Archives of Toxicology

, Volume 92, Issue 4, pp 1551–1561 | Cite as

Irritant-induced asthma to hypochlorite in mice due to impairment of the airway barrier

  • Sofie Van Den Broucke
  • Lore Pollaris
  • Greetje Vande Velde
  • Erik Verbeken
  • Benoit Nemery
  • Jeroen Vanoirbeek
  • Peter Hoet
Organ Toxicity and Mechanisms


Inhalation of commonly present irritants, such as chlorine and chlorine derivatives, can cause adverse respiratory effects, including irritant-induced asthma (IIA). We hypothesize that due to airway barrier impairment, exposure to hypochlorite (ClO-) can result in airway hypersensitivity. C57Bl/6 mice received an intra-peritoneal (i.p.) injection of the airway damaging agent naphthalene (NA, 200 mg/kg body weight) or vehicle (mineral oil, MO). In vivo micro-computed tomography (CT) images of the lungs were acquired before and at regular time points after the i.p. treatment. After a recovery period of 14 days an intranasal (i.n.) challenge with 0.003% active chlorine (in ClO-) or vehicle (distilled water, H2O) was given, followed by assessment of the breathing frequency. One day later, pulmonary function, along with pulmonary inflammation was determined. Lung permeability was assessed by means of total broncho-alveolar lavage (BAL) protein content and plasma surfactant protein (SP)-D levels. In vivo micro-CT imaging revealed enlargement of the lungs and airways early after NA treatment, with a return to normal at day 14. When challenged i.n. with ClO-, NA-pretreated mice immediately responded with a sensory irritant response. Twenty-four hours later, NA/ClO- mice showed airway hyperreactivity (AHR), accompanied by a neutrophilic and eosinophilic inflammation. NA administration followed by ClO- induced airway barrier impairment, as shown by increased BAL protein and plasma SP-D concentrations; histology revealed epithelial denudation. These data prove that NA-induced lung impairment renders the lungs of mice more sensitive to an airway challenge with ClO-, confirming the hypothesis that incomplete barrier repair, followed by irritant exposure results in airway hypersensitivity.


Irritants Asthma Airway damage Airway epithelium 



This project was supported by a grant from the University of Leuven Research Council (GOA/14/011, STG/15/024 and C24/17/061) and a grant of the Flemish Research Foundation (Research Grant 1504912N).

Compliance with ethical standards

Conflict of interest

The authors declare no conflict of interest.

Supplementary material

204_2018_2161_MOESM1_ESM.docx (27 kb)
Supplementary material 1 (DOCX 26 KB)
204_2018_2161_MOESM2_ESM.pptx (76 kb)
Supplementary material 2 (PPTX 76 KB)


