, Volume 31, Issue 3, pp 167–179 | Cite as

IL-17 Producing γδ T Cells are Required for a Controlled Inflammatory Response after Bleomycin-induced Lung Injury

  • Ruedi K. BraunEmail author
  • Christina Ferrick
  • Paul Neubauer
  • Michael Sjoding
  • Anja Sterner-Kock
  • Martin Kock
  • Lei Putney
  • David A. Ferrick
  • Dallas M. Hyde
  • Robert B. Love


γδ T cells play a key role in the regulation of inflammatory responses in epithelial tissue, and in adaptive immunity, as γδ T cell deficient mice have a severely impaired capacity to clear lung pathogens. γδ T cells regulate the initial inflammatory response to microbial invasion and thereby protect against tissue injury. Here we examined the response of γδ T cells to lung injury induced by bleomycin, in an effort to study the inflammatory response in the absence of any adaptive immune response to a pathogen.


After lung injury by bleomycin, we localized the γδ T cells to the lung lesions. γδ T cells were the predominant source of IL-17 (as detected by flow cytometry and real-time PCR). Moreover, γδ T cell knockout mice showed a significant reduction in cellular infiltration into the airways, reduced expression of IL-6 in the lung, and a significant delay in epithelial repair.


Mouse γδ T cells produce IL-17 in response to lung injury and are required for an organized inflammatory response and epithelial repair. The lack of γδ T cells correlates with increased inflammation and fibrosis.

Key words

γδ T cell bleomycin lung fibrosis IL-17 



We acknowledge the help of Kathy Schell and Patricia Simms for the cell sorting. This research was supported by a grant from the NHLBI–NIH # RO1 HL 56262 and by a grant from the Department of Surgery, Loyola University of Chicago.

Authors’ Contributions

RKB designed the study, carried out the flow cytometry studies, participated in the bleomycin instillation, drafted the manuscript, and performed the statistical analysis.

CF performed the staining of the tissue sections and helped with the bleomycin instillation. PN carried out the molecular analysis of the IL-17 expression. MS participated in the flow cytometry studies and the bleomycin instillation. ASK carried out the pathology analysis of the mouse lungs. MK participated in the preparation of the mouse lungs and the pathology analysis. LP carried out the confocal microscopy. DAF participated in the design of the study and helped to draft the manuscript. DMH performed the morphometric analysis of the epithelial repair. RBL participated in the design of the study and helped to draft the manuscript.

Competing Interests

The authors declare that they have no competing interests.


