tubular myelin formation in human fetal lung in vitro. Am J Physiol Lung Cell Mol Physiol 282:L386–L393PubMedGoogle Scholar
  15. Koslowski R, Dobbs LG, Wenzel KW, Schuh D, Muller M, Kasper M (1998) Loss of immunoreactivity for RTI40, a type I cell-specific protein in the alveolar epithelium of rat lungs with bleomycin-induced fibrosis. Eur Respir J 12:1397–1403CrossRefPubMedGoogle Scholar
  16. Kuniyasu H, Chihara Y, Takahashi T (2003) Co-expression of receptor for advanced glycation end products and the ligand amphoterin associates closely with metastasis of colorectal cancer. Oncol Rep 10:445–448PubMedGoogle Scholar
  17. Le Prieur E, Vaz E, Bion A, Dionnet F, Morin JP (2000) Toxicity of diesel engine exhausts in an in vitro model of lung slices in biphasic organotypic culture: induction of a proinflammatory and apoptotic response. Arch Toxicol 74:460–466CrossRefPubMedGoogle Scholar
  18. Lin CJ, Yang PC, Hsu MT, Yew FH, Liu TY, Shun CT, Tyan SW, Lee TC (1998) Induction of pulmonary fibrosis in organ-cultured rat lung by cadmium chloride and transforming growth factor-beta 1. Toxicology 127:157–166CrossRefPubMedGoogle Scholar
  19. McBride S, Rannie D, Harrison DJ (2000) Gene transfer to adult human lung tissue ex vivo. Gene Ther 7:675–678CrossRefPubMedGoogle Scholar
  20. Morin JP, Fouquet F, Monteil C, Le Prieur E, Vaz E, Dionnet F (1999) Development of a new in vitro system for continuous in vitro exposure of lung tissue to complex atmospheres: application to diesel exhaust toxicology. Cell Biol Toxicol 15:143–152PubMedGoogle Scholar
  21. Nakanishi K, Maruyama M, Shibata T, Morishima N (2001) Identification of a caspase-9 substrate and detection of its cleavage in programmed cell death during mouse development. J Biol Chem 276:41237–41244CrossRefPubMedGoogle Scholar
  22. Rahman I, Skwarska E, Henry M, Davis M, O’Connor CM, Fitzgerald MX, Greening A, MacNee W (1999) Systemic and pulmonary oxidative stress in idiopathic pulmonary fibrosis. Free Radic Biol Med 27:60–68CrossRefPubMedGoogle Scholar
  23. Shapiro P, Absher PM, Posada JP, Evans JN (1997) Activation of ERK and JNK1 MAP kinases in cultured lung tissue. Am J Physiol 273:L459–L467PubMedGoogle Scholar
  24. Shih CM, Wu JS, Ko WC, Wang LF, Wei YH, Liang HF, Chen YC, Chen CT (2003) Mitochondria-mediated caspase-independent apoptosis induced by cadmium in normal human lung cells. J Cell Biochem 89:335–347CrossRefPubMedGoogle Scholar
  25. Towne JE, Harrod KS, Krane CM, Menon AG (2000) Decreased expression of aquaporin (AQP)1 and AQP5 in mouse lung after acute viral infection. Am J Respir Cell Mol Biol 22:34–44PubMedGoogle Scholar
  26. Towne JE, Krane CM, Bachurski CJ, Menon AG (2001) Tumor necrosis factor-alpha inhibits aquaporin 5 expression in mouse lung epithelial cells. J Biol Chem 276:18657–18664CrossRefPubMedGoogle Scholar
  27. Wohlsen A, Uhlig S, Martin C (2001) Immediate allergic response in small airways. Am J Respir Crit Care Med 163:1462–1469PubMedGoogle Scholar
  28. Wright AF, Green TP, Robson RT, Niewola Z, Wyatt I, Smith LL (1987) Specific polyclonal and monoclonal antibody prevents paraquat accumulation into rat lung slices. Biochem Pharmacol 36:1325–1331Google Scholar

Copyright information

© Springer-Verlag 2004

Authors and Affiliations

  • M. Kasper
    • 1
    Email author
  • D. Seidel
    • 2
  • L. Knels
    • 1
  • N. Morishima
    • 3
  • A. Neisser
    • 1
  • S. Bramke
    • 1
  • R. Koslowski
    • 2
  1. 1.Institute of Anatomy, Medical FacultyTechnical University of DresdenDresdenGermany
  2. 2.Institute of Physiological Chemistry, Medical FacultyTechnical University of DresdenDresdenGermany
  3. 3.Bioarchitect Research Group and Cellular and Molecular Biology LaboratoryRIKENSaitamaJapan

Personalised recommendations