  1. Alarie Y, Ferguson JS, Stock MF, Weyel DA, Schaper M (1987) Sensory and pulmonary irritation of methyl isocyanate in mice and pulmonary irritation and possible cyanidelike effects of methyl isocyanate in guinea pigs. Environ Health Perspect 72(June):159–167CrossRefPubMedPubMedCentralGoogle Scholar
  2. Benfante A, Battaglia S, Principe S, Mitri CD, Paternò A, Spatafora M, Scichilone N (2016) Asthmatics with high levels of serum surfactant protein D have more severe disease. Eur Respir J 47(6):1864–1867. CrossRefPubMedGoogle Scholar
  3. Bernstein JA, Alexis N, Barnes C, Bernstein IL, Nel A, Peden D, Diaz-Sanchez D, Tarlo SM, Williams PB, Bernstein JA (2004) Health effects of air pollution. J Allergy Clin Immunol 114(5):1116–1123. CrossRefPubMedGoogle Scholar
  4. Brooks SM, Bernstein IL (2011) Irritant-induced airway disorders. Immunol Allergy Clin N Am 31(4):747–768. CrossRefGoogle Scholar
  5. Brooks SM, Weiss MA, Bernstein IL (1985) Reactive airways dysfunction syndrome (RADS). Persistent asthma syndrome after high level irritant exposures. Chest 88(3):376–384CrossRefPubMedGoogle Scholar
  6. Buckpitt A, Chang AM, Weir A, Van Winkle L, Duan X, Philpot R, Plopper C (1995) Relationship of cytochrome P450 activity to clara cell cytotoxicity. IV. Metabolism of naphthalene and naphthalene oxide in microdissected airways from mice, rats, and hamsters. Mol Pharmacol 47(1):74–81PubMedGoogle Scholar
  7. Buckpitt A, Boland B, Isbell M, Morin D, Shultz M, Baldwin R, Chan K, Karlsson A, Lin C, Taff A, West J, Fanucchi M, Van Winkle L, Plopper C (2002) Naphthalene-induced respiratory tract toxicity: metabolic mechanisms of toxicity. Drug Metab Rev 34(4):791–820. CrossRefPubMedGoogle Scholar
  8. Casas L, Zock J-P, Torrent M, García-Esteban R, Gracia-Lavedan E, Hyvärinen A, Sunyer J (2013) Use of household cleaning products, exhaled nitric oxide and lung function in children. Eur Respir J 42(5):1415–1418. CrossRefPubMedGoogle Scholar
  9. Crouch EC (2000) Surfactant protein-D and pulmonary host defense. Respir Res 1 (Aug):6. CrossRefGoogle Scholar
  10. Devos FC, Maaske A, Robichaud A, Pollaris L, Seys S, Lopez CA, Verbeken E, Tenbush M, Lories R, Nemery B, Hoet P, Vanoirbeek, J (2017) Forced expiration measurements in mouse models of obstructive and restrictive lung diseases. Respir Res 18(1):123. CrossRefPubMedPubMedCentralGoogle Scholar
  11. Evans RB (2005) Chlorine: state of the art. Lung 183(3):151–167. CrossRefPubMedGoogle Scholar
  12. Fujita M, Shannon JM, Ouchi H, Voelker DR, Nakanishi Y, Mason RJ (2005) Serum surfactant protein D is increased in acute and chronic inflammation in mice. Cytokine 31(1):25–33. CrossRefPubMedGoogle Scholar
  13. Georas SN, Rezaee F (2014) Epithelial barrier function: at the frontline of asthma immunology and allergic airway inflammation. J Allergy Clin Immunol 134(3):509–520. CrossRefPubMedPubMedCentralGoogle Scholar
  14. Greeley MA, Van Winkle LS, Edwards PC, Plopper CG (2010) Airway trefoil factor expression during naphthalene injury and repair. Toxicol Sci 113(2):453–467. CrossRefPubMedGoogle Scholar
  15. Heijink IH, Noordhoek JA, Timens W, van Oosterhout AJ, Postma DS (2014) Abnormalities in airway epithelial junction formation in chronic obstructive pulmonary disease. Am J Respir Crit Care Med 189(11):1439–1442. CrossRefPubMedGoogle Scholar
  16. Holgate ST (2007) Epithelium dysfunction in asthma. J Allergy Clin Immunol 120(6):1233–1244–1246. CrossRefPubMedGoogle Scholar
  17. Hox V, Vanoirbeek JA, Callebaut I, Bobic S, De Vooght V, Ceuppens J, Hoet P, Nemery B, Hellings PW (2011) Airway exposure to hypochlorite prior to ovalbumin induces airway hyperreactivity without evidence for allergic sensitization. Toxicol Lett 204(2–3):101–107. CrossRefPubMedGoogle Scholar