  1. 1.
    Langhorne, J., P. Mombaerts, and S. Tonegawa. 1995. alpha beta and gamma delta T cells in the immune response to the erythrocytic stages of malaria in mice. Int. Immunol. 7:1005–1011.PubMedCrossRefGoogle Scholar
  2. 2.
    Mombaerts, P., J. Arnoldi, F. Russ, S. Tonegawa, and S. H. Kaufmann. 1993. Different roles of alpha beta and gamma delta T cells in immunity against an intracellular bacterial pathogen. Nature 365:53–56.PubMedCrossRefGoogle Scholar
  3. 3.
    Korn, T., M. Oukka, and E. Bettelli. 2007. Th17 cells: effector T cells with inflammatory properties. Semin. Immunol. 19:362–371.Google Scholar
  4. 4.
    Park, H., Z. Li, X. O. Yang, S. H. Chang, R. Nurieva, Y. H. Wang, Y. Wang, L. Hood, Z. Zhu, Q. Tian, and C. Dong. 2005. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17. Nat. Immunol. 6:1133–1141.PubMedCrossRefGoogle Scholar
  5. 5.
    Roark, C. L., J. D. French, M. A. Taylor, A. M. Bendele, W. K. Born, and R. L. O’Brien. 2007. Exacerbation of collagen-induced arthritis by oligoclonal, IL-17-producing gamma delta T cells. J. Immunol. 179:5576–5583.PubMedGoogle Scholar
  6. 6.
    Lockhart, E., A. M. Green, and J. L. Flynn. 2006. IL-17 production is dominated by gammadelta T cells rather than CD4 T cells during Mycobacterium tuberculosis infection. J. Immunol. 177:4662–4669.PubMedGoogle Scholar
  7. 7.
    Kelly, M. N., J. K. Kolls, K. Happel, J. D. Schwartzman, P. Schwarzenberger, C. Combe, M. Moretto, and I. A. Khan. 2005. Interleukin-17/interleukin-17 receptor-mediated signaling is important for generation of an optimal polymorphonuclear response against Toxoplasma gondii infection. Infect. Immun. 73:617–621.PubMedCrossRefGoogle Scholar
  8. 8.
    Ye, P., P. B. Garvey, P. Zhang, S. Nelson, G. Bagby, W. R. Summer, P. Schwarzenberger, J. E. Shellito, and J. K. Kolls. 2001. Interleukin-17 and lung host defense against Klebsiella pneumoniae infection. Am. J. Respir. Cell Mol. Biol. 25:335–340.PubMedGoogle Scholar
  9. 9.
    Huang, W., L. Na, P. L. Fidel, and P. Schwarzenberger. 2004. Requirement of interleukin-17A for systemic anti-Candida albicans host defense in mice. J. Infect. Dis. 190:624–631.PubMedCrossRefGoogle Scholar
  10. 10.
    Ye, P., F. H. Rodriguez, S. Kanaly, K. L. Stocking, J. Schurr, P. Schwarzenberger, P. Oliver, W. Huang, P. Zhang, J. Zhang, J. E. Shellito, G. J. Bagby, S. Nelson, K. Charrier, J. J. Peschon, and J. K. Kolls. 2001. Requirement of interleukin 17 receptor signaling for lung CXC chemokine and granulocyte colony-stimulating factor expression, neutrophil recruitment, and host defense. J. Exp. Med. 194:519–527.PubMedCrossRefGoogle Scholar
  11. 11.
    Langrish, C. L., Y. Chen, W. M. Blumenschein, J. Mattson, B. Basham, J. D. Sedgwick, T. McClanahan, R. A. Kastelein, and D. J. Cua. 2005. IL-23 drives a pathogenic T cell population that induces autoimmune inflammation. J. Exp. Med. 201:233–240.PubMedCrossRefGoogle Scholar
  12. 12.
    Bi, Y., G. Liu, and R. Yang. 2007. Th17 cell induction and immune regulatory effects. J. Cell. Physiol. 211:273–278.PubMedCrossRefGoogle Scholar
  13. 13.
    Born, W. K., C. L. Reardon, and R. L. O’Brien. 2006. The function of gammadelta T cells in innate immunity. Curr. Opin. Immunol. 18:31–38.PubMedCrossRefGoogle Scholar
  14. 14.