  18. Hox V, Vanoirbeek JA, Alpizar YA, Voedisch S, Callebaut I, Bobic S, Sharify A et al (2013) Crucial role of transient receptor potential ankyrin 1 and mast cells in induction of nonallergic airway hyperreactivity in mice. Am J Respir Crit Care Med 187(5):486–493. CrossRefPubMedGoogle Scholar
  19. Hoyle GW, Svendsen ER (2016) Persistent effects of chlorine inhalation on respiratory health. Ann N Y Acad Sci. PubMedPubMedCentralGoogle Scholar
  20. Jonasson S, Koch B, Bucht A (2013) Inhalation of chlorine causes long-standing lung inflammation and airway hyperresponsiveness in a murine model of chemical-induced lung injury. Toxicology 303(January):34–42. CrossRefPubMedGoogle Scholar
  21. Karagiannis TC, Li X, Tang MM, Orlowski C, El-Osta A, Tang MLK, Royce SG (2012) Molecular model of naphthalene-induced DNA damage in the murine lung. Hum Exp Toxicol 31(1):42–50. CrossRefPubMedGoogle Scholar
  22. López IP, Piñeiro-Hermida S, Pais RS, Torrens R, Hoeflich A, Pichel JG (2016) Involvement of Igf1r in bronchiolar epithelial regeneration: role during repair kinetics after selective club cell ablation. PloS One 11(11):e0166388. CrossRefPubMedPubMedCentralGoogle Scholar
  23. Maestrelli P, Boschetto P, Fabbri LM, Mapp CE (2009) Mechanisms of occupational asthma. J Allergy Clin Immunol 123(3):531–542–544. CrossRefPubMedGoogle Scholar
  24. Martin JG, Campbell HR, Iijima H, Gautrin D, Malo JL, Eidelman DH, Hamid Q, Maghni K (2003) Chlorine-induced injury to the airways in mice. Am J Respir Crit Care Med 168(5):568–574. CrossRefPubMedGoogle Scholar
  25. McGovern TK, Powell WS, Day BJ, White CW, Govindaraju K, Karmouty-Quintana H, Lavoie N, Tan J, Martin JG (2010) Dimethylthiourea protects against chlorine induced changes in airway function in a murine model of irritant induced asthma. Respir Res 11(Oct):138. CrossRefPubMedPubMedCentralGoogle Scholar
  26. McGovern TK, Goldberger M, Allard B, Farahnak S, Hamamoto Y, O’Sullivan M, Hirota N, Martel G, Rousseau S, Martin JG (2015) Neutrophils mediate airway hyperresponsiveness after chlorine-induced airway injury in the mouse. Am J Respir Cell Mol Biol 52(4):513–522. CrossRefPubMedGoogle Scholar
  27. Pan T, Nielsen LD, Allen MJ, Shannon KM, Shannon JM, Selman M, Mason RJ (2002) Serum SP-D is a marker of lung injury in rats. Am J Physiol Lung Cell Mol Physiol 282(4):L824–L832. CrossRefPubMedGoogle Scholar
  28. Plopper CG, Cranz DL, Kemp L, Serabjit-Singh CJ, Philpot RM (1987) Immunohistochemical demonstration of cytochrome P-450 monooxygenase in clara cells throughout the tracheobronchial airways of the rabbit. Exp Lung Res 13(1):59–68. CrossRefPubMedGoogle Scholar
  29. Plopper CG, Suverkropp C, Morin D, Nishio S, Buckpitt A (1992) Relationship of cytochrome P-450 activity to clara cell cytotoxicity. I. Histopathologic comparison of the respiratory tract of mice, rats and hamsters after parenteral administration of naphthalene. J Pharmacol Exp Ther 261(1):353–363PubMedGoogle Scholar
  30. Rezaee F, Georas SN (2014) Breaking barriers. New insights into airway epithelial barrier function in health and disease. Am J Respir Cell Mol Biol 50(5):857–869. CrossRefPubMedPubMedCentralGoogle Scholar
  31. Royce SG, Li X, Tortorella S, Goodings L, Chow BS, Giraud AS, Tang ML, Samuel CS (2014a) Mechanistic insights into the contribution of epithelial damage to airway remodeling. Novel therapeutic targets for asthma. Am J Respir Cell Mol Biol 50(1):180–192. PubMedGoogle Scholar