    Nanno, M., T. Shiohara, H. Yamamoto, K. Kawakami, and H. Ishikawa. 2007. gammadelta T cells: firefighters or fire boosters in the front lines of inflammatory responses. Immunol. Rev. 215:103–113.PubMedCrossRefGoogle Scholar
  15. 15.
    Moore, T. A., B. B. Moore, M. W. Newstead, and T. J. Standiford. 2000. Gamma delta-T cells are critical for survival and early proinflammatory cytokine gene expression during murine Klebsiella pneumonia. J. Immunol. 165:2643–2650.PubMedGoogle Scholar
  16. 16.
    Takano, M., H. Nishimura, Y. Kimura, Y. Mokuno, J. Washizu, S. Itohara, Y. Nimura, and Y. Yoshikai. 1998. Protective roles of gamma delta T cells and interleukin-15 in Escherichia coli infection in mice. Infect. Immun. 66:3270–3278.PubMedGoogle Scholar
  17. 17.
    Nakamura, T., G. Matsuzaki, and K. Nomoto. 1999. The protective role of T-cell receptor Vgamma1+ T cells in primary infection with Listeria monocytogenes. Immunology 96:29–34.PubMedCrossRefGoogle Scholar
  18. 18.
    Blum, R. H., S. K. Carter, and K. Agre. 1973. A clinical review of bleomycin–a new antineoplastic agent. Cancer 31:903–914.PubMedCrossRefGoogle Scholar
  19. 19.
    Jones, A. W. 1978. Bleomycin lung damage: the pathology and nature of the lesion. Br. J. Dis. Chest 72:321–326.PubMedCrossRefGoogle Scholar
  20. 20.
    Braun, R. K., A. Sterner-Kock, P. J. Kilshaw, D. A. Ferrick, and S. N. Giri. 1996. Integrin alpha E beta 7 expression on BAL CD4+, CD8+, and gamma delta T-cells in bleomycin-induced lung fibrosis in mouse. Eur. Respir. J. 9:673–679.PubMedCrossRefGoogle Scholar
  21. 21.
    Thrall, R. S., and R. W. Barton. 1984. A comparison of lymphocyte populations in lung tissue and in bronchoalveolar lavage fluid of rats at various times during the development of bleomycin-induced pulmonary fibrosis. Am. Rev. Respir. Dis. 129:279–283.PubMedGoogle Scholar
  22. 22.
    Zhu, J., D. A. Cohen, S. N. Goud, and A. M. Kaplan. 1996. Contribution of T lymphocytes to the development of bleomycin-induced pulmonary fibrosis. Ann. N.Y. Acad. Sci. 796:194–202.PubMedCrossRefGoogle Scholar
  23. 23.
    Raisfeld, I. H. 1980. Pulmonary toxicity of bleomycin analogs. Toxicol. Appl. Pharmacol. 56:326–336.PubMedCrossRefGoogle Scholar
  24. 24.
    Creemers, L. B., D. C. Jansen, A. Veen-Reurings, B. T. van den, and V. Everts. 1997. Microassay for the assessment of low levels of hydroxyproline. Biotechniques 22:656–658.PubMedGoogle Scholar
  25. 25.
    Bolender, R. P., D. M. Hyde, and R. T. Dehoff. 1993. Lung morphometry: a new generation of tools and experiments for organ, tissue, cell, and molecular biology. Am. J. Physiol. 265:L521–L548.PubMedGoogle Scholar
  26. 26.
    Hyde, D. M., D. J. Magliano, E. Reus, N. K. Tyler, S. Nichols, and W. S. Tyler. 1992. Computer-assisted morphometry: point, intersection, and profile counting and three-dimensional reconstruction. Microsc. Res. Tech. 21:262–270.PubMedCrossRefGoogle Scholar
  27. 27.
    Yang, H., P. A. Antony, B. E. Wildhaber, and D. H. Teitelbaum. 2004. Intestinal intraepithelial lymphocyte gammadelta-T cell-derived keratinocyte growth factor modulates epithelial growth in the mouse. J. Immunol. 172:4151–4158.PubMedGoogle Scholar
  28. 28.
    D’Souza, C. D., A. M. Cooper, A. A. Frank, R. J. Mazzaccaro, B. R. Bloom, and I. M. Orme. 1997. An anti-inflammatory role for gamma delta T lymphocytes in acquired immunity to Mycobacterium tuberculosis. J. Immunol. 158:1217–1221.PubMedGoogle Scholar
  29. 29.
    King, D. P., D. M. Hyde, K. A. Jackson, D. M. Novosad, T. N. Ellis, L. Putney, M. Y. Stovall, L. S. Van Winkle, B. L. Beaman, and D. A. Ferrick. 1999. Cutting edge: protective response to pulmonary injury requires gamma delta T lymphocytes. J. Immunol. 162:5033–5036.PubMedGoogle Scholar
  30. 30.
    Afzali, B., G. Lombardi, R. I. Lechler, and G. M. Lord. 2007. The role of T helper 17 (Th17) and regulatory T cells (Treg) in human organ transplantation and autoimmune disease. Clin. Exp. Immunol. 148:32–46.PubMedCrossRefGoogle Scholar
  31. 31.
    Shin, H. C., N. Benbernou, S. Esnault, and M. Guenounou. 1999. Expression of IL-17 in human memory CD45RO+ T lymphocytes and its regulation by protein kinase A pathway. Cytokine 11:257–266.PubMedCrossRefGoogle Scholar
  32. 32.
    Shibata, K., H. Yamada, H. Hara, K. Kishihara, and Y. Yoshikai. 2007. Resident Vdelta1+ gammadelta T cells control early infiltration of neutrophils after Escherichia coli infection via IL-17 production. J. Immunol. 178:4466–4472.PubMedGoogle Scholar
  33. 33.
    Ferrick, D. A., D. P. King, K. A. Jackson, R. K. Braun, S. Tam, D. M. Hyde, and B. L. Beaman. 2000. Intraepithelial gamma delta T lymphocytes: sentinel cells at mucosal barriers. Springer Semin. Immunopathol 22:283–296.PubMedCrossRefGoogle Scholar
  34. 34.
    Tam, S., D. P. King, and B. L. Beaman. 2001. Increase of {gamma}{delta} T Lymphocytes in murine lungs occurs during recovery from pulmonary infection by nocardia asteroides. Infect. Immun. 69:6165–6171.PubMedCrossRefGoogle Scholar
  35. 35.
    Hsieh, B., M. D. Schrenzel, T. Mulvania, H. D. Lepper, L. DiMolfetto-Landon, and D. A. Ferrick. 1996. In vivo cytokine production in murine listeriosis. Evidence for immunoregulation by gamma delta+ T cells. J. Immunol. 156:232–237.PubMedGoogle Scholar
  36. 36.
    McKenzie, B. S., R. A. Kastelein, and D. J. Cua. 2005. Understanding the IL-23-IL-17 immune pathway. Trends Immunol. 27:17–23.PubMedCrossRefGoogle Scholar
  37. 37.
    Bowman, E. P., A. A. Chackerian, and D. J. Cua. 2006. Rationale and safety of anti-interleukin-23 and anti-interleukin-17A therapy. Curr. Opin. Infect. Dis. 19:245–252.PubMedCrossRefGoogle Scholar
  38. 38.
    Rohn, T. A., G. T. Jennings, M. Hernandez, P. Grest, M. Beck, Y. Zou, M. Kopf, and M. F. Bachmann. 2006. Vaccination against IL-17 suppresses autoimmune arthritis and encephalomyelitis. Eur. J. Immunol. 36:2857–2867.PubMedCrossRefGoogle Scholar
  39. 39.
    Schrier, D. J., S. H. Phan, and P. A. Ward. 1982. Cellular sensitivity to collagen in bleomycin-treated rats. J. Immunol. 129:2156–2159.PubMedGoogle Scholar
  40. 40.
    Kravis, T. C., A. Ahmed, T. E. Brown, J. D. Fulmer, and R. G. Crystal. 1976. Pathogenic mechanisms in pulmonary fibrosis: collagen-induced migration inhibition factor production and cytotoxicity mediated by lymphocytes. J. Clin. Invest. 58:1223–1232.PubMedCrossRefGoogle Scholar
  41. 41.
    Nakos, G., A. Adams, and N. Andriopoulos. 1993. Antibodies to collagen in patients with idiopathic pulmonary fibrosis. Chest 103:1051–1058.PubMedCrossRefGoogle Scholar