  32. Royce SG, Patel KP, Samuel CS (2014b) Characterization of a novel model incorporating airway epithelial damage and related fibrosis to the pathogenesis of asthma. Lab Investig 94(12):1326–1339. CrossRefPubMedGoogle Scholar
  33. Shaykhiev R, Crystal RG (2014) Early events in the pathogenesis of chronic obstructive pulmonary disease. Smoking-induced reprogramming of airway epithelial basal progenitor cells. Ann Am Thorac Soc 11 Suppl 5(Dec):S252–S258. CrossRefPubMedGoogle Scholar
  34. Soyka MB, Wawrzyniak P, Eiwegger T, Holzmann D, Treis A, Wanke K, Kast JI, Akdis CA (2012) Defective epithelial barrier in chronic rhinosinusitis: the regulation of tight junctions by IFN-γ and IL-4. J Allergy Clin Immunol 130(5):1087–1096.e10. CrossRefPubMedGoogle Scholar
  35. Tuck SA, Ramos-Barbón D, Campbell H, McGovern T, Karmouty-Quintana H, Martin JG (2008) Time course of airway remodelling after an acute chlorine gas exposure in mice. Respir Res 9(Aug):61. CrossRefPubMedPubMedCentralGoogle Scholar
  36. Van Winkle LS, Buckpitt AR, Nishio SJ, Isaac JM, Plopper CG (1995) Cellular response in naphthalene-induced Clara cell injury and bronchiolar epithelial repair in mice. Am J Physiol 269(6 Pt 1):L800–L818PubMedGoogle Scholar
  37. Van Winkle LS, Johnson ZA, Nishio SJ, Brown CD, Plopper CG (1999) Early events in naphthalene-induced acute Clara cell toxicity: comparison of membrane permeability and ultrastructure. Am J Respir Cell Mol Biol 21(1):44–53. CrossRefPubMedGoogle Scholar
  38. Vandenplas O, Wiszniewska M, Raulf M, de Blay F, Gerth R, van Wijk G, Moscato B, Nemery et al (2014) EAACI position paper: irritant-induced asthma. Allergy 69(9):1141–1153. CrossRefPubMedGoogle Scholar
  39. Velde GV, Poelmans J, Langhe E, Hillen A, Vanoirbeek J, Himmelreich U, Lories RJ (2016) Longitudinal micro-CT provides biomarkers of lung disease that can be used to assess the effect of therapy in preclinical mouse models, and reveal compensatory changes in lung volume. Dis Models Mech 9(1):91–98. CrossRefGoogle Scholar
  40. White CW, Martin JG (2010) Chlorine gas inhalation: human clinical evidence of toxicity and experience in animal models. Proc Am Thorac Soc 7(4):257–263. CrossRefGoogle Scholar
  41. Wigenstam E, Elfsmark L, Koch B, Bucht A, Jonasson S (2016) Acute respiratory changes and pulmonary inflammation involving a pathway of TGF-β1 induction in a rat model of chlorine-induced lung injury. Toxicol Appl Pharmacol 309(Oct):44–54. CrossRefPubMedGoogle Scholar
  42. Winkler C, Atochina-Vasserman EN, Holz O, Beers MF, Erpenbeck VJ, Krug N, Roepcke S, Lauer G, Elmlinger M, Hohlfeld JM (2011) Comprehensive characterisation of pulmonary and serum surfactant protein D in COPD. Respir Res 12(Mar):29. CrossRefPubMedPubMedCentralGoogle Scholar
  43. Xiao C, Puddicombe SM, Field S, Haywood J, Broughton-Head V, Puxeddu I, Haitchi HM et al (2011) Defective epithelial barrier function in asthma. J Allergy Clin Immunol 128(3):549–556.e12. CrossRefPubMedGoogle Scholar
  44. Yildirim AO, Veith M, Rausch T, Müller B, Kilb P, Van Winkle LS, Fehrenbach H (2008) Keratinocyte growth factor protects against Clara cell injury induced by naphthalene. Eur Respir J 32(3):694–704. CrossRefPubMedPubMedCentralGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Centre for Environment and Health, Department of Public Health and Primary CareUniversity of LeuvenLeuvenBelgium
  2. 2.Biomedical MRI Unit, Department of Imaging and PathologyUniversity of LeuvenLeuvenBelgium
  3. 3.Translational Cell and Tissue Research Unit, Department of Imaging and PathologyUniversity of LeuvenLeuvenBelgium

Personalised recommendations