  42. 42.
    Collard, H. R., B. B. Moore, K. R. Flaherty, K. K. Brown, R. J. Kaner, T. E. King Jr., J. A. Lasky, J. E. Loyd, I. Noth, M. A. Olman, G. Raghu, J. Roman, J. H. Ryu, D. A. Zisman, G. W. Hunninghake, T. V. Colby, J. J. Egan, D. M. Hansell, T. Johkoh, N. Kaminski, D. S. Kim, Y. Kondoh, D. A. Lynch, J. Muller-Quernheim, J. L. Myers, A. G. Nicholson, M. Selman, G. B. Toews, A. U. Wells, and F. J. Martinez. 2007. Acute exacerbations of idiopathic pulmonary fibrosis. Am. J. Respir. Crit. Care Med. 176:636–643.PubMedCrossRefGoogle Scholar
  43. 43.
    Adamson, I. Y. 1984. Drug-induced pulmonary fibrosis. Environ. Health Perspect. 55:25–36.PubMedCrossRefGoogle Scholar
  44. 44.
    Ghosh, S., T. Mendoza, L. A. Ortiz, G. W. Hoyle, C. D. Fermin, A. R. Brody, M. Friedman, and G. F. Morris. 2002. Bleomycin sensitivity of mice expressing dominant-negative p53 in the lung epithelium. Am. J. Respir. Crit. Care Med. 166:890–897.PubMedCrossRefGoogle Scholar
  45. 45.
    Serrano-Mollar, A., M. Nacher, G. Gay-Jordi, D. Closa, A. Xaubet, and O. Bulbena. 2007. Intratracheal transplantation of alveolar Type II cells reverse bleomycin-induced lung fibrosis. Am. J. Respir. Crit. Care Med. 176:1261–1268.Google Scholar
  46. 46.
    Watanabe, M., M. Ebina, F. M. Orson, A. Nakamura, K. Kubota, D. Koinuma, K. Akiyama, M. Maemondo, S. Okouchi, M. Tahara, K. Matsumoto, T. Nakamura, and T. Nukiwa. 2005. Hepatocyte growth factor gene transfer to alveolar septa for effective suppression of lung fibrosis. Mol. Ther. 12:58–67.PubMedCrossRefGoogle Scholar
  47. 47.
    Yi, E. S., S. T. Williams, H. Lee, D. M. Malicki, E. M. Chin, S. Yin, J. Tarpley, and T. R. Ulich. 1996. Keratinocyte growth factor ameliorates radiation- and bleomycin-induced lung injury and mortality. Am. J. Pathol. 149:1963–1970.PubMedGoogle Scholar
  48. 48.
    Jameson, J., and W. L. Havran. 2007. Skin gammadelta T-cell functions in homeostasis and wound healing. Immunol. Rev. 215:114–122.PubMedCrossRefGoogle Scholar
  49. 49.
    Chen, Y., K. Chou, E. Fuchs, W. L. Havran, and R. Boismenu. 2002. Protection of the intestinal mucosa by intraepithelial gamma delta T cells. Proc. Natl. Acad. Sci. U. S. A. 99:14338–14343.PubMedCrossRefGoogle Scholar
  50. 50.
    Koohsari, H., M. Tamaoka, H. R. Campbell, and J. G. Martin. 2007. The role of gamma delta T cells in airway epithelial injury and bronchial responsiveness after chlorine gas exposure in mice. Respir. Res. 8:21.PubMedCrossRefGoogle Scholar
  51. 51.
    Inoue, D., M. Numasaki, M. Watanabe, H. Kubo, T. Sasaki, H. Yasuda, M. Yamaya, and H. Sasaki. 2006. IL-17A promotes the growth of airway epithelial cells through ERK-dependent signaling pathway. Biochem. Biophys. Res. Commun. 347:852–858.PubMedCrossRefGoogle Scholar
  52. 52.
    Manoury, B., S. Nenan, I. Guenon, V. Lagente, and E. Boichot. 2007. Influence of early neutrophil depletion on MMPs/TIMP-1 balance in bleomycin-induced lung fibrosis. Int. Immunopharmacol. 7:900–911.PubMedCrossRefGoogle Scholar
  53. 53.
    Kolls, J. K., and A. Linden. 2004. Interleukin-17 family members and inflammation. Immunity. 21:467–476.PubMedCrossRefGoogle Scholar
  54. 54.
    Hyde, D. M., L. A. Miller, R. J. McDonald, M. Y. Stovall, V. Wong, K. E. Pinkerton, C. D. Wegner, R. Rothlein, and C. G. Plopper. 1999. Neutrophils enhance clearance of necrotic epithelial cells in ozone-induced lung injury in rhesus monkeys. Am. J. Physiol. 277:L1190–L1198.PubMedGoogle Scholar
  55. 55.
    Liu, W., A. L. Putnam, Z. Xu-Yu, G. L. Szot, M. R. Lee, S. Zhu, P. A. Gottlieb, P. Kapranov, T. R. Gingeras, B. F. de St Groth, C. Clayberger, D. M. Soper, S. F. Ziegler, and J. A. Bluestone. 2006. CD127 expression inversely correlates with FoxP3 and suppressive function of human CD4(+) T reg cells. J. Exp. Med. 203:1701–1711.PubMedCrossRefGoogle Scholar
  56. 56.
    Gunnarsson, M., S. M. Walther, T. Seidal, G. D. Bloom, and S. Lennquist. 1998. Exposure to chlorine gas: effects on pulmonary function and morphology in anaesthetised and mechanically ventilated pigs. J. Appl. Toxicol. 18:249–255.PubMedCrossRefGoogle Scholar
  57. 57.
    Crouch, E. 1990. Pathobiology of pulmonary fibrosis. Am. J. Physiol. 259:159–184.Google Scholar
  58. 58.
    Eickelberg, O., A. Pansky, R. Mussmann, M. Bihl, M. Tamm, P. Hildebrand, A. P. Perruchoud, and M. Roth. 1999. Transforming growth factor-beta1 induces interleukin-6 expression via activating protein-1 consisting of JunD homodimers in primary human lung fibroblasts. J. Biol. Chem. 274:12933–12938.PubMedCrossRefGoogle Scholar
  59. 59.
    Xing, Z., J. Gauldie, G. Cox, H. Baumann, M. Jordana, X. F. Lei, and M. K. Achong. 1998. IL-6 is an antiinflammatory cytokine required for controlling local or systemic acute inflammatory responses. J. Clin. Invest. 101:311–320.PubMedCrossRefGoogle Scholar
  60. 60.
    Gurujeyalakshmi, G., Y. Wang, and S. N. Giri. 2000. Taurine and niacin block lung injury and fibrosis by down-regulating bleomycin-induced activation of transcription nuclear factor-kappaB in mice. J. Pharmacol. Exp. Ther. 293:82–90.PubMedGoogle Scholar
  61. 61.
    Tabata, C., Y. Kadokawa, R. Tabata, M. Takahashi, K. Okoshi, Y. Sakai, M. Mishima, and H. Kubo. 2006. All-trans-retinoic acid prevents radiation- or bleomycin-induced pulmonary fibrosis. Am. J. Respir. Crit. Care Med. 174:1352–1360.PubMedCrossRefGoogle Scholar
  62. 62.
    Sener, G., N. Topaloglu, A. O. Sehirli, F. Ercan, and N. Gedik. 2007. Resveratrol alleviates bleomycin-induced lung injury in rats. Pulm. Pharmacol. Ther. 20:642–649.PubMedCrossRefGoogle Scholar
  63. 63.
    Tabata, C., R. Tabata, Y. Kadokawa, S. Hisamori, M. Takahashi, M. Mishima, T. Nakano, and H. Kubo. 2007. Thalidomide prevents bleomycin-induced pulmonary fibrosis in mice. J. Immunol. 179:708–714.PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2008

Authors and Affiliations

  • Ruedi K. Braun
    • 1
    • 5
    Email author
  • Christina Ferrick
    • 2
  • Paul Neubauer
    • 1
  • Michael Sjoding
    • 1
  • Anja Sterner-Kock
    • 3
  • Martin Kock
    • 3
  • Lei Putney
    • 2
  • David A. Ferrick
    • 4
  • Dallas M. Hyde
    • 2
  • Robert B. Love
    • 1
  1. 1.Department of Thoracic and CV SurgeryLoyola UniversityMaywoodUSA
  2. 2.California National Primate Research CenterUniversity of California, DavisDavisUSA
  3. 3.Veterinary PathologyFreie Universität BerlinBerlinGermany
  4. 4.Seahorse BioscienceNorth BillericaUSA
  5. 5.Department of Thoracic and CV SurgeryLoyola UniversityMaywoodUSA

Personalised recommendations